Table of contents :

bone marrow

Blood / haema / hema / sanguis : the fluid that circulates through the heart, arteries, capillaries, and veins, carrying nutriment and oxygen to the body cells. It consists of the plasma, a pale yellow liquid containing the microscopically visible formed elements of the blood: the erythrocytes, or red blood corpuscles; the leukocytes, or white blood corpuscles; and the platelets, or thrombocytes. Its amount in a normal adult is ~ 5 L (~ 7% of body volume or 8.5-9.0% of body mass) :

• arterial blood : oxygenated blood, found in the pulmonary veins, the left chambers of the heart, and the systemic arteries; it is bright red in color
• venous blood : deoxygenated blood, found in the systemic veins, the right chambers of the heart, and the pulmonary arteries; it is dark red in color.
• cord blood : blood contained within the umbilical vessels at the time of delivery of the infant.
• peripheral blood : blood obtained from acral areas, or from the circulation remote from the heart, as from earlobe, fingertip, or heel pad (in a child), or from the antecubital vein; the blood in the systemic circulation.
• whole human blood : blood from which none of the elements have been removed.  2. blood that has been drawn from a selected donor under strict aseptic conditions, containing citrate ion or heparin as an anticoagulant; used as a blood replenisher
• citrated blood : blood treated with sodium citrate or citric acid to prevent its coagulation.
• defibrinated blood : whole blood from which fibrin was separated during the clotting process.
• laky blood : blood that has undergone laking and contains at least some lysed erythrocytes.
• occult blood : blood present in such small quantities that, while it is not visible to the naked eye, it can be detected only by chemical tests of suspected material, e.g., feces.
• sludged blood : blood in which the red cells have become aggregated into masses

• blood islands : aggregations of mesenchyme cells in the angioblast of the early embryo, as in the wall of the yolk sac; they subsequently develop into vascular endothelium and blood corpuscles.
• 40-45 % is represented by figurated elements (cells and platelets) arising from hematopoiesis :
• primitive hematopoiesis (only erythrocytes and megakaryocytes are produced) :
• from foetal week ? up to foetal week 12 in yolk sac (and intraembryonic splanchnopleura ?)
• definitive hematopoiesis from (pluripotent) hematopoietic stem cell ((P)HSC)
• from foetal week ? up to foetal week ? in aorta-gonads-mesonephros (AGM) region (HSCs are probably generated within the layer of endothelial cells that lines the wall of the dorsal aorta, so sharing a common precursor with endothelial cells)
• from foetal week 6 up to foetal week 38 in liver
• from foetal week 12 up to foetal week 38 in spleen and lymph nodes
• from foetal week 20 up in bone marrow, decreasing with age (in long bones no hematopoiesis occur after age 20, except for the proximal portion of humeruses and tibias => only in flat bones (vertebrae, sternum, ribs and iliac wings)). Each day an adult produces approximately 200 billion erythrocytes (3 x 10⁹/kg), 100 billion leukocytes (0.8 x 10⁹/kg) , and 100 billion platelets (1.5 x 10⁹/kg). Moreover, these rates can increase by a factor or 10 or more when the demand for blood cells increases.

HSC localization to the bone marrow depends on CXCL12 / SDF-1 (produced by bone marrow stromal cells)-- CXCR4 interaction. The bone marrow microenvironment has a crucial supporting role in the growth, differentiation and survival of HSC. Without bone-marrow stromal cells, HSCs cannot be maintained in vitro, even when cultured with a cocktail of growth factors. KIRRE / NEPH2 protein is accumulated in the areas of contact to maintain HSCs in an undifferentiated, proliferative state. TIE1 and TIE2 receptor tyrosine kinases are not required for fetal haematopoiesis, but they are requried during postnatal bone-marrow hematopoiesis^ref. Unique features of bone that contribute to a microenvironmental niche for stem cells might include the known high concentration of calcium ions at the HSC-enriched endosteal surface. Cells respond to extracellular ionic calcium concentrations through calcium-sensing receptor (CaSR), expressed on HSCs. Through the CaSR, the simple ionic mineral content of the niche may dictate the preferential localization of adult mammalian haematopoiesis in bone. Antenatal mice deficient in CaSR had primitive haematopoietic cells in the circulation and spleen, whereas few were found in bone marrow. CaSR^-/- HSCs from fetal liver were normal in number, in proliferative and differentiative function, and in migration and homing to the bone marrow. Yet they were highly defective in localizing anatomically to the endosteal niche, behaviour that correlated with defective adhesion to the extracellular matrix protein, collagen I. CaSR has a function in retaining HSCs in close physical proximity to the endosteal surface and the regulatory niche components associated with it^ref. HSCs can be isolated efficiently from bone marrow cells by following Gata2-directed GFP fluorescence, and that they can also be monitored in vivo. Each individual GFP⁺ cell lay in a G₀/G₁ cell cycle state, in intimate contact with osteoblasts beside the endosteum, at the edge of the bone marrow. The HSC niche is composed of solitary cells and that adult bone marrow HSC are not clustered^ref
• diseases that cause bone marrow replacement can cause hematopoiesis to revert to the liver, even in adulthood.
Normal myelogram in adulthood :
• granulocyte series (56%)
• myeloblast (1.0%)
• promyelocyte (3.4%)
• myelocyte (11.9%)
• metamyelocyte (18%)
• band form (11%)
• neutrophils (10.7%)
• eosinophils (3.2%)
• basophils (< 0.1%)
• erythroid series (21.5%)
• proerythroblast (0.6%)
• basophilic erythroblast (2.0%)
• polychromatophilic erythroblast (12.4%)
• orthochromatic erythroblast (6.5%)
• megakaryocytes (< 0.1%)
• lymphocytes (15.8%)
• plasma cells (1.8%)
• monocytes (1.8%)
The initial divisions of HSC are likely to involve cytokine-independent stochastic fate choices (promiscuous gene expression provides a framework for stochastic fluctuations in expression of signaling or transcriptional complexes, which ultimately are amplified or repressed to cement lineage choice), while maturation of terminally differentiated cells from progenitor and precursor cells depends on instructive cytokines.
• colony-forming unit (CFU) : any of several hematopoietic stem cells identified by their ability to give rise to monoclonal colonies in the spleen when transplanted into isogeneic, lethally irradiated mice.
• unitarian or monophyletic theory / monophyletism : the theory that all forms of blood cells have their origin in a single type of cell, the blast cell / hematoblast / hematocytoblast / hemoblast / hemocytoblast / hematogone / hematohistioblast / hemohistioblast (which develops into a pluripotential stem cell), with the different types of cells arising from there by a process of differentiation


• polyphyletic theory / polyphyletism : the theory that the various blood cells have their origin from two or more types of stem cells
• dualistic theory / dualism : a variant of the polyphyletic theory that holds that blood cells arise from 2 distinct types of stem cells, the myeloblasts and lymphoblasts
• trialistic theory / trialism : a variant of the polyphyletic theory that holds that blood cells arise from 3 distinct types of stem cells, the myeloblasts, lymphoblasts, and monocytes
Hematopoietins at COPE
• synclitism / syncliticism : normal, synchronous maturation of the nucleus and cytoplasm of blood cells
IL-3, SCF and Epo induce expression of the anti-apoptotic protein anamorsin, essential for late stages of definitive hematopoiesis and terminal differentiation (deficient embryos die in late gestation due to anemia with reduces by 50% the number of peripheral RBCs compared with littermate controls, and a marked proportion of cells in the fetal liver, which was < 1/3 of the size of that in anamorsin-sufficient embryos, are apoptotic)^ref
MicroRNAs (miRNAs) influence hematopoietic lineage commitment but do not completely block differentiation to other lineages : in mouse 3 miRNAs show preferential expression in haematopoietic tissue^ref
• miR-181 is preferentialy expressed by the B-lymphoid lineage compared with undifferentiated progenitor cells or other lineages
• miR-223 is preferentially expressed by myeloid lineage cells
• miR-142 is most highly expressed by B cells and myeloid cells

