Academic literature on the topic 'Megakaryocytes'
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Journal articles on the topic "Megakaryocytes"
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Levene, R. B., J. M. Lamaziere, H. E. Broxmeyer, L. Lu, and E. M. Rabellino. "Human megakaryocytes. V. Changes in the phenotypic profile of differentiating megakaryocytes." Journal of Experimental Medicine 161, no. 3 (March 1, 1985): 457–74. http://dx.doi.org/10.1084/jem.161.3.457.
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Human megakaryocytes were studied for phenotypic changes occurring throughout differentiation using a panel of monoclonal antibodies raised against marrow megakaryocytes and blood platelets. 11 monoclonal antibody preparations were selected for restricted specificity against megakaryocytes and/or platelets after screening by immunofluorescence, complement-mediated cytolysis, and solid phase enzyme-linked immunosorbent assay. The expression of the cellular epitopes recognized by these reagents enabled the identification of three levels of megakaryocyte maturation characterized by distinct immunologic phenotypes. Based upon their reactivities against megakaryocytic cells at different ontogenetic levels, monoclonal antibodies were operationally categorized into three groups. Group A consisted of six different monoclonal antibodies that recognized antigens on the colony-forming unit-megakaryocyte (CFU-Mk), in vitro grown colony megakaryocytes, and early immature marrow megakaryocytes, only, and did not detect their respective epitopes on either mature megakaryocytes or platelets. A monoclonal antibody categorized in group B detected a cell antigen expressed by megakaryocytic cells at all maturational levels, but which is lost or suppressed during terminal differentiation and is not expressed on blood platelets. Group C included four different monoclonal antibodies raised against platelets that recognized antigenic determinants expressed on the CFU-Mk, colony megakaryocytes, early and mature megakaryocytes, and platelets. Three group C monoclonal antibodies (PC-1, PC-3, and PC-4) were specific for platelet glycoprotein IIb/IIIa. Additionally, group C monoclonal antibody PC-2 was unique in that it showed partial reactivity against the clonable progenitor for the erythroid series (BFU-E). Recognition of discrete phenotypic changes in differentiating megakaryocytes will enable multiparameter analyses of these cells as well as the study of factors regulating the dynamics of megakaryocytopoiesis in health and disease.
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Pokharel, S., P. Upadhyaya, S. Karki, P. Paudyal, B. Pradhan, and P. Poudel. "Megakaryocytic alterations in thrombocytopenia: A bone marrow aspiration study." Journal of Pathology of Nepal 6, no. 11 (March 17, 2016): 914–21. http://dx.doi.org/10.3126/jpn.v6i11.15673.
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Background: Megakaryocyte morphology plays an important role in thrombopoiesis. A defect in any stage of megakaryocytopoiesis can lead to dysmegakaryocytopoiesis and thrombocytopenia. This study was conducted to understand megakaryocytic alterations and their contribution in the diagnosis of cases of thrombocytopenia.Materials and Methods: This was a cross-sectional study was conducted on all consecutive cases of bone marrow aspirates of thrombocytopenia over a duration of one year in BPKIHS. Megakaryocyte morphology was studied with a 100X objective. Data were entered into Microsoft excel 10 and analysed with SPSS version 11.5. Descriptive statistics charted and Chi-square tests were done for inferential statistics to find any association at 95% Confidence Interval.Results: Among the 38 subjects, megakaryocytic thrombocytopenia (44.7%) was the most common cause of thrombocytopenia. Hypolobated megakaryocytes (63.2%), bare megakaryocytic nuclei (57.9%) were the common morphological changes in megakaryocytes. Odds of increased megakaryocyte count in megakaryocytic thrombocytopenia was found to be 12.5 times than for other causes of thrombocytopenia and the presence of bare megakaryocytic nuclei in MTP was statistically significant. (p –value<0.05)Conclusion: Many similarities were observed in megakaryocytic morphology among different hematological diseases. However, increased megakaryocyte count and presence of bare megakaryocytic nuclei, hypolobated forms were significant in megakaryocytic thrombocytopenia.
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Rivière, Christel, Frédéric Subra, Karine Cohen-Solal, Véronique Cordette-Lagarde, Remi Letestu, Christian Auclair, William Vainchenker, and Fawzia Louache. "Phenotypic and Functional Evidence for the Expression of CXCR4 Receptor During Megakaryocytopoiesis." Blood 93, no. 5 (March 1, 1999): 1511–23. http://dx.doi.org/10.1182/blood.v93.5.1511.
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Abstract The identification of stromal cell–derived factor (SDF)-1 as a chemoattractant for human progenitor cells suggests that this chemokine and its receptor might represent critical determinants for the homing, retention, and exit of precursor cells from hematopoietic organs. In this study, we investigated the expression profile of CXCR4 receptor and the biological activity of SDF-1 during megakaryocytopoiesis. CD34+ cells from bone marrow and cord blood were purified and induced to differentiate toward the megakaryocyte lineage by a combination of stem-cell factor (SCF) and recombinant human pegylated megakaryocyte growth and development factor (PEG-rhuMGDF). After 6 days of culture, a time where mature and immature megakaryocytes were present, CD41+ cells were immunopurified and CXCR4mRNA expression was studied. High transcript levels were detected by a RNase protection assay in cultured megakaryocytes derived from cord blood CD34+ cells as well as in peripheral blood platelets. The transcript levels were about equivalent to that found in activated T cells. By flow cytometry, a large fraction (ranging from 30% to 100%) of CD41+cells showed high levels of CXCR4 antigen on their surface, its expression increasing in parallel with the CD41 antigen during megakaryocytic differentiation. CXCR4 protein was also detected on peripheral blood platelets. SDF-1 acts on megakaryocytes by inducing intracellular calcium mobilization and actin polymerization. In addition, in in vitro transmigration experiments, a significant proportion of megakaryocytes was observed to respond to this chemokine. This cell migration was inhibited by pertussis toxin, indicating coupling of this signal to heterotrimeric guanine nucleotide binding proteins. Although a close correlation between CD41a and CXCR4 expession was observed, cell surface markers as well as morphological criteria indicate a preferential attraction of immature megakaryocytes (low level of CD41a and CD42a), suggesting that SDF-1 is a potent attractant for immature megakaryocytic cells but is less active on fully mature megakaryocytes. This hypothesis was further supported by the observation that SDF-1 induced the migration of colony forming unit–megakaryocyte progenitors (CFU-MK) and the expression of activation-dependent P-selectin (CD62P) surface antigen on early megakaryocytes, although no effect was observed on mature megakaryocytes and platelets. These results indicate that CXCR4 is expressed by human megakaryocytes and platelets. Furthermore, based on the lower responses of mature megakaryocytes and platelets to SDF-1 as compared with early precursors, these data suggest a role for this chemokine in the maintenance and homing during early stages of megakaryocyte development. Moreover, because megakaryocytes are also reported to express CD4, it becomes important to reevaluate the role of direct infection of these cells by the human immunodeficiency virus (HIV)-1 in HIV-1–related thrombocytopenia.
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Rivière, Christel, Frédéric Subra, Karine Cohen-Solal, Véronique Cordette-Lagarde, Remi Letestu, Christian Auclair, William Vainchenker, and Fawzia Louache. "Phenotypic and Functional Evidence for the Expression of CXCR4 Receptor During Megakaryocytopoiesis." Blood 93, no. 5 (March 1, 1999): 1511–23. http://dx.doi.org/10.1182/blood.v93.5.1511.405k02_1511_1523.
