title: Novel technique for generating macrophage foam cells for in vitro reverse cholesterol transport studies.
abstract: Generation of foam cells, an essential step for reverse cholesterol transport studies, uses the technique of receptor-dependent macrophage loading with radiolabeled acetylated LDL. In this study, we used the ability of a biologically relevant detergent molecule, lysophosphatidylcholine (lyso-PtdCho), to form mixed micelles with cholesterol or cholesteryl ester (CE) to generate macrophage foam cells. Fluorescent or radiolabeled cholesterol/lyso-PtdCho mixed micelles were prepared and incubated with RAW 264.7 or mouse peritoneal macrophages. Results showed that such micelles were quite stable at 4°C and retained the solubilized cholesterol during one month of storage. Macrophages incubated with cholesterol or CE (unlabeled, fluorescently labeled, or radiolabeled)/lyso-PtdCho mixed micelles accumulated CE as documented by microscopy, lipid staining, labeled oleate incorporation, and by TLC. Such foam cells unloaded cholesterol when incubated with HDL but not with oxidized HDL. We propose that stable cholesterol or CE/lyso-PtdCho micelles would offer advantages over existing methods.
authors.affiliation: Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, Orlando, FL 32827.
authors.forename: Bhaswati
authors.initials: B
authors.lastname: Sengupta
authors.forename: Chandrakala Aluganti
authors.initials: CA
authors.lastname: Narasimhulu
authors.forename: Sampath
authors.initials: S
authors.lastname: Parthasarathy
mesh_terms: D000818:Animals; D001692:Biological Transport; D002460:Cell Line; D002784:Cholesterol / Q000737:chemistry / Q000378:metabolism*; D003902:Detergents / Q000494:pharmacology; D005487:Foam Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D008075:Lipoproteins, HDL / Q000494:pharmacology; D008244:Lysophosphatidylcholines / Q000494:pharmacology; D051379:Mice; D008823:Micelles; D012995:Solubility; D014867:Water / Q000737:chemistry
chemical_list: D003902:Detergents; D008075:Lipoproteins, HDL; D008244:Lysophosphatidylcholines; D008823:Micelles; D014867:Water; D002784:Cholesterol
keywords: NBD-cholesterol; high density lipoprotein; lysophosphatidylcholine