Stem Cell And Tissue Engineering Song Li Nicolas Lheureux Jennifer Elisseeff Editors
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STEM CELL AND
TISSUE ENGINEERING
World Scientific
edited by
Song Li
University of California, Berkeley, USA
Nicolas L’Heureux
Cytograft Tissue Engineering, USA
Jennifer Elisseeff
Johns Hopkins University, USA
7829tp.indd 2 7/27/10 11:42 AM
N E W J E R S E Y • L O N D O N • S I N G A P O R E • B E I J I N G • S H A N G H A I • H O N G K O N G • T A I P E I • C H E N N A I
TISSUE ENGINEERING
World Scientific
edited by
Song Li
University of California, Berkeley, USA
Nicolas L’Heureux
Cytograft Tissue Engineering, USA
Jennifer Elisseeff
Johns Hopkins University, USA
7829tp.indd 2 7/27/10 11:42 AM
N E W J E R S E Y • L O N D O N • S I N G A P O R E • B E I J I N G • S H A N G H A I • H O N G K O N G • T A I P E I • C H E N N A I
British Library Cataloguing-in-Publication Data
A catalogue record for this book is available from the British Library.
For photocopying of material in this volume, please pay a copying fee through the Copyright
Clearance Center, Inc., 222 Rosewood Drive, Danvers, MA 01923, USA. In this case permission to
photocopy is not required from the publisher.
ISBN-13 978-981-4317-05-4
ISBN-10 981-4317-05-5
Typeset by Stallion Press
Email: [email protected]
All rights reserved. This book, or parts thereof, may not be reproduced in any form or by any means,
electronic or mechanical, including photocopying, recording or any information storage and retrieval
system now known or to be invented, without written permission from the Publisher.
Copyright © 2011 by World Scientific Publishing Co. Pte. Ltd.
Published by
World Scientific Publishing Co. Pte. Ltd.
5 Toh Tuck Link, Singapore 596224
USA office: 27 Warren Street, Suite 401-402, Hackensack, NJ 07601
UK office: 57 Shelton Street, Covent Garden, London WC2H 9HE
Printed in Singapore.
STEM CELL AND TISSUE ENGINEERING
A catalogue record for this book is available from the British Library.
For photocopying of material in this volume, please pay a copying fee through the Copyright
Clearance Center, Inc., 222 Rosewood Drive, Danvers, MA 01923, USA. In this case permission to
photocopy is not required from the publisher.
ISBN-13 978-981-4317-05-4
ISBN-10 981-4317-05-5
Typeset by Stallion Press
Email: [email protected]
All rights reserved. This book, or parts thereof, may not be reproduced in any form or by any means,
electronic or mechanical, including photocopying, recording or any information storage and retrieval
system now known or to be invented, without written permission from the Publisher.
Copyright © 2011 by World Scientific Publishing Co. Pte. Ltd.
Published by
World Scientific Publishing Co. Pte. Ltd.
5 Toh Tuck Link, Singapore 596224
USA office: 27 Warren Street, Suite 401-402, Hackensack, NJ 07601
UK office: 57 Shelton Street, Covent Garden, London WC2H 9HE
Printed in Singapore.
STEM CELL AND TISSUE ENGINEERING
Contents
Contributors xiii
Preface xxiii
1 Tissue Engineering: From Basic Biology to Cell-Based 1
Applications
Robert M. Nerem
1. Introduction 1
2. Cell Source 3
3. Stem Cells 5
4. From Benchtop Science to Cell-Based Applications7
5. Concluding Comments 8
Acknowledgments 8
References 9
2 Recent Advances and Future Perspectives 13
on Somatic Cell Reprogramming
Kun-Yong Kim and In-Hyun Park
1. Introduction 13
2. Nuclear Reprogramming 14
3. Reprogramming by Defined Factors 16
v
Contributors xiii
Preface xxiii
1 Tissue Engineering: From Basic Biology to Cell-Based 1
Applications
Robert M. Nerem
1. Introduction 1
2. Cell Source 3
3. Stem Cells 5
4. From Benchtop Science to Cell-Based Applications7
5. Concluding Comments 8
Acknowledgments 8
References 9
2 Recent Advances and Future Perspectives 13
on Somatic Cell Reprogramming
Kun-Yong Kim and In-Hyun Park
1. Introduction 13
2. Nuclear Reprogramming 14
3. Reprogramming by Defined Factors 16
v
4. Recent Advances in Reprogramming Methods 17
5. Future Perspectives on Reprogramming and iPS Cells 19
Acknowledgments 23
References 23
3 Hematopoietic Stem Cells 31
Jennifer J. Trowbridge
1. Introduction 31
2. Hematopoietic Stem Cell Sources 32
3. Applications 36
4. Challenges for Tissue Engineering 37
Acknowledgments 41
References 42
4 Mesenchymal Stem Cells for Tissue Regeneration 49
Ngan F. Huang and Song Li
1. Introduction 49
2. MSC Sources and Phenotype 50
3. Differentiation of MSCs in vitro 52
4. Tissue Engineering and Regeneration Using Bone 54
Marrow MSCs and ASCs
5. Future Directions 61
Acknowledgments 62
References 62
5 Delivery Vehicles for Deploying Mesenchymal Stem Cells 71
in Tissue Repair
Michael S. Friedman and J. Kent Leach
1. Introduction 71
2. Delivery of MSCs for Repairing Cardiovascular Tissues 72
3. Delivery Vehicles for Deploying Stem Cells in 78
Skin Regeneration
4. Biomaterials for Implanting MSCs for Regenerating 81
Osteochondral Tissues
5. Conclusions 88
References 88
vi Contents
5. Future Perspectives on Reprogramming and iPS Cells 19
Acknowledgments 23
References 23
3 Hematopoietic Stem Cells 31
Jennifer J. Trowbridge
1. Introduction 31
2. Hematopoietic Stem Cell Sources 32
3. Applications 36
4. Challenges for Tissue Engineering 37
Acknowledgments 41
References 42
4 Mesenchymal Stem Cells for Tissue Regeneration 49
Ngan F. Huang and Song Li
1. Introduction 49
2. MSC Sources and Phenotype 50
3. Differentiation of MSCs in vitro 52
4. Tissue Engineering and Regeneration Using Bone 54
Marrow MSCs and ASCs
5. Future Directions 61
Acknowledgments 62
References 62
5 Delivery Vehicles for Deploying Mesenchymal Stem Cells 71
in Tissue Repair
Michael S. Friedman and J. Kent Leach
1. Introduction 71
2. Delivery of MSCs for Repairing Cardiovascular Tissues 72
3. Delivery Vehicles for Deploying Stem Cells in 78
Skin Regeneration
4. Biomaterials for Implanting MSCs for Regenerating 81
Osteochondral Tissues
5. Conclusions 88
References 88
vi Contents
6 Stem Cells for Cardiac Tissue Engineering 95
Jennifer L. Young, Karen L. Christman and Adam J. Engler
1. Cell Therapies for Myocardial Infarction and Heart Failure 95
2. Cellular Cardiomyoplasty Revisited: The Influence of 98
in vitroMechanics
3. Tissue Engineering Approach: Utilizing Biomaterial 102
Scaffolds
References 107
7 Cardiovascular System: Stem Cells in Tissue-Engineered 115
Blood Vessels
Rajendra Sawh-Martinez, Edward McGillicuddy,
Gustavo Villalona, Toshiharu Shin’oka
and Christopher K. Breuer
1. Introduction 115
2. Critical Elements of an Artificial Blood Vessel117
3. Approaches to Creating TEBVs 119
4. Conclusion 127
Acknowledgments 128
References 128
8 Stem Cells for Vascular Regeneration: An 135
Engineering Approach
Laura E. Dickinson and Sharon Gerecht
1. Introduction 135
2. Cell Sources 136
3. Engineering Vascular Differentiation 141
4. Three-Dimensional Space 142
Acknowledgments 152
References 152
9 Stem Cells and Wound Repair 159
Sae Hee Ko, Allison Nauta, Geoffrey C. Gurtner
and Michael T. Longaker
1. Clinical Burden of Wound Healing 159
Contents vii
Jennifer L. Young, Karen L. Christman and Adam J. Engler
1. Cell Therapies for Myocardial Infarction and Heart Failure 95
2. Cellular Cardiomyoplasty Revisited: The Influence of 98
in vitroMechanics
3. Tissue Engineering Approach: Utilizing Biomaterial 102
Scaffolds
References 107
7 Cardiovascular System: Stem Cells in Tissue-Engineered 115
Blood Vessels
Rajendra Sawh-Martinez, Edward McGillicuddy,
Gustavo Villalona, Toshiharu Shin’oka
and Christopher K. Breuer
1. Introduction 115
2. Critical Elements of an Artificial Blood Vessel117
3. Approaches to Creating TEBVs 119
4. Conclusion 127
Acknowledgments 128
References 128
8 Stem Cells for Vascular Regeneration: An 135
Engineering Approach
Laura E. Dickinson and Sharon Gerecht
1. Introduction 135
2. Cell Sources 136
3. Engineering Vascular Differentiation 141
4. Three-Dimensional Space 142
Acknowledgments 152
References 152
9 Stem Cells and Wound Repair 159
Sae Hee Ko, Allison Nauta, Geoffrey C. Gurtner
and Michael T. Longaker
1. Clinical Burden of Wound Healing 159
Contents vii
2. Physiology of Wound Healing 161
3. Stem Cells and Wound Repair 165
4. Conclusion 173
References 174
10 Engineering Cartilage: From Materials to Small 181
Molecules
Jeannine M. Coburn and Jennifer H. Elisseeff
1. Introduction 181
2. Structure of Articular Cartilage of the Knee181
3. Osteoarthritis of the Knee 183
4. Surgical Strategies for Repairing Focal Cartilage Defects 185
5. Scaffolds for Assisting Operative Techniques187
6. Mesenchymal Stem Cells for Cartilage Tissue 192
Engineering
7. Hydrogels for Directed Differentiation of Mesenchymal 193
Stem Cells
8. Fiber-Hydrogel Composites 197
9. Small Molecules for Directing Chondrogenesis199
10. Conclusion 201
Acknowledgments 202
References 202
11 Adult Stem Cells for Articular Cartilage Tissue 211
Engineering
Sushmita Saha, Jennifer Kirkham, David Wood,
Stephen Curran and Xuebin B. Yang
1. Introduction 211
2. Human Bone Marrow Mesenchymal Stem Cells 213
(hBMMSCs)
3. Adipose Derived Mesenchymal Stem Cells (ASCs) 216
4. Periosteum Derived Stem/Progenitor Cells 217
(PDSCs/PDPCs)
5. Synovium Derived Mesenchymal Stem Cells (SMSCs) 218
6. Human Dental Pulp Stem Cells (HDPSCs) 219
7. Umbilical Cord/Cord Blood Derived Stem Cells 220
viii Contents
3. Stem Cells and Wound Repair 165
4. Conclusion 173
References 174
10 Engineering Cartilage: From Materials to Small 181
Molecules
Jeannine M. Coburn and Jennifer H. Elisseeff
1. Introduction 181
2. Structure of Articular Cartilage of the Knee181
3. Osteoarthritis of the Knee 183
4. Surgical Strategies for Repairing Focal Cartilage Defects 185
5. Scaffolds for Assisting Operative Techniques187
6. Mesenchymal Stem Cells for Cartilage Tissue 192
Engineering
7. Hydrogels for Directed Differentiation of Mesenchymal 193
Stem Cells
8. Fiber-Hydrogel Composites 197
9. Small Molecules for Directing Chondrogenesis199
10. Conclusion 201
Acknowledgments 202
References 202
11 Adult Stem Cells for Articular Cartilage Tissue 211
Engineering
Sushmita Saha, Jennifer Kirkham, David Wood,
Stephen Curran and Xuebin B. Yang
1. Introduction 211
2. Human Bone Marrow Mesenchymal Stem Cells 213
(hBMMSCs)
3. Adipose Derived Mesenchymal Stem Cells (ASCs) 216
4. Periosteum Derived Stem/Progenitor Cells 217
(PDSCs/PDPCs)
5. Synovium Derived Mesenchymal Stem Cells (SMSCs) 218
6. Human Dental Pulp Stem Cells (HDPSCs) 219
7. Umbilical Cord/Cord Blood Derived Stem Cells 220
viii Contents
8. Other Potential Cell Sources with a Chondrogenic 220
Potential
9. Conclusion and Future Directions 222
Acknowledgments 222
References 222
12 Stem Cells for Disc Repair 231
Aliza A. Allon, Zorica Buser, Sigurd Berven
and Jeffrey C. Lotz
1. Introduction 231
2. The Demanding Intervertebral Disc Environment 233
3. Evaluating a Stem Cell-Based Therapy 234
4. Non-Stem Cell-Based Regeneration Strategies 236
5. Stem Cells for Disc Repair 237
6. Conclusion 244
References 244
13 Skeletal Tissue Engineering: Progress and Prospects 251
Nicholas J. Panetta, Deepak M. Gupta and
Michael T. Longaker
1. Introduction 251
2. Lessons Learned from Endogenous Skeletal Tissue 255
Development, Healing and Regeneration
3. Progenitor Cell-Based Skeletal Tissue Engineering 257
4. Pro-osteogenic Molecular Biology 261
5. Advances in Skeletal Tissue Engineering Scaffolds 266
6. Summary and Future Directions 268
References 269
14 Clinical Applications of a Stem Cell Based Therapy 277
for Oral Bone Reconstruction
Bradley McAllister and Kamran Haghighat
1. Introduction 277
2. Procurement Methodology for Stem Cell Containing 279
Allograft
Contents ix
Potential
9. Conclusion and Future Directions 222
Acknowledgments 222
References 222
12 Stem Cells for Disc Repair 231
Aliza A. Allon, Zorica Buser, Sigurd Berven
and Jeffrey C. Lotz
1. Introduction 231
2. The Demanding Intervertebral Disc Environment 233
3. Evaluating a Stem Cell-Based Therapy 234
4. Non-Stem Cell-Based Regeneration Strategies 236
5. Stem Cells for Disc Repair 237
6. Conclusion 244
References 244
13 Skeletal Tissue Engineering: Progress and Prospects 251
Nicholas J. Panetta, Deepak M. Gupta and
Michael T. Longaker
1. Introduction 251
2. Lessons Learned from Endogenous Skeletal Tissue 255
Development, Healing and Regeneration
3. Progenitor Cell-Based Skeletal Tissue Engineering 257
4. Pro-osteogenic Molecular Biology 261
5. Advances in Skeletal Tissue Engineering Scaffolds 266
6. Summary and Future Directions 268
References 269
14 Clinical Applications of a Stem Cell Based Therapy 277
for Oral Bone Reconstruction
Bradley McAllister and Kamran Haghighat
1. Introduction 277
2. Procurement Methodology for Stem Cell Containing 279
Allograft
Contents ix
3. Ridge Augmentation 282
4. Sinus Augmentation 284
5. Discussion 289
Acknowledgments 292
References 292
15 Therapeutic Strategies for Repairing the Injured Spinal 297
Cord Using Stem Cells
Michael S. Beattie and Jacqueline C. Bresnahan
1. Introduction 297
2. Secondary Injury and Endogenous Repair After SCI 299
3. Therapeutic Targets for Transplanted Stem and300
Progenitor Cells
4. Animal Models of Spinal Cord Injury 301
5. Types of Stem and Progenitor Cells Used for 305
Transplantation in SCI
6. Evidence for Effects on Regeneration and Sprouting 306
7. Evidence for Effects on Neuroprotection 307
8. Evidence for Replacement of Neurons 307
9. Evidence for Oligodendrocyte Replacement and 308
Remyelination
10. Keys to Future Progress 309
11. Are Stem and Progenitor Cell Therapies Ready for 311
Clinical Trials?
Acknowledgments 312
References 312
16 Potential of Tissue Engineering and Neural Stem 321
Cells in the Understanding and Treatment of
Neurodegenerative Diseases
Caroline Auclair-Daigle and François Berthod
1. Introduction 321
2. Neurodegenerative Diseases and Their Current Treatments 322
3. Tissue Engineering as a Tool to Better Understand 324
Neurodegenerative Diseases
4. Neural Stem Cells to Treat Neurodegenerative Diseases 329
x Contents
4. Sinus Augmentation 284
5. Discussion 289
Acknowledgments 292
References 292
15 Therapeutic Strategies for Repairing the Injured Spinal 297
Cord Using Stem Cells
Michael S. Beattie and Jacqueline C. Bresnahan
1. Introduction 297
2. Secondary Injury and Endogenous Repair After SCI 299
3. Therapeutic Targets for Transplanted Stem and300
Progenitor Cells
4. Animal Models of Spinal Cord Injury 301
5. Types of Stem and Progenitor Cells Used for 305
Transplantation in SCI
6. Evidence for Effects on Regeneration and Sprouting 306
7. Evidence for Effects on Neuroprotection 307
8. Evidence for Replacement of Neurons 307
9. Evidence for Oligodendrocyte Replacement and 308
Remyelination
10. Keys to Future Progress 309
11. Are Stem and Progenitor Cell Therapies Ready for 311
Clinical Trials?
Acknowledgments 312
References 312
16 Potential of Tissue Engineering and Neural Stem 321
Cells in the Understanding and Treatment of
Neurodegenerative Diseases
Caroline Auclair-Daigle and François Berthod
1. Introduction 321
2. Neurodegenerative Diseases and Their Current Treatments 322
3. Tissue Engineering as a Tool to Better Understand 324
Neurodegenerative Diseases
4. Neural Stem Cells to Treat Neurodegenerative Diseases 329
x Contents
5. Conclusion 339
Acknowledgments 339
References 340
17 High-Throughput Systems for Stem Cell Engineering 347
David A. Brafman, Karl Willert and Shu Chien
1. Introduction 347
2. Sources of Stem Cells Suitable for High-Throughput 348
Screening Approaches
3. The Stem Cell Niche: A Cellular Microenvironment 349
That Controls Stem Cell Behavior
4. High-Throughput Intrinsic Systems for Stem Cell 363
Investigations
5. Conclusions and Future Trends 365
References 367
18 Microscale Technologies for Tissue Engineering and 375
Stem Cell Differentiation
Jason W. Nichol, Hojae Bae, Nezamoddin N. Kachouie,
Behnam Zamanian, Mahdokht Masaeli and
Ali Khademhosseini
1. Introduction 375
2. Control of Cellular and Tissue Microarchitecture377
3. Microscale Technologies to Investigate and Control 380
Stem Cell Behavior
4. Assembly Techniques for Creating Engineered Tissues 385
from Microscale Building Blocks
5. Conclusions and Future Directions 391
References 391
19 Quality Control of Autologous Cell- and Tissue-Based 397
Therapies
Nathalie Dusserre, Todd McAllister and Nicolas L’Heureux
1. Introduction 397
2. Regulations Pertaining to Quality Control of Cell- 398
and Tissue-Based Products
Contents xi
Acknowledgments 339
References 340
17 High-Throughput Systems for Stem Cell Engineering 347
David A. Brafman, Karl Willert and Shu Chien
1. Introduction 347
2. Sources of Stem Cells Suitable for High-Throughput 348
Screening Approaches
3. The Stem Cell Niche: A Cellular Microenvironment 349
That Controls Stem Cell Behavior
4. High-Throughput Intrinsic Systems for Stem Cell 363
Investigations
5. Conclusions and Future Trends 365
References 367
18 Microscale Technologies for Tissue Engineering and 375
Stem Cell Differentiation
Jason W. Nichol, Hojae Bae, Nezamoddin N. Kachouie,
Behnam Zamanian, Mahdokht Masaeli and
Ali Khademhosseini
1. Introduction 375
2. Control of Cellular and Tissue Microarchitecture377
3. Microscale Technologies to Investigate and Control 380
Stem Cell Behavior
4. Assembly Techniques for Creating Engineered Tissues 385
from Microscale Building Blocks
5. Conclusions and Future Directions 391
References 391
19 Quality Control of Autologous Cell- and Tissue-Based 397
Therapies
Nathalie Dusserre, Todd McAllister and Nicolas L’Heureux
1. Introduction 397
2. Regulations Pertaining to Quality Control of Cell- 398
and Tissue-Based Products
Contents xi
3. cGMP, cGTP and Quality System 401
4. Core Requirements of a Quality Program 402
5. Tailoring Quality Control to the Manufacturing Process 407
6. Conclusion 417
Acknowledgments 418
References 418
20 Regulatory Challenges for Cell-Based Therapeutics 423
Todd McAllister, Corey Iyican, Nathalie Dusserre
and Nicolas L’Heureux
1. Introduction 423
2. Regulatory Challenges at Each Phase of Clinical425
Development
3. Regional Considerations for Clinical Trials 430
4. Use of a Clinical Research Organization 435
5. Universal Regulatory Considerations for Cell-Based 437
Therapeutics
References 439
Index 441
xii Contents
4. Core Requirements of a Quality Program 402
5. Tailoring Quality Control to the Manufacturing Process 407
6. Conclusion 417
Acknowledgments 418
References 418
20 Regulatory Challenges for Cell-Based Therapeutics 423
Todd McAllister, Corey Iyican, Nathalie Dusserre
and Nicolas L’Heureux
1. Introduction 423
2. Regulatory Challenges at Each Phase of Clinical425
Development
3. Regional Considerations for Clinical Trials 430
4. Use of a Clinical Research Organization 435
5. Universal Regulatory Considerations for Cell-Based 437
Therapeutics
References 439
Index 441
xii Contents
Contributors
Aliza A. Allon
Department of Orthopedic Surgery
University of California San Francisco
San Francisco, CA 94110
Caroline Auclair-Daigle
Laboratoire d’Organogénèse Expérimentale (LOEX)
Centre de recherche FRSQ du CHA universitaire de Québec
Hôpital du Saint-Sacrement, and Département de chirurgie
Faculté de médecine, Université Laval
Québec, Canada
Hojae Bae
Harvard-MIT Division of Health Sciences and Technology
Center for Biomedical Engineering, Department of Medicine
Brigham and Women’s Hospital, Harvard Medical School
Massachusetts Institute of Technology
Cambridge, MA 02139
xiii
Aliza A. Allon
Department of Orthopedic Surgery
University of California San Francisco
San Francisco, CA 94110
Caroline Auclair-Daigle
Laboratoire d’Organogénèse Expérimentale (LOEX)
Centre de recherche FRSQ du CHA universitaire de Québec
Hôpital du Saint-Sacrement, and Département de chirurgie
Faculté de médecine, Université Laval
Québec, Canada
Hojae Bae
Harvard-MIT Division of Health Sciences and Technology
Center for Biomedical Engineering, Department of Medicine
Brigham and Women’s Hospital, Harvard Medical School
Massachusetts Institute of Technology
Cambridge, MA 02139
xiii
Michael S. Beattie
Brain and Spinal Injury Center
Department of Neurological Surgery
University of California San Francisco
San Francisco, CA 94110
François Berthod
Laboratoire d’Organogénèse Expérimentale (LOEX)
Centre de recherche FRSQ du CHA universitaire de Québec
Hôpital du Saint-Sacrement, and Département de chirurgie
Faculté de médecine, Université Laval
Québec, Canada
Sigurd Berven
Department of Orthopedic Surgery
University of California San Francisco
San Francisco, CA 94110
David A. Brafman
Department of Bioengineering
Institute of Engineering in Medicine
University of California San Diego
San Diego, CA 92093
Jacqueline C. Bresnahan
Brain and Spinal Injury Cente
Department of Neurological Surgery
University of California San Francisco
San Francisco, CA 94110
Christopher K. Breuer
Interdepartmental Program in Vascular Biology and Therapeutics
Yale University School of Medicine
New Haven, CT 06520
xiv Contributors
Brain and Spinal Injury Center
Department of Neurological Surgery
University of California San Francisco
San Francisco, CA 94110
François Berthod
Laboratoire d’Organogénèse Expérimentale (LOEX)
Centre de recherche FRSQ du CHA universitaire de Québec
Hôpital du Saint-Sacrement, and Département de chirurgie
Faculté de médecine, Université Laval
Québec, Canada
Sigurd Berven
Department of Orthopedic Surgery
University of California San Francisco
San Francisco, CA 94110
David A. Brafman
Department of Bioengineering
Institute of Engineering in Medicine
University of California San Diego
San Diego, CA 92093
Jacqueline C. Bresnahan
Brain and Spinal Injury Cente
Department of Neurological Surgery
University of California San Francisco
San Francisco, CA 94110
Christopher K. Breuer
Interdepartmental Program in Vascular Biology and Therapeutics
Yale University School of Medicine
New Haven, CT 06520
xiv Contributors
Zorica Buser
Department of Orthopedic Surgery
University of California San Francisco
San Francisco, CA 94122
Shu Chien
Departments of Bioengineering, Cellular and Molecular Medicine,
and Medicine; Institute of Engineering in Medicine
University of California San Diego
San Diego, CA 92093
Karen L. Christman
Department of Bioengineering
University of California San Diego
La Jolla, CA 92093
Jeannine M. Coburn
Department of Chemical and Biomolecular Engineering
Johns Hopkins University
Baltimore, MD 21218
Stephen Curran
Smith & Nephew Research Centre, York Science Park
York YO10 5DF, United Kingdom
Laura E. Dickinson
Department of Chemical and Biomolecular Engineering
Johns Hopkins University
Baltimore, MD 21218
Nathalie Dusserre
Cytograft Tissue Engineering
Novato, CA 94949
Jennifer H. Elisseeff
Department of Biomedical Engineering
John Hopkins University
Baltimore, MD 21218
Contributors xv
Department of Orthopedic Surgery
University of California San Francisco
San Francisco, CA 94122
Shu Chien
Departments of Bioengineering, Cellular and Molecular Medicine,
and Medicine; Institute of Engineering in Medicine
University of California San Diego
San Diego, CA 92093
Karen L. Christman
Department of Bioengineering
University of California San Diego
La Jolla, CA 92093
Jeannine M. Coburn
Department of Chemical and Biomolecular Engineering
Johns Hopkins University
Baltimore, MD 21218
Stephen Curran
Smith & Nephew Research Centre, York Science Park
York YO10 5DF, United Kingdom
Laura E. Dickinson
Department of Chemical and Biomolecular Engineering
Johns Hopkins University
Baltimore, MD 21218
Nathalie Dusserre
Cytograft Tissue Engineering
Novato, CA 94949
Jennifer H. Elisseeff
Department of Biomedical Engineering
John Hopkins University
Baltimore, MD 21218
Contributors xv
Adam J. Engler
Department of Bioengineering
University of California San Diego
La Jolla, CA 92093
Michael S. Friedman
ThermoGenesis Corporation
Rancho Cordova, CA 95742-6303
Sharon Gerecht
Department of Chemical and Biomolecular Engineering
John Hopkins University
Baltimore, MD 21218
Deepak M. Gupta
Department of Surgery
Stanford University School of Medicine
Stanford, CA 94305-5406
Geoffrey C. Gurtner
Hagey Laboratory for Pediatric and Regenerative Medicine
Division of Plastic and Reconstructive Surgery
Department of Surgery
Institute of Stem Cell Biology and Regenerative Medicine
Stanford University School of Medicine
Stanford, CA 94305
Kamran Haghighat
Private Practice
Portland, OR 97205
Ngan F. Huang
Division of Cardiovascular Medicine
Stanford University
Stanford, CA 94305-5406
xvi Contributors
Department of Bioengineering
University of California San Diego
La Jolla, CA 92093
Michael S. Friedman
ThermoGenesis Corporation
Rancho Cordova, CA 95742-6303
Sharon Gerecht
Department of Chemical and Biomolecular Engineering
John Hopkins University
Baltimore, MD 21218
Deepak M. Gupta
Department of Surgery
Stanford University School of Medicine
Stanford, CA 94305-5406
Geoffrey C. Gurtner
Hagey Laboratory for Pediatric and Regenerative Medicine
Division of Plastic and Reconstructive Surgery
Department of Surgery
Institute of Stem Cell Biology and Regenerative Medicine
Stanford University School of Medicine
Stanford, CA 94305
Kamran Haghighat
Private Practice
Portland, OR 97205
Ngan F. Huang
Division of Cardiovascular Medicine
Stanford University
Stanford, CA 94305-5406
xvi Contributors
Corey Iyican
Cytograft Tissue Engineering
Novato, CA 94949
Nezamoddin N. Kachouie
Harvard-MIT Division of Health Sciences and Technology
Center for Biomedical Engineering, Department of Medicine
Brigham and Women’s Hospital, Harvard Medical School
Massachusetts Institute of Technology
Cambridge, MA 02139
Ali Khademhosseini
Harvard-MIT Division of Health Sciences and Technology
Center for Biomedical Engineering, Department of Medicine
Brigham and Women’s Hospital, Harvard Medical School
Massachusetts Institute of Technology
Cambridge, MA 02139
Kun-Yong Kim
Department of Genetics, Yale Stem Cell Center
Yale University School of Medicine
New Haven, CT 06520
Jennifer Kirkham
Biomaterials and Tissue Engineering Group, Leeds Dental Institute
University of Leeds
Leeds LS2 9LU, United Kingdom
Sae Hee Ko
Hagey Laboratory for Pediatric and Regenerative Medicine
Division of Plastic and Reconstructive Surgery
Department of Surgery
Institute of Stem Cell Biology and Regenerative Medicine
Stanford University School of Medicine
Stanford, CA 94305
Contributors xvii
Cytograft Tissue Engineering
Novato, CA 94949
Nezamoddin N. Kachouie
Harvard-MIT Division of Health Sciences and Technology
Center for Biomedical Engineering, Department of Medicine
Brigham and Women’s Hospital, Harvard Medical School
Massachusetts Institute of Technology
Cambridge, MA 02139
Ali Khademhosseini
Harvard-MIT Division of Health Sciences and Technology
Center for Biomedical Engineering, Department of Medicine
Brigham and Women’s Hospital, Harvard Medical School
Massachusetts Institute of Technology
Cambridge, MA 02139
Kun-Yong Kim
Department of Genetics, Yale Stem Cell Center
Yale University School of Medicine
New Haven, CT 06520
Jennifer Kirkham
Biomaterials and Tissue Engineering Group, Leeds Dental Institute
University of Leeds
Leeds LS2 9LU, United Kingdom
Sae Hee Ko
Hagey Laboratory for Pediatric and Regenerative Medicine
Division of Plastic and Reconstructive Surgery
Department of Surgery
Institute of Stem Cell Biology and Regenerative Medicine
Stanford University School of Medicine
Stanford, CA 94305
Contributors xvii
J. Kent Leach
Department of Biomedical Engineering
University of California Davis
Davis, CA 95616
Nicolas L’Heureux
Cytograft Tissue Engineering
Novato, CA 94949
