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At least 19 recordsLinked to original sources
PubMed1979

Human megakaryocytes. I. Characterization of the membrane and cytoplasmic components of isolated marrow megakaryocytes.

Human marrow megakaryocytes have been isolated with high purity and yield by processing marrow cells sequentially through density centrifugation and velocity sedimentation. Analysis of the isolated cells for various platelet-associated components by immunofluorescence demonstrated that fibrinogen, plasma factor VIII antigen (factor VIII:AGN) platelet myosin, platelet glycoproteins I and III are present on the membrane and in the cytoplasm of over 90% of marrow megakaryocytes. Parallel studies of human and mouse megakaryocytes and platelets for IgG receptor (FcR), complement receptor type one (CR1) (C3b receptor), complement receptor type two (CR2) (C3d receptor), and Ia antigen by fluorescence and (or) rosette formation methods were performed. FcR were present on most human megakaryocytes and platelets. The Ia antigen was detected on a proportion (10-15%) of human megakaryocytes but it was undetectable on human platelets. CR1 was found on 20-40% of mouse megakaryocytes and also on a proportion of mouse platelets. These differentiation markers may be of use in monitoring megakaryocyte maturation.

Animals↗
PubMed1978

Studies on splenic megakaryocytes. I. Recovery of megakaryocytes from the splenic venous outflow.

Spleens from 14 healthy adult cats were isolated and perfused with cell-free Ringer's solution. Histological studies on the splenic megakaryocytes, as well as a search for splenic megakaryocytes in samples of the splenic venous outflow, were performed. The samples of the splenic venous outflow contained a lower cellular concentration when perfusion was prolonged, but the number of megakaryocytes appearing in the venous samples increased. At least five different forms of megakaryocytes were observed from smears of the venous outflow: (i) unipolar, (ii) multipolar, (iii) foamy, (iv) mobile and (v) nuclear types. These types were also found in splenic venous outflow of dogs, kittens and bled cats. It is suggested that not only platelets but also the larger megakaryocytes can pass out of the splenic circulation. This may imply that megakaryocytes can be culled, matured in the splenic red pulp, and ultimately released into peripheral blood.

Animals↗
PubMed1977

Chronic megakaryocytic-granulocytic myelosis--an electron microscopic study. I. Megakaryocytes and thrombocytes.

The fine structure of the bone marrow in chronic megakaryocytic-granulocytic myelosis (CMGM) was studied in 5 nontreated patients to investigate possible malignant proliferation of megakaryocytes and the role of megakaryopoiesis in fibrillogenesis, terminating in osteomyelofibrosis. In comparison with normal megakaryopoiesis there is an enormous increase of the megakaryocytic cell line and many immature and atypical forms are seen. Most conspicuous are microforms, nuclear-cytoplasmic disorganization and nuclear inclusions. Asynchrony of maturation causes abnormal thrombocytogenesis with premature detachment of platelets resulting in immature and peculiar giant forms of thrombocytes. Besides megakaryocytes appearing superficially normal the maturation anarchy of many cells is so severe that by analogy with observations in other leukaemic cells these abnormalities are thought to be representative of a malignant growth. Moreover, there is a striking accumulation of microfibrils and single collagen fibres around megakaryoblasts. Since these cells contain all those organelles commonly associated with fibre production the initial step for fibrillogenesis may therefore arise from the megakaryoblasts prior to platelet release, or any fibroblast proliferation.

Blood Platelets↗
PubMed1977

The functional capacity of guinea pig megakaryocytes. I. Uptake of 3H-serotonin by megakaryocytes and their physiologic and morphologic response to stimuli for the platelet release reaction.

