Characterization of the platelet-activating factor acetylhydrolase from human plasma by heterologous expression in Xenopus laevis oocytes.
Characterization of the platelet-activating factor acetylhydrolase from human plasma by heterologous expression in Xenopus laevis oocytes.
复制标题
通过在非洲爪蟾卵母细胞中异源表达来表征人血浆中的血小板激活因子乙酰水解酶。
DOI：
10.1073/pnas.91.22.10320
复制
发表时间：
1994
影响因子：
11.1
通讯作者：
Prescott,SM
中科院分区：
文献类型：
--
作者：
Yamada,Y;Stafforini,DM;Imaizumi,T;Zimmerman,GA;McIntyre,TM;Prescott,SM
血小板活化因子（PAF）是炎症和动脉粥样硬化的介质。PAF乙酰水解酶是血浆中的一种特异性降解酶，在PAF引起的多种病理生理过程中起重要作用。人巨噬细胞和Hep G2细胞分泌PAF乙酰水解酶，其特征与血浆活性相同。其他研究者报道载脂蛋白B可能具有磷脂酶A2活性，提示载脂蛋白B可能是PAF乙酰水解酶的酶原。然而，虽然巨噬细胞表达PAF乙酰水解酶活性，但我们在这些细胞的cDNA文库中未检测到载脂蛋白B的cDNA，表明巨噬细胞不表达该蛋白。与此相反，Hep G2细胞具有高水平的载脂蛋白B的cDNA，正如预期的那样。接下来，我们用从培养的人巨噬细胞和Hep G2细胞中提取的poly（A）+ RNA注射非洲爪蟾卵母细胞。25%~ 50%的爪蟾卵母细胞注射来自巨噬细胞或Hep G2细胞的poly（A）+ RNA后，分泌PAF乙酰水解酶活性（1.0-7.8 nmol/ml/h），该活性也利用合成的氧化磷脂作为底物。poly（A）+RNA注射的卵母细胞分泌的活性与脂蛋白相关，并以pH依赖性方式在颗粒之间转移，非常像血浆活性。这些实验证实，从poly（A）+RNA注射的卵母细胞释放的酶的性质与PAF乙酰水解酶的血浆形式的性质相同，并且检测到的活性不是载脂蛋白B中结构域的表达。
Platelet-activating factor (PAF) has been implicated as a mediator of inflammation and atherosclerosis. A specific degradative enzyme found in plasma, PAF acetylhydrolase, plays important roles in various pathophysiological events induced by PAF. Human macrophages and Hep G2 cells secrete PAF acetylhydrolase with characteristics identical to the plasma activity. Other investigators reported that apolipoprotein B may possess phospholipase A2 activity, which suggested that apolipoprotein B might be a zymogen for PAF acetylhydrolase. However, while macrophages express PAF acetylhydrolase activity, we did not detect cDNAs for apolipoprotein B in a cDNA library from these cells, indicating that macrophages do not express this protein. In contrast, Hep G2 cells had high levels of cDNA for apolipoprotein B, as expected. We next injected Xenopus laevis oocytes with poly(A)+ RNA extracted from cultured human macrophages and Hep G2 cells. Twenty-five to 50% of Xenopus oocytes injected with poly(A)+ RNA from macrophages or Hep G2 cells secreted a PAF acetylhydrolase activity (1.0-7.8 nmol/ml per h) that also utilized a synthetic oxidized phospholipid as substrate. The activity secreted by poly(A)+ RNA-injected oocytes associated with lipoproteins and transferred between the particles in a pH-dependent manner, much like the plasma activity. These experiments establish that the properties of the enzyme released from poly(A)+ RNA-injected oocytes are identical to those of the plasma form of PAF acetylhydrolase and that the activity detected is not the expression of a domain in apolipoprotein B.