[a viral homologue of  CD200 encoded by the K14 ORF of HHV8 interacts with the CD200R with the same kinetics and low affinity as the host protein, although it has only 40% sequence identity. Cells expressing CD200 or K14 inhibits the secretion of pro-inflammatory cytokines by activated macrophages, indicating that infected cells might deliver downmodulatory signals to host myeloid cells through the CD200 receptor to evade elimination^ref]

+ ? :
• precursor of myeloid cells (p-M)

+ ? :
• common myeloerythroid progenitor (p-ME)
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+ SCF, IL-1b, IL-3, IL-4, IL-6 :
• MEMeg
• erythroid cells => erythropoiesis: the erythron is found near bone marrow blood vessels and its maturation requires ~ 5 days. Functional reserve allows increasing output by 6-8 folds. Uneffective erythropoiesis normally accounts for 10-15%. It consists of :
• burst-forming unit–erythroid (P-BFU-E) : the earliest erythrocyte precursor in the erythrocytic series, detectable mainly in vitro and having a high requirement for erythropoietin; it gets its name from the fact that its growth is composed of subcolonies resembling bursts

‡
+ GM-CSF, IL-1b, IL-3, IL-4; need GATA1 :
• M-BFU-E

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• colony-forming unit–erythroid (CFU-E) : an erythrocyte precursor in the erythrocyte series that follows the burst-forming unit–erythroid and precedes the proerythroblast; detectable mainly in vitro (CD71 / TfR⁺)

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+ Epo, IL-9 :
• nucleated RBC (NRBC)
• proerythroblast / pronormoblast / lymphoid hemoblast of Pappenheim / rubriblast (CD71 / TfR⁺235a / glycophorin A^-235b / glycophorin B^-235R / glycophorin C^-) : 20-25 mm diameter; basophilic cytoplasm with large, distinct nucleoli


They begin synthesis of hemoglobins (Hb), the red oxygen-carrying pigment of erythrocytes, formed by developing erythrocytes in bone marrow. It is a type of hemoprotein that contains 4 heme groups and globin and has the property of reversible oxygenation. A molecule of hemoglobin contains 4 polypeptide globin chains, composed of between 141 and 146 amino acids; those most often found are a and b chains, with g and d chains seen somewhat less often. Different types of hemoglobins are determined by different combinations of chains, with the number of chains of each type in the molecule being indicated by a subscript. Hundreds of hemoglobins with differing electrophoretic mobilities and characteristics have been reported; the first ones were given capital letters, such as S, C, D, E, G, H, I, J, K, L, M, N, and Q. As refined biochemical techniques led to the discovery of many additional hemoglobins, newer standards for nomenclature were devised: those with electrophoretic mobility equal to one of the lettered hemoglobins could be named with that letter using the place of discovery as a subscript, such as hemoglobin M_Saskatoon or hemoglobin M_Milwaukee. New hemoglobins with unique electrophoretic mobilities are now often named simply for the laboratory, hospital, or town where they were discovered, such as hemoglobin Chesapeake or hemoglobin Gun Hill. When known, the number of each amino acid substituting in each polypeptide in the molecule should be indicated by the appropriate superscript numeral
• 4 chains of globin(s) (synthetized in cytoplasm)
• globins genes are arranged as follows :
• on chromosome 16p13.3 : 5'-z-y_z-y_a2-y_a1-a₂-a₁-q₁-3'. a₂ and a₁ cds are identical but genes differ in 5'UTR, intron, and 3'UTR


• on chromosome 11p15.5 : 5'-e-g_G-g_A-y_b-d-b-3'
• in embryo :
• Gower hemoglobin / hemoglobin Gower : present in early embryonic life and disappearing before birth
• Hb Gower-1 (z₂e₂)
• Hb Gower-2 (a₂e₂)
• occasionally e₄
• in fetus :
• HbF / fetal hemoglobin (a₂g₂^F) : > 50%;becomes the dominant fetal hemoglobin at 2 months' gestation. HbF binds oxygen more tightly than HbA, enhancing delivery of oxygen from maternal oxyhemoglobin to fetal deoxyhemoglobin at the placenta.
• Hb Portland (z₂g₂) present in the fetus late in the first trimester of pregnancy; it disappears in utero.
• HbA₂ (a₂d₂)
• in adults : KLF1 / EKLF and PYR complex modulate a shift from g- to b-globin binding to the b-globin promoter but not to the repeated CCTTG domain in the g-globin promoter.
• HbA (a₂^Ab₂^A or a₂b₂) (97%)
• HbA₀ (93-95%)
• HbA₁ / glycated hemoglobin (GHb) / glycohemoglobin (5-7%) (sometimes unproperly^ref named glycosylated or glucosylated haemoglobin) (3.9-6.1%) : any of various hemoglobins to which glucose is bound by glycation.
• HbA_1a (0.5%)
• HbA_1b (0.5%)
• HbA_1c (4-6%) has a hexose attached to the N terminal of its b chain. HbA_1c is formed through the progressive glycosylation of Hb during the 120-day lifetime of erythrocytes^ref. It forms when haemoglobin is exposed to glucose and undergoes a sequence of nonenzymatic reactions. The first is the rapid but reversible formation of an aldimine (or Schiff base), followed by the considerably slower formation of a stable ketoamine via a process known as the Amadori rearrangement^ref. The ketoamine steadily accumulates over the life of the red cell and forms the bulk of the glycated haemoglobin measured by laboratories. Therefore, both the erythrocyte age and blood glucose concentration are important determinants of the HbA_1c value. Young erythrocytes have lower HbA_1c values than old; its levels are increased in poorly controlled diabetes mellitus
• HbA₂ (a₂^Ad₂^A2 or a₂^Ad₂) (1.5-3%)
• HbF (a₂g₂) (0.5-2%) : abnormally elevated in aplastic anemia, leukemia, and certain types of thalassemia

Coordinated expression of the genes in each cluster at all stages of development is dependent on critical regulatory elements located upstream of the genes. In the ß cluster there are 5 such elements, collectively referred to as the locus control region (LCR) while in the a cluster, there is a single known element referred to as HS-40. In normal individuals, the synthesis of a and ß globin chains is finely balanced during terminal erythroid differentiation, giving rise to red cells of consistent size (MCV) and hemoglobin content (MCH).
The synthesis of haemoglobin A (HbA) is exquisitely coordinated during erythrocyte development to prevent damaging effects from individual a- and b-subunits. The a-haemoglobin-stabilizing protein (AHSP) / erythroid associated factor (ERAF) binds a-haemoglobin (aHb), inhibits the ability of aHb to generate reactive oxygen species and prevents its precipitation on exposure to oxidant stress. The structure of AHSP bound to ferrous aHb is thought to represent a transitional complex through which aHb is converted to a non-reactive, hexacoordinate ferric form. The crystal structure of this ferric aHb-AHSP complex has been reported at 2.4 Å resolution. There is a striking bis-histidyl configuration in which both the proximal and the distal histidines coordinate the haem iron atom. To attain this unusual conformation, segments of aHb undergo drastic structural rearrangements, including the repositioning of several a-helices. Moreover, conversion to the ferric bis-histidine configuration strongly and specifically inhibits redox chemistry catalysis and haem loss from aHb. The observed structural changes, which impair the chemical reactivity of haem iron, explain how AHSP stabilizes aHb and prevents its damaging effects in cells^ref.
Function of globins :
• prevent Fe²⁺ oxidation by carried O₂
• blood pH buffering
• 4 heme molecules (one per globin), synthetized in mitochondrial matrix and cytoplasm in the following steps :
• glycine + succinyl-CoA

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• a-amino-b-ketoadipic acid

‡
• d-aminolevulinic acid :