Song Li
Department of Bioengineering
University of California Berkeley
Berkeley, CA 94720-1762
Michael T. Longaker
Hagey Laboratory for Pediatric and Regenerative Medicine
Division of Plastic and Reconstructive Surgery
Department of Surgery
Institute of Stem Cell Biology and Regenerative Medicine
Stanford University School of Medicine
Stanford, CA 94305
Jeffrey C. Lotz
Department of Orthopedic Surgery
University of California San Francisco
San Francisco, CA 94110
Mahdokht Masaeli
Department of Electrical and Computer Engineering
Northeastern University
Boston, MA 02115
Bradley McAllister
Department of Periodontology
Oregon Health Sciences University
Portland, OR 97239
xviii Contributors
Department of Biomedical Engineering
University of California Davis
Davis, CA 95616
Nicolas L’Heureux
Cytograft Tissue Engineering
Novato, CA 94949
Song Li
Department of Bioengineering
University of California Berkeley
Berkeley, CA 94720-1762
Michael T. Longaker
Hagey Laboratory for Pediatric and Regenerative Medicine
Division of Plastic and Reconstructive Surgery
Department of Surgery
Institute of Stem Cell Biology and Regenerative Medicine
Stanford University School of Medicine
Stanford, CA 94305
Jeffrey C. Lotz
Department of Orthopedic Surgery
University of California San Francisco
San Francisco, CA 94110
Mahdokht Masaeli
Department of Electrical and Computer Engineering
Northeastern University
Boston, MA 02115
Bradley McAllister
Department of Periodontology
Oregon Health Sciences University
Portland, OR 97239
xviii Contributors
Todd McAllister
Cytograft Tissue Engineering
Novato, CA 94949
Edward McGillicuddy
Interdepartmental Program in Vascular Biology and Therapeutics
Yale University School of Medicine
New Haven, CT 06520
Allison Nauta
Hagey Laboratory for Pediatric and Regenerative Medicine
Division of Plastic and Reconstructive Surgery
Department of Surgery
Institute of Stem Cell Biology and Regenerative Medicine
Stanford University School of Medicine
Stanford, CA 94305
Georgetown University Hospital, Washington DC 20007
Robert M. Nerem
The Georgia Tech/Emory Center (GTEC) for Regenerative Medicine
Emory University
Atlanta, GA 30322
Jason W. Nichol
Harvard-MIT Division of Health Sciences and Technology
Center for Biomedical Engineering, Department of Medicine
Brigham and Women’s Hospital, Harvard Medical School
Massachusetts Institute of Technology
Cambridge, MA 02139
Nicholas J. Panetta
Department of Surgery
Stanford University School of Medicine
Stanford, CA 94305-5406
Contributors xix
Cytograft Tissue Engineering
Novato, CA 94949
Edward McGillicuddy
Interdepartmental Program in Vascular Biology and Therapeutics
Yale University School of Medicine
New Haven, CT 06520
Allison Nauta
Hagey Laboratory for Pediatric and Regenerative Medicine
Division of Plastic and Reconstructive Surgery
Department of Surgery
Institute of Stem Cell Biology and Regenerative Medicine
Stanford University School of Medicine
Stanford, CA 94305
Georgetown University Hospital, Washington DC 20007
Robert M. Nerem
The Georgia Tech/Emory Center (GTEC) for Regenerative Medicine
Emory University
Atlanta, GA 30322
Jason W. Nichol
Harvard-MIT Division of Health Sciences and Technology
Center for Biomedical Engineering, Department of Medicine
Brigham and Women’s Hospital, Harvard Medical School
Massachusetts Institute of Technology
Cambridge, MA 02139
Nicholas J. Panetta
Department of Surgery
Stanford University School of Medicine
Stanford, CA 94305-5406
Contributors xix
In-Hyun Park
Department of Genetics, Yale Stem Cell Center
Yale University School of Medicine
New Haven, CT 06520
Sushmita Saha
Biomaterials and Tissue Engineering Group, Leeds Dental Institute
University of Leeds
Leeds LS2 9LU, United Kingdom
Rajendra Sawh-Martinez
Interdepartmental Program in Vascular Biology and Therapeutics
Yale University School of Medicine
New Haven, CT 06520
Toshiharu Shin’oka
Interdepartmental Program in Vascular Biology and Therapeutics
Yale University School of Medicine
New Haven, CT 06520
Jennifer J. Trowbridge
Department of Pediatric Oncology, Dana-Farber Cancer Institute
Division of Hematology/Oncology, Children’s Hospital Boston
Harvard Stem Cell Institute, Harvard Medical School
Boston, MA 02215
Gustavo Villalona
Interdepartmental Program in Vascular Biology and Therapeutics
Yale University School of Medicine
New Haven, CT 06520
Karl Willert
Department of Cellular and Molecular Medicine
Institute of Engineering in Medicine
University of California San Diego
San Diego, CA 92093
xx Contributors
Department of Genetics, Yale Stem Cell Center
Yale University School of Medicine
New Haven, CT 06520
Sushmita Saha
Biomaterials and Tissue Engineering Group, Leeds Dental Institute
University of Leeds
Leeds LS2 9LU, United Kingdom
Rajendra Sawh-Martinez
Interdepartmental Program in Vascular Biology and Therapeutics
Yale University School of Medicine
New Haven, CT 06520
Toshiharu Shin’oka
Interdepartmental Program in Vascular Biology and Therapeutics
Yale University School of Medicine
New Haven, CT 06520
Jennifer J. Trowbridge
Department of Pediatric Oncology, Dana-Farber Cancer Institute
Division of Hematology/Oncology, Children’s Hospital Boston
Harvard Stem Cell Institute, Harvard Medical School
Boston, MA 02215
Gustavo Villalona
Interdepartmental Program in Vascular Biology and Therapeutics
Yale University School of Medicine
New Haven, CT 06520
Karl Willert
Department of Cellular and Molecular Medicine
Institute of Engineering in Medicine
University of California San Diego
San Diego, CA 92093
xx Contributors
David Wood
Biomaterials and Tissue Engineering Group, Leeds Dental Institute
University of Leeds
Leeds LS2 9LU, United Kingdom
Xuebin B. Yang
Biomaterials and Tissue Engineering Group, Leeds Dental Institute
University of Leeds
Leeds LS2 9LU, United Kingdom
Jennifer L. Young
Department of Bioengineering
University of California San Diego
La Jolla, CA 92093
Behnam Zamanian
Harvard-MIT Division of Health Sciences and Technology
Center for Biomedical Engineering, Department of Medicine
Brigham and Women’s Hospital, Harvard Medical School
Massachusetts Institute of Technology
Cambridge, MA 02139
Contributors xxi
Biomaterials and Tissue Engineering Group, Leeds Dental Institute
University of Leeds
Leeds LS2 9LU, United Kingdom
Xuebin B. Yang
Biomaterials and Tissue Engineering Group, Leeds Dental Institute
University of Leeds
Leeds LS2 9LU, United Kingdom
Jennifer L. Young
Department of Bioengineering
University of California San Diego
La Jolla, CA 92093
Behnam Zamanian
Harvard-MIT Division of Health Sciences and Technology
Center for Biomedical Engineering, Department of Medicine
Brigham and Women’s Hospital, Harvard Medical School
Massachusetts Institute of Technology
Cambridge, MA 02139
Contributors xxi
Preface
Cells are the building blocks of tissues and organs. Therefore, cell source
is a critical issue for tissue engineering. An ideal cell source should be suf-
ficient in quantity, compatible with the immune system of the recipient
and free of pathogens or contamination. Depending on the specific tissue
engineering application, the cell source can be autologous, allogeneic or
xenogenic. Traditionally, fully differentiated cell types are used to engi-
neer tissues. However, for many cell types, differentiated cells from adult
tissues often have little or no proliferation potential.
In the past few years, the advancement of stem cell biology has opened
a new avenue for tissue engineering. Stem cells can be isolated from adult
tissues, fetal tissues or embryos, are highly expandable, and can be
directed to differentiate into specific cell types. Furthermore, recent
breakthroughs in cell reprogramming make it possible to take tissue biop-
sies from patients and reprogram the cells into pluripotent stem cells or
specific cell types such as neurons or cardiomyocytes. This progress has
allowed tissue engineers to have access to unlimited, immune-acceptable
cell sources. To fully harness the therapeutic potential of stem cells, we
need to understand how stem cells respond to microenvironmental factors
including both biochemical and biophysical cues. This is not only
required for controlling cell fate in vitro, but it is also important for the
design of scaffolds and tissue constructs that can maximize the recruitment
xxiii
Cells are the building blocks of tissues and organs. Therefore, cell source
is a critical issue for tissue engineering. An ideal cell source should be suf-
ficient in quantity, compatible with the immune system of the recipient
and free of pathogens or contamination. Depending on the specific tissue
engineering application, the cell source can be autologous, allogeneic or
xenogenic. Traditionally, fully differentiated cell types are used to engi-
neer tissues. However, for many cell types, differentiated cells from adult
tissues often have little or no proliferation potential.
In the past few years, the advancement of stem cell biology has opened
a new avenue for tissue engineering. Stem cells can be isolated from adult
tissues, fetal tissues or embryos, are highly expandable, and can be
directed to differentiate into specific cell types. Furthermore, recent
breakthroughs in cell reprogramming make it possible to take tissue biop-
sies from patients and reprogram the cells into pluripotent stem cells or
specific cell types such as neurons or cardiomyocytes. This progress has
allowed tissue engineers to have access to unlimited, immune-acceptable
cell sources. To fully harness the therapeutic potential of stem cells, we
need to understand how stem cells respond to microenvironmental factors
including both biochemical and biophysical cues. This is not only
required for controlling cell fate in vitro, but it is also important for the
design of scaffolds and tissue constructs that can maximize the recruitment
xxiii
of adult stem cells following implantation. In addition, to cultivate cells
for clinical applications, quality control and FDA requirements must be
fulfilled.
Tissue engineering using stem cells is an emerging and fast-growing
field. There is a pressing need for a book that provides a comprehensive
introduction to the field and summarizes its recent progress. We have
invited experts in their respective fields to provide insightful reviews of
specific topics on stem cells and tissue engineering. We hope that this
book is timely and useful for researchers and students. Chapters 1 to 4 of
this book introduce tissue engineering (Chapter 1) and discuss different
types of stem cells (Chapters 2 to 4). Chapters 5 to 8 discuss the use of
stem cells and biomaterials for the regeneration of cardiac tissue, blood
vessels and the vascular network. Chapter 9 reviews the role of stem cells
in general wound repair. Chapters 10 to 14 focus on skeletal tissue engi-
neering, including cartilage, intervertebral disc and bone. Chapters 15 and
16 review the use of stem cells to treat spinal cord injury and neurode-
generative diseases. Chapters 17 and 18 illustrate state-of-art technologies
used in stem cell engineering, including high-throughput systems and
microtechnologies. Chapters 19 and 20 discuss quality control and regu-
latory issues. Although this book does not have the capacity to cover the
use of stem cells for all tissues and organs, we hope that the general con-
cepts and approaches illustrated in this book are helpful for researchers
who are interested in other tissues and organs that are not discussed here.
We thank all the contributors for their hard work and valuable contri-
butions. We also thank Joy Quek and the other staff of World Scientific
Inc. for their tremendous effort in editing and organizing this book.
Song Li, Ph.D.
University of California, Berkeley
Nicolas L’Heureux, Ph.D.
Cytograft, Inc.
Jennifer Elisseeff, Ph.D.
Johns Hopkins University
xxiv Preface
for clinical applications, quality control and FDA requirements must be
fulfilled.
Tissue engineering using stem cells is an emerging and fast-growing
field. There is a pressing need for a book that provides a comprehensive
introduction to the field and summarizes its recent progress. We have
invited experts in their respective fields to provide insightful reviews of
specific topics on stem cells and tissue engineering. We hope that this
book is timely and useful for researchers and students. Chapters 1 to 4 of
this book introduce tissue engineering (Chapter 1) and discuss different
types of stem cells (Chapters 2 to 4). Chapters 5 to 8 discuss the use of
stem cells and biomaterials for the regeneration of cardiac tissue, blood
vessels and the vascular network. Chapter 9 reviews the role of stem cells
in general wound repair. Chapters 10 to 14 focus on skeletal tissue engi-
neering, including cartilage, intervertebral disc and bone. Chapters 15 and
16 review the use of stem cells to treat spinal cord injury and neurode-
generative diseases. Chapters 17 and 18 illustrate state-of-art technologies
used in stem cell engineering, including high-throughput systems and
microtechnologies. Chapters 19 and 20 discuss quality control and regu-
latory issues. Although this book does not have the capacity to cover the
use of stem cells for all tissues and organs, we hope that the general con-
cepts and approaches illustrated in this book are helpful for researchers
who are interested in other tissues and organs that are not discussed here.
We thank all the contributors for their hard work and valuable contri-
butions. We also thank Joy Quek and the other staff of World Scientific
Inc. for their tremendous effort in editing and organizing this book.
Song Li, Ph.D.
University of California, Berkeley
Nicolas L’Heureux, Ph.D.
Cytograft, Inc.
Jennifer Elisseeff, Ph.D.
Johns Hopkins University
xxiv Preface
1
Tissue Engineering: From Basic
Biology to Cell-Based Applications
Robert M. Nerem
1. Introduction
With the advent of the 21st century, the use of tissue engineering-based
therapies to treat a variety of diseases and/or injuries has moved from
being a dream of what might be possible in the future to the realm of the
achievable. Even so, there is still much that needs to be done, much still
to be learned; however, it is clear that major advances have been made in
the last two decades.
The term tissue engineering is used here to describe a wide variety of
approaches. This includes the replacement, the repair, and/or the regener-
ation of tissues and organs. Different terms have been used to describe this
harnessing of the intrinsic biological abilities of the human body and of
living cells. Although research in this general area goes back nearly half a
century and the possibilities of such approaches was described even ear-
lier, it was in 1987 that the term tissue engineering was introduced.
1
Then
in the 1990s the term regenerative medicine came into use. For some these
terms are interchangeable. For others tissue engineering is used for
1
Tissue Engineering: From Basic
Biology to Cell-Based Applications
Robert M. Nerem
1. Introduction
With the advent of the 21st century, the use of tissue engineering-based
therapies to treat a variety of diseases and/or injuries has moved from
being a dream of what might be possible in the future to the realm of the
achievable. Even so, there is still much that needs to be done, much still
to be learned; however, it is clear that major advances have been made in
the last two decades.
The term tissue engineering is used here to describe a wide variety of
approaches. This includes the replacement, the repair, and/or the regener-
ation of tissues and organs. Different terms have been used to describe this
harnessing of the intrinsic biological abilities of the human body and of
living cells. Although research in this general area goes back nearly half a
century and the possibilities of such approaches was described even ear-
lier, it was in 1987 that the term tissue engineering was introduced.
1
Then
in the 1990s the term regenerative medicine came into use. For some these
terms are interchangeable. For others tissue engineering is used for
1
approaches that are aimed at fabricating substitute tissues outside of the
body that then can be implanted into the body as replacements. For some
the term regenerative medicine means stem cell technology. For this
author, however, tissue engineering has a much broader meaning. Thus, to
minimize any confusion associated with the choice of terms, it is the term
tissue engineering that will be used in this introductory chapter and it will
be used in the broadest sense to include replacement, repair, and regener-
ation, i.e. the wide variety of approaches that harness the intrinsic
biological ability of the body and the use of living cells, whether of exoge-
nous or endogenous origin.
It should be noted that over the past two decades the industry associ-
ated with this field has had its “ups and downs.” There were products
being developed in the 1990s, and these were largely skin substitutes. The
leading companies were Advanced Tissue Sciences (ATS) and
Organogenesis (OI). As we entered the 21st century, however, we encoun-
tered what might be called “the sobering years.” Both ATS and OI entered
bankruptcy. Today ATS no longer exists, but OI has reinvented itself and
is a profitable company. In fact, in the last five years there has been a
resurgence of the industry. In 2007, the last year for which there is data
available, total industrial activity was US$2.4. billion with more than half
of this being the sale of commercial products.
2
For development stage
funding the largest component is that of stem cells.
Whatever the approach being used in tissue engineering, a critical
issue is the source of the cells to be employed. This thus will be
addressed in the next section. One possibility of course is to use stem
cells, and since the early reports of human stem cells a decade ago
3–7
there has been a surge of activity. As stem cells and tissue engineering
are the focus of this book, a brief introduction to stem cells is
provided. To employ stem cells in a cell-based therapy will require,
however, the translation of the basic benchtop science to the variety of
applications that are possible. This is an area that has been largely
overlooked, certainly not addressed to the extent necessary, and in the
next to last section of this introductory chapter the issues that need to
be addressed as one moves from the basic stem cell biology research
to applications will be briefly discussed. The chapter then ends with
some concluding comments.
2 R. M. Nerem
body that then can be implanted into the body as replacements. For some
the term regenerative medicine means stem cell technology. For this
author, however, tissue engineering has a much broader meaning. Thus, to
minimize any confusion associated with the choice of terms, it is the term
tissue engineering that will be used in this introductory chapter and it will
be used in the broadest sense to include replacement, repair, and regener-
ation, i.e. the wide variety of approaches that harness the intrinsic
biological ability of the body and the use of living cells, whether of exoge-
nous or endogenous origin.
It should be noted that over the past two decades the industry associ-
ated with this field has had its “ups and downs.” There were products
being developed in the 1990s, and these were largely skin substitutes. The
leading companies were Advanced Tissue Sciences (ATS) and
Organogenesis (OI). As we entered the 21st century, however, we encoun-
tered what might be called “the sobering years.” Both ATS and OI entered
bankruptcy. Today ATS no longer exists, but OI has reinvented itself and
is a profitable company. In fact, in the last five years there has been a
resurgence of the industry. In 2007, the last year for which there is data
available, total industrial activity was US$2.4. billion with more than half
of this being the sale of commercial products.
2
For development stage
funding the largest component is that of stem cells.
Whatever the approach being used in tissue engineering, a critical
issue is the source of the cells to be employed. This thus will be
addressed in the next section. One possibility of course is to use stem
cells, and since the early reports of human stem cells a decade ago
3–7
there has been a surge of activity. As stem cells and tissue engineering
are the focus of this book, a brief introduction to stem cells is
provided. To employ stem cells in a cell-based therapy will require,
however, the translation of the basic benchtop science to the variety of
applications that are possible. This is an area that has been largely
overlooked, certainly not addressed to the extent necessary, and in the
next to last section of this introductory chapter the issues that need to
be addressed as one moves from the basic stem cell biology research
to applications will be briefly discussed. The chapter then ends with
some concluding comments.
2 R. M. Nerem
2. Cell Source
Whatever the tissue engineering approach, whether it be one of replace-
ment or that of repair and regeneration, a critical issue is that of cell
source, i.e. from where will the cells come that are to be employed in the
treatment or therapy. In addressing this issue, there are several questions
that need to be asked. These are as follows.
•Will the source of cells be endogenous or exogenous?
•Will one use undifferentiated stem cells, progenitor cells, or fully
differentiated somatic cells?
•Will one employ an autologous cell strategy or an allogeneic or even
xenogeneic strategy?
•Are there differences associated with the age of the donor or with the
disease state?
•Are there sex differences that must be taken into account?
Let us consider these one by one.
To start with, is the strategy one of recruiting cells from within the
patient or one using an exogenous source? If the former, then the approach
is an autologous one, and the challenge is how to recruit the cells. If the
latter, then one must proceed to a series of additional questions.
This book focuses on stem cells; however, it also includes the use of
progenitor cells that in fact can be derived from stem cells. Furthermore,
and as will be discussed in the next section, there are different types of
stem cells, e.g. embryonic versus adult, and these may be different in their
ability to be differentiated into the particular type of cell to be used in the
therapy. Even starting with a stem cell, however, does one use the stem
cell directly in the therapy, does one use a progenitor cell derived from the
stem cell, or does one use a fully differentiated cell?
One question for any clinical therapeutic strategy is that of autolo-
gous vs. allogeneic vs. xenogeneic cells. Although the use of autologous
cells is attractive from the viewpoint of immunogenicity, the use of
autologous cells does not in general provide for off-the-shelf availabil-
ity to the clinician. Why is off-the-shelf availability important? For
surgeries that must be carried out on short notice, e.g. following a heart
Tissue Engineering and Stem Cells 3
Whatever the tissue engineering approach, whether it be one of replace-
ment or that of repair and regeneration, a critical issue is that of cell
source, i.e. from where will the cells come that are to be employed in the
treatment or therapy. In addressing this issue, there are several questions
that need to be asked. These are as follows.
•Will the source of cells be endogenous or exogenous?
•Will one use undifferentiated stem cells, progenitor cells, or fully
differentiated somatic cells?
•Will one employ an autologous cell strategy or an allogeneic or even
xenogeneic strategy?
•Are there differences associated with the age of the donor or with the
disease state?
•Are there sex differences that must be taken into account?
Let us consider these one by one.
To start with, is the strategy one of recruiting cells from within the
patient or one using an exogenous source? If the former, then the approach
is an autologous one, and the challenge is how to recruit the cells. If the
latter, then one must proceed to a series of additional questions.
This book focuses on stem cells; however, it also includes the use of
progenitor cells that in fact can be derived from stem cells. Furthermore,
and as will be discussed in the next section, there are different types of
stem cells, e.g. embryonic versus adult, and these may be different in their
ability to be differentiated into the particular type of cell to be used in the
therapy. Even starting with a stem cell, however, does one use the stem
cell directly in the therapy, does one use a progenitor cell derived from the
stem cell, or does one use a fully differentiated cell?
One question for any clinical therapeutic strategy is that of autolo-
gous vs. allogeneic vs. xenogeneic cells. Although the use of autologous
cells is attractive from the viewpoint of immunogenicity, the use of
autologous cells does not in general provide for off-the-shelf availabil-
ity to the clinician. Why is off-the-shelf availability important? For
surgeries that must be carried out on short notice, e.g. following a heart
Tissue Engineering and Stem Cells 3
attack, off-the-shelf availability of the cells to be employed in the
therapy is essential; however, even when the time of surgery is elective,
one can argue that only with off-the-shelf availability will the wider
patient population that is in need be served. There is of course one
exception to the above generalization, and this is if the cells to be
employed are to be recruited from within the body of the patient. In this
case what is needed by the clinician off-the-shelf is in fact not the cells
themselves, but perhaps only an acellular implant to be used in recruit-
ing the cells and/or to serve as a target for the cells. In contrast,
allogeneic cells or even xenogeneic cells do provide for off-the-shelf
availability. Here the challenge is that of immunogenicity. The problem
of achieving immune acceptance with xenogeneic cells is particularly
severe; however, even for allogeneic cells for at least some cell types,
e.g. vascular endothelial cells, a strategy for creating immune accep-
tance would have to be used.
Finally, there are the questions of differences due to age, due to the
disease state of the patient, or due to the sex of the donor. Although largely
unexplored, these can be significant issues, and it is important that future
research addresses these questions. For example, there is a report that
patient characteristics affects the number of human cardiac progenitor
cells that will be available.
8
There also is evidence that the disease state
can have influence. In this case an example is that in patients with coro-
nary artery disease there was observed a functional impairment of the
hematopoietic progenitor cells.
9
There also are sex differences in the basic characteristics of cells
whether they be stem cells, progenitor cells, or fully differentiated cells.
This is a very important area, one which gives rise to a variety of ques-
tions. For example, if the cells are to be cultured, are different culturing
protocols required for female cells as compared to male cells? For the
clinical therapy itself, will the outcome be different depending on sex of
the donor versus the sex of the patient? In this latter case, there are reports
in the literature documenting differences.
