<p>Psoriasis is a chronic, systemic immune-mediated skin disease. Although many new strategies for psoriasis treatment have been developed, there is great need in clinic for treating psoriasis. Gentiopicroside (GPS), derived from <i>Gentiana manshurica Kitagawa</i>, has multiple pharmacological activities including anti-inflammatory, anti-oxidative and antiviral activities. In this study, we investigated the potential effects of GPS in imiquimod (IMQ)-induced psoriasis mouse model and the underlying mechanisms. The mice were sensitized on their shaved back with IMQ cream for 7 days with or without topical application of 1% or 2% GPS cream. We showed that the application of GPS cream significantly ameliorated psoriasis-like skin lesions; GPS effect was better than that of calcipotriol. GPS rectified the immune cells infiltration and keratinocytes activation in the skin lesions, and significantly inhibited TNF-α/IFN-γ stimulated human keratinocyte (HaCaT) activation in vitro. Proteomic analysis from keratinocytes with and without GPS treatment prompted that GPS regulated the Keap1-Nrf2 pathway, which was the most important pathway in regulating oxidative stress and inflammation. We demonstrated that GPS regulated the protein expression of p62 and Keap1, induced Nrf2 nuclear translocation followed by transcription of Nrf2 downstream antioxidant genes in HaCaT cells. Furthermore, the antioxidant effects of GPS were abolished in Nrf2^-/- keratinocytes. Simultaneously, Nrf2^-/- mice showed increased psoriasiform symptoms with a diminished protective effect in response to GPS treatment. Collectively, the study discloses that GPS inhibits keratinocyte activation and ameliorates psoriasis-like skin lesions in an Nrf2-dependent manner.</p>