UBC Theses and Dissertations
Investigation of the function of lipoprotein-associated phospholipase A₂ in lipid metabolism Yang, Ming
Lipoprotein-associated phospholipase A₂ (Lp-PLA₂) has been recognized in many population studies as an independent risk factor for cardiovascular and cerebrovascular disease. However, it remains unclear if there is a direct mechanistic link between Lp-PLA₂ function and atherogenesis. The complexity of this issue is related to the fact that the catalytic products of the enzymatic reaction catalyzed by Lp-PLA₂ namely lysophosphatidylcholine (lysoPC) and oxidized fatty acids, as well as the substrates of Lp-PLA₂, including oxidized phospholipids and platelet-activating factor that can be considered to be proatherogenic. Lp-PLA₂ is secreted by macrophages, hepatocytes and platelets, and circulates primarily with LDL, HDL, and Lp(a). Whether these native and oxidized forms of lipoproteins affect Lp-PLA₂ expression from macrophages and hepatocytes is largely unknown. Furthermore, the specific role of Lp-PLA₂ in lipoprotein metabolism remains unclear. In the present study, we observed that oxidized phospholipids, oxysterols and oxidized fatty acids decreased Lp-PLA₂ activity without affecting Lp-PLA₂ mRNA and protein expression from human monocyte-derived macrophages (MDM) or HepG2 cells. For the investigation of the role of Lp-PLA₂ in lipid metabolism, we constructed Lp-PLA₂ Gain-of-Function and Loss-of-Function lentivirus, and purified 2 types of human recombinant Lp-PLA₂ proteins, active Lp-PLA₂ and catalytically inactive Lp-PLA₂. Human Lp-PLA₂ protein decreased the cellular association of oxLDL and oxLp(a) by human MDM and HepG2 cells whereas the inactive form of Lp-PLA₂ did not significantly change oxidized lipoprotein cellular association by either cell type. OxLDL pretreated with Pefabloc (serine esterase inhibitor) increased oxLDL uptake by MDM and HepG2 cells compared to untreated oxLDL. To assess whether the reduced cellular association mediated by Lp-PLA₂ was due to the hydrolysis of the scavenger receptor ligands, i.e. oxPCs by Lp-PLA₂, we measured the concentration of lysophosphatidylcholine (lysoPC) in lipoprotein fractions after Lp-PLA₂ treatment. LysoPC was increased by active Lp-PLA₂ compared to inactive Lp-PLA₂ for oxLDL and oxLp(a), respectively. In summary, we report the ability of endogenous and exogenous Lp-PLA₂ to reduce oxidized lipoprotein cellular association in human-derived macrophages and HepG2 cells, a phenomenon likely resulting from the removal of ligands from lipoproteins for cell surface scavenger receptors.
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