TLR3-dependent induction of nitric oxide synthase in RAW 264.7 macrophage-like cells via a cytosolic phospholipase A2/cyclooxygenase-2 pathway
TLR3-dependent induction of nitric oxide synthase in RAW 264.7 macrophage-like cells via a cytosolic phospholipase A2/cyclooxygenase-2 pathway
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DOI：
10.4049/jimmunol.179.7.4821
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发表时间：
2007-10-01
影响因子：
4.4
通讯作者：
Balboa, Maria A.
中科院分区：
文献类型：
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作者：
Pindado, Jose;Balsinde, Jesus;Balboa, Maria A.
dsRNA是病毒复制的副产物，当被吞噬细胞识别时能够诱导炎症反应。在这项研究中，我们确定了IVA组胞质磷脂酶A（2）（cPLA（2）α）作为抗病毒反应的效应物。用dsRNA类似物聚肌苷酸：聚胞苷酸（poly-IC）处理RAW 264.7小鼠巨噬细胞样细胞，促进游离花生四烯酸的释放，随后通过新合成的环氧合酶-2（考克斯-2）酶将其转化为PGE（2）。这些过程被选择性cPLA（2）α抑制剂吡咯苯酮阻断，指出cPLA（2）α是参与的效应子。与这一观察结果一致，在用poly-IC处理细胞后，cPLA（2）α磷酸化状态增加。通过小干扰RNA（siRNA）或吡咯苯酮抑制cPLA（2）α表达和活性导致诱导型NO合酶（iNOS）基因表达的抑制。此外，考克斯-2衍生的PGE（2）产生似乎参与iNOS表达，因为siRNA抑制考克斯-2也导致抑制iNOS，后者通过外源性加入PGE 2而恢复。最后，通过siRNA去除TLR 3抑制考克斯-2表达、PGE 2产生和多聚IC诱导的iNOS。总的来说，这些发现提示了一种巨噬细胞对dsRNA反应的活化模型，其中TLR 3的参与导致cPLA（2）α介导的花生四烯酸动员和考克斯-2介导的PGE（2）产生，它们共同诱导iNOS的表达。
dsRNA is a by-product of viral replication capable of inducing an inflammatory response when recognized by phagocyte cells. In this study, we identify group IVA cytosolic phospholipase A(2) (cPLA(2)alpha) as an effector of the antiviral response. Treatment of RAW 264.7 murine macrophage-like cells with the dsRNA analog polyinosinic:polycytidylic acid (poly-IC) promotes the release of free arachidonic acid that is subsequently converted into PGE(2) by the de novo-synthesized cyclooxygenase-2 (COX-2) enzyme. These processes are blocked by the selective cPLA(2)alpha inhibitor pyrrophenone, pointing out to cPLA(2)alpha as the effector involved. In keeping with this observation, the cPLA(2)alpha phosphorylation state increases after cellular treatment with poly-IC. Inhibition of cPLA(2)alpha expression and activity by either small interfering RNA (siRNA) or pyrrophenone leads to inhibition of the expression of the inducible NO synthase (iNOS) gene. Moreover, COX-2-derived PGE(2) production appears to participate in iNOS expression, because siRNA inhibition of COX-2 also leads to inhibition of iNOS, the latter of which is restored by exogenous addition of PGE2. Finally, cellular depletion of TLR3 by siRNA inhibits COX-2 expression, PGE2 generation, and iNOS induction by poly-IC. Collectively, these findings suggest a model for macrophage activation in response to dsRNA, whereby engagement of TLR3 leads to cPLA(2)alpha-mediated arachidonic acid mobilization and COX-2-mediated PGE(2) production, which cooperate to induce the expression of iNOS.