本文選題：骨髓間充質(zhì)干細胞 + 內(nèi)皮細胞　； 參考：《天津醫(yī)科大學(xué)》2006年博士論文

【摘要】：人骨髓間充質(zhì)干細胞向血管內(nèi)皮細胞誘導(dǎo)分化的實驗研究及差異表達基因的微陣列分析 骨髓間充質(zhì)干細胞是多能干細胞,已有實驗證明在體內(nèi)和體外細胞因子作用下,可分化為骨、軟骨和脂肪細胞,在體內(nèi)參與組織更新、損傷的修復(fù)和重建。本研究探討人骨髓間充質(zhì)干細胞誘導(dǎo)分化為內(nèi)皮細胞的可能性。 1.材料和方法 1.1 人骨髓間充質(zhì)干細胞的分離培養(yǎng) 人骨髓標本來源于39歲男性健康志愿者。按骨髓穿刺的常規(guī)方法,抽取骨髓液5ml。肝素抗凝。DMEM+F12培養(yǎng)液稀釋后,用人淋巴細胞分離液Ficoll(比重為1.077)密度梯度離心法,分離出單個核細胞。用DMEM+F12洗滌后,加入DMEM+F12與MCBD的混合培養(yǎng)液(3:2,V/V),并加入10%胎牛血清(FCS)、2ng/ml堿性成纖維細胞生長因子(bFGF)、10ng/ml表皮生長因子(EGF)、10ng/ml胰島素樣生長因子(IGF)和雙抗。24小時后,吸取培養(yǎng)液,連同未貼壁細胞棄去。貼壁細胞繼續(xù)培養(yǎng),每四天更換半量的培養(yǎng)液,12天后細胞融合時,用0.25%胰蛋白酶/0.02%EDTA消化液消化后,細胞傳代,每天觀察生長情況。 1.2.誘導(dǎo)分化 細胞傳至第四代后,在培養(yǎng)液中加量bFGF至10ng/ml,加10ng/mlVEGF培養(yǎng),每四天半量換液,定期觀察細胞生長形態(tài),于第0、14、24天分析細胞的表型和功能。 1.3 流式細胞分析 細胞收集后,用多聚甲醛固定后用流式細胞分析儀,分析CD34、CD45、CD54、CD106、HLA-DR、vWF及KDR的表達情況。
[Abstract]:Experimental study on differentiation of human bone marrow mesenchymal stem cells into vascular endothelial cells and microarray analysis of differentially expressed genesBone marrow mesenchymal stem cells (BMSCs) are pluripotent stem cells. It has been proved that bone cartilage and adipocytes can be differentiated into bone cartilage and adipocytes under the action of cytokines in vivo and in vitro. Bone marrow mesenchymal stem cells participate in tissue renewal repair and reconstruction of injury in vivo.The aim of this study was to investigate the possibility of differentiation of human bone marrow mesenchymal stem cells into endothelial cells.1.Materials and methods1.1 isolation and Culture of Human Bone Marrow Mesenchymal Stem cellsHuman bone marrow specimens were derived from 39-year-old male healthy volunteers.According to the routine method of bone marrow puncture, 5 ml of bone marrow fluid was extracted.After dilution of heparin anticoagulant, DMEM F12 culture medium, human lymphocytes were separated by density gradient centrifugation with Ficoll1 (specific gravity 1.077), and mononuclear cells were isolated.After washing with DMEM F12, adding the mixture of DMEM F12 and MCBD (3: 2 V / V), and adding 10% fetal bovine serum to 2ng / ml of basic fibroblast growth factor (bFGF-1) and 10ng / ml EGF (10ng / ml) and double antibody (.24h),With unattached cells discarded.The adherent cells were cultured continuously. After 12 days of cell fusion, the cells were digested with 0.25% trypsin / 0.02TA digestible solution, and then the cells were subcultured, and the growth was observed every day.1.2.Induced differentiationAfter the cells were transferred to the fourth passage, bFGF was added to the culture medium to 10 ng / ml, and 10ng/mlVEGF was added to the culture medium. The cell growth morphology was observed regularly every four and a half days. The phenotypes and functions of the cells were analyzed on day 1424.1.3 flow cytometry analysisAfter cell collection, the expression of HLA-DRV WF and KDR was analyzed by flow cytometry.
【學(xué)位授予單位】：天津醫(yī)科大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2006
【分類號】：R329.2

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