554256626-tissue-engineering-final-ppt.ppt

This is a full presentation about the Tissue Engineering

1. SUPERVISED BY:-
DR.GEETA RAJPUT
DEPT OF PROSTHODONTICS
PRESENTED BY
SARAH ASIF(JR1)
(DEPT OF ORTHODONTICS)
CO-SUPERVISED BY:-
DR.ABHINAV
DEPT OF PROSTHODONTICS

2. DEFINITION
HISTORY
REQUIREMENTS OF TISSUE ENGINEERING
APPLICATIONS
BIOMATERIALS
NANOTECHNOLOGY AND TISSUE
ENGINEERING
CHALLENGES
THE FUTURE

3. Creation of a functional biological substitute using living cells and
a matrix to maintain, improve or restore damage to tissues and
organs (Atala, A. Engineering tissues, organs and cells. 2007 J
Tissue Eng Regen Med 1: 83-96)
Bringing together the fields of medicine, biology, engineering and
biotechnology

4. Tissue Engineering is an interdisciplinary field
that applies the principles of engineering and the
life sciences towards the development of
biological substitutes that
RESTORE
MAINTAIN
IMPROVE TISSUE FUNCTION.

5. Prior to 1950 references to the concept of tissue
engineering may be found in the literature.
In the 1970s and 1980s research on what we now call
tissue engineering emerged.
In 1987 the term tissue engineering was coined.
In 1988 the first meeting called tissue engineering was
held at Lake Tahoe.
In the 1990s research accelerated and an industry
began to emerge.

6. Failing tissues and organs.
Shortfalls of current options Autologous
tissues ,Allogeneic tissues, Xenogeneic tissues,
Synthetic materials.
To create products that improve tissue function or heal
tissue defects.
Replace diseased or damaged tissue.
Because donor tissues and organs are in short supply;we
want to minimize immune system response by using our
own cells or novel ways to protect transplant.

7. Tissue from the same patient
The ideal option
Biocompatible
No immune response
Decreasing availability of healthy tissue
from patients with disease

8. Tissue from another person
More practical than autologous harvest
Severe shortage of donors
Increased immune response to foreign material
Immunosuppressant drugs required
Unpleasant side effects
Expensive

9. Tissue harvested from animals
Potentially readily available
Immune response from host to foreign material
Risk of disease transmission from animal to human
Significant ethical considerations

10. Biocompatible
Made with the patient’s own cells
Engineered to fill the exact role required
Degradation rate,
Composition,
Size,
Mechanical properties
Off-the-shelf availability

11. Remove cells from the
body.
Expand number in culture
Seed onto an appropriate
scaffold with suitable growth
factors and cytokines
Place into culture
Re-implant engineered
tissue repair damaged
site

12. Tissue
Engineering
Cells Scaffolds
Biorectors Signals
Differentiated cells
Adult stem cells
Embryonic stem cells
Dynamic cell seeding
Improved mass transfer
Mechanical stimuli
Hydrogels
Nanofibrous scaffolds
Self-assembling scaffolds
Solid freeform fabricated scaffolds
Small molecules
Growth factors/polypeptides
Nucleic acids (DNA, RNA and
antisense oligonucleotides)

14. Stem cells are immature, unspecialized cells in the
body that are able to grow into specialized cell
types by a process known as “differentiation.”

There are two primary sources of stem cells:
Embryonic stem cells and Adult stem cells
Adult stem cells are found in many tissues
including dental pulp .

15. Two sources
◦Fertilized egg from in vitro fertilization
◦Ovum that has had nucleus removed and nuclear
material injected from intended recipient of final tissue
product
(reproductive/therapeutic cloning)

16. Found in:
◦Umbilical cord blood/tissue
◦Adult brain,
◦Blood cornea,
◦Retina,
◦Heart,
◦Fat,
◦Skin,
◦Dental pulp,
◦Bone marrow,
◦Blood vessels, Skeletal muscle and Intestines

17. Stem cells:
 Undifferentiated cells
Develop
 into differentiated cells
with distinctive
 features and
functions
Ameloblasts Odontoblasts
Enamel dentin
Christopherson and Nesti Stem Cell Research &
Therapy 2011 
Dental tissue
formation

