应用EBM-2培养犬脐血间充质干细胞的实验研究

目的应用EBM-2培养犬脐血间充质干细胞(MSC),为MSC的研究提供新的细胞培养方法。方法取犬的脐血,分为四组进行细胞培养。第1组用EBM-2培养;第2组用添加有EGM-2MV的EBM-2在6孔板上培养。第3组用添加有EGM-2MV的EBM-2在纤连蛋白包被的6孔板上培养。第4组用添加有EGM-2MV的EBM-2在25cm2细胞培养瓶中培养。应用免疫组化法检测细胞抗原CD11a、CD11b、CD34、CD29及CD71的表达。结果各组均有纤维母细胞样细胞培养出。第1组细胞形态不良,增殖缓慢;第2组细胞增殖旺盛;第3组细胞克隆不稳定,容易老化;同时出现另一种细胞克隆。第4组细胞培养的结果与第3组相似。免疫组化法检测抗原显示:CD11a(-)、CD11b(-)、CD34(-)、CD29( )、CD71( )。结论在不用纤连蛋白包被的6孔板上,用添加有EGM-2MV的EBM-2细胞培养液可以培养出生长良好、增殖旺盛的MSC。

Culturing Mesenchymal Stem Cells in Cord Blood of Dogs with EBM-2

Objective To culture mesenchymal stem cells(MSC) in cord blood of dogs with EBM-2, providing a new method to culture cells for research in MSC. Methods Collect cord blood from dogs, and divide them into four groups. Culture the cells with EBM-2 in the first group; with EBM-2 adding EGM-2 MV in the 6-well cell culture cluster in the second group; with EBM-2 adding EGM-2 MV in the 6-well cell culture cluster coating with fibronectin in the third group; with EBM-2 adding EGM-2 MV in the 25 cm~ 2 culturing flask in the fourth group. Detect antigen CD11a, CD11b, CD34, CD29 and CD71 with immunohistochemistry method. Results Fibroblast-like cells could grow up in each group. The cells grew badly in morphology and proliferated slowly in the first group, while they proli- ferated rapidly in the second group. The cell clones were instable and inclined to aging in the third group, with a new cell clone being found. The cells in the fourth group were similar to those in the third group. Surface antigens detected with immunohistochemistry method showed CD11a(-), CD11b(-), CD34(-), CD29( ), CD71( ). Conclusion The MSC could be cultured with EBM-2 adding EGM-2 MV in the 6-well cell culture cluster.