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Human CD34+ Cells (human + cd34+_cell)
Selected AbstractsInteraction of human herpesvirus 6 with human CD34 positive cellsJOURNAL OF MEDICAL VIROLOGY, Issue 3 2003Hiroki Isomura Abstract We reported previously that human herpesvirus 6 (HHV-6) suppresses hematopoietic colony formation of erythroid (BFU-E), granulocyte/macrophage (CFU-GM), and megakaryocyte (CFU-Meg) lineages in vitro. Here we describe the interaction between HHV-6 and human CD34+ cells, which are a major source of hematopoietic progenitor cells. CD34+ cells were immunomagnetically isolated from cord blood mononuclear cells using anti-CD34+ antibodies coated onto either DynabeadsÔ or MACS beads. The CD34+ population selected with Dynabeads showed a broad range of fluorescence. The population selected with MACS beads showed a narrow range of fluorescence. After infection with HHV-6, two transcripts of the immediate early genes were detected with both cell populations. HHV-6 suppressed colony formation of BFU-E, CFU-GM, and CFU-Meg. HHV-6 suppressed cell growth after 3 to 7 days culture in the presence of thrombopoietin (TPO). More differentiated CD34+ cells were more susceptible to the effects of HHV-6. These data indicate that the targets for hematopoietic suppression by HHV-6 are the differentiated cells. J. Med. Virol. 70:444,450, 2003. © 2003 Wiley-Liss, Inc. [source] Probing platelet factor 4 ,-granule targetingJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 12 2004V. Briquet-Laugier Summary., The storage mechanism of endogenous secretory proteins in megakaryocyte ,-granules is poorly understood. We have elected to study the granule storage of platelet factor 4 (PF4), a well-known platelet ,-granule protein. The reporter protein green fluorescent protein (GFP), PF4, or PF4 fused to GFP (PF4-GFP), were transfected in the well-characterized mouse pituitary AtT20 cell line, and in the megakaryocytic leukemic DAMI cell line. These proteins were also transduced using a lentiviral vector, in human CD34+ cells differentiated into megakaryocytes in vitro. Intracellular localization of expressed proteins, and colocalization studies were achieved by laser scanning confocal microscopy and immuno-electronmicroscopy. In preliminary experiments, GFP, a non-secretory protein (no signal peptide), localized in the cytoplasm, while PF4-GFP colocalized with adrenocorticotropin hormone (ACTH)-containing granules in AtT20 cells. In the megakaryocytic DAMI cell line and in human megakaryocytes differentiated in vitro, PF4-GFP localized in ,-granules along with the alpha granular protein von Willebrand factor (VWF). The signal peptide of PF4 was not sufficient to specify ,-granule storage of PF4, since when PF4 signal peptide was fused to GFP (SP4-GFP), GFP was not stored into granules in spite of its efficient translocation to the ER-Golgi constitutive secretory pathway. We conclude that the PF4 storage pathway in ,-granules is not a default pathway, but rather a regular granule storage pathway probably requiring specific sorting mechanisms. In addition PF4-GFP appears as an appropriate probe with which to analyze ,-granule biogenesis and its alterations in the congenital defect gray platelet syndrome. [source] Insights into human CD34+ hematopoietic stem/progenitor cells through a systematically proteomic survey coupled with transcriptomePROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 9 2006Feng Liu Abstract Hematopoietic stem cells are capable of self-renewal and differentiation into different hematopoietic lineages. To gain a comprehensive understanding of hematopoietic stem/progenitor cells, a systematic proteomic survey of human CD34+ cells collected from human umbilical cord blood was performed, in which the proteins were separated by 1- and 2-DE, as well as by nano-LC, and subsequently identified by MS. A total of 370,distinct proteins identified from those cells provided new insights into the potential of the stem/progenitor cells because the nerve, gonad, and eye-associated proteins were reliably identified. Interestingly, the transcripts of 133 (35.9%) identified proteins were not found by the prevalent transcriptome approaches, although several selected transcripts could be detected by RT-PCR. Moreover, the heterogeneity of 33,proteins identified from 2-DE was attributable primarily to post-translational processes rather than to alternative splicing at transcriptional level. Furthermore, the biosyntheses of 15,proteins identified in this study appears not to be completely interrupted in spite of the fact that corresponding antisense RNAs were found in the existing transcriptome data. The integrated proteomic and transcriptomic analyses employed here provided a unique view of the human stem/progenitor cells. [source] In vitro generation of human CD86+ dendritic cells from CD34+ haematopoietic progenitors by PMA and in serum-free mediumCLINICAL & EXPERIMENTAL IMMUNOLOGY, Issue 2 2001G. Ramadan The cytokine requirements to differentiate CD34+ progenitor cells from different origins either cord blood (CB) or peripheral blood (PB) into dendritic cells (DC) are known to be different. In addition to DC, macrophages and neutrophils are generated. On the other hand, phorbol esters such as PMA induce primary human CD34+ bone marrow (BM) progenitor cells to differentiate into functional DC and no other lineages are generated. In addition, FCS is used as culture supplement in most of the protocols described which contains additional foreign antigens potentially skewing the resulting immune response. Therefore, we evaluated the ability to differentiate CB- and PB-CD34+ progenitor cells into DC with PMA and under serum-free conditions. In this study, we delineate the maturation of cultured human blood DC by analysis of expression co-stimulatory molecule B7,2 (CD86). Human mature DC with typical morphology and surface antigen phenotype (CD1a,, CD83+ and CD86+) were obtained from CB- and PB-CD34+ progenitor cells after 1 week of culture in serum-free medium upon stimulation with PMA alone. The same result was obtained from ex vivo -expanded BM-CD34+ cells. CD86+ yield was increased by PMA compared to cytokine cocktails (28·0% ± 7·0 versus 15·3% ± 5·6 for CB and 44·6% ± 7·5 versus 28·1% ± 7·5 for PB, respectively). CD86 was most up-regulated in the presence of the calcium ionophore ionomycin. However, the number of viable cells after differentiation was decreased by PMA plus ionomycin (P < 0·05) or plus TNF-alpha (P > 0·05) as compared with that in PMA alone. We conclude that PMA is a potent activator to differentiate human CD34+ cells into mature DC in serum-free medium. This may be used for in vitro studies of primed or genetically modified DC against infectious and tumour-associated antigens. [source] | ||||||||||