高糖下腺苷A_（2A）受体对人视网膜色素上皮细胞血管内皮生长因子表达的影响

目的研究高糖下腺苷A2A受体对人视网膜色素上皮（retinal pigment epithelium,RPE）细胞血管内皮生长因子（vascular endothelial growth factor,VEGF）表达的影响。方法体外培养人RPE细胞,应用免疫组化的方法检测腺苷A2A受体在人RPE细胞中的表达;用腺苷A2A受体作用培养的人RPE细胞,将其分为高糖（33mM）＋腺苷脱氨酶（ADA,2U/ml）、高糖＋ADA＋腺苷A2A受体特异性激动剂（CGS21680,50μM）、高糖＋ADA＋CGS21680＋腺苷A2A受体特异性拮抗剂（SCH58261,100μM）,药物组合刺激RPE细胞24h后,裂解细胞提取RNA,同时收集细胞培养液,分别应用real-time PCR和ELISA的方法检测高糖下腺苷A2A受体对人RPE细胞VEGF mRNA和蛋白的影响。结果腺苷A2A受体在RPE细胞中有表达,在高糖条件下,50μMCGS21680可以刺激RPE细胞VEGF mRNA水平升高约2.7倍,蛋白水平升高1.5倍,而再加100μMSCH58261反而使RPE细胞VEGF mRNA较前下降约 40%,蛋白水平较前下降60%。结论在高糖条件下,腺苷A2A受体可以刺激VEGF的合成,因此腺苷A2A受体可能参与糖尿病性视网膜病变新生血管的形成。

Effect of the A2A adenosine receptor on the expression of vascular endothelial growth factor under high glucose conditions in human retinal pigment epithelium cells

Objective To study the effect of the A2A adenosine receptor on the expression of vascular endothelial growth factor（VEGF） under high glucose conditions in human retinal pigment epithelium（RPE） cells.Methods For cultured human RPE cells：immunohistochemistry was used to detect the A2A adenosine receptor in human RPE cells;cultured human RPE cells were treated with high glucose（33 mM）＋adenosine deaminase（ADA,2 U/ml）,high glucose＋ADA＋adenosine A2A receptor agonist（CGS21680,50 μM）,high glucose ＋ADA ＋CGS21680 ＋adenosine A2A receptor antagonists（SCH58261,100 μM） for 24 h.After treatment,total RNA was extracted and cell culture medium was collected,real-time PCR and ELISA were used to measure the expression of VEGF mRNA and protein in human RPE by adenosine A2A receptor under high glucose conditions.Results A2A adenosine receptor expression as found in cultured RPE.The labeling was largely in the cytosol and membrane,but not in the nucleus.Under high glucose conditions,the adenosine A2A receptor agonist CGS（50 μM） stimulates an increase in the expression of VEGF mRNA（2.7 fold） and protein（1.5 fold）,while for the adenosine A2A receptor antagonist SCH（100 μM） in RPE cells,VEGF mRNA decreased by about 40% and protein levels decreased by about 60%.Conclusion Under high glucose conditions,the adenosine A2A receptor can stimulate the synthesis of VEGF.Therefore the adenosine A2A receptor may be involved in the formation of diabetic retinopathy neovascularization.