Background: The mechanism by which niacin increases plasma levels of high-density lipoprotein cholesterol (HDL-C) is not clearly understood yet. Adipocytes contain the largest pool of free cholesterol in the body and might play a significant role in cholesterol metabolism. Despite preferential accumulation in adipose tissue, it is not clear whether the actions of niacin on cholesterol efflux from adipocytes contribute to its HDL-raising effect. Methods: Fully differentiated 3T3-L1 adipocytes were incubated in the medium containing various concentrations of niacin (0–1.0 mmol/l) for 24 h. Reverse transcription polymerase chain reaction was used to evaluate peroxisome proliferator-activated receptor-γ (PPARγ), LXRα and ABCA1 mRNA expression in adipocytes. Cholesterol efflux rate was determined by measuring the release of radioactivity from 3H-cholesterolprelabeled cells into medium containing apolipoprotein A-I (ApoA-I). Results: Niacin dose-dependently stimulated PPARγ, LXRα and ABCA1 mRNA expression and promoted ApoA-I-induced cholesterol efflux in adipocytes. Treatment of PPARγ-selective antagonist GW9662 significantly abolished the niacin-induced increase in LXRα and ABCA1 mRNA expression and cholesterol efflux to ApoA-I. Conclusions: Niacin may promote cholesterol efflux from adipocytes to ApoA-I via activation of the PPARγ-LXRα-ABCA1 pathway. To some extent, this effect might help to explain the possible mechanism by which niacin increases plasma HDL-C levels.