Balgoma David, Montero Olimpio, Balboa María A, Balsinde Jesús
Instituto de Biología y Genética Molecular, Consejo Superior de Investigaciones Científicas and Centro de Investigación Biomédica en Red de Diabetes y Enfermedades Metabólicas Asociadas, Valladolid, Spain.
FEBS J. 2008 Dec;275(24):6180-91. doi: 10.1111/j.1742-4658.2008.06742.x. Epub 2008 Nov 5.
Phagocytic cells exposed to exogenous arachidonic acid (AA) incorporate large quantities of this fatty acid into choline and ethanolamine glycerophospholipids, and into phosphatidylinositol (PtdIns). Utilizing liquid chromatography coupled to MS, we have characterized the incorporation of exogenous deuterated AA ([(2)H]AA) into specific PtdIns molecular species in human monocyte cells. A PtdIns species containing two exogenous [(2)H]AA molecules (1-[(2)H]AA-2-[(2)H]AA-glycero-3-phosphoinositol) was readily detected when human U937 monocyte-like cells and peripheral blood monocytes were exposed to [(2)H]AA concentrations as low as 160 nm to 1 mum. Bromoenol lactone, an inhibitor of Ca(2+)-independent phospholipase A(2) (iPLA(2)), diminished lyso-PtdIns levels, and almost completely inhibited the appearance of 1-[(2)H]AA-2-[(2)H]AA-glycero-3-phosphoinositol, suggesting the involvement of deacylation reactions in the synthesis of this phospholipid. De novo synthesis did not appear to be involved, as no other diarachidonoyl phospholipid or neutral lipid was detected under these conditions. Measurement of the metabolic fate of 1-[(2)H]AA-2-[(2)H]AA-glycero-3-phosphoinositol after pulse-labeling of the cells with [(2)H]AA showed a time-dependent, exponential decrease in the level of this phospholipid. These results identify 1-[(2)H]AA-2-[(2)H]AA-glycero-3-phosphoinositol as a novel, short-lived species for the initial incorporation of AA into the PtdIns class of cellular phospholipids in human monocytes.
暴露于外源性花生四烯酸（AA）的吞噬细胞会将大量这种脂肪酸掺入胆碱和乙醇胺甘油磷脂以及磷脂酰肌醇（PtdIns）中。利用液相色谱与质谱联用技术，我们已对人单核细胞中外源性氘代AA（[(2)H]AA）掺入特定PtdIns分子种类的情况进行了表征。当人U937单核细胞样细胞和外周血单核细胞暴露于低至160 nM至1 μM的[(2)H]AA浓度时，很容易检测到一种含有两个外源性[(2)H]AA分子的PtdIns种类（1-[(2)H]AA-2-[(2)H]AA-甘油-3-磷酸肌醇）。溴代烯醇内酯是一种不依赖Ca(2+)的磷脂酶A(2)（iPLA(2)）抑制剂，它降低了溶血磷脂酰肌醇水平，并几乎完全抑制了1-[(2)H]AA-2-[(2)H]AA-甘油-3-磷酸肌醇的出现，这表明脱酰基反应参与了这种磷脂的合成。在这些条件下未检测到其他二花生四烯酰磷脂或中性脂质，因此似乎不涉及从头合成。在用[(2)H]AA对细胞进行脉冲标记后，对1-[(2)H]AA-2-[(2)H]AA-甘油-3-磷酸肌醇的代谢命运进行测量，结果显示该磷脂水平呈时间依赖性指数下降。这些结果确定1-[(2)H]AA-2-[(2)H]AA-甘油-3-磷酸肌醇是AA最初掺入人单核细胞中细胞磷脂PtdIns类别的一种新型、短命的种类。
