Objective To investigate whether exogenous osteocalcin intervention prevents high fat diet-induced nonalcoholic fatty liver disease (NAFLD) in wild type mice. Methods Five-week-old C57Bl/6J male mice were randomly divided into 4 groups, that is, normal diet, high fat diet, and high fat diet＋osteocalcin 3 or 10ng/(g·d) groups. The body mass, blood glucose, and food intake were monitored during the 24 weeks’ experiment. Serum levels of total cholesterol (TC), triglycerides (TG), free fatty acid (FFA), and tumor necrosis factor α (TNF-α) and liver contents of TC and TG were measured at the end of the study. Oil Red O and hematoxylin-eosin staining were performed to observe the morphology of the livers. Results Compared with high fat diet group, the groups treated by different doses of osteocalcin had significantly decreased total fat mass (including the fat around epididymides and kidneys as well as subcutaneous fat) and total fat mass/body mass ratio (P＜0.001), better fasting blood glucose and glucose tolerance (P＜0.05?0.001), and obviously reduced serum levels of TC, TG, FFA and TNF-α (P＜0.05). Osteocalcin treatment prevented high fat diet-induced liver steatosis and resulted in remarkably decreased liver mass, ratio of liver mass to body mass as well as liver TC and TG levels (P＜0.05). Conclusion Intermittent injection of osteocalcin significantly prevents high fat diet-induced NAFLD, improves glucose-lipids metabolism and reduces serum TNF-α level.