摘要
Objective To identify and validate the key genes of ferroptosis in phospholipase A2 receptor(PLA2R)associated membranous nephropathy through bioinformatics analysis and in vitro experiments,and to explore the potential roleof ferroptosis in PLA2RRassociated membranous nephropathy(PMN).Methods The GSE115857 dataset obtained by retrieving the Gene Expression Omnibus(GEO)database and the ferroptosisrelated genes obtained by retrieving the FerDb database were intersected.The intersected genes were subjected to Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analysis.The key ferroptosis genes associated with PMN were identified by intersectinggenes sseelected1usingSsupport vector machines-recursive feature elimination and least absolute shrinkage and selection operator regression.The results were validate by real-time PCR,cell counting kit-8,Western blotting and immunofluorescence in human renal podocyte line AB 8/13 from both the control group and model group.Results A total of 25 genes related to ferroptosis of PMN were obtained,and GO and KEGG analysis showed that these genes were mainly involved in cell ferroptosis metabolism.The key ferroptosis genes of PMN obtained by machine learning method were activating transcription factor 3(ATF3)and coiled coil domain containing 6(CCDC6).The results of in vitro experiments showed that the human renal podocyte line AB 8/13 in the model group was significantly deformed and retracted compared with the control group.The surface area density of foot processes was significantly reduced,and the podocyte cytoskeleton was allosteric.The morphology of F-actin was disordered and the expression of synaptopodin was decreased.The cell proliferation activity was significantly decreased(P<0.05).The expression of PLA2R protein was increased(P<0.05),and the expression of GPX4 protein was decreased(P<0.01).The protein and mRNA levels of ATF3 and CCDC6 were significantly up-regulated(all P<0.05).Conclusion Ferroptosis may be one of the key mechanisms in the occurrence and development of PMN.In vitro experiments show that ATF3 and CCDC6 are the key genes in the ferroptosis of PMN podocytes,whichprovidesnew insightsand ideas for the pathogenesis of PMN.
