BACKGROUND: Mechanical ventilation (MV) can induce lung injury. Proinflammatory cytokines have been shown to play an important role in the development of ventilator-induced lung injury. Previously, the authors have shown a role for Toll-like receptor 4 signaling. The current study aims to investigate the role of Toll/interleukin-1 receptor domain-containing adapter-inducing interferon-β (TRIF), a protein downstream of Toll-like receptors, in the development of the inflammatory response after MV in healthy mice. METHODS: Wild-type C57BL6 and TRIF mutant mice were mechanically ventilated for 4 h. Lung tissue and plasma was used to investigate changes in cytokine profile, leukocyte influx, and nuclear factor-κB activity. In addition, experiments were performed to assess the role of TRIF in changes in cardiopulmonary physiology after MV. RESULTS: MV significantly increased messenger RNA expression of interleukin (IL)-1β in wild-type mice, but not in TRIF mutant mice. In lung homogenates, MV increased levels of IL-1α, IL-1β, and keratinocyte-derived chemokine in wild-type mice. In contrast, in TRIF mutant mice, only a minor increase in IL-1β and keratinocyte-derived chemokine was found after MV. Nuclear factor-κB activity after MV was significantly lower in TRIF mutant mice compared with wild-type mice. In plasma, MV increased levels of IL-6 and keratinocyte-derived chemokine. In TRIF mutant mice, no increase of IL-6 was found after MV, and the increase in keratinocyte-derived chemokine appeared less pronounced. TRIF deletion did not affect cardiopulmonary physiology after MV. CONCLUSIONS: The current study supports a prominent role for TRIF in the development of the pulmonary and systemic inflammatory response after MV.