Excessive activation of the TLR/MyD88 signaling pathway contributes to several inflammation-related diseases. Previously, our laboratory synthesized a novel thiazaol-aminoramification MyD88 inhibitor named TJ-M2010-5. In this study, we interrogated the role of MyD88, as well as the protective effect of TJ-M2010-5, in a D-gal/LPS-induced acute liver injury mouse model. In order to induce acute liver injury, BALB/c mice received intraperitoneal injection of D-gal and LPS at a dose of 800 mg/kg and 80 mu g/kg body weight, respectively. All mice died within 48 h of injection without intervention. However, pre-treatment with TJ-M2010-5 as well as knockout (KO) of the MyD88 gene significantly improved mouse survival rate to 73.3% and 80% at 48 h, respectively, and both treatments protected liver function. These pathological results demonstrated that TJ-M2010-5 and MyD88 KO reduced the infiltration of inflammatory cells and protected hepatocytes against apoptosis. Furthermore, TJ-M2010-5 remarkably inhibited NF-kappa B and MAPK signaling in vivo. LPS-induced activation of macrophages as well as pro-inflammatory factors were also shown to be decreased after TJ-M2010-5 treatment in vivo and in vitro. Taken together, these results suggested that blockage of the TLR/MyD88 signaling pathway by TJ-M2010-5 has an important role in the prevention of inflammation-related acute liver injury.