目的 探讨视网膜色素上皮细胞(RPE)的钙离子与整合素基因表达的相关性.方法 人RPE细胞经分离培养,免疫细胞化学鉴定后,传至第4~6代;对各组RPE细胞进行5 μmol/L Fluo-3负载30 min,姜黄素分别以0.9375、1.8750、3.7500、7.5000、15.0000 mg/L浓度干预人RPE细胞,利用激光扫描共聚焦显微镜(LSCM)测定各组RPE细胞内早期钙离子的变化.利用MTT法测定不同浓度的姜黄素干预人RPE细胞48 h后的IC50,并观察相应的形态学变化.用Real-Time PCR测定姜黄素干预后的整合素基因的表达.结果 LSCM检测结果:空白对照组、0.9375、1.8750、3.7500、7.5000、15.0000 mg/L姜黄素组RPE细胞内均有钙荧光.空白对照组钙荧光强度明显低于姜黄素各组,且差异有统计学意义(P＜0.05),对照组基因表达量与整合素α4和整合素β5呈正效应,与整合素α3和整合素α5呈负效应.结论 体外培养的人RPE细胞经不同浓度姜黄素干预后,较空白对照组早期细胞内钙离子荧光强度明显增加,且有显著性差异.当细胞受到姜黄素外界信号刺激时,细胞内钙离子浓度能迅速增加,继而调节整合素的活性,继而引起细胞的生物学效应.因此,姜黄素通过调节细胞内的信号传导,可能成为治疗人眼疾,如增殖性玻璃体视网膜病变的一种有效物质.%Objective To explore the association between calcium signal of retinal pigment epithelial cell ( RPE ) and the integrin responses. Methods The RPE cells were separated and cultured to the 4 ~ 6 generations after determination by im-munocytochemistry. RPE cells were treated by Fluo -3(5 μmol/L ) for 30 min. Each group of RPE cells was affected with curcumin of 0. 9375 mg/L, 1. 8750 mg/L, 3. 7500 mg/L, 7. 5000 mg/L, and 15. 000 mg/L, respectively. Intracellular calcium changes of RPE cells were measured by laser scanning confocal microscope ( LSCM ). The RPE cells'IC50 after 48 h incubation in each group was recorded by MTT and related morphology changes were observed. The integrin gene expression was measured by Real - time PCR. Results LSCM results showed calcium fluorescence in RPE cells in the control group and all curcumin groups. The calcium fluorescence in the curcumin groups was significantly greater than that in the control group ( P < 0. 05 ). The gene expression level was positively correlated with integrin α4 and β5 , and negatively correlated with integrin α3 and α5. Conclusion The intracellular calcium fluorescence of curcumin affected human RPE cells is stronger than that of those not affected. The intracellular calcium immediately increases when stimulated by external curcumin, which can regulate integrin gene expression and lead to biological effects. Therefore, curcumin might be a therapeutic agent to modulate the cell signal transduction for certain human ocular disorders such as PVR.
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