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1.
Blood Research ; : 102-102, 2018.
Artigo em Inglês | WPRIM | ID: wpr-715049
3.
China Journal of Chinese Materia Medica ; (24): 1685-1689, 2014.
Artigo em Chinês | WPRIM | ID: wpr-300205

RESUMO

<p><b>OBJECTIVE</b>To investigate the effects of total saponins from Sanguisorba officinalis (DYS) on hematopoietic cell proliferation, differentiation and the expression level of IL-3R and c-kit.</p><p><b>METHOD</b>Baf3 and 32D cells were cultured with or without IL-3, then the cells were exposed to DYS in different concentrations of 5, 10, 20, 30 and 40 mg x L(-1) for 24, 48, 72 and 96 hours separately. After that, the cell proliferation and differentiation capacity were determinated by the methods of CCK8 and Giemsa staining separately. The effects of DYS on the expression level of IL-3 receptor in Baf3 cells and the expression level of c-kit in 32D cells were determinated using RT-PCR.</p><p><b>RESULT</b>DYS promotes alone proliferation of Baf3 cells and 32D cells after 48 h. In contrast to control cells, 32D cells containing DYS without IL-3 form many large clusters. DYS also increases the proliferation when cultured with IL-3. High concentration of DYS induce alone the differentiation of 32D cells and increase alone the number of the polyploidy megakaryocyte. Moreover, DYS increases alone the expression level of IL-3R in Baf3 cells and the expression level of c-kit in 32D cells separately.</p><p><b>CONCLUSION</b>Our data shows DYS can promote alone proliferation and differentiation of megakaryocyte progenitor cells. The proliferative and differentiative effect of DYS on megakaryocyte progenitor cells is correlated to the up-regulation of IL-3 receptor and c-kit expression.</p>


Assuntos
Animais , Camundongos , Diferenciação Celular , Genética , Linhagem Celular , Proliferação de Células , Relação Dose-Resposta a Droga , Expressão Gênica , Interleucina-3 , Farmacologia , Células Progenitoras de Megacariócitos , Metabolismo , Proteínas Proto-Oncogênicas c-kit , Genética , Receptores de Interleucina-3 , Genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sanguisorba , Química , Saponinas , Farmacologia , Fatores de Tempo
4.
Chinese Journal of Hematology ; (12): 187-190, 2014.
Artigo em Chinês | WPRIM | ID: wpr-295678

RESUMO

<p><b>OBJECTIVE</b>To build a protocol of separation and induction of megakaryocytes derived from cord blood mononuclear cells.</p><p><b>METHODS</b>Red blood cells were precipitated by hydroxyethyl starch (HES). Mononuclear cells were obtained by density gradient centrifugation with Ficoll. The inducing efficiencies of megakaryocytes by using of different cytokine cocktails and culture media were analyzed.</p><p><b>RESULTS</b>The best choice for erythrocyte sedimentation and high efficiency of nucleated cells retrieving were obtained by using of 1.5% HES. The isolated cord blood mononuclear cells were cultured with domestic serum-free medium supplemented with 116t (IL-11, IL-6, TPO), st36(SCF, TPO, IL-3, IL-6), pt36 (PDGF,TPO,IL-3,IL-6) or pst36 for 7 days. St36 group (50 ng/ml SCF, 50 ng/ml TPO, 20 ng/ml IL-3 and 50 ng/ml IL-6) yielded the most CD41/CD61 positive [(6.79±1.97)×10⁴]. The cell viability [(82.85 ± 0.64)%] of st36 group by using of imported serum-free medium was better than [(60.90±6.93)%] that in domestic medium on day 7 after induction, and CD41/CD61 positive cells count [(18.60±1.97)×10⁴] were more than domestic serum-free medium group. Therefore, we chose imported serum-free medium containing st36 to induce cord blood mononuclear cells. After a prolonged culture, the total cell numbers increased accompanied with an elevated percentage of CD41/CD61 positive cells, which reached (54.27 ± 6.31)% on day 14. Wright-Giemsa staining showed that different phase cells, such as megakaryoblast, promegakaryocyte and granular megakaryocyte, occurred after 10 days'culture. Clone forming unit-megakarocytes (CFU-MK) assay showed that the colonies count increased with the prolonged incubation. CFU-MK colonies were [1 236.0±32.9] on day 14, which was higher than that in medium without induction (P<0.01). Platelets from megakaryocytes showed agglutination function after 10 days'culture.</p><p><b>CONCLUSION</b>1.5% HES was the best solution to precipitate erythrocytes. The combination of an imported serum-free medium with IL-3, IL-6, SCF and TPO showed better induction efficiency than domestic medium or other cytokine cocktails. Meanwhile, induced megakaryocytes produced functional platelets.</p>