18. The recent discovery of dental stem cells in 2000 by a
researcher at the National Institutes of Health – USA ,
has developed a wide range of possiblities in the
regeneration of the different tissues and organs.
Researches have been carried out the past years
evolving a new branch in dentistry known as
“ regenerative dentistry “
18

19. It is now known that adult stem cells taken from
one area of the body can be transplanted into
another area and grown into a completely different
type of tissue.
Dental stem cells are being studied for a wide
range of diseases due to their ability to form
connective , neural , muscle , bone and dental
tissues .
19

20. 2000:
 
Dental pulp stem cells discovered by a researcher at
the National Institutes of Health.
2003: 
National Institutes of Health announces viable stem
cells are in dental pulp of teeth.
2004 to present:
 
Over 1,000 published studies identifying
therapeutic potential of dental mesenchymal stem cells.
2008: 
Surgeons from Spain announced the world’s first
tissue-engineered whole organ transplant procedure, using
a trachea made with the patient’s own adult mesenchymal
stem cells.

21. 2009:
 
Scientists from Italy announced the first-ever
human clinical application using patients’ own
 dental
stem cells
 to repair mandibular bone defects.
2011 : a study showed that stem cells from teeth can
create islet- like cells which produce insulin ( a potential
therapy for type 1 diabetes ) .
21

22. Dental stem cells are present in various dental
tissues-;

Including both Deciduous and Permanent teeth
Periodontal ligament
Apical papilla
Dental follicle

23. Dental Pulp Stem Cells (DPSCs)
Stem cells from human exfoliated
deciduous teeth (SHEDs)
Periodontal Ligament (PDLSCs)
Dental follicle stem cells (DFSCs)
Apical papilla (SCAPs)

24. Multipotent cells
High proliferation rates
Accessibility
Generate dentin complex
www.newsbrown.edu

26. Dental Pulp SC -Generate bone, dentin complex
-Repair damaged dental tissues

-Induce bone regeneration
Periodontal ligament SC
-Form cementum & alveolar bone

-Cures periodontal lesions in pig
Dental follicle SC -Forms cementum in vivo
SC from apical papilla -Osteoblasts & odondoblasts
-Regenerative endodontic therapy

27. Dental pulp stem cells potential-;

Stem cells derived from the dental pulp can form pulp like
tissue , in future it is possible to replace infected pulp
tissue of a paining tooth with newly generated pulp like
tissue instead of doing RCT ,
Thus preserving the vitality of the tooth “regenerative
endodotics “.
 It also has the ability to form bone that is useful for
the osseointegration of dental implants, thus increaseing
its success rate.

28. Stem cells of Human Exfoliated Deciduous Teeth-
Have higher rate of proliferation.

Have potential to form bone which is useful during
Osseointegration of dental implants.
Have the potential to repair calvarial defects in
immunocompromised mice.

29. Periodontal Ligament Stem Cells-;
Have potentials of regenerating typical cementum
and periodontal ligament like structure .

Tissue of the periodontium made by stem cell can
be used as a treatment modality to replace the
diseased periodontium around teeth so as
treatment to mobility of teeth.

30. Stem Cells from Apical Papilla-;

Can only be isolated at certain specific stages of
the development of tooth.
Dental papilla contain higher number of adult
stem cells than mature dental pulp, thus have a
greater potential for regenerating dentin than
DPSCs.

31. Dental Follicle Precursor Cells -;

Dental follicles contain progenitor cells which have
the capability of differentiating into
Cementum forming cells (cementoblasts),
Osteoblasts of the alveolar bone,
Periodontal ligament fibroblasts.

32. Stem cells can be identified and isolated from mixed cell
populations by four commonly used techniques:
Staining the cells with specific antibody markers and
using a flow cytometer, in a process called fluorescent
antibody cell sorting (FACS);
Immunomagnetic bead selection;
Immunohistochemical staining;
Physiological and histological criteria, including
phenotype (appearance), chemotaxis, proliferation,
differentiation, and mineralizing activity.