The functional capacity of guinea pig megakaryocytes was tested by studying their ability to concentrate serotonin and their response to agents which trigger the platelet release reaction. Megakaryocytes can concentrate 3H-serotonin as demonstrated by autoradiography after exposure to 0.5 muM 3H-serotonin and by quantitative measurement of isotope incorporation within 60 minutes. Uptake of isotope is rapid and linear within the first 30 minutes and tapers off between 30 and 60 minutes. Incorporation of isotope is diminished during exposure to cold, 2 muM reserpine, and 20 muM imiprimine. The following triggering agents: 10(-5) to 10(-3) M ADP, 1 to 100 units of thrombin, 10(-5) to 10(-3)M epinephrine, and 1 to 12 muM ionophore A23187 all produce significant release of stored 3H-serotonin. In the presence of ADP, albumin and serum completely inhibit the release of serotonin. Scanning microscopic studies show that coincident with serotonin release the triggering agents produce marked changes in cell shape. Transmission electron microscopy on these cells shows that there is the appearance of a prominent contraction zone, which is composed of microfilaments, and also variable diminution of cytoplasmic granules. The specifically induced serotonin release from megakaryocytes coupled with shape change and evidence of cell contraction produced by certain agents demonstrate one aspect of the functional similarly between megakaryocytes and platelets.

Adenosine Diphosphate↗
PubMed1977

Freeze-fracture of the normal and pathological megakaryocyte lineage in chronic megakaryocytic-granulocytic myelosis.

Freeze-fracture and thin sections were performed on human bone marrow of chronic megakaryocytic-granulocytic myelosis (CMGM) to study the three-dimensional fine structure and maturation of normal and atypical megakaryocytes and thrombocytes. In the many normally maturing megakaryocytes the development of the demarcation membrane system (DMS) was best investigated by comparison of thin sections with freeze-fracture replicas. The DMS shows no connections with the Golgi apparatus or rough-surfaced endoplasmic reticulum, but originates from tubular infoldings of the plasma membrane. These infoldings are always in continuity with the extracellular space and form an intracellular membranous pool by branching and coalescing of flattened tubules from which finally the perforated cisternae of the DMS arise. Freeze-fracture of the normal thrombocytes confirms earlier findings. The abnormal giant platelets seen in CMGM display extensive areas of smooth membranes of a spongy structure consisting of dense tubules surrounded by the labyrinth of the surface-connected system. Their physiological significance in these atypical platelets remains unsolved.

Blood Platelets↗
PubMed1979

[Acute myeloid leukemia with megakaryocytic predominance and malignant megakaryocyte proliferation. Apropos of 3 cases].

Three cases of acute myeloid leukemia with megakaryocyte predominance are reported. In the first case megakaryocytosis was particularly evident in bone marrow, liver and spleen. In the second case high content of megakaryocytes was observed in the bone marrow and spleen, during the preleukemic phase only. Third case exhibited a strong predominance of megakaryocytes exclusively within the bone marrow. The characteristics of such observations and their nosologic position within myeloproliferative disorders group are discussed, with special reference to modern views concerning myeloid leukemia.

Adult↗
PubMed2025

HIV-1 inhibits IFITM3 expression to promote the infection of megakaryocytes.

Despite an undetectable plasma viral load as a result of antiretroviral therapy, HIV-1-infected individuals with poor immune reconstitution harbor infectious HIV-1 within their platelets. Megakaryocytes, as platelet precursors, are the likely cellular origin of these HIV-1-containing platelets. To investigate the mechanisms that allow megakaryocytes to support HIV-1 infection, we established in vitro models of viral infection using hematopoietic stem cell-derived megakaryocytes and the megakaryocytic MEG-01 cell line. We observed HIV-1 DNA provirus integration into the megakaryocyte cell genome, self-limiting virus production, and HIV-1 protein and RNA compartmentalization, which are hallmarks of HIV-1 infection in myeloid cells. In addition, following HIV-1 infection of megakaryocyte precursors, the expression of interferon-induced transmembrane protein 3 (IFITM3), an antiviral factor constitutively expressed in megakaryocytes, was inhibited in terminally differentiated HIV-1-infected megakaryocytes. IFITM3 knockdown in MEG-01 cells prior to infection led to enhanced HIV-1 infection, indicating that IFITM3 acts as an HIV-1 restriction factor in megakaryocytes. Together, these findings indicate that megakaryocyte precursors are susceptible to HIV-1 infection, leading to terminally differentiated megakaryocytes harboring virus in a process regulated by IFITM3. Megakaryocytes may thus constitute a neglected HIV-1 reservoir that warrants further study in order to develop improved antiretroviral therapies and to facilitate HIV-1 eradication.

