Nicotine is the main addictive substance in tobacco. It has been reported that nicotine can improve obesity and promote body weight loss in humans and rodents. In addition, obesity is associated with many chronic diseases. Many studies have demonstrated that the skeletal muscle regenerative capacity is impaired in obese mice. However, the effect of nicotine on skeletal muscle regeneration under obese conditions remains unclear. Thus, in the present study, we examined the effects of nicotine on the differentiation of C2C12 myoblasts in vitro and on skeletal muscle regeneration in obese mice in vivo. The results showed that nicotine promoted C2C12 myoblast differentiation by upregulating myogenic regulatory factors, including MyoD and Myogenin. Nicotine also activated the PI3K/Akt signaling pathway, while blocking MK with the inhibitor LY294002 abrogated the effects of nicotine on the differentiation of C2C12 cells. Furthermore, nicotine was injected into the cardiotoxin (CTX)-injured skeletal muscles of obese mice. The results showed that the skeletal muscles injected with nicotine regenerated more quickly than the skeletal muscles injected with saline. Taken together, our data suggested that nicotine promoted the differentiation of C2C12 cells through activation of the PI3K/Akt pathway and rescued the impaired skeletal muscle regeneration in obese mice. (C) 2019 Elsevier Inc. All rights reserved.