Is platelet-activating factor (PAF-acether) synthesis by murine peritoneal cells (PC) a two-step process?
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PAF-acether (1-O-alkyl-2-acetyl-sn-glyceryl-3-phosphorylcholine) is released from several cell sources simultaneously with an inactive non-acylated compound (1-O-alkyl-glyceryl-3-phosphorylcholine) (lyso-PAF-acether). Formation of the latter probably results from the activation of a phospholipase A2 (PLA2). Indeed, the PLA2 inhibitors, bromophenacyl bromide (BPB), mepacrine, 874CB (100 μM), and EDTA (5 mM), blocked the zymosan-induced release of PAF-acether from PC. EDTA and BPB also markedly reduced the release of lyso-PAF-acether. PAF-acether formation could involve a metabolic step linking the acetyl moiety to the 2nd carbon of lyso-PAF-acether. To verify this hypothesis, acetyl coenzyme A (acetyl-CoA) was added to stimulated PC. This enhanced the release of PAF-acether in a dose-dependent fashion from 1 μM acetyl-CoA to reach a maximal increase −200%—at 100 μM. Furthermore, using3H acetyl-CoA, incorporation of labelled acetate into PAF-acether was suggested by (1) identical chromatographic patterns of biological activity and radioactivity; (2) disappearance of these activities after treatment with PLA2, but not after exposure to lipase fromRhizopus arrhizus. PAF-acether was also obtained when both acetyl-CoA (100 μM) and synthetic lyso-PAF-acether (0.2 μM) were added to unstimulated PC previously treated with BPB (100 μM for 10 min). These results suggest that the release of PAF-acether is the consequence of at least two different steps: (1) hydrolysis of 1-O-alkyl-2-acyl-glyceryl-phosphorylcholine by PLA2; (2) enzymatic acetylation of the hydrolysis product.
KeywordsLipase Acetyl Hydrolysis Product Cell Source Mepacrine
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