骨化三醇对高糖诱导的肾小管上皮细胞肾素高表达的影响  

Effect of calcitriol on up-regulated expression of renin in renal tubular epithelial cells induced by high glucose

作　　者：赵天涯 林梅[1] 李梦思[1] 贺磊[1] 李垚瑶 高苹[1] 

机构地区：[1]武汉大学中南医院肾内科,武汉430071

出　　处：《临床肾脏病杂志》2016年第6期365-371,共7页Journal Of Clinical Nephrology

基　　金：国家自然科学基金(NO.81000304);湖北省自然科学基金(NO.2010CDB00402)

摘　　要：目的探讨骨化三醇对高糖状态下肾小管上皮细胞肾素表达的影响及其机制。方法按随机数字表法将24只6~8周龄Wistar大鼠随机分为对照组、糖尿病组、骨化三醇组(0.03μg·kg^(-1)·d^(-1)),各组8只。16周后处死各组大鼠,检测尿蛋白、尿肌酐、内生肌酐清除率(endogenous creatinine clearance rate,CCr),并应用实时荧光定量法(real time-polymerase chain reaction,RT-PCR)和蛋白印迹法(Western blot)检测肾脏组织肾素及磷酸化细胞外信号调节激酶(phosphorylated extracellularsignal-regulated kinase,p-ERK)/细胞外信号调节激酶(extracellularsignal-regulated kinase,ERK)表达情况。NRK-52E细胞分组:①正常对照组;②高糖组(糖浓度为25 mmol/L,不同时间点);③高糖+骨化三醇(10~8 mol/L、10~9 mol/L、10~10 mol/L)组;④高糖+ERK抑制剂(FR18020410~5 mol/L)组。RT-PCR法检测肾素mRNA表达,Western印记法检测肾素蛋白、p-ERK/ERK蛋白表达。结果与对照组比较,糖尿病组大鼠尿β2微球蛋白(β2-microglobulin,β2-MG)、尿白蛋白/尿肌酐升高(P<0.05),CCr降低(P<0.05);与糖尿病组比较,骨化三醇组大鼠尿β2-MG、尿白蛋白/肌酐降低(P<0.05),CCr升高(P<0.05)。与对照组比较,糖尿病组大鼠肾组织肾素、p-ERK/总ERK表达均明显升高,骨化三醇能显著减轻糖尿病大鼠上述蛋白的高表达(P<0.05)。高糖可诱导NRK-52E细胞ERK蛋白磷酸化(P<0.05),诱导肾素mRNA及蛋白表达上调,并呈时间依赖性(P<0.05)。骨化三醇可浓度依赖性地降低高糖诱导的NRK-52E细胞ERK蛋白磷酸化及肾素mRNA、蛋白的高表达(P<0.05)。ERK抑制剂能部分抑制高糖诱导的肾素蛋白高表达(P<0.05)。结论骨化三醇降低糖尿病大鼠蛋白尿,具有肾脏保护作用,其机制可能与抑制ERK通路降低高糖诱导的肾素高表达有关。Objective To explore the effect of calcitriol on the expression of renin in renal tubular epithelial cells induced by high glucose and the mechanism. Methods Six-eight-week old Wistar rats were randomly divided imo control group, diabetic group and calcitriol （0. 03μg·kg^-1·day^-1）-treated group. The rats were sacrificed for determination of urinary protein, urinary creatinine, and creatin[ne clearance rate （CCr） at the end of 16 weeks. RT-PCR and Western blotting were used to detect the expression of renin and phosphorylated extracellular signal-regulated kinase/extracellular sig- nal-regulated kinase（p-ERK/ERK） in kidney tissues. Cultured NRK-52E cells were divided into four groups： normal control group, high glucose group（25 mmol/L glucose at different time points respectively）, high glucose＋ calcitriol （10^-8, 10^-9 , and 10^-10 mol/L） group, and high glucose ＋ ERK inhibitor（FR180204, 10^-5 mol/L） group. The expression of renin and p-ERK/ERK was detected by RT-PCR and Western blotting. Results As compared with the control group, urine β2-microglobulin,and urinary Alb/UCr were increased （P〈0. 05）, but CCr decreased（P〈0. 05） in diabetic group. As compared with diabetic group, urinary Alb/UCr was reduced（P%0. 05） and CCr increased （P〈0.05） in caleitriol-treated group. As compared with the control group, the expression of renin and p-ERK/ ERK in kidney tissue of diabetic group was significantly increased, and that was significan.tly reduced in calcitriol-treated group（P〈0. 05）. High glucose could induce the phosphorylation of ERK protein （P〈0. 05）, and time-dependently up-regulate the expression of renin mRNA and protein in NRK-52E cells（P〈0. 05）. Calcitriol suppressed the high expression of p-ERK and renin induced by high glucose concentration-dependently in NRK-52E cells（P〈0. 05）. ERK inhibitor could partially inhibit the high expression of rennin （mRNA and protein） {nduced by high glucose in NRK-52E cells（P〈0. 05）. Conclu

关 键 词：骨化三醇 肾小管上皮细胞 肾素 

分 类 号：R587.2[医药卫生—内分泌] R692.9[医药卫生—内科学]