Alcohol inducing macrophage M2b polarization in colitis by modulating the TRPV1-MAPK/NF-κB pathways.

BACKGROUND: Macrophages exhibit different phenotypes in inflammatory bowel disease (IBD) and promote inflammation or tissue repair depending on their polarization state. Alcohol is a widely used solvent in pharmaceutical formulations, and its consumption is associated with an increased risk of colitis; however, its effects on macrophages in IBD remain poorly understood. PURPOSE: This study aimed to investigate the effect of alcohol on macrophages in dextran sodium sulfate (DSS)-induced colitis and understand the underlying mechanisms. METHODS: DSS-treated C57BL/6 mice were exposed to varying concentrations of alcohol, transient receptor potential vanilloid 1 (TRPV1) antagonist, and 5-aminosalicylic acid. The distal colon was resected, fixed, stained, and histologically analyzed, through hematoxylin and eosin (H&E) staining and immunofluorescence staining. Ratio [Ca2+]i measurements, western blotting, quantitative polymerase chain reaction, cytokine measurements, and RNA sequencing analyses were also performed. Peritoneal macrophages and RAW264.7 cells were used for in vitro experiments, and various assays were performed to evaluate cellular responses, gene expression, and signaling pathways. RESULTS: Alcohol exacerbated DSS-treated mice colitis and promoted the secretion of various inflammatory cytokines from colonic macrophages. Alcohol enhances the calcium ion influx induced by lipopolysaccharide (LPS) in peritoneal macrophages, while the TRPV1 antagonist capsazepine (CPZ) inhibits LPS- and/or alcohol- induced calcium influx in macrophages. Alcohol and LPS activate the MAPK/P38, MAPK/ERK, and NF-κB signaling pathways and induce the macrophage M2b polarization, resulting in the increased expression level of inflammatory cytokines such as Tnf, Il1b, and Il10. Additionally, CPZ can inhibit the facilitatory effects of alcohol or LPS on the abovementioned pathways and inflammatory factors, reversing macrophage M2b polarization and promoting alcohol-induced colitis. The inhibition of nucleotide binding oligomerization domain containing 2 (NOD2) partially suppressed the alcohol and LPS effects on macrophages. CONCLUSION: Alcohol exacerbates experimental colitis and induces M2b polarization of macrophage via TRPV1-MAPK/NF-κB. Our study provides new insights into the potential therapeutic targets for IBD treatment by elucidating the role of TRPV1 in alcohol-exacerbated colitis, using CPZ as a potential therapeutic option. The identification of transient receptor potential ankyrin subtype 1 (TRPA1) as a therapeutic target expands the scope of future research.

TRPM8 inhibits substance P release from primary sensory neurons via PKA/GSK-3beta to protect colonic epithelium in colitis.

Transient receptor potential melastatin 8 (TRPM8) is a cold sensory receptor in primary sensory neurons that regulates various neuronal functions. Substance P (SP) is a pro-inflammatory neuropeptide secreted by the neurons, and it aggravates colitis. However, the regulatory role of TRPM8 in SP release is still unclear. Our study aimed to investigate TRPM8's role in SP release from primary sensory neurons during colitis and clarify the effect of SP on colonic epithelium. We analyzed inflammatory bowel disease patients' data from the Gene Expression Omnibus dataset. Dextran sulfate sodium (DSS, 2.5%)-induced colitis in mice, mouse dorsal root ganglion (DRG) neurons, ND7/23 cell line, and mouse or human colonic organoids were used for this experiment. Our study found that TRPM8, TAC1 and WNT3A expression were significantly correlated with the severity of ulcerative colitis in patients and DSS-induced colitis in mice. The TRPM8 agonist (menthol) and the SP receptor antagonist (Aprepitant) can attenuate colitis in mice, but the effects were not additive. Menthol promoted calcium ion influx in mouse DRG neurons and inhibited the combination and phosphorylation of PKAca from the cAMP signaling pathway and GSK-3ß from the Wnt/ß-catenin signaling pathway, thereby inhibiting the effect of Wnt3a-driven ß-catenin on promoting SP release in ND7/23 cells. Long-term stimulation with SP inhibited proliferation and enhanced apoptosis in both mouse and human colonic organoids. Conclusively, TRPM8 inhibits SP release from primary sensory neurons by inhibiting the interaction between PKAca and GSK-3ß, thereby inhibiting the role of SP in promoting colonic epithelial apoptosis and relieving colitis.