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The macrophage-like cell line, J774, and its variant, CT2, were used to study the effect of verapamil on metabolism of esterified cholesterol. The cells had been labelled with 3H-cholesterol for 24 hours and thereafter acetylated low density lipoprotein (LDL) or d less than 1.019 g/ml fraction of hypercholesterolemic rabbit plasma was added. After an additional 24 hours, 60% or 40% of the label, respectively, was recovered in esterified cholesterol in control dishes, but only 2% to 15%, in the presence of 50 microM verapamil. Enhancement of esterification of cellular 3H-cholesterol was also obtained by addition of liposomes with a cholesterol/phosphatidylcholine molar ratio of 2:1. This reaction was almost completely inhibited by compound 58-035, an acyl-CoA:cholesterol acyltransferase (ACAT) inhibitor, while addition of verapamil resulted in a dose-dependent partial inhibition which was evident after 4 hours. To determine whether the reduction in cholesterol esterification was due to a direct inhibition of ACAT or to an effect of verapamil on the transport of cholesterol to the site of ACAT activity, the cells were incubated with acetylated LDL labelled with 3H-cholesteryl ester, and the amount of uptake, lysosomal hydrolysis, and cytoplasmic reesterification was determined. In control cells and in cells exposed to the ACAT inhibitor, more than 98% of the 3H-cholesteryl ester taken up had been hydrolyzed. Reesterification of 3H-cholesterol was negligible in the presence of the ACAT inhibitor and amounted to 37.1% in control cells. When verapamil was present, reesterification of 3H-cholesterol to cholesteryl ester was only 4%, while the amount of 3H-cholesteryl ester hydrolyzed was 58% of that taken up.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of verapamil on cholesteryl ester hydrolysis and reesterification in macrophages.