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[摘要]
[已撤稿]目的：研究体外培养的人视网膜色素上皮细胞在人工模拟的缺氧和高糖环境中VEGF165 及VEGF165b 表达的情况。
方法：正常接种并体外培养人视网膜色素上皮细胞，待细胞稳定生长后以150μmol/L CoCl2 和25mmol/L 葡萄糖浓度分别模拟细胞缺氧和高糖环境，将接种细胞分为正常组(5.56mmol/L 葡萄糖)、缺氧组(5.56mmol/L 葡萄糖+150μmol/L CoCl2)、高糖组(25mmol/L 葡萄糖，不加CoCl2)、联合组(25mmol/L 葡萄糖+150μmol/L CoCl2)共四组。每组分别于12、24、36、48h提取RNA。MTT 比色法检测细胞活力及增长趋势； RT-PCR 法检测不同时间点四组RPE 细胞VEGF165 mRNA 和VEGF165b mRNA 的相对表达量。
结果：缺氧和高糖环境下RPE 细胞分裂增殖受限，细胞活力降低。同一组细胞不同时间点比较，正常组VEGF165和VEGF165b随时间变化的表达差异无统计学意义(P >0.05)，而缺氧组、高糖组、联合组VEGF165和VEGF165b的表达则随时间变化而变化，差异有统计学意义(P<0.05)。同一时间点各组相互比较，在缺氧模型建立后12h 各组细胞间VEGF165和VEGF165b的表达差异无统计学意义(P >0.05)，而24、36、48h 时各组细胞VEGF165和VEGF165b的表达差异有统计学意义(P<0.05)。
结论：体外培养RPE细胞能够正常表达VEGF165b，缺氧和高糖是在转录水平诱导VEGF165 mRNA的表达上调，同时降低了VEGF165b mRNA的表达量。
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[Abstract]
[THIS ARTICLE HAS BEEN RETRACTED]AIM: Through the expression of VEGF
165 and VEGF
165b in human retinal pigment epithelial cells
in vitro in artificial simulated hypoxia and high glucose environment, to discuss their roles in the development of diabetic retinopathy and the relationship between each other.
METHODS: After normal inoculation and cultivation of human retinal pigment epithelial cells(RPE)in vitro, the cells was divided into the normal group(5.56mmol/L glucose, without CoCl2), the hypoxia group(5.56mmol/L glucose + 150μmol/L CoCl2), the high glucose group(25mmol/L glucose, without CoCl2), the combination group(25mmol/L glucose + 150μmol/L CoCl2), a total of four groups. The RNA of each group was extracted respectively in 12h, 24h, 36h, and 48h. We used the MTT colorimetry to detect cell vitality and growth trend; RT-PCR method to detect VEGF165 and VEGF165b relative expression of mRNA of RPE cells in four different time points.
RESULTS: Hypoxia and high sugar environment limited proliferation of RPE cell division and cell vitality. After comparing cells of the same group in different time points, in the normal group there was no statistically significant different expression over time(P>0.05); the expression in the hypoxia group, the high glucose group and the combination group increased over time, the difference was statistically significant(P<0.05). At the same time, differences of the expression between groups was not statistical significant in 12h(P>0.05); the difference was statistically significant in 24h, 36h, 48h(P<0.05).
CONCLUSION: Cultured RPE cells can express VEGF165b normal. Lack of oxygen and high glucose can induce the increase of VEGF165 mRNA, at the same time reduces the VEGF165b mRNA expression.
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