Synthesis of platelet activating factor and metabolism of related lipids in embryonic cells

Abstract

Primary cultures of mouse embryo palate mesenchyme (MEPM) cells incubated with 1-O-[3H]alkyl-2-lyso-sn-glycero-3-phosphocholine ([3H])lyso-PAF) incorporated radiolabel into 1-radyl-2-acyl-sn-glycero-3-phosphocholine (PC) and -phosphoethanolamine (PE). The radiolabeled PC was insensitive to hydrolysis with HCl fumes, whereas at least 82% of the 3H found in the PE was hydrolyzed to 3H-aldehydes by such treatment. Treatment of the PC with Vitride produced [3H]alkylglycerol; similar treatment of the PE produced [3H]alk-1-enylglycerol. None of the radiolabeled products yielded fatty alcohol upon reduction with Vitride. These findings indicate the radiolabeled PC was 1-O-alkyl-linked whereas the PE contained predominantly 1-O-alk-1'-enyl species with smaller amounts of 1-O-alkyl species. Homogenates of MEPM cells which had been prelabeled with [3H]lyso-PAF and [14C]arachidonic acid produced 14C-fatty acid, [3H]lyso-PC, and [3H]alkylglycerol when incubated at selected values of pH and concentrations of calcium. There was no accumulation of [3H]lyso-PE in the various incubation mixtures. Stimulation of MEPM cells with the ionophore A23187 in the presence of calcium and [3H]acetate resulted in the production of 3H-platelet-activating factor (PAF), identified by its migration with authentic PAF and its conversion to 1-O-[3H]alkyl-2,3-diacetylglycerol upon treatment with phospholipase C and acetic anhydride. These studies demonstrate that: (i) MEPM cells are able to incorporate [3H]lyso-PAF into 1-O-alkyl-2-acyl-sn-glycero-3-phosphocholine, the storage form of PAF, and into 1-O-alk-1'-enyl-2-acyl-sn-glycero-3-phosphoethanolamine (PE plasmalogen); (ii) endogenous 1-O-[3H]alkyl-2-acyl-sn-glycero-3-phosphocholine can serve as a substrate for phospholipase A2 in homogenates; and (iii) MEPM cells have the ability to synthesize PAF, thus raising the possibility that this compound may play a role in modulating the physiology of these embryonic cells.