马兜铃酸A致大鼠肾病的肾毒性作用机理研究

以大鼠为靶动物进行亚慢性毒性试验,探究马兜铃酸A的肾毒性作用机理。将56只健康雄性Wistar大鼠适应性饲养1周,随机分为马兜铃酸A高剂量组(40 mg/kg BW)、马兜铃酸A中剂量组(8 mg/kg BW)、马兜铃酸A低剂量组(4 mg/kg BW)和空白组4组,试验持续28 d,后进行14 d恢复性试验。通过监测体质量,计算肾脏指数,检测血尿素氮(BUN)、尿酸(UA)和肌酐(SCr)的含量,检测丙二醛(MDA)含量、超氧化物歧酶(SOD)活性、谷胱甘肽过氧化物酶(GSH-Px)活性和总抗氧化能力(T-AOC),观察肾脏的组织病理学变化,采用实时荧光定量PCR检测相关基因mRNA的表达情况,来综合评价马兜铃酸A的肾毒性作用。结果分析显示,从第16天开始马兜铃酸A高剂量组大鼠体质量显著低于空白组(P<0.05);肾脏指数各组间差异不显著;试验第28天,高剂量组BUN、UA、CA含量均显著高于空白组(P<0.05),中剂量组BUN、UA含量均显著高于空白组(P<0.05),低剂量组UA含量显著高于空白组(P<0.05),试验第42天,高剂量组的BUN、UA、CA含量仍然均显著高于空白组(P<0.05),中剂量组CA含量显著高于空白组(P<0.05);试验第28天,高剂量组MDA含量显著高于空白组(P<0.05),SOD、GSH-Px活性均显著低于空白组(P<0.05),中剂量组和低剂量组GSH-Px活性显著低于空白组(P<0.05),试验第42天,高剂量组MDA含量仍然显著高于空白组(P<0.05),SOD、GSH-Px活性和T-AOC均显著低于空白组(P<0.05),中剂量组SOD、GSH-Px活性和T-AOC均显著低于空白组(P<0.05),低剂量组MDA含量显著高于空白组(P<0.05),GSH-Px活性显著低于空白组(P<0.05);肾脏组织病理学观察发与组织和细胞层面的损伤程度大体呈剂量相关性;试验第28天和第42天,高剂量组和中剂量组TGF-β1、smad3、collagen1、collagen3的mRNA表达量显著高于空白组(P<0.05),低剂量组TGF-β1、smad3的mRNA表达量显著高于空白组(P<0.05)。说明马兜铃酸A对大鼠肾脏有毒性作用,肾功能被损害,引起一定程度的肾纤维化,这种损害作用在一定时间内不能自行恢复。马兜铃酸A的毒性作用可能与氧化应激与TGF-β1/smad通路作用有关。

Study on the mechanism of nephrotoxicity of aristolochic acid A in rat nephropathy

A subchronic toxicity test was conducted with rats as the target animal to explore the mechanism of nephrotoxicity of aristolochic acid A.Fifty-six healthy male Wistar rats were adaptively reared for one week and randomly divided into aristolochic acid A high-dose group(40 mg·kg^-1 BW),aristolochic acid A medium-dose group(8 mg·kg^-1 BW),and equine Toulic acid A low-dose group(4 mg·kg^-1 BW) and blank group 4 groups,the test lasted 28 days,and then a 14-day recovery test.By monitoring body weight,calculating kidney index,detecting blood urea nitrogen(BUN),uric acid(UA)and creatinine(CA) content,detecting malondialdehyde(MDA) content,superoxide dismutase(SOD) activity,glutathione Glycine peroxidase(GSH-Px) activity and total antioxidant capacity(T-AOC),observe the histopathological changes of kidney tissue,and use real-time fluorescent quantitative PCR to detect the expression of related genes mRNA to comprehensively evaluate aristolochia nephrotoxic effects of cervic acid A.The analysis of the results showed that the body weight of rats in the high-dose aristolochic acid A group was significantly lower than that in the blank group from day 16(P<0.05);there was no significant difference in renal index between the groups;on the 28 th day,the high-dose group BUN The contents of UA,CA were significantly higher than those in the blank group(P<0.05),the contents of BUN and UA in the medium-dose group were significantly higher than those in the blank group(P<0.05),and the UA content in the low-dose group was significantly higher than that in the blank group(P<0.05),on the 42 nd day of the test,the contents of BUN,UA,and CA in the highdose group were still significantly higher than those in the blank group(P<0.05),and the CA content in the middle-dose group was significantly higher than that in the blank group(P<0.05);on the 28 th day of the test the MDA content of the high-dose group was significantly higher than that of the blank group(P<0.05),the activities of SOD and GSH-Px were significantly lower than those of the blank group(P<0.05),and the GSH-Px activity of the medium-dose group and low-dose group was significantly lower than that of the blank group Group(P<0.05),on the 42 nd day of the test,the MDA content of the high-dose group was still significantly higher than that of the blank group(P<0.05),SOD,GSH-Px activity and T-AOC were significantly lower than the blank group(P<0.05),SOD,GSH-Px activity and T-AOC of the middle dose group were significantly lower than the blank group(P<0.05),the MDA content of the low dose group was significantly higher than that of the blank group(P<0.05),and the GSH-Px activity was significantly lower than the blank group(P<0.05);the pathological histology of kidney tissues showed that the degree of damage at the tissue and cell level was generally dose-dependent;on the28 th and 42 nd day of the test,the mRNA expression levels of TGF-β1,smad3,collagen1,and collagen3 in the high-dose and medium-dose groups were significantly higher than those in the blank group(P<0.05),and the mRNA expression of TGF-β1 and smad3 in the low-dose group was significantly higher than that of the blank group(P<0.05).It showed that aristolochic acid A has a toxic effect on the kidneys of rats,and the renal function is damaged,causing a certain degree of renal fibrosis.This damage can not be restored by itself within a certain period of time.The toxic effect of aristolochic acid A may be related to the effect of oxidative stress and TGF-β1/smad pathway.