Plasma levels of high density lipoprotein (HDL) cholesterol and its major apolipoprotein (apo), apo A-I, are inversely correlated with the incidence of ischemic cardiovascular diseases. Reverse cholesterol transport is likely the main mechanism underlying the atheroprotective effects of HDL. Here, we investigated whether increased HDL cholesterol following hepatocyte-directed adenoviral rabbit apo A-I (AdrA-I) or rabbit lecithin:cholesterol acyltransferase (LCAT) (AdrLCAT) transfer may induce cholesterol unloading in complex atherosclerotic lesions in heterozygous low density lipoprotein deficient rabbits fed a 0.15% cholesterol diet for 420 days before and for 120 days after transfer. HDL cholesterol levels increased 2.0-fold (p<0.001) and 1.9-fold (p<0.001) in the 120 days after transfer with AdrA-I and AdrLCAT, respectively, compared to levels just before transfer whereas non-HDL cholesterol remained unchanged. Increased HDL cholesterol following AdrA-I and AdrLCAT transfer resulted in a 31% (p<0.05) reduction of the intima/media ratio in comparison with the control progression group. Compared to the baseline group sacrificed after 420 days of cholesterol diet, AdrA-I and AdrLCAT transfer reduced the percentage of Oil Red O area 1.6-fold (p<0.001) and 1.4-fold (p<0.001), respectively. In conclusion, increased HDL cholesterol after AdrA-I and AdrLCAT transfer inhibits progression of atherosclerosis and induces cholesterol unloading in complex lesions in rabbits.