‡ porphobilinogen synthase / aminolevulinate dehydratase : The enzyme is inhibited by minute quantities of lead poisoning. Genetic deficiency of the enzyme causes a porphyria similar to acute intermittent porphyria but with high urinary levels of d-aminolevulinic acid, coproporphyrinogen III, and protoporphyrin but not porphobilinogen. It is an enzyme of the lyase class that catalyzes the condensation of 2 molecules of d-aminolevulinate to form ...
• porphobilinogen (PBG) : the immediate precursor of the porphyrins, a pyrrole ring with acetyl, propionyl, and aminomethyl side chains; 4 molecules of porphobilinogen are condensed to form one molecule of uroporphyrinogen III, which is then converted successively to coproporphyrinogen III, protoporphyrin IX, and heme. Porphobilinogen is produced in excess and excreted in the urine in acute intermittent porphyria and several other porphyrias

‡
• porphyrinogen : the reduced form of a porphyrin, containing pyrrole rings linked by methylene (—CH₂—) instead of methylidyne bridges. The porphyrinogen forms are the functional intermediates in the biosynthesis of heme and if oxidized to their corresponding porphyrins, such as occurs in porphyrias, are irreversibly removed from the biosynthetic pathway and accumulate in tissues. Their nomenclature corresponds to that of the porphyrins.
‡
• porphyrin : any of a group of compounds containing the porphin structure, four pyrrole rings connected by methylidyne (—CH=) bridges in a cyclic configuration, to which a variety of side chains may be attached. The nature of the side chains is indicated by a prefix, such as coproporphyrin, deuteroporphyrin, etioporphyrin (each pyrrole ring has one methyl and one ethyl side chain), hematoporphyrin (2 pyrrole rings have 1 methyl and 1 propionate side chain and the other 2 pyrrole rings have 1 methyl and 1 1-hydroxyethyl side chain), mesoporphyrin (2 pyrrole rings each have 1 methyl and 1 propionate side chain and the other 2 pyrrole rings each have 1 methyl and 1 ethyl side chain), protoporphyrin, or uroporphyrin.. Structural isomers are indicated by roman numerals. Free porphyrins are rarely found in tissues except in disorders of heme biosynthesis (porphyrias), but they do occur in the prosthetic groups of hemoglobin, myoglobin, and cytochromes, complexed with metal ions. The term is sometimes used to include porphin or to denote porphin specifically


(A) : pyrrole ring; (B) : porphin ring; (C) : protoporphyrin IX.
• porphyrinogen : the reduced form of a porphyrin, containing pyrrole rings linked by methylene (—CH₂—) instead of methylidyne bridges. The porphyrinogen forms are the functional intermediates in the biosynthesis of heme and if oxidized to their corresponding porphyrins, such as occurs in porphyrias, are irreversibly removed from the biosynthetic pathway and accumulate in tissues. Their nomenclature corresponds to that of the porphyrins

‡
• uroporphyrinogen : a porphyrinogen in which each pyrrole ring has one acetate side chain and one propionate side chain; it is formed by condensation 4 four molecules of porphobilinogen. 4 isomers are possible but only 2 exist naturally, types I and III; the latter is a functional intermediate in heme biosynthesis while the former is produced in an abortive side reaction.

‡
• uroporphyrin : the porphyrin produced by oxidation of the methylene bridges in uroporphyrinogen. Excessive amounts of uroporphyrin I are excreted in congenital erythropoietic porphyria, and both types I and III are excreted in porphyria cutanea tarda
• coproporphyrinogen : a porphyrinogen in which each pyrrole ring has one methyl side chain and one propionate side chain; it is formed by oxidative decarboxylation of uroporphyrinogen. 4 isomers are possible but only 2 exist naturally, types I and III; the latter is a functional intermediate in heme biosynthesis while the former is produced in an abortive side reaction.

‡
• coproporphyrin : the porphyrin produced by oxidation of the methylene bridges in coproporphyrinogen. Coproporphyrin III is excreted in the feces and urine in hereditary coproporphyria and in variegate porphyria, particularly during acute attacks; coproporphyrin I is excreted in the feces and urine in congenital erythropoietic porphyria
• harderoporphyrin : an intermediate in heme biosynthesis, formed in the conversion of coproporphyrinogen to protoporphyrinogen and excreted excessively in the feces in harderoporphyria

‡
• protoporphyrinogen : a porphyrinogen in which 2 pyrrole rings each have one methyl and one propionate side chain and the other two pyrrole rings each have one methyl and one vinyl side chain. 15 isomers are possible but only one, type IX, occurs naturally, produced by oxidative decarboxylation of coproporphyrinogen; it is an intermediate in heme biosynthesis.

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• protoporphyrin : the porphyrin produced by oxidation of the methylene bridge of protoporphyrinogen. Protoporphyrin IX is the only naturally occurring isomer; it is an intermediate in heme biosynthesis, combining with ferrous iron to form protoheme IX, the heme prosthetic group of hemoglobin. It is accumulated and excreted excessively in the feces in protoporphyria and variegate porphyria
• free erythrocyte protoporphyrin (FEP) : protoporphyrin free in the blood rather than incorporated into erythrocytes; most is actually bound to zinc as zinc protoporphyrin. The amount increases greatly in iron-deficiency anemia, lead poisoning, and protoporphyria
• zinc protoporphyrin : free erythrocyte protoporphyrin that is bound to zinc.
• protoporphyrin IX, which binds to ... :
• Fe²⁺ : erythroblasts must modify the "standard" post-transcriptional feedback regulation, balancing expression of ferritin (iron storage) versus TfR1 (iron uptake) via specific mRNA binding of iron regulatory proteins (IRPs) (IRP-1 / aconitase and IRP2). Although erythroid differentiation involves high levels of incoming iron, TfR1 mRNA stability must be sustained and Fer mRNA translation must not be activated because iron storage would counteract hemoglobinization. Furthermore, translation of the erythroid-specific form of aminolevulinic acid synthase (ALAS-E) mRNA, catalyzing the first step of heme biosynthesis and regulated similarly as Fer mRNA by IRPs, must be ensured. Strong inhibition of Fer mRNA translation and efficient ALAS-E mRNA translation in differentiating erythroblasts Moreover, in contrast to self-renewing cells, TfR1 stability and IRP mRNA binding were no longer modulated by iron supply^ref
Each hemoglobin molecule can bind 4 O₂ molecules. Each gram of hemoglobin can bind 1.39 mL O₂. Without such a buffering, plasma could carry only 0.29 mL O₂ / dL.
Oxygen-hemoglobin dissociation curves : a graphic curve representing the normal variation in the amount of O₂ that combines with hemoglobin (oxidized or oxygenated hemoglobin / oxyhemoglobin = 1 - reduced hemoglobin / deoxyhemoglobin) as a function of the partial pressure of oxygen (P_O2)

• shift to the right (the Bohr effect) when less than a normal amount of oxygen is taken up by the blood at a given P_O2
• decreased pH, increased temperature and increased P_CO2, such as occur in the capillaries in metabolically active tissue
• increased concentration of 2,3-DPG
• presence of carbon monoxide
• certain disease states
• shift to the left (the Haldane effect) when more than a normal amount is taken up.
• increased pH, decreased temperature and decreased P_CO2, such as occurs in the alveolar capillaries of the lungs
• decreased 2,3-DPG concentration
It is proposed that the bond between nitric oxide (NO) and the Hb thiol Cys-b⁹³ (SNOHb) is favored when hemoglobin (Hb) is in the relaxed (R, oxygenated) conformation, and that deoxygenation to tense (T) state destabilizes the SNOHb bond, allowing transfer of NO from Hb to form other (vasoactive) S-nitrosothiols (SNOs). However, it has not previously been possible to measure SNOHb without extensive Hb preparation, altering its allostery and SNO distribution. An assay for SNOHb that uses carbon monoxide (CO) and cuprous chloride (CuCl)-saturated Cys is specific for SNOs and sensitive to 2–5 pmol. Uniquely, it measures the total SNO content of unmodified erythrocytes (RBCs) (SNO_RBC), preserving Hb allostery. In room air, the ratio of SNO_RBC to Hb in intact RBCs is stable over time, but there is a logarithmic loss of SNO_RBC with oxyHb desaturation (slope, 0.043). This decay is accelerated by extraerythrocytic thiol (slope, 0.089; P < 0.001). SNO_RBC stability is uncoupled from O₂ tension when Hb is locked in the R state by CO pretreatment. Also, SNO_RBC is increased 20-fold in human septic shock (P = 0.002) and the O₂-dependent vasoactivity of RBCs is affected profoundly by SNO content in a murine lung bioassay. These data demonstrate that SNO content and O₂ saturation are tightly coupled in intact RBCs and that this coupling is likely to be of pathophysiological significance^ref.
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• basophilic erythroblast / basophilic normoblast / prorubricyte (CD71 / TfR⁺235a / glycophorin A⁺235b / glycophorin B⁺235R / glycophorin C⁺). 16-18 mm diameter; The cytoplasm is intensely basophilic; indistinct nucleoli. As the cell matures, chromatin condensation or "clumping" becomes more prominent.


