PLATELET-PRODUCTIONSTRUCTURE-AND-FUNCTION.pdf

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1. PLATELET PRODUCTION,STRUCTURE AND
FUNCTION
DEBBIE KATES R. SANTISTEBAN,RMT

2. MEGAKARYOCYTOPOIESIS
PLATELETS ARE NON NUCLEATED BLOOD CELLS THAT
CIRCULATE AT 150-400 X10⁹/l
AVERAGE PLATELET COUNT SLIGHTLY HIGHER IN WOMEN
THAN IN MEN.
LOWER IN BOTH SEXES WHEN OVER 65 YEARS OLD.

3. PLATELETS
•TRIGGER PRIMARY HOMOSTASIS UPON EXPOSURE TO
SUBENDOTHELIAL COLLAGEN OR ENDOTHELIAL CELLS
INFLAMMATORY PROTEIN AT A TIME OF INJURY.

4. PLATELETS-WRIGHT STAINED WEDGE
PREPARATION BLOOD FILM
•PLATELETS ARE DISTRIBUTED AT 7 -21 CELLS PER 100
X FIELD,DISTRIBUTED THROUGHOUT THE RED BLOOD
CELL MONOLAYER.

5. BLOOD FILM
•PLATELETS HAVE AN
AVERAGE DIAMETER OF
2.5 UM,
CORRESPONDING TO A
MEAN PLATELET
VOLUME(MPV) OF 8 TO
10 Fl WHEN MEASURED
BY IMPEDANCE IN A
BUFFERED ISOTONIC
SUSPENSION-USING LAB
INSTRUMENTS
PROFILING

6. PLATELET
IMPEDANC
E

7. PLATELET AGGREGOMETRY
ELECTRICAL IMPEDANCE

10. PLATELET
INTERNAL
STRUCTUR
E

11. PLATELET
RECEPTORS

12. PLATELETS
ARISE FROM UNIQUE BONE MARRIW
CELLS CALLED MEGAKARYOCYTES

14. MEGAKARYOCYTE
•LARGEST CELLS
IN THE BONE
MARROW
•POLYPLOID,POSS
ESING MULTIPLE
CHROMOSOME
COPIES

18. WRIGHT STAINED BONE MARROW
SMEAR ASPIRATE SMEAR
•EACH MEGAKARYOCYTE IS 30 -50 UM IN
DIAMETER WITH A MULTILOBED
NUCLEUS AND ABUNDANT GRANULAR
CYTOPLASM

19. MEGAKARYOCYTES
•ACCOUNT FOR LESS THAN 0.5% OF ALL BONE MARROW
CELLS

20. BONE MARROW
ASPIRATE NORMAL
WRIGHT STAINED
SMEAR
•2-4 MEGAKARYOCYTES PER 10 X
LPF

21. HEALTHY INTACT BONE MARROW
TISSUE
MEGAKARYOCYTES UNDER
THE INFLUENCE OF AN
ARRAY OF STROMAL CELL
CYTOKINES,CLUSTER WITH
HEMATOPOIETIC STEM
CELLS IN VASCULAR
NICHES ADJACENT TO
VENOUS SINUSOID
ENDOTHELIAL CELLS

22. MEGAKARYOCYTES

23. THROMBOPOIETIN

24. MEGAKARYOCYTE PROGENITORS
•RECRUITED FROM COMMON MYELOID PROGENITORS
•SUBSEQUENTLY DIFFERENTIATE THROUGH SEVERAL
MATURATION STAGES.
•THEY EXTEND PROPLATELET PROCESSES,PROTECTIONS
THAT RESEMBLE STRINGS OF BEADS, THROUGH OR
BETWEEN THE ENDOTHELIAL CELLS AND INTO THE
VENOUS SINUSES,RELEASING PLATELETS FROM THE
TIPS OF THE PROCESSES INTO THE CIRCULATION.
•MEGAKARYOCTES ARE ALSO FOUND IN THE LUNGS.

