白蛋白对肾小管上皮细胞血管紧张素转换酶2表达的抑制作用

Inhibitory effect of albumin on the expression of angiotensin-converting enzyme 2 in human proximal tubular cells

目的观察白蛋白对人近端肾小管上皮细胞株(HK-2)血管紧张素转换酶2(ACE2)mRNA和蛋白表达的影响,探讨蛋白尿激活肾脏局部RAS的机制。方法采用不同浓度的牛血清白蛋白(BSA)作用HK-2细胞不同时间。以实时RT-PCR和Western印迹检测ACE2 mRNA和蛋白的表达。采用放射免疫法检测细胞上清液中血管紧张素Ⅱ(ATⅡ)含量。采用激光共聚焦显微镜观察ACE2蛋白的表达。结果与对照组(相对表达量为0)相比,BSA使ACE2 mRNA表达显著减少(2.5mg/ml:—1.05±0.12;5mg/ml:—1.30±0.11;10mg/ml:—2.54±0.44;P均<0.05)。同时,BSA使ACE2蛋白表达显著降低(对照组:0.90±0.10;2.5mg/ml:0.66±0.09;5mg/ml:0.50±0.07;10mg/ml:0.35±0.05;P均<0.05)。其中BSA(10mg/m1)使ACE2 mRNA及蛋白表达减少呈时间依赖性(P均<0.05)。与对照组相比,不同浓度BSA组的细胞上清液中的ATⅡ均显著升高(P均<0.05)。激光共聚焦显微镜可见ACE2主要分布于细胞膜。结论BSA可显著抑制HK-2细胞ACE2 mRNA和蛋白的表达,此作用所导致的近曲小管间质液ATⅡ浓度升高可能与间质纤维化相关。

Objective To investigate the influence of albumin on the expression of angiotensin-converting enzyme 2 （ACE2） in cultured human proximal tubular cells （HK-2）. Methods The cells were incubated with 2.5,5,10mg/ml of bovine serum albumin （BSA） for 6h and 12h respectively. The expressions of ACEZ was detected by real-time RT-PCR and Western blot. AT Ⅱ concentration in the supernatant was detected by radioimmunoassay. Confocal laser scanning microscope was used to detect the distribution of ACE2. Results BSA led to a significantly decreased expression of ACE2 mRNA compared to the control group （control： 0, 2.5mg/mh-1.05±0. 12, 5mg/ml：-1.3±0.11, 10mg/ml：-2. 54±0.44, P〈0. 05）. While ACE2 protein significantly decreased. Furthermore, expression of ACE2 mRNA and protein significantly decreased in the time-dependent manner （P〈0. 05）. ATⅡ concentration in the supernatant significantly increased （P〈0. 05）. By LSCM, the strong staining for ACE2 could be detected in cell menbrane. Conclusions BSA could inhibit HK-2 cells to express ACE2, which may cause the attenuation of the degradation of ATⅡ and finally contribute to the intrarenal activation of renin angiotensin system.