Y-27632对小鼠骨髓间充质干细胞凋亡的影响

发布时间：2018-12-21 18:28

【摘要】：目的观察ROCK抑制剂Y-27632对体外缺血清培养的小鼠骨髓间充质干细胞凋亡的影响。方法原代分离培养小鼠骨髓单个核细胞,应用流式细胞术对细胞CD29、CD44、CD90、CD34表面标志进行检测。成骨、成脂肪、成软骨分化后分别进行茜素红、油红O及甲苯胺蓝染色。将第3代细胞随机分为3组:对照组、模型组(无血清)、Y-27632组(无血清+10μmol/L Y-27632),分别于培养后24、48、72 h收集细胞进行流式细胞仪TUNEL法观察各组细胞凋亡率,培养后24 h Western blot法检测ROCK1及cleaved caspase 3表达。结果分离的骨髓单个核细胞CD29(+)、CD44(+)、CD90(+)、CD34(-),三系分化后茜素红染色(+),油红O染色(+),甲苯胺蓝染色(+)。Y-27632能明显降低由于缺血清培养导致的BMSCs的凋亡率(P0.01),Y-27632能够明显降低ROCK及cleaved-caspase 3表达(P0.01)。结论分离的骨髓单个核细胞就是骨髓间充质干细胞。Y-27632能够抑制由于缺血清培养导致的小鼠骨髓间充质干细胞的凋亡。
[Abstract]:Objective to observe the effect of Y-27632, a ROCK inhibitor, on apoptosis of bone marrow mesenchymal stem cells (MSCs) cultured in vitro. Methods Murine bone marrow mononuclear cells were isolated and cultured in primary culture. The surface markers of CD29,CD44,CD90,CD34 were detected by flow cytometry. After osteogenesis, fat formation and cartilage differentiation, alizarin red, oil red O and toluidine blue were stained respectively. The third passage cells were randomly divided into three groups: control group, model group (without serum) and Y-27632 group (without serum 10 渭 mol/L Y-27632). The apoptosis rate of each group was observed by flow cytometry (TUNEL) at 24 渭 mol/L 872 h after culture. The expression of ROCK1 and cleaved caspase 3 were detected by 24 h Western blot method. Results CD29 (), CD44 (), CD90 (), CD34 (-) was isolated from bone marrow mononuclear cells and (), was stained by alizarin red (), oil red O after differentiation. Toluidine blue staining with (). Y-27632 significantly decreased the apoptosis rate of BMSCs induced by ischemia culture (P0.01), and Y-27632 significantly decreased the expression of ROCK and cleaved-caspase 3 (P0.01). Conclusion the isolated bone marrow mononuclear cells are bone marrow mesenchymal stem cells and Y-27632 can inhibit the apoptosis of mouse bone marrow mesenchymal stem cells induced by ischemia culture.
【作者单位】：云南省第一人民医院康复医学科;昆明医科大学第一附属医院神经外二科;昆明医科大学第一附属医院骨科;
【基金】：云南省卫计委内设研究机构项目(编号:2011WS0033) 西南医科大学校内基金(编号:15041)
【分类号】：R329.2

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