25. MEGAKARYOCYTE DIFFERENTIATION AND
PROGENITORS
This Photoby Unknown Author is licensed under CC BY-SA

26. MEGAKARYOCYT
E
PROGENITORS
•ARISE FROM COMMON MYELOID
PROGENITOR UNDER THE INFLUENCE
OF THE TRANSCRIPTION GENE
PRODUCT –GATA 1,REGULATED BY
COFACTOR FOG1.

27. MEGAKARYOCYTE
DIFFERENTIATION
•SUPRESSED BY ANOTHER
TRANSCRIPTION GENE PRODUCT-MYB-
GATA1 AND MYB ACT IN OPPOSITION
TO BALANCE MEGAKARYOPOIESIS IN
ONE ARM WITH DIFFERENTIATION TO
THE RED BLOOD CELL LIME IN
ANOTHER ARM-ERYTHROPOIESIS.
This Photoby Unknown Author is licensed under CC BY
This Photoby Unknown Author is licensed under CC BY-SA

31. COMMON MYELOID PROGENITOR
•ARISE 3 MEGAKARYOCYTE LINEAGE -COMMITTED
PROGENITOR STAGES
•DEFINED BY THEIR IN VTRO CULTURE COLONY
CHARACTERISTICS

32. IN ORDER OF DIFFERENTIATION
•LEAST/MATURE BURST-FORMING UNIT(BFU-MEG)
•INTERMEDIATE COLONY-FORMING UNIT(CFU-MEG)
•MORE MATURE PROGENITOR LIGHT DENSITY CFU(LD -
CFU-MEG)
•ALL 3 PROGENITOR STAGES RESEMBLE LYMPHOCYTES
•CANNOT BE DISTINGUISHED BY LIGHT STAINED
MICROSCOPY

33. BFU AND
CFU MEG

34. BFU AND CFU MEG
•DIPLOID AND PARTICIPATE IN NORMAL
MITOSIS MAINTAINING A VIABLE POOL
OF MEGAKARYOCYTE PROGENITORS
•THEIR PROLIFERATIVE PROPERTIES
ARE REFLECTED IN THEIR ABILITY TO
FORM HUNDREDS ( BFU –MEG) OR
SCORES (CFUMEG) OF COLONIES IN
CULTURE)

35. 3
RD
STAGE
•LD-CFU MEG-LOSES ITS
CAPACITY TO DIVIDE BUT
RETAINS ITS DNA REPLICATION
AND CYTOPLASMIC MATURATION -
PARTIALLY CHARACTERIZED
FORM OF MITOSIS UNIQUE TO
MEGAKARYOCYTES KNOWN
ENDOMITOSIS.

37. ENDOMITOSIS
•A FORM OF MITOSIS THAT
LACKS TELOPHASE AND
CYTOKINE (SEPARATION INTO
DAUGHTER CELLS)

38. GATA-1 AND FOG1
•TRANSCRIPTION SLOWS,ANOTHER
TRANSCRIPTION FACTOR RUNX1.
•RUNX1MEDIATES THE SWITCH FROM
MITOSIS TO ENDOMITOSIS BY
SUPRESSING THE RHO/ROCK SIGNALING
PATHWAY-WHICH SUPPRESS THE
ASSEMBLY OF THE ACTIN
CYTOSKELETON.

40. REDUCED RHO/ROCK SIGNAL
•IN RESPONSE,SIGNAL INADEQUATE
LEVELS OF ACTIN AND MYOSIN
(MUSCLE FIBER LIKE MOLECULES)
•ASSEMBLE IN CYTOPLASMIC
CONSTRICTIONS WHERE SEPARATION
WOULD OTHERWISE OCCUR PREVENTING
CYTOKINES

41. OTHER TRANSCRIPTION FACTOR
•NF-E2,DNA REPLICATION PROCEEDS TO
THE PRODICTION OF 8N,16N EVEN32
N PLOIDY WITH DUPLICATED
CHROMOSOME SETS
•SOME MEGAKARYOCYTE
NUCLEI,REPLICATE 5 TIMES,REACHING
128 N–UNUSUAL LEVEL OF PLOIDY -
MAY SIGNAL HEMATOLOGIC DISEASE

