BACKGROUND: Oxidative DNA damage occurs during mucosal inflammation, with such lesions stalling RNA polymerase II and impairing transcription fidelity, thereby driving genomic instability, a hallmark of cancer. TCEA2 (TFIIS) is a transcription elongation factor that rescues stalled polymerases by stimulating transcript cleavage, enabling lesion bypass and transcription-coupled repair. Dysregulated TCEA2 may worsen transcription stress and mutagenesis under inflammatory conditions. Farnesoid X receptor (FXR), a bile acid–activated nuclear receptor, preserves intestinal barrier integrity, prevents mucosal inflammation and colitis-associated colorectal cancer by regulating immune signaling and epithelial homeostasis. This suggests FXR may influence transcriptional stress pathways, including TCEA2 regulation, linking bile acid signaling to genome stability. AIM: To determine whether FXR activation modulates TCEA2 expression in intestinal epithelial cells. METHODS: Differentiated and undifferentiated ileal and colonic enteroids were grown as monolayers and treated with the FXR agonist, obeticholic acid (OCA; 10 µM) for 6 – 24 hrs. Changes in gene expression were analysed by bulk RNASeq and were expressed as fold change in Reads Per Kilobase of Transcript (RPKM) compared to untreated controls (n = 3 throughout). TCEA2 expression in T84 cell monolayers was analyzed by qPCR. RESULTS: Treatment with OCA increased TCEA2 expression in UD colonic enteroids by 17.7 ± 10.9 fold after 6 hrs and 53.0 ± 33.1 fold after 24 hrs compared to unstimulated cells (n = 9; p< 0.001). Similarly, in DF colonic enteroids OCA treatment increased TCEA2 expression by 24.8 ± 5.4 and 33.4 ± 7.3 fold over controls at 6 and 24 hrs, respectively. OCA also stimulated TCEA2 expression in ileal enteroids with maximal responses occurring after 24 hrs of 127.5 ± 41.5 and 782.2 ± 303.2 fold of control in UD and DF cells, respectively (n = 3; p < 0.05). The effects of OCA were specific to TCEA2 as the agonist did not increase expression of the related elongation factors, TCEA1 or TCEA3. In cultured monolayers of T84 colonic epithelial cells the synthetic FXR agonist, GW4064 (5 µM; 3 - 48 hrs), also increased TCEA2 mRNA expression with a maximal effect of 390.3 ± 75.4 fold over controls occurring after 24 hrs of treatment (n = 4; p < 0.05). The effects of GW4064 on TCEA2 expression were significantly inhibited by pretreatment with the NF-κB inhibitor, BMS 345541, from 405.3 ± 60.7 to 96.4 ± 34.1 fold of controls (n = 5; p < 0.01). CONCLUSION: These data demonstrate FXR activation to strongly upregulate TCEA2 mRNA expression in colonic epithelial cells, with effects confirmed across enteroid and T84 cell models. The response is specific to TCEA2 and, at least partially, NF-κB–dependent, suggesting a novel FXR–NF-κB–TCEA2 regulatory axis. These findings indicate FXR may enhance transcriptional resilience under conditions of stress, an effect that may contribute to its anti-inflammatory and anti-cancer effects in vivo. This abstract was presented at the American Physiology Summit 2026 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.

These studies demonstrate for the first time that FXR activation inhibits cytokine-induced necroptosis in vitro, an effect that may underlie protection against dysregulated barrier function in the setting of intestinal inflammation. These data support the potential for targeting FXR to promote epithelial barrier function in treatment of IBD.

