title: 2H2O-based high-density lipoprotein turnover method for the assessment of dynamic high-density lipoprotein function in mice.
abstract: OBJECTIVE
  High-density lipoprotein (HDL) promotes reverse cholesterol transport from peripheral tissues to the liver for clearance. Reduced HDL-cholesterol (HDLc) is associated with atherosclerosis; however, as a predictor of cardiovascular disease, HDLc has limitations because it is not a direct marker of HDL functionality. Our objective was to develop a mass spectrometry-based method for the simultaneous measurement of HDLc and ApoAI kinetics in mice, using a single (2)H2O tracer, and use it to examine genetic and drug perturbations on HDL turnover in vivo.
  
  
  RESULTS
  Mice were given (2)H2O in the drinking water, and serial blood samples were collected at different time points. HDLc and ApoAI gradually incorporated (2)H, allowing experimental measurement of fractional catabolic rates and production rates for HDLc and ApoAI. ApoE(-/-) mice displayed increased fractional catabolic rates (P<0.01) and reduced production rates of both HDLc and ApoAI (P<0.05) compared with controls. In human ApoAI transgenic mice, levels and production rates of HDLc and human ApoAI were strikingly higher than in wild-type mice. Myriocin, an inhibitor of sphingolipid synthesis, significantly increased both HDL flux and macrophage-to-feces reverse cholesterol transport, indicating compatibility of this HDL turnover method with the macrophage-specific reverse cholesterol transport assay.
  
  
  CONCLUSIONS
  (2)H2O-labeling can be used to measure HDLc and ApoAI flux in vivo, and to assess the role of genetic and pharmacological interventions on HDL turnover in mice. Safety, simplicity, and low cost of the (2)H2O-based HDL turnover approach suggest that this assay can be scaled for human use to study effects of HDL targeted therapies on dynamic HDL function.
authors.affiliation: Department of Gastroenterology and Hepatology, Case Western Reserve University School of Medicine, Cleveland Clinic, Cleveland, OH 44195, USA.
authors.forename: Takhar
authors.initials: T
authors.lastname: Kasumov
authors.forename: Belinda
authors.initials: B
authors.lastname: Willard
authors.forename: Ling
authors.initials: L
authors.lastname: Li
authors.forename: Min
authors.initials: M
authors.lastname: Li
authors.forename: Heather
authors.initials: H
authors.lastname: Conger
authors.forename: Jennifer A
authors.initials: JA
authors.lastname: Buffa
authors.forename: Stephen
authors.initials: S
authors.lastname: Previs
authors.forename: Arthur
authors.initials: A
authors.lastname: McCullough
authors.forename: Stanley L
authors.initials: SL
authors.lastname: Hazen
authors.forename: Jonathan D
authors.initials: JD
authors.lastname: Smith
mesh_terms: D000818:Animals; D016632:Apolipoprotein A-I / Q000378:metabolism; D001057:Apolipoproteins E / Q000235:genetics; D050197:Atherosclerosis / Q000378:metabolism*; D002784:Cholesterol / Q000378:metabolism; D003903:Deuterium / Q000493:pharmacokinetics; D004326:Drinking / Q000502:physiology; D005229:Fatty Acids, Monounsaturated / Q000494:pharmacology; D005260:Female; D006801:Humans; D007166:Immunosuppressive Agents / Q000494:pharmacology; D008075:Lipoproteins, HDL / Q000378:metabolism*; D008099:Liver / Q000378:metabolism; D008264:Macrophages / Q000378:metabolism; D008297:Male; D013058:Mass Spectrometry / Q000379:methods* / Q000592:standards*; D051379:Mice; D008822:Mice, Transgenic; D014867:Water / Q000378:metabolism
chemical_list: D016632:Apolipoprotein A-I; D001057:Apolipoproteins E; D005229:Fatty Acids, Monounsaturated; D007166:Immunosuppressive Agents; D008075:Lipoproteins, HDL; D014867:Water; D002784:Cholesterol; D003903:Deuterium; C001996:thermozymocidin
keywords: apolipoprotein A-I; cholesterol; deuterium oxide; lipoproteins, HDL; mass spectrometry; protein biosynthesis