烟酰胺腺嘌呤二核苷酸磷酸氧化酶亚单位4在草酸钙结石患者肾乳头中的表达及其机制  

Expression of nicotinamide adenine dinucleotide phosphate oxidase subunit 4 in renal papilla of patients with calcium oxalate stones and its mechanism

作　　者：寻阳 王少刚[1] Xun Yang;Wang Shaogang(Department of Urology,Tongji Hospital,Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430030,China)

机构地区：[1]华中科技大学同济医学院附属同济医院泌尿外科,武汉430030

出　　处：《中华实验外科杂志》2020年第8期1425-1427,共3页Chinese Journal of Experimental Surgery

基　　金：国家自然科学基金面上项目(81770704)。

摘　　要：目的探讨烟酰胺腺嘌呤二核苷酸磷酸氧化酶亚单位4(Nox4)在草酸钙结石患者肾乳头中的表达及其作用机制。方法通过GEO数据库检索Randall钙斑芯片,按照GSE73680的样本描述,将样本分为无结石对照组(6例)和草酸钙结石组(24例)。采用GEO2R分析正常人肾乳头和草酸钙肾结石患者肾乳头组织中Nox4及成骨相关蛋白基因的表达差异变化。采用高钙处理大鼠肾小管上皮细胞(NRK-52E),免疫荧光技术(IF)检测Nox4的表达,蛋白质印迹法(Western blot)检测Nox4和骨形态发生蛋白(BMP-2)的表达,化学发光法测定氧化应激标志物丙二醛(MDA)和超氧化物歧化酶(SOD)的活性。组间比较采用t检验。结果GEO2R分析结果显示Nox4在草酸钙结石组表达升高,差异表达倍数为0.726倍(t=2.040,P<0.05),基质Gla蛋白(MGP)在草酸钙结石组表达下降,差异表达倍数为-1.228倍(t=-1.230,P>0.05),骨钙素(OCN)在草酸钙结石组表达升高,差异表达倍数为0.439倍(t=1.241,P>0.05),骨保护素(OPG)在草酸钙结石组表达升高,差异表达倍数为0.639倍(t=0.888,P>0.05),骨桥蛋白(OPN)在草酸钙结石组表达升高,差异表达倍数为1.664倍(t=1.171,P>0.05)。免疫荧光结果显示高钙处理的NRK-52E细胞中Nox4的平均荧光吸光度明显高于对照组(0.651、1.446、1.480,t=8.023、6.477,P<0.05)。Western blot显示相对于未处理的正常对照组,高钙处理的NRK-52E细胞中Nox4的表达明显增加(0.426、0.594、0.734,t=28.954、53.082,P<0.01),BMP-2表达也呈增高趋势(0.242、0.472、0.676,t=12.765、14.257,P<0.01)。此外,高钙处理NRK-52E细胞后,MDA含量增加(0.188、0.390、0.650,t=2.679、6.686,P<0.05),SOD活性下降(40.630、38.060、36.230,t=4.079、10.390,P<0.05)。结论Nox4在肾结石患者肾乳头中表达明显增加,Nox4介导的氧化应激可能在肾结石形成过程中发挥重要作用。Objective To explore the expression of nicotinamide adenine dinucleotide phosphate oxidase subunit 4(NOX4)in the renal papilla of patients with calcium oxalate stones and its potential mechanism.Methods The Randall calcium spot chip was searched through the GEO database,and the samples were divided into a Non-stone control group and a calcium oxalate stone group according to the sample description of GSE73680.There were 6 samples in the control group without stones and 24 samples in the calcium oxalate stones group.GEO2R was used to analyze the differences in the expression of Nox4 and osteogenic related protein genes in the renal papilla tissues of patients with normal renal papilla and calcium oxalate renal stones.High calcium treatment of rat renal tubular epithelial cells(NRK-52E),immunofluorescence technique(IF)was used to detect the expression of Nox4,Western blotting was used to detect the expression of Nox4 and bone morphogenetic protein-2(BMP-2),chemiluminescence Method was used to determine the activity of oxidative stress markers malondialdehyde(MDA)and superoxide dismutase(SOD).SPSS 24.0 software was used to analyze the data by t test.Results The results of GEO2R analysis showed that the expression of Nox4 increased in calcium oxalate stone group,and the differential expression multiple in the two groups was 0.726 times(t=2.040,P<0.05);the expression of matrix Gla protein(MGP)decreased in calcium oxalate stone group,and the differential expression multiple was-1.228 times(t=-1.230,P>0.05);the expression of osteocalcin(OCN)increased in calcium oxalate stone group,and the differential expression multiple was 0.439 times(t=1.241,P>0.05);the expression of osteoprotegerin(OPG)increased in calcium oxalate stone group,and the differential expression multiple was 0.639 times(t=0.888,P>0.05);the expression of osteopontin(OPN)increased in calcium oxalate stone group,and the differential expression multiple was 1.664 times(t=1.171,P>0.05).Immunofluorescence results showed that the average fluorescence density of

关 键 词：草酸钙结石 烟酰胺腺嘌呤二核苷酸磷酸氧化酶亚单位4 氧化应激 成骨相关蛋白 

分 类 号：R692.4[医药卫生—泌尿科学]