BACKGROUND Bacterial meningitis and meningoencephalitis are associated with devastating neuroinflammation. We and others have demonstrated the importance of glial cells in the initiation of immune responses to pathogens invading the central nervous system (CNS). These cells use a variety of pattern recognition receptors (PRRs) to identify common pathogen motifs and the cytosolic sensor retinoic acid inducible gene-1 (RIG-I) is known to serve as a viral PRR and initiator of interferon (IFN) responses. Intriguingly, recent evidence indicates that RIG-I also has an important role in the detection of bacterial nucleic acids, but such a role has not been investigated in glia. METHODS In this study, we have assessed whether primary or immortalized human and murine glia express RIG-I either constitutively or following stimulation with bacteria or their products by immunoblot analysis. We have used capture ELISAs and immunoblot analysis to assess human microglial interferon regulatory factor 3 (IRF3) activation and IFN production elicited by bacterial nucleic acids and novel engineered nucleic acid nanoparticles. Furthermore, we have utilized a pharmacological inhibitor of RIG-I signaling and siRNA-mediated knockdown approaches to assess the relative importance of RIG-I in such responses. RESULTS We demonstrate that RIG-I is constitutively expressed by human and murine microglia and astrocytes, and is elevated following bacterial infection in a pathogen and cell type-specific manner. Additionally, surface and cytosolic PRR ligands are also sufficient to enhance RIG-I expression. Importantly, our data demonstrate that bacterial RNA and DNA both trigger RIG-I-dependent IRF3 phosphorylation and subsequent type I IFN production in human microglia. This ability has been confirmed using our nucleic acid nanoparticles where we demonstrate that both RNA- and DNA-based nanoparticles can stimulate RIG-I-dependent IFN responses in these cells. CONCLUSIONS The constitutive and bacteria-induced expression of RIG-I by human glia and its ability to mediate IFN responses to bacterial RNA and DNA and nucleic acid nanoparticles raises the intriguing possibility that RIG-I may be a potential target for therapeutic intervention during bacterial infections of the CNS, and that the use of engineered nucleic acid nanoparticles that engage this sensor might be a method to achieve this goal.

中文翻译：

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维甲酸诱导的基因-I介导的人类小胶质细胞中细菌核酸的检测。

背景技术细菌性脑膜炎和脑膜脑炎与破坏性神经炎症相关。我们和其他人已经证明了神经胶质细胞在针对入侵中枢神经系统（CNS）的病原体的免疫应答中的重要性。这些细胞使用多种模式识别受体（PRR）来识别常见的病原体基序，并且已知胞质传感器视黄酸诱导基因1（RIG-1）可作为病毒PRR和干扰素（IFN）反应的引发剂。有趣的是，最近的证据表明，RIG-I在细菌核酸的检测中也具有重要作用，但尚未在神经胶质细胞中研究这种作用。方法在这项研究中 我们通过免疫印迹分析评估了原发性或永生化的人和鼠神经胶质细胞组成性表达或在细菌或它们的产物刺激下表达RIG-I。我们已经使用捕获ELISA和免疫印迹分析来评估人小胶质干扰素调节因子3（IRF3）激活和细菌核酸和新型工程核酸纳米粒子引发的IFN产生。此外，我们已经利用RIG-I信号的药理抑制剂和siRNA介导的敲低方法来评估RIG-I在此类反应中的相对重要性。结果我们证明RIG-I由人和鼠小胶质细胞和星形胶质细胞组成性表达，并在细菌感染后以病原体和细胞类型特异性方式升高。另外，表面和胞质PRR配体也足以增强RIG-1表达。重要的是，我们的数据表明细菌RNA和DNA均可触发人小胶质细胞中RIG-I依赖的IRF3磷酸化和随后的I型干扰素产生。使用我们的核酸纳米颗粒已经证实了这种能力，我们证明了基于RNA和DNA的纳米颗粒都可以刺激这些细胞中RIG-1依赖性的IFN反应。结论人胶质细胞本构和细菌诱导的RIG-I表达及其介导IFN对细菌RNA和DNA以及核酸纳米颗粒的应答能力增加了令人感兴趣的可能性，即RIG-I可能是细菌感染期间治疗干预的潜在靶标中枢神经系统