Recently, nonalcoholic steatohepatitis (NASH) was found to be correlated with cardiovascular disease events independently of the metabolic syndrome. The aim of this study was to investigate whether an atherogenic (Ath) diet induces the pathology of steatohepatitis necessary for the diagnosis of human NASH and how cholesterol and triglyceride alter the hepatic gene expression profiles responsible for oxidative stress. We investigated the liver pathology and plasma and hepatic lipids of mice fed the Ath diet. The hepatic gene expression profile was examined with microarrays and real-time polymerase chain reactions. The Ath diet induced dyslipidemia, lipid peroxidation, and stellate cell activation in the liver and finally caused precirrhotic steatohepatitis after 24 weeks. Cellular ballooning, a necessary histological feature defining human NASH, was observed in contrast to existing animal models. The addition of a high-fat component to the Ath diet caused hepatic insulin resistance and further accelerated the pathology of steatohepatitis. A global gene expression analysis revealed that the Ath diet up-regulated the hepatic expression levels of genes for fatty acid synthesis, oxidative stress, inflammation, and fibrogenesis, which were further accelerated by the addition of a high-fat component. Conversely, the high-fat component down-regulated the hepatic gene expression of antioxidant enzymes and might have increased oxidative stress. Conclusion: The Ath diet induces oxidative stress and steatohepatitis with cellular ballooning. The high-fat component induces insulin resistance, down-regulates genes for antioxidant enzymes, and further aggravates the steatohepatitis. This model suggests the critical role of lipids in causing oxidative stress and insulin resistance leading to steatohepatitis.

Various peripheral tissues show circadian rhythmicity, which is generated at the cellular level by their own core oscillators that are composed of transcriptional/translational feedback loops involving a set of clock genes. Although the circulating levels of some adipocytokines, i.e. bioactive substances secreted by adipocytes, are on a 24-h rhythmic cycle, it remains to be elucidated whether the clock gene system works in adipose tissue. To address this issue, we investigated the daily mRNA expression profiles of the clock genes and adipocytokines in mouse perigonadal adipose tissues. In C57BL/6J mice, all transcript levels of the clock genes (Bmal1, Per1, Per2, Cry1, Cry2, and Dbp) and adipocytokines (adiponectin, resistin, and visfatin) clearly showed 24-h rhythms. On the other hand, the rhythmic expression of these genes was mildly attenuated in obese KK mice and greatly attenuated in more obese, diabetic KK-A^y mice. Obese diabetes also diminished the rhythmic expression of the clock genes in the liver. Interestingly, a 2-wk treatment of KK and KK-A^y mice with pioglitazone impaired the 24-h rhythmicity of the mRNA expression of the clock genes and adipocytokines despite the antidiabetic effect of the drug. In contrast, pioglitazone improved the attenuated rhythmicity in the liver. These findings suggest that the intracellular clock gene system acts in visceral adipose tissues as well as liver and is influenced by the conditions of obesity/type 2 diabetes and pioglitazone treatment.