中文摘要：

目的通过骨髓间充质干细胞（bone marrow mesenchymal stem cells,BMSCs）与膀胱平滑肌细胞（bladder smooth muscle cells,BSMCs）接触共培养探讨组蛋白乙酰化对BMSCs分化能力的影响。方法分别采用密度梯度离心法和胶原酶消化法培养大鼠BMSCs及BSMCs至第3代,并对BMSCs进行了鉴定;以单独培养的BMSCs作为对照组,接触共培养的BMSCs为实验组;运用免疫荧光化学染色法分别对2组BMSCs中的平滑肌标志性蛋白平滑肌α肌动蛋白（α-smooth muscle actin,α-SMA）、平滑肌肌球蛋白重链（smooth muscle-myosin heavy chain,SM-MHC）、Calponin进行检测,Western blot检测α-SMA蛋白、Calponin蛋白在BMSCs中的表达,并用微球菌核酸酶（micrococcal nuclease,MNase）对平滑肌3个标志性基因的启动子区进行了染色质开放性的检测,同时运用了ChIP技术分析了α-SMA、SM-MHC启动子区组蛋白乙酰化水平对BMSCs向平滑肌分化的影响。结果大鼠原代BMSCs流式检测显示CD29表达阳性,阳性率为99.8%,CD31、CD45表达阴性,阳性率为6.30%、1.04%;免疫荧光显示平滑肌3种标志性蛋白的表达均随时间的延长而增加;Western blot结果显示随着共培养天数的增加α-SMA蛋白、Calponin蛋白在实验组中的表达相比对照组有明显的增加;MNase显示实验组的BMSCs启动子区染色质对核酸酶的敏感性明显高于对照组,ChIP显示实验组中BMSCs平滑肌标志性基因α-SMA、SM-MHC、Calponin启动子区H4乙酰化水平较未分化前明显增加（P〈0.05）。结论组蛋白乙酰化程度的增加促进了BMSCs向BSMCs分化。

英文摘要：

Objective To investigate the effect of histone acetylation on the differentiation capacity of bone marrow mesenchymal stem cells（BMSCs） directly co-cultured with bladder smooth muscle cells（BSMCs）.Methods Density gradient centrifugation and collagenase digestion were used to train the rat BMSCs and BSMCs to the 3rd passage.After the cells were identified,BMSCs co-cultured with BSMCs were regarded as experimental cells,while the cells cultured alone as the control cells.Immunofluorescence assay was used to detect the expression of BMSCs landmark proteins,α-smooth muscle actin（α-SMA）,smooth muscle myosin heavy chain（SM-MHC）,and calponin in the 2 kinds of BMSCs.Western blotting was adopted to detect the protein levels of α-SMA and calponin.Micrococcal nuclease（MNase） was used to detect the promoter regions of the 3 marker genes for open chromatin,while ChIP technical analysis was used to detect the histone acetylation levels in the promoter regions of α-SMA and SM-MHC,and the effects of the levels on BMSCs to smooth muscle differentiation were determined.Results Our obtained primary rat BMSCs were positive to CD29（99.8%）,negative to CD31 and CD45（positive rate of 6.30% and 1.04%）.Immunofluorescence assay showed the expression of the 3 BMSCs landmark proteins was increased with time elapsing.Western blotting showed that the protein levels of α-SMA and calponin were significantly higher in experimental cells than in control cells with the elapse of co-culture time.MNase assay displayed that experimental BMSCs had significantly higher chromatin nuclease sensitivity in promoter regions than the control cells.ChIP showed the experimental BMSCs had significantly higher H4 acetylation level in the promoter regiong of α-SMA,SM-MHC and calponin when compared with the undifferentiated cells（P0.05）.Conclusion The enhanced histone acetylation promotes BMSCs differentiation to BSMCs.