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• polychromatic or polychromatophil erythroblast or normoblast / rubricyte : immature erythrocytes 12-15 mm diameter staining with both acid and basic stains so that their color is a diffuse mixture of blue-gray and pink (CD71 / TfR⁺)


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• orthochromatic (i.e. acidophilic) erythroblast or normoblast / metarubricyte (CD71 / TfR⁺) : 9-11 mm diameter; pyknotic nucleus


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• up-regulation of survivin^ref
• cell-cycle exit and nucleus exocytosis (enucleation) : definitive erythropoiesis usually occurs in the bone marrow or fetal liver, where erythroblasts are associated with a central macrophage in anatomical units called 'blood islands'. Late in erythropoiesis, nuclei are expelled from the erythroid precursor cells and engulfed by the macrophages in the blood island. The nuclei are engulfed by macrophages only after they are disconnected from reticulocytes, and that phosphatidylserine, which is often used as an 'eat me' signal for apoptotic cells, is also used for the engulfment of nuclei expelled from erythroblasts. When late-stage erythroblasts from the spleens of phlebotomized mice were cultured, the nuclei protruded spontaneously from the erythroblasts. A weak physical force could disconnect the nuclei from the reticulocytes. The released nuclei contained an undetectable level of ATP, and quickly exposed phosphatidylserine on their surface. Fetal liver macrophages efficiently engulfed the nuclei; masking the phosphatidylserine on the nuclei with the dominant-negative form of milk-fat-globule EGF8 (MFG-E8) prevented this engulfment^ref
• adult red blood cells have no mitochondria : the cells have no Krebs cycle and meet their energy requirements by glycolysis
• reticulocyte (CD71 / TfR⁺) circulates for only 1 to 2 days; 7-9 mm diameter


Using special stains such as methylene blue or brilliant cresyl blue, reticulocytes stain with dark blue granules (mitochondria and cytoplasmid ribonucleoproteins) whereas mature erythrocytes evenly stain pale blue.

• reticulocyte count : 0.5÷1.5% of total blood cells, recorded as ...
• the percentage of the erythrocyte count
• an absolute number absolute reticulocyte count (ARC) = %[reticulocyte] ^. RBC = 35,000÷40,000 / mL
• correction for anemia : %[reticulocyte] ^. (HCT of the patient)/(normal HCT)
• reticulocyte index = %[reticulocyte] ^. (patient's HCT) / (normal HCT) ^. 0.5 = 1-2.5
• > 2.5 : good erythropoietic response to hemolytic anemia
• < 1 : aplastic crisis
It provides a means of assessing the erythropoietic activity of the bone marrow.
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• erythrocyte / red blood cell (RBC) / ozonophore (CD14^-15^-35 / CR1⁺46^-47⁺55 / DAF⁺75⁺233 / Diego blood group / band 3⁺234 / Duffy antigen / Fy-glycoprotein⁺235a / glycophorin A⁺235b / glycophorin B⁺235R / glycophorin C⁺238 / Kell⁺241 / Rh Ag⁺carbonic anhydrase I⁺). 7-8 mm diameter; The main function of RBCs is to avoid filtration of 3% hemoglobin into urine.

A model depicting the major proteins of the erythrocyte membrane^ref :

Major erythrocyte membrane proteins :

band	designation	molecular weight	chromosomal locus	function	interactions	localization	associated disease
1	a-spectrin (SPTA1)	281	1q22-q23	membrane skeleton	band 3, ankyrin	peripheral	hereditary elliptocytosis, hereditary pyropoikylocytosis, hereditary spherocytosis
1	b-spectrin (SPTB)	246	14q23-q24.1				hereditary spherocytosis
2.1	ankyrin 1 (ANK1)	206	8p11.2	binds membrane protien and cytoskeleton	band 3, b-spectrin	peripheral
2.9	a-adducin / adductin 1 (ADD1)	81	4p16.3	binds actin filaments	spectrin, b-actin	peripheral
2.9	b-adducin / adducin 2 (ADD2)	80		binds actin filaments	spectrin, b-actin	peripheral
3	SLC4A1 / erythrocyte membrane protein band 3 / anion exchange protein 1 (AE1) / CD233	102	17q12-q21	anion transport	ankyrin, spectrin, protein 4.1	integral	hereditary elliptocytosis, hereditary spherocytosis, Southeast Asian ovalocytosis
4.1	erythrocyte membrane protein band 4.1 (EPB41)	66	1p33-p34.1	membrane skeleton	spectrin, b-actin, GPC	peripheral	hereditary elliptocytosis
4.2	pallidin	77	5q15-q21	binds cytoskeleton to lipid bilayer	band 3, ankyrin	peripheral
4.9	dematin / EPB49 (a, b)	43+46	8p21.2-21.1	actin-binding protein	b-actin, lipids	peripheral
4.9	p55 / MPP1	53	Xq28		protein 4.1, GPC	peripheral
5	b-actin	42	7pter-q22	membrane skeleton	spectrin, protein 4.1	peripheral
5	tropomodulin	41	9q22	blocks growth of actin filaments	tropomyosin, b-actin	peripheral
6	GADPH	36	12q13		band 3	peripheral
7	stomatin	32				integral	hereditary stomatocytosis
7	tropomyosin	28	1q31	stabilization of actin filaments	b-actin	peripheral
8	protein 8	23				peripheral
GPA	glycophorin A (GYPA) / MNS blood group	14	4q31	receptor for Plasmodium falciparum	negatively-charged phospholipids	integral	none
GPB	CD235b / glycophorin B	8	4q31	anchor for membrane skeleton	protein 4.1	integral	none
GPC	CD235R / glycophorin C (GYPC)	14	2q14-q21		protein 4.1, p55	integral	hereditary elliptocytosis
GPD	glycophorin D	11	2q14-q21			integral	hereditary elliptocytosis
GPE	glycophorin E (GYPE)	6	4q31			integral