10,11
There thus are a variety of questions to be answered, and although there
is developing a rich literature, further research is required. In the next
section, however, we move on to a very brief discussion of stem cells.
4 R. M. Nerem
therapy is essential; however, even when the time of surgery is elective,
one can argue that only with off-the-shelf availability will the wider
patient population that is in need be served. There is of course one
exception to the above generalization, and this is if the cells to be
employed are to be recruited from within the body of the patient. In this
case what is needed by the clinician off-the-shelf is in fact not the cells
themselves, but perhaps only an acellular implant to be used in recruit-
ing the cells and/or to serve as a target for the cells. In contrast,
allogeneic cells or even xenogeneic cells do provide for off-the-shelf
availability. Here the challenge is that of immunogenicity. The problem
of achieving immune acceptance with xenogeneic cells is particularly
severe; however, even for allogeneic cells for at least some cell types,
e.g. vascular endothelial cells, a strategy for creating immune accep-
tance would have to be used.
Finally, there are the questions of differences due to age, due to the
disease state of the patient, or due to the sex of the donor. Although largely
unexplored, these can be significant issues, and it is important that future
research addresses these questions. For example, there is a report that
patient characteristics affects the number of human cardiac progenitor
cells that will be available.
8
There also is evidence that the disease state
can have influence. In this case an example is that in patients with coro-
nary artery disease there was observed a functional impairment of the
hematopoietic progenitor cells.
9
There also are sex differences in the basic characteristics of cells
whether they be stem cells, progenitor cells, or fully differentiated cells.
This is a very important area, one which gives rise to a variety of ques-
tions. For example, if the cells are to be cultured, are different culturing
protocols required for female cells as compared to male cells? For the
clinical therapy itself, will the outcome be different depending on sex of
the donor versus the sex of the patient? In this latter case, there are reports
in the literature documenting differences.
10,11
There thus are a variety of questions to be answered, and although there
is developing a rich literature, further research is required. In the next
section, however, we move on to a very brief discussion of stem cells.
4 R. M. Nerem
3. Stem Cells
As noted earlier, since the early reports in the late 1990s there has been a
surge of activity and this increases at an ever accelerating level. Further
details will be found in the literature
12,13
and in subsequent chapters. Thus
this section of this introductory chapter will only attempt to provide a
broad and very brief overview of the different types of stem cells and
other pluripotent cells available for use in tissue engineering. From an
overall point of view, however, one may consider three general types of
stem cells as follows.
3.1Embryonic stem cells (ESCs)
These can be isolated from the inner cell mass of pre-implantation
embryos during the blastocyst stage.
3,4
They have the ability to differenti-
ate into virtually all specialized cell types and thus are considered
pluripotent. They also have the ability to proliferate in an undifferentiated
state, i.e. they have the ability to self renew. Since different human ESC
lines have been derived from different embryos, it is not surprising that
different lines will exhibit different gene expression characteristics.
14
Within this general category of stem cells there are in addition to those
derived from embryos, those called embryonic germ cells (ECGs). These
are derived from the gonadal ridge of a fetus that is five to ten weeks
old,
5–15
and these are primordial germ cells that in vivogive rise to eggs or
sperm in the adult.
3.2Induced pluripotent stem (iPS)cells
These iPS cells are the result of the transformation of an adult, somatic
cell through reprogramming into a pluriopotent stem cell.
16,17
Although
pluriopotent, these cells are not necessarily identical to ES cells even
though they are similar. The reprogramming initially has been through
retroviral transfection although there are a number of efforts in progress
to carry out the reprogramming without the use of transfection. Like
ESCs, iPS cells can lead to teratoma formation.
Tissue Engineering and Stem Cells 5
As noted earlier, since the early reports in the late 1990s there has been a
surge of activity and this increases at an ever accelerating level. Further
details will be found in the literature
12,13
and in subsequent chapters. Thus
this section of this introductory chapter will only attempt to provide a
broad and very brief overview of the different types of stem cells and
other pluripotent cells available for use in tissue engineering. From an
overall point of view, however, one may consider three general types of
stem cells as follows.
3.1Embryonic stem cells (ESCs)
These can be isolated from the inner cell mass of pre-implantation
embryos during the blastocyst stage.
3,4
They have the ability to differenti-
ate into virtually all specialized cell types and thus are considered
pluripotent. They also have the ability to proliferate in an undifferentiated
state, i.e. they have the ability to self renew. Since different human ESC
lines have been derived from different embryos, it is not surprising that
different lines will exhibit different gene expression characteristics.
14
Within this general category of stem cells there are in addition to those
derived from embryos, those called embryonic germ cells (ECGs). These
are derived from the gonadal ridge of a fetus that is five to ten weeks
old,
5–15
and these are primordial germ cells that in vivogive rise to eggs or
sperm in the adult.
3.2Induced pluripotent stem (iPS)cells
These iPS cells are the result of the transformation of an adult, somatic
cell through reprogramming into a pluriopotent stem cell.
16,17
Although
pluriopotent, these cells are not necessarily identical to ES cells even
though they are similar. The reprogramming initially has been through
retroviral transfection although there are a number of efforts in progress
to carry out the reprogramming without the use of transfection. Like
ESCs, iPS cells can lead to teratoma formation.
Tissue Engineering and Stem Cells 5
3.3Adult stem cells
Adult stem cell populations have been found in many tissues of the human
body. They are believed to be important to the repair mechanism intrinsic
to many tissues and organs. They also in general are tissue specific; how-
ever, there are some exceptions. One of the exceptions to the believed
tissue specificity of adult stem cells is the mesenchymal stem cell.
6,7,18,19
This type of adult stem cell is derived from bone marrow stroma. In fact,
one could view bone marrow transplantation as the earliest cell-based
therapy. In vitrothe MSC can differentiate into a variety of cell types. It
is thus viewed as multipotent, but not pluriopotent or totipotent. Another
exception to the general specificity of adult stem cells are the amniotic-
fluid and placental derived stem cells.
20
They have been shown to have the
capacity for self renewal like ESCs, can give rise to numerous cell types,
and have been characterized as having properties somewhere in between
those of ESCs and adult stem cells. Finally, adult stem cells may be found
in adipose tissue
21
and also in the umbilical cord.
22,23
There clearly is much more that needs to be done to understand the
characteristics of these different stem cell types and the factors involved
in determining the differentiation pathway down which a stem cell can be
directed. Just as the functional characteristics of a fully differentiated cell
is orchestrated by a symphony of signals, the same can be said for the
fate of a stem cell. This symphony includes soluble molecules, cell-cell
contact, and the substrate/extracellular matrix to which the cell is adher-
ent. It also includes what is of particular interest to this author and that is
the role of physical or mechanical forces in modulating stem cell behav-
ior. This includes the role of the physical force environment in the
regulation or modulation of stem cell fate. As an example, we have
shown that mouse ESCs in an early state of differentiation and when
exposed to laminar flow and the associated shear stress exhibit an upreg-
ulation of endothelial cell phenotypic markers.
24
This suggests that such
physical forces, acting as part of a stem cell’s microenvironment, can
participate in the direction of the differentiation process, perhaps even
accelerate it. Taking a different approach, it has been demonstrated that
mechanical stain inhibits the differentiation of human ESCs.
25
Geometry
is a different physical characteristic that can influence stem cell
6 R. M. Nerem
Adult stem cell populations have been found in many tissues of the human
body. They are believed to be important to the repair mechanism intrinsic
to many tissues and organs. They also in general are tissue specific; how-
ever, there are some exceptions. One of the exceptions to the believed
tissue specificity of adult stem cells is the mesenchymal stem cell.
6,7,18,19
This type of adult stem cell is derived from bone marrow stroma. In fact,
one could view bone marrow transplantation as the earliest cell-based
therapy. In vitrothe MSC can differentiate into a variety of cell types. It
is thus viewed as multipotent, but not pluriopotent or totipotent. Another
exception to the general specificity of adult stem cells are the amniotic-
fluid and placental derived stem cells.
20
They have been shown to have the
capacity for self renewal like ESCs, can give rise to numerous cell types,
and have been characterized as having properties somewhere in between
those of ESCs and adult stem cells. Finally, adult stem cells may be found
in adipose tissue
21
and also in the umbilical cord.
22,23
There clearly is much more that needs to be done to understand the
characteristics of these different stem cell types and the factors involved
in determining the differentiation pathway down which a stem cell can be
directed. Just as the functional characteristics of a fully differentiated cell
is orchestrated by a symphony of signals, the same can be said for the
fate of a stem cell. This symphony includes soluble molecules, cell-cell
contact, and the substrate/extracellular matrix to which the cell is adher-
ent. It also includes what is of particular interest to this author and that is
the role of physical or mechanical forces in modulating stem cell behav-
ior. This includes the role of the physical force environment in the
regulation or modulation of stem cell fate. As an example, we have
shown that mouse ESCs in an early state of differentiation and when
exposed to laminar flow and the associated shear stress exhibit an upreg-
ulation of endothelial cell phenotypic markers.
24
This suggests that such
physical forces, acting as part of a stem cell’s microenvironment, can
participate in the direction of the differentiation process, perhaps even
accelerate it. Taking a different approach, it has been demonstrated that
mechanical stain inhibits the differentiation of human ESCs.
25
Geometry
is a different physical characteristic that can influence stem cell
6 R. M. Nerem
differentiation. This has been demonstrated for MSCs.
26
Interestingly,
there also are reports that the mechanical properties of the extracellular
matrix can influence stem cell fate.
27
There also is the role of epigenetics
in the regulation of stem cell fate. An example of this epigenetic role is
the regulation of gene expression through histone modification or DNA
methylation.
28–30
Thus, there are a variety of ways in which the microen-
vironment can orchestrate stem cell fate, and we know very little about
how to design the symphony of signals so as to optimize the outcome of
this orchestration.
4. From Benchtop Science to Cell-Based Applications
If in fact stem cells are to be employed in a particular therapy or treatment,
then one must address the translation of the benchtop research through a
process that will result in the number of cells required for a specific appli-
cation and one that will achieve regulatory approval.
31,32
There are a
number of aspects that must be considered, and in December 2008
Georgia Tech hosted a workshop on Stem Cell Biomanufacturing, bring-
ing together a group of industry and academic researchers as participants.
Some of the issues identified are as follows.
•The sourcing and isolation of the stem cell.
•The monitoring of stem cell phenotype.
•Possible inhomogeneity in the starting population.
•The expansion and propagation of the cells.
•The control of stem cell fate.
•Methods to assess genetic and epigenetic stability.
•Meaningful real time in-process assays.
All of this would need to be done with the quality control required
for regulatory approval. This leads one to the concept of an automated
cell processing facility. To develop such a facility would require
research that leads to a knowledge base for process design. This would
include having the ability to measure process variables in real time and
experiments designed to determine functional relationships between
process variables and what might be called product quality. There would
Tissue Engineering and Stem Cells 7
26
Interestingly,
there also are reports that the mechanical properties of the extracellular
matrix can influence stem cell fate.
27
There also is the role of epigenetics
in the regulation of stem cell fate. An example of this epigenetic role is
the regulation of gene expression through histone modification or DNA
methylation.
28–30
Thus, there are a variety of ways in which the microen-
vironment can orchestrate stem cell fate, and we know very little about
how to design the symphony of signals so as to optimize the outcome of
this orchestration.
4. From Benchtop Science to Cell-Based Applications
If in fact stem cells are to be employed in a particular therapy or treatment,
then one must address the translation of the benchtop research through a
process that will result in the number of cells required for a specific appli-
cation and one that will achieve regulatory approval.
31,32
There are a
number of aspects that must be considered, and in December 2008
Georgia Tech hosted a workshop on Stem Cell Biomanufacturing, bring-
ing together a group of industry and academic researchers as participants.
Some of the issues identified are as follows.
•The sourcing and isolation of the stem cell.
•The monitoring of stem cell phenotype.
•Possible inhomogeneity in the starting population.
•The expansion and propagation of the cells.
•The control of stem cell fate.
•Methods to assess genetic and epigenetic stability.
•Meaningful real time in-process assays.
All of this would need to be done with the quality control required
for regulatory approval. This leads one to the concept of an automated
cell processing facility. To develop such a facility would require
research that leads to a knowledge base for process design. This would
include having the ability to measure process variables in real time and
experiments designed to determine functional relationships between
process variables and what might be called product quality. There would
Tissue Engineering and Stem Cells 7
need to be robust strategies for the interrogation and evaluation of the
variables affecting the processing of cells. There also would need to be
an implementation of closed loop control methods. All of this would
need to be supported by the use of multivariable statistical analysis to
determine the variables affecting product quality. In addition, mathe-
matical models relating the product and the process variables would
need to be developed.
This of course means a different type of research on stem cells than
what today is largely appearing in the literature. If we are to translate all
the exciting advances taking place in our understanding of stem cells to
applications including patient therapies, however, this is what will be
required.
5. Concluding Comments
Tissue engineering, including replacement, repair, and regeneration,
offers the hope that in the future we will be able to develop new therapies
and treatments. This will be particularly important for diseases and
injuries where currently there are no adequate treatments available. A crit-
ical issue is that of cell source, and here the variety of stem cells available
have the potential of providing the answer. For each type of stem cell,
however, there are issues that need to be addressed. Also, beyond the basic
science there also will need to be research aimed at understanding how to
optimize the processing of stem cells. Only with the combination of
research on basic stem cell biology and research on stem cell processing
will it be possible to translate the basic benchtop science into future appli-
cations and patient therapies.
Acknowledgments
The author acknowledges the support provided by the Georgia
Tech/Emory Center for Regenerative Medicine and his part-time visit-
ing professorship at Chonbuk National University in Jeonju, South
Korea. He also thanks his colleagues and students who have taught him
so much.
8 R. M. Nerem
variables affecting the processing of cells. There also would need to be
an implementation of closed loop control methods. All of this would
need to be supported by the use of multivariable statistical analysis to
determine the variables affecting product quality. In addition, mathe-
matical models relating the product and the process variables would
need to be developed.
This of course means a different type of research on stem cells than
what today is largely appearing in the literature. If we are to translate all
the exciting advances taking place in our understanding of stem cells to
applications including patient therapies, however, this is what will be
required.
5. Concluding Comments
Tissue engineering, including replacement, repair, and regeneration,
offers the hope that in the future we will be able to develop new therapies
and treatments. This will be particularly important for diseases and
injuries where currently there are no adequate treatments available. A crit-
ical issue is that of cell source, and here the variety of stem cells available
have the potential of providing the answer. For each type of stem cell,
however, there are issues that need to be addressed. Also, beyond the basic
science there also will need to be research aimed at understanding how to
optimize the processing of stem cells. Only with the combination of
research on basic stem cell biology and research on stem cell processing
will it be possible to translate the basic benchtop science into future appli-
cations and patient therapies.
Acknowledgments
The author acknowledges the support provided by the Georgia
Tech/Emory Center for Regenerative Medicine and his part-time visit-
ing professorship at Chonbuk National University in Jeonju, South
Korea. He also thanks his colleagues and students who have taught him
so much.
8 R. M. Nerem
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R. M. Nerem and R. Lanza Academic Press, (2007); pp. 28–47.
13. A. Vats, R. C. Bielby, N. S. Tolley, R. M. Nerem and J. M. Polak, Stem cells,
Lancet366: 592–602 (2005).
14. R. R. Rao, J. D. Calhoun, X. Qin, R. Rekaya, J. K. Clark and S. L. Stice,
Comparative transcriptional profiling of two human embryonic stem cell
lines,Biotechnol Bioeng88: 273–286 (2004).
15. M. J. Shamblott, J. Axelman, J. W. Littlefield, P. D. Blumenthal,
G. R. Huggins, Y. Cui, L. Cheng and J. D. Gearhart, Human embryonic germ
cell derivates express a broad range of developmentally distinct markers and
proliferate extensively in vitro, Proc Natl Acad Sci USA98: 113–118 (2001).
16. T. Kazutoshi and S. Yamanaka, Induction of pluriopotent stem cells from
mouse embryonic and adult fibroblast cultures by defined factors, Cell
126: 663–676 (2006).
17. S. Yamanaka, A fresh look at iPS cells, Cell137: 13–17 (2009).
18. Y. Jiang, B. N. Jahagirdar, R. L. Reinhardt, R. E. Schartz, C. D. Keene, X. R.
Ortiz-Gonzalez, M. Reyes, T. Lenvik, T. Lund and M. Blackstad,
Pluripotency of mesenchymal stem cells derived from adult marrow, Nature
418: 41–49 (2002).
19. R. G. Edwards, Stem cells today: B1. Bone marrow stem cells, Reprod
Biomed9: 541–593 (2004).
20. P. De Coppi, G. Bartsch, Jr., M. M. Siddiqui, T. Xu, C. C. Santo, L. Erin,
G. Mostoslavsky, A. C. Sierre, E. Y. Synder, J. J. Yoo, M. E. Furth, S. Soker
and A. Atala, Isolation of amniotic stem cell lines with potential for therapy,
Nat Biotechnol 25: 100–106 (2007).
21 P. A. Zuk, M. Zhur, P. Ashjian, D. A. Ugarte, J. I. Huang, H. Mizuno,
Z. C. Alfonso, J. J. Fraser, P. Benhaim and M. H. Hedrick, Human adipose tis-
sue is a source of multipotent stem cells, Mol Biol Cell13: 4279–4295 (2002).
10 R. M. Nerem
22. H. E. Broxmery, G. W. Douglas, G. Hangoc, S. Cooper, J. Bard, D. English,
M. Arny, L. Thomas and E. A. Boyse, Human umbilical cord blood as a
potential source of transplantable hematopoietic stem/progenitor cells, Proc
Natl Acad Sci USA86: 3828–3832 (1989).
23. A. Erices, P. Conget and J. J. Minguell, Mesenchymal progenitor cells in
human umbilical cord blood, Br J Haematol 109: 235–242 (2000).
24. T. Ahsan and R. M. Nerem, Effect of fluid shear stress on embryonic stem-
derived cells, Tissue Eng(in press).
25. S. Sahi, L. Ji, J. J. de Pablo and S. P. Palecek, Inhibition of human embry-
onic stem cell differentiation by mechanical strain, J Cell Physiol 206:
126–137 (2006).
26. K. A. Kilian, B. Bugarija, B. T. Lahn and M. Mrksich, Geometric cues for
directing the differentiation of mesenchymal stem cells, Proc Natl Acad Sci
USA107: 4872–4877 (2010).
27. D. E. Disher, D. J. Mooney and P. W. Zandstra, Growth factors, matrices, and
forces combine and control stem cells, Science324: 1673–1677 (2009).
28. V. T. Lunyak and M. G. Rosenfeld, Epigenetic regulation of stem cell fate,
Hum Mol Genet17: 28–36 (2008).
29. B. Keenen and I. L. De La Serna, Chromatin remodeling in embryonic stem
cells: regulating the balance between pluripotency and differentiation, J Cell
Physiol219: 1–7 (2009).
30. E. Meshorer and T Misteli, Chromatin in pluriopotent embryonic stem cells
and differentiation, Nat Rev Mol Cell Biol7: 540–546 (2006).
31. J. M. Polak and S. Mantalaris, Stem cell bioprocessing: an important mile-
stone to move regenerative medicine research into the clinical arena, Pediatr
Res63: 461–466 (2008).
32. D. C. Kirouac and P .W. Zandstra, The systematic production of cells for cell
therapies,Cell Stem Cell3: 369–381 (2008).
Tissue Engineering and Stem Cells 11
M. Arny, L. Thomas and E. A. Boyse, Human umbilical cord blood as a
potential source of transplantable hematopoietic stem/progenitor cells, Proc
Natl Acad Sci USA86: 3828–3832 (1989).
23. A. Erices, P. Conget and J. J. Minguell, Mesenchymal progenitor cells in
human umbilical cord blood, Br J Haematol 109: 235–242 (2000).
24. T. Ahsan and R. M. Nerem, Effect of fluid shear stress on embryonic stem-
derived cells, Tissue Eng(in press).
25. S. Sahi, L. Ji, J. J. de Pablo and S. P. Palecek, Inhibition of human embry-
onic stem cell differentiation by mechanical strain, J Cell Physiol 206:
126–137 (2006).
26. K. A. Kilian, B. Bugarija, B. T. Lahn and M. Mrksich, Geometric cues for
directing the differentiation of mesenchymal stem cells, Proc Natl Acad Sci
USA107: 4872–4877 (2010).
27. D. E. Disher, D. J. Mooney and P. W. Zandstra, Growth factors, matrices, and
forces combine and control stem cells, Science324: 1673–1677 (2009).
28. V. T. Lunyak and M. G. Rosenfeld, Epigenetic regulation of stem cell fate,
Hum Mol Genet17: 28–36 (2008).
29. B. Keenen and I. L. De La Serna, Chromatin remodeling in embryonic stem
cells: regulating the balance between pluripotency and differentiation, J Cell
Physiol219: 1–7 (2009).
30. E. Meshorer and T Misteli, Chromatin in pluriopotent embryonic stem cells
and differentiation, Nat Rev Mol Cell Biol7: 540–546 (2006).
31. J. M. Polak and S. Mantalaris, Stem cell bioprocessing: an important mile-
stone to move regenerative medicine research into the clinical arena, Pediatr
Res63: 461–466 (2008).
32. D. C. Kirouac and P .W. Zandstra, The systematic production of cells for cell
therapies,Cell Stem Cell3: 369–381 (2008).
Tissue Engineering and Stem Cells 11
2
Recent Advances and Future
Perspectives on Somatic Cell
Reprogramming
Kun-Yong Kim and In-Hyun Park
1. Introduction
Embryonic stem (ES) cells are pluripotent cells derived from the inner
mass of mammalian blastocysts. They have the capacity to self-renew,
while maintaining pluripotency: the ability to generate all cell types of the
three germinal layers. Therefore, a variety of clinical applications have
been proposed for ES cells, as well as in vitrostudies of basic disease
mechanisms, screens for drug discovery, and tissue engineering for degen-
erative diseases. However, ES cells are generic cells that are unrelated to
the patient requiring treatment and the usage of human embryonic stem
cells continues to be a contentious ethical issue.
Embryonic development and cellular differentiation are presumed
to be unidirectional processes and cells undergo a progressive loss of
pluripotency during cell fate specification. In the 1950s, classical exper-
iments demonstrated that differentiated cells retain the genetic
information required to revert to pluripotency
1
and attempts were made
to generate pluripotent stem cells functionally equivalent to ES cells from
13
Recent Advances and Future
Perspectives on Somatic Cell
Reprogramming
Kun-Yong Kim and In-Hyun Park
1. Introduction
Embryonic stem (ES) cells are pluripotent cells derived from the inner
mass of mammalian blastocysts. They have the capacity to self-renew,
while maintaining pluripotency: the ability to generate all cell types of the
three germinal layers. Therefore, a variety of clinical applications have
been proposed for ES cells, as well as in vitrostudies of basic disease
mechanisms, screens for drug discovery, and tissue engineering for degen-
erative diseases. However, ES cells are generic cells that are unrelated to
the patient requiring treatment and the usage of human embryonic stem
cells continues to be a contentious ethical issue.
Embryonic development and cellular differentiation are presumed
to be unidirectional processes and cells undergo a progressive loss of
pluripotency during cell fate specification. In the 1950s, classical exper-
iments demonstrated that differentiated cells retain the genetic
information required to revert to pluripotency
1
and attempts were made
to generate pluripotent stem cells functionally equivalent to ES cells from
13
differentiated cells (reprogramming) — including by somatic cell nuclear
transfer, fusion of differentiated cells with pluripotent cells, application of
pluripotent stem cell extracts to somatic cells, and direct in vitroadapta-
tion of germ cells.
2
Ultimately in 2006, Takahashi and Yamanaka showed
that enforced expression of four transcription factors, Oct4, Sox2, Klf4
and c-Myc reprogrammed murine somatic cells to pluripotency.
3
After this
discovery, reprogramming drew enormous attention not just from stem
cell biologists, but also from clinicians, bioengineers, geneticists, and
developmental biologists. In this chapter, we review recent advances in
the reprogramming field and discuss the potential future applications of
induced pluripotent stem (iPS) cells.
2. Nuclear Reprogramming
Briggs and King showed that nuclei from Rana pipiensblastula cells have
the ability to undergo normal cleavage and develop into complete
embryos when transplanted into enclosed oocytes.
1
This was the first
demonstration that the oocyte cytoplasm contains factors that can repro-
gram the nuclei of differentiated cells back to a pluripotent state that allow
donor cells to normally divide and develop into complete embryos. This
finding was extended by a study by Gurdon and colleagues, who reported
that even fully differentiated intestinal cells from Xenopuscould be repro-
grammed by frog oocytes.
4
Although the efficiency was very low, these
oocytes could give rise to adult animals indicating that the pluripotent
state could be reacquired. Somatic cell nuclear transfer (SCNT) was not
confirmed in other species until 1997, when Dolly the sheep was cloned.
5
The discovery of reprogramming by SCNT led to further advances in the
understanding of the epigenetic processes governing self-renewal and
differentiation in murine and human ES cells.
Several groups demonstrated that fusing a somatic cell with an ES cell
could induce pluripotency.
6,7
This reprogramming can be accomplished
when embryonal carcinoma (EC) cells, embryonic germ (EG) cells, or te-
ratocarcinoma cells are fused with somatic cells. At the tetraploidy stage of
the ensuing cell, the molecular program of the original somatic cell can be
erased and epigenetically reprogrammed to that of an ES cells. With SCNT
methods, reprogramming to a pluripotent state occurs within a few days and
14 K.-Y. Kim and I.-H. Park
transfer, fusion of differentiated cells with pluripotent cells, application of
pluripotent stem cell extracts to somatic cells, and direct in vitroadapta-
tion of germ cells.
2
Ultimately in 2006, Takahashi and Yamanaka showed
that enforced expression of four transcription factors, Oct4, Sox2, Klf4
and c-Myc reprogrammed murine somatic cells to pluripotency.
3
After this
discovery, reprogramming drew enormous attention not just from stem
cell biologists, but also from clinicians, bioengineers, geneticists, and
developmental biologists. In this chapter, we review recent advances in
the reprogramming field and discuss the potential future applications of
induced pluripotent stem (iPS) cells.
2. Nuclear Reprogramming
Briggs and King showed that nuclei from Rana pipiensblastula cells have
the ability to undergo normal cleavage and develop into complete
embryos when transplanted into enclosed oocytes.
1
This was the first
demonstration that the oocyte cytoplasm contains factors that can repro-
gram the nuclei of differentiated cells back to a pluripotent state that allow
donor cells to normally divide and develop into complete embryos. This
finding was extended by a study by Gurdon and colleagues, who reported
that even fully differentiated intestinal cells from Xenopuscould be repro-
grammed by frog oocytes.
4
Although the efficiency was very low, these
oocytes could give rise to adult animals indicating that the pluripotent
state could be reacquired. Somatic cell nuclear transfer (SCNT) was not
confirmed in other species until 1997, when Dolly the sheep was cloned.
5
The discovery of reprogramming by SCNT led to further advances in the
understanding of the epigenetic processes governing self-renewal and
differentiation in murine and human ES cells.
Several groups demonstrated that fusing a somatic cell with an ES cell
could induce pluripotency.