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The identification of stromal cell–derived factor (SDF)-1 as a chemoattractant for human progenitor cells suggests that this chemokine and its receptor might represent critical determinants for the homing, retention, and exit of precursor cells from hematopoietic organs. In this study, we investigated the expression profile of CXCR4 receptor and the biological activity of SDF-1 during megakaryocytopoiesis. CD34+ cells from bone marrow and cord blood were purified and induced to differentiate toward the megakaryocyte lineage by a combination of stem-cell factor (SCF) and recombinant human pegylated megakaryocyte growth and development factor (PEG-rhuMGDF). After 6 days of culture, a time where mature and immature megakaryocytes were present, CD41+ cells were immunopurified and CXCR4mRNA expression was studied. High transcript levels were detected by a RNase protection assay in cultured megakaryocytes derived from cord blood CD34+ cells as well as in peripheral blood platelets. The transcript levels were about equivalent to that found in activated T cells. By flow cytometry, a large fraction (ranging from 30% to 100%) of CD41+cells showed high levels of CXCR4 antigen on their surface, its expression increasing in parallel with the CD41 antigen during megakaryocytic differentiation. CXCR4 protein was also detected on peripheral blood platelets. SDF-1 acts on megakaryocytes by inducing intracellular calcium mobilization and actin polymerization. In addition, in in vitro transmigration experiments, a significant proportion of megakaryocytes was observed to respond to this chemokine. This cell migration was inhibited by pertussis toxin, indicating coupling of this signal to heterotrimeric guanine nucleotide binding proteins. Although a close correlation between CD41a and CXCR4 expession was observed, cell surface markers as well as morphological criteria indicate a preferential attraction of immature megakaryocytes (low level of CD41a and CD42a), suggesting that SDF-1 is a potent attractant for immature megakaryocytic cells but is less active on fully mature megakaryocytes. This hypothesis was further supported by the observation that SDF-1 induced the migration of colony forming unit–megakaryocyte progenitors (CFU-MK) and the expression of activation-dependent P-selectin (CD62P) surface antigen on early megakaryocytes, although no effect was observed on mature megakaryocytes and platelets. These results indicate that CXCR4 is expressed by human megakaryocytes and platelets. Furthermore, based on the lower responses of mature megakaryocytes and platelets to SDF-1 as compared with early precursors, these data suggest a role for this chemokine in the maintenance and homing during early stages of megakaryocyte development. Moreover, because megakaryocytes are also reported to express CD4, it becomes important to reevaluate the role of direct infection of these cells by the human immunodeficiency virus (HIV)-1 in HIV-1–related thrombocytopenia.
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Kaluzhny, Yulia, Guangyao Yu, Shishinn Sun, Paul A. Toselli, Bernhard Nieswandt, Carl W. Jackson, and Katya Ravid. "BclxL overexpression in megakaryocytes leads to impaired platelet fragmentation." Blood 100, no. 5 (September 1, 2002): 1670–78. http://dx.doi.org/10.1182/blood-2001-12-0263.
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Fragmentation of polyploid megakaryocytes into platelets has great relevance for blood homeostasis. Apoptotic cell death is a highly regulated genetic program, which has been observed in mature megakaryocytes fragmenting into platelets. The antiapoptotic protein BclxL has been reported as up-regulated during megakaryocytic differentiation in vitro, but absent during late megakaryopoiesis. Our study focused on examining BclxL levels in megakaryocytes in vivo and in assessing the effect of its overexpression in transgenic mice (via the platelet factor 4 [PF4] promoter) on megakaryocyte development and platelet fragmentation. Interestingly, in the wild-type and less in PF4-driven transgenic mice, BclxL was not detected in a fraction of the large mature megakaryocytes, suggesting a regulation on the protein level. BclxL overexpression was associated with a moderate increase in megakaryocyte number, with no significant change in ploidy level or platelet counts. When the mice were challenged by induction of immune thrombocytopenia, the rate of platelet recovery was significantly slower in the transgenic mice as compared with controls. Moreover, proplatelet formation in vitro by transgenic megakaryocytes was limited. Transgenic megakaryocytes displayed poorly developed platelet demarcation membranes and cell margin extensions. Our study indicates that regulated expression of BclxL in megakaryocytes is important for the development of cells with a high potential to fragment into platelets.
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Wen, Qiang Jeremy, Benjamin Goldenson, Sebastien Malinge, Priya Koppikar, Ross L. Levine, Ayalew Tefferi, and John Crispino. "Induction of Megakaryocyte Polyploidization in Combination with JAK Inhibition As a Novel Therapeutic Strategy for Myeloproliferative Neoplasms." Blood 118, no. 21 (November 18, 2011): 64. http://dx.doi.org/10.1182/blood.v118.21.64.64.
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Abstract Abstract 64 Megakaryocytes are one of the few cell types that undergo a modified form of the cell cycle termed endomitosis, in which cells skip the late stages of mitosis to become polyploid. Murine and human megakaryocytes commonly reach modal ploidy states of 32N and 16N, respectively, and can sometimes achieve DNA contents as high as 64N. Polyploidization is associated with upregulation of megakaryocyte lineage specific genes, proplatelet formation and expression of genes related to apoptosis. RNA expression array studies have shown that high ploidy states are strongly correlated with megakaryocyte differentiation and maturation. Importantly, the choice of a megakaryocyte to undergo polyploidization and differentiation is inextricably linked to exit from the proliferative cell cycle. Given that megakaryocytes in patients with essential thrombocythemia are hyperproliferative and that those in primary myelofibrosis fail to undergo normal differentiation or polyploidization, we hypothesized that small molecule inducers of polyploidization would drive these cells to exit the proliferative cell cycle and undergo terminal differentiation or death. In collaboration with the Broad Institute, we performed a high throughput screen and identified small molecules that induce polyploidization and proliferative arrest of malignant megakaryocytes, including those that express MPLW515L and JAK2 V617F. We have shown that these compounds, including the Rho kinase inhibitor dimethylfasudil (diMF), selectively increase polyploidization, expression of megakaryocyte cell surface markers, and apoptosis of murine and human megakaryocytic cell lines and primary cells. Furthermore, diMF blocked the growth of primary human AMKL blasts both in vitro and in vivo. With respect to MPNs, diMF showed potent activity against megakaryocytic cell lines and primary cells expressing either JAK2 V617F or MPL W515L alleles commonly associated with these disorders. diMF inhibited proliferation, induced polyploidization and upregulation of lineage specific markers CD41 and CD42, and increased apoptosis of megakaryocytes transduced with JAK2 V617F or MPLW515L. diMF also significantly reduced megakaryocyte colony forming units (CFU-MK), and induced polyploidization and differentiation of bone marrow and fetal liver megakaryocytes from Gata1 mutant mice, which develop a PMF-like disease. Given that diMF induces growth arrest, polyploidization and apoptosis of cells that express activated mutants of JAK2 and MPL, we predicted that it, as well as other small molecule inducers of polyploidy, would be efficient at restraining aberrant megakaryocyte proliferation in both PMF and ET. To assay the effectiveness of diMF in these disorders, we treated peripheral blood mononuclear cells from patients with either PMF or ET with diMF and monitored growth and maturation of megakaryocytes. We discovered that diMF induced polyploidization and subsequent apoptosis of both types of MPN primary samples. diMF also reduced CFU-MK of these MPN patient samples. Next, we assessed the activity of diMF in a model of MPN in which congenic recipients of MPLW515L transduced Balb/C bone marrow cells develop a rapid MPN characterized by leukocytosis, thrombocytosis, bone marrow fibrosis, and death. diMF led to a significant decrease of fibrosis in the bone marrow, diminished infiltration of megakaryocytes and granulocytes in the liver, and a profound reduction in the numbers of megakaryocytes within the spleen of a mouse model of PMF. diMF also led to a significant reduction in the platelet count and a trend towards decreased white cell count, with no effect on hematocrit. Overall, diMF results were comparable to intermediate doses of INCB16562. These encouraging results strongly suggest that diMF induces a decline in megakaryocyte lineage, which leads to reduction in platelet count, and support pre-clinical development of diMF for megakaryocytic subtypes of MPNs. Of note, diMF did not inhibit the phosphorylation of Stat5 or Stat3, suggesting that it acts through a mechanism distinct from JAK2 inhibitors. Interestingly, combination of diMF with a selective JAK2 inhibitor greatly enhanced the efficacy of diMF to inhibit proliferation and induce apoptosis in MPLW515L transduced megakaryocytic cell line. These data support combining JAK inhibition and induction of megakaryocyte polyploidy as a new therapeutic strategy for MPNs. Disclosures: No relevant conflicts of interest to declare.
