Apolipoprotein A-IMilano (apoA-IM), a natural variant of apolipoprotein A-I (apoA-I), confers to the carriers a significant protection against vascular disease. The antiatherogenic activity of a recombinant disulfide-linked apoA-IM dimer (rA-IM/A-IM) was analyzed in vivo by evaluating its effect on neointimal formation induced by periarterial manipulation in 1% cholesterol-fed rabbits. A flexible collar was applied around the carotid artery 21 days after the beginning of the dietary regimen, and animals were killed 10 days later. Rabbits were injected five times with reconstituted high-density lipoprotein containing egg phosphatidylcholine (EPC) and rA-IM/A-IM (119 mg EPC + 40 mg protein per dose) or with EPC liposomes (119 mg EPC per dose) beginning either 5 days before or at the day of collar positioning. Neither treatment affected plasma cholesterol levels. A significant intimal thickening was observed in control animals; the intima-to-media (I/M) ratio was 0.63 +/- 0.11 versus 0.03 +/- 0.05 for the sham-operated contralateral arteries. Neointimal formation was markedly inhibited in animals pretreated with rA-IM/A-IM before lesion induction (I/M, 0.26 +/- 0.19) but not in those in which treatment began the day of collar insertion (I/M, 0.74 +/- 0.14). EPC liposomes did not affect neointimal formation (I/M, 0.50 +/- 0.14 and 0.51 +/- 0.07 in the two treatment groups). Proliferation of smooth muscle cells, assessed by direct incorporation of bromo-2'-deoxyuridine (BrdU) into replicating DNA, was reduced by approximately 30% and 75% in the intimal and medial tissues of rA-IM/A-IM-pretreated rabbits.(ABSTRACT TRUNCATED AT 250 WORDS)