Ca/sup 2 +/-dependent and Ca/sup 2 +/-independent pathways for release of arachidonic acid from phosphatidylinositol in endothelial cells
The pathways for degradation of phosphatidylinositol (PI) were investigated in sonicated suspensions prepared from confluent cultures of bovine pulmonary artery endothelial cells. The time courses of formation of /sup 3/H-labeled and /sup 14/C-labeled metabolites of phosphatidyl-(/sup 3/H)inositol ((/sup 3/H)Ins-PI) and 1-stearoyl-2-(/sup 14/C) arachidonoyl-PI were determined at 37/sup 0/C and pH 7.5 in the presence of 2 mM EDTA with or without a 2 mM excess of Ca/sup 2 +/. The rates of formation of lysophosphatidyl-(/sup 3/H)inositol ((/sup 3/H)Ins-lyso-PI) and 1-lyso-2-(/sup 14/C) arachidonoyl-PI were similar in the presence and absence of Ca/sup 2 +/, and the absolute amounts of the two radiolabeled lyso-PI products formed were nearly identical. This indicated that lyso-PI was formed by phospholipase A1, and phospholipase A2 was not measurable. In the presence of EDTA, (/sup 14/C)arachidonic acid release from 1-stearoyl-2-(/sup 14/C)arachidonoyl-PI paralleled release of glycerophospho-(/sup 3/H)inositol ((/sup 3/H)GPI) from (/sup 3/H)Ins-PI. Formation of (/sup 3/H)GPI was inhibited by treatment with the specific sulfhydryl reagent, 2,2'-dithiodipyridine, and this was accompanied by an increase in (/sup 3/H)Ins-lyso-PI. In the presence of Ca/sup 2 +/, (/sup 14/C) arachidonic acid release from 1-stearoyl-2-(/sup 14/C)arachidonoyl-PI was increased 2-fold and was associated with Ca/sup 2 +/-dependent phospholipase C activity. Under these conditions, (/sup 3/H)inositol monophosphate production exceeded formation of (/sup 14/C)arachidonic acid-labeled phospholipase C products, diacylglycerol plus monoacylglycerol, by an amount that was equal to the amount of (/sup 14/C)arachidonic acid formed in excess of (/sup 3/H)GPI. Low concentrations of phenylmethanesulfonyl fluoride (15-125 microM) inhibited Ca/sup 2 +/-dependent (/sup 14/C)arachidonic acid release, and the decrease in (/sup 14/C) arachidonic acid formed was matched by an equivalent increase in /sup 14/C label in diacylglycerol plus monoacyclglycerol.
- Research Organization:
- St. Louis Univ. School of Medicine, MO
- OSTI ID:
- 5856940
- Journal Information:
- J. Biol. Chem.; (United States), Vol. 262:27
- Country of Publication:
- United States
- Language:
- English
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ARACHIDONIC ACID
SECRETION
LIPASE
ENZYME ACTIVITY
PHOSPHOLIPIDS
METABOLISM
ARTERIES
BIOLOGICAL PATHWAYS
CALCIUM COMPOUNDS
CARBON 14 COMPOUNDS
CATIONS
CATTLE
CELL CULTURES
ENDOTHELIUM
INHIBITION
TRACER TECHNIQUES
TRITIUM COMPOUNDS
ALKALINE EARTH METAL COMPOUNDS
ANIMAL TISSUES
ANIMALS
BLOOD VESSELS
BODY
CARBOXYLESTERASES
CARBOXYLIC ACIDS
CARDIOVASCULAR SYSTEM
CHARGED PARTICLES
DOMESTIC ANIMALS
ENZYMES
ESTERASES
ESTERS
HYDROLASES
IONS
ISOTOPE APPLICATIONS
LABELLED COMPOUNDS
LIPIDS
MAMMALS
MONOCARBOXYLIC ACIDS
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC PHOSPHORUS COMPOUNDS
ORGANS
RUMINANTS
TISSUES
VERTEBRATES
550201* - Biochemistry- Tracer Techniques