Abstract 281: Resveratrol Enhances Cholesterol Efflux Through PPAR-γ--Dependent Signaling Pathways in Human Macrophages

Abstract

Introduction: We previously reported that resveratrol, a plant derived polyphenol with numerous cardioprotective properties, improves cholesterol homeostasis by upregulating the expression of reverse cholesterol transport (RCT) proteins critical for preventing lipid overload and macrophage foam cell formation. However, the mechanism(s) by which resveratrol exerts this effect are unclear. The present study explores a possible mechanism for the atheroprotective actions of resveratrol on cholesterol metabolism. Hypothesis: We hypothesize that the anti-atherogenic effects of resveratrol are mediated through PPAR-γ dependent signaling pathways leading to upregulation of RCT proteins and enhanced cholesterol efflux. Methods: THP-1 macrophages, a pertinent model for atherosclerosis, were incubated for 20h with resveratrol (10μM) with/without the specific PPAR-γ antagonist GW6992 (1μM). Alternatively, cells were transfected with 100 nM of human PPAR-γ small interfering RNA (siRNA) for gene silencing prior to resveratrol treatment. Total RNA was isolated, converted to cDNA, and evaluated by QRT-PCR. Each reaction was done in triplicate. Western blot analysis was performed to confirm results of gene expression. Results: Resveratrol significantly increased expression of PPAR-γ, ABCA1, and 27-hydroxylase mRNA (mean±SEM, 136.2±8.5%, 168.3±9.2%, and 171.7±11.0% of control respectively, P<0.001). PPAR-γ gene silencing and PPAR-γ antagonist GW6992 effectively reduced PPAR-γ message by 90.4% and 54.7%, respectively (P<0.001). Both pharmacological blockade and gene knockdown of PPAR-γ nullified resveratrol effects on cholesterol efflux proteins. In cells treated with GW6992, mRNA levels of ABCA1 and 27-hydroxylase were decreased to 66.1±3.3% and 55.0±2.9% of control, respectively (P<0.001). Similarly, PPAR-γ silencing resulted in downregulation of ABCA1 and 27-hydroxylase expression to the level of control, which is significantly lower than in resveratrol treated cells (P<0.001). Data from gene expression studies were subsequently confirmed by Western blot analysis. Conclusions: We propose here a mechanistic model for the atheroprotective effects of resveratrol. Our data strongly suggests that resveratrol regulates cholesterol efflux and intracellular cholesterol processing via the PPAR-γ/LXR-α pathway.