Therefore, EGFR inhibition we fed the mice HFHFr or ATRA-supplemented HFHFr (ATRA

+ HFHFr) diets for another 4 weeks to investigate the effects of ATRA on NAFLD-associated insulin resistance (Fig. 2A). Although daily consumption did not differ between mice fed the HFHFr and ATRA + HFHFr diets, consumption was significantly lower in these two groups than in the control group (Fig. 2B), indicating that ATRA does not induce an anorexigenic effect and that leptin therefore likely does not affect the central nervous system in NAFLD mice. ATRA significantly decreased body weight, liver-to-body weight ratio, visceral fat tissue weight, serum alanine aminotransferase and aspartate aminotransferase levels in NAFLD mice (Fig. 2C-E, Supporting Fig. 3A,B). In agreement with a significant decrease in hepatic lipid levels

in the ATRA + HFHFr group as shown by biochemical assays, quantitative real-time polymerase chain reaction (qPCR) demonstrated that ATRA significantly up-regulated the lipolytic transcription factors PPARα and PPARβ, with concomitant down-regulation of lipogenic transcription factors, PPARγ and sterol regulatory element-binding protein 1 (SREBP1), and fatty acid synthase, as described24 (Fig. 3A, Supporting Fig. 3C-E). Note that the primers for detecting SREBP1 were designed to detect splicing variants SREBP1a and SREBP1c, both of which play central roles in regulating lipid synthesis, although each variant differs MCE in some aspects.28 Moreover, ATRA significantly improved hepatic

histology as indicated MK-2206 cost by decreased numbers and area of lipid droplets, and ballooned hepatocytes (Supporting Fig. 3F, Supporting Table 1). The histological study also revealed remarkably diminished periportal macrovesicular steatosis, suggesting enhanced lipid metabolism in the livers of the ATRA + HFHFr group. This finding was consistent with the expression of lipid metabolism-related genes (Fig 3A-E). However, the efficacy of ATRA for treating patients with nonalcoholic steatohepatitis remains to be determined, as the mice in our present study did not develop severe inflammation and fibrosis (Supporting Table 1). We then explored the effect of ATRA on insulin resistance in a mouse model of HFHFr diet-induced NAFLD. As observed in KK-Ay mice, ATRA significantly normalized hyperglycemia, hyperinsulinemia, glucose intolerance, insulin resistance (homeostatic model assessment of insulin resistance and intraperitoneal insulin tolerance test), as well as hyperleptinemia (Fig. 4A-D, Supporting Fig. 4). In contrast to leptin, the HFHFr diet did not affect circulating levels of adiponectin, another antidiabetic adipokine, or tumor necrosis factor-α, which is known to impair insulin sensitivity (data not shown). IRS1 expression was significantly increased in association with increased tyrosine phosphorylation in ATRA + HFHFr group, compared with those in the HFHFr group (Fig. 4E, Supporting Fig. 5A,B).