Assuntos
Humanos , Técnicas de Cultura de Células , Diferenciação Celular , Divisão Celular , Separação Celular , Métodos , Células Cultivadas , Meios de Cultura Livres de Soro , Sangue Fetal , Biologia Celular , Células Progenitoras de Megacariócitos , Biologia Celular
5.
Pakistan Journal of Medical Sciences. 2010; 26 (2): 352-357
em Inglês | IMEMR | ID: emr-97979

RESUMO

To observe the K562 cell line derived from a patient of chronic myelogenous leukemia differentiated into megakaryocytes by growing in the presence of phorbol myristate acetate [PMA]. The differentiation process of K562 cells was monitored by the expression of a platelet cell marker, CD61 through immunocytochemistry using mouse alkaline phosphatase antialkaline phosphatase [APAAP] complex employing fast red TR as substrate, crystal violet and MTT assay used for cell growth analysis. The crystal in the presence of PMA, cells obtained were of large size and less in number as compared to cells incubated without PMA where they were of smaller size and more in number and immunochemical reaction used to detect the presence of CD61, a platelet cell marker that is expressed during differentiation of K562 cells to megakaryocytes. The results showed that the addition of PMA to the growing culture of K562 cell lines induced differentiation, observed through CD61 expression and increase in cell size and cessation of proliferation


Assuntos
Humanos , Células K562/efeitos dos fármacos , Acetato de Tetradecanoilforbol , Células Progenitoras de Megacariócitos/efeitos dos fármacos , Leucemia Mielogênica Crônica BCR-ABL Positiva
6.
Journal of Experimental Hematology ; (6): 1121-1125, 2008.
Artigo em Chinês | WPRIM | ID: wpr-234286

RESUMO

This study was purposed to investigate the biological characteristics and immunogenicity changes of ex vivo expanded megakaryocyte progenitors from human umbilical cord blood CD34(+) cells in order to provide experimental basis for clinical application of ex vivo expanded umbilical cord blood megakaryocyte progenitor cells. Mononuclear cells (MNCs) were obtained from umbilical cord blood by Ficoll-Hyapaque density gradient separation. CD34(+) cells were enriched by magnetic cell sorting (MACS). The selected CD34(+) cells were seeded in serum-free medium stimulated with thrombopoietin (TPO, 50 ng/ml), interleukin 11 (IL-11, 50 ng/ml), and heparin (25 U/ml) for 14 days. The immunophenotyping (CD34(+), CD41a(+), CD61(+), CD34(+) CD41a(+) and CD34(+) CD61(+) cells) of amplificated products, matured megakaryocyte apoptosis, and expression of human leukocyte antigen (HLA) class I and class II molecules were measured by fluorescence-activated cell sorter (FACS). The number of colony-forming units-megakaryocyte (CFU-Mk) was also evaluated by CFU-Mk assay. The results showed that the umbilical cord blood CD34(+) mononuclear cells could be effectively differentiated into megakaryocytes. The peak expression ratios of CD41a(+) and CD61(+) cells were all at 14th days, while that of CD34(+) CD41(+) and CD34(+) CD61(+) cells were at 7th day [(3.41 +/- 2.80)% and (1.89 +/- 1.43)%, respectively]. The expansion times of large and small CFU-Mk reached peak at 7th day (20.66 +/- 32.79) and 10th day (435.62 +/- 482.65), respectively. The apoptotic rates of megakaryocytes at 7th, 10th, 14th day were (19.48 +/- 9.64)%, (26.87 +/- 9.03)%, and (52.46 +/- 11.74)%, respectively. The apoptotic rate of megakaryocytes had no significant difference in 7 and 10 days culture (p > 0.05), while that significantly increased in culture for 14 day culture, compared with culture for 7 and 10 days (p < 0.05, respectively). The expression of HLA class I and class II molecules on megakaryocytes decreased along with the prolongation of expansion time and sharply decreased in 0 to 10 days. It is concluded that the cytokines of TPO, IL-11, and heparin can promote the expansion of megakaryocyte progenitors from umbilical cord blood CD34(+) mononuclear cells effectively in vitro. The peaked expansion times of large CFU-Mk, the peaked expression ratios of CD34(+) CD41(+) and CD34(+) CD61(+) cells were all at 7th day. So the culture for 7 days appeared to be the optimal duration of expanding megakaryocyte progenitors.


