Modulating lipoprotein transcellular transport and atherosclerotic plaque formation in apoe-/- mice via nanoformulated lipid-methotrexate conjugates

Macrophage inflammation and maturation into foam cells, following the engulfment of oxidized low-density lipoproteins (oxLDL), are major hallmarks in the onset and progression of atherosclerosis. Yet, chronic treatments with anti-inflammatory agents, such as methotrexate (MTX), failed to modulate disease progression, possibly for the limited drug bioavailability and plaque deposition. Here, MTX-lipid conjugates, based on 1,2-distearoyl-sn-glycero-3-phosphoethanolamine (DSPE), were integrated in the structure of spherical polymeric nanoparticles (MTX-SPN) or intercalated in the lipid bi-layer of liposomes (MTX-LIP). Although, both nanoparticles were colloidally stable with an average diameter of ∼ 200 nm, MTX-LIP exhibited a higher encapsulation efficiency (> 70%) and slower release rate (∼50% at 10h) compared to MTX-SPN. In primary bone marrow derived macrophages (BMDM), MTX-LIP modulated the transcellular transport of oxLDL more efficiently than free MTX mostly by inducing a 2-fold overexpression of ABCA1 (regulating oxLDL efflux); while the effect on CD36 and SRA-1 (regulating oxLDL influx) was minimal. Furthermore, in BMDM, MTX-LIP showed a stronger anti-inflammatory activity than free MTX reducing the expression of IL-1β by 3-fold, IL-6 by 2-fold and also moderately of TNF-α. In 28 days high-fat-diet fed apoE-/- mice, MTX-LIP reduced the mean plaque area by 2-fold and the hematic amounts of RANTES by half as compared to free MTX. These results would suggest that the nano-enhanced delivery to vascular plaques of the anti-inflammatory DSPE-MTX conjugate could effectively modulate the disease progression by halting monocytes’ maturation and recruitment already at the onset of atherosclerosis