Objective To establish a mouse model of diethylnitrosamine(DEN)-induced hepatocellular carcinoma (HCC), and to explore the effects of two different diet formulas on the establishment of DEN-induced HCC model. Methods SPF C57BL/6 mice (8 males and 8 females) were injected intraperitoneally with 25 mg/kg DEN at day 14 to establish a HCC model. The mice were divided into two groups after weaning. One group was fed with the SPF class rodents cereal-based diet, another group was fed with AIN-93G formula diet. The mice were sacrificed at the age of 9 months. The livers were weighed and the growth of liver cancer was observed and recorded. Results All the mice in the cereal-based diet group developed HCC as expected. The body weight and liver mass of the mice in the AIN-93G diet group were significantly lower than that of the cereal-based diet group. The incidence of HCC, and the number and size of tumor nodules were also significantly lower in the AIN-93G diet group than that in the cereal-based diet group. Conclusions DEN-induced HCC model has been successfully established in mice fed with cereal-based diet, while mice fed with AIN93-G diet prevented the development of DEN-induced HCC, and their body weight was decreased significantly, suggesting that dietary factors play a key role in establishment of animal disease models.