〔Abstract〕 Objective To investigate the protective effect of melatonin(MT) on formaldehyde(FA) inhalation-induced acute lung injury(ALI) in rats and its mechanism through the regulation of nuclear factor E2-related factor 2(Nrf2) signaling pathway. Methods Fifty female Wistar rats were randomly divided into Control group, FA group, FA+MT 5 mg/kg group, FA+MT 10 mg/kg group and FA+MT 20 mg/kg group, with 10 rats in each group. Except for the Control group, all other groups inhaled 3 mg/m3FA daily for 21 d consecutively to construct the tainted model, and then treated with different MT doses for 14 d. The tainting was continued during the MT treatment. Hematoxylin-eosin(HE) staining was used to observe the histopathological changes in lung tissue, lung water content and lung coefficient were weighed and measured, glutathione(GSH), superoxide dismutase(SOD) and 8-hydroxydeoxyguanosine(8-OHdG) levels were measured by absorbance photometric method, and enzyme linked immunosorbent assay(ELISA) was used to measure the levels of tumor necrosis factor-alpha(TNF-α), interleukin(IL)-6, and IL-1β concentrations, Western blot to detect the protein expression levels of Nrf2, heme oxygenase-1(HO-1), nuclear factor-κB(NF-κB), and phosphorylated nuclear factor-κB(p-NF-κB) in lung tissues, and quantitative polymerase chain reaction(qPCR) to detect the Nrf2, HO-1, and Kelch-like ECH-associated protein 1(Keap1) mRNA expression levels. Results Compared with the control group, lung injury was obvious in rats in the FA group; lung tissue GSH and SOD levels were reduced, and 8-OHdG levels were elevated(P<0.05); alveolar lavage fluid TNF-α, IL-6, and IL-1β levels were elevated(P<0.05); Nrf2 and HO-1 protein expression levels were reduced in the lung tissue( P<0. 05),and p-NF-κB protein expression levels were was elevated( P<0. 05); the relative mRNA expression of Nrf2 and HO-1 in lung tissue was decreased,and the relative mRNA expression of Keap1 was elevated( P<0. 05). Compared with the FA group,the lung injury of rats in the MT group was improved; the levels of GSH and SOD in the lung tissue were increased( P<0. 05),and the level of 8-OHd G was decreased( P<0. 05); the levels of TNF-α,IL-6,and IL-1β in the alveolar lavage fluid were decreased( P<0. 05); and the expression levels of the Nrf2 and HO-1 proteins in the lung tissue were increased( P<0. 05). p-NF-κB protein expression level was decreased( P<0. 05); the relative mRNA expression levels of Nrf2 and HO-1 in lung tissues were increased( P<0. 05),and the relative mRNA expression level of Keap1 was decreased( P<0. 05) in lung tissues,and all of them were in a dose-dependent manner. Conclusion MT can alleviate oxidative stress and inflammatory responses and mitigate FA exposure-induced acute lung injury by regulating the Nrf2/Keap1/HO-1 signaling pathway.