Effects Of Changes Of TNF-a/VEGF And BFGF Secreted By Retinal Microglia On Retinal Microvessel Degeneration

Objectives:To investigate the effect of TNF-a/VEGF and b FGF secreted by retinal microglia on retinal microvessel degeneration,understanding the pathogenesis of diabetic retinopathy in terms of cellular and molecular aspects,to provide support for the prevention and treatment of new.Methods:1.Animal experiment:(1)the healthy SD rats were randomly divided into five groups:normal control group,DM1 month group,DM2 month group,DM3 month group and DM4 group,and DM group was injected with STZ to establish type 1 diabetic rat mod-el.(2)To detecte the dynamic changes of TNF-a,VEGF,bFGF,IL-1? and IL-6 by imm-unohistochemistry,QPCR and western blotting.(3)To detecte the permeability of reti-nal vascular by Evans blue leakage experiment.Cell experiments:(1)cultured rat retinal microglia,etinal pigment epithelial cells,retinal capillary endothelial cells,to identificate them by immunofluorescence staining.(2)co-cultured retinal microglia,retinal pigment epithelial cells,retinal capillary endothelial cells,according to the medium with the concentration of glucose and LPS were divided into six groups,groupA:5mM glucose,group B:10mMglucose,group C:20mM glucose,group D:30mM glucose,group E:5mM glucose+100ug/L LPS,group F:30mM glucose+100ug/L LPS,culture 72h.(3)To detecte the changes of TNF-a,VEGF,bFGF,IL-1? and IL-6 by western blotting and immunofluorescence staining.Results:1.animal experiments:(1)EB leakage experiment results show that:with the extension of the course,the content of EB of retina in diabetic rats compared with normal control group gradually increased,the retinal vascular permeability increased(P<0.05)(2)QPCR and western blotting results show that:with the extension of the course,TNF-a,VEGF,bFGF,IL-1? and IL-6 were increased gradually(P<0.05),The expression of four time points were increased compared with the normal control group(P<0.05).2.Cell experiment:(1)Western blotting results show that:with the increase of glucose concentratiorn,TNF-a,VEGF,bFGF,IL-1? and IL-6 were increased gradually(P<0.05),the expression of 30mM+LPS group was higher than that of 30mM group(P<0.05),the expression of 5mM + LPS group was higher than that of 5mM group(P<0.05).(2)Immunofluorescence staining show that:TNF-a,VEGF,bFGF,IL-1?,IL-6 staining in the cytoplasm of the cells,30mM group was higher than 5mM group,30mM+LPS group was significantly higher than 30mM group and 5mM group.Conclusions:1.The blood retinal barrier function of diabetic rats has been damaged in the early stage of the disease,and its severity increased with the progression of the disease.2.TNF-?,VEGF,bFGF,IL-1? and IL-6 are involved in the pathogenesis of diabetic retinopathy.3.High glucose and lipopolysaccharide stimulate the activation of microglia to prod-uce TNF-?,IL-1?,IL-6 and other proinflammatory factors.4.Activated microglia secrete TNF-?,IL-1?,IL-6 and other inflammatory factors,up-regulated the expression of VEGF and bFGF,destruction the blood retinal barrier function.