Hepatic steatosis is present in insulin-resistant obese rodents and is concomitant with active lipogenesis. Hepatic lipogenesis depends on the insulin-induced activation of the transcription factor SREBP-1c. Despite prevailing insulin resistance, SREBP-1c is activated in the livers of genetically and diet-induced obese rodents. Recent studies have reported the presence of an ER stress response in the livers of obese ob/ob mice. To assess whether ER stress promotes SREBP-1c activation and thus contributes to lipogenesis, we overexpressed the chaperone glucose-regulated protein 78 (GRP78) in the livers of ob/ob mice using an adenoviral vector. GRP78 overexpression reduced ER stress markers and inhibited SREBP-1c cleavage and the expression of SREBP-1c and SREBP-2 target genes. Furthermore, hepatic triglyceride and cholesterol contents were reduced, and insulin sensitivity improved, in GRP78-injected mice. These metabolic improvements were likely mediated by restoration of IRS-2 expression and tyrosine phosphorylation. Interestingly, GRP78 overexpression also inhibited insulin-induced SREBP-1c cleavage in cultured primary hepatocytes. These findings demonstrate that GRP78 inhibits both insulin-dependent and ER stress-dependent SREBP-1c proteolytic cleavage and explain the role of ER stress in hepatic steatosis in obese rodents.

Obesity is a metabolic disorder often associated with type 2 diabetes, insulin resistance, and hepatic steatosis. Leptin-deficient (ob/ob) mice are a well-characterized mouse model of obesity in which increased hepatic lipogenesis is thought to be responsible for the phenotype of insulin resistance. We have recently demonstrated that carbohydrate responsive element-binding protein (ChREBP) plays a key role in the control of lipogenesis through the transcriptional regulation of lipogenic genes, including acetylCoA carboxylase and fatty acid synthase. The present study reveals that ChREBP gene expression and ChREBP nuclear protein content are significantly increased in liver of ob/ob mice. To explore the involvement of ChREBP in the physiopathology of hepatic steatosis and insulin resistance, we have developed an adenovirus-mediated RNA interference technique in which short hairpin RNAs (shRNAs) were used to inhibit ChREBP expression in vivo. Liver-specific inhibition of ChREBP in ob/ob mice markedly improved hepatic steatosis by specifically decreasing lipogenic rates. Correction of hepatic steatosis also led to decreased levels of plasma triglycerides and nonesterified fatty acids. As a consequence, insulin signaling was improved in liver, skeletal muscles, and white adipose tissue, and overall glucose tolerance and insulin sensitivity were restored in ob/ob mice after a 7-day treatment with the recombinant adenovirus expressing shRNA against ChREBP. Taken together, our results demonstrate that ChREBP is central for the regulation of lipogenesis in vivo and plays a determinant role in the development of the hepatic steatosis and of insulin resistance in ob/ob mice.

Despite its importance in terms of energy homeostasis, the role of AMP-activated protein kinase in adipose tissue remains controversial. Initial studies have described an anti-lipolytic role for AMP-activated protein kinase, whereas more recent studies have suggested the converse. Thus we have addressed the role of AMP-activated protein kinase in adipose tissue by modulating AMP-activated protein kinase activity in primary rodent adipocytes using pharmacological activators or by adenoviral expression of dominant negative or constitutively active forms of the kinase. We then studied the effects of AMPactivated protein kinase activity modulation on lipolytic mechanisms. Finally, we analyzed the consequences of a genetic deletion of AMP-activated protein kinase in mouse adipocytes. AMP-activated protein kinase activity in adipocytes is represented mainly by the ␣ 1 isoform and is induced by all of the stimuli that increase cAMP in adipocytes, including fasting. When AMP-activated protein kinase activity is increased by 5-aminoimidazole-4-carboxamide-riboside, phenformin, or by the expression of a constitutively active form, isoproterenol-induced lipolysis is strongly reduced. Conversely, when AMP-activated protein kinase activity is decreased either by a dominant negative form or in AMP-activated protein kinase ␣ 1 knock-out mice, lipolysis is increased. We present data suggesting that AMP-activated protein kinase acts on hormone-sensitive lipase by blocking its translocation to the lipid droplet. We conclude that, in mature adipocytes, AMP-activated protein kinase activation has a clear anti-lipolytic effect.