Differential Effect of Genistein on the Stimulation of Cholesterol Production by Basic Protein II in Normal and HyperapoB Fibroblasts

Abstract

Abstract-We studied further the basis for the abnormal effect of human serum basic protein II (BP II) on cholesterol production in hyperapobetalipoproteinemia (hyperapoB) fibroblasts and whether this effect involves protein tyrosine kinase phosphorylation (TKP). Genistein, a specific inhibitor of TKP was used as a probe. Compared with normal cells, BP II stimulated significantly the cellular mass of total cholesterol (6.4-fold), unesterified cholesterol (3.6-fold), and esterified cholesterol (6.7-fold) in hyperapoB fibroblasts. The addition of genistein to BP II in hyperapoB cells markedly inhibited these abnormal stimulatory effects of BP II on cell sterol mass. In normal cells, the addition of genistein to BP II produced an opposite effect: a marked stimulation in the mass of total (5.5-fold) and esterified (18.3-fold) cholesterol and a decrease in unesterified cholesterol (3.4-fold). These effects of genistein on the formation of cellular cholesterol by BP II were both time and concentration dependent. The inhibition of the stimulatory effect of BP II on cholesterol production by genistein in hyperapoB cells may be mediated through 3-hydroxy 3-methylglutaryl coenzyme A reductase, the rate-limiting enzyme of cholesterol biosynthesis, since the rate of incorporation of [ 14 C]acetate, but not [ 3 H]mevalonate, into unesterified cholesterol was decreased by genistein in the hyperapoB cells. When the mass of cell total cholesterol in the cells treated with BP II was subtracted from those treated with BP II plus genistein, a negative number was produced in each of the six hyperapoB cell lines, while each of the normal cell lines retained a positive number. The mean difference for the mass of total cholesterol between the hyperapoB and normal fibroblasts under these conditions was 128.2 nmol/mg cell protein, a difference that was separated by Ͼ3 SD. This study supports further the tenet that there is a defect in the response of hyperapoB cells to BP II and that this defect results in an abnormality in cholesterol metabolism that appears mediated through a protein TKP-mediated process. (Arterioscler Thromb Vasc Biol. 1998;18:57-64.) Key Words: tyrosine kinase phosphorylation Ⅲ familial combined hyperlipidemia Ⅲ atherosclerosis H yperapobetalipoproteinemia is a lipoprotein disorder that is prevalent in patients with premature CAD.1,2 HyperapoB is characterized by an increased number of small, dense LDL particles, a phenotype shared with familial combined hyperlipidemia, LDL subclass pattern B, familial dyslipidemic hypertension, and syndrome X.2,3 Two metabolic defects have been described in patients with hyperapoB. First, there is overproduction of apolipoprotein B and VLDL particles. 3,4 Second, the clearance of postprandial triglyceride-rich particles is delayed, accompanied by an abnormal removal of FFA. 5,6 The stimulation of the incorporation of FFA into triglycerides of adipocytes by a low-molecular-weight serum BP (called ASP) is deficient in hyperapoB cells.7 It has been postulated that this defect leads to ...

Cholesterol Depletion Inhibits Epidermal Growth Factor Receptor Transactivation by Angiotensin II in Vascular Smooth Muscle Cells

“…After saponification, the lipids were extracted with 2 ml of hexane. The hexane was evaporated under nitrogen and radioactivity was then quantified by a ␤-counter as described previously (18).…”