氧诱导条件下自发性视网膜色素变性模型rdf小鼠视网膜新生血管相关生长因子的表达和相互作用OA

Objective To investigate the expression and interaction of growth factors associated with retinal neovas-cularization in the rdf mouse model with spontaneous retinitis pigmentosa under oxygen-induced conditions.Methods A total of 42 seven-day-old mice from each of the C57BL/6J and rdf strains were selected.Mice of each strain were randomly divided into a normoxia group and a hyperoxia group,resulting in four experimental groups:C57BL/6J normoxia group,C57BL/6J hyperoxia group,rdf normoxia group,and rdf hyperoxia group,with 18 mice per group.Six mice from each strain(12 mice in total)were reserved for the subsequent isolation of primary retinal microvascular endothelial cells.The retinop-athy model was established under hyperoxic conditions for the mice in the hyperoxia group,while the mice in the normoxia group were reared in a normal atmospheric environment.Retinal structural changes were observed via hematoxylin and eo-sin(HE)staining.Protein expression levels of vascular endothelial growth factor(VEGF),pigment epithelium-derived fac-tor(PEDF),hypoxia-inducible factor(HIF)-1α,and erythropoietin(EPO)in the mouse retinas were detected by Western blot.Co-immunoprecipitation was used to analyze the interaction between PEDF and VEGF proteins in mouse retinal tis-sues.Immunofluorescence staining was performed to analyze the expression of VEGF and PEDF in primary retinal endothe-lial cells isolated from the different mouse strains.Results HE staining revealed well-organized retinal layers without neo-vascular endothelial cells penetrating the inner limiting membrane in mice in the C57BL/6J normoxia group.In contrast,mice in the C57BL/6J hyperoxia group exhibited disordered retinal structure with numerous neovascular endothelial cells penetrating the inner limiting membrane.Mice in both the rdf normoxia and hyperoxia groups maintained relatively well-or-ganized retinal layers overall,with occasional neovascular endothelial cells penetrating the inner limiting membrane in the rdf hyperoxia group.Western blot analysis demonstrated that,under both normoxic and hyperoxic conditions,the protein expression levels of pro-angiogenic factors(VEGF,EPO,and HIF-1α)in retinal tissues of rdf mice were significantly lower than those in C57BL/6J mice.Conversely,the protein expression level of the anti-angiogenic factor PEDF was significantly higher in rdf mice,with all differences being statistically significant(all P<0.05).Co-immunoprecipitation assays confirmed an interaction between the pro-angiogenic factor VEGF and the anti-angiogenic factor PEDF in mouse retinal tissues.Immu-nofluorescence staining further showed co-localization of PEDF and VEGF in primary retinal endothelial cells from both C57BL/6J and rdf mice.The VEGF/PEDF optical density ratios in primary retinal endothelial cells were4.55±1.35 for the C57BL/6J normoxia group,26.39±7.58 for the C57BL/6J hyperoxia group,0.52±0.38 for the rdf normoxia group,and 1.76±0.46 for the rdf hyperoxia group.The differences among all groups were statistically significant(all P<0.05).Conclusion In the retinas of rdf mice with retinitis pigmentosa,the VEGF/PEDF balance shifts significantly towards the anti-angiogenic direction.The low expression of pro-angiogenic factors and the high expression of anti-angiogenic factors in the mouse retina collectively suppress the pathological neovascularization following oxygen induction.