Humans↗
PubMed1979

Growth of human megakaryocyte colonies in culture from fetal, neonatal, and adult peripheral blood cells: ultrastructural analysis.

Megakaryocyte colonies can be grown in culture from human blood cells and fetal liver cells in plasma clot containing erythropoietin. Megakaryocyte progenitors were found in a fraction of mononuclear cells isolated by Ficoll density gradient centrifugation from adult, neonatal, and fetal blood. Megakaryocytes were identified by their morphology and particularly by their polylobulated nucleus when examined by light microscopy. The megakaryocytic nature of large cells was clearly confirmed by the presence of platelet peroxidase, demarcation membranes, and alpha-granules detected by electron microscopy; in addition mature small megakaryocytes were recognized. Megakaryocyte colonies were seen after 9 days of culture and consisted of 2 to 20 cells. The colonies were pure or mixed with the burst erythroblasts. The mixed colonies were numerous in fetal and neonatal cultures, while pure megakaryocyte colonies were seen three times more frequently in those from adult blood. The total number of colonies was also much lower in adult cultures. In colonies derived from neonatal and fetal cells, megakaryocytes often reached a more complete maturation than in those from the adults, proceeding as far as platelet shedding. This study demonstrates for the first time that a megakaryocyte committed cell present in human blood can develop megakaryocyte colonies in culture.

Blood Cells↗
PubMed1977

Culture in vitro of isolated guinea pig megakaryocytes: recovery, survival, morphologic changes, and maturation.

Isolated guinea pig megakaryocytes were maintained in liquid cultures for up to 4 days. Megakaryocytes were incubated in siliconized glass vials in Dulbecco's Modified Eagle Medium with 5%-10% guinea pig serum and 2.3% bovine serum albumin. Cultured megakaryocytes did not adhere to glass vials and were almost entirely recovered by aspiration. No reproduction or cell division of megakaryocytes occurred. A small decline in viability occurred promptly on placing the freshly isolated cells in culture medium and could be attributed to reexposure to calcium. On incubation there was little further cell death. Up to 2 days in culture the megakaryocytes remained morphologically intact and appeared similar to megakaryocytes in situ. Megakaryocytes matured in culture with a loss of cytoplasmic basophilia, an increase in granule content, and progressive changes in nuclear configuration. The most mature megakaryocytes developed pseudopod formation but large-scale platelet liberation was not seen. The ability to culture megakaryocytes in vitro will allow more extensive biochemical and physiologic studies of this cell than previously possible.

Animals↗
PubMed1976

Isolation of intact megakaryocytes from guinea pig femoral marrow. Successful harvest made possible with inhibitions of platelet aggregation; enrichment achieved with a two-step separation technique.

Methods have been devised to harvest megakaryocytes from guinea pig femoral marrow and to isolate them in high yield. When marrow tissue was disaggregated the megakaryocytes underwent degenerative changes characterized by the loss of cytoplasmic granules and alterations in membrane topography, similar to the changes seen in aggregating platelets. These morphologic changes were interpreted to mean that megakaryocytes possessed functional attributes of platelets. The use of agents which inhibit platelt aggregation (0.38% sodium citrate. 10(-3) M adenosine, and 2 x 10(-3) M theophylline) in a medium free of bivalent cations prevented these changes. This solution resulted in both an excellent morphologic preservation and a significantly increased recovery of megakaryocytes from marrow tissue. A two-step purification of the intact megakaryocytes was carried out on the basis of their low density and large size, with equilibrium density gradient centrifugation followed by velocity sedimentation. This sequence gave approximately a 100-fold enrichment of megakaryocytes, significantly better than that achieved with either method alone. These techniques for harvesting and concentrating megakaryocytes make it possible for the first time to study megakaryocytes in vitro.

Adenosine↗
PubMed1978

Occurrence of megakaryocytes in various vessels and their retention in the pulmonary capillaries in man.

