Catz S D, Steŕin-Speziale N B
Department of Biological Chemistry, Faculty of Pharmacy and Biochemistry, University of Buenos Aires, IQUIFIB, CONICET, Argentina.
J Leukoc Biol. 1996 Apr;59(4):591-7. doi: 10.1002/jlb.59.4.591.
In response to formyl-Met-Leu-Phe (fMLP), human neutrophils (PMN) generate superoxide anion (O2-) by the enzyme complex NADPH oxidase. The modulation of phosphoinositide (PPI) turnover and the activation of phospholipases C (PLC) and D (PLD) have been shown to be early steps in the oxidative response of fMLP-stimulated PMN. Although the physiological nonapeptide bradykinin (BK) is involved in inflammation, its participation in PMN activation has not been properly studied. In this work, activation of signal transduction pathways that mediate the oxidative response, and the modulation of the NADPH oxidase activity by BK, are analyzed. A direct comparison between the signal transduction pathway induced by BK and fMLP is also made. BK was not able to elicit O2- production by PMN. Nevertheless, several signal transduction pathways associated with PMN activation were triggered by BK. The nonapeptide induced the phosphorylation of prelabeled membrane PPI. This phenomenon was imitated by PMA and inhibited by H7 and staurosporine, thus suggesting the participation of protein kinase c (PKC). A loss of labeled [32P]PPI was triggered by fMLP. The fact that both PMA and fMLP stimulated O2- production but modulated PPI turnover in different ways, indicates that PPI labeling does not correlate with the oxidative response. Because PKC activation seemed to be a prerequisite for BK-induced modulation of PPI turnover, PLC activation could act as an intermediate step in this mechanism. Our results show that BK activated a PIP2-PLC measured as the release of [3H]IP3. On the contrary, a PC-PLD was highly stimulated by fMLP but not by BK. The fact that BK induced PLC activity but neither that of PLD nor NADPH oxidase, whereas fMLP triggered the activation of both phospholipases and evoked the PMN respiratory burst, suggests that diacylglycerol (DAG) from PIP2 as well as PA or PA-derived DAG, synergize to trigger the PMN oxidative response. Finally, BK inhibited O2- production by fMLP-activated PMN in a time-dependent manner. Since BK did not induce NO production by PMN, the inhibitory effect on the oxidative function was not due to ONOO- formation. These data show that BK plays an important role in inflammation by modulating the PMN function.
为响应甲酰甲硫氨酸-亮氨酸-苯丙氨酸（fMLP），人类中性粒细胞（PMN）通过酶复合物NADPH氧化酶产生超氧阴离子（O2-）。磷酸肌醇（PPI）周转的调节以及磷脂酶C（PLC）和D（PLD）的激活已被证明是fMLP刺激的PMN氧化反应的早期步骤。尽管生理性九肽缓激肽（BK）参与炎症反应，但其在PMN激活中的作用尚未得到充分研究。在这项工作中，分析了介导氧化反应的信号转导途径的激活以及BK对NADPH氧化酶活性的调节。还对BK和fMLP诱导的信号转导途径进行了直接比较。BK不能诱导PMN产生O2-。然而，BK触发了与PMN激活相关的几种信号转导途径。该九肽诱导了预先标记的膜PPI的磷酸化。这种现象可被佛波酯（PMA）模拟，并被H7和星形孢菌素抑制，因此提示蛋白激酶c（PKC）的参与。fMLP触发了标记的[32P]PPI的减少。PMA和fMLP均刺激O2-产生，但以不同方式调节PPI周转，这一事实表明PPI标记与氧化反应无关。由于PKC激活似乎是BK诱导的PPI周转调节的先决条件，PLC激活可能是该机制的中间步骤。我们的结果表明，BK激活了一种以[3H]IP3释放来衡量的磷脂酰肌醇-4,5-二磷酸-磷脂酶C（PIP2-PLC）。相反，fMLP强烈刺激磷脂酰胆碱-磷脂酶D（PC-PLD），而BK则无此作用。BK诱导PLC活性，但不诱导PLD或NADPH氧化酶活性，而fMLP触发两种磷脂酶的激活并引发PMN呼吸爆发，这一事实表明来自PIP2的二酰基甘油（DAG）以及磷脂酸（PA）或PA衍生的DAG协同触发PMN氧化反应。最后，BK以时间依赖性方式抑制fMLP激活的PMN产生O2-。由于BK不诱导PMN产生一氧化氮（NO），因此对氧化功能的抑制作用不是由于过氧亚硝酸根（ONOO-）的形成。这些数据表明，BK通过调节PMN功能在炎症中起重要作用。
