The regulation of platelet-activating factor production in endothelial cells. The role of calcium and protein kinase C.
Endothelial cells (EC) synthesize platelet-activating factor (PAF) when stimulated with agonists that bind to cell-surface receptors. We examined events that link receptor binding to synthesis of PAF by EC. Bovine EC stimulated with agonists that interact with specific cell-surface receptors accumulated PAF only in the presence of extracellular calcium. Hormonal stimulation of EC resulted in Ca2+ entry characteristic of that seen with receptor-operated calcium channels; Indo-1 measurements demonstrated that this inward flux of Ca2+ caused prolonged elevated levels of intracellular Ca2+. EC were exposed to melittin or theta toxin from Clostridium perfringens (pore-forming peptides that increase the permeability of the plasma membrane for small molecules) resulting in an inward flux of Ca2+ and accumulation of PAF. Ca2+ appears to be regulatory for PAF production at the level of phospholipase A2-mediated production of the PAF precursor 1-O-alkyl-2-lyso-sn-glycero-3-phosphocholine, as Ca2+ was required for the stimulated hydrolysis of 1-O-alkyl-2-acyl-sn-glycero-3-phosphocholine. PAF accumulation in EC is also regulated by protein kinase C. Pretreatment of EC with phorbol esters that activate protein kinase C or with dioctanoylglycerol, followed by stimulation, resulted in a 2-fold increase in stimulated PAF production. The regulatory effect of protein kinase C also appears to be at a phospholipase A2-mediated hydrolysis of 1-O-alkyl-2-acyl-sn-glycero-3-phosphocholine.
Whatley
R E
RE
Department of Medicine, University of Utah School of Medicine, Salt Lake City 84112.
Nelson
P
P
Zimmerman
G A
GA
Stevens
D L
DL
Parker
C J
CJ
McIntyre
T M
TM
Prescott
S M
SM
D000227:Adenine Nucleotides / Q000494:pharmacology; D000818:Animals; D001427:Bacterial Toxins / Q000494:pharmacology; D001920:Bradykinin / Q000494:pharmacology; D002118:Calcium / Q000502:physiology*; D002413:Cations, Divalent / Q000494:pharmacology; D002417:Cattle; D003593:Cytoplasm / Q000502:physiology; D004727:Endothelium / Q000502:physiology*; D006460:Hemolysin Proteins; D066298:In Vitro Techniques; D010741:Phospholipases A / Q000378:metabolism; D054467:Phospholipases A2; D010972:Platelet Activating Factor / Q000502:physiology*; D011493:Protein Kinase C / Q000502:physiology*; D013110:Sphingosine / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D000227:Adenine Nucleotides; D001427:Bacterial Toxins; D002413:Cations, Divalent; D006460:Hemolysin Proteins; D010972:Platelet Activating Factor; C020452:Clostridium perfringens theta-toxin; D011493:Protein Kinase C; D010741:Phospholipases A; D054467:Phospholipases A2; D013110:Sphingosine; D013755:Tetradecanoylphorbol Acetate; D001920:Bradykinin; D002118:Calcium