6,7
This reprogramming can be accomplished
when embryonal carcinoma (EC) cells, embryonic germ (EG) cells, or te-
ratocarcinoma cells are fused with somatic cells. At the tetraploidy stage of
the ensuing cell, the molecular program of the original somatic cell can be
erased and epigenetically reprogrammed to that of an ES cells. With SCNT
methods, reprogramming to a pluripotent state occurs within a few days and
14 K.-Y. Kim and I.-H. Park
can be greatly enhanced by co-expression of pluripotency-associated genes
such as Oct4, Sox2, c-Myc, Klf4, Nanog, and Sall4. Yet, each of these meth-
ods has clinical limitations. SCNT has the advantage of using intrinsic
reprogramming machinery in oocytes, but it requires the donation of human
oocytes that can be in limited supply and raise ethical concerns. In somatic
cell fusion, there are no ethical problems and no limitations on the supply
of cells, but the processes result in the formation of the undesired tetraploid
that is genetically unstable, and the overexpression of transcription factors
increases the frequency of tumorigenesis in the resulting progeny.
In recent years, nuclear and cytoplasmic extracts from undifferentiated
cells have been tried to induce the pluripotency of differentiated somatic
cells. Extracts from pluripotent EG, EC cells or ES cells that contain the reg-
ulatory components needed for reprogramming can induce the pluripotency
of somatic cells.
8
The reprogramming extracts reversibly permeabilize the
somatic cell and then induce transcriptional reprogramming, leading to epi-
genetic modification of the somatic cells. Even interspecies reprogramming
via cell extracts showed some success in de-differentiating somatic cells.
The extracts of regenerated new limbs induced partial dedifferentiation of
c2c12-derived mouse myotubes into myoblasts and the extracts of Xenopus
eggs induced the expression of the pluripotency genes Oct4 and Sox2 in
mammalian somatic cells.
9,10
When fibroblast cells were reversibly perme-
abilized and transiently exposed to extracts from mouse EC cells, they lead
to Oct4 biphasic activation. Incubation of extracts from mouse ES cells with
reversibly permeabilized NIH3T3 cells induced partial reprogramming.
This approach to inducing pluripotency using pluripotent cell extracts could
be an excellent alternative to nuclear transfer reprogramming due to the fact
that eggs are not required and the resultant cells are diploid. However, the
intrinsic difficulty to continue the exposure of the somatic cells to pluripo-
tent cell extract holds yet the successful derivation of completely
reprogrammed pluripotent stem cells.
Hence, a variety of studies have been proposed to better understand the
reprogramming of pluripotent stem cells. However, the principal limita-
tion to all the above technologies for cell reprogramming is that the
generated pluripotent stem cells are unrelated to the patient who requires
clinical treatment, and the use of human ES cells remains a contentious
ethical issue.
Reprogramming and iPS Cells 15
such as Oct4, Sox2, c-Myc, Klf4, Nanog, and Sall4. Yet, each of these meth-
ods has clinical limitations. SCNT has the advantage of using intrinsic
reprogramming machinery in oocytes, but it requires the donation of human
oocytes that can be in limited supply and raise ethical concerns. In somatic
cell fusion, there are no ethical problems and no limitations on the supply
of cells, but the processes result in the formation of the undesired tetraploid
that is genetically unstable, and the overexpression of transcription factors
increases the frequency of tumorigenesis in the resulting progeny.
In recent years, nuclear and cytoplasmic extracts from undifferentiated
cells have been tried to induce the pluripotency of differentiated somatic
cells. Extracts from pluripotent EG, EC cells or ES cells that contain the reg-
ulatory components needed for reprogramming can induce the pluripotency
of somatic cells.
8
The reprogramming extracts reversibly permeabilize the
somatic cell and then induce transcriptional reprogramming, leading to epi-
genetic modification of the somatic cells. Even interspecies reprogramming
via cell extracts showed some success in de-differentiating somatic cells.
The extracts of regenerated new limbs induced partial dedifferentiation of
c2c12-derived mouse myotubes into myoblasts and the extracts of Xenopus
eggs induced the expression of the pluripotency genes Oct4 and Sox2 in
mammalian somatic cells.
9,10
When fibroblast cells were reversibly perme-
abilized and transiently exposed to extracts from mouse EC cells, they lead
to Oct4 biphasic activation. Incubation of extracts from mouse ES cells with
reversibly permeabilized NIH3T3 cells induced partial reprogramming.
This approach to inducing pluripotency using pluripotent cell extracts could
be an excellent alternative to nuclear transfer reprogramming due to the fact
that eggs are not required and the resultant cells are diploid. However, the
intrinsic difficulty to continue the exposure of the somatic cells to pluripo-
tent cell extract holds yet the successful derivation of completely
reprogrammed pluripotent stem cells.
Hence, a variety of studies have been proposed to better understand the
reprogramming of pluripotent stem cells. However, the principal limita-
tion to all the above technologies for cell reprogramming is that the
generated pluripotent stem cells are unrelated to the patient who requires
clinical treatment, and the use of human ES cells remains a contentious
ethical issue.
Reprogramming and iPS Cells 15
3. Reprogramming by Defined Factors
Reprogramming somatic cells by using a defined set of factors is a novel
concept and opens extraordinary possibilities for producing pluripotent
stem cells without raising the ethical concerns associated with destroying
human embryos.
5,6
while allowing for patients to be treated with their
own reprogrammed somatic cells. In their original report, Takahashi and
Yamanaka retrovirally transduced different combinations of 24 candidate
reprogramming genes into mouse embryonic fibroblasts derived from
transgenic mice containing a neomycin resistance gene knocked into the
Fbx15 locus.
3
Cells that recovered neomycin resistance from the reactivation
of the reporter gene were selected. They found that the retrovirus-mediated
introduction of transcription factors was sufficient to reprogram mouse
fibroblasts back to a pluripotent state. Specifically, the combined intro-
duction of the four transcription factors, Oct4, Sox2, Klf4, and c-Myc
were identified as sufficient to give rise to pluripotent cells, termed
induced pluripotent stem (iPS) cells.
Selected iPS cells showed properties very similar to murine ES cells in
morphology, proliferative characteristics, and expression of pluripotency
markers. Moreover, iPS cells differentiated into all three germ layers
in vitro, and were able to form teratomas when injected into nude mice,
confirming their pluripotency. Although Fbx15-selected iPS cells formed
chimeras, they did not result in germ line transmission, raising the concern
that iPS cells are not truly equivalent to ES cells. However, several studies
have reported that iPS cells selected using Nanog or Oct4 are more
epigenetically related to ES cells and can produce chimeras capable of
germ line transmission.
11,12
However, these iPS cells did not pass the most
stringent pluripotency test: tetraploid complementation. Albeit negligible,
the continuous expression of reprogramming genes seems to be responsi-
ble for the limited differentiation potential of the iPS cells, since the iPS
cells produced by inducible lentiviral reprogramming factors succeeded to
generate full-term mouse after tetracomplementation.
13,14
After the identification of murine iPS cells was reported, using the
same four transcription factors, Oct4, Sox2, Klf4 and c-Myc or combined
with novel factors, Nanog and Lin28, three different groups isolated
human iPS cells.
15–17
Human iPS cells from either combination of factors
16 K.-Y. Kim and I.-H. Park
Reprogramming somatic cells by using a defined set of factors is a novel
concept and opens extraordinary possibilities for producing pluripotent
stem cells without raising the ethical concerns associated with destroying
human embryos.
5,6
while allowing for patients to be treated with their
own reprogrammed somatic cells. In their original report, Takahashi and
Yamanaka retrovirally transduced different combinations of 24 candidate
reprogramming genes into mouse embryonic fibroblasts derived from
transgenic mice containing a neomycin resistance gene knocked into the
Fbx15 locus.
3
Cells that recovered neomycin resistance from the reactivation
of the reporter gene were selected. They found that the retrovirus-mediated
introduction of transcription factors was sufficient to reprogram mouse
fibroblasts back to a pluripotent state. Specifically, the combined intro-
duction of the four transcription factors, Oct4, Sox2, Klf4, and c-Myc
were identified as sufficient to give rise to pluripotent cells, termed
induced pluripotent stem (iPS) cells.
Selected iPS cells showed properties very similar to murine ES cells in
morphology, proliferative characteristics, and expression of pluripotency
markers. Moreover, iPS cells differentiated into all three germ layers
in vitro, and were able to form teratomas when injected into nude mice,
confirming their pluripotency. Although Fbx15-selected iPS cells formed
chimeras, they did not result in germ line transmission, raising the concern
that iPS cells are not truly equivalent to ES cells. However, several studies
have reported that iPS cells selected using Nanog or Oct4 are more
epigenetically related to ES cells and can produce chimeras capable of
germ line transmission.
11,12
However, these iPS cells did not pass the most
stringent pluripotency test: tetraploid complementation. Albeit negligible,
the continuous expression of reprogramming genes seems to be responsi-
ble for the limited differentiation potential of the iPS cells, since the iPS
cells produced by inducible lentiviral reprogramming factors succeeded to
generate full-term mouse after tetracomplementation.
13,14
After the identification of murine iPS cells was reported, using the
same four transcription factors, Oct4, Sox2, Klf4 and c-Myc or combined
with novel factors, Nanog and Lin28, three different groups isolated
human iPS cells.
15–17
Human iPS cells from either combination of factors
16 K.-Y. Kim and I.-H. Park
shared defining characteristics with human ES cells, including gene
expression profiles, morphology, proliferation, patterns of DNA methyla-
tion and histone modification, as well as telomerase activities. Human iPS
cells were also able to differentiate into three germ layers in vitroas
embryonic bodies and to form teratomas after injection into immune
deficient mice.
4. Recent Advances in Reprogramming Methods
When the first iPS cells were isolated, many questions regarding the tech-
niques to generate iPS cells were raised, such as how to identify iPS cells,
how to derive iPS cells without potentially tumorigenic oncogenes, and
how to generate iPS cells without using retro/lentivirus. Over the last
three years, there have been many advances that have begun to overcome
these hurdles.
Initially, Takahashi and Yamanaka created the first iPS cells using the
neomycin resistant gene regulated by the pluripotent cell specific Fbx15
locus and claimed that reprogramming was a very inefficient process.
3
Selection tools seemed essential to identify the iPS cells, and Nanog or
Oct4 promoter-based drug resistant genes were used to help isolate
murine iPS cells.
11,12,18
Lately, murine and human iPS cells were derived
by using the morphologies unique to ES cells.
15–17
Furthermore, silencing
of the retro- and lentivirus could be retrospectively used to identify the
reprogrammed cells from the partially reprogrammed or transformed
cells.
19
The induction of the gene that mediates retroviral silencing in ES
cells, Trim28, seems to be responsible for retroviral silencing during
reprogramming in iPS cells.
20
The usage of oncogenenic c-Myc and Klf4 in generating iPS cells is a
major hurdle for potential clinical applications. One of the four repro-
gramming factors, c-Myc, is a well-established oncogene, andits continued
expression and/or potential reactivation in iPS cell derivatives is not suit-
able for future iPS cell therapies. Indeed, head and neck tumor formation
has been observed in iPS cell-derived chimeric mice possibly due to the
reactivation of c-Myc.
11
However, c-Myc is dispensable for reprogram-
ming murine and human somatic cells, although its absence significantly
reduces the reprogramming efficiency.
21
Some somatic cells highly express
Reprogramming and iPS Cells 17
expression profiles, morphology, proliferation, patterns of DNA methyla-
tion and histone modification, as well as telomerase activities. Human iPS
cells were also able to differentiate into three germ layers in vitroas
embryonic bodies and to form teratomas after injection into immune
deficient mice.
4. Recent Advances in Reprogramming Methods
When the first iPS cells were isolated, many questions regarding the tech-
niques to generate iPS cells were raised, such as how to identify iPS cells,
how to derive iPS cells without potentially tumorigenic oncogenes, and
how to generate iPS cells without using retro/lentivirus. Over the last
three years, there have been many advances that have begun to overcome
these hurdles.
Initially, Takahashi and Yamanaka created the first iPS cells using the
neomycin resistant gene regulated by the pluripotent cell specific Fbx15
locus and claimed that reprogramming was a very inefficient process.
3
Selection tools seemed essential to identify the iPS cells, and Nanog or
Oct4 promoter-based drug resistant genes were used to help isolate
murine iPS cells.
11,12,18
Lately, murine and human iPS cells were derived
by using the morphologies unique to ES cells.
15–17
Furthermore, silencing
of the retro- and lentivirus could be retrospectively used to identify the
reprogrammed cells from the partially reprogrammed or transformed
cells.
19
The induction of the gene that mediates retroviral silencing in ES
cells, Trim28, seems to be responsible for retroviral silencing during
reprogramming in iPS cells.
20
The usage of oncogenenic c-Myc and Klf4 in generating iPS cells is a
major hurdle for potential clinical applications. One of the four repro-
gramming factors, c-Myc, is a well-established oncogene, andits continued
expression and/or potential reactivation in iPS cell derivatives is not suit-
able for future iPS cell therapies. Indeed, head and neck tumor formation
has been observed in iPS cell-derived chimeric mice possibly due to the
reactivation of c-Myc.
11
However, c-Myc is dispensable for reprogram-
ming murine and human somatic cells, although its absence significantly
reduces the reprogramming efficiency.
21
Some somatic cells highly express
Reprogramming and iPS Cells 17
endogenous reprogramming genes and may not need the expression of all
reprogramming factors. Indeed, neural progenitor cells (NPCs) that highly
express endogenous Sox2, and can be reprogrammed without the ectopic
expression of Sox2.
22
Furthermore, even introduction of Oct4 alone was
shown to be sufficient to generate iPS cells from NPGs.
23,24
Similarly,
meningiocytes and keratinocytes appear to be particularly prone to repro-
gramming due to their relatively high endogenous expression of Sox2,
Myc and Klf4.
25,26
These results indicate that the reprogramming cells like
NPCs would require a minimal genetic modification in reprogramming
and result in most desirable safe iPS cells.
Limitation to the current methods in generating patient specific iPS
cells is the residual presence of the reprogramming factors in chromo-
some. Retro/lentiviral integration into the genome carries a risk of tumor
formation when random integration activates pathways for cell prolifera-
tion or inhibits the tumor suppressor pathways. Using methods based on
non-integrating vectors or direct exposure to reprogramming proteins is
desirable. Stadtfeld and colleagues were able to generate iPS cells from
adult mouse fibroblast and liver cells using non-integrating adenoviral
vectors.
27
They demonstrated that adenoviral-mediated transient expres-
sion of the exogenous reprogramming factors eliminated the risk of
insertional mutagenesis. Okita and colleagues also successfully generated
iPS cells without using any viral vectors but multiple transient transfec-
tion of the reprogramming factors.
28
Lately, oriP/EBNA1-based episomal
vector was successfully used to generate human iPS cells.
29
All the non-
integrating vector methods provide a safer iPS cells but suffer from the
extremely low reprogramming efficiency.
Usage of three or four transcription factors for reprogramming results
in the high incidence of multiple integration of reprogramming factors into
chromosome. Combining the reprogramming factors with picornaviral 2A
sequences allowed the expression of multiple genes in one backbone.
30,31
This method generated iPS cells through less number of retro- or lentiviral
insertion into the genome, as opposed to using separate viral vectors for
each reprogramming factor. Because the single viral copy may also
be removed from the iPS cell genome after reprogramming (e.g. by
loxP/Cre technology) the authors successfully generated safer iPS cells.
32
Similarly, Kaji and colleagues developed a piggyBac transposon-based
18 K.-Y. Kim and I.-H. Park
reprogramming factors. Indeed, neural progenitor cells (NPCs) that highly
express endogenous Sox2, and can be reprogrammed without the ectopic
expression of Sox2.
22
Furthermore, even introduction of Oct4 alone was
shown to be sufficient to generate iPS cells from NPGs.
23,24
Similarly,
meningiocytes and keratinocytes appear to be particularly prone to repro-
gramming due to their relatively high endogenous expression of Sox2,
Myc and Klf4.
25,26
These results indicate that the reprogramming cells like
NPCs would require a minimal genetic modification in reprogramming
and result in most desirable safe iPS cells.
Limitation to the current methods in generating patient specific iPS
cells is the residual presence of the reprogramming factors in chromo-
some. Retro/lentiviral integration into the genome carries a risk of tumor
formation when random integration activates pathways for cell prolifera-
tion or inhibits the tumor suppressor pathways. Using methods based on
non-integrating vectors or direct exposure to reprogramming proteins is
desirable. Stadtfeld and colleagues were able to generate iPS cells from
adult mouse fibroblast and liver cells using non-integrating adenoviral
vectors.
27
They demonstrated that adenoviral-mediated transient expres-
sion of the exogenous reprogramming factors eliminated the risk of
insertional mutagenesis. Okita and colleagues also successfully generated
iPS cells without using any viral vectors but multiple transient transfec-
tion of the reprogramming factors.
28
Lately, oriP/EBNA1-based episomal
vector was successfully used to generate human iPS cells.
29
All the non-
integrating vector methods provide a safer iPS cells but suffer from the
extremely low reprogramming efficiency.
Usage of three or four transcription factors for reprogramming results
in the high incidence of multiple integration of reprogramming factors into
chromosome. Combining the reprogramming factors with picornaviral 2A
sequences allowed the expression of multiple genes in one backbone.
30,31
This method generated iPS cells through less number of retro- or lentiviral
insertion into the genome, as opposed to using separate viral vectors for
each reprogramming factor. Because the single viral copy may also
be removed from the iPS cell genome after reprogramming (e.g. by
loxP/Cre technology) the authors successfully generated safer iPS cells.
32
Similarly, Kaji and colleagues developed a piggyBac transposon-based
18 K.-Y. Kim and I.-H. Park
vector expressing four genes enabling the creation of transgene-free iPS
cells following removal of the transposon.
33
Despite these advancements,
the concern lingers that all of these factors have links to tumorigenesis.
An alternative approach that allows the complete avoidance of the use
of oncogenes altogether is to use small molecules instead. Shi et al. gener-
ated iPS cells from NPCs by transduction of Oct4 and Klf4, and found that
simultaneous treatment with G9a inhibitor, BIX-01294, remarkably
increased the reprogramming efficiency.
34
Subsequent studies have
confirmed that epigenetic modification or the activation of self-renewing
signaling through small molecules can improve reprogramming.
Additionally, the histone deacetylase (HDAC) inhibitor valproic acid (VPA)
and Trichostatin A (TSA), as well as the DNA methyltransferase inhibitor,
5-aza-cytidine have been shown to increase reprogramming efficiency.
35,36
The Wnt signaling component WNT3a, or the L-channel calcium channel
agonist Bayk8644 (BayK) also can increase the reprogramming
efficiency.
22
In another study, the inhibition of mitogen-activated protein
(MAP) kinase signaling and synthase kinase-3 (GSK3), allowed repro-
gramming to occur, even in the absence of Sox2 and c-Myc.
37
Recently,
tumor suppressor pathways, including p53, Ink4a/Arf and p21, were shown
to play a role as a barrier to reprogramming.
38–41
These several signaling
pathways are well known to facilitate the ES cell self-renewal or cell
proliferation and seem to increase the reprogramming efficiency.
Ultimately, iPS cells have been generated without using vectors at all,
but rather by directly introducing proteins into fibroblasts. Zhou and
colleagues purified recombinant reprogramming factors fused with the
poly-arginine (i.e.11R) protein transduction domain of the C-termini and
were able to generate iPS cells from mouse embryonic fibroblasts.
42
Kim
and colleagues established 293T cell lines stably expressing reprogramming
factors fused with the poly-arginine tag and demonstrated the formation of
human iPS cells by exposure of the cell extract to fibroblasts.
43
5. Future Perspectives on Reprogramming and iPS Cells
Dr. Yamanaka’s reports of generating iPS cells using defined factors
revolutionized the approach to manipulating the cellular identity. Initiated
by the finding that fibroblasts became induced to the myogenic lineage by
Reprogramming and iPS Cells 19
cells following removal of the transposon.
33
Despite these advancements,
the concern lingers that all of these factors have links to tumorigenesis.
An alternative approach that allows the complete avoidance of the use
of oncogenes altogether is to use small molecules instead. Shi et al. gener-
ated iPS cells from NPCs by transduction of Oct4 and Klf4, and found that
simultaneous treatment with G9a inhibitor, BIX-01294, remarkably
increased the reprogramming efficiency.
34
Subsequent studies have
confirmed that epigenetic modification or the activation of self-renewing
signaling through small molecules can improve reprogramming.
Additionally, the histone deacetylase (HDAC) inhibitor valproic acid (VPA)
and Trichostatin A (TSA), as well as the DNA methyltransferase inhibitor,
5-aza-cytidine have been shown to increase reprogramming efficiency.
35,36
The Wnt signaling component WNT3a, or the L-channel calcium channel
agonist Bayk8644 (BayK) also can increase the reprogramming
efficiency.
22
In another study, the inhibition of mitogen-activated protein
(MAP) kinase signaling and synthase kinase-3 (GSK3), allowed repro-
gramming to occur, even in the absence of Sox2 and c-Myc.
37
Recently,
tumor suppressor pathways, including p53, Ink4a/Arf and p21, were shown
to play a role as a barrier to reprogramming.
38–41
These several signaling
pathways are well known to facilitate the ES cell self-renewal or cell
proliferation and seem to increase the reprogramming efficiency.
Ultimately, iPS cells have been generated without using vectors at all,
but rather by directly introducing proteins into fibroblasts. Zhou and
colleagues purified recombinant reprogramming factors fused with the
poly-arginine (i.e.11R) protein transduction domain of the C-termini and
were able to generate iPS cells from mouse embryonic fibroblasts.
42
Kim
and colleagues established 293T cell lines stably expressing reprogramming
factors fused with the poly-arginine tag and demonstrated the formation of
human iPS cells by exposure of the cell extract to fibroblasts.
43
5. Future Perspectives on Reprogramming and iPS Cells
Dr. Yamanaka’s reports of generating iPS cells using defined factors
revolutionized the approach to manipulating the cellular identity. Initiated
by the finding that fibroblasts became induced to the myogenic lineage by
Reprogramming and iPS Cells 19
expressing a given myogenic factor,
44
dedifferentiation or reprogramming
to different cellular fates have been explored, including hematopoietic,
pancreatic and cardiac lineages.
45–47
Pluripotent iPS cells generated via
reprogramming will have a broader applicability, since they can differen-
tiate into any cellular lineage. They can be used to investigate the disease
progressionin vitro, and to provide a platform to screening chemicals for
genetic disorders (Fig. 1). Recent advancement of high throughput
sequencing, when combined with iPS cells, will allow us to investigate the
early development of genetically defined, personalized cellsin vitro.
48
After human ES cells were isolated, they have been used to make
models of human diseases.
49,50
Human ES cells genetically modified by
either overexpression or knock-down of genes of interest was proposed to
mimic the early embryonic cells of human diseases in vitro. In vitrodiffer-
entiation of ES cells were presumed to follow the early developmental
program of normal embryonic development.
51
iPS cells derived directly
20 K.-Y. Kim and I.-H. Park
Fig. 1.Recent advances and future perspectives of reprogramming. Since the Yamanaka
lab reported the generation of iPS cells in murine fibroblasts, there has been an explosion
of research on factor-based reprogramming. 1) iPS cells were generated without using
integrating retro- and lentiviral vectors. Small molecules were identified that increased
reprogramming efficiency. 2) Reprogrammed iPS cells will be used to generate in vitro
disease model. 3) In vivotransdifferentiation will be a practical alternative to pluripotent
stem cells for genetic or non-genetic degenerative diseases. Shown in red are challenges
that will lead to successful utility of the iPS cells in regenerative medicine.
44
dedifferentiation or reprogramming
to different cellular fates have been explored, including hematopoietic,
pancreatic and cardiac lineages.
45–47
Pluripotent iPS cells generated via
reprogramming will have a broader applicability, since they can differen-
tiate into any cellular lineage. They can be used to investigate the disease
progressionin vitro, and to provide a platform to screening chemicals for
genetic disorders (Fig. 1). Recent advancement of high throughput
sequencing, when combined with iPS cells, will allow us to investigate the
early development of genetically defined, personalized cellsin vitro.
48
After human ES cells were isolated, they have been used to make
models of human diseases.
49,50
Human ES cells genetically modified by
either overexpression or knock-down of genes of interest was proposed to
mimic the early embryonic cells of human diseases in vitro. In vitrodiffer-
entiation of ES cells were presumed to follow the early developmental
program of normal embryonic development.
51
iPS cells derived directly
20 K.-Y. Kim and I.-H. Park
Fig. 1.Recent advances and future perspectives of reprogramming. Since the Yamanaka
lab reported the generation of iPS cells in murine fibroblasts, there has been an explosion
of research on factor-based reprogramming. 1) iPS cells were generated without using
integrating retro- and lentiviral vectors. Small molecules were identified that increased
reprogramming efficiency. 2) Reprogrammed iPS cells will be used to generate in vitro
disease model. 3) In vivotransdifferentiation will be a practical alternative to pluripotent
stem cells for genetic or non-genetic degenerative diseases. Shown in red are challenges
that will lead to successful utility of the iPS cells in regenerative medicine.
from patients provide an in vitromodel that will more closely mimic the
pathology of the disease. Following are examples of the rigorous attempts
to generate human disease models using murine and human iPS cells.
Murine iPS cells have been used to illustrate the therapeutic potential
of iPS cells in vivo . For example, Hanna and colleagues showed that trans-
genic mice engineered to have human sickle cell anemia could be
successfully treated with hematopoietic progenitor cells produced from
autologous iPS cells and genetically repaired.
52
Wernig et al. showed that
iPS cell-derived neuronal cells could integrate into fetal brains and ame-
liorate the symptoms of rats with Parkinson’s disease.
53
Since then, more
studies have reported the derivation of therapeutically relevant cell types
from iPS cells, including human insulin-secreting cells, and functional
mouse and human cardiomyocytes, as well as mouse endothelial cells that
successfully treated mice with coagulation disorder hemophilia A.
54–56
Human iPS cells from patients’ fibroblasts were generated to investi-
gate human diseases in vitro. Our group has generated iPS cells derived
from a variety of genetic disorders, including adenosine deaminase
deficiency-related severe combined immunodeficiency (ADA-SCID),
Shwachman-Bodian-Diamond syndrome (SBDS), Gaucher disease (GD)
type III, Duchenne (DMD) and Becker muscular dystrophy (BMD),
Parkinson disease (PD), Huntington disease (HD), juvenile-onset, type 1
diabetes mellitus (JDM), Down syndrome (DS)/ trisomy 21 and the car-
rier state of Lesch-Nyhan syndrome.
57
iPS cells generated from patients
of neuromuscular diseases, including amyotrophic lateral sclerosis
(ALS) and spinal muscular atrophy (SMA), were shown to differentiate
into motor neurons, providing a novel in vitromotoneuron disease
model.
58,59
A range of iPS cells from Parkinson’s patients was also gen-
erated as an invaluable resource for studying the disease.
60
These cells do
not contain transgenic reprogramming factors in their genomes as the
factors have been excised by Cre recombinase from the integrated
lentiviral vector. Recently, Lee and colleagues extended the iPS cell
technology to model autosomal recessive familial dysautonomia (FD)
in vitro, and demonstrated the FD-related mis-splicing of IKBKAP and
concurrent defects in neurogenic differentiation and migration behavior.