Introduction: The nuclear bile acid receptor, farnesoid X receptor (FXR), is a critical regulator of colonic epithelial transport and barrier function. Downregulated expression of the receptor is associated with ulcerative colitis (UC) and colorectal cancer (CRC), while studies in animal models show FXR activation to be protective in these conditions. Previous studies have also shown certain plant-derived extracts to be protective against the development of colonic inflammation and cancer, while others have shown that phytochemicals present within such extracts have the capacity to act as ligands of FXR. With this in mind, the current study set out to investigate the potential for using dietary plant extracts to modulate FXR activity in colonic epithelial cells. Methods: A lipid extract (designated QE2) was prepared by supercritical CO2 extraction from a dietary plant grain ( identity withheld ). Studies were carried out on T84 colonic epithelial cells grown as polarised monolayers on semipermeable supports. FXR expression was measured by RT-PCR and Western blotting, while activity was assessed by measuring basolateral secretion of the FXR target protein, FGF-19, by ELISA. Results: Apical treatment of T 84 cells with QE2 alone (24 hrs) was without effect on FGF-19 mRNA expression or protein secretion, indicating naturally-occurring agonists of the receptor were not present in the extract. However, QE2 (6 – 24 hrs) potently increased mRNA expression of FXR to 9.95 ± 2.8 fold of that in control cells (n = 7; p < 0.05). Furthermore, QE2 treatment enhanced FGF-19 secretion in response to subsequent exposure to the FXR agonist, GW4064 (5 μM), to 144.8 ± 31.1 pg/ml compared to 29.3 ± 7.7 pg/ml in cells treated with GW4064 alone (n = 6; p = 0.01). HPLC analysis of the lipid extract revealed it to predominantly contain the polyunsaturated fatty acids (PUFAs), a-linolenic acid (ALA, 41%) and linoleic acid (LA, 52%). Similar to QE2, treatment of T 84 cells with ALA enhanced FXR mRNA and protein expression to 11.43 ± 2.1 fold (n = 7, p < 0.01) and 6.4 ± 1.2 fold (n = 6, p < 0.05) of that in untreated cells, respectively. ALA treatment also enhanced the effects of GW4064 on both mRNA and protein expression of FGF19 by 4.4 ± 1.6 (n = 7) and 6.5 ± 1.4 fold (n = 8, p < 0.05), respectively. LA exerted similar effects on FXR expression and activity (data not shown). Treatment with ALA (3 hrs) also increased expression of the PPAR target gene, ANGPTL4, to 66.3 ± 9.5 fold (n = 7; p < 0.01) of that in untreated cells. Furthermore, the effects of ALA on FXR were mimicked by the PPARγ and PPARα agonists, rosiglitazone (1 μM) and WY14643 (10 μM), respectively. Finally, the PPARγ and PPARα antagonists, GW9662 (20 µM) and GW6471 (1 µM), both inhibited ALA-induced FXR mRNA expression to 0.5 ± 0.1 fold (n = 11; p < 0.001) and 0.29 ± 0.1 fold (n = 4; p < 0.05) of that in control cells, respectively. Conclusion: These data indicate that lipid-based plant extracts have the capacity to enhance FXR expression and activity in colonic epithelial cells and that this effect is likely mediated, at least in part, by PUFAs acting through PPARs. Our findings suggest that dietary plant extracts containing PUFAs have potential for development as a new class of “FXR-targeted nutraceutical” for the treatment and prevention of UC ad CRC. This work was supported by an Investigator Award (Grant #: 16-A-4445) from Science Foundation Ireland. This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.

The complex microbial community residing in the human gut has long been understood to regulate gastrointestinal physiology and to participate in digestive diseases, but its extraintestinal actions and influences are increasingly recognized. This article discusses bidirectional interactions between the gut microbiome and athletic performance, metabolism, longevity and the ability of the gut-brain axis to influence cognitive function and mental health.

