Intracellular regulation of lipoprotein lipase in human monocyte-derived macrophages.

The intracellular pathway of lipoprotein lipase (LPL) has been examined in human monocyte-derived macrophages in culture. These cells were previously shown to synthesize and constitutively secrete LPL. The secretion is dependent on new enzyme synthesis. 6-d-old human monocytes have stores of mRNA for linear release of LPL up to 24 h. Enzyme activity in cells and in culture medium was almost completely inhibited by 24 h treatment with tunicamycin, an inhibitor of glycosylation. In monensin-treated cells a pronounced increase in enzyme activity was found, whereas the secreted activity was markedly reduced. This indicates that LPL in human monocytes is processed through a pH sensitive part of the Golgi complex and that the terminal glycosylation is not needed for the expression of its catalytic activity. Our results suggest that lysosomal function is not important in secretion of the enzyme, whereas vesicular transport seem to be involved in regulating LPL in human monocyte-derived macrophages in culture.

Effect of high-density lipoproteins on cholesterol efflux and esterification in lipid-enriched human skin fibroblasts.

The ability of high-density lipoprotein (HDL) to reduce the cholesterol content was studied in cultured fibroblasts enriched with cholesterol esters. Incubation of cholesterol-enriched cells with HDL in a final concentration of 1 g protein/l for 24 h reduced the total and esterified cholesterol content by 23% as compared with control fibroblasts incubated with albumin. Similar cholesterol efflux was obtained with HDL isolated from lecithin:cholesterol acyltransferase (LCAT)-deficient plasma. The HDL3 subfraction isolated by rate-zonal ultracentrifugation contained the major part of the cholesterol-depleting effect. HDL or HDL3 decreased CoA:cholesterol acyltransferase (ACAT) activity to 5% of the level found in control fibroblasts within 8 h of incubation. These findings suggest that ACAT activity is sensitive to a pool of intracellular cholesterol, which can be mobilized by the addition of HDL to the culture medium, and that ACAT activity is a useful measure of cholesterol efflux from cultured fibroblasts.

Synthesis and secretion of lipoprotein lipase by human monocyte-derived macrophages.

Human monocytes isolated from either defibrinated blood or buffy coat were shown to produce and secrete lipoprotein lipase during culture. The secretion occurred constitutively. Low levels of enzyme activity in the medium from freshly isolated cells increased with time of incubation, and maximal activity was attained after 9 days. The addition of heparin resulted in a substantial increase of enzyme activity in the culture medium. The optimal concentration of heparin was about 2 U/ml. The production of lipoprotein lipase was dependent on the presence of serum in the culture medium, and the optimal supplementation of serum was 25-50%.