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TL1A is a member of the TNF superfamily and its expression is increased in the mucosa of inflammatory bowel disease patients. Moreover, a subset of Crohn's disease (CD) patients with the risk TL1A haplotype is associated with elevated TL1A expression and a more severe disease course. To investigate the in vivo role of elevated TL1A expression, we generated two transgenic (Tg) murine models with constitutive Tl1a expression in either lymphoid or myeloid cells. Compared to wildtype (WT) mice, constitutive expression of Tl1a in either lymphoid or myeloid cells showed mild patchy inflammation in the small intestine, which was more prominent in the ileum. In addition, mice with constitutive Tl1a expression exhibited enhanced intestinal and colonic fibrosis compared to WT littermates. The percentage of T cells expressing the gut homing chemokine receptors CCR9 and CCR10 was higher in the Tl1a Tg mice compared to WT littermates. Sustained expression of Tl1A in T cells also lead to increased Foxp3+ Treg cells. T cells or antigen presenting cells (APC) with constitutive expression of Tl1a were found to have a more activated phenotype and mucosal mononuclear cells exhibit enhanced Th1 cytokine activity. These results indicated an important role of TL1A in mucosal T cells and APC function and showed that up-regulation of TL1A expression can promote mucosal inflammation and gut fibrosis.

The action of highly purified Clostridium difficile toxin A was studied in the jejunum of rats in vivo. C. difficile toxin A reversed dose-dependently net fluid absorption into net fluid secretion, accompanied by an increase in prostaglandin E2 but not 5-hydroxytryptamine output into the gut lumen. Accordingly, indomethacin but not the 5-hydroxytryptamine receptor antagonists ketanserin plus tropisetron were able to inhibit toxin A-induced fluid secretion. Atropine and hexamethonium were without effect on the action of toxin A, such excluding a nervous mechanism. The cyclic nucleotides cyclic AMP and cyclic GMP appear not to be involved in the mediation of the secretory response. The reduced cyclic GMP levels are most likely the result of a complete destruction of the villus membranes, where the guanylate cyclase is located. Histological studies revealed massive damage to intestinal villi, whereas the majority of the crypts seem to be unaffected. In conclusion, toxin A-induced intestinal fluid secretion appears to be caused mainly by severe mucosal damage. PGE2-release may be the consequence of the inflammation accompanying this damage. The mechanism seems to be completely different to those of cholera toxin or Escherichia coli heat stable enterotoxin. © 1992 Wiley-Liss, Inc.

G A A b st ra ct s (untreated), P F). The beneficial effects of E2 appeared to be mediated through ERb since DPN (ERb agonist), but not PPT (ERa agonist) appeared to decrease epithelial permeability in SAMP mice. Conclusions: These results indicate that estrogen/ERb interaction with IEC may modulate gut epithelial barrier function, potentially by regulating TJ protein expression, and subsequent chronic intestinal inflammation characteristic of CD.

G A A b st ra ct s proximal colon by day 7, had no effect on mucosa from mid-colon, and caused a timedependent decrease (47% by day 7) in mucosa from distal colon. In muscle, there was a time-dependent decrease (35% by day 7) in NT-3 in proximal colon but a time-dependent increase in muscle from mid-colon (105%) and distal colon (43%) by day 7. In conclusion, NGF, BDNF, and NT-3 are expressed in distinct region-and tissue-specific patterns in naive rats. Colonic inflammation results in differential changes in these patterns which are neurotrophin-specific.

The contribution of toxin B to Clostridium difficile-associated infection is undefined. Toxin B induces dramatic phenotypic alterations (cytotoxic effects) in cultured mesenchymal and nonintestinal epithelial cells, yet its effects on intestinal epithelial cells are not clearly understood. The alterations induced by toxin B in nonintestinal cells appear to be secondary to toxin-induced redistribution of filamentous actin. It has not been determined whether toxin B exerts similar effects on cultured intestinal epithelial cells or whether such phenotypic alterations are of any physiological consequence. To address these questions, we examined the effect of C. difficile toxin B on the phenotype and barrier function of T84 cell monolayers. Our studies show that the cytotoxic effects of toxin B, i.e., cell rounding, do extend to cultured intestinal epithelial cells (T84). In addition, toxin B dramatically reduces the barrier function of T84 monolayers grown on collagencoated filters. Toxin B-induced redistribution of filamentous actin appears to be responsible for the alterations in both intestinal epithelial cell phenotype and barrier function. Specifically, filamentous actin comprising the perijunctional actomyosin ring, known to be important in regulating tight junction permeability, is condensed into discrete plaques. Flux studies confirm that the permeability defect is at the level of the tight junction. We conclude that toxin-induced changes in actin distribution perturb intercellular junctional contacts and thereby ablate epithelial barrier function. There was no evidence of cell death as determined by lactate dehydrogenase release assays.