Assuntos
Humanos , Antígenos CD34 , Alergia e Imunologia , Diferenciação Celular , Divisão Celular , Separação Celular , Células Cultivadas , Sangue Fetal , Biologia Celular , Alergia e Imunologia , Células Progenitoras de Megacariócitos , Biologia Celular
7.
Journal of Experimental Hematology ; (6): 1398-1402, 2008.
Artigo em Chinês | WPRIM | ID: wpr-234225

RESUMO

The purpose of this study was to explore the differences in megakaryocyte progenitor ex vivo expansion between CD34+ cells derived from human umbilical cord blood (CB) and bone marrow (BM). Mononuclear cells (MNCs) were obtained from CB or BM by Ficoll-Hypaque density gradient separation. CD34+ cells were purified by magnetic cell sorting (MACS). The selected CD34+ cells were seeded in serum-free conditions stimulated with thrombopoietin (TPO), TPO+interleukin 11 (IL-11), or TPO+IL11+heparin for 14 days. Amplification product (CD34+, CD41a+, and CD34+ CD41a+ cells) immunophenotypes, megakaryocyte apoptosis rates and the DNA content were measured by fluorescence-activated cell sorting (FACS). The colony-forming units of granulocytes and monocytes (CFU-GM), burst-forming units of erythrocytes (BFU-E), and colony-forming units of megakaryocytes (CFU-Mk) were also evaluated by the colony-forming units (CFU) assay. The results indicated that CD34+ cells derived from CB showed higher expansion ability of total cell counts, CD41a+ and CD34+ CD41a+ cells than those derived from BM for all days 14 of culture (p<0.05, respectively). There were no significant differences in CFU-GM, BFU-E, and total CFU-Mk counts between CB and BM-derived CD34+ cells on day 0 (p>0.05, respectively), but CB-derived CFU-Mk seemed mainly large colonies, and the number of large colonies was higher than that from BM (p<0.05) on day 0. There were no significant differences in expansion ability of CFU-GM between CB and BM-derived cells on days 7, 10, and 14 of culture (p > 0.05, respectively), but the expansion ability of BFU-E and CFU-Mk derived from CB cells was higher than that from BM (p<0.05, respectively). There were no significant differences in apoptosis rates of megakaryocyte from two source cells for days 14 of culture. Megakaryocytes derived from CB mostly showed the 2N DNA content (>90%) for days 14 of culture, while those cells derived from BM showed the increased DNA content, and 4N, 8N or more ploidy cells gradually increased with prolonging of culture time. It is concluded that CB-derived CD34+ cells have a greater proliferation potential than that derived from BM, which is therefore proven to be a better cell source for megakaryocyte progenitor expansion in vitro.


Assuntos
Humanos , Antígenos CD34 , Células da Medula Óssea , Biologia Celular , Alergia e Imunologia , Técnicas de Cultura de Células , Métodos , Diferenciação Celular , Divisão Celular , Separação Celular , Células Cultivadas , Células Precursoras Eritroides , Biologia Celular , Sangue Fetal , Biologia Celular , Alergia e Imunologia , Células Progenitoras de Megacariócitos , Biologia Celular , Alergia e Imunologia
8.
Chinese Journal of Hematology ; (12): 615-618, 2008.
Artigo em Chinês | WPRIM | ID: wpr-239972

RESUMO

<p><b>OBJECTIVE</b>To study the expansion potential of megakaryocyte progenitor cells (MPC) from human placenta tissue CD133+ (PT-CD133+) cells.</p><p><b>METHODS</b>PT-CD133+ cells were purified from mononuclear cells (MNC) by magnetic activated cell sorting (MACS) and seeded in serum-free liquid culture medium supplemented with thrombopoietin (TPO), interleukin-3 (IL-3), and stem cell factor (SCF) to expand MPC. At day 7, 10 and 14, the total cell number was counted and the dynamic changes of CD133, CD34, and CD41 antigens expression during ex-vivo expansion were analyzed by flow cytometry (FCM). PT-CD133+ cells at different expansion time were collected and cultured in collagen semisolid medium for colony forming units-megakaryocyte (CFU-MK) assay.</p><p><b>RESULTS</b>PT-CD133+ cells could be optimally expanded at day 7 by 13 +/- 2 fold increase in serum-free liquid culture systems and the total cell number was expanded by 160 fold at day 14. With the expansion time going on, the expression of CD133, CD34 decreased and that of CD41 increased. The expanded megakaryocytes were immature and no sign of platelet formation. Both PT-CD133+ cells before and after expansion could form CFU-MK, the total number of CFU-MK peaked at day 10 of expansion by 54 +/- 10 fold increase.</p><p><b>CONCLUSION</b>Human PT-CD133+ cells have a high capacity of MPC expansion, 10 days culture could give rise to the maximum number of CFU-MK.</p>


