Department of Pediatrics and Communicable Diseases, University of Michigan, Ann Arbor, Michigan, USA.
J Virol. 2014 Apr;88(7):3705-18. doi: 10.1128/JVI.03039-13. Epub 2014 Jan 15.
Barrier dysfunction of airway epithelium may increase the risk for acquiring secondary infections or allergen sensitization. Both rhinovirus (RV) and polyinosinic-polycytidilic acid [poly(I·C)], a double-stranded RNA (dsRNA) mimetic, cause airway epithelial barrier dysfunction, which is reactive oxygen species (ROS) dependent, implying that dsRNA generated during RV replication is sufficient for disrupting barrier function. We also demonstrated that RV or poly(I·C)-stimulated NADPH oxidase 1 (NOX-1) partially accounts for RV-induced ROS generation. In this study, we identified a dsRNA receptor(s) contributing to RV-induced maximal ROS generation and thus barrier disruption. We demonstrate that genetic silencing of the newly discovered dsRNA receptor Nod-like receptor X-1 (NLRX-1), but not other previously described dsRNA receptors, abrogated RV-induced ROS generation and reduction of transepithelial resistance (R(T)) in polarized airway epithelial cells. In addition, both RV and poly(I·C) stimulated mitochondrial ROS, the generation of which was dependent on NLRX-1. Treatment with Mito-Tempo, an antioxidant targeted to mitochondria, abolished RV-induced mitochondrial ROS generation, reduction in R(T), and bacterial transmigration. Furthermore, RV infection increased NLRX-1 localization to the mitochondria. Additionally, NLRX-1 interacts with RV RNA and poly(I·C) in polarized airway epithelial cells. Finally, we show that NLRX-1 is also required for RV-stimulated NOX-1 expression. These findings suggest a novel mechanism by which RV stimulates generation of ROS, which is required for disruption of airway epithelial barrier function.
Rhinovirus (RV), a virus responsible for a majority of common colds, disrupts the barrier function of the airway epithelium by increasing reactive oxygen species (ROS). Poly(I·C), a double-stranded RNA (dsRNA) mimetic, also causes ROS-dependent barrier disruption, implying that the dsRNA intermediate generated during RV replication is sufficient for this process. Here, we demonstrate that both RV RNA and poly(I·C) interact with NLRX-1 (a newly discovered dsRNA receptor) and stimulate mitochondrial ROS. We show for the first time that NLRX-1 is primarily expressed in the cytoplasm and at the apical surface rather than in the mitochondria and that NLRX-1 translocates to mitochondria following RV infection. Together, our results suggest a novel mechanism for RV-induced barrier disruption involving NLRX-1 and mitochondrial ROS. Although ROS is necessary for optimal viral clearance, if not neutralized efficiently, it may increase susceptibility to secondary infections and alter innate immune responses to subsequently inhaled pathogens, allergens, and other environmental factors.
气道上皮屏障功能障碍可能会增加继发感染或变应原致敏的风险。鼻病毒（RV）和聚肌苷酸-聚胞苷酸[poly（I·C）]，一种双链 RNA（dsRNA）类似物，均可导致气道上皮屏障功能障碍，这与活性氧物种（ROS）有关，这意味着 RV 复制过程中产生的 dsRNA 足以破坏屏障功能。我们还表明，RV 或 poly（I·C）刺激的 NADPH 氧化酶 1（NOX-1）部分解释了 RV 诱导的 ROS 产生。在这项研究中，我们确定了一种 dsRNA 受体（s），该受体有助于 RV 诱导的最大 ROS 产生，从而破坏屏障。我们证明，新发现的 dsRNA 受体 Nod-like receptor X-1（NLRX-1）的基因沉默，而不是其他先前描述的 dsRNA 受体，消除了 RV 诱导的 ROS 产生和极化气道上皮细胞中跨上皮电阻（R（T））的降低。此外，RV 和 poly（I·C）均刺激线粒体 ROS 的产生，该产生依赖于 NLRX-1。用抗氧化剂 Mito-Tempo 处理，该抗氧化剂靶向线粒体，可消除 RV 诱导的线粒体 ROS 产生，降低 R（T）和细菌迁移。此外，RV 感染增加了 NLRX-1 向线粒体的定位。另外，NLRX-1 在极化的气道上皮细胞中与 RV RNA 和 poly（I·C）相互作用。最后，我们表明 NLRX-1 也需要 RV 刺激的 NOX-1 表达。这些发现表明 RV 通过刺激 ROS 的产生来破坏气道上皮屏障功能的新机制，这对于破坏气道上皮屏障功能是必需的。
鼻病毒（RV）是引起大多数普通感冒的病毒，通过增加活性氧物种（ROS）来破坏呼吸道上皮的屏障功能。多聚肌苷酸-聚胞苷酸（poly（I·C）），一种双链 RNA（dsRNA）类似物，也会引起 ROS 依赖性的屏障破坏，这意味着 RV 复制过程中产生的 dsRNA 中间体足以完成此过程。在这里，我们证明 RV RNA 和 poly（I·C）均与 NLRX-1（一种新发现的 dsRNA 受体）相互作用，并刺激线粒体 ROS。我们首次表明，NLRX-1 主要在细胞质和顶表面表达，而不在线粒体中，并且在 RV 感染后 NLRX-1 向线粒体易位。总的来说，我们的研究结果表明，NLRX-1 和线粒体 ROS 参与了 RV 诱导的屏障破坏的新机制。尽管 ROS 对于最佳病毒清除是必需的，但是如果不能有效地中和，则可能会增加继发感染的风险，并改变对随后吸入的病原体，过敏原和其他环境因素的先天免疫反应。
