Cholesterol Metabolism in the Macrophage

Abstract

Macrophage membrane cholesterol is present in two subcellular cholesterol pools, a rapidly exchanging compartment comprising about two-thirds of the total cholesterol, and a slowly exchanging compartment comprising one-third of the total. The morphological identification of the kinetically distinguishable pools proceeded by alteration of each compartment. Trypsin treatment markedly decreased the rate of cholesterol exchange without removing cholesterol from the membrane. Recovery of normal exchange rates took more than 7 hr and required protein synthesis. This suggested that a plasma membrane receptor is involved in positioning of lipoproteins for exchange, and is consistent with the plasma membrane localization of the rapidly exchanging compartment. Extensive pinocytosis by nondegradable dextran, dextran sulfate, or sucrose resulted in the accumulation of many secondary lysosomes, thus increasing the relative proportion of intracellular membranes. The measurable granule membrane area, cholesterol content, phospholipid content, and the relative size of the slowly exchanging cholesterol compartment all increased. The amount of intracellular membrane altered by extensive phagocytosis of latex particles also increased the size of the slowly exchanging cholesterol compartment. This suggested that the slowly exchanging pool of cholesterol represented the intracellular membranes primarily of lysosomal origin. Rabbit alveolar macrophages and thioglycollate-stimulated peritoneal macrophages contain many secondary lysosomes as a result of multiple bouts of in vivo phagocytosis and pinocytosis. In both of these cells the fast and slow pools are equal in size. The increased cholesterol content was attributable to the increase in the relative size of the slowly exchanging compartment. L-cells and melanoma cells also exchange their cholesterol with that of serum lipoproteins. Both cells contain few cholesterol-rich intracellular membranes, and had lower cellular cholesterol contents. In these cells the slowly exchanging pool was a minor contribution to cell cholesterol. Studies with these cells provided further evidence for the lysosomal membrane and plasma membrane localization of the slowly and rapidly exchanging cholesterol compartments.

Cholesterol Metabolism in the Macrophage

“…This value is quite high as compared with other cells (21,46), remains constant during in vitro cultivation, and is independent of the cholesterol content of the extraceUular medium (18,51,52). The macrophage does not divide under these conditions but gradually increases in size, protein, enzyme content, and the amount of total free cholesterol.…”

“…Inhibitors of pinocytosis and protein synthesis had no effect on cholesterol incorporation during a 4 hr exposure in the presence of isotope. Puromycin and cycloheximide had significant effects after 5 hr of continuous exposure (46). Both subcellular compartments were labeled in the presence of the inhibitors.…”

“…Both subcellular compartments were labeled in the presence of the inhibitors. This was shown by a washout experiment performed after labeling of the cells in the presence of sodium fluoride, and also by fractionation after labeling in the presence of fluoride (46).…”

Cholesterol Metabolism in the Macrophage

“…However, there are a number of storage diseases in which cholesterol accumulates in tissues, although the lesion is not known to involve cholesterol metabolism. We have demonstrated that net increases in both cholesterol and phospholipid can be considerable in cells containing storage granules, the contents of which are free of lipid (2). These increases are due to the membrane of the secondary lysosomes.…”

“…Since the value of the rate constant k32 describing the relative movement of cholesterol from the lysosomal membrane to the plasma membrane during exchange is similar in value to k~, the rate-limiting reaction probably occurs at the same step for both exchange and excretion. In the absence of membrane apposition or translocation, an intracellular soluble "carrier" may mediate the transfer of cholesterol between the lysosomal membrane and the plasma membrane (2). Cholesterol may therefore move from within the lysosome to the lysosomal membrane readily, whereas the transfer to the intracellular carrier, or its concentration may be rate limiting.…”