Joseph B. Ward

The bile acid receptor, TGR5, regulates basal and cholinergic-induced secretory responses in rat colon.

Bile acids (BA) are becoming increasingly appreciated as enteric hormones that regulate many aspects of intestinal physiology. The BA receptor, TGR5, has been recently shown to be expressed on enteric nerves and enterochromaffin cells (ECs), where its activation regulates small intestinal and colonic motility. Here, we show that TGR5 is also expressed on colonic epithelial cells and that its activation decreases basal secretory tone and inhibits cholinergic‐induced secretory responses. Our data demonstrate a new role for TGR5 in regulating colonic fluid and electrolyte transport and suggest that the receptor represents a good therapeutic target for intestinal transport disorders.

Farnesoid X receptor agonists attenuate colonic epithelial secretory function and prevent experimental diarrhoea in vivo

Objective Bile acids are important regulators of intestinal physiology, and the nuclear bile acid receptor, farnesoid X receptor (FXR), is emerging as a promising therapeutic target for several intestinal disorders. Here, we investigated a role for FXR in regulating intestinal fluid and electrolyte transport and the potential for FXR agonists in treating diarrhoeal diseases. Design Electrogenic ion transport was measured as changes in short-circuit current across voltage-clamped T84 cell monolayers or mouse tissues in Ussing chambers. NHE3 activity was measured as BCECF fluorescence in Caco-2 cells. Protein expression was measured by immunoblotting and cell surface biotinylation. Antidiarrhoeal efficacy of GW4064 was assessed using two in vivo mouse models: the ovalbumin-induced diarrhoea model and cholera toxin (CTX)-induced intestinal fluid accumulation. Results GW4064 (5 μmol/L; 24 h), a specific FXR agonist, induced nuclear translocation of the receptor in T84 cells and attenuated Cl− secretory responses to both Ca2+ and cAMP-dependent agonists. GW4064 also prevented agonist-induced inhibition of NHE3 in Caco-2 cells. In mice, intraperitoneal administration of GW4064 (50 mg/mL) also inhibited Ca2+ and cAMP-dependent secretory responses across ex vivo colonic tissues and prevented ovalbumin-induced diarrhoea and CTX-induced intestinal fluid accumulation in vivo. At the molecular level, FXR activation attenuated apical Cl− currents by inhibiting expression of cystic fibrosis transmembrane conductance regulator channels and inhibited basolateral Na+/K+-ATPase activity without altering expression of the protein. Conclusions These data reveal a novel antisecretory role for the FXR in colonic epithelial cells and suggest that FXR agonists have excellent potential for development as a new class of antidiarrheal drugs.

Ursodeoxycholic acid attenuates colonic epithelial secretory function.

•  Although diarrhoeal diseases represent a significant health and economic burden to society, therapeutic options remain limited. •  While several bile acids are known to stimulate epithelial Cl− secretion, the major driving force for fluid secretion in the intestine, the effects of ursodeoxycholic acid (UDCA) on epithelial transport function are not well described. •  We report that in contrast to other bile acids, UDCA exerts anti‐secretory actions on colonic epithelial cells in vitro. •  In contrast, in vivo administration of UDCA enhances epithelial secretory function, an affect we ascribe to being due to its bacterial metabolism to lithocholic acid. In keeping with this hypothesis, in vivo administration of a metabolically stable analogue of UDCA, 6α‐methyl‐UDCA, was anti‐secretory. •  Our findings reveal novel anti‐secretory actions of UDCA and suggest that metabolically stable analogues of bile acid may be useful for development as a new class of anti‐diarrhoeal drug.

Oxygen in the regulation of intestinal epithelial transport

The transport of fluid, nutrients and electrolytes to and from the intestinal lumen is a primary function of epithelial cells. Normally, the intestine absorbs approximately 9 l of fluid and 1 kg of nutrients daily, driven by epithelial transport processes that consume large amounts of cellular energy and O2. The epithelium exists at the interface of the richly vascularised mucosa, and the anoxic luminal environment and this steep O2 gradient play a key role in determining the expression pattern of proteins involved in fluid, nutrient and electrolyte transport. However, the dynamic nature of the splanchnic circulation necessitates that the epithelium can evoke co‐ordinated responses to fluctuations in O2 availability, which occur either as a part of the normal digestive process or as a consequence of several pathophysiological conditions. While it is known that hypoxia‐responsive signals, such as reactive oxygen species, AMP‐activated kinase, hypoxia‐inducible factors, and prolyl hydroxylases are all important in regulating epithelial responses to altered O2 supply, our understanding of the molecular mechanisms involved is still limited. Here, we aim to review the current literature regarding the role that O2 plays in regulating intestinal transport processes and to highlight areas of research that still need to be addressed.