42. MEGAKARYOCYTES
•EMPLOY THEIR MULTIPLE DNA COPIES TO
SYNTHESIZE ABUNDANT CYTOPLASM WHICH
DIFFERENTIATES INTO PLATELETS.
•SINGLE MEGAKARYOCYTE -SHED 2000-4000
PLATELETS-”THROMBOCYTOPOIESIS OR
THROMBOPOIESIS”
•AVERAGE SIZE HEALTHY HUMAN=10 ⁸
MEGAKARYOCYTES PRODUCING 10 ¹¹PLATELETS
PER DAY-TOTAL TURN OVER RATE OF 8 -9 DAYS.

43. HIGH PLATELET CONSUMPTION
•IMMUNE THROMBOCYTOPENIC
PURPURA,PLATELET PRODUCTION RISES
BY TEN FOLD.

44. TERMINAL MEGAKARYOCYTE
DIFFERENTIATION
•AS ENDOMITOSIS
PROCEEDS,MEGAKARYOCYTE PROGENITORS
LEAVE THE PROLIFERATE PHASE ENTER TO
TERMINAL DIFFERENTIATION.

45. TERMINAL DIFFERENTIATION
•A SERIES OF STAGES WHICH
MICROSCOPISTS BECOME ABLE TO
RECOGNIZE THEIR UNIQUE WHRIGHT
STAINED MORPHOLOGY IN BONE MARROW
ASPIRATE FILMS OR HEMATOXYLIN AND
EOSIN-STAINED BONE MARROW BIOPSY
SECTIONS.
•MORPHOLOGIST CALL THE LEAST
DIFFERENTIATED MEGAKARYOCYTE
PRECURSOR-MK-1 STAGE OR
MEGAKARYOBLAST

46. MEGAKARYOBLASTS
•ALTHOUGH THEY NO LONGER LOOK LIKE LYMPHOCYTES -
MEGAKARYOBLASTS CANNOT BE RELIABLY
DISTINGUISHED FROM BONE MARROW MYELOBLAST OR
PRONORMOBLASTS (RUBRIBLAST)
•MORPHOLOGIST MAY SEE A VAGUE CLUE -PLASMA
MEMBRANE BLEBS,BLUNT PROJECTIONS FROM THE
MARGIN THAT RESEMBLE PLATELET.

47. MEGAKARYOBLASTS
•BEGINS TO DEVELOP MOST ITS
CYTOPLASMIC ULTRASTRUCTURE-
PROCOAGULANT LADEN ALPHA
GRANULES,DENSE GRANULES(DENSE
BODIES),DEMERCATION SYSTEM (DMS)
•CONTENT AND FUNCTION OF ALPHA AND
DENSE GRANULES ARE DESCRIBED IN THE
SUBSEQUENT SECTION ON MATURE
PLATELET ULTRASTRUCTURE AND
FUNCTION.

48. DMS
•A SERIES OF MEMBRANE LINED CHANNELS
THAT INVADE FROM THE PLASMA MEMBRANE
AND GROW INWARD TO SUBDIVIDE THE
ENTIRE CYTOPLASM.
•BIOLOGICALLY IDENTICAL TO THE
MEGAKARYOCYTE PLASMA MEMBRANE AND
ULTIMATELY DELINEATES THE INDIVIDUAL
PLATELELTS DURING
THROMBOCYTOPOIESIS.

49. NUCLEAR LOBULARITY
•1
ST
BECOMES APPARENT AS AN INDENTATION OF THE 4N
REPLICATION STAGE, RENDERING THE CELL
IDENTIFIERS AS AN MKII STAGE OR
PROMEGAKARYOCYTES BY LIGHT MICROSCOPY.
•MORPHOLOGIST SELDOM DISTINGUISHES MK I,
MKII,MKIII STAGES DURING THE ROUTINE
EXAMINATION OF BONE MARROW ASPIRATE.
•PROMEGAKARYOCYTE REACHES ITS FULL PLOIDY LEVEL
BY THE END OF THE MKII STAGE.
•MOST ABUNDANT MKIII STAGE-MEGAKARYOCYTE IS
EASILY RECOGNIZED AT 10X MAGNIFICATION ON THE
BASIS OF 30-50 UM IN DIAMETER.

