Measurement of Reverse Cholesterol Transport Pathways in Humans: In Vivo Rates of Free Cholesterol Efflux, Esterification, and Excretion

Abstract

Background--Reverse cholesterol transport from peripheral tissues is considered the principal atheroprotective mechanism of high-density lipoprotein, but quantifying reverse cholesterol transport in humans in vivo remains a challenge. We describe here a method for measuring flux of cholesterol though 3 primary components of the reverse cholesterol transport pathway in vivo in humans: tissue free cholesterol (FC) efflux, esterification of FC in plasma, and fecal sterol excretion of plasma-derived FC.Methods and Results--A constant infusion of [2,[3][4][5][6][7][8][9][10][11][12][13] C 2 ]-cholesterol was administered to healthy volunteers. Three-compartment SAAM II (Simulation, Analysis, and Modeling software; SAAM Institute, University of Washington, WA) fits were applied to plasma FC, red blood cell FC, and plasma cholesterol ester 13 C-enrichment profiles. Fecal sterol excretion of plasma-derived FC was quantified from fractional recovery of intravenous [2,[3][4][5][6][7][8][9][10][11][12][13] C 2 ]-cholesterol in feces over 7 days. We examined the key assumptions of the method and evaluated the optimal clinical protocol and approach to data analysis and modeling. A total of 17 subjects from 2 study sites (n=12 from first site, age 21 to 75 years, 2 women; n=5 from second site, age 18 to 70 years, 2 women) were studied. Tissue FC efflux was 3.79±0.88 mg/kg per hour (mean ± standard deviation), or %8 g/d. Red blood cell-derived flux into plasma FC was 3.38±1.10 mg/kg per hour. Esterification of plasma FC was %28% of tissue FC efflux (1.10±0.38 mg/kg per hour). Recoveries were 7% and 12% of administered [2,[3][4][5][6][7][8][9][10][11][12][13] C 2 ]-cholesterol in fecal bile acids and neutral sterols, respectively.Conclusions--Three components of systemic reverse cholesterol transport can be quantified, allowing dissection of this important function of high-density lipoprotein in vivo. Effects of lipoproteins, genetic mutations, lifestyle changes, and drugs on these components can be assessed in humans. ( J Am Heart Assoc. 2012;1:e001826 doi: 10.1161/JAHA.112.001826)Key Words: cholesterol efflux • esterification • reverse cholesterol transport • isotope labeling, stable • sterol excretion T he regulation of cellular cholesterol homeostasis is crucial for membrane function and cell survival and is maintained by multiple mechanisms, including control of uptake, synthesis, storage, and efflux. Compared to the pathways of cellular uptake and de novo synthesis of cholesterol, however, less information exists about the control of flux though pathways that remove cholesterol from cells and from the whole organism, 1,2 particularly in humans. [3][4][5] These pathways collectively have been termed reverse cholesterol transport (RCT). RCT is postulated to play a fundamental role in cholesterol homeostasis and distribution among tissues 5 and thereby in the development and reversal of atherosclerosis. 6,7 The atheroprotective effects of high-density lipoprotein cholesterol (HDL-C) in both human and animal studies often hav...

Cholesterol metabolism in cholestatic liver disease and liver transplantation: From molecular mechanisms to clinical implications

“…Under normal circumstances, the liver is the primary site of cholesterol biosynthesis and storage [12] . The liver is also the principal site of cholesterol excretion, converting cholesterol to bile acids and removing free cholesterol as neutral sterols via biliary excretion [4,5,13,14] . Since the liver plays a central role in cholesterol metabolism, liver disease can impact cholesterol metabolism, depending on the type of liver injury (parenchymal, cholestatic, or mixed) [15] .…”

In vivo tissue cholesterol efflux is reduced in carriers of a mutation in APOA1

“…Two and seventeen hours after start of the infusion, two standardized light meals were served. For clinical applicability, the infusion protocol originally developed by Turner et al ( 18 ) was slightly modifi ed regarding the duration of 13 C-C infusion. Turner et al ( 18 ) examined the effect of infusion time on the kinetics: small but systematic increases in fl uxes 1 and 3 were observed as the infusion time was shortened.…”

High-Density Lipoprotein

“…In clinical studies, measurement of fecal sterol excretion or the recently described isotopic dilution method does not distinguish macrophage-specific RCT from general RCT. 59 …”