Combined effects of sphingomyelin and cholesterol on the hydrolysis of emulsion particle triolein by lipoprotein lipase

Sphingomyelin (SM) is one of the major lipids in lipoproteins. However, its function in lipoprotein metabolism is unknown. In an attempt to understand the role that this lipid plays in modulation of lipoprotein lipase (LPL)-mediated hydrolysis, triolein-based emulsion particles containing 15% (physiological concentration) and 30% of the phospholipid content as SM together with phosphatidyl choline were used as substrate for the enzyme. Using a continuous fluorescence displacement assay to measure triglyceride (triolein) hydrolysis, it is shown that LPL activity was not modified by physiological concentrations of SM. However, under these assay conditions the presence of 30% SM inhibited LPL hydrolysis. SM and cholesterol (a normal component of the lipoprotein surface monolayer) become closely associated in phospholipid monolayers and bilayers. Incorporation of cholesterol into emulsion particles containing only PC increased LPL activity, but this increase was reduced by the additional presence of a physiological concentration (15%) of SM. These model studies suggest that the ratio, cholesterol:SM, in the monolayer may regulate the hydrolytic activity of the LPL. The production of ceramide by sphingomyelinase pre-treatment of emulsion particles containing SM leads to a two- to three-fold increase in LPL activity. This effect was dependent on sphingomyelinase concentration and time of pre-incubation and was not seen with cholesterol containing substrates. The ability of apolipoprotein CII to enhance LPL-catalysed triolein hydrolysis was not affected by the presence of SM; however, the stimulatory effect of this apolipoprotein was attenuated by pre-treatment of emulsion particles with sphingomyelinase. In summary, physiological concentrations of SM can inhibit the hydrolysis of cholesterol-containing emulsion particles; while pre-treatment of SM containing emulsion particles with sphingomyelinase in the absence of cholesterol can increase LPL-mediated triglyceride hydrolysis. (C) 1997 Elsevier Science B.V.