50. NUCLEAR LOBULARITY
•NUCLEUS IS INTENSELY INDENTED OR LOBULATED AND THE
DEGREE OF LOBULATION IS IMPRESSIVELY PROPORTIONAL
TO PLOIDY.
•PLOIDY LEVELS ARE MEASURED WITH MEPACRINE.
•MEPACRINE-NUCLEIC ACID DYE IN MEGAKARYOCYTE FLOW
CYTOMETRY.
•CHROMATIN IS VARIABLY CONDENSED WITH LIGHT AND
DARK PATCHES.
•CYTOPLASM-AZUROPHILIC(LAVENDER),GRANULAR AND
PLATELET BECAUSE OF THE SPREAD OF THE DMS AND
ALPHA GRANULES.
•FULL MATURATION-PLATELET SHEDDING AND
THROMBOCYTOPOIESIS PROCEEDS .

51. MEGAKARYOCYTE MEMBRANE
RECEPTORS AND MARKERS
•IDENTIFY VISUALLY INDISTINGUISHABLE
MEGAKARYOCYTE PROGENITORS IN HEMATOLOGIC
DISEASE:
IMMUNOSTAINING OF FIXED TISSUE
FLOW CYTOMETRY WITH IMMUNOLOGIC
PROBES
FLUORESCENT IN SITU
HYBRIDIZATION(FISH)

52. SEVERAL FLOW CYTOMETRIC
MEGAKARYOCYTE MEMBRANE MARKERS
MPL-TPO RECEPTOR SITE PRESENT AT ALL MATURATION
STAGES AND STEM CELL AND COMMON MYELOID
PROGENITOR MARKER CD34.
CD34 MARKER-DISAPPEARS AS DIFFERENTIATION
PROCEEDS
PLATELET MEMBRANE GLYCOPROTEIN IIB AND III
A(CD41 MARKER LOCATED ON THE IIB PORTION) -
FIRST APPEARS ON MEGAKARYOCYTIC PROGENITORS AND
REMAINS PRESENT THROUGHOUT MATURATION,ALONG
WITH IMMUNOLOGICMARKERS-CD36,CD42,CD61 AND
CD62.

53. SEVERAL FLOW CYTOMETRIC
MEGAKARYOCYTE MEMBRANE MARKERS
CYTOPLASMIC COAGULATION FACTOR VIII,VON
WILLEBRAND FACTOR(VWF) AND FIBRINOGEN -
DETECTED BY IMMUNOSTAINING IN THE FULLY
DEVELOPED MEGAKARYOCYTE.

54. THROMBOCYTOPOIESIS(PLATELET
SHEDDING)
•THROMBOCYTOPOIESIS LEAVES BEHIND A
NAKED MEGAKARYOCYTE NUCLEI TO BE
CONSUMED BY MARROW MACROPHAGES.

55. HORMONES AND CYTOKINES OF
MEGAKARYOCYTOPOIESIS
GROWTH FACTOR TPO-70,000 DALTON MOLECULE
THAT POSSESS 23% HOMOLOGY WITH THE RED
BLOOD CELL PRODUCING HORMONE
ERYTHROPOIETIN.
mRNA for TPO-kidney,liver,stromal cells
and smooth muscle cells
Liver-has the most copies and considered
as the primary source .