Assuntos
Feminino , Humanos , Gravidez , Antígeno AC133 , Antígenos CD , Diferenciação Celular , Células Cultivadas , Glicoproteínas , Células Progenitoras de Megacariócitos , Biologia Celular , Peptídeos , Placenta , Biologia Celular
9.
Chinese Journal of Pediatrics ; (12): 346-349, 2006.
Artigo em Chinês | WPRIM | ID: wpr-309203

RESUMO

<p><b>OBJECTIVE</b>Idiopathic thrombocytopenic purpura (ITP) is a hemorrhagic disease in children with blood platelets redundant destruction caused by chaotic immunological mechanism. However, some patients with ITP with negative platelet-associated antibody and ineffective adrenal cortical hormone therapy probably have special pathogenesis. It is indicated that the human cytomegalovirus (HCMV) can incubate in haemopoietic stem cell/ancestral cell to inhibit its generation and differentiation. Therefore, the study was designed to investigate HCMV-late mRNA expression in megakaryoblast for the purpose of examining the pathogenesis of ITP and to examine the effectiveness of ganciclovir on ITP.</p><p><b>METHODS</b>Colony forming unit-megakaryocyte (CFU-MK) of 46 ITP patients with HCMV infection were incubated. Reverse transcription-polymerase chain reaction (RT-PCR) was subsequently used for HCMV-late mRNA detection. Ganciclovir therapy was given to both positive group and negative group for comparison of therapeutic effectiveness.</p><p><b>RESULTS</b>Nineteen out of 46 CFU-MK culture cell specimens with positive HCMV-DNA by PCR or positive CMV-IgM by enzyme linked immunosorbent assay (ELISA) from serum of peripheral blood showed positive for HCMV-late mRNA. While, the remaining 27 were negative. Sixteen positive responders to ganciclovir therapy were observed amongst those with positive HCMV-DNA. Whereas, only 4 positive responders to ganciclovir therapy were noticed amongst those with negative HCMV-DNA. The curative effectiveness in positive group was significantly higher than that in negative group (P < 0.01).</p><p><b>CONCLUSION</b>HCMV can directly infect CFU-MK, which might be one of the mechanisms responsible for ITP. Ganciclovir is an effective therapy resulting in an increase in thrombocyte in ITP patients whose HCMV-late mRNA was positive in their CFU-MK.</p>


Assuntos
Adolescente , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Masculino , Anticorpos Antivirais , Alergia e Imunologia , Antivirais , Usos Terapêuticos , Plaquetas , Alergia e Imunologia , Células Cultivadas , Citomegalovirus , Genética , Virulência , Infecções por Citomegalovirus , Tratamento Farmacológico , Genética , Alergia e Imunologia , Ensaio de Imunoadsorção Enzimática , Ganciclovir , Usos Terapêuticos , Imunoglobulina M , Alergia e Imunologia , Células Progenitoras de Megacariócitos , Metabolismo , Púrpura Trombocitopênica Idiopática , Tratamento Farmacológico , Alergia e Imunologia , RNA Mensageiro , RNA Viral , Metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa
10.
Acta Physiologica Sinica ; (6): 247-253, 2005.
Artigo em Chinês | WPRIM | ID: wpr-334179

