PLATELET PRODUCTION,STRUCTURE AND

FUNCTION
DEBBIE KATES R. SANTISTEBAN,RMT
 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.
PLATELETS
• TRIGGER PRIMARY HOMOSTASIS UPON EXPOSURE TO
SUBENDOTHELIAL COLLAGEN OR ENDOTHELIAL CELLS
INFLAMMATORY PROTEIN AT A TIME OF INJURY.
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.
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
 PLATELET
IMPEDANCE
PLATELET AGGREGOMETRY
ELECTRICAL IMPEDANCE
 PLATELET
INTERNAL
STRUCTURE
PLATELET
RECEPTORS
PLATELETS

ARISE FROM UNIQUE BONE MARRIW

CELLS CALLED MEGAKARYOCYTES
MEGAKARYOCY
TE

• LARGEST CELLS
IN THE BONE
MARROW
• POLYPLOID,POSS
ESING MULTIPLE
CHROMOSOME
COPIES
WRIGHT STAINED BONE MARROW
SMEAR ASPIRATE SMEAR
• EACH MEGAKARYOCYTE IS 30-50 UM IN
DIAMETER WITH A MULTILOBED
NUCLEUS AND ABUNDANT GRANULAR
CYTOPLASM
MEGAKARYOCYTES
• ACCOUNT FOR LESS THAN 0.5% OF ALL BONE MARROW
CELLS
BONE MARROW
ASPIRATE NORMAL
WRIGHT STAINED SMEAR

• 2-4 MEGAKARYOCYTES PER 10 X

LPF
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
MEGAKARYOCYT
ES
THROMBOPOIETIN
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.
MEGAKARYOCYTE DIFFERENTIATION AND PROGENITORS

This Photo by Unknown Author is licensed under CC BY-SA

 • ARISE FROM COMMON MYELOID
MEGAKARYOC PROGENITOR UNDER THE INFLUENCE
YTE OF THE TRANSCRIPTION GENE
PROGENITORS PRODUCT –GATA 1,REGULATED BY
COFACTOR FOG1.
MEGAKARYOCYTE
DIFFERENTIATION

This Photo by Unknown Author is licensed under CC BY

• 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 Photo by Unknown Author is licensed under CC BY-SA

COMMON MYELOID PROGENITOR
• ARISE 3 MEGAKARYOCYTE LINEAGE-COMMITTED
PROGENITOR STAGES
• DEFINED BY THEIR IN VTRO CULTURE COLONY
CHARACTERISTICS
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
BFU AND CFU
MEG
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)
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.
ENDOMITOSIS
•A FORM OF MITOSIS THAT LACKS
TELOPHASE AND CYTOKINE
(SEPARATION INTO DAUGHTER
CELLS)
GATA-1 AND FOG1
• TRANSCRIPTION SLOWS,ANOTHER
TRANSCRIPTION FACTOR RUNX1.
• RUNX1 MEDIATES THE SWITCH FROM
MITOSIS TO ENDOMITOSIS BY
SUPRESSING THE RHO/ROCK SIGNALING
PATHWAY-WHICH SUPPRESS THE
ASSEMBLY OF THE ACTIN CYTOSKELETON.
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
OTHER TRANSCRIPTION FACTOR
• NF-E2,DNA REPLICATION PROCEEDS TO
THE PRODICTION OF 8N,16N EVEN 32 N
PLOIDY WITH DUPLICATED CHROMOSOME
SETS
• SOME MEGAKARYOCYTE
NUCLEI,REPLICATE 5 TIMES,REACHING 128
N– UNUSUAL LEVEL OF PLOIDY-MAY
SIGNAL HEMATOLOGIC DISEASE
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 PLATELETS PER
DAY-TOTAL TURN OVER RATE OF 8-9 DAYS.
HIGH PLATELET CONSUMPTION
• IMMUNE THROMBOCYTOPENIC
PURPURA,PLATELET PRODUCTION RISES
BY TEN FOLD.
TERMINAL MEGAKARYOCYTE
DIFFERENTIATION
• AS ENDOMITOSIS
PROCEEDS,MEGAKARYOCYTE PROGENITORS
LEAVE THE PROLIFERATE PHASE ENTER TO
TERMINAL DIFFERENTIATION.
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
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.
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.
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.
NUCLEAR LOBULARITY
• 1ST 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.
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.
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)
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 IMMUNOLOGIC MARKERS-CD36,CD42,CD61 AND
CD62.
SEVERAL FLOW CYTOMETRIC
MEGAKARYOCYTE MEMBRANE
MARKERS
CYTOPLASMIC COAGULATION FACTOR VIII,VON
WILLEBRAND FACTOR(VWF) AND FIBRINOGEN-
DETECTED BY IMMUNOSTAINING IN THE FULLY
DEVELOPED MEGAKARYOCYTE.
THROMBOCYTOPOIESIS(PLATELET
SHEDDING)
• THROMBOCYTOPOIESIS LEAVES BEHIND A
NAKED MEGAKARYOCYTE NUCLEI TO BE
CONSUMED BY MARROW MACROPHAGES.
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.
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.
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-2ND NONPEPTIDE MPL RECEPTOR
AGONIST;BINDS AND ACTIVATES MPL SITE SEPARATE FROM
ROMIPLOSTIM.
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
OTHER CYTOKINES
IL-11 POLYPEPTIDE MIMETIC-OPREIVEKIN
-STIMULATES PLATELET PRODUCTION IN
PATIENTS WITH CHEMOTHERAPHY INDUCED
THROMBOCYTOPENIA
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
OTHER CYTOKINES& HORMONES THAT PARTICIPATE
SYNERGISTICALLY WITH TPO AND INTERLEUKINS
PLATELET FACTOR 4(PF4)
B-THROMBOGLOBULIN INDICATES THAT
NEUTROPHIL ACTIVATING THEY MAY HAVE A
PEPTIDE 2
ROLE IN CONTROL
IL-8
OTHER FACTORS INHIBIT OF
IN VITRO MEGAKARYOCYT
MEGAKARYOCYTE
GROWTH OPOIESIS IN VIVO.
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 DECREASES AFTER 65 YEARS OLD TO 122-350 X 10⁹/L
IN MEN AND 140-379 X 10⁹/L IN WOMEN.
SCANNING ELECTRON MICROGRAPHS

RESTING PLATELET TEMPERATURE

ACTIVATED
PLATELET
HEMOSTASIS
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.
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.
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.
RESTING
AND
ACTIVATE
D
PLATELET
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.
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.
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.
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.