56. HORMONES AND CYTOKINES OF
MEGAKARYOCYTOPOIESIS
TPO
-CIRCULATES AS A HORMONE IN PLASMA AND IS THE LIGAND
THAT BINDS THE MEGAKARYOCYTE AND PLATELET MEMBRANE
RECEPTOR PROTEIN IDENTIFIED ABOVE.
INDUCES PROLIFERATION AND MATURATION OF MEGAKARYOCYTES
AND INDUCES THROMBOCYTOPOIESIS OR PLATELET RELEASE.
MPL-NAMED FOR V-MPL,A VIRAL ONCOGENE ASSOCIATED WITH
MURINE MYELOPROLIFERATIVE LEUKEMIA
*PLASMA CONC. OF TPO IS INVERSELY PROPORTIONAL TO
PLATELET MEGAKARYOCYTE MASS —IMPLYFYINNG-MEMBRANE
BINDING AND CONSEQUENT REMOVAL OF TPO BY PLATELETS -
PRIMARY PLATELET COUNT CONTROL MECHANISM.

57. HORMONES AND CYTOKINES OF
MEGAKARYOCYTOPOIESIS
SYNTHETIC TPO MIMETICS -ELEVATE PH COUNT IN
PATIENTS TREATED FOR A VARIETY OF CANCERS
INCLUDING ACUTE LEUKEMIA.
ROMIPLOSTIM(1 COMMERCIAL MPL RECEPTOR
ANTAGONIST)-
NON IMMUNOLOGIC OLIGOPEPTIDE -EFFECTIVE IN
RAISING PLATELET COUNT IN IMMUNE
THROMBOCYTOPENIC PURPURA
ELTROMBOPAG-2
ND
NONPEPTIDE MPL RECEPTOR
AGONIST;BINDS AND ACTIVATES MPL SITE SEPARATE
FROM ROMIPLOSTIM.

58. OTHER CYTOKINES
•-FUNCTION WITH TPO
•STIMULATE MEGAKARYOCYTPOIESIS
•INCLUDE INTERLEUKIN 3(IL3),IL -6 AND IL-11
IL3-ACT IN SYNERGY WITH TPO TO INDUCE THE EARLY
DIFFERENTIATION OF STEM CELLS.
IL6 AND IL 11-ACT IN THE PRESENCE OF TPO TO
ENHANCE PHENOMENA OF ENDOMITOSIS,MEGAKARYOCYTE
MATURATION AND THROMBOCYTOPOIESIS

59. OTHER CYTOKINES
IL-11 POLYPEPTIDE MIMETIC -OPREIVEKIN
-STIMULATES PLATELET PRODUCTION IN
PATIENTS WITH CHEMOTHERAPHY INDUCED
THROMBOCYTOPENIA

60. OTHER CYTOKINES& HORMONES THAT
PARTICIPATE SYNERGISTICALLY WITH
TPO AND INTERLEUKINS
STEM CELL FACTOR-KIT LIGAND OR MAST CELL
GROWTH FACTOR
GRANULOCYTE MACROPHAGE COLONY
STIMULATING FACTOR(GM -CSF)
GRANULOCYTE COLONY STIMULATING FACTOR(G -
CSF)
ACETYLCHOLINESTERASE -DERIVED
MEGAKARYOCYTE GROWTH STIMULATING PEPTIDE

61. OTHER CYTOKINES& HORMONES THAT
PARTICIPATE SYNERGISTICALLY WITH
TPO AND INTERLEUKINS
PLATELET FACTOR 4(PF4)
B-THROMBOGLOBULIN
NEUTROPHIL ACTIVATING
PEPTIDE 2
IL-8
OTHER FACTORS INHIBIT
IN VITRO MEGAKARYOCYTE
GROWTH
INDICATES THAT
THEY MAY HAVE A
ROLE IN CONTROL
OF
MEGAKARYOCYTOPO
IESIS IN VIVO.

62. PLATELETS
•PROPLATELET PROCESS SHEDS PLATELETS,CELLS
CONSISTING OF GRANULAR CYTOPLASM WITH A
MEMBRANE BUT NO NUCLEAR MATERIAL INTO THE
VENOUS SINUS OF BONE MARROW.
•CIRCULATING OR RESTING PLATELETS ARE
BICONVEX;ALTHOUGH THE PLATELETS COLLECTED IN
EDTA TEND TO “ROUND UP”.
•NORMAL PERIPHERAL BLOOD PLATELET COUNT IS 150-
400 X 10⁹/L
•COUNT DECREASESAFTER 65 YEARS OLD TO 122-350 X
10⁹/L IN MEN AND 140-379 X 10⁹/L IN WOMEN.