33. FACS together with the protein marker CD34 is widely
used to separate human stem cells expressing CD34 from
peripheral blood, umbilical cord blood, and cell cultures.

The most studied dental stem cells are those of the dental
pulp.

Human pulp stem cells express von Willebrand factor
CD146, alpha-smooth muscle actin, and 3G5 proteins.
Human pulp stem cells also have a fibroblast phenoptype,
with specific proliferation, differentiation, and mineralizing
activity pattern.

34. More research required
DFSCs and SCAPs isolation: needs 3
rd
molar
Use of embryonic stem cells
Not ethical, not easily applicable
Expensive

35. Allow cell attachment and migration
Deliver and retain cells and biochemical factors
Enable diffusion of vital cell nutrients and
expressed products.
Exert certain mechanical and biological
influences to modify the behaviour of the cell
phase.

36. Temporary structural support
◦Maintain shape
Cellular microenvironment
◦High surface area/volume
◦ECM secretion
◦Integrin expression
◦Facilitate cell migration
Surface
coating
Structural

37. 3-dimensional
Cross-linked
Porous
Biodegradable
Proper surface chemistry
Matching mechanical strength
Biocompatible
Promotes natural healing
Accessibility
Commercial Feasibility

38. Cell attachment & migration
Permit delivery of growth factors
Enable influx of oxygen
Porosity is critical
Degradation is fundamental
Should not be toxic
Ceramics, natural or synthetic
polymers

39. Surface chemistry
Matrix topography
◦Cell organization, alignment
◦Fiber alignment -> tissue development
Rigidity
◦5-23 kPa
Porosity
◦Large interconnected
◦small disconnected

40. collagen
Hydroxyapatite

3Dhydrogel
Porous ceramics

41. Polymers
◦Collagen
◦Laminin
◦Fibrin
◦Matrigel
◦Decellularized matrix
Ceramics
◦Hydroxyapatite
◦Calcium phosphate
◦Bioglass

42. Growth factors directing cellular activity
Designed to expose cells to physical stimuli and/or
maintain desired conditions.
Growth factors are proteins that bind to receptors on the
cell and induce cellular proliferation and/or differentiation.
Many growth factors are quite versatile, stimulating
cellular division in numerous cell types, while others are
more cell specific.

44. Growth factors, especially those of the transforming
growth factor beta (TGF) family, are important in cellular
signaling for odontoblast differentiation and stimulation
of dentin matrix secretion.

These growth factors are secreted by odontoblasts and
deposited within the dentin matrix , where they remain
protected in an active form through interaction with other
components of the dentin matrix.

The addition of purified dentin protein fractions has
stimulated an increase in tertiary dentin matrix secretion.

45. Another important family of growth factors in tooth development and regeneration
consists of the bone morphogenic protein (BMPs) .
Recombinant human BMP2 stimulates differentiation of adult pulp stem cells into an
odontoblastoid morphology in culture
 Recombinant BMP-2, -4, and -7 induce formation of reparative dentin in vivo.
The application of recombinant human insulin-like growth factor-1 together with
collagen has been found to induce complete dentin bridging and tubular dentin
formation .
This indicates the potential of adding growth factors before pulp capping, or
incorporating them into restorative and endodontic materials to stimulate dentin and
pulp regeneration.
In the longer term, growth factors will likely be used in conjunction with postnatal
stem cells to accomplish the tissue engineering replacement of diseased tooth pulp.

46. In vitro differentiation
◦Construct tissues outside body before transplantation
◦Ultimate goal
Most economical
Least waiting time
In situ methodology
◦Host remodeling of environment
Ex vivo approach
◦Excision and remodeling in culture

48. ADVANTAGES
This approach seeks to overcome the problem
of limited implant lifetimes by regenerating
tissue in culture.
Alleviates problem of donor shortage by using
a few culture cells to regenerate entire tissue

49. DISADVANTAGES
May face same rejection issues as
transplanted tissues
Long term stability of tissues generated in this
manner is still unknown
Approaches to generate complex tissue types
still very experimental

