Background and AimsThe clinical onset and severity of intestinal disorders in humans and animals can be profoundly impacted by early life stress. Here we investigated the impact of early weaning stress in pigs on intestinal physiology, clinical disease, and immune response to subsequent challenge with enterotoxigenic F18 E. coli (ETEC).MethodologyPigs weaned from their dam at 16 d, 18 d, and 20 d of age were given a direct oral challenge of F18 ETEC at 26 d of age. Pigs were monitored from days 0 to 4 post-infection for clinical signs of disease. On Day 4 post-ETEC challenge, ileal barrier function, histopathologic and inflammatory cytokine analysis were performed on ileal mucosa.ResultsEarly weaned pigs (16 d and 18 d weaning age) exhibited a more rapid onset and severity of diarrhea and reductions in weight gain in response to ETEC challenge compared with late weaned pigs (20 d weaning age). ETEC challenge induced intestinal barrier injury in early weaned pigs, indicated by reductions in ileal transepithelial electrical resistance (TER) and elevated FD4 flux rates, in early weaned pig ileum but not in late weaned pigs. ETEC-induced marked elevations in IL-6 and IL-8, neutrophil recruitment, and mast cell activation in late-weaned pigs; these responses were attenuated in early weaned pigs. TNF levels elevated in ETEC challenged ileal mucosa from early weaned pigs but not in other weaning age groups.ConclusionsThese data demonstrate the early weaning stress can profoundly alter subsequent immune and physiology responses and clinical outcomes to subsequent infectious pathogen challenge. Given the link between early life stress and gastrointestinal diseases of animals and humans, a more fundamental understanding of the mechanisms by which early life stress impacts subsequent pathophysiologic intestinal responses has implications for the prevention and management of important GI disorders in humans and animals.

Background and Aims Psychological stress is a predisposing factor in the onset and exacerbation of important gastrointestinal diseases including irritable bowel syndrome (IBS) and the inflammatory bowel diseases (IBD). The pathophysiology of stress-induced intestinal disturbances is known to be mediated by corticotropin releasing factor (CRF) but the precise signaling pathways remain poorly understood. Utilizing a porcine ex vivo intestinal model, the aim of this study was to investigate the mechanisms by which CRF mediates intestinal epithelial barrier disturbances. Methodology Ileum was harvested from 6–8 week-old pigs, mounted on Ussing Chambers, and exposed to CRF in the presence or absence of various pharmacologic inhibitors of CRF-mediated signaling pathways. Mucosal-to-serosal flux of 4 kDa-FITC dextran (FD4) and transepithelial electrical resistance (TER) were recorded as indices of intestinal epithelial barrier function. Results Exposure of porcine ileum to 0.05–0.5 µM CRF increased (p<0.05) paracellular flux compared with vehicle controls. CRF treatment had no deleterious effects on ileal TER. The effects of CRF on FD4 flux were inhibited with pre-treatment of tissue with the non-selective CRF 1/2 receptor antagonist Astressin B and the mast cell stabilizer sodium cromolyn (10 −4 M). Furthermore, anti-TNF-α neutralizing antibody (p<0.01), protease inhibitors (p<0.01) and the neural blocker tetrodotoxin (TTX) inhibited CRF-mediated intestinal barrier dysfunction. Conclusion These data demonstrate that CRF triggers increases in intestinal paracellular permeability via mast cell dependent release of TNF-α and proteases. Furthermore, CRF-mast cell signaling pathways and increases in intestinal permeability require critical input from the enteric nervous system. Therefore, blocking the deleterious effects of CRF may address the enteric signaling of mast cell degranulation, TNFα release, and protease secretion, hallmarks of IBS and IBD.

Life stress is a major risk factor in the onset and exacerbation of mast cell-associated diseases, including allergy/anaphylaxis, asthma, and irritable bowel syndrome. Although it is known that mast cells are highly activated upon stressful events, the mechanisms by which stress modulates mast cell function and disease pathophysiology remains poorly understood. Here, we investigated the role of corticotropin-releasing factor receptor subtype 1 (CRF) in mast cell degranulation and associated disease pathophysiology. In a mast cell-dependent model of IgE-mediated passive systemic anaphylaxis (PSA), prophylactic administration of the CRF-antagonist antalarmin attenuated mast cell degranulation and hypothermia. Mast cell-deficient mice engrafted with CRF bone marrow-derived mast cells (BMMCs) exhibited attenuated PSA-induced serum histamine, hypothermia, and clinical scores compared with wild-type BMMC-engrafted mice. mice engrafted with CRF BMMCs also exhibited suppressed in vivo mast cell degranulation and intestinal permeability in response to acute restraint stress. Genetic and pharmacologic experiments with murine BMMCs, rat RBL-2H3, and human LAD2 mast cells demonstrated that although CRF activation did not directly induce MC degranulation, CRF signaling potentiated the degranulation responses triggered by diverse mast cell stimuli and was associated with enhanced release of Ca from intracellular stores. Taken together, our results revealed a prominent role for CRF signaling in mast cells as a positive modulator of stimuli-induced degranulation and in vivo pathophysiologic responses to immunologic and psychologic stress.

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