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Noh, Ji-Yoon, Shilpa Gandre-Babbe, Yuhuan Wang, Vincent Hayes, Yu Yao, Paul Gadue, Spencer Sullivan, et al. "Inducible Gata1 Suppression As a Novel Strategy to Expand Physiologic Megakaryocyte Production from Embryonic Stem Cells." Blood 124, no. 21 (December 6, 2014): 3846. http://dx.doi.org/10.1182/blood.v124.21.3846.3846.
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Abstract Embryonic stem (ES) and induced pluripotent stem (iPS) cells represent potential sources of megakaryocytes and platelets for transfusion therapy. However, most current ES/iPS cell differentiation protocols are limited by low yields of hematopoietic progeny, including platelet-releasing megakaryocytes. Mutations in the mouse and human genes encoding transcription factor GATA1 cause accumulation of proliferating, developmentally arrested megakaryocytes. Previously, we reported that in vitro differentiation of Gata1-null murine ES cells generated self-renewing hematopoietic progenitors termed G1ME cells that differentiated into erythroblasts and megakaryocytes upon restoration of Gata1 cDNA by retroviral transfer. However, terminal maturation of Gata1-rescued megakaryocytes was aberrant with immature morphology and no proplatelet formation, presumably due to non-physiological expression of GATA1. We now engineered wild type (WT) murine ES cells that express doxycycline (dox)-regulated Gata1 short hairpin (sh) RNAs to develop a strategy for Gata1-blockade that upon its release, restores physiologic GATA1 expression during megakaryopoiesis. In vitro hematopoietic differentiation of control scramble shRNA-expressing ES cells with dox and thrombopoietin (TPO) produced megakaryocytes that underwent senescence after 7 days. Under similar differentiation conditions, Gata1 shRNA-expressing ES cells produced immature hematopoietic progenitors, termed G1ME2 cells, which replicated continuously for more than 40 days, resulting in ~1013-fold expansion (N=4 separate experiments). Upon dox withdrawal with multi-lineage cytokines present (EPO, TPO, SCF, GMCSF and IL3), endogenous GATA1 expression was restored to G1ME2 cells followed by differentiation into erythroblasts and megakaryocytes, but no myeloid cells. In clonal methylcellulose assays, dox-deprived G1ME2 cells produced a mixture of erythroid, megakaryocytic and erythro-megakaryocytic colonies. In liquid culture with TPO alone, dox-deprived G1ME2 cells formed mature megakaryocytes in 5-6 days, as determined by morphology, ultrastructure, acetylcholinesterase staining, upregulated megakaryocytic gene expression (Vwf, Pf4, Gp1ba, Selp, Ppbp), CD42b surface expression, increased DNA ploidy and proplatelet production. Compared to G1ME cells rescued with Gata1 cDNA retrovirus, dox-deprived G1ME2 cells exhibited more robust megakaryocytic maturation, similar to that of megakaryocytes produced from cultured fetal liver. Importantly, G1ME2 cell-derived megakaryocytes generated proplatelets in vitro and functional platelets in vivo (~40 platelets/megakaryocyte with a circulating half life of 5-6 hours). These platelets were actively incorporated into growing arteriolar thrombi at sites of laser injury and subsequently expressed the platelet activation marker p-selectin (N=3-4 separate experiments). Our findings indicate that precise timing and magnitude of a transcription factor is required for proper terminal hematopoiesis. We illustrate this principle using a novel, readily reproducible strategy to expand ES cell-derived megakaryocyte-erythroid progenitors and direct their differentiation into megakaryocytes and then into functional platelets in clinically relevant numbers. Disclosures No relevant conflicts of interest to declare.
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Ghai, Shuchi, and Sharada Rai. "Megakaryocytic morphology in Janus kinase 2 V617F positive myeloproliferative neoplasm." South Asian Journal of Cancer 06, no. 02 (April 2017): 075–78. http://dx.doi.org/10.4103/2278-330x.208854.
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Abstract Context: Alterations in megakaryocyte morphology are the hallmark of myeloproliferative neoplasms (MPNs). These neoplasm are also associated with Janus kinase 2 (JAK2) V617F mutation in nearly 95% patients with polycythemia vera (PV), 40% patients of essential thrombocythemia (ET) and 50% patients of myelofibrosis (MF). The utility of megakaryocyte morphology in these disorders in correlation with JAK2 V617F remains unresolved. Aims: The aim of the study was to assess the morphology of megakaryocytes in bone marrow aspirates (BMAs) and bone marrow biopsies of patients of BCR-ABL negative MPNs with JAK2 V617F mutation. Settings and Design: This study was a retrospective and prospective, hospital-based study undertaken for a period ranging from January 2011 to April 2015. Subjects and Methods: Assessment of morphological features of megakaryocytes in 15 BMAs and their respective biopsies which included seven cases of PV, three cases of ET, and five cases of MF with JAK2 V617F mutation. Statistical Analysis Used: Chi-square test and Fisher exact test were used to compare the different features of megakaryocytes. Software version SPSS 13.0 was used. Results: Megakaryocytes in ET were found to have characteristically large size with staghorn multinucleated nuclei and exhibiting large amount of cytoplasm. MF showed dense clustering of megakaryocytes with staghorn nucleus along with sinusoidal dilatation and intrasinusoidal hematopoiesis. PV showed loose and dense clustering of megakaryocytes with a predominance of cloud-like nuclei. Few of the megakaryocytic morphologic features showed overlap between MF and PV and between ET and early MF. Conclusions: Megakaryocytic morphology can aid in the accurate diagnosis of the different subcategories of MPNs. This would help in categorization of clinically suspicious patients of JAK2 V617F negative patients.
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Ghai, Shuchi, and Sharada Rai. "Megakaryocytic morphology in Janus kinase 2 V617F positive myeloproliferative neoplasm." South Asian Journal of Cancer 06, no. 02 (April 2017): 075–78. http://dx.doi.org/10.4103/2278-330x.208854.
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Abstract Context: Alterations in megakaryocyte morphology are the hallmark of myeloproliferative neoplasms (MPNs). These neoplasm are also associated with Janus kinase 2 (JAK2) V617F mutation in nearly 95% patients with polycythemia vera (PV), 40% patients of essential thrombocythemia (ET) and 50% patients of myelofibrosis (MF). The utility of megakaryocyte morphology in these disorders in correlation with JAK2 V617F remains unresolved. Aims: The aim of the study was to assess the morphology of megakaryocytes in bone marrow aspirates (BMAs) and bone marrow biopsies of patients of BCR-ABL negative MPNs with JAK2 V617F mutation. Settings and Design: This study was a retrospective and prospective, hospital-based study undertaken for a period ranging from January 2011 to April 2015. Subjects and Methods: Assessment of morphological features of megakaryocytes in 15 BMAs and their respective biopsies which included seven cases of PV, three cases of ET, and five cases of MF with JAK2 V617F mutation. Statistical Analysis Used: Chi-square test and Fisher exact test were used to compare the different features of megakaryocytes. Software version SPSS 13.0 was used. Results: Megakaryocytes in ET were found to have characteristically large size with staghorn multinucleated nuclei and exhibiting large amount of cytoplasm. MF showed dense clustering of megakaryocytes with staghorn nucleus along with sinusoidal dilatation and intrasinusoidal hematopoiesis. PV showed loose and dense clustering of megakaryocytes with a predominance of cloud-like nuclei. Few of the megakaryocytic morphologic features showed overlap between MF and PV and between ET and early MF. Conclusions: Megakaryocytic morphology can aid in the accurate diagnosis of the different subcategories of MPNs. This would help in categorization of clinically suspicious patients of JAK2 V617F negative patients.
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James, Chloé, Emma C. Josefsson, Michael J. White, Katya J. Henley, and Benjamin T. Kile. "Deletion of Bcl-X in the Megakaryocyte Compartment Results in Profound Thrombocytopenia Caused by Impaired Platelet Production and Survival." Blood 114, no. 22 (November 20, 2009): 1495. http://dx.doi.org/10.1182/blood.v114.22.1495.1495.
