Separation of and cholesterol synthesis by human lymphocytes and monocytes

Abstract

We have devised techniques for the isolation of human monocytes which do not require the adherence of the cells to a surface. In 15 consecutive experiments using density-gradient and counterflow centrifugations, a population of mononuclear cells that was 75 +/- 11% monocytes was obtained within 2 hours of venipuncture. These cells had never been pelleted and represented approximately three-fourths of the monocytes that had been present in the whole blood. In another 22 consecutive experiments using sedimentation in gelatin followed by counterflow and density-gradient centrifugations, a population of lymphocytes that was 99.5 +/- 0.5% pure and a population of monocytes that was 94 +/- 3% pure were obtained within 3 hours of venipuncture. When these freshly isolated cells were incubated in the lipoprotein-deficient fraction of serum (d > 1.21 g/ml) or in solvent-extracted serum, the monocytes incorporated 10-20 times more [2-(14)C]acetate into sterols than did the lymphocytes. Monocytes were seen to constitute between 6 and 46% of the mononuclear cells isolated from normal individuals by the usual density-gradient centrifugation of whole blood on Ficoll-Hypaque. We conclude that future studies of cholesterol metabolism utilizing human mononuclear cells must take into account this large variation in the percentage of monocytes and their disproportionately greater activity during short-term incubations in media that induce sterol synthesis.-Fogelman, A. M., J. Seager, M. Hokom, and P. A. Edwards. Separation of and cholesterol synthesis by human lymphocytes and monocytes.