Hydroxylase inhibition attenuates colonic epithelial secretory function and ameliorates experimental diarrhea

Hydroxylases are oxygen‐sensing enzymes that regulate cellular responses to hypoxia. Transepithelial Cl− secretion, the driving force for fluid secretion, is dependent on O2 availability for generation of cellular energy. Here, we investigated the role of hydroxylases in regulating epithelial secretion and the potential for targeting these enzymes in treatment of diarrheal disorders. Ion transport was measured as short‐circuit current changes across voltage‐clamped monolayers of T84 cells and mouse colon. The antidiarrheal efficacy of dimethyloxallyl glycine (DMOG) was tested in a mouse model of allergic disease. Hydroxylase inhibition with DMOG attenuated Ca2+‐ and cAMP‐dependent secretory responses in voltage‐clamped T84 cells to 20.2 ± 2.6 and 38.8 ± 6.7% (n= 16; P<0.001) of those in control cells, respectively. Antisecretory actions of DMOG were time and concentration dependent, being maximal after 18 h of DMOG (1 mM) treatment. DMOG specifically inhibited Na+/K+‐ATPase pump activity without altering its expression or membrane localization. In mice, DMOG inhibited agonist‐induced secretory responses ex vivo and prevented allergic diarrhea in vivo. In conclusion, hydroxylases are important regulators of epithelial Cl− and fluid secretion and present a promising target for development of new drugs to treat transport disorders.—Ward, J. B. J., Lawler, K., Amu, S., Taylor, C. T., Fallon, P. G., Keely, S. J. Hydrox‐ylase inhibition attenuates colonic epithelial secretory function and ameliorates experimental diarrhea. FASEB J. 25, 535–543 (2011). www.fasebj.org

Ursodeoxycholic acid inhibits TNFα-induced IL-8 release from monocytes

Monocytes are critical to the pathogenesis of inflammatory bowel disease (IBD) as they infiltrate the mucosa and release cytokines that drive the inflammatory response. Ursodeoxycholic acid (UDCA), a naturally occurring bile acid with anti-inflammatory actions, has been proposed as a potential new therapy for IBD. However, its effects on monocyte function are not yet known. Primary monocytes from healthy volunteers or cultured U937 monocytes were treated with either the proinflammatory cytokine, TNFα (5 ng/ml) or the bacterial endotoxin, lipopolysaccharide (LPS; 1 μg/ml) for 24 h, in the absence or presence of UDCA (25-100 μM). IL-8 release into the supernatant was measured by ELISA. mRNA levels were quantified by qPCR and changes in cell signaling proteins were determined by Western blotting. Toxicity was assessed by measuring lactate dehydrogenase (LDH) release. UDCA treatment significantly attenuated TNFα-, but not LPS-driven, release of IL-8 from both primary and cultured monocytes. UDCA inhibition of TNFα-driven responses was associated with reduced IL-8 mRNA expression. Both TNFα and LPS stimulated NFκB activation in monocytes, while IL-8 release in response to both cytokines was attenuated by an NFκB inhibitor, BMS-345541. Interestingly, UDCA inhibited TNFα-, but not LPS-stimulated, NFκB activation. Finally, TNFα, but not LPS, induced phosphorylation of TNF receptor associated factor (TRAF2), while UDCA cotreatment attenuated this response. We conclude that UDCA specifically inhibits TNFα-induced IL-8 release from monocytes by inhibiting TRAF2 activation. Since such actions would serve to dampen mucosal immune responses in vivo, our data support the therapeutic potential of UDCA for IBD.

Chronic regulation of colonic epithelial secretory function by activation of G protein-coupled receptors.

Background  Enteric neurotransmitters that act at G protein‐coupled receptors (GPCRs) are well known to acutely promote epithelial Cl− and fluid secretion. Here we examined if acute GPCR activation might have more long‐term consequences for epithelial secretory function.