Lysosomal accumulation of cholesterol and sphingomyelin: evidence for inhibition of acid sphingomyelinase

Abstract

The subcellular site of accumulation of non-esterified cholesterol and sphingomyelin in lipid-laden livers and spleens from rats given multiple intravenous injections with liposomes made up of these lipids were investigated by morphological and biochemical techniques. The subcellular fractionation of liver homogenates from cholesterol-sphingomyelin treated rats followed by lipid and enzymatic analyses of the fractions revealed that most of the accumulating lipid was present in very low density lysosomes floating in the post-microsomal supernatant fraction. The low density lysosomes exhibited good latency and had a very much lower relative activity of sphingomyelinase compared with values for N-acetyl-beta-glucosaminidase. Multiple injections of sphingomyelin-cholesterol liposomes resulted in splenomegaly. The spleen homogenates of the treated rats showed a many-fold increase in the concentration of sphingomyelin, of non-esterified cholesterol and in the activity of N-acetyl-beta-glucosaminidase over corresponding values for rats injected with saline. Electron microscopy of liver and spleen sections from treated rats revealed distinctive polymorphic intracellular inclusions bound by a membrane and containing numerous osmiophilic bodies. Structures identical to the storage inclusions seen in fixed liver sections from treated rats were also seen by electron microscopy in the post-microsomal fraction of these livers. The results suggest that sphingomyelin and non-esterified cholesterol accumulate in lysosomes when they occur in cells in excess of that structurally associated with cellular membranes.