Transforming growth factor beta 1 (TGFβ1) is a key player in skeletal muscle degenerative and regenerative processes. We previously showed that conditionally overexpressing TGFβ1 in skeletal muscles caused myofiber atrophy and endomysial fibrosis in mice. However, the disease severity varied significantly among individual mice. While 40% of mice developed severe muscle pathology and lost body weight within 2 weeks of TGFβ1 transgene induction in muscles, the rest showed milder or no phenotype. This study aims at determining whether signal transducer and activator of transcription 3 (STAT3) plays a role in the phenotypic difference and whether it can be activated by TGFβ1 directly in muscle cells. Our results show that while total STAT3 was not differentially expressed between the two groups of mice, there was significantly higher pSTAT3 (Tyr705) in the muscles of the mice with severe phenotype. Immunohistochemistry showed that pSTAT3 (Tyr705) was localized in approximately 50% of the nuclei of the muscles. We further showed that TGFβ1 induced Tyr705 phosphorylation of STAT3 in C2C12 cells within 30 minutes of treatment while total STAT3 was not affected. Our findings suggest that TGFβ1 alone can induce Tyr705 phosphorylation of STAT3 in skeletal muscle cells and contribute to disease severity in transgenic TGFβ1 mice.