61
Most of iPS cells used to model human diseases still contained the retro-
viral vectors in their chromosomes. With the advent of reprogramming
Reprogramming and iPS Cells 21
pathology of the disease. Following are examples of the rigorous attempts
to generate human disease models using murine and human iPS cells.
Murine iPS cells have been used to illustrate the therapeutic potential
of iPS cells in vivo . For example, Hanna and colleagues showed that trans-
genic mice engineered to have human sickle cell anemia could be
successfully treated with hematopoietic progenitor cells produced from
autologous iPS cells and genetically repaired.
52
Wernig et al. showed that
iPS cell-derived neuronal cells could integrate into fetal brains and ame-
liorate the symptoms of rats with Parkinson’s disease.
53
Since then, more
studies have reported the derivation of therapeutically relevant cell types
from iPS cells, including human insulin-secreting cells, and functional
mouse and human cardiomyocytes, as well as mouse endothelial cells that
successfully treated mice with coagulation disorder hemophilia A.
54–56
Human iPS cells from patients’ fibroblasts were generated to investi-
gate human diseases in vitro. Our group has generated iPS cells derived
from a variety of genetic disorders, including adenosine deaminase
deficiency-related severe combined immunodeficiency (ADA-SCID),
Shwachman-Bodian-Diamond syndrome (SBDS), Gaucher disease (GD)
type III, Duchenne (DMD) and Becker muscular dystrophy (BMD),
Parkinson disease (PD), Huntington disease (HD), juvenile-onset, type 1
diabetes mellitus (JDM), Down syndrome (DS)/ trisomy 21 and the car-
rier state of Lesch-Nyhan syndrome.
57
iPS cells generated from patients
of neuromuscular diseases, including amyotrophic lateral sclerosis
(ALS) and spinal muscular atrophy (SMA), were shown to differentiate
into motor neurons, providing a novel in vitromotoneuron disease
model.
58,59
A range of iPS cells from Parkinson’s patients was also gen-
erated as an invaluable resource for studying the disease.
60
These cells do
not contain transgenic reprogramming factors in their genomes as the
factors have been excised by Cre recombinase from the integrated
lentiviral vector. Recently, Lee and colleagues extended the iPS cell
technology to model autosomal recessive familial dysautonomia (FD)
in vitro, and demonstrated the FD-related mis-splicing of IKBKAP and
concurrent defects in neurogenic differentiation and migration behavior.
61
Most of iPS cells used to model human diseases still contained the retro-
viral vectors in their chromosomes. With the advent of reprogramming
Reprogramming and iPS Cells 21
methods not relying on integrating virus, iPS cells are expected to be
more close to ES cells and make better disease models.
43
Although it is highly hoped to utilize iPS cells in various applications
mentioned above, there are several issues to be resolved (Fig. 1). First of
all, methods to generate clinically useful iPS cells should be improved.
Non-integrating virus, direct protein transduction, and nuclear transfer
will make genetically non-modified iPS cells. But, their reprogramming
efficiency is extremely low, and will not be practical to produce iPS cells
in a reliable manner. Improvement in viral vectors, or screening small
molecules will be essential to optimize the reprogramming methods.
62,63
In order to reach the final goal of regenerative medicine using pluripo-
tent stem cells, it is crucial to make them acquire the desired lineage
in vitro. Directed differentiation of pluripotent stem cells relied on the pre-
vious knowledge of cell specification obtained from developmental
biology.
51
Treatment of cytokines, growth factors, chemicals, and small
molecules have been attempted to differentiate pluripotent stem cells.
Ectopic expression of lineage specific transcription factors is a reliable
approach to direct the differentiating cells into specific lineage.
64,65
Cell
surface markers were used to isolate the cells of interests. When mouse ES
cell lines expressing fluorescent proteins of lineage specific markers are
available, they provide an effective way to isolate pure cell population.
The isolation of lineage specific cells from human ES cells was facilitated
by the transgenic human ES cell lines with lineage markers. Teratoma
formed by the incompletely differentiated pluripotent stem cells continu-
ously impedes the utility of cells differentiated from ES cells.
Improvement of negative selection of partially or non-differentiated cells,
or rigorous isolation of completely differentiated cells are required.
In vivotransdifferentiation will be an alternative to iPS cells of great prac-
tical importance. Most of the degenerative disorders show loss in
parenchymal or functional cells in tissues: loss of endocrine beta cells in
type I diabetes, dopaminergic neurons in Parkinson’s diseases and
motoneuron in ALS. Despite the degeneration of these cell types, the stro-
mal or adjacent cells surrounding cells are still intact. When expressed
with a combination of genes important for the development of the same
lineages of the degenerated cells, the surviving cells in the damaged tis-
sue will change their cellular fate to the cells of the interest and will help
22 K.-Y. Kim and I.-H. Park
more close to ES cells and make better disease models.
43
Although it is highly hoped to utilize iPS cells in various applications
mentioned above, there are several issues to be resolved (Fig. 1). First of
all, methods to generate clinically useful iPS cells should be improved.
Non-integrating virus, direct protein transduction, and nuclear transfer
will make genetically non-modified iPS cells. But, their reprogramming
efficiency is extremely low, and will not be practical to produce iPS cells
in a reliable manner. Improvement in viral vectors, or screening small
molecules will be essential to optimize the reprogramming methods.
62,63
In order to reach the final goal of regenerative medicine using pluripo-
tent stem cells, it is crucial to make them acquire the desired lineage
in vitro. Directed differentiation of pluripotent stem cells relied on the pre-
vious knowledge of cell specification obtained from developmental
biology.
51
Treatment of cytokines, growth factors, chemicals, and small
molecules have been attempted to differentiate pluripotent stem cells.
Ectopic expression of lineage specific transcription factors is a reliable
approach to direct the differentiating cells into specific lineage.
64,65
Cell
surface markers were used to isolate the cells of interests. When mouse ES
cell lines expressing fluorescent proteins of lineage specific markers are
available, they provide an effective way to isolate pure cell population.
The isolation of lineage specific cells from human ES cells was facilitated
by the transgenic human ES cell lines with lineage markers. Teratoma
formed by the incompletely differentiated pluripotent stem cells continu-
ously impedes the utility of cells differentiated from ES cells.
Improvement of negative selection of partially or non-differentiated cells,
or rigorous isolation of completely differentiated cells are required.
In vivotransdifferentiation will be an alternative to iPS cells of great prac-
tical importance. Most of the degenerative disorders show loss in
parenchymal or functional cells in tissues: loss of endocrine beta cells in
type I diabetes, dopaminergic neurons in Parkinson’s diseases and
motoneuron in ALS. Despite the degeneration of these cell types, the stro-
mal or adjacent cells surrounding cells are still intact. When expressed
with a combination of genes important for the development of the same
lineages of the degenerated cells, the surviving cells in the damaged tis-
sue will change their cellular fate to the cells of the interest and will help
22 K.-Y. Kim and I.-H. Park
to recover the function. Given the great opportunity of manipulating cell
fates, it becomes possible to conquer the devastating diseases, but there
are still much to overcome.
Acknowledgments
We thank Brian Adams for discussion and critical reading for the manuscript.
References
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cells into enucleated frogs’ eggs, Proc Natl Acad Sci USA38: 455–463
(1952).
2. K. Hochedlinger and R. Jaenisch, Nuclear reprogramming and pluripotency,
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mouse embryonic and adult fibroblast cultures by defined factors, Cell126:
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Reprogramming and iPS Cells 23
fates, it becomes possible to conquer the devastating diseases, but there
are still much to overcome.
Acknowledgments
We thank Brian Adams for discussion and critical reading for the manuscript.
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mouse embryonic and adult fibroblast cultures by defined factors, Cell126:
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24 K.-Y. Kim and I.-H. Park
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B. E. Bernstein and R. Jaenisch, In vitroreprogramming of fibroblasts into a
pluripotent ES-cell-like state, Nature448: 318–324 (2007).
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Ma, L. Wang, F. Zeng and Q. Zhou, iPS cells produce viable mice through
tetraploid complementation, Nature461: 86–90 (2009).
14. M. J. Boland, J. L. Hazen, K. L. Nazor, A. R. Rodriguez, W. Gifford,
G. Martin, S. Kupriyanov and K. K. Baldwin, Adult mice generated from
induced pluripotent stem cells, Nature461: 91–94 (2009).
15. K. Takahashi, K. Tanabe, M. Ohnuki, M. Narita, T. Ichisaka, K. Tomoda and
S. Yamanaka, Induction of pluripotent stem cells from adult human fibrob-
lasts by defined factors, Cell131: 861–872 (2007).
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17. I. H. Park, R. Zhao, J. A. West, A. Yabuuchi, H. Huo, T. A. Ince, P. H. Lerou,
M. W. Lensch and G. Q. Daley, Reprogramming of human somatic cells to
pluripotency with defined factors, Nature451: 141–146 (2008).
18. N. Maherali, R. Sridharan, W. Xie, J. Utikal, S. Eminli, K. Arnold,
M. Stadtfeld, R. Yachechko, J. Tcheiu, R. Jaenisch, K. Plath and
K. Hochedlinger, Directly reprogrammed fibroblasts show global epigenetic
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(2007).
19. E. M. Chan, S. Ratanasirintrawoot, I. H. Park, P. D. Manos, Y. H. Loh,
H. Huo, J. D. Miller, O. Hartung, J. Rho, T. A. Ince, G. Q. Daley and
T. M. Schlaeger, Live cell imaging distinguishes bona fide human iPS cells
from partially reprogrammed cells, Nat Biotechnol27: 1033–1037 (2009).
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24 K.-Y. Kim and I.-H. Park
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26. T. Aoi, K. Yae, M. Nakagawa, T. Ichisaka, K. Okita, K. Takahashi, T. Chiba
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27. M. Stadtfeld, M. Nagaya, J. Utikal, G. Weir and K. Hochedlinger, Induced
pluripotent stem cells generated without viral integration, Science322:
945–949 (2008).
28. K. Okita, M. Nakagawa, H. Hyenjong, T. Ichisaka and S. Yamanaka,
Generation of mouse induced pluripotent stem cells without viral vectors,
Science322: 949–953 (2008).
29. J. Yu, K. Hu, K. Smuga-Otto, S. Tian, R. Stewart, Slukvin, II and J. A.
Thomson, Human induced pluripotent stem cells free of vector and transgene
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R. Jaenisch, Reprogramming of murine and human somatic cells using a
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cells induced from adult neural stem cells by reprogramming with two fac-
tors,Nature454: 646–650 (2008).
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and H. R. Scholer, Direct reprogramming of human neural stem cells by
OCT4,Nature461: 649–653 (2009).
25. D. Qin, Y. Gan, K. Shao, H. Wang, W. Li, T. Wang, W. He, J. Xu, Y. Zhang,
Z. Kou, L. Zeng, G. Sheng, M. A. Esteban, S. Gao and D. Pei, Mouse menin-
giocytes express Sox2 and yield high efficiency of chimeras after nuclear
reprogramming with exogenous factors, J Biol Chem283: 33730–33735 (2008).
26. T. Aoi, K. Yae, M. Nakagawa, T. Ichisaka, K. Okita, K. Takahashi, T. Chiba
and S. Yamanaka, Generation of pluripotent stem cells from adult mouse
liver and stomach cells, Science321: 699–702 (2008).
27. M. Stadtfeld, M. Nagaya, J. Utikal, G. Weir and K. Hochedlinger, Induced
pluripotent stem cells generated without viral integration, Science322:
945–949 (2008).
28. K. Okita, M. Nakagawa, H. Hyenjong, T. Ichisaka and S. Yamanaka,
Generation of mouse induced pluripotent stem cells without viral vectors,
Science322: 949–953 (2008).
29. J. Yu, K. Hu, K. Smuga-Otto, S. Tian, R. Stewart, Slukvin, II and J. A.
Thomson, Human induced pluripotent stem cells free of vector and transgene
sequences,Science324: 797–801 (2009).
30. B. W. Carey, S. Markoulaki, J. Hanna, K. Saha, Q. Gao, M. Mitalipova and
R. Jaenisch, Reprogramming of murine and human somatic cells using a
single polycistronic vector, Proc Natl Acad Sci USA106: 157–162 (2009).
31. C. A. Sommer, M. Stadtfeld, G. J. Murphy, K. Hochedlinger, D. N. Kotton
and G. Mostoslavsky, iPS cell generation using a single lentiviral stem cell
cassette,Stem Cells18: 18 (2008).
Reprogramming and iPS Cells 25
Other documents randomly have
different content
different content
party, or as interferes less with the parties' territorial and personal
supremacy, or as contains less general restrictions upon the parties.
(5) Previous treaties between the same parties, and treaties
between one of the parties and third parties, may be alluded to for
the purpose of clearing up the meaning of a stipulation.
(6) If there is a discrepancy between the clear meaning of a
stipulation, on the one hand, and, on the other, the intentions of one
of the parties declared during the negotiations preceding the signing
of a treaty, the decision must depend on the merits of the special
case. If, for instance, the discrepancy was produced through a mere
clerical error or by some other kind of mistake, it is obvious that an
interpretation is necessary in accordance with the real intentions of
the contracting parties.
(7) In case of a discrepancy between the clear meaning of a
stipulation, on the one hand, and, on the other, the intentions of all
the parties unanimously declared during the negotiations preceding
the signing of the treaty, the meaning which corresponds to the real
intentions of the parties must prevail over the meaning of the text.
If, therefore—as in the case of the Declaration of London of 1909—
the Report of the Drafting Committee contains certain interpretations
and is unanimously accepted as authoritative by all the negotiators
previous to the signing of the treaty, their interpretations must
prevail.
(8) If two meanings of a stipulation are admissible according to
the text of a treaty, such meaning is to prevail as the party
proposing the stipulation knew at the time to be the meaning
preferred by the party accepting it.
(9) If it is a matter of common knowledge that a State upholds a
meaning which is different from the generally prevailing meaning of
a term, and if nevertheless another State enters into a treaty with
the former in which such term is made use of, such meaning must
prevail as is upheld by the former. If, for instance, States conclude
commercial treaties with the United States of America in which the
most-favoured-nation clause
[912]
occurs, the particular meaning
which the United States attribute to this clause must prevail.
supremacy, or as contains less general restrictions upon the parties.
(5) Previous treaties between the same parties, and treaties
between one of the parties and third parties, may be alluded to for
the purpose of clearing up the meaning of a stipulation.
(6) If there is a discrepancy between the clear meaning of a
stipulation, on the one hand, and, on the other, the intentions of one
of the parties declared during the negotiations preceding the signing
of a treaty, the decision must depend on the merits of the special
case. If, for instance, the discrepancy was produced through a mere
clerical error or by some other kind of mistake, it is obvious that an
interpretation is necessary in accordance with the real intentions of
the contracting parties.
(7) In case of a discrepancy between the clear meaning of a
stipulation, on the one hand, and, on the other, the intentions of all
the parties unanimously declared during the negotiations preceding
the signing of the treaty, the meaning which corresponds to the real
intentions of the parties must prevail over the meaning of the text.
If, therefore—as in the case of the Declaration of London of 1909—
the Report of the Drafting Committee contains certain interpretations
and is unanimously accepted as authoritative by all the negotiators
previous to the signing of the treaty, their interpretations must
prevail.
(8) If two meanings of a stipulation are admissible according to
the text of a treaty, such meaning is to prevail as the party
proposing the stipulation knew at the time to be the meaning
preferred by the party accepting it.
(9) If it is a matter of common knowledge that a State upholds a
meaning which is different from the generally prevailing meaning of
a term, and if nevertheless another State enters into a treaty with
the former in which such term is made use of, such meaning must
prevail as is upheld by the former. If, for instance, States conclude
commercial treaties with the United States of America in which the
most-favoured-nation clause
[912]
occurs, the particular meaning
which the United States attribute to this clause must prevail.
[912] See below, § 580.
(10) If the meaning of a stipulation is ambiguous and one of the
contracting parties, at a time before a case arises for the application
of the stipulation, makes known what meaning it attributes to the
stipulation, the other party or parties cannot, when a case for the
application of the stipulation occurs, insist upon a different meaning.
They ought to have previously protested and taken the necessary
steps to secure an authentic interpretation of the ambiguous
stipulation. Thus, when in 1911 it became obvious that Germany and
other continental States attributed to article 23(h) of the Hague
Regulations respecting the Laws and Usages of War on Land a
meaning different from the one preferred by Great Britain, the
British Foreign Office made the British interpretation of this article
known.
(11) It is to be taken for granted that the parties intend the
stipulations of a treaty to have a certain effect and not to be
meaningless. Therefore, such interpretation is not admissible as
would make a stipulation meaningless or inefficient.
(12) All treaties must be interpreted so as to exclude fraud and so
as to make their operation consistent with good faith.
(13) The rules commonly applied by the Courts as regards the
interpretation and construction of Municipal Laws are in so far only
applicable to the interpretation and construction of treaties, and in
especial of law-making treaties, as they are general rules of
jurisprudence. If, however, they are particular rules, sanctioned only
by the Municipal Law or by the practice of the Courts of a particular
country, they may not be applied.
(14) If a treaty is concluded in two languages, for instance, a
treaty between Great Britain and France in English and French, and
if there is a discrepancy between the meaning of the two different
texts, each party is only bound by the text of its own language. But
a party cannot claim any advantage from the text of the language of
the other party.
(10) If the meaning of a stipulation is ambiguous and one of the
contracting parties, at a time before a case arises for the application
of the stipulation, makes known what meaning it attributes to the
stipulation, the other party or parties cannot, when a case for the
application of the stipulation occurs, insist upon a different meaning.
They ought to have previously protested and taken the necessary
steps to secure an authentic interpretation of the ambiguous
stipulation. Thus, when in 1911 it became obvious that Germany and
other continental States attributed to article 23(h) of the Hague
Regulations respecting the Laws and Usages of War on Land a
meaning different from the one preferred by Great Britain, the
British Foreign Office made the British interpretation of this article
known.
(11) It is to be taken for granted that the parties intend the
stipulations of a treaty to have a certain effect and not to be
meaningless. Therefore, such interpretation is not admissible as
would make a stipulation meaningless or inefficient.
(12) All treaties must be interpreted so as to exclude fraud and so
as to make their operation consistent with good faith.
(13) The rules commonly applied by the Courts as regards the
interpretation and construction of Municipal Laws are in so far only
applicable to the interpretation and construction of treaties, and in
especial of law-making treaties, as they are general rules of
jurisprudence. If, however, they are particular rules, sanctioned only
by the Municipal Law or by the practice of the Courts of a particular
country, they may not be applied.
(14) If a treaty is concluded in two languages, for instance, a
treaty between Great Britain and France in English and French, and
if there is a discrepancy between the meaning of the two different
texts, each party is only bound by the text of its own language. But
a party cannot claim any advantage from the text of the language of
the other party.
CHAPTER III
IMPORTANT GROUPS OF TREATIES
I
IMPORTANT LAW-MAKING TREATIES
Important Law-making Treaties a product of the Nineteenth Century.
§ 555. Law-making treaties
[913]
have been concluded ever since
International Law came into existence. It was not until the
nineteenth century, however, that such law-making treaties existed
as are of world-wide importance. Although at the Congress at
Münster and Osnabrück all the then existing European Powers, with
the exception of Great Britain, Russia, and Poland, were
represented, the Westphalian Peace of 1648, to which France,
Sweden, and the States of the German Empire were parties, and
which recognised the independence of Switzerland and the
Netherlands, on the one hand, and, on the other, the practical
sovereignty of the then existing 355 States of the German Empire,
was not of world-wide importance, in spite of the fact that it
contains various law-making stipulations. And the same may be said
with regard to all other treaties of peace between 1648 and 1815.
The first law-making treaty of world-wide importance was the Final
Act of the Vienna Congress, 1815, and the last, as yet, is the
Declaration of London of 1909. But it must be particularly noted that
not all of these are pure law-making treaties, since many contain
other stipulations besides those which are law-making.
[913] Concerning the conception of law-making treaties, see above, §§ 18 and 492.
Final Act of the Vienna Congress.
§ 556. The Final Act of the Vienna Congress,
[914]
signed on June 9,
1815, by Great Britain, Austria, France, Portugal, Prussia, Russia,
Spain, and Sweden-Norway, comprises law-making stipulations of
world-wide importance concerning four points—namely, first, the
perpetual neutralisation of Switzerland (article 118, No. 11);
IMPORTANT GROUPS OF TREATIES
I
IMPORTANT LAW-MAKING TREATIES
Important Law-making Treaties a product of the Nineteenth Century.
§ 555. Law-making treaties
[913]
have been concluded ever since
International Law came into existence. It was not until the
nineteenth century, however, that such law-making treaties existed
as are of world-wide importance. Although at the Congress at
Münster and Osnabrück all the then existing European Powers, with
the exception of Great Britain, Russia, and Poland, were
represented, the Westphalian Peace of 1648, to which France,
Sweden, and the States of the German Empire were parties, and
which recognised the independence of Switzerland and the
Netherlands, on the one hand, and, on the other, the practical
sovereignty of the then existing 355 States of the German Empire,
was not of world-wide importance, in spite of the fact that it
contains various law-making stipulations. And the same may be said
with regard to all other treaties of peace between 1648 and 1815.
The first law-making treaty of world-wide importance was the Final
Act of the Vienna Congress, 1815, and the last, as yet, is the
Declaration of London of 1909. But it must be particularly noted that
not all of these are pure law-making treaties, since many contain
other stipulations besides those which are law-making.
[913] Concerning the conception of law-making treaties, see above, §§ 18 and 492.
Final Act of the Vienna Congress.
§ 556. The Final Act of the Vienna Congress,
[914]
signed on June 9,
1815, by Great Britain, Austria, France, Portugal, Prussia, Russia,
Spain, and Sweden-Norway, comprises law-making stipulations of
world-wide importance concerning four points—namely, first, the
perpetual neutralisation of Switzerland (article 118, No. 11);
secondly, free navigation on so-called international rivers (articles
108-117); thirdly, the abolition of the negro slave trade (article 118,
No. 15); fourthly, the different classes of diplomatic envoys (article
118, No. 16).
[914] Martens, N.R. II. p. 379. See Angeberg, "Le congrès de Vienne et les traités de
1815" (4 vols., 1863).
Protocol of the Congress of Aix-la-Chapelle.
§ 557. The Protocol of November 21 of the Congress of Aix-la-
Chapelle,
[915]
1818, signed by Great Britain, Austria, France, Prussia,
and Russia, contains the important law-making stipulation
concerning the establishment of a fourth class of diplomatic envoys,
the so-called "Ministers Resident," to rank before the Chargés
d'Affaires.
[915] Martens, N.R. IV. p. 648. See Angeberg, op. cit.
Treaty of London of 1831.
§ 558. The Treaty of London
[916]
of November 15, 1831, signed by
Great Britain, Austria, France, Prussia, and Russia, comprises in its
article 7 the important law-making stipulation concerning the
perpetual neutralisation of Belgium.
[916] Martens, N.R. XI. p. 390. See Descamps, "La neutralité de la Belgique" (1902).
Declaration of Paris.
§ 559. The Declaration of Paris
[917]
of April 13, 1856, signed by
Great Britain, Austria, France, Prussia, Russia, Sardinia, and Turkey,
is a pure law-making treaty of the greatest importance, stipulating
four rules with regard to sea warfare—namely, that privateering is
abolished; that the neutral flag covers enemy goods with the
exception of contraband of war; that neutral goods, contraband
excepted, cannot be confiscated even when sailing under the enemy
flag; that a blockade must be effective to be binding.
[917] Martens, N.R.G. XV. p. 767.
Through accession during 1856, the following other States have
become parties to this treaty: Argentina, Belgium, Brazil, Chili,
Denmark, Ecuador, Greece, Guatemala, Hayti, Holland, Peru,
108-117); thirdly, the abolition of the negro slave trade (article 118,
No. 15); fourthly, the different classes of diplomatic envoys (article
118, No. 16).
[914] Martens, N.R. II. p. 379. See Angeberg, "Le congrès de Vienne et les traités de
1815" (4 vols., 1863).
Protocol of the Congress of Aix-la-Chapelle.
§ 557. The Protocol of November 21 of the Congress of Aix-la-
Chapelle,
[915]
1818, signed by Great Britain, Austria, France, Prussia,
and Russia, contains the important law-making stipulation
concerning the establishment of a fourth class of diplomatic envoys,
the so-called "Ministers Resident," to rank before the Chargés
d'Affaires.
[915] Martens, N.R. IV. p. 648. See Angeberg, op. cit.
Treaty of London of 1831.
§ 558. The Treaty of London
[916]
of November 15, 1831, signed by
Great Britain, Austria, France, Prussia, and Russia, comprises in its
article 7 the important law-making stipulation concerning the
perpetual neutralisation of Belgium.
[916] Martens, N.R. XI. p. 390. See Descamps, "La neutralité de la Belgique" (1902).
Declaration of Paris.
§ 559. The Declaration of Paris
[917]
of April 13, 1856, signed by
Great Britain, Austria, France, Prussia, Russia, Sardinia, and Turkey,
is a pure law-making treaty of the greatest importance, stipulating
four rules with regard to sea warfare—namely, that privateering is
abolished; that the neutral flag covers enemy goods with the
exception of contraband of war; that neutral goods, contraband
excepted, cannot be confiscated even when sailing under the enemy
flag; that a blockade must be effective to be binding.
[917] Martens, N.R.G. XV. p. 767.
Through accession during 1856, the following other States have
become parties to this treaty: Argentina, Belgium, Brazil, Chili,
Denmark, Ecuador, Greece, Guatemala, Hayti, Holland, Peru,
Portugal, Sweden-Norway, and Switzerland. Japan acceded in 1886,
Spain and Mexico in 1907.
Geneva Convention.
§ 560. The Geneva Convention
[918]
of August 22, 1864, and that of
July 6, 1906, are pure law-making treaties for the amelioration of the
conditions of the wounded of armies in the field. The Geneva
Convention of 1864 was originally signed only by Switzerland,
Baden, Belgium, Denmark, France, Holland, Italy, Prussia, and Spain,
but in time all other civilised States have acceded except Costa Rica,
Lichtenstein, and Monaco. A treaty
[919]
containing articles additional
to the Geneva Convention of 1864 was signed at Geneva on October
20, 1868, but was not ratified. A better fate was in store for the
Geneva Convention
[920]
of 1906, which was signed by the delegates
of thirty-five States, many of which have already granted ratification.
Colombia, Costa Rica, Cuba, Nicaragua, Turkey, and Venezuela have
already acceded. It is of importance to emphasise that the
Convention of 1864 is not entirely replaced by the Convention of
1906, in so far as the former remains in force between those Powers
which are parties to it without being parties to the latter. And it must
be remembered that the Final Act of the First as well as of the
Second Peace Conference contains a convention for the adaptation
to sea warfare of the principles of the Geneva Convention.
[918] Martens, N.R.G. XVIII. p. 607. See Lueder, "Die Genfer Convention" (1876), and
Münzel, "Untersuchungen über die Genfer Convention" (1901).
[919] Martens, N.R.G. XVIII. p. 612.
