miR-182在多氯联苯(PCB_(1254))暴露后视网膜神经节细胞内的表达及意义

Expression and effects of mi R-182 in retinal ganglion cells exposed to PCB_(1254)

目的:研究多氯联苯(polychlorinated biphenyls,PCBs)PCB1254引起视网膜神经节细胞毒性作用的可能机制。方法:分别配制浓度为0.125、0.250、0.500、1.000 mg/L的PCB1254对RGC-5细胞进行暴露处理,并设立空白对照及0.01%甲醇对照组,测定细胞内miR-182表达水平;利用miR-182类似物或抑制剂转染RGC-5细胞,使miR-182上调或下调,分别检测细胞凋亡、细胞增殖功能和细胞周期;使用0.5、1.0 mg/L的PCB1254刺激RGC-5细胞,再转染miR-182类似物,分别检测细胞凋亡、细胞增殖功能。结果:随着PCB1254浓度的增高,RGC-5细胞内miR-182的表达逐渐被抑制,当浓度≥0.5 mg/L时,与对照组的差异有统计学意义(P<0.05)。在RGC-5细胞内沉默miR-182可导致Caspase-3活性增高(P<0.05),表明miR-182沉默促进细胞凋亡;CCK-8法显示miR-182沉默会导致RGC-5细胞活力降低(P<0.05),表明miR-182沉默会抑制细胞增殖;Mi R-182沉默对RGC-5细胞周期无显著影响(P>0.05)。使用miR-182类似物干预0.5、1.0 mg/L PCB1254暴露后的RGC-5细胞,发现细胞内Caspase-3活性下降(P<0.05)、细胞活力增加(P<0.05),表明miR-182可改善PCB1254对RGC-5细胞凋亡及增殖功能的影响。结论:PCB1254可能通过抑制miR-182的表达来影响RGC-5细胞的增殖和凋亡,进而导致视功能损害。

Objective： To study the mechanism of toxic effect of polychlorinated biphenyls PCB1254 on retinal ganglion ceils （RGC- 5）. Methods： After RGC-5 cells were exposed to 0.125, 0.250, 0.500, 1.000 mg/L PCB 1254 , 0.01% methanol, and pure water, the level of miR-182 was detected in every group. In addition, by using the miR-182 mimics or miR-182 inhibitor to up-regulate or down- regulate the expression of miR-182 in RGC-5, apoptosis, proliferation, and cell cycle were observed. After the treatment with 0.5 mg/ L and 1.0 mg/L PCB1254 , miR-182 mimics were transfected into RGC-5 cells to observe the apoptosis and proliferation. Results： （1） With the increasing concentration of PCB1254 , the expression of miR-182 was declined. When the concentration was ≥0.5 mg/L, there were significant differences between experimental groups and the control group （P〈0.05）. （2） MiR-182 silencing led to increased Caspase-3 activity in RGC-5 cells, which indicated that miR-182 silencing promoted apoptosis （P〈0.05）. CCK-8 assay showed that the cell viability was lower in miR-182 inhibitor group than that in the negative control group, which indicated that miR-182 silencing inhibited cell proliferation （P〈0.05）. There were no significant differences in cell cycle between the negative control group and the miR-182 inhibitor group （P〉0.05）. （3） When miR-182 were tranfected in RGC-5 exposed to 0.5 and 1.0 mg/L PCB1254 , the caspase-3 activity deereased（P〈0.05）, and cell viability increased （P〈0.05）. It showed that miR-182 could improve the toxic effect of PCB1254 in cell apoptosis and proliferation. Conclusion： Our findings suggest that the PCB1254 may affect the proliferation and apoptosis of RGC-5 by inhibiting the expression of miR-182, which may lead to the damage of visual function.