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Abstract Abstract 1495 Poster Board I-518 Recent studies have suggested a role for the intrinsic apoptosis pathway in megakaryocytic differentiation and platelet shedding. The intrinsic pathway is regulated by the Bcl-2 family of pro- and anti- cell death proteins. We recently demonstrated that platelet life span is controlled by an intrinsic cell death pathway, whereby the anti-apoptotic protein Bcl-xL constrains the pro-apoptotic activity of Bak to maintain platelet survival. As Bcl-xL is expressed in megakaryocytes, we investigated whether this protein is required for megakaryocyte survival, differentiation and/or platelet shedding. We specifically deleted the Bcl-x gene in the megakaryocyte lineage by crossing mice carrying a floxed allele of Bcl-x with mice carrying a platelet factor 4-regulated Cre transgene. Bcl-xfl/flCre+ mice were profoundly thrombocytopenic (26 ± 5 × 103/μl, n=14) compared with Bcl-xfl/flCre− animals (1157 ± 202 × 103/μl, n=13). Platelet life span in these mice was reduced to only 5 hours, as compared to 5 days in wild type littermates. This result confirmed that Bcl-xL is absolutely required for platelet survival. To determine whether Bcl-x deletion has an impact on platelet production, we analyzed the megakaryocyte compartment in Bcl-xfl/flCre+ and Bcl-xfl/flCre− mice. We observed that the number of megakaryocyte progenitors, and number of megakaryocytes in the bone marrow were increased in Bcl-xfl/flCre+ mice (23 ± 9 megakaryocyte progenitors vs 11 ± 5, and 51 ± 9 megakaryocytes vs 12 ± 1). This result suggested that Bcl-xL is not required for the survival of megakaryocytes or their progenitors. To determine whether Bcl-xL is required for the last step of megakaryocyte differentiation, i.e. platelet shedding, we cultured fetal liver cells with thrombopoietin. Large megakaryocytes were isolated after 3 days of differentiation on discontinuous bovine serum albumin gradient. They were cultured for 3 more days in the same media and the percentage of megakaryocytes displaying proplatelets was determined each day. Interestingly, Bcl-xfl/flCre+ megakaryocytes died much more quickly than Bcl-xfl/flCre− megakaryocytes, and almost none of those that survived were able to form proplatelets. Our study indicates that Bcl-xL is not only essential for platelet survival, but it is also required for the survival of mature megakaryocytes at the stage of platelet shedding. Disclosures: No relevant conflicts of interest to declare.
Dissertations / Theses on the topic "Megakaryocytes"
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Melford, Steven K. "Calcium signalling in megakaryocytes." Thesis, University of Oxford, 1997. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.364104.
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Hamlet, Isla. "GATA1 protein partners in megakaryocytes." Thesis, University of Oxford, 2007. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.442463.
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McGrath, Catherine Jane. "Glutamate release mechanisms from megakaryocytes." Thesis, University of York, 2007. http://etheses.whiterose.ac.uk/9950/.
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Cardiovascular disease (CVD) is one of the main causes of death in western society. Platelet activation, thrombus formation and plaque rupture are all central events in the pathogenesis of acute coronary syndromes, therefore therapies targeted at controlling platelet numbers and aggregation are likely to be beneficial in the treatment of CVD. Megakaryocytes (MKs) which are the precursors to platelets are an ideal target for these therapies, however the intrinsic factors that regulate the production and shedding of platelet precursors are poorly understood. Recent studies identified that MKs express functional NMDA-type glutamate receptors similar to those found in the CNS and that antagonism of these receptors prevents normal MK differentiation and platelet function. This thesis investigates glutamate signalling within MKs further, focusing on the glutamate release capability of MK cells and the mechanisms involved. Using molecular and cellular techniques it was demonstrated that MK cells expressed numerous regulatory proteins required for vesicular glutamate release, including core SNARE proteins, VAMP, SNAP-23 and syntaxin; specific glutamate-loading vesicle proteins, VGLUTI and VGLUT2; and glutamate transporters, EAATI and EAAT2. Active vesicle recycling was observed in MK cells using a fluorescent reporter and an enzyme-linked fluorimetric assay confirmed that MK cells constitutively released glutamate and that glutamate release levels increased significantly following MK differentiation. Transient transfection of the human cell line MEG-Ol with tetanus toxin, which disables vesicle recycling, induced a 30% decrease (P
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Cowart, Miles. "Exploring the role of NIPSNAP4, CIB1, and PLK3 in megakaryocyte maturation." Access to citation, abstract and download form provided by ProQuest Information and Learning Company; downloadable PDF file, 80 p, 2007. http://proquest.umi.com/pqdweb?did=1253509361&sid=2&Fmt=2&clientId=8331&RQT=309&VName=PQD.
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Kuhl, Christiane. "Structure / function analysis of GATA1 in megakaryocytes." Thesis, University of Oxford, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.418523.
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Sobolewski, S. "A new function of megakaryocytes in malignancies." Thesis, University of Bradford, 1986. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.375083.
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Hitchcock, Ian Stuart. "Functional activity of NMDA receptors on megakaryocytes." Thesis, University of York, 2003. http://etheses.whiterose.ac.uk/9856/.
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Dalby, Amanda Louise. "Forward programming of human pluripotent stem cells to a megakaryocyte-erythrocyte bi-potent progenitor population : an in vitro system for the production of platelets and red blood cells for transfusion medicine." Thesis, University of Cambridge, 2018. https://www.repository.cam.ac.uk/handle/1810/273749.
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There exists a need to produce platelets in vitro for use in transfusion medicine, due to increased platelet demands and short shelf life. Our lab uses human induced pluripotent stem cells (iPSCs), as an attractive alternative supply, as iPSCs can be cultured indefinitely and differentiate into almost any cell type. Using a technique called forward programming, we over express three key haematological transcription factors (TFs), pushing iPSCs towards the megakaryocyte lineage, to produce mature megakaryocytes, the platelet precursor cell type. A major limitation of the forward programming technique is a reliance of lentiviral transduction to overexpress the three TFs, which leads to a number of issues including heterogeneity and high experimental costs. To overcome this, I have developed an inducible iPSC line by inserting the forward programming TFs into a genomic safe harbour, using genome editing techniques. TF expression is strictly controlled, with the TFs expressed only after chemical induction. Inducing forward programming is an efficient method for producing mature megakaryocytes and these cells maintain higher purity in long-term cultures, when compared to cells produced by the lentiviral method. Removing the requirement of lentiviral transduction is a major advancement, making forward programming more amenable to scaling-up, thus moving this technology closer towards our goal of producing in vitro platelets for use in transfusion medicine. I have also shown that forward programming generates a bi-potent progenitor population, from which erythroblasts can be generated, by altering only media conditions. As for megakaryocyte cultures, inducing forward programming improves the purity of erythroblasts produced, compared to the lentiviral method. I have developed single cell progenitor assays combined with index sorting of different cell surface markers, to allow retrospective analysis of cells which successfully generate colonies. The aim of this work is to better characterise the progenitor cells produced by forward programming, to allow further study of this cell type. Single cell RNA-seq of megakaryocytes revealed heterogeneity in long-term cultures and also identified novel candidate surface markers that may help to further characterise the progenitor cell population.
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Wagner, Leonard [Verfasser], and Andreas [Gutachter] Geier. "Zinc homeostasis in megakaryocytes / Leonard Wagner ; Gutachter: Andreas Geier." Würzburg : Universität Würzburg, 2020. http://d-nb.info/1220634220/34.
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Abdalla, Sarah. "Identification of the Regions in Factor V Mediating its Edocytosis by Megakaryocytes to Form the Unique Platelet-Derived Cofactor Molecule." ScholarWorks @ UVM, 2013. http://scholarworks.uvm.edu/graddis/6.
