小鼠EPCs和RAW264.7细胞株体外联合培养体系的建立.docVIP

PAGE PAGE 1 小鼠EPCs和RAW264.7细胞株体外联合培养体系的建立 付生龙，庞 浩，吴雪晖，许建中 (400038 重庆，第三军医大学西南医院骨科，全军矫形外科中心) [摘要] 目的 体外建立小鼠血管内皮祖细胞（endothelial progenitor cells, EPCs）和破骨细胞前体细胞RAW264.7细胞株共培养体系。方法 EPCs和RAW264.7细胞株共培养为实验组，加了等量细胞培养液但没有加入EPCs细胞作为对照组。利用分离式培养小室进行细胞联合培养，Cell Counting Kit-8（CCK-8）试剂盒检测细胞的增殖，通过检测吸光度绘制共培养过程中RAW264.7细胞生长曲线；逆转录 PCR检测共培养1、3、5和7天后RAW264.7细胞VEGFR-2和CXCR4 mRNA水平的表达；TRAP染色检测破骨细胞活性。结果 和EPCs细胞的联合培养能加快RAW264.7细胞的增殖速率，和RAW264.7单独生长比较，共培养3、5、7 d后，两组的吸光度有显著性的差异（P0.01）；共培养促进RAW264.7细胞分化为破骨细胞。结论 在体外联合培养体系中，EPCs具有促进宿主破骨细胞前体细胞增殖与分化的作用。 [关键词] 小鼠EPCs；RAW264.7；联合培养 [中图法分类号] [文献标志码] A ［基金项目］ 国家自然科学基金(0081071465) ［通信作者］ 吴雪晖，电话:（023 E-mail: HYPERLINK mailto:fatbear7217@ fatbear7217@ 许建中，电话:（023 E-mail: HYPERLINK mailto:xjzslw@163.com xjzslw@163.com Establishment of co-culture system of mouse endothelial progenitor cells and RAW264.7 cells Fu Shenglong, Pang Hao, Wu Xuehui, Xu Jianzhong (Center of Orthopedics, Department of Orthopedics, Southwest Hospital, Third Military Medical University, Chongqing, 400038, China) Abstract Objective：To establish mouse endothelial progenitor cells (EPCs) and RAW264.7 co-culture system. Methods: endothelial progenitor cells (EPCs) and RAW264.7 cells co-cultured for the experimental group, and Add the equivalent cells nutrient solution but not join EPCs cells as control group.Transwell was used to co-culture EPCs and RAW264.7. Cell Counting Kit-8 was used to measure cell proliferation and absorption values were detected to depicter cellular growth curve. Reverse transcript PCR was applied to detect VEGFR-2 and CXCR4 expression in mRNA level.TRAP staining was used to detect osteoclast activity. Results: Co-culture of mouse endothelial progenitor cells (EPCs) and RAW264.7 cells could facilitate RAW264.7 cells growth. In comparison with single growth of RAW264.7 cells, co-culturing for 3 days significantly increased cell growth. Co-culture also increased vWF and VEGFR-2 expression and increased osteoclas