Cholesterol and lipoprotein metabolism of human promyelocytic leukemic HL-60 induced macrophages.

Abstract

Human promyelocytic leukemic cells (HL-60) possess well regulated expression of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, acylCoA:cholesterol acyltransferase (ACAT), and receptor-mediated low density lipoprotein (LDL) catabolism, but lack receptor-mediated acetyl-LDL processing. Differentiation of HL-60 cells with tetramyristic phorbol acetate (TPA) is accompanied by the loss of receptor-mediated LDL degradation and no expression of a functionally active scavenger receptor. 1,25-Dihydroxyvitamin D₃ (D₃)-induced HL-60 macrophages possess specific and saturable receptor-mediated binding for LDL, with an apparent K(d) of 29 μg/ml and a B(max) of 219 ng/mg. Receptor-mediated LDL degradation is specific for apoB and apoE containing lipoproteins; it is calcium dependent, and is inhibited by pronase and chloroquine. Differentiation of HL-60 cells with D₃ for 2 days induces a 45-fold increase in acetyl-LDL degradation rate compared to undifferentiated cells. Receptor-mediated degradation of acetyl-LDL is specific for acetyl-LDL, calcium independent, inhibited by chloroquine, pronase and fucoidin treatment, and is not regulated by cellular cholesterol. Acetyl-LDL binding studies demonstrated a K(d) of 36 μg/ml and a B(max) 313 ng/mg. Delivery of cholesterol via receptor-mediated catabolism of LDL or acetyl-LDL results in significant suppression of sterol synthesis and HMG-CoA reductase activity, and significant induction of ACAT activity relative to macrophages incubated with LPDS (P < 0.001). However, receptor-mediated degradation of acetyl-LDL, but not LDL, significantly increases cholesteryl ester content (P < 0.001). D₃-induced HL-60 macrophages incubated with or without LDL for 48 hr exhibited large empty vacuoles with little or no lipid stainable material. In contrast, macrophages incubated with acetyl-LDL exhibited a dramatic increase in lipid stainable material which imparted the macrophages with a foamy appearance. In conclusion, HL-60 cells treated with D₃ for 48 hr undergo activation differentiation assuming the structural and functional characteristics of human monocyte-derived macrophages. Thus, D₃-induced HL-60 macrophages are a suitable in vitro system to study lipoproteins and cholesterol regulation as related to macrophage involvement in atherosclerosis.