Supplementary Material for: Identification and verification of B4GALNT2 as an epigenetic marker in ulcerative colitis

Background: Ulcerative colitis represents an inflammatory bowel disease characterized with a multifaceted pathogenesis, which may be attributed to influenced by genetic factors. This study aimed to identify and validate novelmarkers associated with ulcerative colitis, with a specific focuc on their regulation through DNA methylation. Methods: Gene expression and DNA methylation profiling of intestinal mucosal tissues from ulcerative colitis and healthy controls was retrieved from the GEO repository. Differentially expressed and methylated genes were examined in ulcerative colitis. Subsequently, overlapped analyses were performed to identify highly expressed and hypomethylated genes, as well as lowly expressed and hypermethylated genes. Functional annotation, transcription factor-mRNA network analysis and protein-protein interaction (PPI) network analysis were conducted for above genes. DSS-induced LOVO and Caco-2 cells were established to stimulate ulcerative colitis injury. The expression and methylation of B4GALNT2 was verified by RT-qPCR and MSP. CCK-8, flow cytometry, western blot, and ELISA were used to measure cell survival, apoptosis, and cytokine levels after B4GALNT2 overexpression. Results: Our study screened 1 down-regulated and hypermethylated gene (B4GALNT2) and 114 up-regulated and hypomethylated genes in ulcerative colitis. They were markedly associated with immune response. Totally, 10 potential transcription factors were predicted. The PPI network revealed their complex interactions. B4GALNT2 was confirmed to be down-regulated and hypermethylated in DSS-induced intestinal epithelial cells. B4GALNT2 overexpression enhanced cell viability and weakened apoptosis and cytokine production and release of DSS-induced intestinal epithelial cells. Conclusion: Collectively, this study integrally analyzed DNA methylation and gene expression in ulcerative colitis as well as identified and verified B4GALNT2 as a key epigenetic marker.