Osteoarthritis (OA) is a chronic degenerative disease associated with inflammation and degradation of articular cartilage and is typically considered an age-related disease. In OA patients, persisting pain reduces quality of life. However, the mechanism underlying OA development remain unclear. Recent studies have reported that mitochondrial dysfunction occurs with aging and might contribute to aggravation of inflammation. Hence, we examined whether mitochondrial dysfunction has effects on inflammatory responses in chondrocytes. Primary cultured chondrocytes were prepared from knee articular cartilage of neonatal Wistar rats. Chondrocytes were co-treated with rotenone (Rote), an electron transport complex I inhibitor, and interleukin (IL)-1β. Co-treatment of cultured chondrocytes with Rote and IL-1β potentiated the expression of inflammatory mediators such as matrix metalloproteinase 3, IL-6, and inducible nitric oxide synthase compared to each treatment alone. The potentiation was suppressed by blockade of transforming growth factor beta-activated kinase 1 (TAK1), c-Jun N- terminal kinase (JNK), or nuclear factor-κB (NF-κB). These results suggest that mitochondrial dysfunction potentiates inflammatory responses via the TAK1/JNK/NF-κB pathway in chondrocytes. These responses might contribute to the induction of OA pathogenesis.