Osteoarthritis (OA) is an inflammatory disorder dealing with the focal degradation of articular cartilage. Oxidative stress and inflammation are the major events in OA. The present study aimed at identifying the mechanism of the potent antioxidant, plumbagin, in protecting against hydrogen peroxide (H2O2)-induced chondrocyte oxidative stress and inflammatory signaling. Oxidative stress was determined by measuring reactive oxygen species, lipid peroxidation, non-enzymic (glutathione; GSH) and enzymic antioxidant activities (GSH, glutathione S-transferase, glutathione peroxidase, superoxide dismutase, catalase). Expression levels of nuclear factor (erythroid-derived 2)-like 2 (Nrf-2), heme oxygenase 1 (HO-1), NAD(P)H: quinone oxidoreductase 1 (NQO-1), nuclear factor-kappa B (NF-kappa B), cyclooxygenase-2 (COX-2) and inducible NO synthase (iNOS) were determined by western blot analysis. Pro-inflammatory cytokine expression levels were assessed using ELISA. Results from reactive oxygen species generation, lipid peroxidation content and antioxidant enzyme activities demonstrated that plumbagin significantly inhibited oxidative stress status in H2O2-induced chondrocytes. In addition, plumbagin modulated transcription factors involved in redox and inflammation regulation, including NF-kappa B and Nrf-2, by nuclear expression. plumbagin enhanced antioxidant status by increasing the expression levels of Nrf-2 target genes, including HO-1 and NQO-1. An anti-inflammatory effect against chondrocyte-induced inflammation was demonstrated by downregulating COX-2, iNOS and pro-inflammatory cytokine expression levels (tumor necrosis factor-alpha, interleukin (IL)-6 and IL-8). The present study identified strong evidence for a protective role of plumbagin against H2O2-induced oxidative stress and inflammation in chondrocytes by modulating redox signaling transcription factors.