Insulin resistance and type 2 diabetes are pathogenetically linked to increased risk of cardiovascular disease. While insulin resistance is defined by a dysregulation in hepatic insulin signaling, it is unclear how this impairment relates to the development of cardiovascular disease. Recently, there has been evidence showing that in insulin resistant individuals, cardiovascular disease is associated with a defect in reverse cholesterol transport - the cardioprotective process by which excess cholesterol is removed from the periphery by high- density lipoproteins (HDL), and returned to the liver for biliary excretion. The proposed project seeks to establish a better understanding of HDL metabolism and reverse cholesterol transport, as they relate to key transcription factors that mediate hepatic insulin signaling, namely the insulin-repressible forkhead transcription factors, FoxO1, FoxO3, and FoxO4 (FoxOs). Preliminary data have shown that hepatic FoxO ablation in a liver-specific triple FoxO knock-out mouse (L-FoxO1,3,4) causes elevated circulatory levels of HDL-cholesterol, as well as dysregulation in multiple genes required for HDL metabolism and function. Thus, in Aim 1, hepatic FoxO expression will be restored in L-FoxO1,3,4 mice in order to verify that the previously identified HDL metabolism genes are in fact targets of hepatic FoxOs. In parallel, these dysregulated HDL genes will also be restored in order to establish a causal link between hepatic FoxO deficiency and impaired reverse cholesterol transport. In Aim 2, L-FoxO1,3,4 mice will be crossed to mice expressing a human transgene of cholesteryl ester transfer protein (CETP), an enzyme that mediates the transfer of cholesterol from HDL particles to low- density lipoprotein (LDL) particles in humans, thereby allowing HDL-cholesterol to be cleared through the LDL receptor pathway. As a result, this aim will determine the contribution of hepatic FoxOs to reverse cholesterol transport in a setting more relevant to human physiology. Taken together, the proposed research seeks to uncover a novel mechanism by which HDL metabolism is regulated by an insulin signaling pathway. Moreover, the findings of this work can be extrapolated to provide insight into the pathophysiology of cardiovascular disease in the setting of insulin resistance, which may lead to the discovery of new therapeutic targets for the prevention of cardiovascular disease in insulin resistant and type 2 diabetic patients.