Normal values :
• erythrocyte or red blood cell count (RBC) (blood that has been diluted in an isotonic solution, done with an automatic counter such as a flow cytometer) = 3^.10⁹RBCs / weight Kg ; 98% of total blood cells
• 4.5÷5.9 ^. 10⁶ RBCs / mL blood in males
• 4.1÷5.1 ^. 10⁶ RBCs / mL blood in females
• average life : 120 days (established with ⁵¹Cr release test).
• hematocrit (HCT) : 42÷50% in males ; 36÷45% in females
• microhematocrit : a hematocrit determination done on an extremely small quantity of blood, by use of a capillary tube and a high speed centrifuge.
• hemoglobin (HGB) : 13÷17.5 g / dL whole blood in males, 12÷15.5 g / dL in females. So 21 (in males) or 19 (in females) mL of O₂ can be bound for every dL of whole blood.
• hemoglobin distribution width (HDW) : 2.3÷3.2 g / dL whole blood
• red blood cell or erythrocyte indices :
• mean corpuscular volume (MCV) : 10 ^. HCT / RBC = 80÷95 fL=mm³
• mean corpuscular hemoglobin (MCH) : 10 ^. HGB / RBC = 27.5÷33 pg per red cell
• mean corpuscular hemoglobin concentration (MCHC) (assuming plasma hemoglobin is very low) : 100 ^. HGB / HCT = 33÷35 g / dL RBC (~ 34 % ^v/_v)
• RBC distribution width (RDW)
• Price-Jones curve : a frequency distribution curve of erythrocyte diameters, calculated electronically with a Coulter counter, a flow cytometer, or a similar instrument; it can detect conditions such as macrocytic anemia and microcytic anemia
• standard deviation (SD) : width of the distribution curve when frequency = 20% : 38.7÷45.1 fL=mm³
• coefficient of variation (CV) : 100 ^. SD / MCV = 11.5÷14.5 %
• volume of red blood cells (VRBC) is used routinely in the diagnostic work-up of polycythemia, in assessing the efficacy of erythropoietin administration, and to study factors affecting oxygen transport. VRBC, ⁵¹Cr and MHb, CO are the best measures for research on blood-related changes in oxygen transport. With care, VRBC, Evans is suitable for clinical applications of blood-volume measurement. Techniques based on^ref :
• Evans blue (VRBC, Evans)
• ⁵¹chromium-labelled red cells (VRBC, ⁵¹Cr)
• carbon-monoxide rebreathing (VRBC, CO)
• hemoglobin mass with the carbon-monoxide method (MHb, CO)
• erythrocyte sedimentation rate (ESR) of whole blood + anticoagulant : the rate at which erythrocytes precipitate out from a well-mixed specimen of venous blood, measured by the distance the top of the column of erythrocytes falls in a given time interval under specified conditions. The traditional methods of calculating ESR are the :

	males	females
Wintrobe method : EDTA-anticoagulated whole blood is placed in a Wintrobe hematocrit tube	1-13 mm / hr	1-20 mm / hr
Westergren method (the most common method) : 4 volumes of whole blood are mixed with 1 volume of sodium citrate anticoagulant-diluent solution and placed in a Westergren tube graduated in millimeters from 0 to 200, filling to the 0 mark	15 mm / hr	20 mm / hr