Lithocholic acid (LCA) is a secondary bile acid formed in the colon by bacterial metabolism of chenodeoxycholic acid. Our previous studies have shown the bile acid to be protective under conditions of colonic inflammation. Here, we sought to determine if LCA might also play a role in regulating fluid and electrolyte transport. T84 cell monolayers were mounted in Ussing chambers for measurements of Cl− secretion, the primary driving force for fluid secretion into the colon. qRT-PCR and Western blotting were used to analyze mRNA and protein expression. Results were expressed as mean ± SEM and data were analyzed by one way ANOVA and Tukey’s post hoc test or by mixed-effects analysis and Dunnett's post hoc test. To assess the effects of LCA on CFTR promoter activity, we used a luciferase promoter/reporter system, where HEK293 cells were transfected with a plasmid containing the firefly luciferase gene under control of the CFTR promoter. Co-transfections were carried out with 2 additional plasmids expressing either the nuclear bile acid receptor, FXR, or its dimerization partner, RXR. Pretreatment of T84 cell monolayers with LCA inhibited subsequent Cl− secretory responses to the cAMP-dependent agonist, forskolin (FSK; 10 μM), in a concentration (1 – 10 μM) and time-dependent (3 - 24 hrs) manner. Maximal effects of LCA were observed at a concentration of 10 μM after treatment for 24 hrs, when responses to FSK were reduced to 50.9 ± 8.5% of those in controls (n = 6; p < 0.01). LCA (10 μM; 24 hrs) also inhibited responses to the Ca2+-dependent secretagogues, thapsigargin (2 μM) and histamine (100 μM), by 59.4 ± 2.4% (n = 4; p < 0.001) and 52.2 ± 1.9% (n = 5; p < 0.001), respectively. In further experiments, using nystatin-permeabilized T84 monolayers to isolate apical Cl− conductances, LCA (10 μM; 24 hrs) reduced FSK-stimulated responses to 72.7 ± 6.6% (n = 17; p < 0.001) of those in control cells. Analysis of CFTR expression, the primary exit pathway for Cl− in colonic epithelial cells, revealed that LCA treatment reduced mRNA and protein expression of the channel to 0.65 ± 0.05 fold (n = 7; p < 0.01) and 0.43 ± 0.06 fold (n = 6; p < 0.001), respectively. In CFTR promoter assays, LCA (10 μM) reduced CFTR promoter activity to 0.7 ± 0.02 fold of that in control cells (n = 5; p < 0.01), with co-expression of FXR being required for this effect to be observed. Finally, while LCA activated both FXR and the vitamin D receptor (VDR) in T84 cells, its effects in downregulating CFTR expression and Cl− conductances were mimicked by the FXR agonist, GW4064, but not by the VDR agonist, calcitriol. In conclusion, LCA, at physiologically relevant concentrations, inhibits Cl− secretion across colonic epithelial cells, likely through a mechanism involving FXR activation and inhibition of CFTR expression. These data add to the growing pool of knowledge regarding important regulatory actions of LCA in the colon and highlight its potential role as a target for the treatment of intestinal disorders. This work was supported by a grant from Science Foundation Ireland. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Introduction: The nuclear bile acid receptor, farnesoid X receptor (FXR), is expressed on intestinal epithelial cells where its potential as a new drug target for treatment of intestinal and metabolic disorders, such as chronic diarrhoea, colitis, colon cancer, obesity and diabetes, has been well-established. We have previously shown that pentacyclic triterpenes (PCTs), a class of dietary phytochemical, enhance the expression and activity of colonic epithelial FXR. Here, we investigated if another common class of dietary phytochemical, polyunsaturated fatty acids (PUFAs), also modulate FXR signalling in these cells. Furthermore, since peroxisome proliferator-activated receptors (PPARs) have