Hepatic nuclear receptors play an important role in the regulation of lipid metabolism, storage, transport and elimination in response to nutrient and hormonal cues. Dysfunction of these receptors is linked to a number of age-associated metabolic diseases, including diabetes, obesity and atherosclerosis (Chawla et al., 2001
). In the present study, we identify PPARα signaling as a central pathway affected by hepatic deletion of SIRT1. We observed that blunted PPARα signaling in SIRT1 LKO mice resulted in decreased fatty acid oxidation and ketogenesis (), contributing to the development of hepatic steatosis, inflammation, and ER stress on a high-fat diet ( and ).
Several lines of evidence suggest a link between SIRT1 and PPARα. For instance, both SIRT1 and PPARα are activated by fasting and food restriction (Cohen et al., 2004
; Hashimoto et al., 2000
; Kersten et al., 1999
; Rodgers et al., 2005
). PGC-1α, a key coactivator for PPARα signaling (Li et al., 2008
; Vega et al., 2000
), is a direct substrate of SIRT1 (Rodgers et al., 2005
). Here we show that PPARα signaling is significantly impaired in SIRT1 LKO mice, while increased SIRT1 levels stimulate PPARα activity (), providing a direct link between SIRT1 and PPARα. Furthermore, our data suggest that SIRT1 regulates PPARα signaling primarily through the activation of PGC-1α (). In SIRT1 deficient hepatocytes, PGC-1α is still recruited to the PPREs on the promoter of fatty acid oxidation genes (). However, it remains in a hyper-acetylated state () that inhibits its ability to promote transcription (). How acetylation affects the activity of PGC-1α is still not well understood. Since PGC-1α is highly accumulated on the promoter regions of PPARα target genes in SIRT1 deficient cells (), it is possible that SIRT1 is required to efficiently remove less-activated PGC-1α from the promoters to allow recruitment of active PGC-1α for additional rounds of transcription. This mechanism has been well described for other SIRT1-mediated transcriptional activations (Kitamura et al., 2005
; Li et al., 2007
; Pagans et al., 2005
In addition to fatty acid oxidation, PGC-1α also stimulates hepatic gluconeogenesis in response to fasting (Herzig et al., 2001
; Yoon et al., 2001
). The decreased coactivation activity of PGC-1α in SIRT1 LKO mice suggests that gluconeogenesis may be impaired. However, we failed to observe notable defects in the hepatic gluconeogenesis in the SIRT1 LKO mice (Figure S5). Fasting glucose levels in SIRT1 LKO mice were normal under a chow diet, and were slightly increased under the western diet or a high-fat diet (Figure S5A). mRNA levels of two rate-limiting enzymes in the hepatic gluconeogenesis pathway, phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase), were also normal (Figure S5B). These observations are not surprising, given the fact that SIRT1 also deacetylates and represses TORC2 (Liu et al., 2008
) and FOXO1 (Motta et al., 2004
), two additional key factors involved in promoting gluconeogenesis in the early and late fasting phases, respectively. Therefore, the net effect of loss of SIRT1 on gluconeogenesis is determined by the complex compensatory patterns of multiple factors.
An intriguing observation in the present study is that the effect of SIRT1 on fatty acid metabolism is coupled with altered cholesterol metabolism. Not only do SIRT1 LKO mice accumulate massive amounts of cholesterol when fed a western diet (), they also accumulate significantly higher hepatic cholesterol when fed a high-fat diet without cholesterol (Figure S4). These observations suggest increased de novo synthesis is the major source of hepatic cholesterol. However, the expression levels of two key mediators of cholesterol synthesis, SREBP2 and HMGCoA reductase, were normal in SIRT1 LKO mice on the western diet. Hence, the increased synthesis may be due to an increase in the availability of substrates through accumulated hepatic free fatty acids ().
The metabolic phenotypes observed in SIRT1 LKO mice are in line with several previous reports. For example, activation of SIRT1 by the polyphenol resveratrol and several synthetic pharmacologic activators has been shown to protect against high-fat induced obesity and metabolic derangements (Baur et al., 2006
; Lagouge et al., 2006
; Milne et al., 2007
). Manipulation of SIRT1 levels in the liver has been reported to affect the expression of a number of genes involved in glucose and lipid metabolism (Rodgers and Puigserver, 2007
). Additionally, recent studies demonstrated that modest overexpression of SIRT1 resulted in a protective effect against high-fat induced hepatic steatosis and glucose intolerance (Banks et al., 2008
; Pfluger et al., 2008
). Our observations from SIRT1 LKO mice suggest that hepatic SIRT1 mediates a fine balance between energy influx and energy expenditure in the liver. However, it is important to note that although insulin signaling is impaired in the livers of SIRT1 LKO mice (), they show normal insulin sensitivity and fuel metabolism in white adipose tissue and muscle, and thus do not develop systemic glucose intolerance (Figure S5C).
Several of the observed metabolic alterations in the SIRT1 LKO mice are in contrast from those observed in some SIRT1 knockout animal studies (unpublished observations; (Chen et al., 2008
)). For example, SIRT1 LKO mice gain significantly more weight than wild type mice and develop hepatic steatosis when fed with high-fat diets ( and Figure S4). However, whole body SIRT1 knockout mice are protected from high-fat diet induced obesity and fatty liver (Li and Guarente, unpublished observations). A distinguishing difference between these two knockout models is their overall growth condition. SIRT1 LKO mice have no obvious phenotypic abnormalities under normal dietary conditions, whereas whole body SIRT1 knockouts suffer severe growth retardation, which can likely be attributed to a number of developmental defects (Cheng et al., 2003
; McBurney et al., 2003
). In addition, defective SIRT1 function in other tissues is likely to systemically affect energy metabolism in the whole body SIRT1 knockout mice, making it difficult to dissect hepatic specific function of SIRT1.
In summary, we have shown that hepatic SIRT1 plays an important role in the regulation of lipid metabolism in response to nutrient and hormonal signals. While it has been reported that SIRT1 systemically regulates energy homeostasis in many metabolic tissues by modulating a variety of signaling pathways, we show that in the liver, a major target of this sirtuin is the PPARα/PGC-1α signaling pathway and fatty acid oxidation. Our findings provide a direct link between SIRT1 and hepatic fatty acid metabolism, and suggest that therapeutic strategies designed to modulate SIRT1 activity may be beneficial for the treatment of hepatic diseases as well as obesity-associated metabolic syndrome.