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Papers In Press, published online ahead of print December 3, 2005
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Children's Hospital of Philadelphia, Philadelphia, PA 19104
Corresponding Author: rothblat{at}email.chop.edu
Abstract: We developed an assay that quantitates bidirectional cholesterol flux between cells and lipoproteins. Incubating Fu5AH cells with increasing concentrations of human serum resulted in increased influx and efflux, however, influx was 2-3 fold greater at all serum concentrations. With apo B-depleted serum the ratio of influx to efflux (I/E) was close to 1, indicating cholesterol exchange. The apo B fraction of serum induced influx and little efflux, with I/E greater than 1. Using BLT-1 to block SR-BI-mediated flux with different acceptors we determined 50% to 70% of efflux was via SR-BI. With HDL, 90% of influx was via SR-BI, whereas with LDL or serum 20% of influx was SR-BI-mediated. Cholesterol-enriched hepatoma cells produced increased efflux without change in influx, resulting in reduced I/E. The assay was applied to cholesterol normal and enriched mouse peritoneal macrophages exposed to serum or LDL. The enrichment enhanced efflux without shifts in influx. With cholesterol-enriched macrophages HDL efflux was enhanced while influx was greatly reduced. With all lipoproteins cholesterol enrichment of MPM led to a reduced I/E. We conclude that this assay can simultaneously and accurately quantitate the cholesterol bidirectional flux and can be applied to a variety of cells exposed to isolated lipoproteins or serum.
Revised on December 2, 2005
Accepted on December 2, 2005
Measurement of cholesterol bidirectional flux between cells and lipoproteins
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