Hepatocyte nuclear factor 4α (HNF4α) is a transcription factor that plays a key role in hepatocyte differentiation and the maintenance of hepatic function, but its role in hepatocarcinogenesis has yet to be examined. Here, we report evidence of a suppressor role for HNF4α in liver cancer. HNF4α expression was progressively decreased in the diethylinitrosamine-induced rat model of liver carcinogenesis. In human liver tissues, HNF4α expression was decreased in cirrhotic tissue and further decreased in hepatocarcinoma relative to healthy tissue. Notably, an inverse correlation existed with epithelial-mesenchymal transition (EMT). Enforced expression of HNF4α attenuated hepatocyte EMT during hepatocarcinogenesis, alleviated hepatic fibrosis, and blocked hepatocellular carcinoma (HCC) occurrence. In parallel, stem cell marker gene expression was inhibited along with cancer stem/progenitor cell generation. Further, enforced expression of HNF4α inhibited activation of β-catenin, which is closely associated with EMT and hepatocarcinogenesis. Taken together, our results suggest that the inhibitory effect of HNF4α on HCC development might be attributed to suppression of hepatocyte EMT and cancer stem cell generation through an inhibition of β-catenin signaling pathways. More generally, our findings broaden knowledge on the biological significance of HNF4α in HCC development, and they imply novel strategies for HCC prevention through the manipulation of differentiation-determining transcription factors in various types of carcinomas.

Peroxisome proliferator-activated receptor ␥ (PPAR␥) is a nuclear receptor that plays a pivotal role in obesity and diabetes. PPAR␥ has two isoforms, PPAR␥1 and PPAR␥2. We investigated the functional differences between PPAR␥1 and PPAR␥2 by selectively disrupting PPAR␥2 in mice. In contrast to the embryonic lethality of PPAR␥-deficient mice, PPAR␥2 ؊/؊ mice survived. Although normal development was identified in other tissues we examined, PPAR␥2 ؊/؊ mice exhibited an overall reduction in white adipose tissue, less lipid accumulation, and decreased expression of adipogenic genes in adipose tissue. In addition, insulin sensitivity was impaired in male PPAR␥2 ؊/؊ mice, with dramatically decreased expression of insulin receptor substrate 1 and glucose transporter 4 in the skeletal muscle, but thiazolidinediones were able to normalize this insulin resistance. Consistent with in vivo data, PPAR␥2 ؊/؊ mouse embryonic fibroblasts showed a dramatically reduced capacity for adipogenesis in vitro compared with wildtype mouse embryonic fibroblasts. Taken together, our data demonstrate that PPAR␥2 deficiency impairs the development of adipose tissue and insulin sensitivity. PPAR␥2 ؊/؊ mice may provide a tool to study the role of PPAR␥2 in obesity and diabetes.adipogenesis ͉ obesity ͉ diabetes

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Disruption of the intestinal barrier is a causal factor in the development of alcoholic endotoxemia and hepatitis. This study was undertaken to determine whether zinc deficiency is related to the deleterious effects of alcohol on the intestinal barrier. Mice were pair fed an alcohol or isocaloric liquid diet for 4 wk, and hepatitis was detected in association with elevated blood endotoxin level. Alcohol exposure significantly increased the permeability of the ileum but did not affect the barrier function of the duodenum or jejunum. Reduction of tight-junction proteins at the ileal epithelium was detected in alcohol-fed mice although alcohol exposure did not cause apparent histopathological changes. Alcohol exposure significantly reduced the ileal zinc concentration in association with accumulation of reactive oxygen species. Caco-2 cell culture demonstrated that alcohol exposure increases the intracellular free zinc because of oxidative stress. Zinc deprivation caused epithelial barrier disruption in association with disassembling of tight junction proteins in the Caco-2 monolayer cells. Furthermore, minor zinc deprivation exaggerated the deleterious effect of alcohol on the epithelial barrier. In conclusion, epithelial barrier dysfunction in the distal small intestine plays an important role in alcohol-induced gut leakiness, and zinc deficiency attributable to oxidative stress may interfere with the intestinal barrier function by a direct action on tight junction proteins or by sensitizing to the effects of alcohol.

Alcoholic steatosis is a fundamental metabolic disorder in the progression of alcoholic liver disease. Zinc deficiency is one of the most consistently observed biochemical/nutritional manifestations of alcoholic liver disease. The purpose of this study is to determine whether dietary zinc supplementation to mice previously exposed to alcohol could reverse alcoholic steatosis. Male 129S mice were pair-fed an alcohol or isocaloric maltose dextrin liquid diet for 16 weeks with or without dietary zinc supplementation for the last 4 weeks. Zinc supplementation significantly attenuated alcohol-mediated increases in hepatic triglyceride, cholesterol, and free fatty acids in association with accelerated hepatic fatty acid oxidation and very low density lipoproteins (VLDL) secretion. Hepatic genes related to fatty acid oxidation and VLDL secretion were up-regulated by zinc supplementation, which was accompanied by restoring activity of hepatocyte nuclear factor-4␣ (HNF-4␣) and peroxisome proliferators

Nevertheless, a study by Yu et al. has revealed that another combination of four factors (Oct3/4, Sox2, NANOG [Nanog ho-meobox], and LIN28 [lin-28 homolog]) is also sufficient to induce pluripotent stem cells from human somatic cells, 4 indicating that more genes are involved in the establishment or maintenance of pluripotency. In this case, the therapy of HCC directing to one or several of these genes may be insufficient to achieve satisfactory efficacy. On the other hand, a recent report by Gupta et al. showed that salinomycin might selectively eliminate breast CSCs and inhibit metastasis by inducing the differentiation of CSCs. They also suggest that it is preferable to treat cancer using agents that target both the CSCs and non-CSCs, because non-CSCs might transform into CSCs and thus eradication of CSCs alone may not obtain complete regression of an established tumor. 5 Interestingly, our study indicated that HNF4 could induce the differentiation of both hepatoma cells and its CSCs. The suppression of CSCs was accompanied by the inhibition of a cluster of genes which contribute to the pluripotency of human stem cells, including-catenin, Oct3/4, SMO(smoothened homolog), Bmi, Sox2, NANOG, c-Myc, Klf4, LIN28, and ESG1 (enhancer of split groucho 1). 3 More recently, we also demonstrated that up-regulation of HNF4 remarkably ameliorated hepatic fibrosis 6 and prevented the development of HCC in rats accompanied by the revision of epithelial-mesenchymal transition (unpublished data). Thus, we believe that differentiation therapy with HNF4, a central regulator for hepatocyte differentiation, might be an ideal strategy for the treatment of human HCC. Moreover, this strategy may be extended to other cancer types through the induction of differentiation using their corresponding key transcription factors.