Changes in gene expression of TGF-beta family members and their receptors in response to treatment with H(2)O(2) and a calcium ionophore, A23187, were examined in C2C12 myoblasts and myotubes. The expression of Myf5, an initial regulator of myogenesis, was increased by A23187, and H(2)O(2) inhibited the up-regulation of Myf5. Treatment with H(2)O(2) decreased the expression of MHC IIb, a protein component of the myofibrils, irrespective of the presence of A23187, suggesting an inhibitory role of oxidative stress for myogenesis. Expression of ligands and receptors for the TGF-beta family was modulated in response to H(2)O(2) and A23187. Treatment with H(2)O(2) decreased expression of TGF-beta3, BMP-4, ALK4, ALK5, and ActRIIB, and increased expression of inhibin alpha and inhibin betaA in either the myoblast stage or the myotube stage, or both. A23187 potentiated down-regulation of BMP-4 and ALK4 expression, and up-regulation of TGF-beta1, TGF-beta2, inhibin alpha, inhibin betaA, ALK2, and ALK3 expression. These results indicate that oxidative stress and Ca(2+) influx affect expression of the TGF-beta family in C2C12 myoblasts and myotubes.