been shown to regulate FXR expression in other cell types, we investigated their potential in mediating the effects of PCTs and PUFAs on FXR expression in the colonic epithelium. Methods: All experiments were conducted on monolayers of T 84 colonic epithelial cells grown on permeable supports. Alpha-linolenic acid (ALA; 100 μM) and hederagenin (HG; 5 μM) were used as prototypical PUFAs or PCTs, respectively. Rosiglitazone (1 μM) and WY14643 (1 – 20 μM) were employed as PPARγ and PPARα agonists, whereas GW9662 (20 μM) and GW6471 (1 μM) were used as PPARγ and PPARα antagonists, respectively. Levels of FXR, FGF19, an index of FXR activation, and ANGPTL4, a PPAR target gene, were measured by qRT-PCR, western blotting, or ELISA. Results: Similar to our previous studies with HG, treatment of T 84 cells with ALA upregulated FXR mRNA and protein expression to 11.43 ± 2.1 fold (n = 7, p < 0.01) and 6.4 ± 1.2 fold (n = 6, p < 0.05) of that in control cells, respectively. ALA treatment also enhanced the effects of an FXR agonist, GW4064 (5 μM), on FGF19 mRNA and protein expression by 4.4 ± 1.6 (n = 7) and 6.5 ± 1.4 fold (n = 8, p < 0.05), respectively. Treatment with HG and ALA (3 hrs) also increased mRNA expression of the PPAR target gene, ANGPTL4, to 5.7 ± 1.0 (n = 5; p < 0.05) and 66.3 ± 9.5 fold (n = 7; p < 0.01) of that in untreated cells, respectively. Furthermore, the effects of the phytochemicals on FXR expression were mimicked by the PPARγ and PPARα agonists, rosiglitazone and WY14643, respectively. The PPARγ antagonist, GW9662, inhibited ALA-induced FXR mRNA expression by 0.5 ± 0.1 fold (n = 11; p < 0.001) but did not alter responses to HG. In contrast, the PPARα antagonist, GW6471, inhibited both ALA and HG-induced FXR mRNA expression by 0.71 ± 0.1 and 0.58 ± 0.1 fold (n = 4; p < 0.05) of that in control cells, respectively. Conclusion: The dietary phytochemicals, HG and ALA, upregulate colonic epithelial FXR through mechanisms that appear to involve differential activation of PPARs. By virtue of their ability to upregulate FXR expression and activity, foods or food supplements, rich in such PCTs and PUFAs have excellent potential for development as a new class of “FXR-targeted nutraceutical” for treatment and prevention of intestinal and metabolic disorders. Science Foundation Ireland This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Background: Increased epithelial cell death leading to compromised intestinal barrier function is a key contributor to the pathogenesis of inflammatory bowel disease. Previously published studies suggest that the nuclear bile acid receptor, farnesoid X receptor (FXR), promotes intestinal barrier function and is protective against colonic inflammation. Here, we investigated potential mechanisms involved. Methods: Mucosal inflammation was induced in mice by adding 2.5% dextran sulfate sodium (DSS) to their drinking water, either with or without daily oral gavage with the FXR agonist, obeticholic acid (OCA; 10 mg/kg). After 6 days, mice were administered FITC‐dextran (6 mg/kg) and then sacrificed 24hrs later. The severity of colonic mucosal inflammation was assessed by disease activity index (DAI) and mucosal permeability to FITC. Epithelial apoptosis was assessed by immunohistochemical imaging of cleaved caspase 3. To model the effects of cytokine-induced barrier dysfunction in vitro, we employed polarized monolayers of T 84 colonic epithelial cells. Results were expressed as mean ± SEM and data were analyzed by one-way ANOVA, two-way ANOVA, and the Tukey’s post hoc test. Results: In mice, OCA treatment decreased the DSS-induced DAI score from 11.8 to 9.0 ± 0.7 (*p ≤ 0.05, n=6), increases in mucosal FITC flux by 62.3 ± 0.4% (*p ≤ 0.05, n=6), and epithelial caspase 3 cleavage by 68.1 ± 8.5% (n=6). In vitro studies revealed that treatment