高胆红素血症诱导氧化应激引起的大鼠肾小球损伤  

Hyperbilirubinemia induced oxidative stress resulting in glomerular injury in rats

作　　者：刘惠[1] 吕琳[1] 褚海辰[1] 祝琳[1] 刘爱杰[1] 董河[1] Liu Hui;Lyu Lin;Chu Haichen;Zhu Lin;Liu Aijie;Dong He(Department of Anesthesiology,the Affiliated Hospital of Qingdao University,Qingdao 266003,Shandong,China)

机构地区：[1]青岛大学附属医院麻醉科,山东青岛266003

出　　处：《中华危重病急救医学》2022年第1期64-69,共6页Chinese Critical Care Medicine

基　　金：国家自然科学基金(81902001)。

摘　　要：目的观察高胆红素血症对模型大鼠肾小球的影响,并探讨其量效反应及机制。方法将24只成年雄性SD大鼠按随机数字表法分为4组,每组6只。采用每12 h腹腔注射1次胆红素、共4次制备高胆红素血症大鼠模型,小、中、大剂量胆红素组(分别为LB组、MB组、HB组)剂量分别为50、100、200 mg/kg;阴性对照组(NC组)给予不含胆红素粉末的相同溶剂。各组给药结束后收集24 h尿液,检测总蛋白(TP)水平;处死大鼠取心脏血,用全自动生化分析仪检测总胆红素(TBil)和直接胆红素(DBil)水平;取肾组织,过碘酸希夫(PAS)染色后光镜下观察肾小球形态,并进行肾小球损伤评分;醋酸铀-枸橼酸铅双染后,透射电镜下观察足细胞形态;采用比色法检测氧化应激指标超氧化物歧化酶(SOD)活性和丙二醛(MDA)含量;采用蛋白质免疫印迹试验(Western blotting)检测足细胞特异性标志物肾母细胞瘤蛋白1(WT-1)的表达水平。结果随着胆红素剂量增加,大鼠24 h尿TP、血TBil、血DBil、肾组织MDA含量逐渐升高,肾组织SOD活性和WT-1表达量逐渐降低,LB组、MB组、HB组与NC组比较差异均有统计学意义〔24 h尿TP(mg):24.85±2.22、52.57±3.66、56.84±3.49比7.50±1.33,血TBil(μmol/L):37.75±2.19、81.37±2.13、125.13±9.96比5.53±0.41,血DBil(μmol/L):15.50±1.96、37.88±1.05、64.53±2.89比2.38±0.35,肾组织MDA(μmol/g):3.14±0.65、5.01±0.28、7.50±1.08比2.30±0.20,肾组织SOD(kU/g):95.91±10.43、57.06±15.90、37.12±11.72比113.91±12.16,肾组织WT-1蛋白(WT-1/GAPDH):0.280±0.006、0.239±0.006、0.198±0.001比0.361±0.005,均P<0.05〕。光镜下显示,不同剂量胆红素各组大鼠肾小球系膜和基底膜厚度不均匀,部分伴有增生、增宽表现,且肾小球损伤评分随胆红素剂量增加而逐渐升高,LB组、MB组、HB组与NC组比较差异均有统计学意义(分:17.50±1.05、25.00±1.41、34.00±1.41比11.67±0.82,均P<0.05);透射电镜下显示,随着胆红素剂量的�Objective To observe the effect of hyperbilirubinemia on glomerulus of rats,and to explore its dose-response and mechanism.Methods Twenty-four adult male Sprague-Dawley(SD)rats were divided into four groups according to the random number table method,with 6 rats in each group.Hyperbilirubinemia rat model was reproduced by intraperitoneal injection of bilirubin once every 12 hours for 4 times,at doses of 50,100,and 200 mg/kg in low,medium,and high dose bilirubin group(LB group,MB group,HB group),respectively.The rats in negative control group(NC group)were given the same solvent without bilirubin powder.Urine was collected 24 hours after administration,and total protein(TP)level was detected.Then the rats were sacrificed,the blood was collected by cardiac puncture,and the total bilirubin(TBil)and direct bilirubin(DBil)levels were measured by automatic biochemical analyzer.The renal tissue was collected and stained with periodic acid-Schiff(PAS)staine,the glomerular morphology was observed under light microscope,and the glomerular injury score was performed.Podocyte morphology was observed by transmission electron microscopy after uranium acetate and lead citrate double staining.The activity of superoxide dismutase(SOD)and content of malondialdehyde(MDA)were determined by colorimetric method.The expression level of podocyte specific marker Wilms tumor protein-1(WT-1)was determined by Western blotting.Results With the increase of bilirubin dose,the contents of 24-hour urine TP,blood TBil,blood DBil and MDA content in kidney tissue were gradually increased,and the SOD activity and WT-1 expression in kidney tissue were gradually decreased.The differences between LB group,MB group,HB group and NC group were statistically significant[24-hour urine TP(mg):24.85±2.22,52.57±3.66,56.84±3.49 vs.7.50±1.33;blood TBil(μmol/L):37.75±2.19,81.37±2.13,125.13±9.96 vs.5.53±0.41;blood DBil(μmol/L):15.50±1.96,37.88±1.05,64.53±2.89 vs.2.38±0.35;kidney MDA(μmol/g):3.14±0.65,5.01±0.28,7.50±1.08 vs.2.30±0.20;kidney SOD(k

关 键 词：高胆红素血症 肾小球 氧化应激 足细胞 

分 类 号：R459.7[医药卫生—急诊医学]