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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 clarify the effect of hypoxia on the expression of vascular endothelial growth factor (VEGF) in human renal interstitial fibroblasts. Methods Primary cultured human renal interstitial fibroblasts, passed to the third generation for this experiment. The cells were treated with hypoxia for 24 hours or normal culture. The expression of VEGF mRNA was detected by semi-quantitative RT-PCR. The VEGF protein content was detected by flow cytometry. VEGF protein content was detected by double-antibody sandwich ELISA. Results (1) Hypoxia could significantly increase the expression of VEGF mRNA in human renal interstitial fibroblasts (0.34 ± 0.04 vs 0.10 ± 0.01, n = 4, P <0.01). (2) After hypoxia, the cell surface VEGF protein content increased significantly (average fluorescence intensity: 1.053 ± 0.055 vs 0.763 ± 0.057, n = 4, P <0.01). (3) The content of VEGF in the supernatant of hypoxic cultured cells was significantly higher than that of the normal control group [(2.009 ± 0.144) mg / L vs (1.647 ± 0.175) mg / L, n = 4, P <0.05]. Conclusion Hypoxia can induce the expression of VEGF in human renal interstitial fibroblasts, which may play a role in pathological changes caused by hypoxia.