Apolipoprotein-E (
apoE) protects against
coronary artery disease via hepatic removal of atherogenic remnant
lipoproteins, sequestration of
cholesterol from vessel walls and local
anti-oxidant, anti-platelet and anti-inflammatory actions.
ApoE gene transfer may thus ameliorate a hyperlipidaemic profile and have beneficial effects at lesion sites to prevent or regress
atherosclerosis, a concept endorsed by adenoviral-mediated hepatic expression studies. Here, using plasmid vectors expressing allelic human
apoE2 or
apoE3 isoforms, skeletal muscle was evaluated as an effective secretory platform for
apoE gene augmentation. Transfected myoblasts and myotubes were found to efficiently secrete recombinant
apoE in vitro as spherical 10-16 nm
lipoprotein particles with pre-beta mobility. Intramuscular plasmid injection in
apoE(-/-) mice, which develop spontaneous
atherosclerotic plaque and
xanthoma resulted in expression and secretion of
apoE. Human
apoE mRNA was detected by RT-PCR in injected muscles and, although concentrations of
apoE3, which is rapidly cleared from plasma, were near ELISA detection limits, levels of plasma
apoE2 were measurable (17.5 +/- 4.3 ng/ml). To assess whether muscle-based expression of
apoE2 could inhibit
atherogenesis, long-term follow-up studies were conducted. Although hyperlipidaemia was not reduced in treated animals, end-point pathology showed clear retardation of atherosclerotic and xanthomatous lesions. Up to 9 months following a single
apoE2 plasmid administration, atherosclerotic lesion coverage in proximal aorta was significantly reduced by 20-30% (P < 0.01), whereas development of gross dorsal
xanthoma (>5 mm diameter) was effectively reduced to zero. We conclude that expression of
apoE from ectopic muscle sites has therapeutic potential to limit progression of
atherosclerosis.