Cigarette smoke exposure is the major cause of chronic obstructive pulmonary disease (COPD), which is characterized by enhanced oxidative stress, inflammatory responses and tissue destruction. Acetylshikonin has been reported to exhibit anti-oxidative and anti-inflammatory effect. We hypothesized that acetylshikonin could protect against cigarette smoke induced acute lung injury via its anti-oxidative and anti-inflammatory properties. Acetylshikonin was given by tail vein injection to C57BL/6 mice daily 2 h before 4% cigarette smoke exposure for 1 h over five consecutive days. The lung tissues and bronchoalveolar lavage (BAL) fluid were collected and the tissue damage, inflammatory cytokines and anti-oxidative activity were analyzed. Meanwhile, the RAW264.7 cells were cultured in cigarette smoke treated medium and employed to study the mechanisms of action of acetylshikonin. Acetylshikonin could attenuate smoke-induced lung pathological changes, tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), interleukin-1 beta (IL-1 beta), and monocyte chemoattractant protein 1 (MCP-1) productions, and tissue damages caused by oxidative stress. Furthermore, acetylshikonin was found to enhance the expression of Nrf2 and Nur77-mediated COX-2 in vivo and in vitro. Our results indicated that acetylshikonin exhibited an ameliorative effect on smoke-induced acute lung injury and the possible mechanism was involved in increasing the expression of Nrf2 and Nur77-mediated COX-2.