Involvement of cytosolic phospholipase A2, and the subsequent release of arachidonic acid, in signalling by rac for the generation of intracellular reactive oxygen species in rat-2 fibroblasts.
Involvement of cytosolic phospholipase A2, and the subsequent release of arachidonic acid, in signalling by rac for the generation of intracellular reactive oxygen species in rat-2 fibroblasts.
复制标题
胞浆磷脂酶 A2 的参与以及随后花生四烯酸的释放，参与 rac 信号传导，以在rat-2成纤维细胞中产生细胞内活性氧。
作者：
C. Woo;Z. Lee;B. C. Kim;K. Ha;J. Kim
尽管最近有许多关于Rac在活性氧（ROS）产生中的作用的研究，但信号通路的细节仍不清楚。在本研究中，我们分析了胞浆磷脂酶A（2）的激活和花生四烯酸（AA）的释放在多大程度上参与了Rac介导的ROS产生。RacV 12是Rac 1的组成型活性形式，用RacV 12转染Rat-2细胞可诱导ROS水平升高，这可通过增加2 '，7'-二氯荧光素的H（2）O（2）-敏感荧光来反映。抑制磷脂酶A（2）或5-脂氧合酶可阻断上述作用，但抑制环氧化酶则不能阻断。外源AA的应用增加了ROS水平，但这种作用依赖于AA通过5-脂氧合酶进一步代谢为白三烯C（4）/D（4）/E（4）。事实上，外源性应用白三烯C（4）/D（4）/E（4）的混合物引起ROS水平的短暂升高，而ROS水平的升高被过氧化氢酶阻断。这些发现表明，磷脂酶A（2）和随后的AA代谢5-脂氧合酶作为下游介质的Rac信号通路，导致ROS的产生。
Although there have been a number of recent studies on the role of Rac in the generation of reactive oxygen species (ROS), details of the signalling pathway remain unclear. In the present study we analysed the extent to which the activation of cytosolic phospholipase A(2) and the resultant release of arachidonic acid (AA) are involved in the Rac-mediated generation of ROS. Transfection of Rat-2 cells with RacV12, a constitutively active form of Rac1, induced elevated levels of ROS, as reflected by increased H(2)O(2)-sensitive fluorescence of 2', 7'-dichlorofluorescein. These effects could be blocked by inhibiting phospholipase A(2) or 5-lipoxygenase but not by inhibiting cyclo-oxygenase. The application of exogenous AA increased levels of ROS but the effect was dependent on the further metabolism of AA to leukotrienes C(4)/D(4)/E(4) by 5-lipoxygenase. Indeed, the exogenous application of a mixture of leukotrienes C(4)/D(4)/E(4) elicited transient elevations in the levels of ROS that were blocked by catalase. These findings indicate that phospholipase A(2) and subsequent AA metabolism by 5-lipoxygenase act as downstream mediators in a Rac signalling pathway leading to the generation of ROS.