单宁酸对高糖下大鼠肾小球系膜细胞增殖及细胞周期的影响

Effects of gallotannin on proliferation and cell cycle of rat glomerular mesangial cells under condition of high glucose

目的:探讨单宁酸(GLTN)对高糖诱导的大鼠肾小球系膜细胞(GMC)增殖的抑制作用及对细胞周期的影响,阐明GLTN对糖尿病肾病(DN)发生发展的延缓作用。方法:实验细胞分成正常糖对照组(D-葡萄糖5.5mmol·L^(-1),NC组)、高糖组(D-葡萄糖30mmol·L^(-1),HC组)、高糖+5mmol·L^(-1) 3-氨甲苯甲酰胺(3-AB,AB组)、高糖+20μmol·L^(-1) GLTN(G20组)和高糖+40μmol·L^(-1) GLTN(G40组)。MTT法观察不同时间点(4、8、24、48和72h)各组GMC的增殖情况,流式细胞术观察细胞周期,Western blotting法检测转化生长因子β1(TGF-β1)和结缔组织生长因子(CTGF)的蛋白表达情况。结果:与NC组比较,高糖组GMC增殖水平升高(P<0.01),于48h时达高峰;S期细胞百分比明显升高(P<0.01)。与HC组比较,各给药组GMC增殖水平均明显下降(P<0.01),S期细胞百分比降低(P<0.01)。与NC组比较,各给药组S期细胞百分比明显升高(P<0.05或P<0.01),TGF-β1和CTGF蛋白表达水平升高(P<0.05或P<0.01),但与HC组比较明显下降(P<0.01)。结论:单宁酸可通过阻滞细胞周期,下调TGF-β1和CTGF的表达,抑制高糖诱导的大鼠GMC的增殖,从而对DN的发生发展起到一定的延缓作用。

Objective： To explore the inhibitory effect of gallotannin （GLTN） on the proliferation of rat glomerular mesangial cells （GMC）induced by high sugar stimulation and the influence in the cell cycle of the rats, and to clarify the prevention effect of GLTN in the occurrence and development of diabetic nephropathy （DN）. Methods：The experimental cells were divided into normal control group （D-glucose 5.5 mmol·L-1 ,NC group）, high glucose group （D-glucose 30 mmol· L-1 ,HC group）,high glucose ＋ 5 mmol· L-1 3 - AB group （AB group）,high glucose ＋ 20 μmol·L-1 GLTN group （G20 group）,high glucose ＋ 40 μmol· L-1 GLTN group （G40 group）.The proliferation of GMC in different groups at different time points （4,8,24,48 and 72 h）was＆amp;nbsp;observed by MTT assay.The changes of cell cycle of GCM under different culture conditions were examined by flow cytometry,and the expression levels of TGF-β1 and CTGF were detected by Western blotting method. Results：Compared with NC group,the proliferation levels of GMC in HC group were increased （P 〈0.01）,and reached the peak at 48 h ;the percentage of S phase cells was increased （P 〈0.01）.Compared with HC group,the proliferation levels of GMC in 3-AB group and GLTN group were significantly decreased （P 〈 0.01 ）,and the percentages of S phase cells were decreased （P 〈0.01）.Compared with NC group,the protein expression levels of TGF-β1 and CTGF in each drug group were increased （P 〈 0.05 or P 〈 0.01）,but they were significantly lower than those in HC group （P 〈 0.01）.Conclusion：GLTN can inhibit the proliferation of GMC under high sugar stimulation through arresting the cell cycle and down-regulating the expressions of TGF-β1 and CTGF and delay the occurrence and development of DN.