Background: Probiotic VSL#3 is used to treat ulcerative colitis. This study examines the effect of VSL#3 in non-alcoholic steatohepatitis (NASH) that has liver carcinogenic potential. Methods: Weste diet (WD)-fed wild-type (WT) mice that do not have hepatic inflammation with lymphocyte infiltration and carcinogenic potential were used for baseline comparison. Age-, sex-, and diet-matched bile acid (BA) receptor faesoid X receptor (FXR) knockout (KO) mice, which developed severe NASH and had the potential for liver cancer development, were supplemented with and without VSL#3 for 7 months. All the mice were euthanized when they were 10 months old. Results: Supplementation with VSL#3 completely abolished hepatic lymphocyte infiltration, reduced hepatic fat content, and improved insulin sensitivity in WD-fed FXR KO mice. In addition, VSL#3 normalized dysregulated BA homoeostasis by inhibiting the classical BA synthesis pathway, inducing the alteative BA pathway, and activating ileal G-protein coupled BA receptor 1 (GPBAR1)-regulated signaling. Moreover, VSL#3 reconstructed the gut microbiota by reducing Bacteroidaceae , Porphyromonadaceae , and Helicobacteraceae as well as increasing Lachnospiraceae. Further, VSL#3 enriched the abundance of Ruminococcus and Faecalibacterium , which generate butyrate, at the genus level. It also increased the copy number of the butyrate-producing genes bcoA and buk, suggesting their anti-inflammatory and metabolic effects. Conclusions: VSL#3 is useful in reversing NASH that occurred due to dysregulated BA synthesis and dysbiosis, suggesting its potential in liver cancer prevention.