葛根素对肾脏缺血再灌注大鼠肾脏氧化应激损伤的影响及其机制研究

摘要

目的：研究葛根素对肾脏缺血再灌注大鼠肾脏氧化应激损伤的影响，并探讨其可能的作用机制。方法将120只SD大鼠随机分为假手术组、模型组、葛根素25 mg／kg组、葛根素50 mg／kg组、葛根素100 mg／kg组、银杏叶提取物（100 mg／kg）组；除假手术组外，其余各组均通过夹闭双侧肾蒂血管45 min后松夹恢复血流再灌注的方法制备肾缺血再灌注模型；再灌注后各给药组立即腹腔注射给药，每天1次，连续2周，假手术组和模型组给予等体积生理盐水。末次给药后，测定血清尿素氮（ BUN）、肌酐（ SCr）、尿酸（ UA）及总抗氧化能力（ T－AOC）水平；通过苏木精－尹红（ HE）染色观察肾脏组织病理形态学变化；通过原位末端标记法（ TUNEL）观察肾小球细胞凋亡状况并计算凋亡指数；检测肾脏组织匀浆中超氧化物歧化酶（ SOD）、谷胱甘肽过氧化物酶（ GSH－Px）、过氧化氢酶（ CAT）活性和丙二醛（ MDA）含量；通过RT－PCR法测定肾脏组织中bax和bcl－2 mRNA表达量，并计算bax／bcl－2比值。结果葛根素50，100 mg／kg组血清BUN、SCr、UA含量均明显低于模型组（P均＜0．05）；各给药组大鼠肾脏组织病理形态学变化和肾小球细胞凋亡状况呈不同程度改善，以葛根素100 mg／kg组效果最为显著；葛根素50，100 mg／kg组大鼠肾小球细胞凋亡指数及肾脏组织中bax mRNA表达量、MDA含量及bax／bcl－2比值均明显低于模型组（P均＜0．05），肾脏组织中SOD、CAT活性及bcl－2 mRNA表达量均明显高于模型组（P均＜0．05），其中葛根素100 mg／kg组血清T－AOC水平和肾脏组织中GSH－Px活性均显著高于模型组（ P均＜0．05）。结论葛根素对肾脏缺血再灌注大鼠肾脏氧化应激和细胞凋亡具有抑制作用；作用机制可能与其能够改善肾脏组织抗氧化酶活性、促进抗凋亡基因bcl－2表达、抑制促凋亡基因bax表达并降低bax／bcl－2比值有关。%Objective It is to observe the effect of puerarin on oxidative stress induced by renal ischemic-reperfusion in rats, and investigate its mechanism.Methods One hundred and twenty SD rats were randomly divided into six groups: sham operation group, model group, puerarin 25, 50, 100 mg/kg group and ginkgo biloba extract (EGb) 100 mg/kg group;except sham operation group, the rats in other groups were all made renal ischemic-reperfusion rat models by gripping bilateral renal pedicle vascular for 45min , the drugs were given immediately after reperfusion, once a day for 2 weeks.Two weeks later, the content of BUN, SCr, UA and the level of T-AOC in serum were detected;the renal tissue histopathological changes was ob-served by HE staining;the apoptosis of renal cells was observed by TUNEL staining, and the apoptosis index ( AI) was calcu-lated;the activity of SOD, GSH-Px, CAT and the content of MDA in the renal tissue were measured;the expression level of Bax, bcl-2 mRNA were detected by RT-PCR, and the bax/bcl -2 was calculated.Results Compared with the model group, the content of BUN, SCr, UA in serum of puerarin 50, 100 mg/kg group were significantly decreased ( all P<0.05);the histopathological changes and the apoptosis of the renal tissue in puerarin 25, 50 100 mg/kg group and EGB 100 mg/kg group were significantly improved, the effect of puerarin 100 mg/kg group was the most significant; in the puerarin 50, 100 mg/kg group, the AI and bax mRNA expression, MDA content and bax/bcl-2 ratio in renal tissue were significantly lower than those in the model group (all P<0.05), and the expression levels of SOD, CAT and Bcl-2 mRNA in renal tissues were significantly higher than those in the model group ( all P<0.05) , the serum T-AOC level and GSH-Px activity in renal tis-sue of puerarin 100 mg/kg group were significantly higher than those in the model group (all P<0.05).Conclusion Puerarin has inhibitive effects on renal oxidative stress and apoptosis, the mechanism might be related to improving the activity of an-tioxidant enzymes in renal tissue, promoting the expression of anti-apoptotic gene Bcl-2, inhibiting the expression of pro-apoptotic gene Bax and reducing the ratio of bax/bcl-2.