玻璃体条件培养液对视网膜色素上皮细胞P-ERK的影响

目的:探讨人玻璃体条件培养液(vitreous-conditioned medium,VCM)对体外培养的人视网膜色素上皮(human retinal pigment epithelium,HRPE)细胞内磷酸化的细胞外调节蛋白激酶(phosphorylated-extracellular signal-regulated kinase,P-ERK)的影响。方法:将HRPE细胞培养在含100,250,500,1000mL/L人玻璃体的DMEM条件培养液(VCM)里,用免疫荧光细胞化学技术,Western blot观察VCM作用不同时间及不同体积比的VCM作用下P-ERK的表达和分布情况。结果:VCM促使RPE细胞内ERK发生了磷酸化,且P-ERK发生了由细胞质到细胞核的转位。ERK的磷酸化水平在一定范围内与VCM的玻璃体体积成正比,250mL/L的VCM刺激ERK磷酸化的效果最强。ERK在加入VCM后5min开始磷酸化,10min磷酸化水平达到峰值且持续约2h。结论:一定体积分数的VCM使RPE内的ERK发生了磷酸化,激活后的P-ERKs由细胞质进入细胞核,参与转录的调节。提示MEK/MAPK信号传导途径可能参与了增生性玻璃体视网膜病变(proliferative vitreoretinopthy,PVR)。

Effect of human vitreous-conditioned medium on phosphorylated-extracellular signal-regulated kinase in human retinal pigment epithelium

AIM:To investigate the role of human vitreous-conditioned medium(VCM)on phosphorylated-extracellu-lar signal-regulated kinase (P-ERK) in human retinal pigment epithelium (HRPE) cells.·METHODS: HRPE cells were treated with VCM including 100,250,500 and 1 000mL/L human vitreous respectively. After treatment with different time or volume of human VCM,immunofluorescence staining and western blot were used to observe the expression level and distribution of P-ERK in HRPE cells.·RESULTS: ERK was activated after VCM treatment and relocated from cytoplasm to nucleus. Human VCM stimulated P-ERK in HRPE cells in a concentration-dependent manner,and the optimal concentration of VCM was 250mL/L. ERK was activated after 5 minutes,reaching maximal phosphorylated level at 10 minutes and lasting for 2 hours.·CONCLUSION: Some concentration of VCM can activate ERK and induce activated-ERK nuclear translocation,which play an important role in regulation gene transcrip-tion. Such a conclusion prompts MEK/MAPK signal cascade may participate in the pathoprocess of proliferative vitreo retinopathy (PVR).