Abstract

Background: Bile acid (BA) homeostasis is critical for hepatic integrity, with ATP-binding cassette (ABC) transporters Mrp2 (Abcc2) and Bcrp (Abcg2) playing key roles in BA transport. Mrp2 functions as the primary canalicular exporter of conjugated BAs, while Bcrp exerts auxiliary effects; however, their age-specific functional differences, crosstalk, and contributions to cholestatic liver injury remain incompletely elucidated. This study aimed to address these knowledge gaps using genetic knockout models.
Methods: Four mouse genotypes (wild-type [WT], Mrp2 ^⁻/⁻ , Bcrp ^⁻/⁻ , and Mrp2 ^⁻/⁻ -Bcrp ^⁻/⁻ ) were analyzed across three age cohorts (10, 20, and 40 weeks). Organ weights and indices were calculated; BA profiles in liver, bile, plasma and feces, and the expression of BA synthetic/metabolic enzymes (Cyp7a1, Cyp27a1, Cyp8b1, Baat, Bacs) was quantified using UHPLC-MS/MS method; and liver histopathology was evaluated via hematoxylin-eosin (H&E) staining.
Results: Mrp2 ^⁻/⁻ mice exhibited significantly increased liver weight and liver index across all ages, accompanied by marked hepatic accumulation of conjugated BAs (e.g., 13.4-fold elevation of taurochenodeoxycholic acid [T-CDCA] at 20 weeks) and upregulated expression of Cyp27a1, Cyp8b1, Baat, and Bacs (1.4–1.9-fold vs. WT). They also developed progressive liver injury, including macrovesicular steatosis and focal inflammatory infiltration, which worsened at 40 weeks. In contrast, Bcrp ^⁻/⁻ mice showed only transient liver weight increase at 10 weeks and minimal alterations in BA profiles or enzyme expression. Notably, Mrp2 ^⁻/⁻ -Bcrp ^⁻/⁻ mice displayed milder hepatic pathology than Mrp2 ^⁻/⁻ mice. BA dysregulation was age-dependent, with peak perturbations at 10 weeks and partial compensation (e.g., via upregulated basolateral transporters) in later cohorts.
Conclusions: We constructed double gene knockout mice and established a UHPLC-MS/MS method for detecting the expression of enzymes related to BA synthesis and metabolism in mice. The results indicate that Mrp2 is indispensable for canalicular export of conjugated BAs and maintenance of gut-liver axis homeostasis, as its deficiency drives toxic BA accumulation and progressive liver injury. Bcrp plays a secondary, age-restricted role in BA transport and may exert protective effects in Mrp2-deficient contexts. These findings highlight Mrp2 as a potential therapeutic target for cholestatic liver diseases such as primary biliary cholangitis (PBC).