Osteoporosis (OP) is a common skeletal disease involving low bone mineral density (BMD) that often leads to fragility fracture, and its development is affected by multiple cellular pathologies and associated with marked epigenetic alterations of osteogenic genes. Proper physical exercise is beneficial for bone health and OP and reportedly possesses epigenetic modulating capacities; however, whether the protective effects of exercise on OP involve epigenetic mechanisms is unclear. Here, we report that epigenetic derepression of nuclear factor erythroid derived 2-related factor-2 (Nrf2), a master regulator of oxidative stress critically involved in the pathogenesis of OP, mediates the significant osteoprotective effects of running exercise (RE) in a mouse model of OP induced by ovariectomy. We showed that Nrf2 gene knockout (Nfe2l2-/-) ovariectomized mice displayed a worse BMD reduction than the controls, identifying Nrf2 as a critical antiosteoporotic factor. Further, femoral Nrf2 was markedly repressed with concomitant DNA methyltransferase (Dnmt) 1/Dnmt3a/Dnmt3b elevations and Nrf2 promoter hypermethylation in both patients with OP and ovariectomized mice. However, daily 1-h treadmill RE significantly corrected epigenetic alterations, recovered Nrf2 loss and improved the femur bone mass and trabecular microstructure. Consistently, RE also normalized the adverse expression of major osteogenic factors, including osteoblast/osteoclast markers, Nrf2 downstream antioxidant enzymes and proinflammatory cytokines. More importantly, the RE-conferred osteoprotective effects observed in the wild-type control mice were largely abolished in the Nfe2l2-/- mice. Thus, Nrf2 repression due to aberrant Dnmt elevation and subsequent Nrf2 promoter hypermethylation is likely an important epigenetic feature of the pathogenesis of OP, and Nrf2 derepression is essential for the antiosteoporotic effects of RE.