地塞米松加强体外培养的大鼠视网膜血管内皮细胞间的紧密连接

Dexamethasone intensify the tight junctions in rat retinal vascular endothelial cells in vitro

目的 探讨地塞米松对体外培养的大鼠视网膜血管内皮细胞间紧密连接蛋白表达、分布及其功能的影响。方法首先使用CD31抗体包被的免疫磁珠对体外培养的大鼠视网膜血管内皮细胞进行分离培养及鉴定。然后取P4代细胞，加入含有地塞米松的培养液对细胞进行处理，并设立对照组，通过检测跨细胞电阻、细胞间紧密连接蛋白免疫荧光染色和RT-PCR方法，从地塞米松对大鼠视网膜血管内皮细胞紧密连接的功能、蛋白分布以及蛋白mRNA表达水平几方面的影响进行研究。结果细胞经鉴定后证实为大鼠视网膜血管内皮细胞。分组处理2d后，地塞米松组电阻值与对照组电阻值之间差异有统计学意义（P〈0．05）。通过对紧密连接蛋白claudin-1的免疫荧光染色发现地塞米松组较对照组细胞间紧密连接蛋白分布向胞质的外周聚集。RT-PCR证实地塞米松组细胞间紧密连接蛋白claudin-1mRNA表达水平，均对照组细胞升高。结论地塞米松可以增加大鼠视网膜血管内皮细胞间紧密连接蛋白claudin-1的表达，并促进其聚集于细胞外周，并增强细胞间紧密连接的密封性。从而推测，糖皮质激素治疗黄斑水肿的药物作用机制可能与其可以加强视网膜血管内皮细胞间紧密连接有关。（中华腰群系右，2007，43：646-650）

Objective To investigate the effect of dexamethasone on the expression, distribution and function of tight junctions in rat retinal vascular endothelial cells in vitro. Try to explain the mechanism of glucocorticoid in the treatment of macular edema from a new viewpoint. Methods Rat retinal vascular endothelial cells were isolated and purified by magnetic beads coated with anti-CD31. Cells were identified by vW factor indirect immunofluorescence staining. The fourth-passage cells were used to investigate the effect of dexamethasone on the tight junctions in rat retinal vascular endothelial cells. Cells were separated into two groups, one was treated with 500 nmol/L dexamethasone and the other was used as the control. Transepithelial electrical resistance （TER） was measured to estimate the changes in the treated group. Indirect immunofluorescent stain and RT-PCR were used to observe the difference of tight junction protein distribution and mRNA expression level between these two groups. Results Rat retinal vascular endothelial cell monolayer showed positive immunofluorescent staining for vW factor. The dexamethasone treated group showed greater TER than that of the control （P 〈 0.01 ）. Tight junction protein in the dexamethasone treated group localized closer to the borders of retinal vascular endothelial cells than that of the control. Claudin-1 mRNA level of the dexamethasone treated cells were greater than that of the control. Conclusions Dexamethasone intensifies the tight junctions in rat retinal vascular endothelial cells. Therefore this is one of the mechanisms of treatment of macular edema by glucocorticoid. （Chin J Ophtbalmol, 2007,43：646- 650）