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Expression of Proliferating Cell Nuclear Antigen in Cord Blood CD34~+ Hematopoietic Stem/Progentior Cells and Its Significance

Author: ZhangDongQing
Tutor: LiuYuanSheng
School: Shantou University
Course: Internal Medicine
Keywords: Umbilical cord blood CD34 ~ cells Proliferating cell nuclear antigen Cell Cycle
CLC: R329.2
Type: Master's thesis
Year: 2004
Downloads: 26
Quote: 1
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Abstract


Human umbilical cord blood is rich in hematopoietic stem / progenitor cells, is an alternative to bone marrow and peripheral blood hematopoietic cell source has broad application prospects. In recent years, umbilical cord blood transplantation has been widely used in the treatment of childhood leukemia and malignant tumors. However, due to the limited amount of single cord blood collection, which contains hematopoietic stem / progenitor cell number is difficult to meet the needs of older children and adult patients. Hematopoietic stem cells in vitro amplification technology is expected to overcome this obstacle. Cord blood hematopoietic stem / progenitor cells has a strong proliferation and differentiation, proliferation in the hematopoietic cytokine stimulation, but in the amplification of hematopoietic stem / progenitor cells to differentiation and maturation, thus affecting their self-renewal capacity and pluripotent reserved. So how can both amplified to retain as much as possible the number of hematopoietic cells of hematopoietic cell performance is the focus of current research. In recent years, cytokines programs large number of studies have not yet established a precise and effective way, and very little for the study of cell cycle regulation, but as the body's cells, hematopoietic stem cell proliferation and differentiation is still the fundamental evolution of the cell cycle, hematopoietic stem cell proliferation and differentiation process not only controlled by the cytokine, is closely related to cell cycle regulators. PCNA (proliferating cell nuclear antigen, PCNA) is an important cell cycle regulatory proteins, can induce cells into S phase by the Gl. In this study, flow cytometry PCNA expression levels of cord blood CD34 - cells in the different culture time and under different culture conditions and cell cycle were determined to reveal the biological significance of PCNA in the regulation of hematopoiesis for further study of how to control the proliferation and differentiation of hematopoietic stem cells through the regulation of PCNA provide some basis for further study. 1. Cord blood CD34 ~ cells in vitro amplification of PCNA expression profiles were analyzed using Mini MACS (magnetic activated cell sorting) isolated CD34 ~ cells, Shantou University Medical Journal cultured in IMDM (Iseove containing different cytokine combinations, 5 modified Dulbeeeo, smedium, IMDM) culture system. Experimental groups: (1) a group of cytokines that blank control group; ② stem cell factor (stem eell faetor, SCF) interleukin 6 (IL a 6) interleukin-3 (xL a 3) group; ③ SCF xL a 6 IL a 3 small plate of IfII EPO (thrombopoietin, TPO) group; ④ Flt3 a ligand Flt3 a ligand (FL) the TPO SCF, IL a 6 IL-3 group. In 3 days, 5 days, 7 days, cells were collected and PCNA expression levels by flow cytometry. It was found that freshly isolated cord blood of CD3 Kuang cells in low expression of PCNA-positive rate (H .6 ± 5.2)%, short-term culture in vitro, cell-free combination of factors PCNA expression levels gradually decreased, and in the presence of each the cytokine combinations culture system, PCNA expression increased and to FL TPO SCF IL a 6 IL-3 combined effect of the most significant. Cord blood CD34 cells in vitro amplification cell cycle. Using Mini MACS separation of CD34 cells, in containing different cell factor combinations (① SEF IL a 6 IL-3 group; ② SCF IL a 6 IL a 3 Tpo group; ③ FL TPO SCF IL a 6 IL-3 group) the training system training 7 days, 0 days, 3 days, 7 days, cells were collected, propidium iodide (propidium iodide, PI) staining using flow cytometry to detect cell cycle. It was found that cord blood CD34 cells freshly isolated from 95.1% in G 0 / Gl period, only 4.9% in S phase, and cultured for 3 days, cell proliferation and differentiation into the proliferation of different cytokine combinations, 7 days S / GZ / M phase was significantly increased. The study suggests that the CD34 ten hematopoietic stem / progenitor cells in the hematopoietic growth factor role of PCNA protein expression was significantly higher, thus contributing to the proliferation of cells from the quiescent proliferation.

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