摘要
Abstract
·AIM:To investigate the role of the ABCA4 gene in the pathogenesis of dry age-related macular degeneration(ARMD)through bioinformatics and cellular experiments.
·METHODS:Integrating bioinformatics and cellular experiments,differential expression genes(DEGs)associated with ARMD were screened based on the GEO dataset GSE29801.Hub genes were identified through PPI network analysis(using the STRING database)and topological parameter screening.A lentivirus-mediated stable ABCA4 knockdown cell line in human retinal pigment epithelial cells(ARPE-19;ABCA4-KD group)and a negative control were constructed.An oxidative stress model was established using NaIO3,and the cells were divided into four treatment groups:NC group(negative control lentivirus+PBS),NaIO3+NC group(negative control lentivirus+oxidative stress injury),ABCA4-KD group(ABCA4 knockdown lentivirus+PBS),NaIO3+ABCA4-KD group(ABCA4 knockdown lentivirus+oxidative stress injury).Knockdown efficiency was verified via Western blot,cell viability was assessed using the CCK-8 assay,and the apoptosis rate was measured by Hoechst 33342 and Annexin V-FITC/PI double staining.
·RESULTS:Bioinformatics analysis identified 5 069 DEGs(2 493 upregulated/2 576 downregulated),of which 118 key genes were obtained by intersecting with ARMD disease targets.PPI network analysis identified the top 5 hub genes(ABCA4,RPE65,PRPH2,RHO,PDE6B),with ABCA4 showing the highest degree centrality(Degree=58).ROC curve analysis demonstrated that ABCA4 had excellent discriminative efficacy for ARMD(AUC=0.986).Further cellular experiments revealed that ABCA4 protein expression in the ABCA4-KD group was significantly lower than that in the NC group(P<0.05).Under oxidative stress conditions,the NaIO3+ABCA4-KD group exhibited lower cell viability compared to the NaIO3+NC group(P<0.01),while the apoptosis rate was significantly increased(all P<0.01).In the absence of oxidative stress,knockdown of ABCA4 alone did not affect cell survival(P>0.05).
·CONCLUSION:Loss of ABCA4 function contributes to the pathology of ARMD by exacerbating oxidative stress-induced RPE cell apoptosis,and is expected to serve as a novel therapeutic target for dry ARMD.关键词
年龄相关性黄斑变性/ABCA4/氧化应激/凋亡Key words
age-related macular degeneration/ABCA4/oxidative stress/apoptosis
