The ex vivo effects of passive mechanical stretch on the activation of nuclear factor‐kappaB (NF‐κB) pathways in skeletal muscles from normal and mdx mouse, a model of Duchenne muscular dystrophy (DMD), were investigated. The NF‐κB/DNA binding activity of the diaphragm muscle was increased by the application of axial mechanical stretch in a time‐dependent manner. The increased activation of NF‐κB was associated with a concomitant increase in I‐kappaB (IκB) kinase activity and the degradation of IκBμ protein. Pretreatment of the muscles with nifedipine (a Ca²⁺ channel blocker) and gadolinium(III) chloride (a stretch‐activated channel blocker) did not alter the level of activation of NF‐κB, ruling out involvement of Ca²⁺ influx through these channels. Furthermore, N‐acetyl cysteine, a free radical inhibitor, blocked the mechanical stretch‐induced NF‐κB activation, suggesting the involvement of free radicals. Compared with normal diaphragm, the basal level of NF‐κB activity was higher in muscles from mdx mice, and it was further enhanced in mechanically stretched muscles. Furthermore, activation of NF‐κB and increased expression of inflammatory cytokines IL‐1 β and tumor necrosis factor β in the mdx mouse precede the onset of muscular dystrophy. Our results show that mechanical stretch activates the classical NF‐κB pathway and this pathway could be predominately active in DMD. FASEB J. 17, 386–396 (2003)