mRNA
and
miRNA
Sequencing Identifies Increased Oxidative Stress Pathways in a Mouse Model of Ethanol Exacerbation of Colitis
Madison M. Tschann, Mary Grace Murray, Abigail R. Cannon, Paulius V. Kuprys, Keit Dine, Joshua M. Eberhardt, Xianzhong Ding, Nan Miles Xi, Joel R. Pekow, Mashkoor A. Choudhry ABSTRACT
Background
Alcohol consumption has been found to worsen the pathology of colitis. To elucidate the underlying mechanism, we performed both miRNA and mRNA sequencing of the large intestinal epithelial cells derived from a murine model of alcohol‐induced colitis.
Methods
Mice received Dextran Sulfate Sodium (DSS) for 5 days to induce colitis and were gavaged with alcohol (~3 g/kg) or water for 3 days. Three hours after the final gavage, mice were euthanized, and colon lengths were measured, followed by epithelial cell isolation. RNA, including miRNA, was isolated from the colonic epithelial cells and sent to Novogene for mRNA and miRNA sequencing.
Results
Analysis comparing DSS Ethanol vs. Sham Vehicle mice showed 3931 significantly upregulated genes and 3788 significantly downregulated genes. Functional enrichment analysis of GO and KEGG pathways showed enrichment of positive regulation of superoxide anion generation and HIF‐1 signaling pathways, both related to oxidative stress. DSS Ethanol compared to DSS Vehicle resulted in 921 significantly upregulated and 1072 significantly downregulated genes ( P adj < 0.05), with functional enrichment analysis showing the term with the highest fold enrichment was superoxide anion generation. We further confirmed the RNA expression of NOS2, CYBB, and NCF1, which are involved in nitric oxide and superoxide production. miRNA sequencing revealed 84 differentially expressed miRNAs comparing DSS Ethanol to DSS Vehicle. Functional enrichment analysis of predicted targets of these miRNAs also showed enrichment in superoxide anion generation.
Conclusion
These results suggest that combined ethanol and DSS exposure may increase the production of reactive oxygen species, which further damages the intestinal lining and potentially contributes to the ethanol exacerbation of colitis.