The Role Of ASK1 Regulating VEGF Signalling Pathway In The Pathogenesis Of Diabetic Retinopathy And Its Intervention

Part I The role of ASK1 regulating VEGF signalling pathway in the pathogenesis of diabetic retinopathyObjective:To investigate the role of the ASK1 signalling pathway in high glucose-exposed human retinal microvascular endothelial cells(HRMECs),explore the regulatory mechanism of VEGF signalling pathway under high glucose conditions.Method:HRMECs were incubated with Low glucose(5mM)or high glucose(30mM)with or without ASK1 shRNA or NQDI-1(100nM)or MCC950(50nM).T-ASK1、p-ASKl、p-p38、T-p38、VEGF、Caspase-1 and Clevaed-Caspase-3 in the cells of each group were examined by Western blotting,and NLRP3、p53、IL-1β and IL-18 by Real-time PCR and Western blotting.The TUNEL method and MTT assay was performed to detect the apoptotic cells and cell viability.Graphpad Prism 5.0 soft ware was used for statistical analysis.Results:Compared with the control group,high glucose-induced p-ASK1and p-p38 protein expression in HRMECs was significantly increased(P<0.05),and NLRP3 mRNA and protein expression in high glucose incubated group were significantly higher than low glucose incubated group(P<0.05),while the ASK1 gene silencing and NQDI-1 intervention significantly inhibited these changes(P<0.05);IL-1β、IL-18、VEGF、p53、Caspase-1 and Clevaed-Caspase-3 protein expression in high glucose-incubated HRMECs were significantly increased(P<0.05)than the low glucose incubated group,and MCC950 intervention significantly inhibited increased protein expression of IL-1β、IL-18、VEGF、p53、Caspase-1 and Clevaed-Caspase-3 and mRNA expression of P53、IL-1β and IL-18mRNA;Compared with the low glucose incubated group,high glucose-induced apoptosis in HRMECs was significantly increased(P<0.01)and cell viability decreased(P<0.05),while the ASK1 gene silencing and NQDI-1 intervention significantly inhibited these changes(P<0.05),and inhibited increased protein expression of IL-1β、IL-18、VEGF、p53、Caspase-1 and Cleaved-Caspase-3 and mRNA expression of P53、IL-1β and IL-18mRNA(P<0.05).GraphPad Prism 5.0 software was used for statistical analysis.Conclusions:ASK1/p38MAPK is activated in high glucose incubated HRMECs,then induce NLRP3 inflammasome and VEGF activation,which in turn triggers inflammation and apoptotic signaling through induce NLRP3 inflammasome activation.High glucose-induced apoptosis and inflammation was alleviated by inhibition of Askl,block Askl by shRNA or target specific inhibitor NQDI-1 respectively.ASK1 may be a new potential target for the prevention and treatment of diabetic retinopathy.Part Ⅱ Comparison of anti-VEGF drugs with modified-ETDRS laser photocoagulation for diabetic macular edemaObjective:To evaluate the efficacy of intravitreal conbercept(IVC)injection versus intravitreal ranibizumab(IVR)injection,and compared with modified-ETDRS laser photocoagulation(MEP)alone for treatment of diabeticmacular edema(DME).Method:24 patients(24 eyes)with DME were randomly(1:1:1)assigned to receive IVC,IVR or MEP.Best corrected visual acuity(BCVA),optical coherence tomography(OCT)and fundus photography were obtained at baseline and each month after treated with 12 months of follow-up.Results:There was no significant difference in the mean age,sex,duraion of diabetes,fasting blood-glucose,glycosylated hemoglobin,type of DME,BCVA and CRT among three groups before treatment(.P>0.05).At last visit,6 patients(6 eyes)were treated with I VC(IVC group),7 patients(7 eyes)with IVR(IVR group),and 8 patients(8 eyes)with MEP(MEP group).At month 12,the mean BCVA letter score improvement was 9.50±1.75 and 7.29±1.77 and the mean central retinal thickness(CRT)reduction was 220.17±48.95 um and 226.29±37.44um in the IVC group and the IVR group,respectively.The BCVA letter score and CRT were significantly improved 12 months after injection in comparison with before injection in the IVC group and the IVR group(P<0.05).The BCVA letter score improvement was 4.00±2.65 and the mean CRT reduction was 59.38±43.972 um in the MEP group at 12th month,but have no difference with before injection(P>0.05).There was no statistically significant difference of improvement in BCVA(P>0.05)among the three groups at each visit,and evident difference was found in CRT(P<0.05)among the three groups except the postoperative 1 month visit.However,no significant difference was found between IVC group and IVR group for BCVA,CRT and intravitreal times(P>0.05).Conclusions:IVR or IVC appeared to be more effective for treatment of DME to reduce CRT than MEP alone.IVR or IVC remarkably ameliorate the macular edema and improve the visual acuity,achieving the similar clinical efficacy.Intravitreal injection of anti VEGF drugs is currently the most effective treatment for DME.