Characterization of triglyceride lipase activities in rat heart

Abstract

Triglyceride lipase activity was determined in particulate and soluble fractions from rat heart homogenates. Lipoprotein lipase activity was identified in the 100 000 × g supernatant fraction by the following in vitro criteria: stimulation of lipase activity by serum; maximal activity at alkaline pH (pH 8.2 to 8.4); and inhibition of lipase activity with NaCl. Triglyceride lipase activity was also determined under conditions in which the contribution of lipoprotein lipase to the assay was minimal. Particulate fractions (17 000 × g and 100 000 × g pellets) exhibited both an acid (pH 4.5 to 5) and a neutral (pH 7.5 to 8) pH optimum; the soluble (100 000 × g supernatant) fraction was characterized as having a single pH optimum of 7.5. The acid pH optimum for triglyceride lipase in the particulate fractions suggests that this activity originated in cardiac lysosomes, however the distribution of lipase activity determined at pH 5 did not correlate well with the lysosomal marker enzyme, N-acetyl-β-glucosaminidase. Chloroquine inhibited the acid triglyceride lipase (half maximal inhibition at 3 to 4 m m). In contrast, low concentrations (2 to 5 m m) of chloroquine stimulated lipase activity determined at pH 7.5; higher concentrations then inhibited enzyme activity. Similar results were obtained with NaCl; acid lipase activity was inhibited by all concentrations of NaCl whereas neutral lipase activity was first stimulated (25 to 150 m m NaCl) and then inhibited (200 to 500 m m NaCl). The addition of MgCl 2 (2 to 25 m m) resulted in an inhibition of lipase activity determined at pH 5 and a stimulation of activity determined at pH 7.5. Preincubation of the various rat heart fractions with ATP, cyclic AMP and protein kinase resulted in no change in lipase activity determined at either pH 5 or pH 7.5.