Acute liver failure (ALF) is a potentially life-threatening disorder without any effective treatment strategies. d-Galactosamine (GalN)/lipopolysaccharide (LPS)-induced ALF is a widely used animal model to identify novel hepato-protective agents. In the present study, we investigated the potential of a cannabinoid receptor 2 (CB2) agonist, JWH-133 [(6aR,10aR)-3-(1,1-dimethylbutyl)-6a,7,10,10a-tetrahydro-6,6,9-trimethyl-6H-dibenzo[b,d]pyran], in the amelioration of GalN/LPS-induced ALF. JWH-133 treatment protected the mice from ALF-associated mortality, mitigated alanine transaminase and proinflammatory cytokines, suppressed histopathological and apoptotic liver damage, and reduced liver infiltration of mononuclear cells (MNCs). Furthermore, JWH-133 pretreatment of M1/M2-polarized macrophages significantly increased the secretion of anti-inflammatory cytokine interleukin-10 (IL-10) in M1 macrophages and potentiated the expression of M2 markers in M2-polarized macrophages. In vivo, JWH-133 treatment also suppressed ALF-triggered expression of M1 markers in liver MNCs, while increasing the expression of M2 markers such as Arg1 and IL-10. microRNA (miR) microarray analysis revealed that JWH-133 treatment altered the expression of only a few miRs in the liver MNCs. Gene ontology analysis of the targets of miRs suggested that Toll-like receptor (TLR) signaling was among the most significantly targeted cellular pathways. Among the altered miRs, miR-145 was found to be the most significantly decreased. This finding correlated with concurrent upregulated expression of its predicted target gene, interleukin-1 receptor-associated kinase 3, a negative regulator of TLR4 signaling. Together, these data are the first to demonstrate that CB2 activation attenuates GalN/LPS-induced ALF by inducing an M1 to M2 shift in macrophages and by regulating the expression of unique miRs that target key molecules involved in the TLR4 pathway.