Integrated analysis of RNA-binding proteins in MAFLD: mechanistic insights and diagnostic biomarker potential via alternative splicing.

Metabolic associated fatty liver disease (MAFLD) is a highly prevalent global chronic liver disease. While abnormal expression of RNA-binding proteins (RBPs) has been implicated in MAFLD, their functional roles-particularly in regulating alternative splicing (AS)-remain poorly characterized. This study aimed to investigate the abnormal expression and regulatory mechanism of RBPs in MAFLD. The source data were obtained from the GSE130970 dataset of the Gene Expression Omnibus (GEO) database. Then, we utilized differential expression analysis to acquire the differentially expressed genes (DEGs) between different stages of MAFLD patients and normal patients. Alternative splicing analysis was performed via the ABLas pipeline to explore the alternative splicing events that may enhance and regulate the development of MAFLD. The Gene Ontology (GO) terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were utilized via KOBAS2.0. Finally, quantitative real-time Polymerase Chain Reaction (RT-qPCR) and Western blotting analysis were performed to confirm the expression of significant RBPs. We observed a significant increase in the number of DEGs as the stages of MAFLD progressed. Furthermore, co-expression analysis suggested that abnormally expressed RBPs such as S100A4, CYCS, and JUN might participate in MAFLD development by potentially influencing the AS of downstream metabolism-related genes such as FAT1, SLCO2B1, and C4BPB. Moreover, we confirmed that the expression of the three RBPs (S100A4, CYCS and JUN) is significantly up-regulated in the liver through validation experiments. The abnormal up-regulated expression of RBPs (S100A4, CYCS and JUN) might contribute to the progression of MAFLD and hence they can be further regarded as potential therapeutic targets for MAFLD.