Objective To evaluate the effect of ambroxol on p38 mitogen - activated protein kinase pathway in mice with ventilator-induced lung injury(VILI). Methods Thirty male SD mice, weighing 280～320 g, were equally and randomly divided into 3 groups (n=10 each) using a random number table: control group (group C), ventilator-induced lung injury group (group VILI), and ventilator-induced lung injury + ambroxol group (group AMB). VILI was induced by 4 h mechanical ventilation with tidal volume 40 mL/kg. Ambroxol 50 mg/kg preconditioning was injected intraperitoneally for 3 days in group AMB, while the equal volume of normal saline was given in C and VILI groups. The mice were then sacrificed, the arterial blood gas was detected, and broncho-alveolar lavage fluid (BALF) was collected for determination of the concentrations of total protein, interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), IL-6 and intercellular adhesion molecule-1 (ICAM-1). The lung tissues were removed for determination of wet to dry lung weight ratio (W/D ratio), expression of p-p38 MAPK, IL-1β mRNA, TNF-α mRNA, IL-6 mRNA and ICAM-1 mRNA, and for examination of the pathological changes which were scored. Results Compared with group C, partial pressure of oxygen in arterial blood(PaO₂) was decreased (P＜0.05), W/D ratio, lung injury score, concentrations of total protein, IL-1β, TNF-α, IL-6 and ICAM-1 in BALF, and expression of p-p38 MAPK, IL-1β, TNF-α, IL-6 and ICAM-1 mRNA were significantly increased in VILI and AMB groups (P＜0.05). Compared with group VILI, PaO₂ was increased (P＜0.05), W/D ratio, lung injury score, concentrations of total protein, IL-1β, TNF-α, IL-6 and ICAM-1 in BALF, and expression of p-p38 MAPK, IL-1β, TNF-α, IL-6 and ICAM-1 mRNA were decreased in group AMB (P＜0.05). Conclusion Ambroxol can attenuate VILI probably through inhibiting p38 mitogen - activated protein kinase pathway in mice.