Inflammation of the muscle invariably leads to muscle cell damage and impaired regeneration. Biomechanical signals play a vital role in the regulation of myogenesis in healthy and inflamed muscle. We hypothesized that biomechanical signals counteract the actions of proinflammatory mediators and upregulate myogenesis in inflamed muscle cells. For this purpose, C2C12 cells plated on silastic membranes were subjected to equibiaxial cyclic tensile strain (CTS) in the presence or absence of TNF-, and the myogenic gene induction examined over a period of 72 h. Exposure of cells to CTS resulted in a significant upregulation of mRNA expressions and synthesis of myogenic regulatory factors, MYOD1, MYOG (myogenin), MEF2A, CDKN1A (p21) as well as muscle structural proteins like myosin heavy chain (MYHC) isoforms (MYH1, MYH2 and MYH4) and TPM1 (-tropomyosin), eventually leading to an increase in myotube formation. Contrarily, TNF- suppressed expression of all of the above differentiation-inducing factors in C2C12 cells. Further results revealed that simultaneous exposure of C2C12 cells to CTS and TNF- abrogated the TNF--mediated downregulation of myogenic differentiation. The mRNA expression and protein synthesis of all myogenic factors (Myod1, Myog, Mef2a, Cdkn1a, Myh1, Myh2, Myh4 and Tpm1) were increased in stretched C2C12 cells despite the sustained presence of TNF-. These results demonstrate that mechanotransduction regulates multiple signaling molecules involved in C2C12 cell differentiation. The findings suggest that these signals are potent transducers of myotube phenotype in myoblasts and counteract catabolic actions of proinflammatory cytokine like TNF- and allow expression of myogenic genes to upregulate muscle cell differentiation.