Abstract
The scavenger receptor class B type I (SR-BI) binds to HDL and mediates the selective uptake of cholesterol esters from HDL to cells. SR-BII is an alternatively spliced product of the SR-BI gene that only differs in the C-terminal cytoplasmic domain. Previous studies with male mice demonstrated that SR-BII comprises about 12% of the total SR-BI/SR-BII present in liver. In the current studies we used a liver cell line, HepG2, and a rat estrogen replacement model to examine the effects of estrogen on the expression of SR-BII. HepG2 cells express SR-BI but not SR-BII. SR-BI/SR-BII-blocking antibodies demonstrated that HepG2 cells selectively internalize cholesterol esters in a SR-BI-dependent manner. Incubation of HepG2 cells with 10 pM of 17β-estradiol for 12 h eliminated the expression of SR-BI and promoted the up-regulation of SR-BII. Radiolabeled HDL-binding studies demonstrated that 17β-estradiol increased the number of HDL binding sites by 3-fold in HepG2 cells. However, 17β-estradiol-treated cell internalized approximately 25% less cholesterol ester than vehicle-only-treated cells. The livers obtained from male rats and ovariectomized female rats contained SR-BI and a small amount of SR-BII. In contrast, the livers obtained from intact female rats and ovariectomized female rats receiving estrogen replacement contained SR-BII and a small amount of SR-BI. The amount of SR-BI and SR-BII in adrenal tissue was not affected by any of the experimental treatments. We conclude that estrogen up-regulates SR-BII in HepG2 cells and rat liver. —Graf G. A., K. L. Roswell, and E. J. Smart. 17β-estradiol promotes the up-regulation of SR-BII in HepG2 cells and in rat livers.
Highlights
The scavenger receptor class B type I (SR-BI) binds to HDL and mediates the selective uptake of cholesterol esters from HDL to cells
To determine if alterations in SR-BI and SR-BII levels affected the ability of HepG2 cells to take up cholesterol ester selectively, cells were incubated with 10 pM 17-estradiol for
In the present study we demonstrated that as little as 10 pM of 17-estradiol could induce the complete loss of SR-BI and the dramatic up-regulation of SR-BII in HepG2 cells, a liver-derived cell line
Summary
Degraded was determined by measuring the amount of non-TCA precipitatable 125I in the cell medium. Sample buffer (5ϫ) consisted of 0.31 M Tris (pH 6.8), 2.5% (w/v) SDS, tions were determined by radioimmunoassay as described [19]. 137 mM NaCl. Blotting buffer consisted of TBS plus 0.5% Tween 20 and 5% dry milk. Proteins were diluted in 1ϫ sample buffer plus 1.2% (v/v) -mercaptoethanol and heated to 95ЊC for 5 min immediately prior to loading. Primary antibodies were diluted in blotting buffer and incubated with blocked membranes for 60 min at 22ЊC. Horseradish peroxidase conjugated IgGs directed against the appropriate host IgGs were diluted and incubated with membranes as described for primary antibodies. The selective uptake of cholesterol esters from HDL into cells was determined using dual-labeled HDL 3 as described previously [15]. Uptake was terminated by aspirating the medium and washing the cell monolayers four times with Tris-saline (4ЊC).
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