摘要
目的研究长期缺氧对体外培养的人ARPE-19细胞血管内皮生长因子(vascular endotheial growth factor,VEGF)表达的诱导作用,以及金雀异黄素(genistein,Gen)对其表达的影响和机理。方法使用半定量RT-PCR和酶联免疫吸附试验(ELISA)检测ARPE-19细胞中VEGFmRNA和蛋白的表达,分析不同浓度Gen以及PD98059、TyrphostinA25对ARPE-19细胞缺氧24h诱导的VEGF表达的影响。结果(1)正常对照组有少量VEGF表达,缺氧24h可明显提高ARPE-19细胞VEGFmRNA和蛋白的表达,分别为(9.566±1.528)倍和(2.636±0.499)倍;(2)Gen(50μmol·L-1,100μmol·L-1,200μmol·L-1)与缺氧组对比,分别抑制缺氧组VEGFmRNA表达的55.03%、78.72%、90.69%,蛋白的表达量为(189.60±20.20)ng·L-1(117.70±21.97)ng·L-1、(49.70±13.17)ng·L-1,抑制作用呈浓度依赖性;(3)PD98059、TyrphostinA25也可抑制缺氧诱导的VEGFmRNA表达的67.24%和56.25%,蛋白表达量为(86.33±12.47)ng·L-1、(98.08±2.42)ng·L-1。结论Gen可抑制长期缺氧诱导的ARPE-19细胞VEGF表达,并可能是通过抑制p42/p44MAPK途径和HIF-1途径共同抑制VEGF的表达,提示Gen对视网膜新生血管的形成具有预防和潜在的治疗价值。
Objective To investigate the up-regulation of VEGF expression induced by long-term hypoxia in ARPE-19 cells, the effects of genistein(Gen) on hypoxia-induced VEGF expression, and the mechanisms by which Gen worked. Methods RT-PCR and ELISA were performed to examine the expressions of VEGF mRNA and protein when ARPE-19 cells were exposed to hypoxia for 24 hours, and different concentrations of Gen, PD98059 and Tyr-phostin A25 were used to observe their effects on the expressions. Results Hypoxia could up-regulate the expression of VEGF mRNA and protein(9.566± 1.528) times and (2. 636± 0. 499) times, respectively, as the normal control which showed little expression of VEGF; Gen inhibited the expression of VEGF mRNA induced by hypoxia for 24 hours in a dose-dependent manner, the inhibitory rate were 55.03% ,78.72% and 90.69% , respectively with the expressions of VEGF protein of(189. 60±20.20) ng·L-1, (117.70±21.97) ng·L-1.and (49.70±13.17) ng·L-1 respectively. PD98059 and Tyrphostin A25 could also inhibit the hypoxia-induced expression of VEGF mRNA by 67.24% and 56.25% , respectively,with the protein expressions of(86.33±12.47) ng·L-1 and(98.08±2.42) ng·L-1, respectively. Conclusion Gen could inhibit hypoxia-induced VEGF expression in ARPE-19 cells possibly through the inhibition of p42/ p44MAPK and HIF-1 pathways. It was suggested that Gen might play a latent role in the prevention and treatment of retinal neovascularization.
出处
《眼科新进展》
CAS
2005年第3期219-222,共4页
Recent Advances in Ophthalmology
基金
国家自然科学基金资助(编号:30000207)南京医科大学科技创新基金资助(编号:CX2001003)~~
