Limits...
Effects of acceptor composition and mechanism of ABCG1-mediated cellular free cholesterol efflux.

Sankaranarayanan S, Oram JF, Asztalos BF, Vaughan AM, Lund-Katz S, Adorni MP, Phillips MC, Rothblat GH - J. Lipid Res. (2008)

Bottom Line: ABCG1 did not enhance influx of FC or cholesteryl oleyl ether (COE) when cells were incubated with radiolabeled HDL(3).ABCG1 expression did not increase the association of HDL(3) with cells.In conclusion, composition and particle size determine the acceptor efficiency for ABCG1-mediated efflux.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, Children's Hospital of Philadelphia, Philadelphia, PA, USA.

ABSTRACT
Among the known mechanisms of reverse cholesterol transport (RCT), ATP binding cassette transporter G1 (ABCG1)-mediated free cholesterol (FC) transport is the most recent and least studied. Here, we have characterized the efficiencies of different acceptors using baby hamster kidney (BHK) cells transfected with human ABCG1 cDNA, which is inducible upon treatment with mifepristone. When normalized on particle number and particle surface area, the acceptor efficiency for FC efflux was as follows: small unilamellar vesicles (SUV)>LDL>reconstituted HDL>HDL(2) = HDL(3). Based on phospholipid content, the order was reversed. ABCG1 also mediated phospholipid efflux to human serum and HDL(3). ABCG1-mediated FC efflux correlated significantly with a number of HDL subfractions and components in serum collected from 25 normolipidemic individuals: apolipoprotein A-II (apoA-II) (r(2) = 0.7), apolipoprotein A-I (apoA-I) (r(2) = 0.5), HDL-C (r(2) = 0.4), HDL-PL (r(2) = 0.4), alpha-2 HDL (r(2) = 0.4), and prebeta HDL (r(2) = 0.2). ABCG1 did not enhance influx of FC or cholesteryl oleyl ether (COE) when cells were incubated with radiolabeled HDL(3). ABCG1 expression did not increase the association of HDL(3) with cells. Compared with control cells, ABCG1 expression significantly increased the FC pool available for efflux and the rate constant for efflux. In conclusion, composition and particle size determine the acceptor efficiency for ABCG1-mediated efflux. ABCG1 increases cell membrane FC pools and changes its rate of desorption into the aqueous phase without enhancing the association with the acceptor.

Show MeSH

Related in: MedlinePlus

ABCG1-mediated free cholesterol efflux to phospholipid-enriched HDL3. Radiolabeled BHK cells were treated with or without mifepristone for 18 h and were incubated with HDL3 (25 μg protein/ml), enriched with different levels of either sphingomyelin (SM; 5-, 10-, 20-fold; A), or 1, 2-dimyristoyl-sn-glycerophosphocholine (DMPC; 1.5-, 2-, 4-fold; B) for 4 h (the control HDL3 PL/protein ratio is 0.45 for SM and 0.48 for DMPC). ABCG1-mediated efflux was the difference between the values for untreated and mifepristone-treated cells. The efflux values are mean ± SD of triplicate measurements (the error bars are inside the symbols). Untreated, □; mifepristone, ▴; ABCG1-specific, •.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2636919&req=5

fig2: ABCG1-mediated free cholesterol efflux to phospholipid-enriched HDL3. Radiolabeled BHK cells were treated with or without mifepristone for 18 h and were incubated with HDL3 (25 μg protein/ml), enriched with different levels of either sphingomyelin (SM; 5-, 10-, 20-fold; A), or 1, 2-dimyristoyl-sn-glycerophosphocholine (DMPC; 1.5-, 2-, 4-fold; B) for 4 h (the control HDL3 PL/protein ratio is 0.45 for SM and 0.48 for DMPC). ABCG1-mediated efflux was the difference between the values for untreated and mifepristone-treated cells. The efflux values are mean ± SD of triplicate measurements (the error bars are inside the symbols). Untreated, □; mifepristone, ▴; ABCG1-specific, •.

Mentions: We next examined the effect of enrichment of HDL3 with DMPC or SM on ABCG1-mediated FC efflux (Fig. 2) by enriching HDL3 with SM (5-, 10-, and 20-fold, Fig. 2A), or DMPC (1.5-, 2-, and 4-fold, Fig. 2B). The FC efflux for untreated and mifepristone-treated cells at a fixed HDL protein concentration increased in parallel with increasing degrees of phospholipid enrichment (PC and SM). However, the ABCG1-mediated FC efflux (the difference between the mifepristone-treated and untreated cells) did not change regardless of the type of phospholipid or the level of the enriched phospholipid (Fig. 2A, B).