of T 84 cells with the pro-inflammatory cytokines, IFNγ (10ng/ml) and TNFα (10ng/ml), induced apoptosis, as evidenced by increased levels of the apoptotic markers, cleaved PARP and cleaved caspase 3. However, pre-treatment of the cells with the FXR agonist, GW4064 (5μM), did not prevent cytokine-induced apoptosis. Treatment of T 84 cells with IFNγ, TNFα and the apoptosis inhibitor, Q-VD-OPh, induced a necroptotic response, as evidenced by increased levels of phosphorylated RIP3 (pRIP3). Co-treatment with Q-VD-OPh also enhanced cytokine-induced transepithelial FITC flux to 3.0 ± 0.2 fold of that in control cells, whereas pre-treatment with the necroptosis inhibitor, necrostatin (200μM), reduced pRIP3 expression by 53.2 ± 3.4% (n=3) and FITC flux by 30.9 ± 0.2% (**p ≤ 0.01, n=10) of that in cells treated with Q-VD-OPh/IFNγ/TNFα alone. Similar to necrostatin, FXR activation with GW4064 inhibited both pRIP3 expression and FITC flux by 47.5 ± 9.1% (n=4) and 46.7 ± 0.6% (**p≤0.01, n=6), respectively, in this in vitro model of cytokine-induced necroptosis. Conclusion: Our studies show that FXR activation protects against intestinal inflammation, an effect that is likely due to preservation of epithelial barrier function. The protective effects of FXR activation on epithelial barrier function may be due to inhibition of necroptosis rather than apoptosis. Our data suggest that FXR represents a promising target for the development of new approaches to prevent epithelial barrier dysfunction in conditions of intestinal inflammation. Science Foundation Ireland (SFI) This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Activation of the nuclear bile acid receptor, farnesoid X receptor (FXR), specifically upregulates ACE2 expression in undifferentiated colonic epithelial cells and inhibits virus-induced proinflammatory cytokine release. By virtue of these actions FXR represents a promising target for the development of new approaches to prevent intestinal manifestations of SARS-CoV-2 infection.

Introduction: While better known for its pulmonary symptoms, SARS-CoV-2 also adversely affects the gastrointestinal tract, causing diarrhoea with evidence of inflammation. The nuclear bile acid receptor, farnesoid x receptor (FXR), is expressed in colonic epithelial cells and has been previously shown to inhibit cytokine production and promote barrier function, thereby preventing inflammatory responses in the gut. In the current studies, we set out to investigate a potential role for FXR in modulating epithelial responses to SARS-CoV-2. Methods: Undifferentiated human colonic or ileal enteroids, colonic epithelial T 84 , or Caco-2 cells were all grown as monolayers on permeable supports. Cells were treated with the FXR agonists, obeticholic acid (OCA, 10 μM) or GW4064 (5 μM), or infected with live SARS-CoV-2 (2019-nCoV/USA_WA1/2020). Changes in cellular mRNA, protein or secreted cytokines were measured by qPCR, western blotting, or ELISA. SARS-CoV-2 levels were quantified by qPCR using primers for envelope (E) and nucleocapsid (N) protein. Data are expressed as mean ± SEM and statistical analysis was performed using paired t-tests or one-way ANOVA with Dunnett’s or Tukey’s multiple comparisons tests. Results: Treatment of colonic or ileal enteroids with the FXR agonist, OCA, increased expression of mRNA for the SARS-CoV-2 receptor, ACE2, by 2.1 ± 0.4 (n = 3; p = 0.08) and 2.3 ± 0.2 (n = 3; p < 0.05) fold, respectively. Similarly, treatment of T 84 cells with another FXR agonist, GW4064, increased expression of ACE2 mRNA with a maximal response of 1.8 ± 0.2 fold (n = 8; p < 0.01) occurring after 72 hrs. Increased mRNA was accompanied by a 2.3 ± 0.7 fold (n = 6; p < 0.01) increase in cellular protein expression and a 1.4 ± 0.1 fold (n = 9; p < 0.001) increase in ACE2 protein secretion into the apical