RESUMO

In this study the effects of bone marrow stromal cells conditioned medium on the expansion of mature megakaryocytes and colony forming unit-megakaryocyte (CFU-Meg) in vitro were investigated. The serum-free bone marrow fibroblast conditioned medium (F-CM), bone marrow endothelial cell conditioned medium (E-CM) and bone marrow macrophage conditioned medium (M-CM) were collected and ultrafiltrated by using Centriprep-10. F-CM, E-CM, M-CM, the retentate (>10 kDa F-CM, >10 kDa E-CM and >10 kDa M-CM) contained substances whose molecular weight was more than 10 kDa and the filtrate (<10 kDa F-CM, <10 kDa E-CM and <10 kDa M-CM) contained substances whose molecular weight was less than 10 kDa were added in liquid culture system respectively. The results showed that F-CM, >10 kDa F-CM and E-CM, >10 kDa E-CM significantly promoted the expansion of mature megakaryocytes in liquid culture system, the percentage of mature megakaryocytes compared with 0 h control were (287.33-/+16.77)%, (236.67-/+39.72)%, (141.21-/+17.47)% and (179.03-/+30.98)%. But <10 kDa F-CM, <10 kDa E-CM, M-CM, >10 kDa M-CM and <10 kDa M-CM had no positive effects on the expansion of mature megakaryocytes. The effects of F-CM, E-CM or M-CM on the expansion of CFU-Meg were also investigated. The results indicated that F-CM and E-CM promoted the expansion of CFU-Meg in liquid culture system. M-CM had no positive effect on the expansion of CFU-Meg. the percentage of CFU-Meg compared with 0 h control were (168.18-/+30.24)%, (215.17-/+17.4)% and (85.0-/+7.0)%. The results of reverse transcription-polymerase chain reaction (RT-PCR ) showed that transforming growth factor -beta1 (TGF-beta1) mRNA was expressed in bone marrow endothelial cells and was not expressed in bone marrow fibroblasts. Thrombopoietin (TPO) mRNA was expressed in bone marrow fibroblasts and was not expressed in bone marrow endothelial cells. These results suggest that F-CM, >10kDa F-CM and E-CM, >10kDa E-CM significantly promoted the expansion of mature megakaryocytes and CFU-Meg in liquid culture system. The effect of F-CM on the expansion of mature megakaryocytes is better than that of E-CM.


Assuntos
Animais , Feminino , Masculino , Camundongos , Células da Medula Óssea , Biologia Celular , Diferenciação Celular , Fisiologia , Células Cultivadas , Ensaio de Unidades Formadoras de Colônias , Meios de Cultivo Condicionados , Farmacologia , Células Progenitoras de Megacariócitos , Biologia Celular , Megacariócitos , Biologia Celular , Células Estromais , Biologia Celular
11.
Chinese Journal of Hematology ; (12): 720-723, 2004.
Artigo em Chinês | WPRIM | ID: wpr-229911

RESUMO

<p><b>OBJECTIVE</b>To explore the suppression effect of human cytomegalovirus (HCMV) on megakaryocytes and their precursors and study the antiviral effect of antisense phosphorothioate deoxyoligonucleotide (ASON) against HCMV.</p><p><b>METHODS</b>CD34(+) cells were induced to proliferate and differentiate committedly to megakaryocytes in a semi-solid CFU-MK culture system. Cultured cells and ASON pretreated CD34(+) cells were infected by HCMV of AD169 strain. HCMV immediate early protein (IEP) DNA and mRNA and UL36 mRNA were detected by PCR and reverse transcription-polymerase chain reaction (RT-PCR). Cytotoxicity was evaluated by MTT assay.</p><p><b>RESULTS</b>HCMV AD169 suppressed the proliferation of megakaryocytes significantly. Compared with the mock group, the CFU-MK yields were decreased by 21.6%, 33.8%, and 46.3%, respectively, in 3 different titers of virus infected groups (P < 0.05). The suppression was virus titer dependent. HCMV IEP DNA, HCMV IEP mRNA and UL36 mRNA were detected in the colony cells of viral infection group. Compared with the infected group by HCMV AD169, UL36Anti treatment at 0.08 micromol/L could recover the CFU-MK yields significantly (P < 0.05). In the infected MK, which was pretreated with UL36Anti at 0.08 micromol/L, HCMV UL36 mRNA was undetectable by RT-PCR. The oligonucleotide MM(1) containing a G-to-C substitution in UL36Anti was inactive at 0.08 micromol/L but active at 0.40 micromol/L. The concentration of UL36Anti necessary to significantly affect cell growth was 90.00 micromol/L.</p><p><b>CONCLUSIONS</b>HCMV AD169 infection inhibits the proliferation and differentiation of megakaryocytes and their precursors. There are early transcriptions of HCMV IE and UL36 protein in infected CFU-MK. The specific ASON has a definite anti-HCMV activity.</p>


Assuntos
Humanos , Recém-Nascido , Antígenos Virais , Genética , Antivirais , Farmacologia , Diferenciação Celular , Proliferação de Células , Sobrevivência Celular , Células Cultivadas , Citomegalovirus , Genética , Fisiologia , Sangue Fetal , Biologia Celular , Interações Hospedeiro-Patógeno , Proteínas Imediatamente Precoces , Genética , Células Progenitoras de Megacariócitos , Biologia Celular , Virologia , Oligonucleotídeos Antissenso , Genética , Farmacologia , RNA Mensageiro , Genética , Metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa
12.
Korean Journal of Pediatric Hematology-Oncology ; : 262-268, 2003.
Artigo em Coreano | WPRIM | ID: wpr-194907

