Non-alcoholic fatty liver disease (NAFLD) is defined as an abnormal hepatic triglycerides (HTG) accumulation, associated to obesity or insulin resistance (IR). The pathogenesis of NAFLD is not fully understood. Bile acids (BA), as endogenous ligands for the nuclear hormone receptor farnesoid X receptor have been implicated in glucose metabolism and IR. The effects of BA or its derivatives on NAFLD are still unclear as ursodeoxycholic acid (UDCA) has shown discrepant effects in human NAFLD. On the other hand, BA sequestration has been shown to improve glucose tolerance but its effects on NAFLD are unclear. We studied the effects of BA supplementation and sequestration on liver steatosis development in genetically obese mice. Methods: ob/ob mice (n=4-6), were fed with standard chow (control) or with chow supplemented with UDCA 0.5%, cholic acid (CA)0.5% or cholestiramine (CTM)3%, since 4 to 8 weeks-old (prevention experiments) or since 8 to 12 weeks-old (reversion experiments). Anesthesia: ketamine (100 mg/Kg) plus xylazine (10 mg/Kg) i.p. Experiments were approved by the local ethic review committee on animal experiments (2). We assessed serum and biliary parameters, glucose tolerance test (GTT), HTG content, liver histology and hepatic gene expression. Results: 12 weeks-old mice exhibited marked obesity, hyperinsulinemia, fasting hyperglycemia and decreased bile flow and biliary glutathione secretion. Administration of both UDCA and CA, but no CTM, significantly reduced histological NAFLD development in prevention experiments and prevented HTG accumulation (controls: 111±19 mg/g liver, UDCA-fed: 61.8±11.0 mg/g liver, CA-fed: 25.4±13.6, p<0.05, (mean±SD; Student’s t-test). A marked steatosis reduction and HTG (UDCA-fed: 53.1±1.4 mg/g liver; CA-fed: 54.5±10.8 mg/g liver) was also seen in established NAFLD (reversion experiments). Although all treated animals exhibited decreased basal glucose levels, GTT improved only after CTM treatment. Gene expression studies showed expected repression of 7-α-hydroxylase (rate limiting enzyme in BA synthesis pathway) in CA and UDCA feeding. Conclusion: BA strongly modulate fatty liver development in mice being able to prevent liver steatosis and reverse established NAFLD. UDCA and CA seem to exert this effect through different pathways. On the other hand, CTM positively affects glucose tolerance but had no effects on hepatic steatosis in this experimental model of NAFLD. More studies are needed to demonstrate the efficacy of novel therapies for NAFLD, linked to the effects of BA on HTG accumulation or the effect of CTM in other pathways related to glucose metabolism (FONDECYT1110455).