medium. Effects of FXR on ACE2 expression were confirmed in monolayers of Caco-2 cells where GW4064 induced a 1.5 ± 0.1 fold (n = 3; p < 0.001) increase in mRNA for the receptor. GW4064 treatment did not affect mRNA levels for SARS-CoV-2 E or N protein in the infected Caco-2 cells. However, further experiments revealed that treatment with GW4064 inhibited release of the proinflammatory cytokine, IL-6, from either Caco-2 cells infected with SARS-CoV-2 or from T 84 cells treated with the viral mimic, polyinosinic-polycytidylic acid (poly (I:C)) (25 μg/ml) by 46 ± 12 % (n = 3, p < 0.05) and 35 ± 6 % (n = 8; p < 0.01), respectively. Conclusion: Our data suggest that FXR activation upregulates expression of ACE2 in intestinal and colonic epithelial cells but did not directly affect SARS-CoV-2 cellular levels. Rather, FXR activation inhibits viral-induced proinflammatory cytokine secretion and may therefore represent a good target for the development of new approaches to alleviate inflammatory diarrhoea associated with SARS-CoV-2 infection. This work was funded by grants from Science Foundation Ireland and the National Institutes of Health (DK047987). This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

One of the primary functions of the intestinal epithelium is to transport fluid and electrolytes to and from the luminal contents. Under normal circumstances, absorptive and secretory processes are tightly regulated such that absorption predominates, thereby enabling conservation of the large volumes of water that pass through the intestine each day. However, in conditions of secretory diarrhea, this balance becomes dysregulated, so that fluid secretion, driven primarily by Cl- secretion, overwhelms absorptive capacity, leading to increased loss of water in the stool. Secretory diarrheas are common and include those induced by pathogenic bacteria and viruses, allergens, and disruptions to bile acid homeostasis, or as a side effect of many drugs. Here, we review the cellular and molecular mechanisms by which Cl- and fluid secretion in the intestine are regulated, how these mechanisms become dysregulated in conditions of secretory diarrhea, currently available and emerging therapeutic approaches, and how new strategies to exploit intestinal secretory mechanisms are successfully being used in the treatment of constipation.

INTRODUCTIONBile acids, classically known for their roles in fat digestion, are now recognised as important regulators of many aspects of intestinal physiology, including epithelial proliferation/apoptosis, transport and barrier function. Thus, the nuclear bile acid receptor, farnesoid X receptor (FXR), represents a promising therapeutic target for intestinal disorders, such as inflammatory bowel disease and colorectal cancer. Previous studies suggest dietary plant‐derived phytochemicals may modulate FXR activity.AIMSTo investigate the effects of a phytochemical, denoted KFS1, and a KFS1 rich plant extract, denoted as QE1, on FXR signalling in colonic epithelial cells.METHODST84 colonic epithelial cells, grown as polarised monolayers, were treated bilaterally with the FXR agonist, GW4064 (5 μM), KFS1 (1 – 100 μM) or QE1 (30 – 100 μg/ml) for 24 hr. Expression of FXR and FGF‐19, an index of FXR activation, were measured by qRT‐PCR, western blotting and ELISA. Transepithelial electrical resistance (TEER) was measured as an index of epithelial barrier function. Lactate dehydrogenase (LDH) release and PARP cleavage were used as measurements of necrosis and apoptosis, respectively. FXR activity was also studied using an FXR/luciferase reporter HepG2 cell line. Data are expressed as the mean ± SEM for a series of n experiments.RESULTSGW4064 (5 μM) induced a 719.8 ± 183.3 fold increase in FGF‐19 mRNA expression (n = 20; p<0.001) and a 10.4 ± 4.9 fold increase in