金丝桃素对培养的人视网膜色素上皮细胞钙离子内流的抑制(英文)

Inhibition of hypericin to Ca^(2+) influx in cultured human retinal pigment epithelium

目的 研究蛋白激酶 C(PKC)抑制剂金丝桃素对视网膜色素上皮细胞 (RPE cells)钙离子信号共聚成像的影响 .方法 使用钙荧光指示剂 fluo- 3AM和激光扫描共聚焦显微镜 (L SCM) ,分析在 10 0 nmol·L- 1佛波酯 (PMA)和 (或 )5个浓度的金丝桃素 (1,2 ,3,4和 5μmol· L- 1 )刺激时培养的人 RPE细胞的钙离子信号的变化 .结果 RPE细胞荧光着色很强 ,遍布整个细胞 ,其中细胞核比细胞质更强 .只用PMA或金丝桃素刺激时 ,可以观察到荧光强度迅速下降 ,并且 5个浓度金丝桃素之间的下降曲线无明显差异 .用 PMA预先处理细胞 2 4h后 ,再给予金丝桃素刺激发现荧光强度并没有进一步下降 .结论 fluo- 3AM是人 RPE细胞钙离子良好的指示剂 ,金丝桃素能强力抑制钙离子内流 ,它作为治疗增殖性玻璃体视网膜病变的潜在药物可能是因为能抑制 PKC活性和钙离子内流 .

AIM To investigate the mechanism of the Ca 2+ signaling in cultured human retinal pigment epithelial (RPE) cells with the protein kinase C (PKC) specific inhibitor hypericin stimulation. METHODS Cultured human RPE cells were analyzed using the fluorescence Ca 2+ dye fluo 3 AM and laser scanning confocal microscope (LSCM) after stimulation with 100 nmol·L -1 phorbol 12 myristate 13 acetate (PMA ) and (or) 5 concentrations of hypericin (1,2,3,4 and 5 μmol· L -1 ). RESULTS Normal fluorescence in RPE cells was strong and distributed throughout the cells. The nucleus appeared to be more fluorescent than the cytoplasm. After stimulation with PMA alone or 5 concentrations of hypericin, a rapid decrease in fluorescence intensity was observed. There was no obvious difference in decreased curve among 5 concentrations. However, after stimulation with a 24 h preincubation of PMA and 5 concentrations of hypericin, no further decrease was observed. CONCLUSION Fluo 3 AM appears to be a good indicator of the change in Ca 2+ occurring in RPE cells and hypericin is a strong inhibitor of Ca 2+ influx channel. hypericin has potential as a therapeutic drug for proliferative vitreoretinopathy (PVR), the inhibitory effect on PVR might be caused by blocking the PKC activity and inhibiting Ca 2+ influx pathway.