Hypothesis

Sex‑specific overexpression of the high‑activity bile salt hydrolase subtype BSH‑T1 in aging males drives hepatic accumulation of the FXR antagonist tauro‑β‑muricholic acid (T‑β‑MCA), thereby suppressing FXR signaling and accelerating age‑related liver steatosis and fibrosis.

Mechanistic Rationale

Aging reshapes gut microbiota in a sex‑dependent manner 1. In old males, metagenomic analyses reveal enrichment of BSH‑T1 phylotypes harbored by Blautia and Roseburia 1. Although BSH enzymes deconjugate primary bile acids, kinetic studies show that BSH‑T1 exhibits a strong preference for tauro‑conjugated substrates and is prone to product inhibition by liberated taurine, leading to a net decrease in overall deconjugation activity 2. This kinetic quirk causes a selective buildup of tauro‑β‑muricholic acid (T‑β‑MCA), a potent endogenous FXR antagonist 3. Elevated hepatic T‑β‑MCA competitively inhibits FXR, reducing expression of FXR‑target genes involved in fatty acid oxidation and bile acid homeostasis, thereby promoting steatosis 3. In contrast, aging females exhibit a shift toward BSH phylotypes with lower tauro‑specific activity, resulting in lower hepatic T‑β‑MCA and relatively preserved FXR signaling 1.

Testable Predictions

1. Hepatic T‑β‑MCA concentrations will be significantly higher in old male mice than in old female mice, correlating with hepatic BSH‑T1 abundance.
2. Genetic or pharmacological suppression of BSH‑T1 activity in aged males will lower hepatic T‑β‑MCA, restore FXR target gene expression (e.g., Shp, Cyp7a1), and reduce steatosis and fibrosis markers.
3. Transplanting fecal microbiota enriched for BSH‑T1 from old males into young germ‑free recipients will recapitulate the male‑specific increase in hepatic T‑β‑MCA and impair FXR signaling.
4. In a human aging cohort, fecal metagenomic abundance of BSH‑T1 genes will positively associate with serum T‑β‑MCA levels and non‑invasive fibrosis scores (e.g., FIB‑4), independent of sex after stratification.

Experimental Design

• Mouse study: Use 20‑month‑old C57BL/6J males and females. Quantify fecal BSH‑T1 copies by qPCR targeting the T1‑specific allele 2. Measure hepatic BA pool by LC‑MS/MS, focusing on T‑β‑MCA. FXR activity assessed via reporter luciferase assay in isolated hepatocytes and expression of Shp, Cyp7a1. Intervention: deliver an antisense oligonucleotide or CRISPRi plasmid targeting the bshT1 gene via oral gavage; control groups receive scrambled oligo. Outcomes: histology (Steatosis, fibrosis via Sirius Red), serum ALT/AST, and metabolic profiling.
• Microbiota transplant: Fecal material from old male donors (high BSH‑T1) vs old female donors (low BSH‑T1) transplanted into 8‑week‑old germ‑free males and females; monitor hepatic T‑β‑MCA and FXR readouts over 12 weeks.
• Human pilot: Recruit 120 participants aged 60‑80, stratified by sex. Collect stool for shotgun metagenomics to compute BSH‑T1 RPKM, serum for targeted BA metabolomics (T‑β‑MCA), and liver fibrosis assessment via transient elastography (FibroScan) and serum biomarkers.

Potential Confounds and Falsifiability

Variations in diet, antibiotic exposure, or baseline liver disease could influence BA pools; therefore, all mouse cohorts will be pair‑fed a standardized chow and human participants will complete detailed food frequency questionnaires. If hepatic T‑β‑MCA does not rise despite increased BSH‑T1 abundance, or if BSH‑T1 knockdown fails to modify FXR signaling, the hypothesis would be falsified, prompting investigation of alternative BSH subtypes or host‑mediated BA modifications.