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Factor Va is a plasma protein that plays an important role in the regulation of blood coagulation by serving as the essential cofactor in thrombin generation via the prothrombinase complex. The procofactor, factor V, exists in two whole blood pools with 75-80% found in plasma, and 20-25% stored in the α-granules of platelets. As compared to the plasma procofactor, platelet-derived factor V is physically and functionally distinct, and displays a more procoagulant phenotype. Despite these profound differences, platelet-derived factor V originates via endocytosis of the plasma-derived procofactor by megakaryocytes. Endocytosis is mediated by two receptors: an unidentified, specific factor V receptor, and low density lipoprotein (LDL) receptor related protein-1 (LRP-1), a ubiquitous receptor that plays a role in endocytosis of proteins targeted for lysosomal degradation. These observations represent a novel role for LRP-1 in endocytosis of a protein that is functionally modified, and not targeted for lysosomal degradation. The goal of this study is to define the factor V regions involved in its interactions with the unidentified factor V receptor and LRP-1 expressed on megakaryocytes to begin to elucidate the molecular mechanisms regulating formation of the unique platelet-derived cofactor. Epitope mapping studies were performed using anti-factor V monoclonal antibodies, E9 and anti-factor V #2. Previous observations indicated that these factor Va light chain antibodies inhibited endocytosis of factor V by megakaryocytes. However, subsequent analyses demonstrated that only E9 inhibited both factor V binding and endocytosis. Thus, it was used for these studies. Western blotting of factor V and Va suggested that E9 recognizes a conformation-dependent epitope, which precluded the use of conventional epitope mapping approaches used for linear epitopes. E9 had no effect on factor Va cofactor activity in a plasma-based clotting assay suggesting that it does not perturb factor Va’s interactions with the membrane surface or factor Xa. Cleavage of lipid-bound factor Va by factor Xa at Arg1765 was also not affected by the presence of E9 suggesting that the epitope is not directed against this cleavage site. When E9 was used to immunoprecipitate the factor Xa-generated light chain cleavage products, both the 48/46 and 30 kDa light chain fragments were captured. These observations were confirmed using a solid phase competition assay where factor Xa-cleaved factor Va inhibited binding of 125I-factor V to E9 as well as intact factor V or Va. Limited proteolysis of the factor Va light chain with trypsin or Asp-N, generated products that were no longer detectable in this assay. These combined observations suggest that the anti-factor V light chain antibody, E9, has an epitope that is conformation-dependent and extremely labile. Future directions and alternative approaches are discussed.
Books on the topic "Megakaryocytes"
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Gibbins, Jonathan M., and Martyn P. Mahaut-Smith. Platelets and Megakaryocytes. New Jersey: Humana Press, 2004. http://dx.doi.org/10.1385/1592597823.
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Gibbins, Jonathan M., and Martyn P. Mahaut-Smith. Platelets and Megakaryocytes. New Jersey: Humana Press, 2004. http://dx.doi.org/10.1385/1592597831.
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Gibbins, Jonathan M., and Martyn P. Mahaut-Smith, eds. Platelets and Megakaryocytes. New York, NY: Springer New York, 2012. http://dx.doi.org/10.1007/978-1-61779-307-3.
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Gibbins, Jonathan M., and Martyn Mahaut-Smith, eds. Platelets and Megakaryocytes. New York, NY: Springer New York, 2018. http://dx.doi.org/10.1007/978-1-4939-8585-2.
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M, Gibbins Jonathan, and Mahaut-Smith Martyn P, eds. Platelets and megakaryocytes. Totowa, N.J: Humana Press, 2004.
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Harris, J. R., ed. Megakaryocytes, Platelets, Macrophages, and Eosinophils. Boston, MA: Springer US, 1991. http://dx.doi.org/10.1007/978-1-4757-9531-8.
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R, Harris James, ed. Megakaryocytes, platelets, macrophages, and eosinophils. New York: Plenum Press, 1991.
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W, Colman Robert, and Smith J. Bryan, eds. Methods for studying platelets and megakaryocytes. New York: Liss, 1987.
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F, Levine Richard, ed. Megakaryocyte development and function: Proceedings of an international conference held at the Marine Biological Laboratory, Woods Hole, Massachusetts, September 18-21, 1985. New York: Liss, 1986.
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A, Selivanov V., and Lanin V. N, eds. Reguli͡at͡sii͡a megakariot͡sitopoėza. Pushchino: Pushchinskiĭ nauch. t͡sentr AN SSSR, 1991.
Find full textBook chapters on the topic "Megakaryocytes"
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Morgan, Michael M., MacDonald J. Christie, Thomas Steckler, Ben J. Harrison, Christos Pantelis, Christof Baltes, Thomas Mueggler, et al. "Megakaryocytes." In Encyclopedia of Psychopharmacology, 752. Berlin, Heidelberg: Springer Berlin Heidelberg, 2010. http://dx.doi.org/10.1007/978-3-540-68706-1_805.
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Garzon, Ramiro. "MicroRNA Profiling of Megakaryocytes." In DNA and RNA Profiling in Human Blood, 293–98. Totowa, NJ: Humana Press, 2009. http://dx.doi.org/10.1007/978-1-59745-553-4_19.
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Kaushansky, Alexis, and Kenneth Kaushansky. "Systems Biology of Megakaryocytes." In A Systems Biology Approach to Blood, 59–84. New York, NY: Springer New York, 2014. http://dx.doi.org/10.1007/978-1-4939-2095-2_4.
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Martin, J. F., and P. M. W. Bath. "Platelets and Megakaryocytes in Vascular Disease." In Developments in Cardiovascular Medicine, 49–62. Dordrecht: Springer Netherlands, 1991. http://dx.doi.org/10.1007/978-94-011-3484-2_3.
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Baatout, Sarah. "Isolation of Megakaryocytes Using Magnetic Cell Separation." In Scientific and Clinical Applications of Magnetic Carriers, 261–68. Boston, MA: Springer US, 1997. http://dx.doi.org/10.1007/978-1-4757-6482-6_18.
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Schönrich, Günther, and Martin J. Raftery. "Megakaryocytes and Platelet Production During Viral Infection." In Molecular and Cellular Biology of Platelet Formation, 351–62. Cham: Springer International Publishing, 2016. http://dx.doi.org/10.1007/978-3-319-39562-3_16.
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Bergmeier, Wolfgang, and Ellen C. O’Shaughnessy. "Rap GTPase Signaling in Platelets and Megakaryocytes." In Molecular and Cellular Biology of Platelet Formation, 175–91. Cham: Springer International Publishing, 2016. http://dx.doi.org/10.1007/978-3-319-39562-3_8.
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Dušek, J., K. Indrák, V. Ščudla, and M. Jarošová. "Morphology of Megakaryocytes in Chronic Myeloproliferative Diseases." In Leukemias, 285–88. Berlin, Heidelberg: Springer Berlin Heidelberg, 1993. http://dx.doi.org/10.1007/978-3-642-77083-8_50.
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Duperray, Alain, Rolande Berthier, and Gérard Marguerie. "Biosynthesis and Processing of Platelet Glycoproteins in Megakaryocytes." In Blood Cell Biochemistry, 37–58. Boston, MA: Springer US, 1991. http://dx.doi.org/10.1007/978-1-4757-9531-8_2.
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Pogozhykh, Denys, Rainer Blasczyk, and Constança Figueiredo. "Isolation, Cryopreservation, and Characterization of iPSC-Derived Megakaryocytes." In Cryopreservation and Freeze-Drying Protocols, 539–54. New York, NY: Springer US, 2020. http://dx.doi.org/10.1007/978-1-0716-0783-1_27.
Full textConference papers on the topic "Megakaryocytes"
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Berthier, R., A. Duperray, O. Valiron, M. Prenant, I. Newton, and A. Schweitzer. "MEGAKARYOCYTIC DEVELOPMENT IN LIQUID CULTURES OF CRYOPRESERVED LEUKOCYTE STEM CELL CONCENTRATES FROM CHRONIC MYELOGENOUS LEUKEMIA PATIENTS." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1644622.