[920] Martens, N.R.G. 3rd Ser. II. p. 323.
Treaty of London of 1867.
§ 561. The Treaty of London
[921]
of May 11, 1867, signed by Great
Britain, Austria, Belgium, France, Holland, Italy, Prussia, and Russia,
comprises in its article 2 the important law-making stipulation
concerning the perpetual neutralisation of Luxemburg.
[921] Martens, N.R.G. XVIII. p. 445. See Wampach, "Le Luxembourg Neutre" (1900).
Declaration of St. Petersburg.
Spain and Mexico in 1907.
Geneva Convention.
§ 560. The Geneva Convention
[918]
of August 22, 1864, and that of
July 6, 1906, are pure law-making treaties for the amelioration of the
conditions of the wounded of armies in the field. The Geneva
Convention of 1864 was originally signed only by Switzerland,
Baden, Belgium, Denmark, France, Holland, Italy, Prussia, and Spain,
but in time all other civilised States have acceded except Costa Rica,
Lichtenstein, and Monaco. A treaty
[919]
containing articles additional
to the Geneva Convention of 1864 was signed at Geneva on October
20, 1868, but was not ratified. A better fate was in store for the
Geneva Convention
[920]
of 1906, which was signed by the delegates
of thirty-five States, many of which have already granted ratification.
Colombia, Costa Rica, Cuba, Nicaragua, Turkey, and Venezuela have
already acceded. It is of importance to emphasise that the
Convention of 1864 is not entirely replaced by the Convention of
1906, in so far as the former remains in force between those Powers
which are parties to it without being parties to the latter. And it must
be remembered that the Final Act of the First as well as of the
Second Peace Conference contains a convention for the adaptation
to sea warfare of the principles of the Geneva Convention.
[918] Martens, N.R.G. XVIII. p. 607. See Lueder, "Die Genfer Convention" (1876), and
Münzel, "Untersuchungen über die Genfer Convention" (1901).
[919] Martens, N.R.G. XVIII. p. 612.
[920] Martens, N.R.G. 3rd Ser. II. p. 323.
Treaty of London of 1867.
§ 561. The Treaty of London
[921]
of May 11, 1867, signed by Great
Britain, Austria, Belgium, France, Holland, Italy, Prussia, and Russia,
comprises in its article 2 the important law-making stipulation
concerning the perpetual neutralisation of Luxemburg.
[921] Martens, N.R.G. XVIII. p. 445. See Wampach, "Le Luxembourg Neutre" (1900).
Declaration of St. Petersburg.
§ 562. The Declaration of St. Petersburg
[922]
of November 29,
1868, signed by Great Britain, Austria-Hungary, Belgium, Denmark,
France, Greece, Holland, Italy, Persia, Portugal, Prussia and other
German States, Russia, Sweden-Norway, Switzerland, and Turkey—
Brazil acceded later on—is a pure law-making treaty. It stipulates
that projectiles of a weight below 400 grammes (14 ounces) which
are either explosive or charged with inflammable substances shall
not be made use of in war.
[922] Martens, N.R.G. XVIII. p. 474.
Treaty of Berlin of 1878.
§ 563. The Treaty of Berlin
[923]
of July 13, 1878, signed by Great
Britain, Austria-Hungary, France, Germany, Italy, Russia, and Turkey,
is law-making with regard to Bulgaria, Montenegro, Roumania, and
Servia. It is of great importance in so far as the present phase of the
solution of the Near Eastern Question arises therefrom, although
Bulgaria became full-sovereign in 1908.
[923] Martens, N.R.G. 2nd Ser. III. p. 449. See Mulas, "Il congresso di Berlino" (1878).
General Act of the Congo Conference.
§ 564. The General Act of the Congo Conference
[924]
of Berlin of
February 26, 1885, signed by Great Britain, Austria-Hungary,
Belgium, Denmark, France, Germany, Holland, Italy, Portugal,
Russia, Spain, Sweden-Norway, Turkey, and the United States of
America, is a law-making treaty of great importance, stipulating:
freedom of commerce for all nations within the basin of the river
Congo; prohibition of slave-transport within that basin; neutralisation
of Congo Territories; freedom of navigation for merchantmen of all
nations on the rivers Congo and Niger; and, lastly, the obligation of
the signatory Powers to notify to one another all future occupations
on the coast of the African continent.
[924] Martens, N.R.G. 2nd Ser. X. p. 414. See Patzig, "Die afrikanische Conferenz und
der Congostaat" (1885).
Treaty of Constantinople of 1888.
[922]
of November 29,
1868, signed by Great Britain, Austria-Hungary, Belgium, Denmark,
France, Greece, Holland, Italy, Persia, Portugal, Prussia and other
German States, Russia, Sweden-Norway, Switzerland, and Turkey—
Brazil acceded later on—is a pure law-making treaty. It stipulates
that projectiles of a weight below 400 grammes (14 ounces) which
are either explosive or charged with inflammable substances shall
not be made use of in war.
[922] Martens, N.R.G. XVIII. p. 474.
Treaty of Berlin of 1878.
§ 563. The Treaty of Berlin
[923]
of July 13, 1878, signed by Great
Britain, Austria-Hungary, France, Germany, Italy, Russia, and Turkey,
is law-making with regard to Bulgaria, Montenegro, Roumania, and
Servia. It is of great importance in so far as the present phase of the
solution of the Near Eastern Question arises therefrom, although
Bulgaria became full-sovereign in 1908.
[923] Martens, N.R.G. 2nd Ser. III. p. 449. See Mulas, "Il congresso di Berlino" (1878).
General Act of the Congo Conference.
§ 564. The General Act of the Congo Conference
[924]
of Berlin of
February 26, 1885, signed by Great Britain, Austria-Hungary,
Belgium, Denmark, France, Germany, Holland, Italy, Portugal,
Russia, Spain, Sweden-Norway, Turkey, and the United States of
America, is a law-making treaty of great importance, stipulating:
freedom of commerce for all nations within the basin of the river
Congo; prohibition of slave-transport within that basin; neutralisation
of Congo Territories; freedom of navigation for merchantmen of all
nations on the rivers Congo and Niger; and, lastly, the obligation of
the signatory Powers to notify to one another all future occupations
on the coast of the African continent.
[924] Martens, N.R.G. 2nd Ser. X. p. 414. See Patzig, "Die afrikanische Conferenz und
der Congostaat" (1885).
Treaty of Constantinople of 1888.
§ 565. The Treaty of Constantinople
[925]
of October 29, 1888,
signed by Great Britain, Austria-Hungary, France, Germany, Holland,
Italy, Russia, Spain, and Turkey, is a pure law-making treaty
stipulating the permanent neutralisation of the Suez Canal and the
freedom of navigation thereon for vessels of all nations.
[925] Martens, N.R.G. 2nd Ser. XV. p. 557. See above, § 183.
General Act of the Brussels Anti-Slavery Conference.
§ 566. The General Act of the Brussels Anti-Slavery Conference,
[926]
signed on July 2, 1890, by Great Britain, Austria-Hungary,
Belgium, the Congo Free State, Denmark, France,
[927]
Germany,
Holland, Italy, Persia, Portugal, Russia, Sweden-Norway, Spain,
Turkey, the United States of America, and Zanzibar, is a law-making
treaty of great importance which stipulates a system of measures for
the suppression of the slave-trade in Africa, and, incidentally,
restrictive measures concerning the spirit-trade in certain parts of
Africa. To revise the stipulations concerning this spirit-trade the
Convention of Brussels
[928]
of November 3, 1906, was signed by
Great Britain, Germany, Belgium, Spain, the Congo Free State,
France, Italy, Holland, Portugal, Russia, and Sweden.
[926] Martens, N.R.G. 2nd Ser. XVI. p. 3, and XXV. p. 543. See Lentner, "Der
afrikanische Sklavenhandel und die Brüsseler Conferenzen" (1891).
[927] But France only ratified this General Act with the exclusion of certain articles.
[928] Martens, N.R.G. 3rd Ser. I. p. 722.
Two Declarations of the First Hague Peace Conference.
§ 567. The Final Act of the Hague Peace Conference
[929]
of July 29,
1899, was a pure law-making treaty comprising three separate
conventions—namely, a convention for the peaceful adjustment of
international differences, a convention concerning the law of land
warfare, and a convention for the adaptation to maritime warfare of
the principles of the Geneva Convention of 1864,—and three
Declarations—namely, a Declaration prohibiting, for a term of five
years, the discharge of projectiles and explosives from balloons, a
Declaration concerning the prohibition of the use of projectiles the
only object of which is the diffusion of asphyxiating or deleterious
[925]
of October 29, 1888,
signed by Great Britain, Austria-Hungary, France, Germany, Holland,
Italy, Russia, Spain, and Turkey, is a pure law-making treaty
stipulating the permanent neutralisation of the Suez Canal and the
freedom of navigation thereon for vessels of all nations.
[925] Martens, N.R.G. 2nd Ser. XV. p. 557. See above, § 183.
General Act of the Brussels Anti-Slavery Conference.
§ 566. The General Act of the Brussels Anti-Slavery Conference,
[926]
signed on July 2, 1890, by Great Britain, Austria-Hungary,
Belgium, the Congo Free State, Denmark, France,
[927]
Germany,
Holland, Italy, Persia, Portugal, Russia, Sweden-Norway, Spain,
Turkey, the United States of America, and Zanzibar, is a law-making
treaty of great importance which stipulates a system of measures for
the suppression of the slave-trade in Africa, and, incidentally,
restrictive measures concerning the spirit-trade in certain parts of
Africa. To revise the stipulations concerning this spirit-trade the
Convention of Brussels
[928]
of November 3, 1906, was signed by
Great Britain, Germany, Belgium, Spain, the Congo Free State,
France, Italy, Holland, Portugal, Russia, and Sweden.
[926] Martens, N.R.G. 2nd Ser. XVI. p. 3, and XXV. p. 543. See Lentner, "Der
afrikanische Sklavenhandel und die Brüsseler Conferenzen" (1891).
[927] But France only ratified this General Act with the exclusion of certain articles.
[928] Martens, N.R.G. 3rd Ser. I. p. 722.
Two Declarations of the First Hague Peace Conference.
§ 567. The Final Act of the Hague Peace Conference
[929]
of July 29,
1899, was a pure law-making treaty comprising three separate
conventions—namely, a convention for the peaceful adjustment of
international differences, a convention concerning the law of land
warfare, and a convention for the adaptation to maritime warfare of
the principles of the Geneva Convention of 1864,—and three
Declarations—namely, a Declaration prohibiting, for a term of five
years, the discharge of projectiles and explosives from balloons, a
Declaration concerning the prohibition of the use of projectiles the
only object of which is the diffusion of asphyxiating or deleterious
gases, and a Declaration concerning the prohibition of so-called
dum-dum bullets. All these conventions, however, and the first of
these declarations have been replaced by the General Act of the
Second Hague Peace Conference, and only the last two declarations
are still in force. All the States which were represented at the
Conference are now parties to these declarations except the United
States of America.
[929] Martens, N.R.G. 2nd Ser. XXVI. p. 920. See Holls, "The Peace Conference at the
Hague" (1900), and Mérignhac, "La Conférence internationale de la Paix" (1900).
Treaty of Washington of 1901.
§ 568. The so-called Hay-Pauncefote Treaty of Washington
[930]
between Great Britain and the United States of America, signed
November 18, 1901, although law-making between the parties only,
is nevertheless of world-wide importance, because it neutralises
permanently the Panama Canal, which is in course of construction,
and stipulates free navigation thereon for vessels of all nations.
[931]
[930] Martens, N.R.G. 2nd Ser. XXX. p. 631.
[931] It ought to be mentioned that article 5 of the Boundary Treaty of Buenos Ayres,
signed by Argentina and Chili on September 15, 1881—see Martens, N.R.G. 2nd Ser. XII.
p. 491—contains a law-making stipulation of world-wide importance, because it
neutralises the Straits of Magellan for ever and declares them open to vessels of all
nations. See above, p. 267, note 2, and below, vol. II. § 72.
Conventions and Declaration of Second Hague Peace Conference.
§ 568a. The Final Act of the Second Hague Peace Conference of
October 18, 1907, is a pure law-making treaty of enormous
importance comprising the following thirteen conventions
[932]
and a
declaration:—
[932] Only a greater number of States have as yet ratified the Conventions, but it is to
be expected that many more will grant ratification in the course of time.
(1) Convention for the Pacific Settlement of International Disputes.
All States represented at the Conference signed except Nicaragua,
but some signed with reservations only. Nicaragua acceded later.
(2) Convention respecting the Limitation of the Employment of
Force for the Recovery of Contract Debts, signed by Great Britain,
Germany, the United States of America, Argentina, Austria-Hungary,
dum-dum bullets. All these conventions, however, and the first of
these declarations have been replaced by the General Act of the
Second Hague Peace Conference, and only the last two declarations
are still in force. All the States which were represented at the
Conference are now parties to these declarations except the United
States of America.
[929] Martens, N.R.G. 2nd Ser. XXVI. p. 920. See Holls, "The Peace Conference at the
Hague" (1900), and Mérignhac, "La Conférence internationale de la Paix" (1900).
Treaty of Washington of 1901.
§ 568. The so-called Hay-Pauncefote Treaty of Washington
[930]
between Great Britain and the United States of America, signed
November 18, 1901, although law-making between the parties only,
is nevertheless of world-wide importance, because it neutralises
permanently the Panama Canal, which is in course of construction,
and stipulates free navigation thereon for vessels of all nations.
[931]
[930] Martens, N.R.G. 2nd Ser. XXX. p. 631.
[931] It ought to be mentioned that article 5 of the Boundary Treaty of Buenos Ayres,
signed by Argentina and Chili on September 15, 1881—see Martens, N.R.G. 2nd Ser. XII.
p. 491—contains a law-making stipulation of world-wide importance, because it
neutralises the Straits of Magellan for ever and declares them open to vessels of all
nations. See above, p. 267, note 2, and below, vol. II. § 72.
Conventions and Declaration of Second Hague Peace Conference.
§ 568a. The Final Act of the Second Hague Peace Conference of
October 18, 1907, is a pure law-making treaty of enormous
importance comprising the following thirteen conventions
[932]
and a
declaration:—
[932] Only a greater number of States have as yet ratified the Conventions, but it is to
be expected that many more will grant ratification in the course of time.
(1) Convention for the Pacific Settlement of International Disputes.
All States represented at the Conference signed except Nicaragua,
but some signed with reservations only. Nicaragua acceded later.
(2) Convention respecting the Limitation of the Employment of
Force for the Recovery of Contract Debts, signed by Great Britain,
Germany, the United States of America, Argentina, Austria-Hungary,
Bolivia, Bulgaria, Chili, Columbia, Cuba, Denmark, San Domingo,
Ecuador, Spain, France, Greece, Guatemala, Haiti, Italy, Japan,
Mexico, Montenegro, Norway, Panama, Paraguay, Holland, Peru,
Persia, Portugal, Russia, Salvador, Servia, Turkey, Uruguay; China
and Nicaragua acceded later. Some of the South American States
signed with reservations.
(3) Convention relative to the Opening of Hostilities. All the States
represented at the Conference signed except China and Nicaragua;
both, however, acceded later.
(4) Convention concerning the Laws and Usages of War on Land.
All the States represented at the Conference signed except China,
Spain, and Nicaragua, but Nicaragua acceded later. Some States
made reservations in signing.
(5) Convention concerning the Rights and Duties of Neutral
Powers and Persons in Case of War on Land. All the States
represented at the Conference signed except China and Nicaragua,
but some States made reservations. Both China and Nicaragua
acceded later.
(6) Convention relative to the Status of Enemy Merchantmen at
the Outbreak of Hostilities. All the Powers represented at the
Conference signed except the United States of America, China, and
Nicaragua, but the last named acceded later. Some States made
reservations in signing.
(7) Convention relative to the Conversion of Merchant Ships into
War Ships. All the Powers represented at the Conference signed
except the United States of America, China, San Domingo,
Nicaragua, and Uruguay, but Nicaragua acceded later. Turkey made a
reservation in signing.
(8) Convention relative to the Laying of Automatic Submarine
Contact Mines. The majority of the States represented at the
Conference signed. China, Spain, Montenegro, Nicaragua, Portugal,
Russia, and Sweden have not signed, but Nicaragua acceded later.
Some States made reservations.
Ecuador, Spain, France, Greece, Guatemala, Haiti, Italy, Japan,
Mexico, Montenegro, Norway, Panama, Paraguay, Holland, Peru,
Persia, Portugal, Russia, Salvador, Servia, Turkey, Uruguay; China
and Nicaragua acceded later. Some of the South American States
signed with reservations.
(3) Convention relative to the Opening of Hostilities. All the States
represented at the Conference signed except China and Nicaragua;
both, however, acceded later.
(4) Convention concerning the Laws and Usages of War on Land.
All the States represented at the Conference signed except China,
Spain, and Nicaragua, but Nicaragua acceded later. Some States
made reservations in signing.
(5) Convention concerning the Rights and Duties of Neutral
Powers and Persons in Case of War on Land. All the States
represented at the Conference signed except China and Nicaragua,
but some States made reservations. Both China and Nicaragua
acceded later.
(6) Convention relative to the Status of Enemy Merchantmen at
the Outbreak of Hostilities. All the Powers represented at the
Conference signed except the United States of America, China, and
Nicaragua, but the last named acceded later. Some States made
reservations in signing.
(7) Convention relative to the Conversion of Merchant Ships into
War Ships. All the Powers represented at the Conference signed
except the United States of America, China, San Domingo,
Nicaragua, and Uruguay, but Nicaragua acceded later. Turkey made a
reservation in signing.
(8) Convention relative to the Laying of Automatic Submarine
Contact Mines. The majority of the States represented at the
Conference signed. China, Spain, Montenegro, Nicaragua, Portugal,
Russia, and Sweden have not signed, but Nicaragua acceded later.
Some States made reservations.
(9) Convention respecting Bombardments by Naval Forces in Time
of War. Except China, Spain, and Nicaragua all the States
represented at the Conference signed, but China and Nicaragua
acceded later. Some States made reservations.
(10) Convention for the Adaptation to Naval War of the Principles
of the Geneva Convention. All the Powers represented at the
Conference signed except Nicaragua, but some made reservations.
Nicaragua acceded later.
(11) Convention relative to certain Restrictions on the Exercise of
the Right of Capture in Maritime War. All States represented at the
Conference signed except China, Montenegro, Nicaragua, and
Russia, but Nicaragua acceded later.
(12) Convention relative to the Creation of an International Prize
Court. The majority of the States represented at the Conference
signed. Brazil, China, San Domingo, Greece, Luxemburg,
Montenegro, Nicaragua, Roumania, Russia, Servia, and Venezuela
have not signed, and some of the smaller signatory Powers made a
reservation with regard to the composition of the Court according to
article 15 of the Convention.
(13) Convention concerning the Rights and Duties of Neutral
Powers in Naval War. All the States represented at the Conference
signed except the United States of America, China, Cuba, Spain, and
Nicaragua. Some States made reservations. But the United States of
America, China, and Nicaragua acceded later.
(14) Declaration prohibiting the Discharge of Projectiles and
Explosives from Balloons. Only twenty-seven of the forty-four States
represented at the Conference signed. Germany, Chili, Denmark,
Spain, France, Guatemala, Italy, Japan, Mexico, Montenegro,
Nicaragua, Paraguay, Roumania, Russia, Servia, Sweden, and
Venezuela refused to sign, but Nicaragua acceded later.
The Declaration of London.
§ 568b. The Declaration of London
[933]
of February 26, 1909,
concerning the Laws of Naval War, is a pure law-making treaty of the
greatest importance. All the ten Powers represented at the
of War. Except China, Spain, and Nicaragua all the States
represented at the Conference signed, but China and Nicaragua
acceded later. Some States made reservations.
(10) Convention for the Adaptation to Naval War of the Principles
of the Geneva Convention. All the Powers represented at the
Conference signed except Nicaragua, but some made reservations.
Nicaragua acceded later.
(11) Convention relative to certain Restrictions on the Exercise of
the Right of Capture in Maritime War. All States represented at the
Conference signed except China, Montenegro, Nicaragua, and
Russia, but Nicaragua acceded later.
(12) Convention relative to the Creation of an International Prize
Court. The majority of the States represented at the Conference
signed. Brazil, China, San Domingo, Greece, Luxemburg,
Montenegro, Nicaragua, Roumania, Russia, Servia, and Venezuela
have not signed, and some of the smaller signatory Powers made a
reservation with regard to the composition of the Court according to
article 15 of the Convention.
(13) Convention concerning the Rights and Duties of Neutral
Powers in Naval War. All the States represented at the Conference
signed except the United States of America, China, Cuba, Spain, and
Nicaragua. Some States made reservations. But the United States of
America, China, and Nicaragua acceded later.
(14) Declaration prohibiting the Discharge of Projectiles and
Explosives from Balloons. Only twenty-seven of the forty-four States
represented at the Conference signed. Germany, Chili, Denmark,
Spain, France, Guatemala, Italy, Japan, Mexico, Montenegro,
Nicaragua, Paraguay, Roumania, Russia, Servia, Sweden, and
Venezuela refused to sign, but Nicaragua acceded later.
The Declaration of London.
§ 568b. The Declaration of London
[933]
of February 26, 1909,
concerning the Laws of Naval War, is a pure law-making treaty of the
greatest importance. All the ten Powers represented at the
Conference of London which produced this Declaration signed
[934]
it
—namely, Great Britain, Germany, the United States of America,
Austria-Hungary, Spain, France, Italy, Japan, Holland, and Russia,
but it is not yet ratified.
[933] On account of the opposition to the Ratification of the Declaration of London
which arose in England, the English literature on the Declaration is already very great.
The more important books are the following:—Bowles, "Sea Law and Sea Power" (1910);
Baty, "Britain and Sea Law" (1911); Bentwich, "The Declaration of London" (1911); Bray,
"British Rights at Sea" (1911); Bate, "An Elementary Account of the Declaration of
London" (1911); Civis, "Cargoes and Cruisers" (1911); Holland, "Proposed Changes in
Naval Prize Law" (1911); Cohen, "The Declaration of London" (1911). See also Baty and
Macdonell in the Twenty-sixth Report (1911) of the International Law Association. There
are also innumerable articles in periodicals.
[934] There is no doubt that the majority, if not all, of the States concerned will in time
accede to the Declaration of London.
II
ALLIANCES
Grotius, II. c. 15—Vattel, III. §§ 78-102—Twiss, I. § 246—Taylor, §§ 347-349—
Wheaton, §§ 278-285—Bluntschli, §§ 446-449—Heffter, § 92—Geffcken in
Holtzendorff, III. pp. 115-139—Ullmann, § 82—Bonfils, Nos. 871-881—Despagnet,
No. 459—Mérignhac, II. p. 683—Nys, III. pp. 554-557—Pradier-Fodéré, II. Nos.
934-967—Rivier, II. pp. 111-116—Calvo, III. §§ 1587-1588—Fiore, II. No. 1094,
and Code, Nos. 893-899—Martens, I. § 113—Rolin-Jaequemyns in R.I. XX. (1888),
pp. 5-35—Erich, "Ueber Allianzen und Allianzverhältnisse nach heutigem
Völkerrecht" (1907).
Conception of Alliances.
§ 569. Alliances in the strict sense of the term are treaties of
union between two or more States for the purpose of defending
each other against an attack in war, or of jointly attacking third
States, or for both purposes. The term "alliance" is, however, often
made use of in a wider sense, and it comprises in such cases treaties
of union for various purposes. Thus, the so-called "Holy Alliance,"
concluded in 1815 between the Emperors of Austria and Russia and
the King of Prussia, and afterwards joined by almost all of the
Sovereigns of Europe, was a union for such vague purposes that it
cannot be called an alliance in the strict sense of the term.
[934]
it
—namely, Great Britain, Germany, the United States of America,
Austria-Hungary, Spain, France, Italy, Japan, Holland, and Russia,
but it is not yet ratified.
[933] On account of the opposition to the Ratification of the Declaration of London
which arose in England, the English literature on the Declaration is already very great.
The more important books are the following:—Bowles, "Sea Law and Sea Power" (1910);
Baty, "Britain and Sea Law" (1911); Bentwich, "The Declaration of London" (1911); Bray,
"British Rights at Sea" (1911); Bate, "An Elementary Account of the Declaration of
London" (1911); Civis, "Cargoes and Cruisers" (1911); Holland, "Proposed Changes in
Naval Prize Law" (1911); Cohen, "The Declaration of London" (1911). See also Baty and
Macdonell in the Twenty-sixth Report (1911) of the International Law Association. There
are also innumerable articles in periodicals.
[934] There is no doubt that the majority, if not all, of the States concerned will in time
accede to the Declaration of London.
II
ALLIANCES
Grotius, II. c. 15—Vattel, III. §§ 78-102—Twiss, I. § 246—Taylor, §§ 347-349—
Wheaton, §§ 278-285—Bluntschli, §§ 446-449—Heffter, § 92—Geffcken in
Holtzendorff, III. pp. 115-139—Ullmann, § 82—Bonfils, Nos. 871-881—Despagnet,
No. 459—Mérignhac, II. p. 683—Nys, III. pp. 554-557—Pradier-Fodéré, II. Nos.
934-967—Rivier, II. pp. 111-116—Calvo, III. §§ 1587-1588—Fiore, II. No. 1094,
and Code, Nos. 893-899—Martens, I. § 113—Rolin-Jaequemyns in R.I. XX. (1888),
pp. 5-35—Erich, "Ueber Allianzen und Allianzverhältnisse nach heutigem
Völkerrecht" (1907).
Conception of Alliances.
§ 569. Alliances in the strict sense of the term are treaties of
union between two or more States for the purpose of defending
each other against an attack in war, or of jointly attacking third
States, or for both purposes. The term "alliance" is, however, often
made use of in a wider sense, and it comprises in such cases treaties
of union for various purposes. Thus, the so-called "Holy Alliance,"
concluded in 1815 between the Emperors of Austria and Russia and
the King of Prussia, and afterwards joined by almost all of the
Sovereigns of Europe, was a union for such vague purposes that it
cannot be called an alliance in the strict sense of the term.
History relates innumerable alliances between the several States.
They have always played, and still play, an important part in politics.
At the present time the triple alliance between Germany, Austria,
and Italy since 1879 and 1882, the alliance between Russia and
France since 1899, and that between Great Britain and Japan since
1902, renewed in 1905 and 1911, are illustrative examples.
[935]
[935] The following is the text of the Anglo-Japanese treaty of Alliance of 1911:—
The Government of Great Britain and the Government of Japan, having in view the
important changes which have taken place in the situation since the conclusion of the
Anglo-Japanese agreement of the 12th August 1905, and believing that a revision of that
Agreement responding to such changes would contribute to general stability and repose,
have agreed upon the following stipulations to replace the Agreement above mentioned,
such stipulations having the same object as the said Agreement, namely:—
(a) The consolidation and maintenance of the general peace in the regions of Eastern
Asia and of India;
(b) The preservation of the common interests of all Powers in China by insuring the
independence and integrity of the Chinese Empire and the principle of equal
opportunities for the commerce and industry of all nations in China;
(c) The maintenance of the territorial rights of the High Contracting Parties in the
regions of Eastern Asia and of India, and the defence of their special interests in the said
regions:—
Articäe I.
