Influence Of Discoidin Domain Receptor2on HIF-1Alpha And VEGF Expression Which In RPE Cells Under Hypoxic Condition

Background DDR2is one type of receptor tyrosine kinase.It gets involvedin migration, adhesion, differentiation and transfer of a variety of cells byinteracting with extracellular matrix, and plays an important role in thedevelopment and progress of tumors and autoimmune disease. After activatedby collagen, DDR2could induce receptors to auto-phosphorylate, thensubsequently mediate numerous signal transduction pathways and regulatecells’ biological behaviors. CNV, one of the most important manifestation ofintraocular neovascularization, is associated with many diseases, is one of themain reasons for the blind, at present there is no effective treatment methods.At present about whether DDR2participate in the choroidalneovascularization development is not clear,this experiment is discussingabout the role of DDR2in CNV.Purposes To observe the effect of DDR2on laser induced mice CNVgeneration. The expression of DDR2in hypoxia state of RPE cells and the influence of the expression of actived or suppression HIF-1α and VEGF,the effect of Actived or inhibition DDR2on actived HIF-1a and expression ofVEGF, discussing the effects of DDR2in CNV.Methods It was an experimental study, using200μmol/L CoCl２treated RPEcells to establish chemical hypoxia model, expression of DDR2of cells wereexamed after hypoxia0,2,6,12,24h by immunofluorescence,RT-PCR,Q-PCR and Western blot.In hypoxiaed state with type I collagen’s(10μg／ml in50degrees incubated box) stimulation, the expression ofDDR2in cells were examed after hypoxia0,2,6,12,24h byimmunofluorescence, RT-PCR, Q-PCR and Western blot, using RT-PCR，ELISA to observe the expression of VEGF in cell culture supematant. Inhypoxia state importing siRNA into RPE cells,expression of HIF-1α of cellswere examed after hypoxia0,2,6,12,24h byRT-PCR,Q-PCR and Westernblot,expression of VEGF were examed after hypoxia0,2,6,12,24h byRT-PCR and ELISA.Results Normal RPE cells’ nuclei, cytoplasm and cell membrane had a faintDDR2fluorescence expression. After prolonged hypoxia, DDR2’ fluorescentexpression was reduced, protein and mRNA expression was reduced. Afterprolonged hypoxia, the expression of RPE cells’ HIF-1α and VEGF increasedwith hypoxia time, In hypoxiaed state with type I collagen’s stimulation, theexpression of HIF-1α, VEGF did not significantly increase. In hypoxia statewith type I collagen’s stimulation can surpress hypoxia induced upregulationexpression of HIF-1α and VEGF in RPE cell. In hypoxia state importingsiRNA into RPE cells,expression of VEGF and HIF-1α of cells wereincreasing.Conclusions DDR2had the effect of inhibiting increasing of HIF-1α and VEGF in hypoxiaed state.