Chepenik K P, Wykle R L
Department of Anatomy, Jefferson Medical College, Thomas Jefferson University, Philadelphia, PA 19107.
Biochim Biophys Acta. 1992 Jun 22;1126(2):192-8. doi: 10.1016/0005-2760(92)90290-c.
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.
用1-O-[3H]烷基-2-溶血-sn-甘油-3-磷酸胆碱（[3H]溶血血小板活化因子）孵育的小鼠胚胎腭间充质（MEPM）细胞原代培养物，将放射性标记掺入1-烷基-2-酰基-sn-甘油-3-磷酸胆碱（PC）和磷酸乙醇胺（PE）中。放射性标记的PC对HCl烟雾水解不敏感，而在PE中发现的至少82%的3H经此处理后水解为3H-醛。用二异丁基氢化铝处理PC产生[3H]烷基甘油；对PE进行类似处理产生[3H]烯基甘油。用二异丁基氢化铝还原后，没有一种放射性标记产物产生脂肪醇。这些发现表明，放射性标记的PC是1-O-烷基连接的，而PE主要含有1-O-烯基-1'-烯基物种，1-O-烷基物种含量较少。用[3H]溶血血小板活化因子和[14C]花生四烯酸预标记的MEPM细胞匀浆在选定的pH值和钙浓度下孵育时，产生14C-脂肪酸、[3H]溶血PC和[3H]烷基甘油。在各种孵育混合物中没有[3H]溶血PE的积累。在钙和[3H]乙酸盐存在下，用离子载体A23187刺激MEPM细胞导致产生3H-血小板活化因子（PAF），通过其与 authentic PAF的迁移以及用磷脂酶C和乙酸酐处理后转化为1-O-[3H]烷基-2,3-二乙酰甘油来鉴定。这些研究表明：（i）MEPM细胞能够将[3H]溶血血小板活化因子掺入PAF的储存形式1-O-烷基-2-酰基-sn-甘油-3-磷酸胆碱以及1-O-烯基-1'-烯基-2-酰基-sn-甘油-3-磷酸乙醇胺（PE缩醛磷脂）中；（ii）内源性1-O-[3H]烷基-2-酰基-sn-甘油-3-磷酸胆碱可作为匀浆中磷脂酶A2的底物；（iii）MEPM细胞具有合成PAF的能力，从而增加了这种化合物可能在调节这些胚胎细胞生理学中发挥作用的可能性。
