Abstract 117: The Impact of Apolipoprotein B Depletion Methods on HDL Subspecies Distribution and Function
Recent failures of high density lipoprotein cholesterol (HDL-C) raising therapies have shifted attention on understanding HDL functionality vs. quantity. For example, the ability of HDL to promote cholesterol efflux has been recently shown to be highly correlated with protection from cardiovascular disease - more so than HDL-C. VLDL and LDL, apoB rich lipoproteins (apoB-Lp), are commonly precipitated from plasma and the remaining HDL is assayed in situ. Two reagents for precipitating apoB-Lp’s are polyethylene glycol (PEG), a neutral polymer that influences their solubility and dextran sulfate/MgCl2+ (DS), a polyvalent/ divalent combination that separates apoB-containing particles by charge. Given the recent focus on measuring HDL function under these precipitating conditions, it is critical to be sure that the constellation of HDL subspecies is not being altered by the apoB-Lp precipitation procedure. Using a gel filtration chromatography system, we compared the HDL subspecies profile in plasma before and after apoB-Lp precipitation by these methods. Compared to plasma alone, apoB-Lp depetion by PEG dramatically altered the HDL subspecies distribution with a loss of larger phospholipid-rich HDL and subsequent generation of smaller particles. This effect may result from the persistent association of PEG with HDL in plasma. The ability of these HDL particles to promote cholesterol efflux was also shifted in line with the phospholipid distribution. Conversely, DS precipitation had no effect on the HDL size distribution as determined by phospholipid and cholesterol profiles and cholesterol efflux pattern. Western blots showed significant decreases in HDL apoE content for the PEG method, but less so for DS. Ongoing work is aimed at characterizing the effect of these methods on other proteomic changes and functions (i.e. antioxidation) as well as comparing these methods to the “gold standard” of immunoaffinity isolation of apoB-Lp. Given the complexity of the HDL proteome and the growing realization that the family is composed of many functional subspecies, it is important that future clinical assays be carefully validated to be sure that important HDL functional particles are not being overlooked.
Author Disclosures: A.S. Shah: None. A. Heink: None. W.S. Davidson: None.
- © 2015 by American Heart Association, Inc.