Regulator Of G-protein Signaling 1 Promotes Choroidal Neovascularization In Age-related Macular Degeneration

Background: Age-related macular degeneration(AMD)is one of the common eye diseases closely related to age factors,also known as age-related macular degeneration.choroidal neovascularization(CNV)can cause severe visual impairment in people over 60 years old,which is the main pathological manifestation of wet AMD.The aim of this study is to investigate the effect of regulator of G protein signaling 1(RGS1)on krypton ion laserinduced CNV in mice and human microvascular endothelial cells under hypoxic conditions.HMECs)produce and release vascular endothelial growth factor(VEGF).Methods: 1.Male C57BL/6 mice aged 6-8 weeks were used in the experiment.The PASCAL diode ophthalmic laser system(Topcon,Livermore,CA,Bruch’s membrane(a5-layer structure composed of elastic fibers and collagen fibers between the retinal pigment epithelium basement membrane and the choroid capillary basement membrane)was broken down at 3,6,9 and 12 o ’clock around the optic disc in mice to establish a mouse model of CNV.Mice were randomly divided into three groups: normal group,CNV 7-day group and CNV 14-day group.On day 7,pathologic vascular leakage was detected with the use of fundus angiography(FFA).Then the mice in each group were sacrificed,and their eyeballs were collected.Hematoxylin-eosin(HE)staining was used to make pathological sections,and the location of lesions in the CNV group was analyzed histopathologically.Using quantitative real-time PCR(q RT-PCR),q RT-PCR was used to detect the m RNA expression levels of regulator of G protein signaling 1(RGS1),hypoxia-inducible factor-1(HIF-1α)and vascular endothelial growth factor(VEGF)in the retinal pigment epithelium-choroid complex.Western Blot was used to detect the protein expression levels of regulator of G protein signaling1(RGS1),hypoxia-inducible factor-1(HIF-1α)and vascular endothelial growth factor(VEGF)in the retinal pigment epithelium-choroid complex.2.Human microvascular endothelial cell lines(HMECs)were cultured in complete medium to establish an in vitro hypoxia model of human microvascular endothelial cells.The cells were divided into four groups according to whether they were cultured under hypoxic conditions or not: normal group,hypoxia group,low RGS1 expression group and low RGS1 expression hypoxia group.The m RNA expression levels of RGS1,HIF-1α and VEGF were detected by real-time PCR.The protein expression levels of RGS1,HIF-1α and VEGF in HMECs were detected by Western blot.The proliferation ability of HMECs was detected using Ed U assay.Results: 1.RGS1 is an up-regulated gene in AMD,and its functional enrichment is closely related to inflammation and neovascularization.2.In the krypton ion laser-induced CNV model,the m RNA and protein expression levels of RGS1,HIF-1α and VEGF in the retinal pigment epithelium-choroid complex were significantly increased in the CNV group.3.In vitro,compared with the normal group,the m RNA and protein expression levels of RGS1,HIF-1αand VEGF in HMECs in hypoxia treatment group were significantly higher.When RGS1 was knocked down,the m RNA and protein expression levels of HIF-1α and VEGF were effectively inhibited.4.The proliferation ability of HMECs was significantly reduced after RGS1 knockdown.Conclusions: In age-related macular degeneration(AMD),the expression of RGS1 is increased,which promotes the proliferation of human microvascular cells through the HIF-1α/VEGF signaling pathway,and then promotes the formation of choroidal neovascularization(CNV).This suggests that reducing the expression level of RGS1 may be a potential new drug for the treatment of CNV by inhibiting neovascularization.