Expression of inducible nitric oxide synthase in mast cells contributes to the regulation of inflammatory cytokines in irritable bowel syndrome with diarrhea

Background Nitric oxide (NO) and mast cells (MCs) are possibly involved in the development of irritable bowel syndrome (IBS), but details on their role and interactions still remain undetermined. We aimed to investigate the expression of inducible NO synthase (iNOS) in MCs of the colon of IBS with diarrhea (IBSD), and elucidated a potential role of NO in the differential regulation of cytokines in MCs. Methods Colonic mucosal biopsies of 19 IBS-D patients and 16 healthy controls were collected. The expression of tryptase and iNOS was investigated by immunohistochemistry, Western blotting, and real-time PCR. Effects of NO on the expression of cytokines in rat bone marrow MCs (BMMCs) were examined using a cytokine array by NG-nitro-L-arginine methyl ester (L-NAME) treatment. Key Results Immunohistochemistry for tryptase revealed an increase in number of MCs with extensive iNOS expression in the colonic mucosa of IBS-D. Tryptase, iNOS and interleukin (IL)-1 beta mRNA and protein levels were upregulated in IBSD compared with healthy controls. Specifically, a positive correlation between tryptase and iNOS protein expression was observed in the colon of IBS-D (r = 0.667, p < 0.05). Supernatant from IBS-D increased iNOS expression in BMMCs. Antibody array showed that agrin, beta-nerve growth factor, fractalkine, granulocyte-macrophage colony-stimulating factor, IL-1 beta, IL-1R6, IL-13, leptin, tumor necrosis factor alpha were suppressed, and cytokine-induced neutrophil chemoattractant (CINC)-1, CINC-2 alpha, CINC-3, monocyte chemotactic protein-1, matrix metalloproteinase-8 were strongly produced in L-NAME treated BMMCs, comparable to levels in the control group. Conclusions & Inferences Our findings provide new evidence that NO is able to regulate many cytokines in MCs that may be involved in the development of IBS.