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The proliferation and differentiation of human megakaryocytes in liquid culture has been obtained using cryopreserved light density blood cell concentrates from chronic myelogenous leukemia (CML) patients. These cryopreserved leukocytes concentrates contain a large number of viable granulo-monocytic, erythroid and megakaryocytic committed stem cells. A high number of spontaneous megakaryocytic colonies was observed in semisolid cultures plated with the CML leukocytes concentrates. A liquid culture system using RPMI 1640 supplemented with 20% human plasma (HP) has been defined where maturing megakaryocytes make up 20 to 60% of the total cells after 14 days of incubation. The same cell suspension cultured in medium supplemented with 20% foetal calf serum (FCS) showed poor megakaryocytic cell development. The megakaryocytic nature of the cells produced in HP supplemented cultures was confirmed by cytological studies and indirect immunofluorescence labeling using monoclonal antibodies (MoAb) against membrane platelet GPIb and Ilbllla, and intracellular antigens like fibrinogen and von Willebrand factor.Ploidy of the cultured cells was studied after labeling with propidium iodide and the DNA fluorescence determined using the fluorescence activated cell sorter (FACSIV). Peaks of 8N, 16N and 32N cells were observed from HP supplemented cultures representing about 20% of the cells reacting with a GP11b111 a MoAb, while very few cells greater than 4N were observed in FCS supplemented cultures. The megakaryocytes produced in HP cultures could be further enriched by cell sorting on the FACSIV after labeling with an anti-IIbIIIa MoAb. Depending on the initial megakaryocytic concentration of the cells cultured, one to 2 é 106 megakaryocytes per hour could be harvested. Thus, cryopreserved CML blood stem cell concentrates seem to offer a reproducible source of human megakaryocytes which retain their capacity to proliferate and differentiate in liquid cultures supplemented with human plasma. These megakaryocytes can be used for the study of platelet glycoprotein biosynthesis as well as the regulation of megakaryocytopoiesis.
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Cramer, Elisabeth M., F. John, William Vainchenker, and Janine Breton-Gorius. "PRODUCTION AND LOCALISATION OF ALPHA-GRANULE PROTEINS IN MATURING MEGAKARYOCYTES: AN OVERVIEW ON ULTRA-STRUCTURAL ASPECTS OF MEGAKARYOCYTE MATURATION." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1642952.
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In order to study the production of α- granule proteins in maturing megakaryocytes, we used immunocytochemical techniques performed on cultured and enriched bone marrow megakaryocytes. Cultures were prepared from bone marrow CFU-MK with the methylcellulose and plasma clot techniques. Preparation of bone marrow megakaryocytes was carried out from human or pig rib marrow separated on percoll gradient and counterflow centrifugation. Megakaryocyte preparations were 90$ pure and represented 85$ of those in the whole marrow. Activation was prevented with prostacyclin and prefixation with low concentration glutaraldehyde. A panel of monoclonal and polyclonal antibodies, against different platelet membrane glycoproteins and against cytoplasmic antigens (such as von Willebrand Factor (vWF), fibrinogen (Fg) and thrombospondin (TSP)) was used and observed by immunofluorescence or by immunogold in electron microscopy.The first megakaryocytic precursors, promegakaryoblasts (PMKB) identifiable by these antibodies were found at day 5 of culture. They had the size of lymphocytes, were labelled for GP lib, Ilia, and Ilb-IIIa complex but not for GPIb which appeared later. Platelet peroxidase was also present, otherwise these cells were devoid of α- granules and only a few of them exhibited a diffuse pattern for vWF immunolabelling. One day later membrane GPIb and diffuse cytoplasmic labelling for vWF were detected in the majority of PMKB. At day 9 of culture, this pattern of labelling for vWF became more intense and granular. The same pattern was observed for TSP and platelet factor 4. Immunoelectron microscopy showed that in immature megakaryocytes isolated from human bone marrow, labelling for vWF and TSP was observed in vesicles located in the Golgi region; in addition numerous small granules less than 0.1pm in diameter, round or elongated in shape, were labelled for these antigens. In mature human megakaryocytes, the labelling for these cytoplasmic antigens was restricted to the platelet α- granules in a distribution pattern similar to that of platelet α- granules. However, the labelling for Fg was consistently less intense in the granules of immature and mature megakaryocytes than in platelets.Because in platelets α- granule immunolabelling for vWF is associated with tubular structures which are specially prominent in porcine species, we studied vWF and tubular structures in pig megakaryocytes. Standard and immunoelectron microscopy revealed the simultaneous appearance of both in the small vesicles located in the Golgi area in the small immature α- granules and later in the mature α- granules. In mature megakaryocytes, labelling for vWF was intense and restricted to the α- granules. It was distributed eccentrically as in porcine blood platelets. Gold particles were often eccentrically located at one pole of the α- granule either labelling only its periphery or outlining one side of an elongated granule. Standard electron microscopy showed that tubular structures were very numerous in the mature α-granules, regularly spaced, arranged in parallel and usually located at one side of the granule. On the other hand platelets from pigs with homozygous von Willebrand disease were found to be completely devoid of both tubular structures and immunolabelling for vWF suggesting that the tubules represent the vWF itself.In acute megakaryoblastic leukemia, several phenotypes of PMKB were found in different patients, which corresponded to the stages of maturation of cultured megakaryocytes from CFU-MK.In conclusion, immunolabelling methods combined with megakaryocyte enrichment techniques are useful tools to study the origin of megakaryocyte (and platelet) granular proteins.
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Marquerie, G., A. Duperray, G. Uzan, and R. Berthier. "BIOSYNTHETIC PATHWAYS OF THE PLATELET FIBRINOGEN RECEPTOR IN HUMAN MEGAKARYOCYTES." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1642954.
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Interaction between cells and between cells and extracellular matrices are critical for a number of biological processes, including organ development, cell differenciation, cell motility, and the inimune' response. These interactions are mediated by a family of adhesion receptors that recognize short sequences such as Arg-Gly-Asp (RGD). These receptors share similar structural properties. They are heterodimers composed of a and B subunits and sometime express common epitopes. This suggests that the structural and functional relationship of these receptors may result from the transcription of related genes or may arise from cell specific post-transcriptional events. Thus, analysis of the biosynthesis and processing of these receptors would provide valuable insights into the molecular mechanism which control their expression at the surface,of different cells. Platelet membrane glycoprotein (GP) IIb-IIIa is a member of this receptor family. This protein is a non covalent heterodimer composed of two distinct polypeptides, Glib which consists of two subunits Ilba and IlbB (Mr = 116 kD, Mr = 25 kD) and GPIIIa (Mr = 100 kD, reduced). GPIIb-IIIa functions at site of platelet aggregation and serves as receptor for RGD containing factors including fibrinogen, fibronectin and von Willebrand factor. We report here on the investigation of the biosynthetic pathways of this RGD receptor in human megakaryocytes. High number of megakaryocytic cells from the megakaryoblastic stage to the polyploid mature megakaryocyte were obtained from liquid culture of cryopreserved leukocyte stem cell concentrates from patients with chronic myelogenous leukemia (CML). After sorting, using a FACS IV and indirect immunofluores-cent labeling with monoclonal antibodies anti-GPIIb-IIIa, 95 % of the cells in culture were of the megakaryocytic lineage. These megakarocytes represented an excellent tool to delineate at the molecular level events associated with the biosynthesis of GPIIb-IIIa.Metabolic labeling and pulse-chase experiments indicated that GPIIb and GPIIIa are synthesized from separate mRNA and that the two subunits of GPIIb derive from a common precursor. This was further confirmed by cell-free translation of megakaryocyte mRNA and the identification of separate cDNA containing sequences coding for the pro-GPIIb and for GPIIIa. These cDNA were isolated from a Xgt11 expression library constructed with purified megakaryocyte RNA, and were used to size the messengers coding for the two polypeptides. A single mRNA species of 3.9 kB was found to encode the pro-GPIIb, whereas two different mRNA species of 2.9 kB and 4. 1 kB were identified with the GPIIIa cDNA.The newly synthesized GPIIIa associates early with the pro-GPIIb in the rough endoplasmic reticulum. Examination of the glycosylation pathways with endoglycosidase H, tunicamycin and monensin indicated that high mannose oligosaccharides are added to the GPIIIa and pro-GPIIb polypeptide backbone. The pro-GPIIb is then processed with conversion of high mannose to the complex type carbohydrate, whereas GPIIIa remains endoH sensitive. Glycosylation of pro-GPIIb-IIIa and processing of oligosaccharides are prerequisite for proteolytic maturation of pro-GPIIb and the expression of the mature complex at the surface of the cell. Thus post-translational processing of GPIIb-IIIa requires an early assembly of the complex. This may have important implications in the maturation of megakaryocyte granules and in the molecular mechanism underlying the Glanzmann thrombastenic disease.