A total of 17 patients with hypertension undergoing renal vein or adrenal vein catheterization were investigated in order to ascertain the number of megakaryocytes in blood from the inferior vena cava, the femoral artery and a cubital vein. On an average 11.9, 3.8, and 4.5 megakaryocytes per ml were found, respectively. In blood from the inferior vena cava, 30% of the megakaryocytes had copious cytoplasm, while megakaryocytes in arterial and cubital venous blood had sparse or no visible cytoplasm. It was demonstrated that 2/3 of the megakaryocytes were retained in the pulmonary circulation and that at least 70% of the platelets could derive from megakaryocytes in central venous blood or the pulmonary circulation. It was found that megakaryocytes pass through a life cycle in which the differentiation take place in the bone marrow, platelet release occurs mainly in central venous blood and in the pulmonary circulation and the destruction of the megakaryocyte nucleus take place outside the bone marrow, especially in the pulmonary circulation.

Adult↗
PubMed1978

Circulating megakaryocytes in blood from the antecubital vein in healthy, adult humans.

A total of 21 healthy, adult men and 30 healthy, adult women aged 21-73 years were investigated for circulating megakaryocytes in the antecubital vein using the saponinhaemolysis leucoconcentration technique. In the males, the number of megakaryocytes in 1.5 ml blood varied from 1 to 20, the arithmetic mean being 10 megakaryocytes. In the females the number varied from 0 to 29 megakaryocytes and the arithmetic mean was 7.6 megakaryocytes. No significant difference was found between the values for the two sexes. There was no correlation between the number of megakaryocytes and platelets or leucocytes. 99% of the observed megakaryocytes were naked nuclei or had sparse cytoplasm and 1% had copious cytoplasm. Megakaryocytes without copious cytoplasm may be regarded as normally occurring cells in the peripheral venous blood.

Adult↗
PubMed1979

Megakaryocyte colony formation from human bone marrow precursors.

We report the growth in plasma clot culture of megakaryocyte colonies from adult bone marrow cells with the use of four different sources of erythropoietin (Ep) as the stimulating factor. A major proportion of the megakaryocyte colonies (75%) were pure, while the others were mixed, involving erythroblasts and megakaryocytes. Ultrastructural studies have shown that the maturation of megakaryocytes was essentially normal and that platelet shedding occurred. Megakaryocyte colony formation required a large number of plated cells (greater than 3 X 10(5)/ml). In the absence of erythropoietin, rare spontaneous megakaryocyte colonies could be observed, while no erythroid colonies were present. However, erythropoietin induced a fivefold increase in the total number of colonies. These data suggest that erythropoietin is involved in the differentiation of human megakaryocytes, but that it does not act alone, since another factor related to the number of seeded cells appears essential for the formation of human megakaryocyte colonies.

Animals↗
PubMed1979

Response of pulmonary (circulating) megakaryocytes to experimentally induced consumption coagulopathy in rabbits.

The effects of slow temporary infusion of a tissue thromboplastin solution into the superior vena cava on pulmonary as well as circulating megakaryocytes were studied in 40 rabbits (2-48 hours after infusion) and related to 6 noninfused and 7 infused with normal saline. This is a simple and specific method of inducing a fall in blood platelet levels and thereby an activation of thrombocytopoiesis and megakaryocytopoiesis. The induced intravascular coagulation is probably counterbalanced by an activated fibrinolysis allowing the animals to survive the infusion and thereby offering the possibility of studying the long-term effects. An increase to about 300% of the normal values in circulating as well as pulmonary megakaryocytes was found 20 and 24 hours after the onset of the infusions respectively. The number of circulating and pulmonary megakaryocytes, showing great individual variations, however, dropped to normal levels within 28 hours after onset of the infusions, which means that megakaryocytes remain detectable for less than eight hours in the lungs. No increase was found in pulmonary megakaryocytes in the control (saline infused) group. In our opinion the entrance of megakaryocytes from the bone marrow into the blood circulation in an incidental event, the number in the circulation reflecting the activity of megakarycocytopoiesis. This experiment supports our suggestion that intravascular coagulation is one of the major pathophysiological mechanisms leading to an increase in pulmonary megakaryocytes.

Animals↗
PubMed1978

Identification of young megakaryocytes by immunofluorescence and cytophotometry.

