Obesity is a condition of abnormal adipose tissue storage and recently it has been recognized as a major factor in metabolic syndrome. High-fat diet-induced obesity in the C57BL/6 mouse is an important animal model because of its similarities with human obesity. The aim of the present study was to estimate obesity, liver injury and steatohepatitis, and the distribution of inducible nitric oxide synthase (iNOS) in mice with high-fat diet induced obesity. Three-week-old male C57BL/6J mice were fed either a high-fat diet (D-60: 60 kcal% fat, or D-45: 45 kcal% fat) or a normal diet (D-10: 10 kcal% fat) for 15 weeks. Oral glucose tolerance tests and intraperitoneal glucose tolerance tests showed that the D-60 mice had severely impaired glucose tolerance. In serum chemistry values and histopathological lesions, the D-60 group showed severe steatohepatitis. A distinct positive signal for iNOS was detected by immunohistochemistry in the cytoplasm of hepatocytes around the central vein in the D-45 and D-60 groups. Serum insulin levels and insulin immunohistochemistry in the pancreas showed pancreatic injury and insulin resistance in the D-60 group. We observed the presence of more iNOS in the high-fat diet-induced obese mouse, which has characteristics of non-alcoholic steatohepatitis (NASH) and diabetes, and expect that these background pathological data will be useful in research on obesity, diabetes mellitus, and non-alcoholic fatty liver disease.