Aim: To elucidate the role of Toll-like receptors (TLRs) in gastric cytoprotection after ethanol injury. Methods: C57BL/6J, C3H/HeOuJ and C3H/HeJ mice were used. All mice were killed 4h after ethanol administration. TLR4, cyclooxygenase-2 (Cox-2) and prostaglandin E2 (PGE2) expression were measured by immunohistochemistry, western blotting and enzyme-linked immunosorbent assay (ELISA) separately. Results: The expression of TLR4 increased in C57BL/6J mice stomach 4h after ethanol injury. The cells expressing TLR4 included Cox-2 expressing cells and macrophages. The injury in C3H/HeJ mice was more severe than in C3H/HeOuJ mice 4h after ethanol injury. The expression of Cox-2 and PGE2 only increased in C3H/HeOuJ mice. The number of macrophages and the expression of macrophage-inflammatory protein-2 (MIP-2) also increased only in C3H/HeOuJ mice. Conclusion: TLR4 signal is activated in mice stomach 4h after ethanol injury. The protective effects of TLR4 signalling are mediated through the induction of Cox-2 expression and the production of PGE2. © 2009 Editrice Gastroenterologica Italiana S.r.l.