64. SCANNING ELECTRON MICROGRAPHS
RESTING PLATELET
TEMPERATURE
ACTIVATED
PLATELET

66. HEMOSTASI
S

69. ADHESION
AS BLOOD FLOWS VESSEL
WALLS CREATE STRESS OR
SHEAR FORCE MEASURED IN
UNITS LABELED AS S-1.
INJURY TO VESSEL WALLS
DISRUPTS THE COLLAGEN OF
THE EXTRACELLULAR
MATRIX.

70. AGGREGATION:PLATEL
ETS IRREVERSIBLY
COHERE
IN ADDITION TO COLLAGEN
EXPOSURE AND VWF
SECRETION,BLOOD VESSEL
INJURY RELEASES
CONSTITUITIVE (INTEGRAL)
TISSUE FACTOR FROM
ENDOTHELIAL CELLS.
TISSUE FACTOR TRIGGERS
THE FORMATION OF
THROMBIN.

71. SECRETION
•ACTIVATED PLATELETS
RELEASE GRANULAR
CONTENTS.
•OUTSIDE IN PLATELET
ACTIVATION THROUGH
LIGAND(AGONIST BINDING)
TO INTEGRINS,STRS, AND
THE IG GENE PRODUCT GP VI
TRIGGERS ACTIN
MICROFILAMENT
CONTRACTION.

72. RESTING
AND
ACTIVAT
ED
PLATELE
T

76. PLATELET AGGREGATION
•IT IS THE KEY PART OF PRIMARY
HOMEOSTASIS,WHICH IN ARTERIES MAY END
WITH THE FORMATION OF “ WHITE CLOT”, A
CLOT COMPOSED PRIMARILY OF PLATELETS
AND VWF.
•ALTHOUGH AGGREGATION IS PART OF NORMAL
VESSEL REPAIR, WHITE CLOTS OFTEN IMPLY
INAPPROPRIATE PLATELET ACTIVATION IN
SEEMINGLY UNINJURED ARTERIOLES AND
ARTERIES AND ARE THE PATHOLOGICAL
BASIS OF ARTERIAL THROMBOTIC EVENTS
SUCH AS AMI,PERIPHERAL ARTERY DISEASE
AND STROKES.

77. PLATELET
AGGREGATION
•FIBRIN AND RED BLOOD CELL
DEPOSIT AROUND AND WITHIN
THE PLATELET
SYNCTIUM,FORMING A BULKY
“RED CLOT”
•THE RED CLOT IS ESSENTIAL
TO WOUND REPAIR,BUT IT MAY
ALSO BE A CHARACTERISTIC
OF INAPPROPRIATE
COAGULATION IN VENULES AND
VEINS,RESULTING IN DEEP
VEIN THROMBOSIS AND
PULMONARY EMBOLI.

78. COAGULATION
•SECONDARY HEMOSTASIS
•THE COMBINATION OF POLAR
PHOSPHOLIPID EXPOSURE ON
ACTIVATED PLATELETS,PLATELET
FRAGMENTATION WITH CELLULAR
MICROPARTICLE DISPERSION AND
SECRETION OF PLATELETS ALPHA AND
DENSE GRANULES CONTENTS THAT
TRIGGERS SECONDARY HEMOSTASIS.

81. TXA2
•IT HAS A HALF LIFE OF 30 SECONDS IT
DIFFUSES FROM PLATELET AND BECOMES
SPONTANEOUSLY REDUCED TO THROMBOXANE B2,
A STABLE MEASURABLE PLASMA METABOLITE.
•THROMBOXANE B2 IS ACTED ON A VARIETY OF
LIVER ENZYMES TO PRODUCE AN ARRAY OF
SOLUBLE URINE METABOLITES ,INCLUDING 11 -
DEHYDROTHROMBOXANE B2,WHICH IS STABLE AND
MEASURABLE.