50. ADVANTAGES
Seeks to solve all of the potential problems of
the other approaches
The body’s own cells or growth factors are used for
regeneration, so no autograft of transplanted tissue
is needed
The implant will regenerate natural tissue and the
components of the guiding scaffold are
biodegradable so long term stability is not an issue

51. DISADVANTAGES

Faithful regeneration of complex tissue is an unsolved
problem
In vivo synthesis of skin successfully recovers the
overall structure of the organ but fails to regenerate
organelles such as the sweat glands and hair follicles
Proper scaffolds for many cell types (e.g. nerve) have not
been fully developed yet, primarily because the biological
cues necessary for regeneration are not well understood

52. FIRST APPROACH
Seeding of cells into various gels (collagen gels, fibrin
and other component)
The true integration of cells into gels allow them to
reorganize the surrounding matrix.

53. FIRST APPROACH
Drawback: Week mechanical resistance of the obtained
substitutes. The structural integrity may be sufficient for skin,
but not so for substitutes in vascular or orthopaedic systems
This problem has been addressed with glycation and
magnetic alignment of the collagen fibres

54. SECOND APPROACH
The seeding of cells into scaffolds
The cells thrive in the porous material and secrete
various amount of extracellular matrix depending on
their nature. (developed by Robert Langer’s group MIT)

55. Advantage: of the scaffold approach is the immediate
creation of a three-dimensional structure that already
has significant structural properties
Drawback: The intrinsic nature of most of these
polymers, which are suture materials, entails slow
degradation with an ensuing lowering of pH of
surrounding tissues. This leads to a slow but rather
protracted low-level inflammatory process.

56. THIRD APPROACH
In this approach various types of cells, mostly of
mesencymal origin, are grown in such a fashion within a
culture flask that they literally embedded themselves in
their very oven extracellular matrix.
Among many factors, the addition of sodium ascorbate
allows the significant appearance of the various
components of the extracellular matrix.
These sheets are then either stacked or rolled to obtain
various tissue substitutes.

57. Advantage: The absence of extraneous collagens and any
synthetic material
Drawback: This approach is time consuming

58. In surgery
Transplantation of failing tissues/organs
Aiding tissues in the healing process
In the laboratory
Observing immunological, pathological and healing changes
in human tissue without harming patients
Drug therapies: efficacy and side effects of drugs

60. Regenerative endodontics is the creation and delivery of
tissues to replace diseased, missing, and traumatized pulp.
These potential approaches include
Root-canal revascularization,
Postnatal (adult) stem cell therapy,
Pulp implant,
Scaffold implant,
Three-dimensional cell printing,
Injectable scaffolds,
Gene therapy.

61. These regenerative endodontic techniques will possibly
involve some combination of disinfection or debridement
of infected root canal systems with apical enlargement to
permit revascularization and use of adult stem cells,
scaffolds, and growth factors.

64. Cell-free Approaches
An alternative to transferred stem cells already present in
the patient's body that can be recruited to the site of injury to
stimulate self-healing mechanisms and unlock the innate
powers of regeneration.

Methods of in situ tissue regeneration relying on
endogenous cell homing, functional stimulation, and local
tissue responses hold great promise because recent proof-
of-concept studies document success .

65. Regarding pulp regeneration, this approach involves the
following:
1.Endogenous stem cells from resident tissue, either dental pulp
or the periapical region
2.An injectable, bioactive scaffold, which will eventually undergo
cell-mediated degradation to be replaced with natural
extracellular matrix
3.Potent chemoattractants and growth factors to induce cell
migration, proliferation, and differentiation.
Thus, in situ tissue regeneration of dental pulp follows the
classical tissue engineering approach in contrast to the
”regenerative protocol”

66. Endogenous Stem Cells
In many postnatal tissues, stem cells are present that are
responsible for tissue maintenance during regular turnover as
well as repair after injury.
They reside in a specific stem cell niche, a microenvironment
that regulates their survival, self-renewal, and differentiation.
Upon insult, however, they exit the niche, migrate, proliferate
extensively, and differentiate to regenerate tissue damage.
This holds true also for dental pulp in which localization of
endothelial and mesenchymal stem cell markers has been
shown

67. This includes more reliable methods for the clinician to
distinguish between irreversibly damaged and healthy
pulp tissue, which need to be developed.

