骨化三醇对糖尿病大鼠肾脏血管紧张素转换酶与血管紧张素转换酶2的影响

目的观察血管紧张素转换酶（angiotensinconvertingenzyme，ACE）和血管紧张素转换酶2（angiotensinconvertingenzyme2，ACE2）在糖尿病大鼠肾脏的表达及骨化三醇对其的影响。方法6～8周龄Wistar大鼠被随机分为3组，分别为对照组、糖尿病组、骨化三醇组（O．03μg·kg-1·d-1）。16周后处死各组大鼠，留取标本。放射免疫法检测尿胆微球蛋白（p2-microglobulin，p2-MG）、尿白蛋白（urinaryalbumin，Alb）、血全段甲状旁腺素（intactparathyrin，iPTH）；分光光度法检测尿肌酐（urinecreatinine，UCr）；苦味酸法检测血肌酐（serumcreatinine，SCr）；全自动生化仪检测血钙（Ca）和血磷（P）。PAS染色观察肾脏病理改变并进行半定量统计学分析。实时定量基因扩增荧光（quan—titativepolymerasechainreaction，QPCR）检测肾脏中ACE和ACE2mRNA表达。免疫组织化学法检测肾脏中ACE和ACE2蛋白表达。结果与糖尿病组相比，骨化三醇组尿陀一MG、尿Alb／UCr降低（P〈0．05），内生肌酐清除率（endogenouscreatinineclearancerate，CCr）升高（P〈O．05）。各组大鼠血Ca、血P及iPTH差异无统计学意义（P〉0．05）。PAS病理染色及半定量分析结果提示，与糖尿病组相比，骨化三醇组大鼠肾脏肾小球硬化程度及肾小管间质纤维化程度有所减轻（P〈0．05）。与对照组比较，糖尿病组大鼠肾脏ACE与ACE2mRNA和蛋白表达均升高（P〈O．05），ACE／ACE2升高（P〈0．05）。与糖尿病组比较，骨化三醇组大鼠肾脏ACEmRNA和蛋白的表达降低（P〈O．05），ACE2mRNA和蛋白的表达升高（P〈（）．05），ACE／ACE2降低（P〈0．05）。相关分析结果显示，尿82-MG、Alb／ucr与ACE／AcE2呈正相关（P〈0．05），CCr与ACE／ACE2呈负相关（P〈0．05）。结论骨化三醇降低糖尿病大鼠蛋白尿、升高CCr，改善病理损伤，其肾脏保护作用可能与调节ACE和ACE2的表达有关。

Effects of calcitriol on renal expression of angiotensin converting enzyme and angiotensin converting en- zyme 2 in experimental diabetic rats

Objective To investigate the expression of angiotensin converting enzyme/angioten- sin converting enzyme 2（ACE/ACE2） in the kidney of diabetic rats, and the influences of calcitriol. Methods Six-eight-weeks old Wistar rats were randomly divided into 3 groups： controls, diabetic rats, diabetic rats treated with calcitriol （0. 03 μg. kg-1 .day-1）. Urinary 132 microglobulin（132-MG）/urinary albumin （Alb） and intact parathyrin （iPTH） were determined by radioimmunoassay, urine creatinine （UCr） by spectrophotometry, serum creatinine（SCr） by Picric acid methods, and blood calcium and se- rium inorganic phosphorus by automatic biochemical analyzer. The renal pathological changes were ob- served by PAS staining and lesions semi-quantitative analysis. ACE/ACE2 mRNA expression in the kidney was detected by QPCR, and ACE/ACE2 protein expression by immunohistochemistry 16 weeks after administration. Results The levels of urinary 132-MG and urinary Alb/UCr in rats treated with calcitriol were lower than those in diabetic rats （P〈0. 05）, and CCr in rats treated with calcitriol was higher than that in diabetic rats （P%0. 05）. There was no significant difference in blood calcium and serum inorganic phosphorus （P〉0. 05）. PAS staining results and semi-quantitative analysis indicated that the degrees of glomerular sclerosis and renal tubule interstitial fibrosis in diabetic rats treated with calcitriol were alleviated as compared with those in diabetic rats. The expression levels of ACE/ACE2 （mRNA and protein） and the ratio of ACE/ACE2 in diabetic rats were higher than those in control rats （P〈0. 05）. The expression of ACE（mRNA and protein） and the ratio of ACE/ACE2 in rats trea- ted with calcitriol were lower than those in diabetic rats （P〈0. 05）. The expression levels of ACE2 （mRNA and protein） in rats treated with calcitriol were higher than those in diabetic rats （P〈0. 05）. Correlation analysis revealed that urine β2-MG and urine Alb/UCr were positively correlated with the ratio of ACE/ACE2 （P〈0. 05） ,and CCr was negatively correlated with the ratio of ACE/ACE2 （P〈 0. 05）. Conclusions Calcitriol can improve proteinuria, increase CCr and relieve the renal pathological damages in diabetic rats probably by regulating the ACE/ACE2 expression.