It is agreed that whenever, in the opinion of either Great Britain or Japan, any of the
rights and interests referred to in the preamble of this Agreement are in jeopardy, the
two Governments will communicate with one another fully and frankly, and will consider
in common the measures which should be taken to safeguard those menaced rights or
interests.
Articäe II.
If by reason of unprovoked attack or aggressive action, wherever arising, on the part
of any Power or Powers, either High Contracting Party should be involved in war in
defence of its territorial rights or special interests mentioned in the preamble of this
Agreement, the other High Contracting Party will at once come to the assistance of its
ally, and will conduct the war in common, and make peace in mutual agreement with it.
Articäe III.
The High Contracting Parties agree that neither of them will, without consulting the
other, enter into separate arrangements with another Power to the prejudice of the
objects described in the preamble of this Agreement.
Articäe IV.
Should either High Contracting Party conclude a treaty of general arbitration with a
third Power, it is agreed that nothing in this Agreement shall entail upon such Contracting
Party an obligation to go to war with the Power with whom such treaty of arbitration is in
force.
They have always played, and still play, an important part in politics.
At the present time the triple alliance between Germany, Austria,
and Italy since 1879 and 1882, the alliance between Russia and
France since 1899, and that between Great Britain and Japan since
1902, renewed in 1905 and 1911, are illustrative examples.
[935]
[935] The following is the text of the Anglo-Japanese treaty of Alliance of 1911:—
The Government of Great Britain and the Government of Japan, having in view the
important changes which have taken place in the situation since the conclusion of the
Anglo-Japanese agreement of the 12th August 1905, and believing that a revision of that
Agreement responding to such changes would contribute to general stability and repose,
have agreed upon the following stipulations to replace the Agreement above mentioned,
such stipulations having the same object as the said Agreement, namely:—
(a) The consolidation and maintenance of the general peace in the regions of Eastern
Asia and of India;
(b) The preservation of the common interests of all Powers in China by insuring the
independence and integrity of the Chinese Empire and the principle of equal
opportunities for the commerce and industry of all nations in China;
(c) The maintenance of the territorial rights of the High Contracting Parties in the
regions of Eastern Asia and of India, and the defence of their special interests in the said
regions:—
Articäe I.
It is agreed that whenever, in the opinion of either Great Britain or Japan, any of the
rights and interests referred to in the preamble of this Agreement are in jeopardy, the
two Governments will communicate with one another fully and frankly, and will consider
in common the measures which should be taken to safeguard those menaced rights or
interests.
Articäe II.
If by reason of unprovoked attack or aggressive action, wherever arising, on the part
of any Power or Powers, either High Contracting Party should be involved in war in
defence of its territorial rights or special interests mentioned in the preamble of this
Agreement, the other High Contracting Party will at once come to the assistance of its
ally, and will conduct the war in common, and make peace in mutual agreement with it.
Articäe III.
The High Contracting Parties agree that neither of them will, without consulting the
other, enter into separate arrangements with another Power to the prejudice of the
objects described in the preamble of this Agreement.
Articäe IV.
Should either High Contracting Party conclude a treaty of general arbitration with a
third Power, it is agreed that nothing in this Agreement shall entail upon such Contracting
Party an obligation to go to war with the Power with whom such treaty of arbitration is in
force.
Articäe V.
The conditions under which armed assistance shall be afforded by either Power to the
other in the circumstances mentioned in the present Agreement, and the means by
which such assistance is to be made available, will be arranged by the Naval and Military
authorities of the High Contracting Parties, who will from time to time consult one
another fully and freely upon all questions of mutual interest.
Articäe VI.
The present Agreement shall come into effect immediately after the date of its
signature, and remain in force for ten years from that date.
In case neither of the High Contracting Parties should have notified twelve months
before the expiration of the said ten years the intention of terminating it, it shall remain
binding until the expiration of one year from the day on which either of the High
Contracting Parties shall have denounced it. But if, when the date fixed for its expiration
arrives, either ally is actually engaged in war, the alliance shall, ipso facto, continue until
peace is concluded.
In faith whereof the undersigned, duly authorised by their respective Governments,
have signed this Agreement, and have affixed thereto their Seals.
Done in duplicate at London, the 13th day of July 1911.
Parties to Alliance.
§ 570. Subjects of alliances are said to be full-Sovereign States
only. But the fact cannot be denied that alliances have been
concluded by States under suzerainty. Thus, the convention of April
16, 1877, between Roumania, which was then under Turkish
suzerainty, and Russia, concerning the passage of Russian troops
through Roumanian territory in case of war with Turkey, was
practically a treaty of alliance.
[936]
Thus, further, the former South
African Republic, although, at any rate according to the views of the
British Government, a half-Sovereign State under British suzerainty,
concluded an alliance with the former Orange Free State by treaty of
March 17, 1897.
[937]
[936] See Martens, N.R.G. 2nd Ser. III. p. 182.
[937] See Martens, N.R.G. 2nd Ser. XXV. p. 327.
A neutralised State can be the subject of an alliance for the
purpose of defence, whereas the entrance into an offensive alliance
on the part of such State would involve a breach of its neutrality.
Different kinds of Alliances.
The conditions under which armed assistance shall be afforded by either Power to the
other in the circumstances mentioned in the present Agreement, and the means by
which such assistance is to be made available, will be arranged by the Naval and Military
authorities of the High Contracting Parties, who will from time to time consult one
another fully and freely upon all questions of mutual interest.
Articäe VI.
The present Agreement shall come into effect immediately after the date of its
signature, and remain in force for ten years from that date.
In case neither of the High Contracting Parties should have notified twelve months
before the expiration of the said ten years the intention of terminating it, it shall remain
binding until the expiration of one year from the day on which either of the High
Contracting Parties shall have denounced it. But if, when the date fixed for its expiration
arrives, either ally is actually engaged in war, the alliance shall, ipso facto, continue until
peace is concluded.
In faith whereof the undersigned, duly authorised by their respective Governments,
have signed this Agreement, and have affixed thereto their Seals.
Done in duplicate at London, the 13th day of July 1911.
Parties to Alliance.
§ 570. Subjects of alliances are said to be full-Sovereign States
only. But the fact cannot be denied that alliances have been
concluded by States under suzerainty. Thus, the convention of April
16, 1877, between Roumania, which was then under Turkish
suzerainty, and Russia, concerning the passage of Russian troops
through Roumanian territory in case of war with Turkey, was
practically a treaty of alliance.
[936]
Thus, further, the former South
African Republic, although, at any rate according to the views of the
British Government, a half-Sovereign State under British suzerainty,
concluded an alliance with the former Orange Free State by treaty of
March 17, 1897.
[937]
[936] See Martens, N.R.G. 2nd Ser. III. p. 182.
[937] See Martens, N.R.G. 2nd Ser. XXV. p. 327.
A neutralised State can be the subject of an alliance for the
purpose of defence, whereas the entrance into an offensive alliance
on the part of such State would involve a breach of its neutrality.
Different kinds of Alliances.
§ 571. As already mentioned, an alliance may be offensive or
defensive, or both. All three kinds may be either general alliances, in
which case the allies are united against any possible enemy
whatever, or particular alliances against one or more individual
enemies. Alliances, further, may be either permanent or temporary,
and in the latter case they expire with the period of time for which
they were concluded. As regards offensive alliances, it must be
emphasised that they are valid only when their object is not
immoral.
[938]
[938] See above, § 505.
Conditions of Alliances.
§ 572. Alliances may contain all sorts of conditions. The most
important are the conditions regarding the assistance to be
rendered. It may be that assistance is to be rendered with the whole
or a limited part of the military and naval forces of the allies, or with
the whole or a limited part of their military or with the whole or a
limited part of their naval forces only. Assistance may, further, be
rendered in money only, so that one of the allies is fighting with his
forces while the other supplies a certain sum of money for their
maintenance. A treaty of alliance of such a kind must not be
confounded with a simple treaty of subsidy. If two States enter into
a convention that one of the parties shall furnish the other
permanently in time of peace and war with a limited number of
troops in return for a certain annual payment, such convention is not
an alliance, but a treaty of subsidy only. But if two States enter into
a convention that in case of war one of the parties shall furnish the
other with a limited number of troops, be it in return for payment or
not, such convention really constitutes an alliance. For every
convention concluded for the purpose of lending succour in time of
war implies an alliance. It is for this reason that the above-
mentioned
[939]
treaty of 1877 between Russia and Roumania
concerning the passage of Russian troops through Roumanian
territory in case of war against Turkey was really a treaty of alliance.
[939] See above, § 570.
defensive, or both. All three kinds may be either general alliances, in
which case the allies are united against any possible enemy
whatever, or particular alliances against one or more individual
enemies. Alliances, further, may be either permanent or temporary,
and in the latter case they expire with the period of time for which
they were concluded. As regards offensive alliances, it must be
emphasised that they are valid only when their object is not
immoral.
[938]
[938] See above, § 505.
Conditions of Alliances.
§ 572. Alliances may contain all sorts of conditions. The most
important are the conditions regarding the assistance to be
rendered. It may be that assistance is to be rendered with the whole
or a limited part of the military and naval forces of the allies, or with
the whole or a limited part of their military or with the whole or a
limited part of their naval forces only. Assistance may, further, be
rendered in money only, so that one of the allies is fighting with his
forces while the other supplies a certain sum of money for their
maintenance. A treaty of alliance of such a kind must not be
confounded with a simple treaty of subsidy. If two States enter into
a convention that one of the parties shall furnish the other
permanently in time of peace and war with a limited number of
troops in return for a certain annual payment, such convention is not
an alliance, but a treaty of subsidy only. But if two States enter into
a convention that in case of war one of the parties shall furnish the
other with a limited number of troops, be it in return for payment or
not, such convention really constitutes an alliance. For every
convention concluded for the purpose of lending succour in time of
war implies an alliance. It is for this reason that the above-
mentioned
[939]
treaty of 1877 between Russia and Roumania
concerning the passage of Russian troops through Roumanian
territory in case of war against Turkey was really a treaty of alliance.
[939] See above, § 570.
Casus Fœderis.
§ 573. Casus fœderis is the event upon the occurrence of which it
becomes the duty of one of the allies to render the promised
assistance to the other. Thus in case of a defensive alliance the
casus fœderis occurs when war is declared or commenced against
one of the allies. Treaties of alliance very often define precisely the
event which shall be the casus fœderis, and then the latter is less
exposed to controversy. But, on the other hand, there have been
many alliances concluded without such specialisation, and,
consequently, disputes have arisen later between the parties as to
the casus fœderis.
That the casus fœderis is not influenced by the fact that a State,
subsequent to entering into an alliance, concludes a treaty of
general arbitration with a third State, has been pointed out above, §
522.
III
TREATIES OF GUARANTEE AND OF PROTECTION
Vattel, II. §§ 235-239—Hall, § 113—Phillimore, II. §§ 56-63—Twiss, I. § 249—
Halleck, I. p. 285—Taylor, §§ 350-353—Wheaton, § 278—Bluntschli, §§ 430-439—
Heffter, § 97—Geffcken in Holtzendorff, III. pp. 85-112—Liszt, § 22—Ullmann, § 83
—Fiore, Code, Nos. 787-791—Bonfils, Nos. 882-893—Despagnet, No. 461—
Mérignhac, II. p. 681—Nys, III. pp. 36-41—Pradier-Fodéré, II. Nos. 969-1020—
Rivier, II. pp. 97-105—Calvo, III. §§ 1584-1585—Martens, I. § 115—Neyron, "Essai
historique et politique sur les garanties" (1779)—Milovanovitch, "Des traités de
garantie en droit international" (1888)—Erich, "Ueber Allianzen und
Allianzverhältnisse nach heutigem Völkerrecht" (1907)—Quabbe, "Die
völkerrechtliche Garantie" (1911).
Conception and Object of Guarantee Treaties.
§ 574. Treaties of guarantee are conventions by which one of the
parties engages to do what is in its power to secure a certain object
to the other party. Guarantee treaties may be mutual or unilateral.
They may be concluded by two States only, or by a number of States
jointly, and in the latter case the single guarantors may give their
§ 573. Casus fœderis is the event upon the occurrence of which it
becomes the duty of one of the allies to render the promised
assistance to the other. Thus in case of a defensive alliance the
casus fœderis occurs when war is declared or commenced against
one of the allies. Treaties of alliance very often define precisely the
event which shall be the casus fœderis, and then the latter is less
exposed to controversy. But, on the other hand, there have been
many alliances concluded without such specialisation, and,
consequently, disputes have arisen later between the parties as to
the casus fœderis.
That the casus fœderis is not influenced by the fact that a State,
subsequent to entering into an alliance, concludes a treaty of
general arbitration with a third State, has been pointed out above, §
522.
III
TREATIES OF GUARANTEE AND OF PROTECTION
Vattel, II. §§ 235-239—Hall, § 113—Phillimore, II. §§ 56-63—Twiss, I. § 249—
Halleck, I. p. 285—Taylor, §§ 350-353—Wheaton, § 278—Bluntschli, §§ 430-439—
Heffter, § 97—Geffcken in Holtzendorff, III. pp. 85-112—Liszt, § 22—Ullmann, § 83
—Fiore, Code, Nos. 787-791—Bonfils, Nos. 882-893—Despagnet, No. 461—
Mérignhac, II. p. 681—Nys, III. pp. 36-41—Pradier-Fodéré, II. Nos. 969-1020—
Rivier, II. pp. 97-105—Calvo, III. §§ 1584-1585—Martens, I. § 115—Neyron, "Essai
historique et politique sur les garanties" (1779)—Milovanovitch, "Des traités de
garantie en droit international" (1888)—Erich, "Ueber Allianzen und
Allianzverhältnisse nach heutigem Völkerrecht" (1907)—Quabbe, "Die
völkerrechtliche Garantie" (1911).
Conception and Object of Guarantee Treaties.
§ 574. Treaties of guarantee are conventions by which one of the
parties engages to do what is in its power to secure a certain object
to the other party. Guarantee treaties may be mutual or unilateral.
They may be concluded by two States only, or by a number of States
jointly, and in the latter case the single guarantors may give their
guarantee severally or collectively or both. And the guarantee may
be for a certain period of time only or permanent.
The possible objects of guarantee treaties are numerous.
[940]
It
suffices to give the following chief examples: the performance of a
particular act on the part of a certain State, as the discharge of a
debt or the cession of a territory; certain rights of a State; the
undisturbed possession of the whole or a particular part of the
territory; a particular form of Constitution; a certain status, as
permanent neutrality
[941]
or independence
[942]
or integrity
[943]
; a
particular dynastic succession; the fulfilment of a treaty concluded
by a third State.
[940] The important part that treaties of guarantee play in politics may be seen from a
glance at Great Britain's guarantee treaties. See Munro, "England's Treaties of
Guarantee," in The Law Magazine and Review, VI. (1881), pp. 215-238.
[941] See above, § 95.
[942] Thus Great Britain, France, and Russia have guaranteed, by the Treaty with
Denmark of July 13, 1863, the independence (but also the monarchy) of Greece
(Martens, N.R.G. XVII. Part. II. p. 79). The United States of America has guaranteed the
independence of Cuba by the Treaty of Havana of May 22, 1903 (Martens, N.R.G. 2nd
Ser. XXXII. p. 79), and of Panama by the Treaty of Washington of November 18, 1903
(Martens, N.R.G. 2nd Ser. XXXI. p. 599).
[943] Thus the integrity of Norway is guaranteed by Great Britain, Germany, France,
and Russia by the Treaty of Christiania of November 2, 1907; see Martens, N.R.G. 3rd
Ser. II. p. 9. A condition of this integrity is that Norway does not cede any part of her
territory to any foreign Power.
Effect of Treaties of Guarantee.
§ 575. The effect of guarantee treaties is the creation of the duty
of the guarantors to do what is in their power in order to secure the
guaranteed objects. The compulsion to be applied by a guarantor for
that purpose depends upon the circumstances; it may eventually be
war. But the duty of the guarantor to render, even by compulsion,
the promised assistance to the guaranteed depends upon many
conditions and circumstances. Thus, first, the guaranteed must
request the guarantor to render assistance. When, for instance, the
possession of a certain part of its territory is guaranteed to a State
which after its defeat in a war with a third State agrees as a
condition of peace to cede such piece of territory to the victor
be for a certain period of time only or permanent.
The possible objects of guarantee treaties are numerous.
[940]
It
suffices to give the following chief examples: the performance of a
particular act on the part of a certain State, as the discharge of a
debt or the cession of a territory; certain rights of a State; the
undisturbed possession of the whole or a particular part of the
territory; a particular form of Constitution; a certain status, as
permanent neutrality
[941]
or independence
[942]
or integrity
[943]
; a
particular dynastic succession; the fulfilment of a treaty concluded
by a third State.
[940] The important part that treaties of guarantee play in politics may be seen from a
glance at Great Britain's guarantee treaties. See Munro, "England's Treaties of
Guarantee," in The Law Magazine and Review, VI. (1881), pp. 215-238.
[941] See above, § 95.
[942] Thus Great Britain, France, and Russia have guaranteed, by the Treaty with
Denmark of July 13, 1863, the independence (but also the monarchy) of Greece
(Martens, N.R.G. XVII. Part. II. p. 79). The United States of America has guaranteed the
independence of Cuba by the Treaty of Havana of May 22, 1903 (Martens, N.R.G. 2nd
Ser. XXXII. p. 79), and of Panama by the Treaty of Washington of November 18, 1903
(Martens, N.R.G. 2nd Ser. XXXI. p. 599).
[943] Thus the integrity of Norway is guaranteed by Great Britain, Germany, France,
and Russia by the Treaty of Christiania of November 2, 1907; see Martens, N.R.G. 3rd
Ser. II. p. 9. A condition of this integrity is that Norway does not cede any part of her
territory to any foreign Power.
Effect of Treaties of Guarantee.
§ 575. The effect of guarantee treaties is the creation of the duty
of the guarantors to do what is in their power in order to secure the
guaranteed objects. The compulsion to be applied by a guarantor for
that purpose depends upon the circumstances; it may eventually be
war. But the duty of the guarantor to render, even by compulsion,
the promised assistance to the guaranteed depends upon many
conditions and circumstances. Thus, first, the guaranteed must
request the guarantor to render assistance. When, for instance, the
possession of a certain part of its territory is guaranteed to a State
which after its defeat in a war with a third State agrees as a
condition of peace to cede such piece of territory to the victor
without having requested the intervention of the guarantor, the latter
has neither a right nor a duty to interfere. Thus, secondly, the
guarantor must at the critical time be able to render the required
assistance. When, for instance, its hands are tied through waging
war against a third State, or when it is so weak through internal
troubles or other factors that its interference would expose it to a
serious danger, it is not bound to fulfil the request for assistance. So
too, when the guaranteed has not complied with previous advice
given by the guarantor as to the line of its behaviour, it is not the
guarantor's duty to render assistance afterwards.
It is impossible to state all the circumstances and conditions upon
which the fulfilment of the duty of the guarantor depends, as every
case must be judged upon its own merits. And it is certain that,
more frequently than in other cases, changes in political
constellations and the general developments of events may involve
such vital change of circumstances as to justify
[944]
a State in
refusing to interfere in spite of a treaty of guarantee. It is for this
reason that treaties of guarantee to secure permanently a certain
object to a State are naturally of a more or less precarious value to
the latter. The practical value, therefore, of a guarantee treaty,
whatever may be its formal character, would as a rule seem to
extend to the early years only of its existence while the original
conditions still obtain.
[944] See above, § 539.
Effect of Collective Guarantee.
§ 576. In contradistinction to treaties constituting a guarantee on
the part of one or more States severally, the effect of treaties
constituting a collective guarantee on the part of several States
requires special consideration. On June 20, 1867, Lord Derby
maintained
[945]
in the House of Lords concerning the collective
guarantee by the Powers of the neutralisation of Luxemburg that in
case of a collective guarantee each guarantor had only the duty to
act according to the treaty when all the other guarantors were ready
to act likewise; that, consequently, if one of the guarantors
themselves should violate the neutrality of Luxemburg, the duty to
has neither a right nor a duty to interfere. Thus, secondly, the
guarantor must at the critical time be able to render the required
assistance. When, for instance, its hands are tied through waging
war against a third State, or when it is so weak through internal
troubles or other factors that its interference would expose it to a
serious danger, it is not bound to fulfil the request for assistance. So
too, when the guaranteed has not complied with previous advice
given by the guarantor as to the line of its behaviour, it is not the
guarantor's duty to render assistance afterwards.
It is impossible to state all the circumstances and conditions upon
which the fulfilment of the duty of the guarantor depends, as every
case must be judged upon its own merits. And it is certain that,
more frequently than in other cases, changes in political
constellations and the general developments of events may involve
such vital change of circumstances as to justify
[944]
a State in
refusing to interfere in spite of a treaty of guarantee. It is for this
reason that treaties of guarantee to secure permanently a certain
object to a State are naturally of a more or less precarious value to
the latter. The practical value, therefore, of a guarantee treaty,
whatever may be its formal character, would as a rule seem to
extend to the early years only of its existence while the original
conditions still obtain.
[944] See above, § 539.
Effect of Collective Guarantee.
§ 576. In contradistinction to treaties constituting a guarantee on
the part of one or more States severally, the effect of treaties
constituting a collective guarantee on the part of several States
requires special consideration. On June 20, 1867, Lord Derby
maintained
[945]
in the House of Lords concerning the collective
guarantee by the Powers of the neutralisation of Luxemburg that in
case of a collective guarantee each guarantor had only the duty to
act according to the treaty when all the other guarantors were ready
to act likewise; that, consequently, if one of the guarantors
themselves should violate the neutrality of Luxemburg, the duty to
act according to the treaty of collective guarantee would not accrue
to the other guarantors. This opinion is certainly not correct,
[946]
and
I do not know of any publicist who would or could approve of it.
There ought to be no doubt that in a case of collective guarantee
one of the guarantors alone cannot be considered bound to act
according to the treaty of guarantee. For a collective guarantee can
have the meaning only that the guarantors should act in a body. But
if one of the guarantors themselves violates the object of his own
guarantee, the body of the guarantors remains, and it is certainly
their duty to act against such faithless co-guarantor. If, however, the
majority,
[947]
and therefore the body of the guarantors, were to
violate the very object of their guarantee, the duty to act against
them would not accrue to the minority.
[945] Hansard, vol. 183, p. 150.
[946] See Hall, § 113; Bluntschli, § 440; and Quabbe, op. cit. pp. 149-159.
[947] See against this statement Quabbe, op. cit. p. 158.
Different, however, is the case in which a number of Powers have
collectively and severally guaranteed a certain object. Then, not only
as a body but also individually, it is their duty to interfere in any case
of violation of the object of guarantee.
Pseudo-Guarantees.
§ 576a. Different from real Guarantee Treaties are such treaties as
declare the policy of the parties with regard to the maintenance of
their territorial status quo. Whereas treaties guaranteeing the
maintenance of the territorial status quo engage the guarantors to
do what they can to maintain such status quo, treaties declaring the
policy of the parties with regard to the maintenance of their
territorial status quo do not contain any legal engagements, but
simply state the firm resolution of the parties to uphold the status
quo. In contradistinction to real guarantee treaties, such treaties
declaring the policy of the parties may fitly be called Pseudo-
Guarantee Treaties, and although their political value is very great,
they have scarcely any legal importance. For the parties do not bind
themselves to pursue a policy for maintaining the status quo, they
only declare their firm resolution to that end. Further, the parties do
to the other guarantors. This opinion is certainly not correct,
[946]
and
I do not know of any publicist who would or could approve of it.
There ought to be no doubt that in a case of collective guarantee
one of the guarantors alone cannot be considered bound to act
according to the treaty of guarantee. For a collective guarantee can
have the meaning only that the guarantors should act in a body. But
if one of the guarantors themselves violates the object of his own
guarantee, the body of the guarantors remains, and it is certainly
their duty to act against such faithless co-guarantor. If, however, the
majority,
[947]
and therefore the body of the guarantors, were to
violate the very object of their guarantee, the duty to act against
them would not accrue to the minority.
[945] Hansard, vol. 183, p. 150.
[946] See Hall, § 113; Bluntschli, § 440; and Quabbe, op. cit. pp. 149-159.
[947] See against this statement Quabbe, op. cit. p. 158.
Different, however, is the case in which a number of Powers have
collectively and severally guaranteed a certain object. Then, not only
as a body but also individually, it is their duty to interfere in any case
of violation of the object of guarantee.
Pseudo-Guarantees.
§ 576a. Different from real Guarantee Treaties are such treaties as
declare the policy of the parties with regard to the maintenance of
their territorial status quo. Whereas treaties guaranteeing the
maintenance of the territorial status quo engage the guarantors to
do what they can to maintain such status quo, treaties declaring the
policy of the parties with regard to the maintenance of their
territorial status quo do not contain any legal engagements, but
simply state the firm resolution of the parties to uphold the status
quo. In contradistinction to real guarantee treaties, such treaties
declaring the policy of the parties may fitly be called Pseudo-
Guarantee Treaties, and although their political value is very great,
they have scarcely any legal importance. For the parties do not bind
themselves to pursue a policy for maintaining the status quo, they
only declare their firm resolution to that end. Further, the parties do
not engage themselves to uphold the status quo, but only to
communicate with one another, in case the status quo is threatened,
with a view to agreeing upon such measures as they may consider
advisable for the maintenance of the status quo. To this class of
pseudo-guarantee treaties belong:—
(1) The Declarations
[948]
exchanged on May 16, 1907, between
France and Spain on the one hand, and, on the other hand, between
Great Britain and Spain, concerning the territorial status quo in the
Mediterranean. Each party declares that its general policy with
regard to the Mediterranean is directed to the maintenance of the
territorial status quo, and that it is therefore resolved to preserve
intact its rights over its insular and maritime possessions within the
Mediterranean. Each party declares, further, that, should
circumstances arise which would tend to alter the existing territorial
status quo, it will communicate with the other party in order to
afford it the opportunity to concert, if desired, by mutual agreement
the course of action which the two parties shall adopt in common.
[948] See Martens, N.R.G. 2nd Ser. XXXV. p. 692, and 3rd Ser. I. p. 3.
(2) The Declarations
[949]
concerning the maintenance of the
territorial status quo in the North Sea, signed at Berlin on April 23,
1908, by Great Britain, Germany, Denmark, France, Holland, and
Sweden, and concerning the maintenance of the territorial status
quo in the Baltic, signed at St. Petersburg, likewise on April 23,
1908, by Germany, Denmark, Russia, and Sweden. The parties
declare their firm resolution to preserve intact the rights of all the
parties over their continental and insular possessions within the
region of the North Sea, and of the Baltic respectively. And the
parties concerned further declare that, should the present territorial
status quo be threatened by any events whatever, they will enter
into communication with one another with a view to agreeing upon
such measures as they may consider advisable in the interest of the
maintenance of the status quo.
[949] See Martens, N.R.G. 3rd Ser. I. pp. 17 and 18.
