In-depth investigation of the mechanism in rats with alcoholic acute pancreatitis via DNA methylation, intestinal flora, and fecal metabolomics.

BACKGROUND

Alcohol consumption is the main cause of acute pancreatitis (AP). Alcohol combined with cerulein (CI) exerts direct damage to pancreas, with its mechanism undefined.

METHODS

Rats were randomly divided into three subgroups, with subgroup I including Control and Alcohol groups at weeks 6, 8, and 10, subgroup II including Control and Alcohol groups administered CI (12.5, 25, and 50 µg/kg), and subgroup III including Control and Alcohol groups administered CI (50 µg/kg). Following measurement of liver and kidney function indicators, pathological damage of liver, kidney, lung, and intestine was detected using HE staining. Methylation-Specific PCR, Sequenom MassARRAY methylation, ELISA, Western blot, and correlation analysis of 16S rRNA sequencing and metabolomics were performed.

RESULTS

Compared to Control group, Alcohol + CI group increased pathological damage, serum aspartate aminotransferase, alanine aminotransferase, gamma-glutamyl transferase, blood urea nitrogen, alkaline phosphatase, amylase, lipase, TNF-α, IL-6, and IL-1β levels (P < 0.01). Meanwhile, p-mTOR/mTOR, hypoxia-inducible factor 1 alpha (HIF-1α), and vascular endothelial growth factor were enhanced, with lower creatinine and HIF-1α DNA methylation (P < 0.01). Alcohol at week 8 caused higher HIF-1α-25_CpG_35 methylation (P = 0.0222), which were reduced by further CI (50 µg/kg, P = 0.0034). Moreover, strong correlation was reported between key flora (Methanobrevibacter_A, Ruminococcus_C_59129, Lactobacillus, Eubacterium_Q, and Phascolarctobacterium_A) and metabolites (Homocysteine, Aspartic acid, Agmatine, 6-Ketoprostaglandin e1, Stearic acid, and Palmitic acid).

CONCLUSION

HIF-1α-25_CpG_35 methylation, intestinal flora, and metabolites involved in regulating AP pathogenesis. Methanobrevibacter_A, Ruminococcus_C_59129, Lactobacillus, Eubacterium_Q, Phascolarctobacterium_A, Homocysteine, Aspartic acid, Agmatine, 6-Ketoprostaglandin e1, Stearic acid, and Palmitic acid were the potential biomarkers, offering new insights into the pathogenesis of AP.