缺氧诱导人肾间质成纤维细胞血管内皮生长因子的表达

Hypoxia stimulates vascular endothelial growth factor production in human renal flbroblasts

目的 阐明缺氧对人肾间质成纤维细胞血管内皮生长因子(VEGF)表达的影响。方法 原代培养获得人肾间质成纤维细胞,传至第3代用于本实验。细胞经缺氧处理24 h或正常培养后以半定量RT-PCR法检测VEGF mRNA表达量,以流式细胞术检测细胞表面VEGF蛋白含量,以双抗夹心ELISA法检测培养上清VEGF蛋白含量。结果 (1)缺氧能显著增加人肾间质成纤维细胞VEGF mRNA表达(0.34±0.04 vs 0.10±0.01,n=4,P<0.01)。(2)缺氧后细胞表面VEGF蛋白含量较正常对照显著增加(平均荧光强度:1.053±0.055 vs 0.763±0.057,n=4,P<0.01)。(3)缺氧培养细胞上清VEGF含量显著高于正常对照组[(2.009±0.144)mg/L vs(1.647±0.175)mg/L,n=4,P<0.05]。结论 缺氧能诱导人肾间质成纤维细胞表达VEGF,这在缺氧引起的肾脏病理改变中可能起到一定的作用。

Objective To investigate the effect of hypoxia on vascular endothelial growth factor (VEGF) expression and secretion in human renal fibroblasts. Methods Human renal fibroblasts were primary cultured from donor kidney, and cells in passage 3 were used in this experiment. Hypoxia was induced by incubation in a gas-tight box gassed with 10% CO2/10% H2/80% N2 for 24 hours. VEGF mRNA levels were measured by semi-quantitative RT-PCR method. VEGF proteins expressed on cell surface were measured by flow cytometry. VEGF proteins secreted into supernatants were measured by ELISA method. Results Hypoxia caused a 2. 5-fold increase of VEGF mRNA level in human renal fibroblasts. Hypoxia also induced significant increases of VEGF proteins on cell surfaces (mean fluorescence intensities: 1. 053±0. 055 vs 0.763±0.057, n=4,P <0.01), and in culture supernatants [(2. 009±0. 144)mg/L vs (1. 647±0. 175) mg/L, n=4, P < 0. 05] . Conclusion Hypoxia stimulates VEGF expression and secretion in human renal fibroblasts.