黄芩苷对高糖环境下肾小球系膜细胞缝隙连接蛋白43表达的影响

Effect of Baicalin on Expression of Connexin 43 in Glomerular Mesangial Cells under High-glucose Condition

目的研究黄芩苷对高糖环境下肾小球系膜细胞(glomerular mesangial cell,GMC)中缝隙连接蛋白43(connexin 43,Cx43)表达的影响。方法将体外培养的大鼠GMC分为6组:低糖组,高糖组,黄芩苷低、中、高剂量组,蛋白激酶C(protein kinase C,PKC)抑制剂组。培养72h后,收集细胞并提取细胞总蛋白,Western-blot法检测黄芩苷对高糖环境下GMC中PKC和Cx43表达的影响,用划痕标记染料示踪技术检测黄芩苷对GMC缝隙链接通讯功能(gap junction intercellular communication,GJIC)的影响。结果 Western-blot结果显示,与低糖组比较,高糖组中PKC蛋白表达量显著增加(P<0.01),Cx43蛋白表达量显著减少(P<0.01);黄芩苷呈剂量依赖性降低PKC蛋白表达水平,升高Cx43蛋白表达水平。划痕标记染料示踪实验结果显示:与低糖组比较,高糖组GMC的GJIC功能缺乏,未向邻近细胞传输;与高糖组比较,PKC抑制剂组罗氏黄荧光染料向划痕两侧邻近细胞传输的层数较黄芩苷各剂量组明显增多,黄芩苷各剂量组呈剂量依赖性地增加传递的层数。结论黄芩苷可剂量依赖性地抑制高糖环境下GMC中PKC的异常活化,从而上调Cx43表达,恢复GJIC的功能。

Objective To investigate the effect of baicalin on the expression of connexin 43 （Cx43） in glo- merular mesangial cells （GMCs） under high-glucose condition. Methods The rat GMCs cultured in vitro were divided into six groups： low-glucose group, high-glucose group, low-, middle-, and high-dose baica- lin groups, and protein kinase C （PKC） inhibitor group. After 72 hours of culture, cells were collected,and total protein was extracted. The effect of baicalin on the expression of PKC and Cx43 in GMCs under high-glucose condition was evaluated by Western blot. The effect of baicalin on the gap junction intercellu- lar communication （GJIC） of GMCs was evaluated by scrape-loading and dye transfer. Results The Western blot showed that compared with the low-glucose group, the high-glucose group had significantly increased protein expression of PKC and decreased protein expression of Cx43（P^0.01）~ compared with the high-glucose group, the baicalin groups had decreased protein expression of PKC and increased protein expression of Cx43, with a dose dependence. The scrape-loading and dye transfer experiment showed that compared with the low-glucose group, the high-glucose group had a lack of GJIC, without dye transfer in- to contiguous cells~ compared with the high-glucose group, the PKC inhibitor group had significantly more layers of fluorescent dye Lucifer yellow transfer into contiguous cells, and the baicalin groups had the number of transfer layers increased in a dose-dependent manner. Conclusion Baicalin can inhibit the abnormal activation of GMC PKC under high-glucose condition in a dose-dependent manner, thus upregu- lating the expression of Cx43 and restoring the function of GJIC.