Reconstitution of the lipoprotein cholesteryl ester transfer process using isolated rat ovary plasma membranes.
Steroidogenic cells are able to utilize lipoprotein-derived cholesteryl esters for steroidogenesis without internalizing intact lipoproteins. In the current report, we provide evidence that an early step in this process may be the selective extraction of cholesteryl esters at the cell (plasma membrane) surface. We have used a highly purified plasma membrane preparation from rat luteinized ovaries for incubation with rat- and human-derived high density (HDL) and low density (LDL) lipoproteins. The lipoproteins were modified with residualizing [125I]apoprotein or [3H]cholesteryl ester markers. Following trypsin treatment to remove intact surface-bound apoprotein particles, the membranes were analyzed for transferred radioactive labels. The results show that all the lipoproteins tested could serve as cholesteryl ester donors. Although far more [3H]cholesteryl ester than [125I]apoprotein radioactivity was transferred to plasma membranes in each case, and varied with the ligand used, the total (net) mass of cholesteryl ester transferred was comparable with the different lipoproteins. These data were confirmed using direct chemical methodology. Transfer was found to be specific for cholesteryl esters or ethers and did not involve other lipoprotein core lipids tested. Endomembranes from the same tissue could not substitute for plasma membranes as the primary cholesteryl ester acceptor. These results provide evidence that a reconstituted lipoprotein-plasma membrane system can simulate the cholesteryl ester extraction process described in situ and suggest uses for this methodology in future experiments designed to understand the transfer process.
Shi
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Department of Medicine, Stanford University School of Medicine, Palo Alto, California.
Azhar
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Reaven
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D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002458:Cell Fractionation; D002462:Cell Membrane / Q000737:chemistry; D053480:Cholesterol Ester Transfer Proteins; D002788:Cholesterol Esters / Q000737:chemistry* / Q000235:genetics; D008076:Cholesterol, HDL / Q000737:chemistry* / Q000235:genetics; D008078:Cholesterol, LDL / Q000737:chemistry* / Q000235:genetics; D005260:Female; D006023:Glycoproteins; D006801:Humans; D007700:Kinetics; D008563:Membrane Lipids / Q000737:chemistry* / Q000235:genetics; D010053:Ovary / Q000737:chemistry*; D011499:Protein Processing, Post-Translational; D051381:Rats
C506642:CETP protein, human; D002352:Carrier Proteins; D053480:Cholesterol Ester Transfer Proteins; D002788:Cholesterol Esters; D008076:Cholesterol, HDL; D008078:Cholesterol, LDL; D006023:Glycoproteins; D008563:Membrane Lipids