6A). The recovery of expression of RORα in the liver by tail-vein injection of Ad-RORα

led to restoration of pAMPK levels (Fig. 6B). In addition, mRNA expression of SREBP-1c, FAS, ACC, and SCD1, was significantly lower in the Ad-RORα–infected liver (Fig. 6C). In agreement with these molecular findings, hepatic triglyceride levels were reduced in the Ad-RORα–injected mice (Fig. 6D). Histological examination showed clearly that the hepatocytes of HFD-fed mice were distended by accumulation of lipid droplets. This change was attenuated in the Ad-RORα–injected mice (Fig. 6E). Together, these results indicate that RORα reduces lipid accumulation in vivo by regulating genes that are important in hepatic lipogenesis (Fig. 6F). We synthesized 50 thiourea derivatives PKC412 in vitro based on the structure of CGP52608, an RORα-activating

compound (Fig. 7A). 18 The most active, JC1-38, JC1-40, and JC1-42, induced expression of known endogenous RORα target genes such as SPARC and AGRP (Fig. 7B and Supporting Fig. 5). 23 A docking study using the X-ray crystallographic Lapatinib structure of the RORα complex with CS showed that JC1-42 fitted well into the binding pocket (Fig. 7C). These compounds induced phosphorylation of AMPK/ACC, decreased the expression as well as transcriptional activity of LXRα, and attenuated the FFA mixture–induced lipid accumulation in HepG2 cells (Fig. 7D–F). Finally, we performed in vivo experiments to evaluate the inhibitory effects of JC1-40 and JC1-42 on hepatic lipid accumulation using a safflower oil–enriched HFD model. 27 As expected, oral administration of JCI-40 or JCI-42 led to strong activation of AMPK and repression of LXRα expression in the liver of HFD-fed mice (Fig. 8A). Consistently, hepatic triglyceride levels were significantly lowered in the mice given compound (30

mg/kg) mice (Fig. 8B). Furthermore, body weight was significantly reduced by the compounds, although food intake was not much different among experimental buy Docetaxel groups (Fig. 8C). Oil-red O staining of liver tissues showed clearly that hepatic steatosis in the HFD-fed mice were attenuated by administration of JC1 compounds (Fig. 8D). In the present study, we demonstrated that RORα induced activation of AMPK and inhibition of the lipogenic function of LXRα, which conferred beneficial effects against hepatic steatosis. AMPK is activated by a variety of physiological and pathological stresses that increase the intracellular AMP/ATP ratio, either by increasing ATP consumption or by decreasing ATP production. 6 We showed clearly that RORα expression was associated with a reduction in cellular ATP levels and activation of LKB1 (Fig. 1). Interestingly, we observed that overexpression of constitutively active (CA)-AMPK or treatment with aminoimidazole carboxamide ribonucleotide (AICAR), an activator of AMPK, induced phosphorylation of RORα, which was abolished in the presence of compound C, an inhibitor of AMPK (Supporting Fig. 6A,B).