细胞间接触诱导骨髓间充质干细胞向平滑肌细胞分化的实验研究.docVIP

细胞间接触诱导骨髓间充质干细胞向平滑肌细胞分化的实验研究 刘晓芳　王延洲　徐惠成　梁志清 (400038 重庆 第三军医大学第一附属医院) 【摘要】 目的　通过两种细胞的共培养探讨细胞间接触对间充质干细胞（Mesenchymal Stem Cells, MSCs）增加英文全称分化的影响。方法 分别采用密度梯度离心法和胶原酶消化法分离培养了骨髓间充质干细胞（Bone Marrow Mesenchymal Stem Cells ，BMSCs）增加中英文全称和膀胱平滑肌细胞。并对其进行了鉴定。在此基础上以单独培养的BMSCs和平滑肌细胞分别作为阴性和阳性对照组，将获得的BMSCs和平滑肌细胞在体外通过接触与非接触共培养作为实验组,;对诱导后的BMSCs是否向平滑肌分化通过免疫荧光和western进行鉴定. 结果对BMSCs进行流式细胞术检测结果显示CD90、 CD44表达阳性率分别为99.78%,60.27%，CD45表达阴性，阳性率为0.21%。在BMSCs与平滑肌细胞共同培养的实验中，western结果显示：BMSCs本身表达少量calponin蛋白，接触组calponin的表达随培养时间的延长逐渐增多，非接触组无明显变化；免疫荧光显示：接触组在共培养第8天部分细胞抗calponin阳性，非接触组与诱导前无明显变化。 结论 初步认为BMSCs与平滑肌细胞接触共同培养能诱导BMSCs向平滑肌分化。 增加英文全称 增加中英文全称 【关键词】 细胞间接触　微环境　骨髓间充质干细胞　平滑肌细胞 The Study of Cell-Cell Contact Inducing Mesenchymal Stem Cell Differentiation to Smooth Muscle Cell Liu Xiaofang，Wang Yangzhou，Xu Huicheng， Liang Zhiqing (The first affiliated hospital of The Third Military University, Chongqing 400038, China) 【Abstract】 Objective To establish the co-culture system between mesenchymal stem cells （MSCs） and smooth muscle cells in vitro, and study the influence of smooth muscle cells on the differentiation of bone marrow mesenchymal stem cells (BMSCs ) into smooth muscle cells in the co-culture system. Methord We used the method of density gradient centrifugation to separate and culture BMSCs and method of Collagenase digestion to separate and culture blander smooth muscle cells, then assayed cells respectively. Afer that, We designed four group ,BMSCs alone was the positive control grope , smooth muscle cells alone was the negative control grope, BMSCs and smooth muscle cells co-culture with contact or non-contact are the two experimental group . The expression of α- SMA, calponin were detected by immunofluorescence and observed with fluorescence microscope in contact co-culture grope. The method of Western-Blot was applied to determine protein expression of calponin. Results Cells with contact co-culture induction grope were observed for the expression of calponin increase gradually. Conc