Objective To investigate the role of cluster of differentiation (CD) 36 in the formation of non-alcoholic fatty liver disease (NAFLD) in mice. Methods Twenty speciifc pathogen free healthy male C57BL/6J mice [8 weeks old, average weight:(18.8±2.3)g] were randomly divided into NAFLD group and control group according to the random number table method with 10 mice in each group. Mice in NAFLD group was fed with high-fat diet for 10 weeks, while mice in control group was fed with normal diet for 10 weeks. The mice were sacriifced after the processing, and samples of cardiac blood and liver tissue were collected. The levels of serum alanine transaminase (ALT), total cholesterol (TC) and triglyceride (TG), levels of TC and TG in liver tissue, pathological changes of liver tissue, expression of CD36 protein and content of its messenger ribonucleic acid (mRNA) in liver tissue of 2 groups were observed. Experimental data of 2 groups were compared using t test. Results The level of serum ALT was (49±6)U/L in NAFLD group and (45±7)U/L in control group, where no signiifcant difference was obsersed (t=1.70, P>0.05). The levels of serum TC and TG were (4.42±0.09), (0.45±0.04) mmol/L in NAFLD group and (2.42±0.05), (0.32±0.03)mmol/L in control group respectively. The levels of serum TC and TG in NAFLD group were signiifcantly higher than those in control group (t=21.90, 8.22;P<0.05). The levels of TC and TG in liver tissue were (1.18±0.09), (1.75±0.08)mmol/L in NAFLD group and (0.55±0.06), (1.28±0.06)mmol/L in control group respectively. The levels of TC and TG in liver tissue in NAFLD group were signiifcantly higher than those in control group (t=18.42, 14.86; P<0.05). Obviously fatty degeneration and ballooning degeneration were observed in hepatocytes of NAFLD group, while hepatocytes were normal in morphology and size in control group. The expression of CD36 protein in liver tissue in NAFLD group was signiifcantly stronger than that in control group. The content of CD36 mRNA in liver tissue was 2.75±0.26 in NAFLD group and 1.00±0.08 in control group. The content of CD36 mRNA in liver tissue in NAFLD group was significantly higher than that in control group (t=21.16, P<0.05). Conclusions CD36 may play a role in the formation of NAFLD in mice. To decrease its expression can be a new target for the prophylaxis and treatment of NAFLD.