ACTHR在介导血管紧张素Ⅱ和氯化钾对醛固酮分泌调控中的实验研究  

ACTHR in regulating aldosterone secretion to angiotensin Ⅱ and potassium chloride stimulation Hu

作　　者：胡东亮[1] 刘同族[1] 倪栋[1] 王行环[1] 

机构地区：[1]武汉大学中南医院泌尿外科,430071

出　　处：《中华内分泌外科杂志》2016年第4期313-316,共4页Chinese Journal of Endocrine Surgery

基　　金：国家自然科学基金资助项目（81200579）

摘　　要：目的研究血管紧张素Ⅱ（AT-Ⅱ）和氯化钾刺激人肾上腺皮质癌H295R细胞后醛固酮合成酶（CYP11B2）mRNA及醛固酮激素的改变情况，并探讨促肾上腺皮质激素受体（ACTHR）在介导该反应中的作用。方法用慢病毒包装的ACTHR高表达载体感染H295R细胞作为实验组。空载体病毒转染组细胞为对照组。应用免疫印迹Westernblot和荧光定量PCR法验证ACTHR表达情况。分别用100nmol／LAT-Ⅱ及16mmol／L氯化钾刺激各组细胞，刺激24h后测定CYPllB2mRNA表达水平，对比2组细胞CYP11B2mRNA增幅情况。用ELISA试剂盒测定醛固酮激素的含量，对比2组细胞醛固酮增幅情况。结果实验组细胞ACTHR在蛋白水平和mRNA水平分别增加2．4倍和18倍（P〈0．05）。AT-Ⅱ刺激24h后实验组细胞CYP11B2 mRNA表达水平较对照组细胞高1．7倍，2组细胞醛固酮激素水平分别为（121．98±8．31）ng／L和（104．05±6．88）ng／L，前者增幅为后者的2．06倍（—P〈0．05）；氯化钾刺激各组细胞24h后，实验组细胞CYP11B2mRNA表达水平较对照组细胞高19．2倍，2组细胞醛固酮激素水平分别为（137．67±10．35）ng／L和（104．05±6．88）ng／L，前者增幅为后者的3．13倍（P〈0．05）。结论ACTHR高表达可增加醛固酮对AT-Ⅱ和氯化钾刺激的敏感性，有望成为研究原发性醛固酮增多症重要突破口。Objective To study CYP11B2 mRNA and aldosterone secretion alteration in human adreno- cortical carcinoma H295R cell after angiotensin Ⅱ（AT-Ⅱ） and potassium chloride stimulation, and to investigate the effect ofadrenocorticotropic hormone receptor （ACTHR） on them. Methods Lenfiviral vector was used to in- crease ACTHR expression. It was transfected into the H295R cells. Similarly, another H295R cells, without AC- THR vector, was used as the control group. ACTHR alteration was measured by Western blot and real-time polymerase chain reaction （RT-PCR）. CYP11B2 mRNA was detected at 24 hours after 100 nmol/L AT-Ⅱ/16 mmol/L KCL stimula- tion, and the amplification of the two groups was compared. Aldosterone was measured by ELISA kit. Results Com- pared with those in control cells, the protein and mRNA level of ACTHR in experimental cells were increased 2.4 times and 18 times respectively （P〈0.05）. CYPllB2 mRNA of experimental cells was 1.7 times higher than con- trol cells after 24 h stimulation of AT-II. Aldosterone production was 121.98±8.31 and 104.05±6.88 ng/L respec- tively. The former amplification was 2.06 times higher than that of the latter （P〈0.05）. Similarly, CYP11 B2 mR- NA of experimental cells was 19.2 times higher than control cells after 24 h stimulation of KCL. Aldosterone pro- duction was 137.67±10.35 and 104.05 ±6.88 ng/L respectively. The former amplification was 3.13 times higher than that of the latter （P〈0.05）. Conclusion Overexpression of ACTHR increases the sensitivity and response of CYP11B2 mRNA and aldosterone to AT-Ⅱ and KCL stimulation, and ACTHR is expected to become a key pro- tein in understanding primary aldosteronism.

关 键 词：促肾上腺皮质激素受体 H295R细胞 血管紧张素Ⅱ 氯化钾 醛固酮合成酶 原发性醛固酮增多症 

分 类 号：R586.24[医药卫生—内分泌]