Abstract 1048: High Density Lipoprotein To Macrophage Free Cholesterol Influx And Transfer To Low Density Lipoprotein - New Assays For Dysfunctional High Density Lipoprotein: The Houston Heart Study

Abstract

Free cholesterol (FC) flux between HDL and macrophages switches from efflux to influx as HDL-particle number (HDL-P) and mol% FC increase. Among normolipidemic subjects, ASCVD is more prevalent among patients with HDL that are dysfunctional acceptors of macrophage-FC efflux and among patients with a very high plasma HDL concentration, i.e., hyperalphalipoproteinemia. HDL receptor deficient mice (Scarb1 -/- ), a robust model of hyperalphalipoproteinemia, are ASCVD-susceptible. Scarb1 -/- and wild-type mouse HDL are equally good acceptors of macrophage-FC efflux but Scarb1 -/- HDL are more potent donors of FC influx . This provoked our hypothesis that high HDL-FC bioavailability = HDL-P x HDL-mol% FC is associated with excess human ASCVD. Here we describe assay-validation for HDL-FC influx into macrophages and transfer to LDL. Methods: HDL-[ 3 H]FC Transfer to J774 Macrophages: HDL-[ 3 H]FC from five volunteers were co-incubated with cells for 2 h at 37°C, cells were washed, lipid-extracted, and the extract β-counted. Influx was expressed as pmol FC uptake/mg cell-protein. HDL-[ 3 H]FC Transfer to LDL: LDL and HDL-[ 3 H]FC, each at 0.5 mg protein/mL, were co-incubated for 2 h at 37°C and remaining HDL-associated FC was determined. FC transfer was expressed as pmol FC transferred to LDL. Results: Within-day and day-to-day CVs for FC transfer to LDL were 2.5% and 2.0%, and for influx, 4.3% and 10.7%. HDL-mol % FC and nmol FC/mg protein differed among five donors and correlated with the donor LDL-FC content. HDL-FC transfer to LDL and macrophages increased linearly with HDL-mol% FC. HDL-FC transfer to LDL correlated with influx to macrophages (Figure). Conclusions: Among putatively normolipidemic blood donors, HDL-FC content varies. This is associated with similar variations in HDL-mol% FC, FC transfer to LDL and FC influx to cells. HDL-FC transfer to LDL is a good surrogate of macrophage influx. These assays will be used to determine if a high HDL-FCBI correlates with human ASCVD.