In cases of advanced stages of pulpitis or necrosis, the
source of stem cells from the dental pulp is lost.
The recruitment of stem cells from the periapical region or
delivered via the blood stream appears more challenging
but not impossible.

68. Research has focused on two main approaches
involving preparations containing biological mediators to
selectively enhance the cells that populate the
periodontal wound.

The first approach utilized semi-purified preparations
such as enamel matrix derivative and autologous
platelet-rich plasma preparations.

The second approach utilized recombinant growth
factors such as recombinant platelet derived growth
factor- BB and recombinant human basic fibroblast
growth factor, and bone morphogenic protein

69. 1. Enamel matrix derivative
It has been effective in the treatment of infrabony defects
and has been shown to be safe for clinical use.

2. Recombinant human platelet derived growth factor
It is commercially available in combination with a tricalcium
phosphate carrier.
Studies suggest that it is easy to use, requires no barrier
membranes and produced results similar or superior to
other regenerative graft materials.

70. 3. Recombinant human fibroblast growth factor-2
Topical application of fibroblast growth factor-2 into intraosseous
defects in alveolar bone induces significant periodontal tissue
regeneration.
4.Application at implant site preparation
The challenge lies in regenerating adequate volume of hard and
possibly soft tissue.
Recombinant human platelet derived growth factor and
Recombinant human bone morphogenic protein-2 may be used
for implant site preparation..

71. Periodontal ligament stem cells from human permanent
teeth (PePDLSCs) have been investigated extensively in
periodontal tissue engineering and regeneration.
However, little knowledge is available on the periodontal
ligament stem cells from human retained deciduous
teeth (DePDLSCs). This study evaluated the potential of
DePDLSCs in periodontal tissue regeneration.

72. DePDLSCs presented a higher proliferation rate and colony-forming
capacity than PePDLSCs in vitro.
During the osteogenic induction, alkaline phosphatase (ALP)
activity, mineralized matrix formation and expression of
mineralization-related genes, including runt-related transcription
factor 2 (RUNX2), ALP, collagen type I (COLI) and osteocalcin
(OCN) were significantly enhanced in DePDLSCs compared with
PePDLSCs.
 Furthermore, DePDLSC cell sheets showed a stronger synthesis of
collagen type I in the extracellular matrix than did PePDLSC cell
sheets.
 After in vivo transplantation, DePDLSC cell sheets recombined with
human dentin blocks were able to generate new
cementum/periodontal ligament-like tissues.

73. A biomaterial is a nonviable material used in a
medical device, intended to interact with
biological systems.
Defined by their application NOT chemical make-
up.
It’s a interdisciplinary field –Bioengineers,material
scientists,immunologists,chemist,biologist and
scientists

75. Physical Requirements
◦Hard Materials.
◦Flexible Material.

Chemical RequirementsChemical Requirements

Must not react with any tissue in the Must not react with any tissue in the
body.body.

Must be non-toxic to the body.Must be non-toxic to the body.

Long-term replacement must not be Long-term replacement must not be
biodegradable.biodegradable.

76. Mechanical
Thermal/Electrical Conductivity
Diffusion
Water Absorption
Biostability
Biocompatibility

77. More than 2000 years ago, Romans, Chinese,
and Aztec’s used gold in dentistry.
Turn of century, synthetic implants become
available.
1937 Poly(methyl methacrylate) (PMMA)
introduced in dentistry.
1958, Rob suggests Dacron Fabrics can be used
to fabricate an arterial prosthetic.

78. 1960 Charnley uses PMMA, ultrahigh-molecular-
weight polyethylend, and stainless steal for total
hip replacement.
Late 1960 – early 1970’s biomaterial field
solidified.
1975 Society for Biomaterials formed.