FGF‐19 protein release (n = 5; p<0.01). KFS1 (5 – 100 μM) did not alter FGF‐19 protein expression and at higher concentrations (50 – 100 μM) reduced TEER. Similarly, GW4064 (5 μM) significantly induced luciferase activity in FXR HepG2 reporter cells by 1580.7 ± 195.7 counts per second (p<0.001; n = 3), whereas treatment with KFS1 (1 – 100 μM) was without effect. Lower concentrations of KFS1 (5 μM), which alone did not alter TEER, increased FXR mRNA and protein expression by 4.2 ± 0.4 (p<0.05; n = 4) and 1.5 ± 0.04 fold (p<0.05; n = 4), respectively. Moreover, pre‐treatment of T84 cells with KFS1 (1, 5, or 10 μM) for 1 hr prior to treatment with GW4064 (5 μM; 24 hr) potentiated GW4064‐induced FGF‐19 mRNA expression by 2.5 ± 0.8, 2.9 ± 0.2, and 4.6 ± 0.5 fold, respectively (p<0.05; n = 6) and FGF‐19 protein expression by 2.5 ± 0.4, 4.1 ± 0.5, and 4.6 ± 0.5 fold, respectively (p<0.05, p<0.001; n = 7). Combined treatment with KFS1 (1, 5, 10 μM) and GW4064 (5 μM) also significantly reduced TEER to 49.8 ± 7.4%, 50.6 ± 8.3%, and 40.6 ± 6.6% of control values, respectively (n = 7). TEER reductions in response to such co‐treatments were not associated with significant LDH release or PARP cleavage (n = 3). Finally, a methanolic extract of plant material known to be rich in KFS1, QE1 (100 μg/ml), increased GW4064‐induced FGF‐19 release in T84 cells by 2.1 ± 0.3 fold (n = 6; p<0.01).CONCLUSIONThese data demonstrate that KFS1, a common dietary phytochemical, induces FXR expression in colonic epithelial cells and prime the cells for agonist‐induced FXR activation. These findings suggest that foods, or food supplements, rich in such plant‐derived phytochemicals have potential for development as FXR‐targeted neutraceuticals.Support or Funding InformationStAR PhD Scholarship from RCSI & Science Foundation Ireland (SFI)

The intestinal epithelium forms the interface between the body and the luminal contents, facilitating fluid and electrolyte transport, while also acting as a barrier to harmful pathogens. Dysregulation of barrier and transport function is associated with the pathogenesis of a number of conditions, including chronic diarrhoeal and inflammatory bowel diseases (IBD). Previous studies from ours, and other laboratories, have identified the nuclear bile acid receptor, farnesoid x receptor (FXR), as an excellent target for the development of new anti‐diarrhoeal and anti‐inflammatory therapeutics. Polyunsaturated fatty acids (PUFAs), such as alpha‐linolenic acid (ALA), are a group of bioactive phytochemicals, which have been shown in other systems to modulate FXR activity. PUFAs are also thought to be ligands for another group of nuclear receptors, known as the peroxisome proliferator‐activated receptors (PPARs), which have been recently shown to modulate FXR expression. Here, we set out to investigate a possible role for ALA in regulating farnesoid x receptor signalling in colonic epithelial cells.

Bile acids (BAs) are known to be important regulators of intestinal motility and epithelial fluid and electrolyte transport. Over the past two decades, significant advances in identifying and characterizing the receptors, transporters, and ion channels targeted by BAs have led to exciting new insights into the molecular mechanisms involved in these processes. Our appreciation of BAs, their receptors, and BA-modulated ion channels as potential targets for the development of new approaches to treat intestinal motility and transport disorders is increasing. In the current review, we aim to summarize recent advances in our knowledge of the different BA receptors and BA-modulated ion channels present in the gastrointestinal system. We discuss how they regulate motility and epithelial transport, their roles in pathogenesis, and their therapeutic potential in a range of gastrointestinal diseases.