RESUMO

PURPOSE: We investigated the expression of vasoactive intestinal peptide (VIP), VIP receptor 1 (VPAC1), VIP receptor 2 (VPAC2) genes in the human umbilical cord blood CD34 cells, and the ability of VIP to stimulate human primitive as well as monopotent hematopoietic progenitors. METHODS: We isolated RNA from umbilical cord blood CD34 cells, and then performed RT-PCR, and sequencing. The umbilical cord blood CD34 cells were cultured with the various concentrations of VIP for burst-forming unit of erythrocyte (BFU-E), colony-forming unit of granulocyte/monocyte (CFU-GM), colony-forming unit of graulocyte/erythrocyte/monocyte/megakaryocyte (CFU-GEMM), and colony-forming unit of megakaryocyte (CFU-Mk). RESULTS: The RNA coding for VPAC1 was detected in human umbilical cord blood CD34 cells. VIP significantly stimulated the growth of CFU-GEMM and CFU-Mk. CONCLUSION: The present results suggest that VIP is an important neuropeptide in the early proliferation of human primitive as well as megakaryocyte progenitors.


Assuntos
Humanos , Codificação Clínica , Eritrócitos , Sangue Fetal , Células Progenitoras de Megacariócitos , Megacariócitos , Células Progenitoras Mieloides , Neuropeptídeos , Receptores de Peptídeo Intestinal Vasoativo , RNA , Células-Tronco , Peptídeo Intestinal Vasoativo
13.
Korean Journal of Anatomy ; : 285-296, 2002.
Artigo em Coreano | WPRIM | ID: wpr-650629

RESUMO

Liver tissuses obtained from 5 human fetuses between 11 weeks and 23 weeks of gestation during the high activity of hepatic hemopoiesis were observed with transmission electron microscope using continuous series of thin sections. The objective of present study was to evaluate ultrastructures of megakaryopoietic cells, the migration of extravascular megakaryocyte into the sinusoidal lumen and the relevence between a migrated megakaryocyte and a Kupffer cell. Immature megakaryocytes were usually observed between growing hepatic laminae and within hepatic sinusoids. A megakaryoblast contained numerous polyribosomes, rather large mitochondria, short tubular elements of rough endoplasmic reticulum and small granules. Moreover, demarcation tubules and a few small specific granules were observed in immature megakaryocytes. The nucleus was mononuclear but frequently indented. With maturation, the nuclei were multilobulated. In the cytoplasm, in contrast to the decrease in polyribosomes and rough endoplasmic reticulum, the numerous specific granules and well -developed demarcation membrane system were predominant. Thereafter cytoplasmic zonation was observed clearly in maturing and mature megakaryocytes. Some megakaryocytes passed through the sinusoidal lining epithelium and into the hepatic sinusoids. The cell to cell interaction was often found as adhesion between migrated megakaryocyte and Kupffer cell, and erythroblasts within megakaryocyte (emperipolesis). These results suggest that intravascular megakaryopoiesis in addition to extravascular megakaryopoiesis occurs to produce platelet during the human fetal liver.


Assuntos
Humanos , Gravidez , Plaquetas , Comunicação Celular , Citoplasma , Emperipolese , Retículo Endoplasmático Rugoso , Epitélio , Eritroblastos , Feto , Fígado , Células Progenitoras de Megacariócitos , Megacariócitos , Membranas , Mitocôndrias , Polirribossomos , Trombopoese
14.
Korean Journal of Hematology ; : 232-240, 2001.
Artigo em Coreano | WPRIM | ID: wpr-720528

RESUMO

BACKGROUND: Rotor off (R/O) fraction obtained by counterflow centrifugal elutriation (CCE) contains small number of T cells and many hematopoietic stem cells. Since megakaryocytes and its progenitors are larger than other cells in bone marrow, it may be easier to be separated by CCE and newly applied to hematopoietic stem cell transplantation (HSCT) for early megakaryocytes reconstitution. METHODS: The marrow cells of BALB/c mice in each group (17, 25, 28mL/min, and R/O fraction) were cultured for quantifying CFU-MK and measured after 10 days. BALB/c mice were lethally irradiated and transplanted with R/O cells. The dosages of transplanted cells were 5x104 in Group A, 5x105 in Group B, and 5x106 in Group C. The platelet counts in peripheral blood were measured up to post-transplant day 14. RESULTS: After CCE, recovery rate of the loaded cells was 82.2% and the R/O fraction was 35.9%. Most CFU-MK were formed in R/O fraction, and Group C showed the fastest recovery. Group A couldn't reach the level of 100x109/L until post-transplant day 14, and Group B showed slower recovery compared to Group C. All 5 mice survived in Group C, but 2 out of 5 mice survived in Group A and B. CONCLUSIONS: R/O fraction contains higher number of megakaryocyte progenitors, and CCE could be an effective method for separating megakaryocyte progenitors essential for reconstituting platelet after HSCT. The cell dose of 5x106 was required for the effective recovery of platelet and the survival of BALB/c mice in syngeneic bone marrow transplantation with R/O cells.


