光损伤后人视网膜色素上皮细胞血管内皮生长因子A及其受体的表达

Expression of vascular endothelial growth factor A and its receptors in light-injured human retinal pigment epithelial cells

目的观察光损伤后人视网膜色素上皮（RPE）细胞血管内皮生长因子A（VEGF-A）及其受体可溶性fm样酪氨酸激酶受体（sFh-1）和包含激酶植入区域受体（KDR）的表达。方法体外培养人RPE细胞，取第8～12代生长良好的传代细胞用于实验。将同代细胞随机分为对照组和光损伤组。以18w冷白色荧光灯作为光源，自制光照器。采用双层高压消毒锡纸包裹对照组细胞，与光损伤组细胞一同置于自制光照器下，控制光照强度在（2200±300）Lux，持续光照12h建立光损伤模型。于光损伤后0、6、12、24h终止培养，采用逆转录聚合酶链反应（RT—PCR）和蛋白免疫印迹法（Westernblot）检测VEGF-A、sFlt-1及KDR的mRNA及蛋白表达。结果光损伤组VEGF—AmRNA及蛋白表达在光损伤后6h时明显增加，12h时达到高峰，明显高于对照组（t=2．74，2．93；P％0．05），随后降低。sFlt-1的mRNA及蛋白表达在光损伤后12h达到高峰，明显高于对照组（t=4．32，P〈0．01）；24h时明显低于对照组（t=2．41，P〈0．05）。KDR的mRNA及蛋白表达在光损伤后6、12h时较对照组无明显变化（t=1．84，P〉0．05），24h时高于对照组（t=2．89，P％0．05）。结论光损伤后6、12h时，RPE细胞VEGF-A及sFh-1的表达明显增高，KDR的表达相对稳定。光损伤后24h时，RPE细胞VEGF-A及sFlt-1的表达降低，KDR的表达增高。

Objective To observe the expression of vascular endothelial growth factor A （VEGF-A） and its receptors sFlt-1, kinase insert domain receptor （KDR） in light-injured human retinal pigment epithelial （RPE） cells. Methods Cultured human RPE cells （8th - 12th generations） were divided into normal control group and light damage group. The cells of two groups were exposed to the 18 W cold white light （2200＋300） Lux for 12 hours to induce light damage responses, but the cells of normal control group were packed by tinfoil with double-deck high pressure disinfection. The VEGF-A, sFlt：l and KDR mRNA and protein expressions were detected by reverse transcription-polymerase chain reaction （RT-PCR） and Western blot at 0, 6, 12, 24 hours after light damage. Results The VEGF：A mRNA and protein expressions in light damage group were significantly increased at 6 hours, and reached its peak at 12 hours after light damage which obviously higher than that in normal group （t=2.74, 2.93; P〈0.05）, and then went down gradually. The sFlt-1 mRNA and protein expressions in light damage group reached its peak at 12 hours after light damage which obviously higher than that in normal group （t= 4.32, P〈0.01）, but obviously lower than that in normal group at 24 hours after light damage （t=2.41, P〈0.05）. The KDR mRNA and protein expressions in light damage group were obviously higher than that in normal group at 24 hours after light damage （t=2.89, P〈0. 05） ,but there was no changes at 6, 12 hours after light damage （t=l. 84, P〈0.05）. Conclusions At 6, 12 hours after light damage, the expressions of VEGF-A and sFlt-1 increases significantly and KDR expression is stable in light：injured RPE cells. At 24 hours after light damage, the expression of VEGF-A and sFlt-1 decreases, but KDR expression increases in light-injured RPE cells.