79. Material Applications
Silicone rubber Catheters, tubing
Dacron Vascular grafts
Cellulose Dialysis membranes
Poly(methyl methacrylate) Intraocular lenses, bone cement
Polyurethanes Catheters, pacemaker leads
Hydogels Opthalmological devices, Drug Delivery
Stainless steel Orthopedic devices, stents
Titanium Orthopedic and dental devices
Alumina Orthopedic and dental devices
Hydroxyapatite Orthopedic and dental devices
Collagen (reprocessed) Opthalmologic applications, wound
dressings

80. Metals
Semiconductor
Materials
Ceramics
Polymers
Synthetic
BIOMATERIALS
Orthopedic
screws/fixation
Dental
Implants
Dental Implants
Heart
valves
Bone
replacements
Biosensors
Implantable
Microelectrod
es
Skin/cartilage
Drug Delivery
Devices
Ocular
implants

81. Specified by physicians using common and
borrowed materials.
Most successes were accidental rather than
by design.
Examples-;
• PMMA dental prosthesis
• steel, gold, ivory, etc., bone plates
• glass eyes and other body parts
• dacron and parachute cloth vascular implants

82. Developed through collaborations of physicians
and engineers.
Engineered implants using common and
borrowed materials.
Built on first generation experiences.
Used advances in materials science (from other
fields).
Examples-;
• titanium alloy dental and orthopaedic implants
• cobalt-chromium-molybdinum orthopaedic implants
• polyethylene bearing surfaces for total joint replacements
• heart valves and pacemakers

83. Bioengineered implants using bioengineered
materials.
Some modified and new polymeric devices.
Many under development.
Example-;
•tissue engineered implants designed to regrow rather than
replace tissues
•Integra LifeSciences artificial skin
•Genzyme cartilage cell procedure
•Resorbable bone repair cements

84. Heart Valve
Artificial Tissue
Dental Implants
Intraocular Lenses
Vascular Grafts
Hip Replacements

85. Thrombosis
Hemolysis
Inflammation
Infection and Sterilization
Carcinogenesis
Hypersensitivity
Systemic Effects

86. Nanofibrous Scaffold
◦Electrospinning
◦Self-Assembly
Nanoporous Scaffold
◦Phase Separation
Carbon Nanotube
Block Copolymer
Printing
◦Nanoimprinting Lithography
◦Organ Printing
◦Contact Printing

87. This process involves the ejection of a charged polymer fluid onto an
oppositely charged surface.
Multiple polymers can be combined
Control over fiber diameter and scaffold architecture

88. This process involves dissolving of a polymer in a solvent at a high
temperature followed by a liquid–liquid or solid–liquid phase
separation induced by lowering the solution temperature
Capable of wide range of geometry and dimensions include pits,
islands, fibers, and irregular pore structures
Simpler than self-assembly
a) powder, b) scaffolds with continuous network, c) foam with closed
pores
SEM of nanofibrous scaffold with interconnected spherical macropores

89. Limited Success: full regeneration of tissues that
do not regenerate spontaneously has not yet
been achieved
◦A lot of success with bone
◦Skin has no glands and hair
◦Engineered cartilage is not articular

90. Unforeseen hurdles in the creation of multicellular
constructs

In the laboratory
Supply of nutrients
Removal of waste
Size
Mechanical stability
In the patient
Availability
Evidence of efficacy
Safety; graft rejection
Cost

91. Micromechanical effects
◦Cell differentiation and growth (especially in
load-bearing tissues) can be affected by
micromechanical stresses transmitted by the
scaffold
Cell function deterioration
Cross-application to other areas (gene therapy,
drug delivery)
Multicellular tissues and organs
◦Complex, multicomponent structures
(vascularized tissues)
◦Regeneration-inducing factors (proteins)

92. To more closely replicate complex tissue
architecture and arrangement in vitro
To better understand extracellular and
intracellular modulators of cell function
To develop novel materials and processing
techniques that are compatible with
biological interfaces
To find better strategies for immune
acceptance

93. Vascularization :The approach of stimulating the
ingrowths of blood vessels into solid organs has
not been successful. Such organs rapidly, within
hours or even minutes, demanded blood
irrigation for survival and proper function.
Innervation

94. Some tissues already in clinical use
Improvements needed to increase availability and safety
For widespread use, reduced cost is essential
Further work should focus on:
vascularisation of new tissue; maintaining nutrient
supply to cells in matrix with increasing size
Achieving full potential of stem cells to differentiate into
desired cell types