Importance of macrophage cholesterol content on the flux of cholesterol mass

Sandhya Sankaranarayanan; Margarita de la Llera-Moya; Denise Drazul-Schrader; Bela F. Asztalos; Ginny L. Weibel; George H. Rothblat

doi:10.1194/jlr.M008441

Importance of macrophage cholesterol content on the flux of cholesterol mass

^*Division of Gastroenterology, Hepatology, and Nutrition, The Children's Hospital of Philadelphia, Philadelphia, PA
^†Lipid Metabolism Laboratory, Tufts University, Boston, MA

Abstract

Net flux of cholesterol represents the difference between efflux and influx and can result in net cell-cholesterol accumulation, net cell-cholesterol depletion, or no change in cellular cholesterol content. We measured radiolabeled cell-cholesterol efflux and cell-cholesterol mass using cholesterol-normal and -enriched J774 and elicited mouse peritoneal macrophage cells. Net cell-cholesterol effluxes were observed when cholesterol-enriched J774 cells were incubated with 3.5% apolipoprotein (apo) B depleted human serum, HDL₃, and apo A-I. Net cell-cholesterol influxes were observed when cholesterol-normal J774 cells were incubated with the same acceptors except apo A-I. When incubated with 2.5% individual sera, cholesterol mass efflux in free cholesterol (FC)-enriched J774 cells correlated with the HDL-cholesterol (HDL-C) concentrations (r² = 0.4; P=0.003), whereas cholesterol mass influx in cholesterol-normal J774 cells correlated with the LDL cholesterol (LDL-C) concentrations (r² = 0.6; P<0.0001) of the individual sera. A positive correlation was observed between measurements of [³H]cholesterol efflux and reductions in cholesterol mass (r² = 0.4; P=0.001) in FC-enriched J774 cells. In conclusion, isotopic efflux measurements from cholesterol-normal or cholesterol-enriched cells provide an accurate measurement of relative ability of an acceptor to remove labeled cholesterol under a specific set of experimental conditions, i.e., efflux potential. Moreover, isotopic efflux measurements can reflect changes in cellular cholesterol mass if the donor cells are enriched with cholesterol.

Footnotes

↵1 To whom correspondence should be addressed. e-mail:rothblat{at}email.chop.edu
Abbreviations:

ABCG1

ATP binding cassette transporter G1

AcLDL

acetylated low density lipoprotein

apo

apolipoprotein

CE

cholesteryl ester

FC

free cholesterol

GLC

gas liquid chromatography

HDL-C

HDL-cholesterol

IMT

intima media thickness

LDL-C

LDL cholesterol

MPM

mouse peritoneal macrophage

PEG

polyethylene glycol

RCT

reverse cholesterol transport

SR-BI

scavenger receptor class B type I

TC

total cholesterol
This work was supported by National Institutes of Health grant, HL-22633 (G.H.R., M.L.L-M., G.L.W., S.S., D.D-S.). Its contents are solely the responsibility of the authors and do not necessarily represent the official views of the National Institutes of Health. Part of the results in this manuscript were presented at a “meeting on high density lipoproteins (HDL)” held on June 19 and 20, 2009, in Newport, RI. The proceedings of this meeting have been published as a chapter in the book, “High Density Lipoproteins, Dyslipidemia, and Coronary Heart Disease” (Schaefer, E.J., editor; Springer Science+Business Media).