Extracapsular Lens Extraction Induces Sever Inflammation Response In Proliferative Diabetic Retinopathy Mice Retina Via Stat1 Phosphorylation Pathway And Activation Of M1 Type Microglia

Purpose:To determine whether the effect of cataract surgery on the expression of pro-inflammatory genes and proteins in the retina are different in normal and diabetic groups and to investigate its effect on diabetic retinopathy and postoperative macular edema by establishing experimental non-proliferative and proliferative diabetic retinopathy mice model.Method:Mice were randomly divided into normal,non-proliferative diabetic retinopathy(NPDR)and proliferative diabetic retinopathy(PDR)groups.NPDR and PDR were induced by streptozotocin.An extracapsular len extraction(ECLE)was performed in one eye of C57BL/6 mice in normal(n=46),NPDR(n=46)and PDR groups(n=46);the contralateral un-operated eyes(F eyes,n=138)as well as eyes from un-operated animals(UC eyes,n=72)served as controls.The neurosensory retinas were collected at postoperatively1d,2d and 7d.The expression of the genes(IL-1?,MCP-1,FGF,TGF-?,etc.)involved in the acute inflammatory response was detected using quantitative polymerase chain reaction(RT-PCR).Based on the results,key inflammatory cytokines were further detected by Western blot and/or immunohistochemistry.To test the function of the pro-inflammatory genes and proteins,Evans Blue exudation and related tight junction proteins were also measured.To determine the major source of the most significantly upregulated MCP-1 in our experimental model,MCP-1 was immunofluorescence co-localized with retinal microglial cells and astrocyte cells.The M1 and M2 polarized markers were also stained to clarify the role of microglial cells in the retinal inflammatory response after ECLE surgery in the PDR group.To further investigate the signaling pathways that cause the up-regulation of inflammatory factors,the protein expressions of p-cjun,p-cfos,p-statl,and p-JNK was detected by western blot in the neurosensory retina of normal,NPDR,and PDR mice after ECLE.Besides,BV2 was treated with Lps and Western blot was used to detect the protein expressions of p-cjun,p-cfos,p-statl,and p-JNK.To verify the mechanism of JNK/cjun,cfos and statl pathways regulating MCP-1 in microglia,SP600125,T-5224 and Fludarabine,the specific inhibitor of JNK/cjun,cfos and statl,were preliminarily added before Lps stimulation.Results:ECLE up-regulated the expression of IL-1?,MCP-1,IL-6,FGF and TGF-? in the retinal neurosensory retina of normal,NPDR and PDR mice with a peak at day 1(p<0.05).Compared to the non-diabetic group and the NPDR group,significant difference was found in the relative fold changes of MCP-1 and IL-1? in PDR group(p<0.05)in which the activation of JNK/c-jun,statl was observed significantly to up-regulate MCP-1 after ECLE,causing significant activation of microglia and increased M1 polarization.Greater Evans blue exudation and the down-regulation of the expressions of ZO-1 and Occuldin were also found in PDR group after ECLE.Conclusion:In mice,ECLE elicited an acute inflammatory genes and protein response in normal,NPDR,and PDR groups,and this goes especially worse in PDR group due to the activation of JNK/cjun,statl and the activation of microglia as well as its polarization to M1 type.A similar response in human might explain the pathogenesis of a higher rate of cataract surgery-associated retinal complications such as cystoid macular edema and progression of diabetic retinopathy in proliferative diabetic patients.