银杏叶提取物对光损伤后视网膜色素上皮细胞蛋白质表达的影响

目的 观察中药银杏叶提取物对光损伤后视网膜色素上皮(RPE)细胞蛋白质表达的影响,探讨银杏叶提取物保护RPE光损伤的分子机制.方法 生长良好的人RPE细胞(ARPE-19)融合生长到80%时,随机分为正常对照组、光损伤组和银杏叶提取物处理组.运用冷白光以(2200±300)1x光照ARPE-19,建立光损伤模型;以100 μg/ml的银杏叶提取物(EGb761)处理光损伤的RPE细胞.提取细胞可溶性蛋白,进行蛋白质双向电泳,ImageMaster凝胶软件分析差异蛋白质点,并选取表达差异蛋白进行质谱鉴定和生物信息学分析.结果 蛋白图谱差异分析显示,光损伤组与正常对照组比较,差异蛋白质有33个,其中蛋白质表达下调10个,蛋白质表达上调23个;银杏叶提取物处理组与光损伤组差异蛋白质有25个,其中,蛋白质表达上调3个,蛋白质表达下调22个.银杏叶提取物处理组与正常对照组比较,差异蛋白质有11个,其中,蛋白质表达上调4个,蛋白质表达下调7个.差异蛋白质的质谱鉴定和生物信息分析,成功鉴定出16个蛋白,包括组织蛋白酶B、热休克蛋白、细胞色素C还原酶等.结论 光损伤RPE细胞与银杏叶提取物处理后的光损伤RPE细胞及正常RPE细胞间的蛋白质表达有差异;银杏叶提取物光损伤保护作用涉及组织蛋白酶B、热休克蛋白、细胞色素C还原酶等多种蛋白.

Effect of Ginkgo biloba extract on protein expression in light-damaged retinal pigment epithelial cells

Objective To investigate the effect and mechanism of Ginkgo biloba extract EGb761 on protein expression in light-damaged retinal pigment epithelial (RPE) cells. Methods The human RPE cells (ARPE-19) were divided into normal control group, light damage group and EGb761 treatment group;the cells of latter 2 groups were exposed to the cold white light (2200±300)1x] to induce light damage responses. The light-damaged RPE cells were treated with or without EGb761 (100 g/ml). The soluble protein of those cells were extracted and separated by two-dimension electrophoresis and stained by silverstaining. Different proteins in the gel were analyzed by ImageMaster and identified by MALDI-TOF-MS, and were further analyzed by mass spectrometry and bioinformatics. Results ImageMaster and MALDITOF-MS identified 25, 33 and 11 different proteins between light damage group and EGb761 treatment group, between normal control and light damage group, between normal control and EGb761 treatment group of RPE cells respectively. Mass spectrometry and bioinformatics analysis successfully identified 16 proteins, including metabolic enzymes, cytoskeleton proteins, anti-oxidation protein and other types of proteins expressed differentially. Conclusion Protein expression profiles are different between normal control group, light damage group and Ginkgo biloba extract treatment group of RPE cells. The mechanism of protective effect of EGb761 may involve cathepsin B, heat shock protein, cytochrome C reductase, and other proteins.