The tube is left standing undisturbed in a vertical position, and the fall of the level of RBCs in 1 hour is recorded in mm/hr. The volume of packed RBCs can then be determined using the same tube
• Katz index (KI) = [(mm in first hr) + (mm in second hr) / 2] / 2
ESR increases during :
• tissue necrosis
• anemias
• pregnancy
• obesity
• elevated levels of plasma proteins which decrease the zeta potential on erythrocytes by dielectric shielding and thus promote rouleau formation
• monoclonal gammopathy
• hypergammaglobulinemia due to inflammatory disease
• hyperfibrinogenemia
• active inflammatory disease
• zeta sedimentation ratio (ZSR) : a measurement comparable to the erythrocyte sedimentation rate, except that it is unaffected by anemia. The packed-cell volume (zetacrit) of a blood specimen is calculated by centrifuging the specimen in a Zetafuge, a specially designed instrument that produces controlled cycles of compaction and dispersion and allows rouleaux to form and sediment rapidly. The zetacrit divided into the true hematocrit gives the zeta sedimentation ratio.
• RBC protoporphyrin :
• 6-24 months : 79.79 +/- 14.3 mg/dL_RBC
• > 2 years : 58.6 +/- 13.6 mg/dL_RBC
Blood group or type : an allotype (or phenotype) of erythrocytes defined by one or more cell surface antigens that are under the control of allelic genes. Antigenic determinants irregularly incite allotypic and sometimes xenotypic immune responses. Human blood groups are identified by agglutination supported by specific human or animal antisera and by lectins extracted from certain plants. An abbreviated classification of human blood groups is given in the accompanying table. Any of certain other characteristics or traits of a cellular or fluid component of blood, considered as the expression (phenotype or allotype) of the actions and interactions of dominant genes; used in medicolegal and other studies of human inheritance. Such characteristics include the antigenic groupings of erythrocytes, leukocytes, platelets, and plasma proteins.
• high frequency blood group : a group containing over 99% of individuals, who have a type of erythrocyte antigens called public antigens.
• low frequency blood group : any small group that has erythrocytic antigens found in < 1% of the population (low frequency or private antigens).
Blood group system : antigenic determinants (symbols within parentheses are those of alternative nomenclatures. Antigenic determinants are systematized according to observed and assumed independent assortment of their responsible genes. Within many systems, alleles are responsible for differing combinations of antigenic determinants)
• ABO blood group : the major human blood group system, dependent on the presence or absence of A and B antigens, which are largely glycolipids on the cell membrane. The gene for A is responsible for synthesis of N-acetyl-a-D-galactosaminyl transferase, whereas that for B is responsible for a-D-galactosyl transferase. Either A or B is created when one of these hexasaccharides is positioned by a specific transferase in 1=>3 linkage to the b-D-galactose of an H-active oligosaccharide. Type O occurs when neither transferase is present or, very rarely (Bombay phenotype), when H antigen or substance (the precursor of the A and B blood group antigens. Normal type O individuals lack enzymes to convert H antigen to A or B antigens. Those individuals having the rare Bombay phenotype lack the ability to make H antigen and thus are phenotypically type O whether or not they possess A or B genes) does not exist. When both transferases are present, type AB results. Differences in degree of transferase activity are determined at the same locus: weak transferase gives rise to weak antigens (A₂, A₃A_x, B₃B_x). Similar oligosaccharides, especially in bacterial cell walls, immunize persons lacking A or B so that their serum contains anti-A or anti-B activity. A and B antigens are on the mucopolysaccharides of secretors; persons with dominant genes have H-active mucoids. A [subgroups A₁, A₂, A₃, A_m, A_o, A_x, A_int, A_end, A_finn, A_e1, A_bantu], B [subgroups B₃, B_x, B_e1]
• Auberger blood group : a blood group consisting of the erythrocytic antigen Au^a, related to the Lutheran blood group
• Bg blood group : a blood group consisting of the erythrocytic HLA antigens Bg^a, Bg^b, Bg^c, DBG, Ho, Ho-like, Ot, and Sto
• Yt or Cartwright blood group : a blood group consisting of the erythrocytic antigens Yt^a and Yt^b
• Colton blood group : a blood group consisting of erythrocytic antigens Co^a, Co^b
• Cost-Sterling blood group : Cs^a, Yk^a
• Cromer blood group : a blood group consisting of erythrocytic antigens Cr^a, Tc^a, Tc^ab, Dr^a, Es^a, WES^b, UMC, and IFC, which are located on the membrane protein called decay accelerating factor (DAF).
• Diego blood group : a blood group consisting of the erythrocytic antigens Di^a and Di^b, determined by allelic genes (SLC4A1 / erythrocyte membrane protein band 3 / CD233). Di^a is most frequent in South American Indians, Japanese, and Chinese
• Dombrock blood group [from the name of the propositus patient first observed in 1965] : a blood group consisting of the erythrocytic antigens Do^a and Do^b, most common in people of European descent
• Duffy blood group : a blood group consisting of the erythrocytic antigens Fy^a (Fy1), Fy^b (Fy2), Fy^ab (Fy3), Fy4, Fy5, determined by allelic genes. Amorphic genes are common in individuals of African descent.
• Gerbich blood group : a blood group consisting of the erythrocytic antigens Ge 1, Ge 2, and Ge 3 (corresponding to glycophorin C and E); although rare in most parts of the world, it has been found often in Papua New Guinea
• H blood group : a blood group consisting of antigen H
• Bombay phenotype : a rare blood phenotype produced by the interaction of genes of the ABO blood group and a rare recessive gene at a different locus, resulting in a complete lack of H antigen; cells of individuals with this phenotype lack A, B, and H antigens, and their serum contains anti-A, anti-B, and anti-H antigen. Associated with leukocyte adhesion deficiency type 2
• Ii blood group : a high frequency blood group involving receptors of most cold reactive hemagglutinins; it is expressed most strongly on cord blood cells : I, I^D, I^F, I^T, i
• Kell blood group : a blood group consisting of multiple erythrocytic antigens, especially 3 pairs of alternates, determined by complex genes at one locus, including an amorph; also regulated by the X chromosome, it is associated with sex-linked chronic granulomatous disease. One antigen, K6, is more frequent in people of African descent. K1 (K), K2 (k), K3 (Kp^a), K4 (Kp^b), K5 (Ku), K6 (Js^a), K7 (Js^b), K8 (kw), K9 (K1) K10 (U1^a), K11 (Côté), K12 (Bøk), K13 (Sgro), K14 (San), K15 (Kx), K16 (K-like), K17 (Wk^a), K18, K19, Kp^c
• McLeod phenotype : a rare blood phenotype with X-linked inheritance in which several antigens of the Kell blood group are weakly expressed; affected individuals sometimes have an anemic condition called McLeod syndrome
• Kidd blood group : a blood group consisting of Jk^a (Jk1), Jk^b (Jk2), and Jk^ab (Jk3) antigens, determined by allelic genes; amorphic genes are most common in those of East Asian descent
• Knops blood group : a blood group consisting of antigens Kn^a, Kn^b, McC^a, Sl^a, and Yk^a, which are located on complement receptor type 1.
• Lewis blood group : a blood group determined by plasma glycolipids that adhere to erythrocytic surfaces. It is based on dominant independent Le genes, but interacts with the H precursor oligosaccharides of A and B. Whereas le/le provides the “double negative” blood type Le(a-b-), Le without H gives rise to Le^a, i.e., blood type Le(a+b-), and that with H gives rise to Le^bH, i.e., blood type Le(a-b+). Le^a (Le1), Le^b (Le2), Le^c (Le5), Le^d, Le^x (L^ab, Le3), Mag (Le4)
• Lutheran blood group : a complex blood group system consisting of antigens Lu^a and Lu^b; it somewhat resembles the Kell group in having pairs of alternative antigens and amorphic genes, but is also subject to a dominant independently segregating repressor. Lu^a (Lu1), Lu^b (Lu2), Lu^ab (Lu3), Lu4, Lu5, Lu6, Lu7, Lu8, Lu9, Lu10, Lu11, Lu12, Lu13, Lu14 (Sw^a)
• MNSs blood group : a complex blood group system consisting principally of two pairs of antigens determined by closely linked genes (crossovers have been observed, but rarely). M and N, determined by allelic genes, depend on sialic (neuraminic) acid residues. S and s are also determined by allelic genes, and an amorphic gene is common in blacks when another antigen (U) is missing. The system also includes numerous low frequency antigens. Cl^a, Far, He, Hill, Hu, M, M1, M^A, M^c, M^e, M^g, M^k, M^r, M^v, M^z, Mi^a, Mt^a, Mur, N, N^A, N^a, N₂, Ny^a, Ri^a, S, S₂, S^B, s, Sj, St^a, Sul, Tm, U, U^B, Vr, Vw, Z
• P blood group : a blood group system originally consisting of only P (now P1) antigen, but later found to include P2 (Tj^a), a very high frequency antigen, and P3 (P^K), a very low frequency antigen. P1 is most common in people of African descent (90%), less so in those of European descent (75%), and least in those of East Asian descent (30%)
• paragloboside : a biosynthetic precursor of the ABH and P1 blood group glycosphingolipids and of one class of gangliosides
• Rh blood group : the most complex of all human blood groups because the genes differ by determining different numbers of antigens (Rhesus (Rh) factors or antigens : any of numerous antigens (agglutinogens) that may be present on the membrane of erythrocytes and that determine the Rh blood group system; the most common ones are called (in one system) Rh 1, Rh 3, Rh 4, Rh 5, and Rh 21) and do so with remarkably different quality; > 40 antigens have been described to date. People of African descent show the greatest degree of diversity and East Asians the least. The major antigen, Rh1 (Rh0, D, or Rh0D), is highly immunogenic and before the development of passive immunization prophylaxis it was responsible for serious hemolytic disease of the newborn (HDN). 2 other pairs of alternative antigens are inherited with or without Rh1; these are Rh21 (rh^G or C^G) and Rh4 (hr' or c), and Rh3 (rh' or E) and Rh5 (hr¢ or e). The most common groups of antigens are R^-1,-3,-21 (in Caucasians), R^1,-3,-21 (in blacks), R^1,-3,21 (in East Asians and Caucasians), and R1,3,-21 (in East Asians and Caucasians). Another antigen Rh10 (hr^v, V) is common in blacks. Rh1 (D, Rh₀), Rh2 (C, rh´) Rh3 (E, rh´), Rh4 (c, hr´), Rh5 (e, hr´), Rh6 (ce, f, hr), Rh7 (Ce, Rhi), Rh8 (C^w, rh^wl), Rh9 (C^x, rh^x), Rh10 (V, ce^S, hr^v), Rh11 (E^w, rh^w2), Rh12 (G, rh^G), Rh13 (Rh^A), Rh14 (Rh^B), Rh15 (Rh^C), Rh16 (Rh^D), Rh17 (Hr₀), Rh18 (Hr), Rh19 (Hr^S), Rh20 (VS, e^s), Rh21 (C^G), Rh22 (CE), Rh23 (D^w), Rh24 (E^T), Rh25 (L^W), Rh26 (c-like), Rh27 (cE), Rh28 (hr^H), Rh29 (RH), Rh30 (D^cor), Rh31 (hr^B), Rh32, Rh33, Rh34 (Hr^B), Rh35, Rh36, Rh37, Rh38, Rh39, Rh40, Rh41, Rh42 (Ce^s)
• Scianna blood group : a blood group consisting of erythrocytic antigens Sc1 (formerly Sm) and Sc2 (formerly Bu^a)
• Stoltzfus blood group : Sf^a
• Vel blood group : a blood group consisting of the erythrocytic antigens Vel 1 and Vel 2
• Wright blood group : a blood group consisting of the erythrocytic antigens Wr^a and Wr^b
• Xg blood group : a blood group consisting of erythrocytic antigen Xg^a, which is determined by a gene on the long arm of the X chromosome
Antigenic determinants that depend on gene interactions :
• ABO/I
• P/ABO
• IP1, IP2 (ITaj), ITP1, iP1
• Fy5,
• P/I
• Xor/Duffy
• ILe^bh
• Rh25 (LW)
• Lewis/I
• Rh/L^W
• A₁Le^b(Seidler)
• Lewis/ABO
• Ih, IA, IB, iH Luke
Selected antigenic determinants not thus far associated with a blood group system :
• Chido-Rodgers blood group : a blood group consisting of antigens Ch^a (Gursha) and Rg^a, antigenic determinants of fragments of the C4 component of complement.
• Lan blood group : a blood group consisting of the erythrocytic antigen Lan.
• Sid blood group : a blood group consisting of those with extra amounts of the public erythrocytic antigen Sd^a, referred to as Sd(a++).
• 754
• An^a
• At^a
• Be^a
• Be^c
• Bi
• Big Charles
• Bpa
• Bra
• Bxa
• By
• Cad
• Car
• Chra
• Cip
• Coates
• Craig
• Dahl
• Donaviesky
• Dp
• Driver
• Duch
• E1
• En^a
• Evans
• Evelyn
• Fin
• Fuerhart
• Fuj
• Gf^a
• Gilbraith
• Gn^a
• Go^b
• Good
• Green
• Gy^a
• Hands
• Hen
• Heibel
• Hill
• Ht^a
• Hy
• Je^a
• Jn^a
• Jo^a
• Jo^b
• Jr
• Kam
• Kelly
• Ken
• Knops (Kn^a)
• Kosis
• Lev
• Lw^a
• McCall
• McCoy (McC^a)
• Man
• Mar
• Mo^a
• MZ443
• Nij
• Ola
• Orr
• Pea
• Pt^a
• Rd^a
• Reid
• Savior
• Sch
• Simon
• Skjelbred
• Ters
• Th^a
• To^a
• Todd
• Tr^a
• Ven
• Vennera
• Wb
• Weeks
• Wil
• Winbourne
• Wu
• Yh^a
• York (Yk^a)
• Za
Bibliography : Blood Groups and Red Cell Antigens by Laura Dean; Bethesda (MD): National Library of Medicine (US), NCBI; 2005 [free at NCBI BookShelf !]
Erythrocyte survival time :
• ⁵¹Cr release assays (gold standard)
• erythrocyte creatine is thought to be a sensitive parameter of erythrocyte age^{ref1, ref2, ref3, ref4}. In hemolytic patients, it has been demonstrated that erythrocyte creatine was closely correlated with the erythrocyte survival time, which was evaluated by the standard procedure involving ⁵¹Cr^ref. The young erythrocytes have higher creatine values than do the old cells^{ref1, ref2}
Erythrocatheresis occurs mainly by macrophages in Schweigger-Seidel sheaths (red pulp of spleen) : as senescent erythrocytes down-regulate CD47 expression, PTPNS1 / SIRPa (an-ITIM containing inhibitory receptor) can no longer prevent phagocytosis.

