Characterization of phospholipase A2 activity in rat aorta smooth muscle cells.
Phospholipase A activity was measured in homogenates and acid extracts of smooth muscle cells from rat aorta and mesenteric artery using [1-14 C]oleate-labeled autoclaved Escherichia coli and 1-[1-14C]stearyl-2-acyl-3-sn-glycerophosphorylethanolamine as substrates. The results demonstrate the presence of neutral-active phospholipase(s) A that exclusively catalyze the release of fatty acid from the 2-position of phospholipids. Optimal activity was at pH 7.5, and there was an absolute requirement for low concentrations of Ca2+. Mg2+ did not substitute for Ca2+, and EGTA inhibited the activity. Phospholipase A2 activity was predominantly membrane-associated and was solubilized by homogenization in 0.18 N H2SO4. Sulfuric acid extracts of rat aortic smooth muscle cells were four times more active than extracts of mesenteric artery (710 vs. 170 nmol/h per mg protein). By comparison, acid extracts of rat lung, heart, and liver were less active (60-75 nmol/h per mg). Indomethacin, sodium meclofenamate, mepacrine and chlorpromazine, but not dexamethasone or aspirin, inhibited acid-solubilized phospholipase(s) A2 between 10(-6) and 10(-3) M in a dose-dependent manner. Preincubation with p-bromophenacyl bromide or diethylpyrocarbonate inhibited phospholipase(s) A2, suggesting the presence of a histidine residue at the active site. An extract from the leaves of feverfew plant (Tanacetum parthenium) was also a potent inhibitor of aortic smooth muscle phospholipase(s) A2.
D000818:Animals; D001011:Aorta / Q000201:enzymology; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008099:Liver / Q000201:enzymology; D008168:Lung / Q000201:enzymology; D008638:Mesenteric Arteries / Q000201:enzymology; D009131:Muscle, Smooth, Vascular / Q000201:enzymology*; D009206:Myocardium / Q000201:enzymology; D009928:Organ Specificity; D010740:Phospholipases / Q000378:metabolism*; D010741:Phospholipases A / Q000378:metabolism*; D054467:Phospholipases A2; D010946:Plants, Medicinal; D051381:Rats; D012717:Sesquiterpenes; D020903:Tanacetum parthenium
D012717:Sesquiterpenes; D010740:Phospholipases; D010741:Phospholipases A; D054467:Phospholipases A2