Assuntos
Animais , Camundongos , Plaquetas , Medula Óssea , Transplante de Medula Óssea , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas , Células Progenitoras de Megacariócitos , Megacariócitos , Contagem de Plaquetas , Linfócitos T , Transplante Isogênico
15.
Korean Journal of Pediatric Hematology-Oncology ; : 138-147, 1998.
Artigo em Coreano | WPRIM | ID: wpr-199965

RESUMO

BACKGROUND: Development of hematopoietic growth factor made it possible to treat anemia and granulocytopenia following intensive chemotherapy and for thrombocytopenia, recently found thrombopoietin(TPO) is being applied experimentally in several countries. The megakaryocyte colony assay can assess the effect of TPO on the thrombocytopenia resulted from cancer chemotherapy or hematopoietic stem cell transplantation. In vitro colony assay procedures for detecting human erythroid and granulocyte macrophage progenitors have been in widespread use for many years. However, reproducible assay methods for human megakaryocyte progenitors have lagged considerably behind especially in Korea. Duration platelet recovery following transplantation depends on the origin of the hematopoietic cells. Usually thrombocyte recovery is delayed following cord blood stem cell transplantation because of the small amount of cells administered. This study was carried out to investigate and establish the megakaryocyte colony assay of hematopoietic stem cells obtained from the various origin of the hematopoietic stem cells with or without TPO. METHOD: Mononuclear cells of bone marrow, peripheral blood and cord blood were collected following Ficoll density gradient centrifugation and megakaryocyte colony assay was done using MegaCultTM(Stem Cell Tech. Inc., Canada). After liquifying the agarose, mononuclear cells were added and then agarose and cell mixture were dispersed into the two wells of the chamber slide. These slides were incubated for 18~21 days at 37oC, 5% CO2. The megakaryocyte colonies were detected by staining of the cells with a primary antibody to the GPIIb/IIIa antigen, secondary antibody, alkaline phosphatase and Evans Blue in order. Changes of CD34 and GPIIb/IIIa positive cells were also analysed in flask culture using flow cytometry. RESULTS: CD34 positive cells were most abundant in the mononuclear cells of the bone marrow, meanwhile the number of CFU-GM and megakaryocyte colony were greater in the mononuclear cells of the cord blood. After administration of TPO, the cell number of megakaryocyte colony was increased dose dependently, but CFU-GM colony did not show any response to TPO. With flask culture, the cell number was decreased with or without TPO. However adding GM-CSF, IL3 and TPO to cord blood mononuclear cell, the number of the cord blood mononuclear cells was increased on the 5 th day. The amount of CD34 positive cells was increased dose dependently to TPO in one of two cord blood and one peripheral blood. The amount of GPIIb/IIIa positive cells was increased dose dependently to TPO following incubation of all the mononuclear cells. CONCLUSION: This study revealed successful result of megakaryocyte colony assay using MegaCultTM in various kinds of mononuclear cells and suggested that TPO was useful for CFU-mega colony formation. The amount of GPIIb/IIIa positive cells was increased with TPO in the flask culture. Therefore TPO could be useful for assessment of CFU- mega, and could be applied for the in vivo and in vitro expansion of megakaryocytes and platelets.


