出版社：American Society for Biochemistry and Molecular Biology
摘要：We developed an assay that quantitates bidirectional cholesterolflux between cells and lipoproteins. Incubating Fu5AH cellswith increasing concentrations of human serum resulted in increasedinflux and efflux; however, influx was 2- to 3-fold greaterat all serum concentrations. With apolipoprotein B (apoB)-depletedserum, the ratio of influx to efflux (I/E) was close to 1, indicatingcholesterol exchange. The apoB fraction of serum induced influxand little efflux, with I/E > 1. Using block lipid transport-1to block scavenger receptor class B type I (SR-BI)-mediatedflux with different acceptors, we determined that 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 effluxwithout a change in influx, resulting in reduced I/E. The assaywas applied to cholesterol-normal and -enriched mouse peritonealmacrophages exposed to serum or LDL. The enrichment enhancedefflux without shifts in influx. With cholesterol-enriched macrophages,HDL efflux was enhanced and influx was greatly reduced. Withall lipoproteins, cholesterol enrichment of murine peritonealmacrophages led to a reduced I/E. We conclude that this assaycan simultaneously and accurately quantitate cholesterol bidirectionalflux and can be applied to a variety of cells exposed to isolatedlipoproteins or serum.Supplementary key words efflux • influx • macrophages • hepatoma • ATP binding cassette transporter A1 • scavenger receptor class B type I • high density lipoprotein