The DNA-content of fluoresceine-labeled platelet antigen containing cells of mouse bone marrow was measured. For immunofluorescence highly specific anti-mouse-platelet-serum and fluoresceine-conjugated antigammaglobuline was used, applying the "sandwich" technique. Three hundred panoptically identifable megakaryocytes served as control group. The DNA-polyploidization pattern of megakaryocytes and immunofluorescence positive cells was almost identical. However, among the immunofluorescence positive cells a considerable amount of cells showed DNA-values lower than 4c, whereas the megakaryocytes of the Pappenheim stained smears revealed no DNA-values lower than 4c. The percentages of diploid and tetraploid cells, respectively, was 6 and 7% compared with 0 and 1% of panoptically identifiable megakaryoctyes. The results suggest that young megakaryocytic cells with diploid and tetraploid DNA-values can be detected by immunofluorescence technique, indicating that the flow from the uncommited to the committed megakaryocytic precursor cell appears at this early stage of megakaryocyte production.

Age Factors↗
PubMed1976

Megakaryocytes and the heterogeneity of circulating platelets.

Megakaryocytes mature to the point of cytoplasmic disintergration in three principal ploidy classes: 8n, 16n and 32n. Cells of each of these ploidy classes have been identified, using both microdensitometry and measurement of cell volume and submitted to morphometric analysis. Mature megakaryocytes of the three ploidy classes have been shown to differ in the concentration of cytoplasmic constituents which would be expected to relate to the buoyant density of their platelet progeny. Density separated platelets have been similarly analysed. Light platelets correspond with the 32n megakarycytes and are more liberally endowed with surface connected canalicular system than the progeny of the common 16n megakaryocytes; it is proposed that they have functional characteristics related to this finding. Dense platelets, which are larger in size, correspond with 8n megakaryocytes and show a greater content of granules and mitochondria than platelets of average density. These platelets most probably show specialized function relating to release of granule constituents. Fragments of cytoplasm released from megakaryocytes represent one form of "megathrombocyte" equated by others with newly formed platelets. The differences in structure between these fragments and circulating platelets are emphasized; each such fragment must undergo further disintergration into a number of platelets. It is suggested that the heterogeneity of circulating platelets with respect to both size and density stems from the origin of platelets of varying density from the different populations of megakaryocyte and their release in the form of large cytoplasmic fragments rather than as platelets.

Animals↗
PubMed1979

Circulating megakaryocytes in patients with pulmonary inflammation and in patients subjected to cholecystectomy.

30 patients with pulmonary inflammation and 17 patients subjected to cholecystectomia à froid were investigated for circulating megakaryocytes in a cubital vein using the saponin-haemolysis leucoconcentration technique. The number of circulating megakaryocytes was significantly higher in patients with pulmonary inflammation than in healthy humans. In 15 patients with bronchitis, bronchopneumonia, and leucocytosis the arithmetic mean was 15.6 megakaryocytes per 1.5 ml blood (range 3 to 47). In 15 patients with bronchitis and a normal leucocyte count the arithmetic mean was similarly 14.1 megakaryocytes per 1.5 ml blood. After cholecystectomy a significant maximum increase in the number of circulating megakaryocytes to 3 times the preoperative value was found on the third postoperative day but not on the other postoperative days. Of the observed megakaryocytes 99% had only a narrow rim of cytoplasm or were naked nuclei.

Adolescent↗
PubMed1978

Regulation of proliferation of murine megakaryocyte progenitor cells by cell cycle.

The extent to which mouse megakaryocyte progenitor cells (colony-forming unit-megakaryocyte, CFU-M) can proliferate in semisolid cultures prior to endomitosis, and conditions that may regulate that differentiation step, were investigated. The proliferative capacity of CFU-M was determined by estimating the number of megakaryocytes per colony. A bimodal distribution was observed (modal values, 10-15 and 25-30 cells/colony), indicating that separate megakaryocyte progenitor cells may be biased in their capacity for proliferation versus endomitosis. Differences were observed in the cell cycle characteristics of CFU-M as determined in vivo and in vitro that suggest that maturation of CFU-M into megakaryocytes may be regulated within the marrow by control of the cell cycle of the megakaryocyte precursor cell.

Animals↗