Lipid oxidation, lipoprotein cell-association and ceroid accumulation in P388D1 macrophage-like cells

Flow cytometry can be used to quantify the accumulation of ceroid in macrophages, the result of cellular handling of certain lipoproteins. Using P388D1 cells, a murine-derived macrophage-like cell line, the effect of the lipophilic antioxidant, DL-alpha-tocopherol, upon the uptake and accumulation of ceroid by the cells was monitored on culture with artificial lipoproteins containing a single lipid species. Ceroid accumulation was greater for artificial lipoprotein composed of BSA complexed with cholesteryl arachidonate, than with cholesteryl linoleate. alpha-Tocopherol inhibited the ceroid accumulation, which was also dependent upon cell density. Thus, since these findings are similar to recent observations in primary cultures of murine peritoneal macrophages, it would appear that macrophage-like cell lines such as P388D1 cells are appropriate for the study of potential agonists and antagonists of lipid oxidation. Culture of P388D1 cells with oxidised human low-density lipoprotein (LDL) also resulted in ceroid formation, shown to be dependent upon the level of LDL oxidation as assessed by thiobarbituric acid-reactivity, the xylenol orange assay of peroxides and gas chromatographic analysis of cholesterol and fatty acid content. Ceroid accumulation reflected changes in the level of LDL oxidation better than did the cell association of oxidised radiolabelled LDL, monitored as that bound and retained by the cell.