Assuntos
Humanos , Agranulocitose , Fosfatase Alcalina , Anemia , Plaquetas , Medula Óssea , Contagem de Células , Centrifugação com Gradiente de Concentração , Transplante de Células-Tronco de Sangue do Cordão Umbilical , Tratamento Farmacológico , Azul Evans , Sangue Fetal , Ficoll , Citometria de Fluxo , Fator Estimulador de Colônias de Granulócitos e Macrófagos , Células Progenitoras de Granulócitos e Macrófagos , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas , Coreia (Geográfico) , Células Progenitoras de Megacariócitos , Megacariócitos , Sefarose , Trombocitopenia
16.
Journal of the Korean Pediatric Society ; : 578-583, 1997.
Artigo em Coreano | WPRIM | ID: wpr-142150

RESUMO

We experienced a case of congenital acute megakaryoblastic leukemia with Down syndrome. The patient was admitted due to characteristic facial figure of Down syndrome and abdominal distension. Acute megakaryoblastic leukemia was diagnosed with abundant megakaryoblast in peripheral blood smear, severe myelofibrosis in bone marrow biopsy and positive platelet glycoprotein III a receptor. On third hospital day, the patient expired due to DIC and pulmonary hemorrhage. Authors report the case with review of literature.


Assuntos
Humanos , Biópsia , Plaquetas , Medula Óssea , Dacarbazina , Síndrome de Down , Glicoproteínas , Hemorragia , Leucemia Megacarioblástica Aguda , Células Progenitoras de Megacariócitos , Mielofibrose Primária
17.
Journal of the Korean Pediatric Society ; : 578-583, 1997.
Artigo em Coreano | WPRIM | ID: wpr-142147

RESUMO

We experienced a case of congenital acute megakaryoblastic leukemia with Down syndrome. The patient was admitted due to characteristic facial figure of Down syndrome and abdominal distension. Acute megakaryoblastic leukemia was diagnosed with abundant megakaryoblast in peripheral blood smear, severe myelofibrosis in bone marrow biopsy and positive platelet glycoprotein III a receptor. On third hospital day, the patient expired due to DIC and pulmonary hemorrhage. Authors report the case with review of literature.


Assuntos
Humanos , Biópsia , Plaquetas , Medula Óssea , Dacarbazina , Síndrome de Down , Glicoproteínas , Hemorragia , Leucemia Megacarioblástica Aguda , Células Progenitoras de Megacariócitos , Mielofibrose Primária
18.
Journal of the Korean Pediatric Society ; : 9-16, 1993.
Artigo em Coreano | WPRIM | ID: wpr-122996

RESUMO

We analyzed the clinical and laboratory features of ten children with acute megakaryoblastic lukemia (M7)and compared the findings with those reported in the literature. The diagnosis was supprted by ultrastructural examination of platelet peroxidase or immunophenotyping for glycoprotein IIb/IIIa. Of the ten children, five were girls and five were boys. The median age at diagnosis was 13 months. Two patients had prominent myelofibrosis and one patient had Down syndrome. Nine patients were treatd with low-dose cytosine arabinoside (10mg/m2)administered intravenously, or subcutaneously, or intramuscularly, twice daily in 21 day courses. Seven patients achieved hematologic response and three patients are alive without evidence of disease. The 4 year event free survival rate was30.0%. It is our impression that the prevalence of acute megakaryoblastic leukemia has been under-estimated, and low-dose cytosine arabinoside treatment may be of value in its management. This approach may be particularily useful in hospitals with scarce well-equipped facilities, since this protocol does not induce profound marrow hypoplasia and intensive supportive measures are not required as they would be with the use of more aggressive drug combination.


Assuntos
Criança , Feminino , Humanos , Plaquetas , Medula Óssea , Citarabina , Diagnóstico , Intervalo Livre de Doença , Síndrome de Down , Glicoproteínas , Imunofenotipagem , Leucemia Megacarioblástica Aguda , Células Progenitoras de Megacariócitos , Peroxidase , Prevalência , Mielofibrose Primária
19.
Yeungnam University Journal of Medicine ; : 209-216, 1991.
Artigo em Coreano | WPRIM | ID: wpr-89728

RESUMO

Acute megakaryoblastic leukemia is a rare and rapidly fatal disease characterized by proliferation of megakaryocyte series and atypical megakaryocytes in the bone marrow. Acute megakaryoblastic leukemia is suspicious when 1) megakaryocyte in peripheral blood, mixture of large and small mononuclear megakaryoblast in the bone marrow 2) cytoplasmic budding in blast 3) myelofibrosis (dense medullary overgrowth of reticulin fibers) 4) PAS (+), ANAE (+), SBB (−), peroxidase (−) and which is confirmed by platelet peroxidase oxidation on electron microscope or monoclonal antibody. A case of acute megakaryoblastic leukemia was studied morphologically and monoclonal antibody.


Assuntos
Plaquetas , Medula Óssea , Citoplasma , Leucemia Megacarioblástica Aguda , Células Progenitoras de Megacariócitos , Megacariócitos , Naftol AS D Esterase , Peroxidase , Mielofibrose Primária , Reticulina
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