IN VITRO AND IN VIVO STRUCTURE/FUNCTION ANALYSIS OF LPL AND HL

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As the major enzyme involved in the hydrolysis of triglycerides and phospholipids present in intermediate and high density lipoproteins hepatic lipase (HL) plays a central role in lipoprotein metabolism. Classically, HL function has been primarily ascribed to lipolysis but recent studies have suggested a role of HL in lipoprotein metabolism independent of the hydrolytic function of the enzyme. To investigate the proposed role of hepatic lipase in mediating the clearance of lipoproteins independent of its lipolytic function in vivo, we used recombinant adenovirus (AdV) to express wild type HL (HL-WT, n=3), catalytically inactive HL (HL145G, n=8), and luciferase (Lucif, n=8) in HL deficient mice with increased plasma concentrations of HDL. Day 4 after i.v. injection of 6x10 to the eighth pfu, similar levels of HL-WT and mutants were detected by ELISA in post heparin plasma (HL-WT: 5.9 plus/minus 1.4 ug/ml; HL145G: 5.0 plus/minus 3.4) while post-heparin plasma HL activity was detected only in mice expressing HL-WT (35.5 plus/minus 6.7 u/mol/ml/min). Compared to baseline (day 0) values (mg/dl): HL deficient mice expressing lipolytically inactive HL145G (day 4) had decreased cholesterol (-35%, p<0.00001), phospholipids (-28%, p<0.00001), HDL-cholesterol (-44%, p<0.005). No significant (p>0.3) lipid changes were observed in mice injected with 6x10 to the eighth pfu of Lucif. HL-WT decreased baseline cholesterol (-85% plus/minus 7%), phospholipids -84 plus/minus 6%), HDL-cholesterol (-100 plus/minus 0%) and apoA-I (-82 plus/minus 7%) more significantly than HL145G. These studies demonstrate that adenovirus mediated expression of catalytically inactive HL145G significantly decreased plasma cholesterol, phospholipids and HDL-cholesterol in HL deficient mice. Thus, lipolysis of HDL lipids does not account for all of the HL-induced metabolic changes in HDL. Our study provides in vivo evidence for a role of HL in HDL metabolism independent of its lipolytic function, a process which may involve HL enhanced uptake of lipoproteins by proteoglycan and/or receptor-mediated mechanisms.