Introduction and AimsThe intestinal epithelium functions to transport nutrients, fluid and electrolytes, while at the same time acting as a barrier to the entry of harmful substances. CFTR is a transmembrane Cl− channel important in regulating intestinal fluid transport and is implicated in the pathogenesis of a number of intestinal diseases. Bile acids, classically known for their roles in lipid digestion, are now also recognised as important enteric hormones that regulate many aspects of epithelial function. Indeed, it is thought that bile acid metabolism is a primary mechanism by which the microbiota communicates with its human host. Here, we set out to investigate the role of the nuclear bile acid receptor, farnesoid x receptor (FXR), in regulating epithelial CFTR expression.MethodsT84 human colonic epithelial cells were cultured as polarised monolayers on permeable supports and treated bilaterally with the FXR agonist, GW4064 (5 μM), over a range of times. Levels of FGF19, an index of FXR activation, CFTR, FXR, FOXA1, and miR‐494 were measured by qRT‐PCR, western blotting, or ELISA.ResultsTreatment of the cells with GW4064 significantly increased FGF19 mRNA expression by 1235 ± 237.3 fold (n = 8; p < 0.05) and protein by 719.6 ± 92.0 pg/mL (n = 5; p < 0.01) after 48 hrs. Moreover, GW4064 downregulated CFTR mRNA to 0.47 ± 0.1 fold after 12 hrs (n = 8; p < 0.01) and protein levels to 0.36 ± 0.1 fold after 48 hrs, compared to vehicle‐treated controls (n = 4; p < 0.05). Transcriptomic analysis confirmed FXR‐induced downregulation of CFTR in primary human colonic enteroids. Studies in Ussing chambers showed that GW4064 treatment for 48 hrs inhibited Cl− secretory responses to the Ca2+‐dependent agonist carbachol (CCh; 100 μM) and the cAMP‐dependent agonist, forskolin (10 μM) by 79.9 ± 7.5 % (n=4; p <0.01) and 74.2 ± 8.9 % (n=4; p < 0.01), respectively. Expression of miR‐494, which is known to target CFTR, was not increased by GW4064 treatment in T84 cells. However, mRNA expression of FOXA1, a transcription factor that regulates CFTR, was inhibited by 33.2 ± 5.2% after 3 hrs (n = 4; p <0.05).ConclusionThe nuclear bile acid receptor, FXR, regulates colonic epithelial CFTR expression. Such actions are likely to be important in the setting of microbial regulation of host intestinal physiology in health and disease. As such, FXR represents an excellent target for development of new drugs to treat intestinal diseases associated with dysregulated CFTR function.Support or Funding InformationThis work was funded by a Science Foundation Ireland Principal Investigator Award to SJK

The nuclear bile acid receptor, farnesoid X receptor (FXR), is an important regulator of intestinal and metabolic function. Previous studies suggest that pentacyclic triterpenes (PCTs), a class of plant-derived bioactive phytochemical, can modulate FXR activity and may therefore offer therapeutic benefits. Here, we investigated the effects of a prototypical PCT, hederagenin (HG), on FXR expression, activity, and antisecretory actions in colonic epithelial cells. T84 cells and murine enteroid-derived monolayers were employed to assess HG effects on FXR expression and activity in colonic epithelia. We measured mRNA levels by qRT-PCR and protein by ELISA and immu-noblotting. Transepithelial Cl-secretion was assessed as changes in short circuit current in Ussing chambers. We determined HG treatment (5-10 mu M) alone did not induce FXR activation but significantly increased expression of the recep-tor, both in T84 cells and murine enteroid-derived monolayers. This effect was accompanied by enhanced FXR activity, as assessed by FGF-15/19 induction in response to the synthetic, GW4064, or natural FXR agonist, chenodeoxycholic acid. Effects of HG on FXR expression and activity were mimicked by another PCT, oleanolic acid. Furthermore, we found FXR-induced downregulation of cystic fibrosis transmembrane conductance regulator Cl- channels and inhibition of trans -epithelial Cl- secretion were enhanced in HG-treated cells. These data demonstrate that dietary PCTs have the capacity to modulate FXR expression, activity, and antisecretory actions in colonic epithelial cells. Based on these data, we propose that plants rich in PCTs, or extracts thereof, have excellent poten-tial for development as a new class of "FXR-targeted nutraceuticals".

Similar to the bile acid receptor, FXR, VDR is expressed in colonic epithelial cells where activation by its natural ligand, calcitriol, downregulates CFTR expression and inhibits agonist-induced Cl- secretory responses. Since Cl- secretion is the primary driving force for intestinal fluid secretion, our data suggest that VDR may serve as a target for the development of new anti-diarrhoeal agents. Future studies will aim to determine the molecular mechanisms involved and investigate a possible role for VDR in mediating anti-secretory actions of luminal bile acids.