There is no doubt that the texts of the Declarations concerning the
status quo in the North Sea and the Baltic stipulate a stricter
communicate with one another, in case the status quo is threatened,
with a view to agreeing upon such measures as they may consider
advisable for the maintenance of the status quo. To this class of
pseudo-guarantee treaties belong:—
(1) The Declarations
[948]
exchanged on May 16, 1907, between
France and Spain on the one hand, and, on the other hand, between
Great Britain and Spain, concerning the territorial status quo in the
Mediterranean. Each party declares that its general policy with
regard to the Mediterranean is directed to the maintenance of the
territorial status quo, and that it is therefore resolved to preserve
intact its rights over its insular and maritime possessions within the
Mediterranean. Each party declares, further, that, should
circumstances arise which would tend to alter the existing territorial
status quo, it will communicate with the other party in order to
afford it the opportunity to concert, if desired, by mutual agreement
the course of action which the two parties shall adopt in common.
[948] See Martens, N.R.G. 2nd Ser. XXXV. p. 692, and 3rd Ser. I. p. 3.
(2) The Declarations
[949]
concerning the maintenance of the
territorial status quo in the North Sea, signed at Berlin on April 23,
1908, by Great Britain, Germany, Denmark, France, Holland, and
Sweden, and concerning the maintenance of the territorial status
quo in the Baltic, signed at St. Petersburg, likewise on April 23,
1908, by Germany, Denmark, Russia, and Sweden. The parties
declare their firm resolution to preserve intact the rights of all the
parties over their continental and insular possessions within the
region of the North Sea, and of the Baltic respectively. And the
parties concerned further declare that, should the present territorial
status quo be threatened by any events whatever, they will enter
into communication with one another with a view to agreeing upon
such measures as they may consider advisable in the interest of the
maintenance of the status quo.
[949] See Martens, N.R.G. 3rd Ser. I. pp. 17 and 18.
There is no doubt that the texts of the Declarations concerning the
status quo in the North Sea and the Baltic stipulate a stricter
engagement of the respective parties than the texts of the
Declarations concerning the status quo in the Mediterranean, but
neither
[950]
of them comprises a real legal guarantee.
[950] Whereas Quabbe (p. 97, note 1), correctly denies the character of a real
guarantee to the Declarations concerning the Mediterranean, he (p. 105) considers the
Declarations concerning the North Sea and the Baltic real Guarantee Treaties.
Treaties of Protection.
§ 577. Different from guarantee treaties are treaties of protection.
Whereas the former constitute the guarantee of a certain object to
the guaranteed, treaties of protection are treaties by which strong
States simply engage to protect weaker States without any
guarantee whatever. A treaty of protection must, however, not be
confounded with a treaty of protectorate.
[951]
[951] See above, § 92.
IV
COMMERCIAL TREATIES
Taylor, 354—Moore, V. §§ 765-769—Melle in Holtzendorff, III. pp. 143-256—Liszt, §
28—Ullmann, § 145—Bonfils, No. 918—Despagnet, No. 462—Pradier-Fodéré, IV.
Nos. 2005-2033—Mérignhac, II. pp. 688-693—Rivier, I. pp. 370-374—Fiore, II.
Nos. 1065-1077, and Code, Nos. 848-854—Martens, II. §§ 52-55—Steck, "Versuch
über Handels- und Schiffahrtsverträge" (1782)—Schraut, "System der
Handelsverträge und der Meistbegünstigung" (1884)—Veillcovitch, "Les traités de
commerce" (1892)—Nys, "Les origines du droit international" (1894), pp. 278-294
—Herod, "Favoured Nation Treatment" (1901)—Calwer, "Die Meistbegünstigung in
den Vereinigten Staaten von Nord-America" (1902)—Glier, "Die
Meistbegünstigungs-Klausel" (1906)—Cavaretta, "La clausola della natiozione più
favorita" (1906)—Barclay, "Problems of International Law and Diplomacy" (1907),
pp. 137-142—Hornbeck, "The Most-Favoured Nation Clause" (1910), and in A.J.
III. (1909), pp. 394-422, 619-647, and 798-827—Lehr in R.I. XXV. (1893), pp. 313-
316—Visser in R.I. 2nd Ser. IV. (1902), pp. 66-87, 159-177, and 270-280—Lehr in
R.I. 2nd Ser. XII. (1910), pp. 657-668—Shepheard in The Journal of the Society of
Comparative Legislation, New Series, III. (1901), pp. 231-237, and V. (1903), pp.
132-136—Oppenheim in The Law Quarterly Review, XXIV. (1908), pp. 328-334.
Commercial Treaties in General.
§ 578. Commercial treaties are treaties concerning the commerce
and navigation of the contracting States and concerning the subjects
Declarations concerning the status quo in the Mediterranean, but
neither
[950]
of them comprises a real legal guarantee.
[950] Whereas Quabbe (p. 97, note 1), correctly denies the character of a real
guarantee to the Declarations concerning the Mediterranean, he (p. 105) considers the
Declarations concerning the North Sea and the Baltic real Guarantee Treaties.
Treaties of Protection.
§ 577. Different from guarantee treaties are treaties of protection.
Whereas the former constitute the guarantee of a certain object to
the guaranteed, treaties of protection are treaties by which strong
States simply engage to protect weaker States without any
guarantee whatever. A treaty of protection must, however, not be
confounded with a treaty of protectorate.
[951]
[951] See above, § 92.
IV
COMMERCIAL TREATIES
Taylor, 354—Moore, V. §§ 765-769—Melle in Holtzendorff, III. pp. 143-256—Liszt, §
28—Ullmann, § 145—Bonfils, No. 918—Despagnet, No. 462—Pradier-Fodéré, IV.
Nos. 2005-2033—Mérignhac, II. pp. 688-693—Rivier, I. pp. 370-374—Fiore, II.
Nos. 1065-1077, and Code, Nos. 848-854—Martens, II. §§ 52-55—Steck, "Versuch
über Handels- und Schiffahrtsverträge" (1782)—Schraut, "System der
Handelsverträge und der Meistbegünstigung" (1884)—Veillcovitch, "Les traités de
commerce" (1892)—Nys, "Les origines du droit international" (1894), pp. 278-294
—Herod, "Favoured Nation Treatment" (1901)—Calwer, "Die Meistbegünstigung in
den Vereinigten Staaten von Nord-America" (1902)—Glier, "Die
Meistbegünstigungs-Klausel" (1906)—Cavaretta, "La clausola della natiozione più
favorita" (1906)—Barclay, "Problems of International Law and Diplomacy" (1907),
pp. 137-142—Hornbeck, "The Most-Favoured Nation Clause" (1910), and in A.J.
III. (1909), pp. 394-422, 619-647, and 798-827—Lehr in R.I. XXV. (1893), pp. 313-
316—Visser in R.I. 2nd Ser. IV. (1902), pp. 66-87, 159-177, and 270-280—Lehr in
R.I. 2nd Ser. XII. (1910), pp. 657-668—Shepheard in The Journal of the Society of
Comparative Legislation, New Series, III. (1901), pp. 231-237, and V. (1903), pp.
132-136—Oppenheim in The Law Quarterly Review, XXIV. (1908), pp. 328-334.
Commercial Treaties in General.
§ 578. Commercial treaties are treaties concerning the commerce
and navigation of the contracting States and concerning the subjects
of these States who are engaged in commerce and navigation.
Incidentally, however, they also contain clauses concerning consuls
and various other matters. They are concluded either for a limited or
an unlimited number of years, and either for the whole territory of
one or either party or only for a part of such territory—e.g., by Great
Britain for the United Kingdom alone, or for Canada alone, and the
like. All full-Sovereign States are competent to enter into commercial
treaties, but it depends upon the special case whether half- and
part-Sovereign States are likewise competent. Although competent
to enter upon commercial treaties, a State may, by an international
compact, be restricted in its freedom with regard to its commercial
policy. Thus, according to articles 1 to 5 of the General Act of the
Berlin Congo Conference of February 26, 1885, all the Powers which
have possessions in the Congo district must grant complete freedom
of commerce to all nations. Again, to give another example, France
and Germany are by article 11 of the Peace of Frankfort of May 10,
1871, compelled to grant one another most-favoured-nation
treatment in their commercial relations, in so far as favours which
they grant to Great Britain, Belgium, Holland, Switzerland, Austria,
and Russia are concerned.
The details of commercial treaties are for the most part purely
technical and are, therefore, outside the scope of a general treatise
on International Law. There are, however, two points of great
importance which require discussion—namely, the meaning of
coasting trade and of the most-favoured-nation clause.
Meaning of Coasting Trade in Commercial Treaties.
§ 579. The meaning of the term coasting-trade
[952]
in commercial
treaties must not be confounded with its meaning in International
Law generally. The meaning of the term in International Law
becomes apparent through its synonym cabotage—that is,
navigation from cape to cape along the coast combined with trading
between the ports of the coast concerned without going out into the
Open Sea. Therefore, trade between Marseilles and Nice, between
Calais and Havre, between London and Liverpool, and between
Incidentally, however, they also contain clauses concerning consuls
and various other matters. They are concluded either for a limited or
an unlimited number of years, and either for the whole territory of
one or either party or only for a part of such territory—e.g., by Great
Britain for the United Kingdom alone, or for Canada alone, and the
like. All full-Sovereign States are competent to enter into commercial
treaties, but it depends upon the special case whether half- and
part-Sovereign States are likewise competent. Although competent
to enter upon commercial treaties, a State may, by an international
compact, be restricted in its freedom with regard to its commercial
policy. Thus, according to articles 1 to 5 of the General Act of the
Berlin Congo Conference of February 26, 1885, all the Powers which
have possessions in the Congo district must grant complete freedom
of commerce to all nations. Again, to give another example, France
and Germany are by article 11 of the Peace of Frankfort of May 10,
1871, compelled to grant one another most-favoured-nation
treatment in their commercial relations, in so far as favours which
they grant to Great Britain, Belgium, Holland, Switzerland, Austria,
and Russia are concerned.
The details of commercial treaties are for the most part purely
technical and are, therefore, outside the scope of a general treatise
on International Law. There are, however, two points of great
importance which require discussion—namely, the meaning of
coasting trade and of the most-favoured-nation clause.
Meaning of Coasting Trade in Commercial Treaties.
§ 579. The meaning of the term coasting-trade
[952]
in commercial
treaties must not be confounded with its meaning in International
Law generally. The meaning of the term in International Law
becomes apparent through its synonym cabotage—that is,
navigation from cape to cape along the coast combined with trading
between the ports of the coast concerned without going out into the
Open Sea. Therefore, trade between Marseilles and Nice, between
Calais and Havre, between London and Liverpool, and between
Dublin and Belfast is coasting-trade, but trade between Marseilles
and Havre, and between London and Dublin is not. It is a universally
recognised rule
[953]
of International Law that every littoral State can
exclude foreign merchantmen from the cabotage within its maritime
belt. Cabotage is the contrast to the over-sea
[954]
carrying trade, and
has nothing to do with the question of free trade from or to a port
on the coast to or from a port abroad. This question is one of
commercial policy, and International Law does not prevent a State
from restricting to vessels of its subjects the export from or the
import to its ports, or from allowing such export or import under
certain conditions only.
and Havre, and between London and Dublin is not. It is a universally
recognised rule
[953]
of International Law that every littoral State can
exclude foreign merchantmen from the cabotage within its maritime
belt. Cabotage is the contrast to the over-sea
[954]
carrying trade, and
has nothing to do with the question of free trade from or to a port
on the coast to or from a port abroad. This question is one of
commercial policy, and International Law does not prevent a State
from restricting to vessels of its subjects the export from or the
import to its ports, or from allowing such export or import under
certain conditions only.
[952] See Oppenheim in The Law Quarterly Review, XXIV. (1908), pp. 328-334.
[953] See above, § 187.
[954] It must be emphasised that navigation and trade from abroad to several ports of
the same coast successively—for instance, from Dover to Calais and then to Havre—is
not coasting-trade but over-sea trade, provided that all the passengers and cargo are
shipped from abroad.
There is no doubt that originally the meaning of coasting-trade in
commercial treaties was identical with its meaning in International
Law generally, but there is likewise no doubt that the practice of the
States gives now a much more extended meaning to the term
coasting-trade as used in commercial treaties. Thus France
distinguishes between cabotage petit and grand; whereas petit
cabotage is coasting-trade between ports in the same sea, grand
cabotage is coasting-trade between a French port situated in the
Atlantic Ocean and a French port situated in the Mediterranean, and
—according to a statute of September 21, 1793—both grand and
petit cabotage are exclusively reserved for French vessels. Thus,
further, the United States of America has always considered trade
between one of her ports in the Atlantic Ocean and one in the Pacific
to be coasting-trade, and has exclusively reserved it for vessels of
her own subjects; she considers such trade coasting-trade even
when the carriage takes place not exclusively by sea around Cape
Horn, but partly by sea and partly by land through the Isthmus of
Panama. Great Britain has taken up a similar attitude. Section 2 of
the Navigation Act of 1849 (12 & 13 Vict. c. 29) enacted "that no
goods or passengers shall be carried coastwise from one part of the
United Kingdom to another, or from the Isle of Man to the United
Kingdom, except in British ships," and thereby declared trade
between a port of England or Scotland to a port of Ireland or the
Isle of Man to be coasting-trade exclusively reserved for British ships
in spite of the fact that the Open Sea flows between these ports.
And although the Navigation Act of 1849 is no longer in force, and
this country now does admit foreign ships to its coasting-trade, it
nevertheless still considers all trade between one port of the United
Kingdom and another to be coasting-trade, as becomes apparent
from Section 140 of the Customs Laws Consolidation Act of July 24,
[953] See above, § 187.
[954] It must be emphasised that navigation and trade from abroad to several ports of
the same coast successively—for instance, from Dover to Calais and then to Havre—is
not coasting-trade but over-sea trade, provided that all the passengers and cargo are
shipped from abroad.
There is no doubt that originally the meaning of coasting-trade in
commercial treaties was identical with its meaning in International
Law generally, but there is likewise no doubt that the practice of the
States gives now a much more extended meaning to the term
coasting-trade as used in commercial treaties. Thus France
distinguishes between cabotage petit and grand; whereas petit
cabotage is coasting-trade between ports in the same sea, grand
cabotage is coasting-trade between a French port situated in the
Atlantic Ocean and a French port situated in the Mediterranean, and
—according to a statute of September 21, 1793—both grand and
petit cabotage are exclusively reserved for French vessels. Thus,
further, the United States of America has always considered trade
between one of her ports in the Atlantic Ocean and one in the Pacific
to be coasting-trade, and has exclusively reserved it for vessels of
her own subjects; she considers such trade coasting-trade even
when the carriage takes place not exclusively by sea around Cape
Horn, but partly by sea and partly by land through the Isthmus of
Panama. Great Britain has taken up a similar attitude. Section 2 of
the Navigation Act of 1849 (12 & 13 Vict. c. 29) enacted "that no
goods or passengers shall be carried coastwise from one part of the
United Kingdom to another, or from the Isle of Man to the United
Kingdom, except in British ships," and thereby declared trade
between a port of England or Scotland to a port of Ireland or the
Isle of Man to be coasting-trade exclusively reserved for British ships
in spite of the fact that the Open Sea flows between these ports.
And although the Navigation Act of 1849 is no longer in force, and
this country now does admit foreign ships to its coasting-trade, it
nevertheless still considers all trade between one port of the United
Kingdom and another to be coasting-trade, as becomes apparent
from Section 140 of the Customs Laws Consolidation Act of July 24,
1876 (39 & 40 Vict. c. 36). Again, Germany declared by a statute of
May 22, 1881, coasting-trade to be trade between any two German
ports, and reserved it for German vessels, although vessels of such
States can be admitted as on their part admit German vessels to
their own coasting-trade. Thus trade between Koenigsberg in the
Baltic and Hamburg in the North Sea is coasting-trade.
These instances are sufficient to demonstrate that an extension of
the original meaning of coasting-trade has really taken place and has
found general recognition. A great many commercial treaties have
been concluded between such countries as established that
extension of meaning and others, and these commercial treaties no
doubt make use of the term coasting-trade in this its extended
meaning. It must, therefore, be maintained that the term coasting-
trade or cabotage as used in commercial treaties has acquired the
following meaning: Sea-trade between any two ports of the same
country whether on the same coast or different coasts, provided
always that the different coasts are all of them the coasts of one and
the same country as a political and geographical unit in
contradistinction to the coasts of Colonial dependencies of such
country.
In spite of this established extension of the term coasting-trade, it
did not include colonial trade until nearly the end of the nineteenth
century.
[955]
Indeed, when Russia, by ukase of 1897, enacted that
trade between any of her ports should be considered coasting trade
and be reserved for Russian vessels, this did not comprise a further
extension of the conception of coasting-trade. The reason is that
Russia, although her territory extends over different parts of the
globe, is a political and geographical unit, and there is one stretch of
territory only between St. Petersburg and Vladivostock. But when, in
1898 and 1899, the United States of America declared trade
between any of her ports and those of Porto Rico, the Philippines,
and the Hawaiian Islands to be coasting-trade, and consequently
reserved it exclusively for American vessels, the distinction between
coasting-trade and over-sea or colonial trade fell to the ground. It is
submitted that this American extension of the conception of
May 22, 1881, coasting-trade to be trade between any two German
ports, and reserved it for German vessels, although vessels of such
States can be admitted as on their part admit German vessels to
their own coasting-trade. Thus trade between Koenigsberg in the
Baltic and Hamburg in the North Sea is coasting-trade.
These instances are sufficient to demonstrate that an extension of
the original meaning of coasting-trade has really taken place and has
found general recognition. A great many commercial treaties have
been concluded between such countries as established that
extension of meaning and others, and these commercial treaties no
doubt make use of the term coasting-trade in this its extended
meaning. It must, therefore, be maintained that the term coasting-
trade or cabotage as used in commercial treaties has acquired the
following meaning: Sea-trade between any two ports of the same
country whether on the same coast or different coasts, provided
always that the different coasts are all of them the coasts of one and
the same country as a political and geographical unit in
contradistinction to the coasts of Colonial dependencies of such
country.
In spite of this established extension of the term coasting-trade, it
did not include colonial trade until nearly the end of the nineteenth
century.
[955]
Indeed, when Russia, by ukase of 1897, enacted that
trade between any of her ports should be considered coasting trade
and be reserved for Russian vessels, this did not comprise a further
extension of the conception of coasting-trade. The reason is that
Russia, although her territory extends over different parts of the
globe, is a political and geographical unit, and there is one stretch of
territory only between St. Petersburg and Vladivostock. But when, in
1898 and 1899, the United States of America declared trade
between any of her ports and those of Porto Rico, the Philippines,
and the Hawaiian Islands to be coasting-trade, and consequently
reserved it exclusively for American vessels, the distinction between
coasting-trade and over-sea or colonial trade fell to the ground. It is
submitted that this American extension of the conception of
coasting-trade as used in her commercial treaties before 1898 is
inadmissible
[956]
and contains a violation of the treaty rights of the
other contracting parties. Should these parties consent to the
American extension of the meaning of coasting-trade, and should
other countries follow the American lead and apply the term
coasting-trade indiscriminately to trade along their coasts and to
their colonial trade, the meaning of the term would then become
trade between any two ports which are under the sovereignty of the
same State. The distinction between coasting-trade and colonial
trade would then become void, and the last trace of the synonymity
between coasting-trade and cabotage would have disappeared.
[955] See details in Oppenheim, loc. cit. pp. 331-332, but it is of value to draw
attention here to a French statute of April 2, 1889. Whereas a statute of April 9, 1866,
had thrown open the trade between France and Algeria to vessels of all nations, article 1
of the statute of April 2, 1889, enacts: La navigation entre la France et l'Algérie ne
pourra s'effectuer que sous pavillon français. This French statute does not, as is
frequently maintained, declare the trade between France and Algeria to be coasting-
trade, but it nevertheless reserves such trade exclusively for French vessels. The French
Government, in bringing the bill before the French Parliament, explained that the statute
could not come into force before February 1, 1892, because art. 2 of the treaty with
Belgium of May 14, 1882, and art. 21 of the treaty with Spain of February 6, 1882—both
treaties to expire on February 1, 1892—stipulated the same treatment for Belgian and
Spanish as for French vessels, cabotage excepted. It is quite apparent that, if France had
declared trade between French and Algerian ports to be coasting-trade in the meaning of
her commercial treaties, the expiration of the treaties with Belgium and Spain need not
have been awaited for putting the law of April 2, 1889, into force.
[956] In the case of Huus v. New York and Porto Rico Steamship Co. (1901), 182
United States 392, the Court was compelled to confirm the extension of the term
coasting-trade to trade between any American port and Porto Rico, because this
extension was recognised by section 9 of the Porto Rican Act, and because in case of a
conflict between Municipal and International Law—see above, § 21—the Courts are
bound to apply their Municipal Law.
Meaning of most-favoured-nation Clause.
§ 580. Most of the commercial treaties of the nineteenth century
contain a stipulation which is characterised as the most-favoured-
nation clause. The wording of this clause is by no means the same in
all treaties, and its general form has therefore to be distinguished
from several others which are more specialised in their wording.
According to the most-favoured-nation clause in its general form, all
favours which either contracting party has granted in the past or will
inadmissible
[956]
and contains a violation of the treaty rights of the
other contracting parties. Should these parties consent to the
American extension of the meaning of coasting-trade, and should
other countries follow the American lead and apply the term
coasting-trade indiscriminately to trade along their coasts and to
their colonial trade, the meaning of the term would then become
trade between any two ports which are under the sovereignty of the
same State. The distinction between coasting-trade and colonial
trade would then become void, and the last trace of the synonymity
between coasting-trade and cabotage would have disappeared.
[955] See details in Oppenheim, loc. cit. pp. 331-332, but it is of value to draw
attention here to a French statute of April 2, 1889. Whereas a statute of April 9, 1866,
had thrown open the trade between France and Algeria to vessels of all nations, article 1
of the statute of April 2, 1889, enacts: La navigation entre la France et l'Algérie ne
pourra s'effectuer que sous pavillon français. This French statute does not, as is
frequently maintained, declare the trade between France and Algeria to be coasting-
trade, but it nevertheless reserves such trade exclusively for French vessels. The French
Government, in bringing the bill before the French Parliament, explained that the statute
could not come into force before February 1, 1892, because art. 2 of the treaty with
Belgium of May 14, 1882, and art. 21 of the treaty with Spain of February 6, 1882—both
treaties to expire on February 1, 1892—stipulated the same treatment for Belgian and
Spanish as for French vessels, cabotage excepted. It is quite apparent that, if France had
declared trade between French and Algerian ports to be coasting-trade in the meaning of
her commercial treaties, the expiration of the treaties with Belgium and Spain need not
have been awaited for putting the law of April 2, 1889, into force.
[956] In the case of Huus v. New York and Porto Rico Steamship Co. (1901), 182
United States 392, the Court was compelled to confirm the extension of the term
coasting-trade to trade between any American port and Porto Rico, because this
extension was recognised by section 9 of the Porto Rican Act, and because in case of a
conflict between Municipal and International Law—see above, § 21—the Courts are
bound to apply their Municipal Law.
Meaning of most-favoured-nation Clause.
§ 580. Most of the commercial treaties of the nineteenth century
contain a stipulation which is characterised as the most-favoured-
nation clause. The wording of this clause is by no means the same in
all treaties, and its general form has therefore to be distinguished
from several others which are more specialised in their wording.
According to the most-favoured-nation clause in its general form, all
favours which either contracting party has granted in the past or will
grant in the future to any third State must be granted to the other
party. But the real meaning of this clause in its general form has
ever been controverted since the United States of America entered
into the Family of Nations and began to conclude commercial
treaties embodying the clause. Whereas in former times the clause
was considered obviously to have the effect of causing all favours
granted to any one State at once and unconditionally to accrue to all
other States having most-favoured-nation treaties with the grantor,
the United States contended that these favours could accrue to such
of the other States only as fulfilled the same conditions under which
these favours had been allowed to the grantee. The majority of the
commercial treaties of the United States, therefore, do not contain
the most-favoured-nation clause in its general form, but in what is
called its conditional, qualified, or reciprocal, form. In this form it
stipulates that all favours granted to third States shall accrue to the
other party unconditionally, in case the favours have been allowed
unconditionally to the grantee, but only under the same
compensation, in case they have been granted conditionally. The
United States, however, has always upheld the opinion, and the
supreme Court of the United States has confirmed
[957]
this
interpretation, that, even if a commercial treaty contains the clause
in its general, and not in its qualified, form, it must always be
interpreted as though it were worded in its qualified form.
[957] See Bartram v. Robertson, 122 United States 116, and Whitney v. Robertson, 124
United States 190.
Now nobody doubts that according to the qualified form of the
clause a favour granted to any State can only accrue to other States
having most-favoured-nation treaties with the grantor, provided they
fulfil the same conditions and offer the same compensations as the
grantee. Again, nobody doubts that, if the clause is worded in its so-
called unconditional form stipulating the accrument of a favour to
other States whether it was allowed to the grantee gratuitously or
conditionally against compensation, all favours granted to any State
accrue immediately and without condition to all the other States.
However, as regards the clause in its general form, what might,
broadly speaking, be called the European is confronted by the
party. But the real meaning of this clause in its general form has
ever been controverted since the United States of America entered
into the Family of Nations and began to conclude commercial
treaties embodying the clause. Whereas in former times the clause
was considered obviously to have the effect of causing all favours
granted to any one State at once and unconditionally to accrue to all
other States having most-favoured-nation treaties with the grantor,
the United States contended that these favours could accrue to such
of the other States only as fulfilled the same conditions under which
these favours had been allowed to the grantee. The majority of the
commercial treaties of the United States, therefore, do not contain
the most-favoured-nation clause in its general form, but in what is
called its conditional, qualified, or reciprocal, form. In this form it
stipulates that all favours granted to third States shall accrue to the
other party unconditionally, in case the favours have been allowed
unconditionally to the grantee, but only under the same
compensation, in case they have been granted conditionally. The
United States, however, has always upheld the opinion, and the
supreme Court of the United States has confirmed
[957]
this
interpretation, that, even if a commercial treaty contains the clause
in its general, and not in its qualified, form, it must always be
interpreted as though it were worded in its qualified form.
[957] See Bartram v. Robertson, 122 United States 116, and Whitney v. Robertson, 124
United States 190.
Now nobody doubts that according to the qualified form of the
clause a favour granted to any State can only accrue to other States
having most-favoured-nation treaties with the grantor, provided they
fulfil the same conditions and offer the same compensations as the
grantee. Again, nobody doubts that, if the clause is worded in its so-
called unconditional form stipulating the accrument of a favour to
other States whether it was allowed to the grantee gratuitously or
conditionally against compensation, all favours granted to any State
accrue immediately and without condition to all the other States.
However, as regards the clause in its general form, what might,
broadly speaking, be called the European is confronted by the
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knowledge seekers. We believe that every book holds a new world,
offering opportunities for learning, discovery, and personal growth.
That’s why we are dedicated to bringing you a diverse collection of
books, ranging from classic literature and specialized publications to
self-development guides and children's books.
More than just a book-buying platform, we strive to be a bridge
connecting you with timeless cultural and intellectual values. With an
elegant, user-friendly interface and a smart search system, you can
quickly find the books that best suit your interests. Additionally,
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