Mesterolone is a synthetic oral anabolic androgenic steroid used to treat hypogonadism. There are frequent reports of mesterolone abuse in human and equine sports to increase muscle mass and strength. However, limited information is available about how this drug exerts its effects on skeletal muscle. Satellite cells (SCs) are mononuclear myogenic stem cells that contribute to postnatal muscle growth and repair. As SC activation and subsequent differentiation to new myonuclei is a major event during muscle hypertrophy, this study investigated the influence of mesterolone on SC distribution within the pectoralis muscle of chickens. Specifically, this study tested the hypotheses that mesterolone induces avian skeletal muscle hypertrophy, and that mesterolone increases the number of SCs in avian skeletal muscle. Robust immunocytochemical techniques and morphometric analyses were used to calculate the numbers of SCs and myonuclei. Also, DNA concentration and Pax7 protein levels were measured to confirm immunocytochemical findings. Mesterolone significantly increased pectoralis mass and fiber size. All SC indices and number of myonuclei increased significantly by mesterolone administration. In addition, greater DNA concentration and Pax7 protein expression were found in mesterolone-treated birds. This study indicates that mesterolone can induce avian skeletal muscle hypertrophy and that this is correlated with increased number of SCs. We suggest that SCs are key cellular intermediaries for mesterolone-induced muscle hypertrophy. Anat Rec,