Abstract 538: Metabolism of Multiple Apolipoproteins Across HDL Size in Humans

Abstract

Introduction: Studying the kinetics of several HDL apolipoproteins together may help elucidate their function in HDL metabolism. We present multicompartmental analysis describing the metabolism of several apolipoproteins across 5 HDL size fractions. Methods and Results: Three participants were infused with a bolus of D3-Leu tracer, and blood was collected for 70 hrs. ApoA-I-HDL was prepared by immunoaffinity purification, separated into 5 size fractions, preβ, α3, α2, α1, and α0, by ND-PAGE, and in-gel trypsinized for mass spectrometry. We monitored 7 proteins that likely affect HDL metabolism - apoA-I, apoA-II, apoE, apoM, apoC-III, apoA-IV, and apoD. We used stable isotope labeled peptide standards to quantify pool size and high resolution parallel reaction monitoring to measure tracer enrichment in these 7 proteins in the 5 HDL size fractions. These data were then used for multicompartmental analysis to describe the kinetic behaviors across HDL size for each apolipoprotein, with the exception of apoD. All size fractions with discernible enrichment curves for at least 2 participants were included in the final model for each apolipoprotein (Fig.1). The majority of apoA-I α HDL originated form the source compartment, presumably the liver and/or small intestine, while apoA-I preβ HDL came primarily from α3. Size expansion pathways (preβ to α1/2, and α3 to α2) contributed only slightly to apoA-I metabolism. Similarly to apoA-I on the α sizes, the majority of the other apolipoproteins appear on each HDL size fraction directly from the source compartment. Only minor flux pathways from smaller to larger HDL sizes were identified: apoA-II, α3 to α2; apoE, α3 to α2, and α3 to α1. Conclusions: This study supports the new model of HDL metabolism in which HDL and its attached apolipoproteins are metabolized mainly within each HDL size fraction. Pathways between HDL sizes only provide a small contribution to the metabolism of each apolipoprotein.