细胞膜电位对大鼠BMMSCs基本生物学行为影响的研究

The influence of membrane potential on fundamental behaviors of BMMSCs

目的:探索BMMSCs膜电位对其基本生物学行为的影响及机制。方法:采用全骨髓贴壁法分离培养BMMSCs,采用电压敏感性染料DiSBAC分析细胞分化中的电位变化;分别使用乌本苷、二氮嗪诱导细胞发生去极化或超极化,采用划痕试验、CCK8、茜素红染色等手段检测细胞迁移、增殖和成骨分化能力等变化;采用Fluo-8 AM试剂盒检测胞浆钙离子浓度变化。结果:在增殖阶段BMMSCs膜电位变化不显著,而一旦换成成骨诱导液后跨膜电位急剧增加;当诱导BMMSCs发生去极化后其迁移、增殖能力显著增强,而成骨分化能力显著抑制,超极化处理后表现与之相反;发生去极化时胞浆钙离子浓度上调,超极化后钙离子浓度则显著下降。结论:BMMSCs膜电位与细胞行为密切相关。

Objective:To investigate the influence of membrane potential on the fundamental behaviors of bone mesenchymal stem cells(BMMSCs).Methods:The rat primary BMMSCs were isolated and cultured by whole bone marrow plating technique.The vol-tage-sensitive dye DiSBAC was used to monitor the membrane potential alteration during early cell proliferation and differentiation.The cell depolarization and hyperpolarization were induced by ouabain and diazoxide respectively.Afterwards,the cell migration,proliferation and osteogenic differentiation were determined by wound healing test,CCK8 assay and alizarin red staining.The cytoplasm calcium was measured by Fluo-8 AM kit.Results:The membrane potential did not change apparently during the proliferation stage,whereas it dramatically increased once the osteogenic medium was applied.The cell migration and proliferation were significantly enhanced while the osteogenic differentiation was inhibited after depolarization.On the contrary,the hyperpolarized BMMSCs showed the opposite behaviors.The cytoplasm calcium was also increased or decreased significantly after depolarization or hyperpolarization.Conclusion:The membrane potential of BMMSCs is closely related to cell behaviors.