‡
• ghost cell / erythroclast / shadow cell : a degenerating or fragmented erythrocyte with no hemoglobin
+ M-CSF
• BFU-Meg => megakaryocytopoiesis. In bone marrow precursors are located in a position intermediate between that of WBC precursors and that of RBC precursors.
‡
+ M-CSF / CSF-1, EPO, IL-3, TPO, thrombopoiesis stimulating factor :
• CFU-Meg (PPo⁺)

‡
+ EPO, IL-3, TPO : TPO supports progenitor cell expansion, whereas CXCL12, FGF-4 enhance VCAM-1 and VLA-4-mediated localization of CXCR4⁺ megakaryocyte  progenitors with the bone marrow endothelial cells (BMECs) allows the progenitors to relocate to a microenvironment that is permissive and instructive for megakaryocyte maturation and thrombopoiesis^ref
• promegakaryoblast

‡
+ EPO, TPO, LIF, OsM, IL-1b, IL-3, IL-4, IL-6, IL-11, thrombopoiesis stimulating factor, megakaryocyte maturation factor (inhibited by IFN-a, IFN-g, TGF-b, CXCL4 / PF4 and b-thromboglobulin). GABP-a, an Ets transcription factor, is required for the early maturation of megakaryocytes and controls the expression of multiple megakaryocyte-specific genes aIIb and cMpl^ref.
• megakaryoblast (CXCL4 / PF4⁺, CD42b / glycocalicin / gpIb_a⁺42c / gpIb_b⁺, vWF⁺, fibrinogen ABg⁺, factor V⁺, thrombospondin⁺, AcP⁺, 4n)


‡
+ IL-6, IL-7, TPO : endomitosis followed by nuclear fusion => autopolyploidy.
• promegakaryocyte or immature megakaryocyte (from 8n up to 128n).


It hasa-granules, which contain :
• CXCL4 / PF4 / antiheparin factor
• b-thromboglobulin => connective tissue-activating peptide III (CTAP-III) (normal serum levels : 1.6-4.8 µM^ref) => CXCL7 / LDGF-PBP / NAP-2 / CTAP-II
• fibronectin
• vitronectin / protein S
• thrombospondin
• fibrinogen ABg
• vWF
• factor V
• factor VIII
• a subunit of factor XIII
• a₁-antitrypsin / a₁-antiplasmin
• PA-I
• PDGF
• PD-ECGF
• CXCL4 / PF4
• TGF
• HGF
• EGF
• PrP^C (platelet activation led to the transient expression of PrPC on the platelet surface and its subsequent release on both microvesicles and exosomes)^ref
‡
• reserve megakaryocyte

‡
down-regulation of survivin => continue to progress through the cell cycle but skip late stages of mitosis to become polyploid cells^ref. Fli-1, an Ets transcription factor, is required for the late maturation of megakaryocytes and controls the expression of multiple megakaryocyte-specific genes^ref.
• mature megakaryocyte (CD42a / p17^gpIX+42b / glycocalicin / gpIb_a⁺42c / gpIb_b(containing HPA-2a / Ko^b and HPA-2b / Ko^a Ags)⁺42d / p82^gpV+)


In 1906, James Wright (Wright JH. The origin and nature of the blood plates. Boston Med Surg J. 1906;154: 643-645) published a morphology study in which he argued that circulating plates (then called platelets) are derived from megakaryocytes, and that megakaryocytes release platelets from elongated, cytoplasmic projections that extend into the marrow sinusoids. Following a series of mitotic cell divisions, megakaryocyte lineage cells enter an endomitotic cycle that generates large, polyploidy megakaryocytes. Cytoplasmic maturation at the end of the endomitotic phase includes the development of the demarcation membrane system (DMS). The DMS, first described by Yamada in 1957^ref, is an extensive membrane network located throughout the cytoplasm of the mature megakaryocyte, except for a thin, uninvolved cortical zone. Staining studies have shown that the DMS is contiguous with the extracellular space, making it effectively an extensive invagination of surface membrane, but its function remains unclear. One early popular theory, which is reflected in its name, is that the DMS "marks" preformed "platelet territories" within the megakaryocyte cytoplasm, which in turn led to the cytoplasmic fragmentation hypothesis for platelet formation. However, this is incompatible with proplatelet formation (thrombocytopoiesis) as the mechanism of platelet biogenesis. Using novel knock-in mice in which the fluorescent protein EYFP is introduced into the CD41 (integrin IIb) locus, the DMS serves as an extensive membrane reservoir to support the explosive proplatelet formation that marks the final hours of the mature megakaryocyte^ref. Key to their studies is a C-terminal myristoylation site that incorporates the EYFP reporter into cellular membranes. The EYFP signal localizes to the outer cell membrane of early megakaryocyte lineage cells, but it becomes diffusely localized throughout the cytoplasm in mature megakaryocytes in a pattern that mirrors the DMS. The EYFP-expressing internal membranes were found to be contiguous with the full outline of emerging proplatelets, and platelets circulating in the EYFP mice have surface EYFP fluorescence, data compatible with the DMS providing an extensive membrane reservoir for proplatelet formation. Using fluorescently tagged plextrin homology domain from PLC1 as a probe, the phospholipid PI-4,5-P₂, which typically resides in the plasma membrane of cells, localizes to the DMS in mature megakaryocytes. siRNAs suggest that the accumulation of PI-4,5-P₂ on the DMS membranes, along with DMS development and the associated increase in cell size, may all be facilitated by the lipid kinase PIP4K. PI-4,5-P₂ signaling from the DMS may trigger actin polymerization via the WASp/WAVE pathway. Reports over the past 5 years have significantly advanced our understanding of the structural and mechanical aspects of proplatelet formation^{ref1, ref2}. From each proplatelet arises more than one ...
‡ The generation of platelets from megakaryoctes is regulated ...
• in the steady state by a variety of cytokines and transcription factors, including thrombopoietin (TPO), GATA-1 and NFE2
• in the setting of stress thrombopoiesis, a pivotal event in the context of cytotoxic chemotherapy, by the transcription factor Scl, which acts upstream of NFE2^ref
• platelet / thromboplastin / Deetjen's body