Effects of acceptor composition and mechanism of ABCG1-mediated cellular free cholesterol efflux.

Sankaranarayanan S, Oram JF, Asztalos BF, Vaughan AM, Lund-Katz S, Adorni MP, Phillips MC, Rothblat GH - J. Lipid Res. (2008)

ABCG1-mediated free cholesterol efflux to phospholipid-enriched HDL3. Radiolabeled BHK cells were treated with or without mifepristone for 18 h and were incubated with HDL3 (25 μg protein/ml), enriched with different levels of either sphingomyelin (SM; 5-, 10-, 20-fold; A), or 1, 2-dimyristoyl-sn-glycerophosphocholine (DMPC; 1.5-, 2-, 4-fold; B) for 4 h (the control HDL3 PL/protein ratio is 0.45 for SM and 0.48 for DMPC). ABCG1-mediated efflux was the difference between the values for untreated and mifepristone-treated cells. The efflux values are mean ± SD of triplicate measurements (the error bars are inside the symbols). Untreated, □; mifepristone, ▴; ABCG1-specific, •.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2636919&req=5

fig2: ABCG1-mediated free cholesterol efflux to phospholipid-enriched HDL3. Radiolabeled BHK cells were treated with or without mifepristone for 18 h and were incubated with HDL3 (25 μg protein/ml), enriched with different levels of either sphingomyelin (SM; 5-, 10-, 20-fold; A), or 1, 2-dimyristoyl-sn-glycerophosphocholine (DMPC; 1.5-, 2-, 4-fold; B) for 4 h (the control HDL3 PL/protein ratio is 0.45 for SM and 0.48 for DMPC). ABCG1-mediated efflux was the difference between the values for untreated and mifepristone-treated cells. The efflux values are mean ± SD of triplicate measurements (the error bars are inside the symbols). Untreated, □; mifepristone, ▴; ABCG1-specific, •.
Mentions: We next examined the effect of enrichment of HDL3 with DMPC or SM on ABCG1-mediated FC efflux (Fig. 2) by enriching HDL3 with SM (5-, 10-, and 20-fold, Fig. 2A), or DMPC (1.5-, 2-, and 4-fold, Fig. 2B). The FC efflux for untreated and mifepristone-treated cells at a fixed HDL protein concentration increased in parallel with increasing degrees of phospholipid enrichment (PC and SM). However, the ABCG1-mediated FC efflux (the difference between the mifepristone-treated and untreated cells) did not change regardless of the type of phospholipid or the level of the enriched phospholipid (Fig. 2A, B).

Bottom Line: ABCG1 did not enhance influx of FC or cholesteryl oleyl ether (COE) when cells were incubated with radiolabeled HDL(3).ABCG1 expression did not increase the association of HDL(3) with cells.In conclusion, composition and particle size determine the acceptor efficiency for ABCG1-mediated efflux.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, Children's Hospital of Philadelphia, Philadelphia, PA, USA.

ABSTRACT
Among the known mechanisms of reverse cholesterol transport (RCT), ATP binding cassette transporter G1 (ABCG1)-mediated free cholesterol (FC) transport is the most recent and least studied. Here, we have characterized the efficiencies of different acceptors using baby hamster kidney (BHK) cells transfected with human ABCG1 cDNA, which is inducible upon treatment with mifepristone. When normalized on particle number and particle surface area, the acceptor efficiency for FC efflux was as follows: small unilamellar vesicles (SUV)>LDL>reconstituted HDL>HDL(2) = HDL(3). Based on phospholipid content, the order was reversed. ABCG1 also mediated phospholipid efflux to human serum and HDL(3). ABCG1-mediated FC efflux correlated significantly with a number of HDL subfractions and components in serum collected from 25 normolipidemic individuals: apolipoprotein A-II (apoA-II) (r(2) = 0.7), apolipoprotein A-I (apoA-I) (r(2) = 0.5), HDL-C (r(2) = 0.4), HDL-PL (r(2) = 0.4), alpha-2 HDL (r(2) = 0.4), and prebeta HDL (r(2) = 0.2). ABCG1 did not enhance influx of FC or cholesteryl oleyl ether (COE) when cells were incubated with radiolabeled HDL(3). ABCG1 expression did not increase the association of HDL(3) with cells. Compared with control cells, ABCG1 expression significantly increased the FC pool available for efflux and the rate constant for efflux. In conclusion, composition and particle size determine the acceptor efficiency for ABCG1-mediated efflux. ABCG1 increases cell membrane FC pools and changes its rate of desorption into the aqueous phase without enhancing the association with the acceptor.

Show MeSH
Related in: MedlinePlus