Laboratory of Immunophysiology, Biophysics Institute Carlos Chagas Filho, Federal University of Rio de Janeiro, Rio de Janeiro 21941-902, Brazil;
J Immunol. 2014 May 15;192(10):4765-73. doi: 10.4049/jimmunol.1301058. Epub 2014 Apr 11.
ATP is an important signaling molecule in the immune system, and it is able to bind the P2X7 purinergic receptor. Recently, our group showed that ATP-treated macrophages eliminate Leishmania amazonensis. It has been reported that leukotriene B4 (LTB4) reduces the parasitic load of infected macrophages. Additionally, it has been demonstrated that the P2X7 receptor can induce PLA2 activation and arachidonic acid mobilization. Based on these findings, we investigated whether LTB4 is produced upon P2X7 receptor activation and examined whether LTB4 modulates parasite elimination. Using macrophages lacking the P2X7 receptor, we observed that ATP was not able to reduce L. amazonensis load. This result suggests a role of the P2X7 purinergic receptor in parasite elimination. In addition, ATP was sufficient to induce LTB4 release from infected control macrophages but not from macrophages lacking the P2X7 receptor. Moreover, we found that ATP failed to decrease the parasitic load in 5-lipoxygenase (LO)-deficient macrophages. Treatment with the 5-LO inhibitor AA861 also impairs the ATP effect on parasitic loads. Furthermore, macrophages from 5-LO knockout mice eliminated L. amazonensis in the presence of exogenous LTB4, and macrophages obtained from P2X7 receptor knockout mice eliminated L. amazonensis when incubated with ionomycin. Finally, we demonstrated that in the presence of CP105696, an antagonist for LTB4 high-affinity receptor, ATP was not able to reduce parasitic load. These results indicate that P2X7 receptor activation leads to LTB4 formation, which is required for L. amazonensis elimination.
三磷酸腺苷（ATP）是免疫系统中的一种重要信号分子，能够与 P2X7 嘌呤能受体结合。最近，我们的研究小组发现，用 ATP 处理的巨噬细胞能够消灭美洲利什曼原虫（Leishmania amazonensis）。据报道，白三烯 B4（LTB4）可减少感染巨噬细胞中的寄生虫载量。此外，已经证明 P2X7 受体可诱导 PLA2 激活和花生四烯酸动员。基于这些发现，我们研究了 P2X7 受体激活后是否会产生 LTB4，并研究了 LTB4 是否调节寄生虫的消除。使用缺乏 P2X7 受体的巨噬细胞，我们观察到 ATP 不能降低 L. amazonensis 的负荷。这一结果表明 P2X7 嘌呤能受体在寄生虫消除中起作用。此外，ATP 足以从感染对照巨噬细胞中诱导 LTB4 释放，但不能从缺乏 P2X7 受体的巨噬细胞中诱导。此外，我们发现，ATP 未能降低 5-脂氧合酶（5-LO）缺陷型巨噬细胞中的寄生虫载量。用 5-LO 抑制剂 AA861 处理也会损害 ATP 对寄生虫负荷的影响。此外，在存在外源性 LTB4 的情况下，5-LO 基因敲除小鼠的巨噬细胞能够消灭 L. amazonensis，而在与离子霉素孵育时，P2X7 受体基因敲除小鼠的巨噬细胞也能消灭 L. amazonensis。最后，我们证明在 CP105696（LTB4 高亲和力受体的拮抗剂）存在的情况下，ATP 不能降低寄生虫负荷。这些结果表明，P2X7 受体的激活导致 LTB4 的形成，这对于消灭 L. amazonensis 是必需的。
