摘要
目的研究蛋白激酶C(protein kinase C,PKC)活化后是否能引起体外培养的兔视网膜色素上皮(retinal pigment epithelium,RPE)细胞内核因子E2相关因子2(nuclear factor enthroid 2-related factor 2,Nrf2)的激活和核转位,探讨PKC活化是否对RPE细胞内的Nrf2表达产生影响。方法将一定浓度的PKC特异激活剂佛波酯(phorbol 12-myristate13-acetate,PMA)作用在体外培养的兔RPE细胞(B组)以及Nrf2抑制剂预处理后的RPE细胞(C组),再培养24 h后,应用免疫荧光技术及Western blot测量Nrf2在胞核内的表达情况,并与空白对照(A)组进行对比分析。结果免疫荧光检测RPE细胞核内的Nrf2蛋白表达结果显示:与A组相比,B、C组RPE细胞核中Nrf2蛋白表达均增加,其中B组增加显著,C组较B组增加幅度低;3组之间差异有统计学意义(P<0.05)。Western blot检测RPE细胞核内的Nrf2蛋白经图像分析处理定量,扫描灰度值差异显著(A组0.286±0.013,B组1.304±0.033,C组0.671±0.087;P<0.05)。结论PKC激活后可使兔RPE细胞浆中Nrf2发生核转位,Nrf2抑制剂能减弱PKC的作用。
Objective To investigate whether activation of protein kinase C （PKC） can induce the activation and nuclear translocation of nuclear factor enthroid 2- related factor 2 （ Nrf2 ） in retinal pigment epithelial （RPE） cells in vitro, and explore whether PKC activation may affect the expression of Nrf2 in RPE cells. Methods PKC-specific activator phorbol ester PMA was used to culture rabbit RPE cells and RPE cells pretreated with Nrf2 inhibitor for 24 hours. Immunofluorescence and Western blot were used to detect Nrf2 in the nucleus of the expression of the situation, the data were obtained for statistical analysis. Results The expression of Nrf2 protein in the nucle- us of PRE ceils was detected by immunofluorescence. Compared with the control group, the expression of Nrf2 protein in the nucleus of RPE ceils increased in the exper- imental group, and the increase of PMA ＋ Nrf2 inhibitor group was lower than that of PMA group. The difference between the two groups was statistically significant （P 〈 0.05）. Western blot analysis showed that the Nrf2 protein in the nucleus of PRE was quantitatively analyzed by image analysis. The gray value of the control group was sig- nificantly different （0.286 ±0.013 in the control group,1. 304 ±0.033 in the PMA group and 0. 671± 0. 087 in the PMA ＋ Nrf2 inhibitor group,P 〈 0. 05）. Conclusion PKC can activate nuclear translocation of Nrf2 in rabbit RPE cytoplasm, and Nrf2 inhibitor can at- tenuate the effect of PKC.
出处
《眼科新进展》
CAS
北大核心
2017年第5期419-422,共4页
Recent Advances in Ophthalmology
基金
黑龙江省卫生计生委科研课题(编号:2016-379)
牡丹江医学院科学技术研究项目(编号:ZSSD201619)
牡丹江医学院2014年度研究生创新科研资金资助(编号:2014YJSCX-11)~~
关键词
蛋白激酶C
佛波酯(PMA)
核因子E2相关因子2
核转位
protein kinase C
phorbol 12-myristate 13-acetate
nuclear factor enthroid 2-related factor 2
nuclear translocation
