Influence Of Ad-slit2shRNA On VEGF Expression Of Hypoxia-inducible Human Retinal Pigment Epithelial Cells

Objective:Observed the impact of the hypoxia-inducible human retinal pigmentepithelial cells(RPE) which transfected by adenovirus-mediated slit2shRNA to the vascular endothelial growth factor(VEGF) expression,Toexplored the possible role of slit2in choroidal neovascularization, andprovide new ideas for the treatment of choroidal neovascularization(CNV).Method：Human ARPE-19cells were cultured in vitro, using RT-PCR andimmunohistoch-emistry to identify the expression of the slit2and robo1inhuman PRE.2. The chemical hypoxia model was established by200umol/lcobalt chloride to unhypoxia RPE cells as a control group. usingReal-TimePCR and Western-blot to observe the changes of slit2, Robo1and VEGF’s expression in hypoxic state.3.the hypoxia RPE cells wererandomly divided into Ad-shRNA group(add slit2shRNA), Ad-null group(add empty adenovirus group) and hypoxia group. The cells werereceived after24hours. using Real-TimePCR and Western-blot to detectethe levels of the mRNA and protein which expressed by slit2, Robo1andVEGF, using ELISA to detevte the expression changes of VGEF’s proteinin clear supernatant liquid.Result：1.The expression of slit2and Robo1is reside in normal human’s RPEcells.2.Compared with the control group,the levels of the mRNA andprotein which expressed by slit2, Robo1and VEGF was significantlyincreased (P<0.05) in the hypoxia group.3.Compared with the hypoxiagroup, the levels of the mRNA and protein which expressed by slit2decreased in Ad-shRNA group, the expression of robo1was reduced too.The expression of VEGF also reduced with the expression slit2-robo1(P<0.05).4.The changes of the slit2,Robo1and VEGF hadn’t statisticallysignificant between hypoxia group and Ad-null group.Conclusion：When slit2shRNA is silencing the slit2in RPE, it can significantlyinhibit the expression of VEGF in hypoxia-induced RPE. That can providea new direction for the treatment of CNV in clinical.