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Schick, Paul K., Barbara P. Schick, and Pat Webster. "THE EFFECT OF OMEGA 3 FATTY ACIDS ON MEGAKARYOCYTE ARACHIDONIC ACID METABOLISM." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1642953.
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Dietary omega 3 polyunsaturated fatty acids are thought to prevent atherosclerosis. It has been proposed that omega 3 fatty acids modify platelet arachidonic acid (20:4) metabolism and platelet function and thereby reduce the incidence of thrombosis. We have previously shown that megakaryocytes (MK), like platelets, contain large amounts of esterified 20:4. The study addresses the following questions: 1) Do omega 3 fatty acids have a primary action on 20:4 metabolism in MK rather than in platelets. 2) Do omega 3 marine oils, docosahexaenoic acid (22:6) and eicosapentaenoic acid (20:5), have a different effect on megakaryocyte 20:4 metabolism than does alpha linolenic acid (18:3), the major omega-3 fatty acid present in normal diets? 3) How do omega-3 fatty acids modify megakaryocyte 20:4 acid metabolism? MK and platelets were isolated from guinea pigs. Isolated cells were incubated with radiolabeled 20:4 acid and unlabeled 18:3, 20:5 or 22:6. Incubations were terminated by lipid extraction, lipid classes were separated by thin-layer chromatography and the incorporation of radiolabeled 20:4 into lipid species was measured by scintillation spectrometry.MK (106) can incorporate about 4 times more 20:4 than 109 platelets. We have previously shown that 20:4 is incorporated into all endogenous pools of 20:4 in MK while platelets appear to have a limited capacity to incorporate 20:4 into phosphatidyl-ethanolamine (PE). Marine oils, 22:6 and 20:5, had similar effects on the incorporation of radiolabeled 20:4 in MK. Both marine oils reduced the total uptake of 20:4 in megakaryocytes but the reduction occured primarily in PE and phosphatidylserine (PS) rather than in phosphatidylcholine (PC) and phosphatidylinositol (PI). Both 20:5 and 22:6 caused a 50% reduction in the incorporation of radiolabeled 20:4 into megakaryocyte PE and PS while only a 20% reduction into PC and PI. There was a striking difference in the effect of 18:3. Even though the incubation of megakaryocytes with 18:3 reduced the uptake of 20:4, the distribution of the incorporated 20:4 in phospholipids of megakaryocytes incubated with 18:3 was similar to that in controls. Thus, 18:3 did not have a selective effect on the incorporation of 20:4 into PE or PS. Whereas megakaryocyte 20:4 metabolism was significantly affected by omega-3 fatty acids, the incubation of guinea pig or human platelets with 22:6, 20:5 or 18:3 did not result in any alteration of the incorporation of 20:4 into platelet phospholipids.20:4 may be initially incorporated into megakaryocyte PC and subsequently transfered to PE and other phospholipids. Omega 3 marine oils, 20:5 and 22:6, appear to have a selective action on the incorporation or transfer of 20:4 into PE and PS. One mechanism for these observations would be an effect of marine oils on megakaryocyte acyltransferase and/or transacylases. Omega 3 linolenic acid appears to reduce the uptake of 20:4 but does not affect the transfer of 20:4 into PE and PS since there was no selective inhibition of uptake into PE or other megakaryocyte phospholipids. The observation that marine oils did not have any effect on 20:4 metabolism in platelets indicated that omega 3 polyunsaturated fatty acids primarily affect megakaryocytes. This phenomenon may result in the production of platelets with abnormal content and compartmentalization of arachidonic acid. The localization of 20:4 in different pools in these platelets could influence the availability of esterified 20:4 for the production of thromboxanes and other eicosanoids. Another implication of the study is that omega 3 fatty acids may have a greater effect on precursor cells than on differentiated cells and tissues and influence cellular maturation.
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Dalby Kristensen, S., K. M. Roberts, and J. F. Martin. "INCREASE IN MEGAKARYOCYTE SIZE AND PLOIUY PRECEDES ACCELERATION OF ATHEROGENESIS IN THE HYPERCHOLESTEROLAEMIC RABBIT." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643412.
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Platelet derived growth factor(s) probably synthesized by the megakaryocyte are important in atherogenesis. In a pilot study destruction of the circulating platelets was induced by injection of goat serum containing a specific platelet antibody (APS) to rabbits fed a high cholesterol diet (2g per day) for 12 weeks. Seven days after APS the percentage of atheroma in the aorta measured by planimetry was increased in these animals compared to control animals on the same diet that had been injected with goat serum (GS) 7 days before. In a new study 15 pairs of male litter mate rabbits on high cholesterol diet were randomised in pairs to treatment with either APS or GS. Five pairs of animals were killed 18 hours after the injection, 5 pairs 4 days after and 5 pairs 7 days after the injection and the platelet count, mean platelet volume, megakaryocyte nuclear, cytoplasmic and total size, megakaryocyte DNA distribution and the percentage of atheroma in the aorta were measured. Comparison of these variables between the 2 groups revealed the following statistically significant findings (p<0.05) : 18 hours after the injection the platelet count was decreased and the mean platelet volume increased in the APS group. At day 4 the platelet count, megakaryocyte nuclear, cytoplasmic and total size and the megakaryocyte DNA content were increased in the APS group. At day 7 the platelet count and the percentage of the atheroma were higher in the APS group. Since platelets produced by big megakaryocytes with high DNA content are more reactive than normal platelets, we suggest that the acceleration of atheroma formation demonstrated 7 days after APS is caused by the large number of platelets with possible high concentrations of growth factor(s) derived from the large megakaryocytes with increased DNA content.
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Ogura, M., N. Tanabe, M. Hamaguchi, T. Hotta, and H. Saito. "BIOSYNTHESIS AND SECRETION OF β-THROMBOGLOBULIN BY A HUMAN MEGAKARYOBLASTIC CELL LINE ( MEG-01 )." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1644618.
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β-Thromboglobulin ( βTG ) is a well known platelet specific a-granular protein, but synthesis of βTG by human megakaryocytes has not been fully proved. A human megakaryoblastic cell line ( MEG-01 ) was investigated for the presence of βJG in the culture medium and cell lysates using a specific radioimmunoassay ( RIA ). The concentration of βTG increased with time in the serum-free culture medium as well as in the cell lysates as shown in the following table.By an indirect immunofluorescent technique using a monospesific rabbit anti serum against human βTG, βTG antigen was detected in MEG-01 cells. Immunoblot analysis of culture medium revealed a single band ( mol wt 8,900 ) that is identical to the band of human plasma βTG. De novo synthesis of βTG was demonstrated by the presence of specific immunoprecipitable radioactivity in the medium after 5 h of labeling of the cells with [35S]-methionine as a 8,900 mol wt protein. These results indicate that human megakaryocytes produce βTG, The production of βTG by MEG-01 cells may be useful for the study of megakaryocyte maturation and differentiation.
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Uzan, G., A. Lajmanovich, M. H. Prandini, Ph Frachet, A. Duperray, and G. Marguerie. "MOLECULAR CLONING OF PLATELET GPIIb FROM HEL CELLS AND HUMAN MEGAKARYOCYTES." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643960.
