44 Abnormal Immune-Mediated (IL1β/LPS) Regulation of Small Intestinal EC Cell Serotonin Release in Inflammatory Bowel Disease

Abstract

Background: Inflammatory bowel disease (IBD) is a common but troublesome disease with limited treatment options. The mechanisms behind IBD-associated mucosal damage and secretory andmotility abnormalities are complex and ill-defined. The gut mucosal enterochromaffin (EC) cell is regarded as a key regulator of intestinal motility and fluid secretion via its secretion of serotonin. It has recently been demonstrated that immune cells are located in close proximity to EC cells in the intestinal mucosa. Furthermore, EC cell numbers are increased in gut inflammation, and as gut hypermobility and hypersecretion are typical of IBD symptoms, we postulated that an activated immune system may lead to EC cell dysregulation and hypersecretion of serotonin. We further hypothesized that both immune derived cytokines (IL1β) and lipopolysaccharides (LPS) from bacteria may induce EC cell activation. LPS signaling is known to involve NFκβ activation through binding to TOLL receptor 4 (TLR4). Methods and results: Pure (>98%) human intestinal EC cells were isolated by FACS sorting from preparations of normal (n=5) and IBD (n=6) mucosa. Using real time PCR, a 5-fold higher TLR4 (p 2-fold (p<0.05) in IBD EC cell cultures compared to normal EC cells. This was not due to cell damage (<1% alteration in Trypan blue or MTT uptake) and was reversible by the TLR4 antagonist, E. coli K12 (IC50=12ng/ml) and the IL1β receptor antagonist (ILRA; IC50=3.4ng/ml) respectively. Addition of IL1β to IBD EC cells also resulted in significant (p<0.05) NFκβ phosphorylation (30-40%). The somatostatin analogue, lanreotide (somatostatin receptor profile: 2,3,5), inhibited IL1β-stimulated secretion in both IBD EC (IC50= 0.61nM) and normal EC cells (IC50=1.8nM). In conclusion, serotonin release from EC cells isolated from IBDmucosa is increased more potently by both interleukins (IL1β) and bacterial products (E. coli LPS) compared to normal EC cells. This occurs through a mechanism that involves NFκβ phosphorylation and can be inhibited by somatostatin analogues and a TLR4 antagonist. Identifying and delineating an abnormal EC cell function is important in elucidating the pathogenesis behind hypersecretion in IBD. In addition, investigating the role of the EC cell will provide the potential for an alternative therapeutic target to reduce symptoms in IBD.