Introduction Hypertrophic mesenteric adipose tissue enwrapping the intestine has been considered as a specific sign of Crohn's disease (CD). The role of this “creeping fat” in CD is unknown. Cartonectin (collagenous repeat-containing sequence of 26 kDa protein, CORS26) is a newly identified secretory protein produced by adipocytes with yet unclear function. Intestinal fibroblasts play an important role in the stricturing and inflammatory process of CD. Aim of this study was to investigate the in-vitro effect of Cartonectin on primary human colonic lamina propria fibroblasts (CLPF) from CD tissue. Materials and Methods: CLPF were isolated from strictured and non-strictured colonic specimens of CD and control patients. For CLPF stimulation increasing concentrations of recombinant human Cartonectin (10, to 500ng/ml) were used alone or for costimulation with LPS or TNF. IL-6, IL-8 and TGF-β in supernatants were quantified by ELISA. mRNA levels of collagen 1, collagen 3 and connective tissue growth factor (CTGF) were detected by real-time PCR. Results: LPSinduced IL-8 secretion fromnormal CLPFwas suppressed by Cartonectin in a dose-dependent fashion (200 ng/ml: p=0.0002 at 8h; p

Background & Aims Chronic psychological stress is associated with visceral hyperalgesia and increased expression of spinal NK1 receptors (NK1Rs). We aimed to identify the role of spinal microglia in this process. Methods Male Wistar rats were exposed to water avoidance (WA) or sham stress 1 hour each day for 10 days and given daily injections of minocycline, the p38 inhibitor SB203580, or saline. Phosphorylation levels of the kinase p38 (P-p38), the microglia marker OX42, NK1R, and IκBα were assessed by immunoblotting and/or immunostaining of spinal samples collected at day 11. The visceromotor response to colorectal distention at baseline and following WA were also assayed in rats given injections of minocycline, SB203580, or vehicle. The effects of fractalkine were assessed on the visceromotor response in rats exposed to minocycline or vehicle. Results P-p38 protein levels and immunoreactivity were increased in stressed rats and colocalized with OX42-positive cells and neurons in the dorsal horn. This increase was reversed by minocycline or SB203580 exposure. Stress-induced increased NK1R expression was blocked by minocycline but not SB203580. WA-induced decreased IκBα expression was blocked by minocycline and SB203580. WA-induced hyperalgesia was blocked by minocycline and SB203580 intrathecally. Fractalkine-induced hyperalgesia was blocked by minocycline. Conclusions This is the first demonstration that stress-induced activation of spinal microglia has a key role in visceral hyperalgesia and associated spinal NK1R up-regulation.

Mucosal sheets of guinea pig ileum mounted in Ussing chambers were used to determine effects of highly purified Clostridium difficile toxin A on intestinal structure and barrier function in the absence of recruited neutrophils and blood flow. With the use of standard electrophysiological and morphological techniques, our results indicate that 5 micrograms/ml mucosal toxin A induces substantial alteration in epithelial permeability and in structure of absorptive cells. Transepithelial fluxes of mannitol (3.6 A) and inulin (11.5 A) increased 20-80 min after toxin A exposure (63 +/- 13 vs. 149 +/- 14 and 2.33 +/- 0.43 vs. 7.03 +/- 1.0 nmol.cm-2.h-1 for mannitol and inulin; controls vs. toxin exposed, respectively, both P less than 0.01). Toxin A exposure diminished short-circuit current (153 +/- 8 vs. 77 +/- 6 microA/cm2 for control and toxin exposed, respectively, P less than 0.001), decreased transepithelial electrical resistance (61.6 +/- 3 vs. 50.1 +/- 3.9 omega.cm2 for control and toxin exposed, respectively, P less than 0.05), increased passive permeation of Na+ (19.1 +/- 0.9 vs. 27.4 +/- 0.9 mumol.cm-2.h-1 for control and toxin exposed, respectively), and induced a Cl- secretory response (net flux 3.65 +/- 3.0 vs. -4.62 +/- 2.6 mumol.cm-2.h-1, for control and toxin exposed, respectively, P less than 0.05) over the 2-h experimental time course. Toxin A-induced structural alterations of villus tip absorptive cells were strikingly similar to those induced by the actin-binding agent cytochalasin D. Specifically, cells displayed constricted subjunctional zones, flared microvillus brush borders, condensation of microfilaments in the zone of the perijunctional actomyosin ring, and breakdown of intercellular tight junctions.(ABSTRACT TRUNCATED AT 250 WORDS)