They don't contain DNA (instead in lower Vertebrates they retain nucleus and are named thrombocytes) but just cytoplasmic enzymes for all metabolic cycles, mitochondria, peroxisomes and glycogen granules.
Bone marrow produces 100,000,000,000 platelets / die, as estimated with ⁷⁵Se-Met and ¹⁴C-5-HT incorporation tests. Normal valeus :
• platelet count (PLT) (either a direct platelet count with a hemacytometer and a microscope or an indirect platelet count in which the ratio of platelets to erythrocytes on a blood smear is determined and the number of platelets is computed from the erythrocyte count) = 1.5 ^. 10⁹ / weight Kg ; 172,000÷500,000 / mL blood ; 0.4 % total blood cells, although they are rather just cell fragments !) : 40,000 / mL are replaced each day
• t_1/2 : 2 days
• average life estimated with ⁵¹Cr or ¹¹¹In release tests = 8÷10 days
• plateletcrit (PCT) = 0.115÷0.32 %
• mean platelet volume (MPV) = PCT / PLT = 9.1÷12.3 fL
• platelet distribution width (PDW) = 10÷16 fL
Apart from a-granules, they also contain :
• d-granules / bull's eye or platelet dense body or granule : an electron-dense granule occurring in blood platelets that stores and secretes  ADP, ATP, GDP, GTP, PP_i, P_i, P, Ca²⁺, Mg²⁺, 5-HT, E and H)
• l-granules / lysosomes (containing AcP, arylsulfatase, cathepsin D, b-glucuronidase and b-galactosidase)
Platelet factors (PF) : factors important in hemostasis which are contained in or attached to the platelets; they act together with coagulation factors.
• platelet factor 1 (PF1) : adsorbed factor V from the plasma.
• platelet factor 2 (PF2) : an accelerator of the thrombin-fibrinogen reaction, attached to platelets.
• platelet factor 3 (PF3) : a lipoprotein, extracted from platelets, which contributes to the interaction of activated plasma coagulation factors IX and VIII to produce activated factor X as well as of activated factors X and V to cleave and activate prothrombin.
• platelet factor 4 (PF4) / CXCL4 : an intracellular protein component of blood platelets, capable of neutralizing the antithrombic activity of heparin in the fibrinogen-fibrin reaction and the inhibitory effect of heparin in the thromboplastin generation test. Normal serum level : 0.4-1.9 µM^ref.
Main membrane proteins are : CD14^-15^-23⁺31 / PECAM-1 / b₁ integrin / p130^gpIIa
+36 / thrombospondin receptor / p95^{gpIV / gpIIIb+}41 / a_IIb integrin / p136^gpIIb (containing HPA-3a / Bak^a / Lek^a and HPA-3b / Bak^b / Lek^b Ags)⁺42a / p17^gpIX+42b / glycocalicin / gpIb_a⁺42c / gpIb_b (containing HPA-2a / Ko^b and HPA-2b / Ko^a Ags)⁺42d / p82^gpV+43^lo46⁺49 / a₂ integrin / p153^gpIa (containing HPA-5a / Br^b and HPA-5b / Br^a / Hc / Zav Ags)⁺61 / b₃ integrin / p95^gpIIIa(containing HPA-1a / PI^A1 / Zw^a, HPA-1b / PI^A2 / Zw^b, HPA4a / Pen^a / Yuk^b and HPA-4b / Pen^b/ Yuk^a Ags)⁺69⁺MHC class I⁺, P2Y₁, P2Y₁₂, P2_T, TP, PAR-3, PAR-4, 5-HT_2A, CD88 / C5aR, AB0, Le, I and P blood group Ags, Gov^a and Gov^bAgs, p148^gpIc.
TLR2 and TLR4 are expressed at low levels on platelets, but TLR agonists LPS and Pam3CSK4 have no direct effect on platelet activation and that platelet TLRs may be a remnant from megakaryocytes^ref.
Receptors for collagen and adenosine diphosphate (ADP) undergo synergistic signaling to integrin a_IIbb₃ for platelet aggregation^ref. CalDAG-GEFI integrates signaling for platelet aggregation and thrombus formation^ref

+ ? :
• precursor of myeloid, B and T cells (p-MTB) or common myelo-lymphoid progenitor (CMLP)

(CD16a / FcgRIIIA⁺32 / FcgRII⁺)
‡
+ ? :
• precursor of myeloid and B cells (p-MB)

(please note all stages in B cell lineage (until mature B-cell stage) are potent APCs and retain the ability to show dendritic morphology, up to convertion into macrophages). The decision to become a B cell or macrophage depends on which transcription factor between C/EBP (C/EBP-a and C/EBP-b : myeloid commitment) and PAX5 / BSAP (B-cell commitment) becomes dominant^ref.

or

+ ? :

• precursor of myeloid and T cells (p-MT)

(please note early thymocytes are able to generate dendritic cells)
• lymphopoiesis => B and T cells(they appear as WBCs, too). Normal blood concentration : 0.16÷0.32 ^. 10⁹ lymphocytes / weight Kg ; 0.88÷4.52 ^. 10³ lymphocytes / mL blood ; 0.32÷0.64 % total blood cells ; 20÷40 % total WBCs.
• small lymphocytes are the most common, ranging in size from 6 to 10 µm. The nucleus is usually round or slightly oval, occasionally showing a small indentation due to the adjacent centrosome. Except in the smallest cells, the nucleus is about 7 µm in diameter, a size that has been convenient for estimating the size of the surrounding erythrocytes. Nuclear chromatin stains a dark reddish-purple to blue with large dark patches of condensed chromatin. The nuclear cytoplasm ratio is 5:1 to 3:1, and the cytoplasm is often seen only as a peripheral ring around part of the nucleus


• myelopoiesis => leukocytes or white blood cells (WBC)

‡
+ GM-CSF
• colony-forming unit–granulocyte-macrophage (CFU-GM) / colony-forming unit–culture (CFU-C) : a precursor cell in the granulocytic series that can grow into a myeloblast in the presence of appropriate stimulators in vitro (CD13⁺33⁺34⁺, MHC II⁺). Within 10 days a myeloblast becomes a granulocyte.

‡
+ G-CSF, IL-3, IL-6, GFI1, PU.1 / SPI1, C/EBPa, C/EBPe :
• CFU-G. The myeloid commitment of hematopoietic progenitors is characterized by the progressive loss of CD34 expression accompanied by the acquisition of expression at high levels of CD33, a member of the sialic acid-binding immunoglobulin-like lectin (Siglec) family. CD33 expression continues along the myelomonocytic pathway of differentiation. Although this expression is maintained on monocytes and its 2 terminal stages of differentiation (ie, tissue macrophages and myeloid DCs), a down-regulation is observed on granulocytes, which, however, continue to express CD33 but only at low levels^{ref1, ref2}

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• myeloblast (CD33⁺34⁺, MHC II⁺) : a myeloblast is the earliest recognizable cell of the neutrophilic series. It is usually 15-20 µm in diameter. The nucleus is round or slightly oval and occupies about 4/5 of the cell. The nucleus is composed of very fine nonaggregated chromatin that stains light blue to reddish-purple with Wright's stain. 2-5 distinct nucleoli are usually present. The nucleus is often bordered at one side by a distinct perinuclear zone. The cytoplasm is usually moderately blue in color, smooth, and nongranular. It occupies one-fifth of the cell. It may be difficult to distinguish myeloblasts from other blasts in the peripheral blood unless one uses special stains or infers their identity from the presence of other immature cells of the same line. A myeloblast can be distinguished from a promyelocyte by its lack of cytoplasmic granulation. It develops into a granulocyte within 10 days


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• neutrophil promyelocyte (CD11b⁺13⁺15⁺33⁺34⁺73 / ecto-5'-nucleotidase⁺, MHC II⁺) :


• primary, azurophilic or aspecific granules (~ lysosomes) contain :
• cathepsin G
• proteinase 3
• myeloperoxidase (MPO : used for histochemical identification !)
• b-galactosidase
• acidic hydrolases
• b-glucuronidase
• acid phosphatase (AcP)
• ...
• arylsulfatase B
• cationic proteins
• defensins
• azurocidin
• BPI / CAP57
• lysozyme
• human leukocytary elastase (HLE) / medullasin
CD63, CD68, and stomatin (STOM) are the only membrane proteins identified so far
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• myelocyte (CD11b⁺13⁺15⁺33⁺34^-, MHC II^-) : disappearance of