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| Y-27632 对小鼠骨髓间充质干细胞成骨分化的影响 |
| Effects of Y-27632 on mouse osteogenesis by bone marrow mesenchymal stem cells |
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| DOI： |
| 中文关键词: 骨髓间充质干细胞 ＲhoA/ＲOCK Y-27632 成骨分化 |
| 英文关键词:Bone marrow mesenchymal stem cells ＲhoA/ＲOCK Y-27632 Osteogenesis |
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| 中文摘要: |
| 目的 探讨 ＲhoA/ＲOCK 通路特异性阻断剂 Y-27632 对小鼠骨髓间充质干细胞(bone marrow mesenchymal stem cells，BMSCs)增殖及成骨分化的影响。方法 自小鼠股骨、胫骨骨髓腔分离培养小鼠骨髓单个核细胞，流式细胞仪细胞表面标记检测及细胞成骨、成脂、成软骨三系分化鉴定小鼠骨髓单个核细胞。将第 3 代小鼠骨髓间充质干细胞随机分为 2 组，单纯成骨诱导液对照组、Y-27632 干预组。分别于细胞培养后 1d、2d、3d、4d 收集细胞，噻唑蓝(Methylthiazolyldiphenyl-tetrazolium bromide，MTT)比色法观察 BMSCs 增殖能力;成骨诱导后 7 d、14 d 收集细胞，可见光比色法检测碱性磷酸酶(ALP)变化;成骨诱导后 24h 收集细胞，western blot 法检测 Ｒho 相关卷曲螺旋形成蛋白激酶 1(Ｒho associated coiled-coil-containing protein kinases1，ＲOCK1)的表达。结果 流式细胞术细胞表面标志检测及成骨、成脂、成软骨三系分化鉴定均证实我们所获细胞为 BMSCs。与对照组相比，Y-27632 能促进 BMSCs 增殖，差异具有统计学意义(P ＜ 0. 05);Y-27632 能降低 ALP 表达，差异具有统计学意义(P ＜0. 05);Y27632 能明显阻断 ＲOCK1 的表达，差异具有统计学意义(P ＜0. 05)。结论 Y-27632 阻断 ＲhoA/ＲOCK 信号通路可以促进 BMSCs 增殖，抑制 BMSCs 成骨分化。 |
| 英文摘要: |
| Objective To investigate the effect of ＲhoA/ＲOCK blocker Y-27632on the proliferation and osteogenesis of mouse marrow mesenchymal stem cells (BMSCs)． Methods Bone marrow mononuclear cells were obtained from the bone marrow cavity of the femur and tibia and identified with flow cytometry for detecting cell surface markers and osteogenic，adipogenic，and chondrogenic differentiation． BMSCs of the passage 3 were randomly divided into 2 groups，control group with osteogenesis induction media only and osteogenesis induction media + Y-27632 group． Proliferation of BMSCs was determined using methyl
thiazolyl tetrazolium (MTT) assay at 1d，2d，3d，and 4d． ALP expression was determined with visible light photocolorimetric method at 7d and 14d after osteogenic induction． ＲOCK1 protein expression was detected with Western blotting at 24h after osteogenic induction． Ｒesults The bone marrow mononuclear cells were identified as BMSCs with flow cytometry and tri-lineage differentiation． Compared with control group，MTT assay showed Y-27632 obviously promoted BMSCs proliferation (P ＜0. 05)． Y-27632 also inhibited ALP expression (P ＜ 0. 05) and the expression of ＲOCK1 (P ＜ 0. 05)． Conclusion Y-27632 promotes proliferation but inhibits ostogenesis of BMSCs by blocking ＲhoA/ＲOCK signaling pathway． |
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