摘要
目的探讨被醛固酮(ALDO)活化的人近端肾小管上皮细胞系(HKC)对人肾间质成纤维细胞(hRIFs)合成Ⅰ型胶原(ColⅠ)的影响。方法利用体外细胞培养及共培养技术进行如下试验。(1)用不同浓度及不同作用时间的外源性ALDO刺激HKC,逆转录多聚酶链反应(RT-PCR)及酶联免疫吸附(ELISA)方法检测转化生长因子-β1(TGF-β1)mRNA及蛋白质表达。(2)用不同浓度及不同作用时间的安体舒通与内皮素-1(ET-1)共同刺激HKC,同上检测TGF-β1mRNA及蛋白质表达。(3)用外源性ALDO活化的HKC与hRIFs共培养(培养液中加或不加抗TGF-β1抗体),ELISA方法检测hRIFs的ColⅠ合成量。结果(1)外源性ALDO使TGF-β1表达呈剂量依赖及时间依赖性增加。1×10-9及1×10-7mol/LALDO刺激组TGF-β1表达量显著高于0mol/LALDO组(P<0.05或0.01);10-7mol/LALDO刺激不同时间后,TGF-β1表达量显著高于0h组(P<0.05或0.01)。(2)安体舒通使TGF-β1表达呈剂量及时间依赖性减少。10-9、10-7mol/L安体舒通组TGF-β1表达量显著低于0mol/L安体舒通组(P<0.05或P<0.01)。10-9mol/LET-1加10-7mol/L安体舒通与单用10-9mol/LET-1刺激不同时间后,两组间TGF-β1表达,差异有显著意义(P<0.05或0.01)。(3)10-7mol/LALDO刺激HKC12h活化细胞后,再与hRIFs共培养48h,hRIFs对ColⅠ的合成量显著增多(P<0.01);1.0、2.0μg/ml抗TGF-β1抗体能部分抑制此反应(P<0.05)。结论外源性ALDO能使HKC合成TGF-β1增加;内皮素刺激HKC产生的内源性ALDO能促进其自身合成TGF-β1;被ALDO活化的HKC能通过“传话”作用促进hRIFs合成Col-1,该作用部分为TGF-β1介导。
Objective Local aldosterone （ALDO） could be synthesized by human proximal tubular epithelial cell lines （HKC） after the stimulation of endothelin-1 （ET-1） in vitro. T to observe the effect of tubular epithelial cells activated by aldosterone （ALDO） on renal interstitial fibroblasts in co-culture system.Methods （1） Human Proxima tubulor epithelial cells of the line HKC were stimulated with ALDO at different concentrations and times, then reverse transcription polymerase chain reaction （RT-PCR） and enzyme-linked immunosorbent assay （ELISA） were performed to detect TGF-β1 expression; （2） HKC were co-stimulated with ET-1 （10^-9 mol/L） and spironolactone at different concentrations and times to evaluate the influence of endogenous ALDO on TGF-β1 expression; （3） HKC which were activated by 10^-7 mol/L ALDO for 12 h, and hRIFs were co-cultured for 48 h with or without anti-TGF-β1 antibody （ 1.0, 2. 0 μg/ml） in the media, then the production of type I collegen （Col-Ⅰ） in the cell layer of hRIFs was detected by ELISA. Results （ 1 ） After stimulation with ALDO, the expression of TGF-β1 by HKC was up-regulated in a dose-and time-dependent manner. With 10^-9 or 10^-7 mol/L ALDO stimulation （mRNA determination at 12 h and protein at 48 h）, the expression of TGF-β1 was significantly increased （ vs 0 mol/L ALDO, P 〈 0. 05 or 0. 01 ）. With 10^-7 mol/L ALDO stimulated at different times （ mRNA determination at 8 h, 12 h and 16 h and protein at 12 h, 24 h and 48 h）, the expression of TGF-β1 was also significantly increased （vs 0 h, P 〈0. 05 or 0. 01 ）. （2） After co-stimulation with ET-1 and spironolatone, the expression of TGF-β1 by HKC was down-regulated in a dose-and-time dependent manner along with spironolatone. The expression of TGF-β1 mRNA and protein was decreased in the 10^-9 or 10^-7 mol./L spironolatone groups compared with 0 mol./L group （P 〈 0.05 or 0.01 ）. The expression of TGF-β1 （mRNA and protein） was significantly decr
出处
《中华医学杂志》
CAS
CSCD
北大核心
2005年第29期2070-2075,共6页
National Medical Journal of China
关键词
醛固酮
肾小管
上皮细胞
转化生长因子Β
成纤维细胞
Aldosterone
Kidney tubules
Epithelial cells
Transforming growth factor beta
Fibroblasts