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Platelet GP IIb-IIIa is an heterodimer which functions as a receptor for fibrinogen, fibronectin and Von Willebrand factor and is implicated in platelet adhesive reactions. To study the structure function relationship of this glycoprotein, a recombinant DNA approach was initiated. cDNA expression libraries were constructed in » gtll vector, from erythro-leukemia cells (HEL) and megakaryocytes mRNA. The human megakaryocytes were isolated from patients with chronic myeloid leukemia. The HEL library was initially screened with polyclonal antibodies anti GPIIb IIIa. One clone, λIIbI, containing a 1.65 kbp insert reacted with a panel of different polyclonal antibodies anti GPIIb IIIa and a monoclonal antibody anti GPIIb. To further characterize this clone the synthesis of the fusion protein was induced by IPTG. The bacterial protein was then blotted onto nitro cellulose and incubated with antisera anti GPIIb-IIIa. Antibodies that specifically bound with the fusion protein were eluted and tested on platelet membrane extracts. The selected antibodies produced a positive signal at the GPIIb position similar to the signal produced by the monoclonal antibody anti GPIIb on the same membrane extract. Finally on western blotting, a protein of Mr= 170kD reacted with the monoclonal antibody anti GPIIb. λIIbI insert was used to screen the megakaryocyte library and 3 clones, λIIb2,λIIb3 and λIIb4 were isolated. The size of HEL cells and megakaryocytes GPIIb mRNA was estimated by northern blotting. Only one species of 3.9 kb was identified in both cells. The four different clones accounted for 50% of the coding sequence of this mRNA.Sequencing of these cDNAs indicated that the plasmatic domain of GPIIb contains a cystein rich region. The sequence of these clones will allow the study of the adhesines genetic diversity in different cellular systems.
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Dalby Kristensen, S., K. M. Roberts, J. Lawry, and J. F. Martin. "SHORT TERM HIGH CHOLESTEROL DIET CAUSES CHANGES IN MEGAKARYOCYTE SIZE AND IN VASCULAR ULTRASTRUCTURE." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643411.
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Platelets produced by megakaryocytes (MK) have a role in atherogenesis. Six pairs of male litter mate rabbits were randomised to feeding with either 2g of cholesterol daily in addition to their normal diet or normal diet alone. After seven days the animals were killed and serum cholesterol, platelet count, MK total, cytoplasmic and nuclear area (microscopic planimetry) and MK DNA content cell distribution (fluorescent activated cell sorting) were measured and compared between the two groups. The results are given in the table as medians with range values in brackets.After perfusion-fixation the aortas were examined by transmission electron microscopy. In the aortas from the animals on high cholesterol diet cells with ultrastructural features resembling smooth muscle cells were found in the intima. Changes in megakaryocyte size are associated with the occurrence of smooth muscle cell proliferation and migration. The platelet-megakaryocyte axis may be activated in early atherogenesis.
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Hoffman, R., B. J. Roth, G. W. Sledge, J. Straneva, and J. Brandt. "ANALYSIS OF PHORBOL ESTER STIMULATED HUMAN MEGAKARYOCYTE DEVELOPMENT." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1642951.
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The events that occur during the terminal maturation of human megakaryocytes are poorly characterized. To examine these events, a recently characterized human megakaryocytic cell line (EST-IU, Cancer Res. 46: 2155-2159, 1986) was exposed to 12-0-tetradecanoyl-phorbol-13-acetate (TPA), as well as 2 non-transforming phorbol esters (4 alpha phorbol and 4 beta phorbol 12 alpha, 13 alpha diacetate) at the identical concentrations. Morphologic changes, including cellular attachment to untreated plastic or glass, occurred within 4 hrs of treatment with TPA. Treatment of EST-IU cells with either of the 2 non-transforming phorbols (4-alpha phorbol, or 4-beta phorbol, 12-beta, 13-alpha diacetate) failed to change morphology, DNA content, or expression of surface membrane glycoproteins or alpha-granule constituents when compared to control cells. TPA treatment resulted, however, in j^rofound changes in adherence to plastic by the EST-IU cells, with an obvious dose-response relationship. At a 5 × 10-8 M TPA, cellular attachment was noted as early as 4 hours following treatment, agd was complete by 16 hours, at which time > 95% of treated cells were attached. Following TPA treatment at 5 × 10-8 M, a number of morphologic changes occurred, including marked cellular flattening, the appearance of extensive cytoplasmic budding, and the development of numerous filopodia. Cells treated with either of the non-transforming phorbols as assessed by propidium iodide staining and flow cytometric analysis failed to exhibit a change in ploidy, although TPA reproducibly altered this parameter of megakaryocyte development. Cells treated with 10-9 M TPA have approximately the same proportion of cells in the 4N and 8N peaks as control cells. Following exposure to 10-9 M and 10-8 M TPA, there was an apparent shift of cells out of the 4N peak to 8N and 16N levels, and even the appearance of a small percentage of 32N cells. The DNA content of TPA-treated cells was also assessed by Feulgen staining and microdensitome try. Those cells (5%) which failed to adhere following TPA treatment were analyzed separately, and showed a very different ploidy distribution than the adherent cell population. Over 85% of adherent cells have a ploidy > 16N, with some cells attaining the 128N level. Treatment of cells with either of the 2 non-transforming phorbols failed to affect the expression of Factor V, Factor VIIIrRAg, beta-throraboglobulin, fibrinogen, or platelet glycoproteins. Cells treated with 5 × 10-8 M TPA similarly do not significantly increse the expression of Factor V, fibrinogen, or beta-thromoglobulin over that observed in control cells. The expression of both Factor VIIIrRAg and platelet glycoproteins however, increase in TPA-treated cells. A similar increase in the expression of platelet glycoprotein Ilb/IIIA using the mouse monoclonal C17 was also observed. Those cells that express the highest levels of Factor VIII:RAg and platelet glycoproteins following phorbol treatment also demonstrated the highest ploidy levels and also are the largest cells as measured by forward angle light scatter during flow cytometry.These studies indicate that TPA treatment of EST-IU cells initiates a cascade of events characterized by cellular adherence, increases in cell size and DNA content, and enhanced expression of platelet glycoproteins and Factor VIIIrRAg. These events appear to occur in concert and closely resemble information that is available concerning maturation of normal rodent and human megakaryocytes. Although it is important to emphasize that EST-IU cells are leukemic and thus intrinsically different from normal human megakaryocytes, their availability and dynamic responses to TPA will provide an appropriate cellular model with which to study megakaryocyte maturation.
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Deschamps, J. F., E. Bodevin, and J. P. Caen. "INCREASED SPONTANEOUS NUMBER OF MEGAKARYOCYTE COLONIES IN ESSENTIAL THROMBOCYTHEMIA (ET)." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643543.
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Megakaryocytes were grown from medullary progenitor cells using the plasma clot technique, with 5 % human serum and with or without 2,5 % PHA-LCM. Using this technique megakaryocyte cultures were done in 5 patients essential thrombocythemia (ET) ( platelet count :0,6 x 106 to 1,4 x 106 /μi) before chemotherapy and antiplatelet agents and in 2 patients with secondary thrombocytosis (ST) (platelet count : 0,65 x lO6 and 0,8 x 106 /μi) corrected after effective anti-bacterial or iron therapy. The results were as followsx number of colonies mean number (in brackets) of cells per colonyIt appears therefore that in ET, megakaryocyte progenitors grow without PHA-LCM and show however a better proliferation in its presence. On the contrary in ST, PHA-LCM is required for obtaining a 6 times increase of MK colonies. The effect of MK growth under chemotherapy is reported in some of the patients studied before treatment and results analyzed and compared to platelet function involvement
Reports on the topic "Megakaryocytes"
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Li, Xin, and Laurie McCauley. Inhibitory Effects of Megakaryocytes in Prostate Cancer Bone Metastasis. Fort Belvoir, VA: Defense Technical Information Center, April 2010. http://dx.doi.org/10.21236/ada542172.
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Mastro, Andrea M. The Role of Megakaryocytes in Breast Cancer Metastasis to Bone. Fort Belvoir, VA: Defense Technical Information Center, May 2014. http://dx.doi.org/10.21236/ada609025.
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Mastro, Andrea M. The Role of Megakaryocytes in Breast Cancer Metastasis to Bone. Fort Belvoir, VA: Defense Technical Information Center, May 2011. http://dx.doi.org/10.21236/ada549461.
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Cohen, Isaac, and Jane N. Winter. Ex vivo Expanded Megakaryocytes for Supportive Care of Breast Cancer Patients. Fort Belvoir, VA: Defense Technical Information Center, October 2001. http://dx.doi.org/10.21236/ada400103.
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