Leyla Valenick

Neurotensin is a proinflammatory neuropeptide in colonic inflammation

The neuropeptide neurotensin mediates several intestinal functions, including chloride secretion, motility, and cellular growth. However, whether this peptide participates in intestinal inflammation is not known. Toxin A, an enterotoxin from Clostridium difficile, mediates pseudomembranous colitis in humans. In animal models, toxin A causes an acute inflammatory response characterized by activation of sensory neurons and intestinal nerves and immune cells of the lamina propria. Here we show that neurotensin and its receptor are elevated in the rat colonic mucosa following toxin A administration. Pretreatment of rats with the neurotensin receptor antagonist SR-48, 692 inhibits toxin A-induced changes in colonic secretion, mucosal permeability, and histologic damage. Exposure of colonic explants to toxin A or neurotensin causes mast cell degranulation, which is inhibited by SR-48,692. Because substance P was previously shown to mediate mast cell activation, we examined whether substance P is involved in neurotensin-induced mast cell degranulation. Our results show that neurotensin-induced mast cell degranulation in colonic explants is inhibited by the substance P (neurokinin-1) receptor antagonist CP-96,345, indicating that colonic mast activation in response to neurotensin involves release of substance P. We conclude that neurotensin plays a key role in the pathogenesis of C. difficile-induced colonic inflammation and mast cell activation.

Regulation of the NK-1 receptor gene expression in human macrophage cells via an NF-κB site on its promoter

We report here that human monocytic/macrophage THP-1 cells express the neurokinin 1 receptor (NK-1R), and that exposure of these cells to the proinflammatory cytokine IL-1β increased the expression of the NK-1R gene at the mRNA and protein levels. Because IL-1β function involves nuclear factor κB (NF-κB) activation, these data suggest that this increase in the expression of the NK-1R gene is mediated by the NF-κB transcription factor. An earlier report noted that the promoter region of the human NK-1R gene contains a putative binding site for NF-κB [Takahashi, K., Tanaka, A., Hara, M. & Nakanishi, S. (1992) Eur. J. Biochem. 204, 1025–1033]. Here we demonstrate that this is indeed a functional NF-κB-binding site, and that NF-κB is responsible for regulating the expression of the NK-1R gene by binding to the promoter region of the NK-1R gene. To further substantiate that the observed NF-κB-dependent IL-1β induction of the human NK-1R gene is regulated via a transcriptional event through this NF-κB site on the NK-1R gene promoter, we transfected THP-1 cells with a luciferase promoter-reporter construct containing the 5′ promoter region of the human NK-1R gene. Exposure of these cells to IL-1β or overexpression of NF-κB cDNAs resulted in a significant increase in the amount of luciferase activity that was diminished greatly in cells transfected with IκBα, the NF-κB inhibitor. These results directly implicate NF-κB in the regulation of the NK-1R gene and provide a molecular mechanism for the increase in expression of the NK-1R gene in responsive cells.

Protective effects of neurokinin-1 receptor during colitis in mice: role of the epidermal growth factor receptor

The role of substance P and its high affinity neurokinin‐1 receptor in colitis has not been fully elucidated. We assessed the participation of neurokinin‐1 receptor in colitis using the 2,4,6,‐trinitrobenzensulphonic acid and dextran sulphate‐induced animal models of colitis and genetically‐engineered, neurokinin‐1 receptor‐deficient mice. Clinical signs, macroscopic and histologic damage associated with 2,4,6,‐trinitrobenzensulphonic acid (12 days) and dextran sulphate (5 days) colitis were more severe in neurokinin‐1 deficient than in wild‐type mice, while immunoreactivities for epidermal growth factor and its receptor were similar in the colon of both mice strains before and after colitis. Substance P, dose‐dependently induced intestinal fibroblast proliferation and enhanced epidermal growth factor‐induced proliferation in intestinal fibroblasts isolated from wild‐type, but not from neurokinin‐1 receptor deficient mice. Substance P‐induced intestinal fibroblast proliferation required the presence of epidermal growth factor receptor with kinase activity. Furthermore, substance P induced epidermal growth factor tyrosine phosphorylation and activation in normal intestinal fibroblasts. Our results indicate that in mice lacking the neurokinin ‐ 1 receptor, substance P plays a protective role in prolonged experimental colitis.