Farnesoid X receptor activation by the novel agonist TC-100 (3α, 7α, 11ß-Trihydroxy-6α-ethyl-5ß-cholan-24-oic Acid) preserves the intestinal barrier integrity and promotes intestinal microbial reshaping in a mouse model of obstructed bile acid flow.

The intestinal tract hosts the gut microbiota (GM), actively shaping health. Bile acids(BAs) are both digestive and signaling molecules acting as hormones via the activation of farnesoid X receptor (FXR). Obstruction of bile flow initiates a cascade of pathological events ultimately leading to intestinal mucosal injury. Administration of BAs in models of obstructed bile flow counteracts these detrimental effects. Objective of this study was to investigate the effects of the novel FXR agonist 3α, 7α, 11ß-Trihydroxy-6α-ethyl-5ß-cholan-24-oic Acid (TC-100) on intestinal mucosa integrity and cecal microbiome composition after surgical bile duct ligation (BDL), a rodent model causing bile flow obstruction. Pharmacological FXR activation was accomplished by daily oral gavage with TC-100 for 5 days. 2 days after treatment initiation, BDL was performed. BAs measurement was carried out and the 16S rDNA (V5-V6 hyper-variable regions) extracted from the cecal content was sequenced. TC-100 activates Fxr in the gut-liver axis and this translated into a significant reduction of serum and bile BA pool size with a shift to a more hydrophilic composition, while signs of intestinal mucosal damage were prevented. Firmicutes:Bacteroidota ratio progressively increased from Sham Operated (SO) mice to TC-100-treated mice. LEfSe analysis showed that Verrucomicrobia, and particularly Akkermansia muciniphila (Amuc) increasingly recognized for improving gut homeostasis and immune functions, were strongly associated to TC-100-treated mice. Intriguingly, Amuc abundance was also negatively associated to cholic acid levels. Collectively, these data indicate that intestinal FXR activation by TC-100 prevents early signs of intestinal mucosal damage by modulating BA homeostasis and GM composition.

Identification and quantification of oxo-bile acids in human faeces with liquid chromatography-mass spectrometry: A potent tool for human gut acidic sterolbiome studies.

Bile acids (BAs) are endogenous steroids involved in the transport of lipids in bile, acting also as molecular signaling hormones. Primary BAs synthesized in the liver undergo several metabolic pathways in the intestine by gut microbiota to produce secondary BAs. Together with secondary BAs, other metabolites have been recovered from human faeces, including many oxo-BA analogues produced in the colon through oxidation of BA hydroxy groups. However, the complete oxo-BA characterization in biospecimens (particularly intestinal content and faeces) has not been reported yet, hampering the assessment of their potential physiological role. Herein, we have developed and validated a new RP-HPLC-ESI-MS/MS method in negative ionization mode for the simultaneous analysis of 21 oxo-BAs and their 7 metabolic BAs precursors in human faeces. The elution was performed in gradient mode and 28 compounds, including primary, secondary BAs, and their oxo-derivatives, were separated within 50 min at 40 °C column temperature. The method is accurate (bias% <13%), precise (CV% <10%), with limits of quantification (LOQ <30 ng/mLextract samples), similar for all the studied compounds. The matrix effect does not significantly affect the analysis accuracy, allowing the use of standard solutions for the quantifications, without matrix-matched protocols. Thanks to the high detectability and the relatively high concentration of oxo-BAs (about µg/gwet faeces), the method does not require a pre-analytical clean-up step. This method was used to identify and quantify oxo-BAs in human faecal samples from healthy subjects, serving as a proof of concept for application in patients with hepatobiliary disease and bacteria overgrowth.