A TLR3-STAT3-miR-155 axis and astrocyte-myeloid crosstalk in viral encephalitis

病毒性脑炎中的 TLR3-STAT3-miR-155 轴和星形胶质细胞-骨髓串扰

• 批准号: 10066377
• 负责人: SONIA NAVAS-MARTIN
• 金额: $ 34.01万
• 依托单位: DREXEL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-01-01 至 2022-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10066377
• 关键词: Ablation; Acute; Adaptor Signaling Protein; Affect; Agonist; Antiviral Response; Astrocytes; Autoimmunity; Automobile Driving; Binding Sites; Blood - brain barrier anatomy; Brain; CCL2 gene; Cells; Central Nervous System Infections; Chemotactic Factors; Clinical; Coronavirus; Coronavirus Infections; Double Stranded RNA Virus; Double-Stranded RNA; Encephalitis; Exhibits; Functional disorder; Genetic; Glial Fibrillary Acidic Protein; Growth Factor; Homeostasis; ITGAM gene; Immune; Immune response; Immunotherapy; In Vitro; Infection; Infiltration; Inflammation; Inflammatory; Interferon-beta; Interferons; Interleukin-1 Receptors; Lead; Leukocytes; MHV-JHM; Maintenance; Matrix Metalloproteinases; Mediating; MicroRNAs; Modeling; Molecular; Morbidity - disease rate; Murine hepatitis virus; Mus; Myelogenous; Myeloid Cells; Necrosis; Neuraxis; Neuroglia; Neuropathogenesis; Neutrophil Infiltration; Pathogenesis; Pattern recognition receptor; Pharmacology; Play; Poly I-C; Poly(A)+ RNA; Protein Tyrosine Kinase; Proteins; RNA Virus Infections; RNA Viruses; Regulation; Resistance; Role; STAT protein; STAT3 gene; Signal Pathway; Signal Transduction; Source; Suppressor of Cytokine Signaling Family Protein; TLR3 gene; Up-Regulation; Viral Encephalitis; Viral Load result; Virus Diseases; Wild Type Mouse; base; blood-brain barrier disruption; brain endothelial cell; cell type; cerebral microvasculature; chemokine; cytokine; exosome; improved; in vivo; macrophage; monocyte; mortality; neuroinflammation; neurotropic; neurotropic virus; novel; novel therapeutic intervention; overexpression; pathogen; promoter; receptor; recruit; response; sensor; transcription factor

ABSTRACT The maintenance of homeostasis in the central nervous system (CNS) during neurotropic viral infections is critical for host survival. Inflammation of the CNS is characterized by the recruitment of leukocytes to the brain and the activation of resident CNS cells, including astrocytes. The pivotal in vivo function of astrocytes during viral encephalitis is poorly defined. Toll like receptor 3 (TLR3) is an interferon-inducing double stranded RNA sensor that senses viral infections. Although it is well known that astrocytes express functional TLR3, its role in response to viral RNA infections of the CNS remains poorly understood. The interplay between transcription factors and microRNAs, and their upstream pattern recognition receptors during viral encephalitis has not been extensively studied. MicroRNA-155 (miR-155) has a key role of in the fine-tuning of TLR responses, and exerts both positive and negative regulation during inflammation. We and others have identified miR-155 as one of the most over- expressed miRNAs after stimulation of TLR3 with the synthetic dsRNA agonist Poly (I:C) in macrophages. In the CNS, miR-155 is also expressed in glial cells. However, a CNS-intrinsic role of miR-155 has yet to be defined in vivo. Signal transducers and activators of transcription (STAT) proteins and their negative regulators, the suppressors of cytokine signaling (SOCS) protein family play central roles in regulating many cytokines and growth factors. Recently, two STAT3 binding sites have been identified in the promoter of miR-155. Furthermore, miR-155 targets SOCS1 and SOCS3 suppressing their negative regulatory effect on JAK/STAT signaling pathway. Coronaviruses (CoVs) are enveloped, positive-sense, single-stranded, polyadenylated RNA viruses. Mouse hepatitis virus JHM (MHV-JHM) is a CNS-tropic strain that induces fatal encephalitis, associated with non-protective, enhanced macrophage and neutrophil infiltration that correlates with exacerbated chemokine and Th1 response and no Th17 involvement. We have identified a TLR3-dependent, miR-155 /STAT3 regulatory loop that promotes exacerbated, detrimental neuroinflammation and blood brain barrier (BBB) disruption in response to MHV-JHM fatal infection. Intriguingly, TRIF, the only TLR3 adaptor known, is not required for TLR3- dependent activation of STAT3 and induction of miR-155 both in vitro and in vivo. We have identified astrocytes and inflammatory monocytes infiltrating the brain as the cell sources driving exacerbated STAT3 activation. Our findings prompted us to investigate the following hypotheses: AIM 1 MHV-JHM infection in astrocytes and macrophages induces STAT3 activation through a TRIF-independent, TLR3-dependent non-canonical axis that is negatively regulated by its adaptor TRIF; AIM 2 Cell type specific genetic ablation of STAT3 and miR-155 expression in astrocytes or myeloid cells will reduce detrimental neuroinflammation and increase host survival after fatal MHV-JHM infection; and AIM 3 TLR3 contributes to BBB disruption through early and sustained upregulation of miR-155 expression in astrocytes, which deliver exosome-miR155 to brain endothelial cells affecting the expression of (to be identified in AIM 3) AJPs, TJPs, and/or MMPs.

抽象的 在神经性病毒感染期间，中枢神经系统（CNS）维持体内平衡是至关重要的 用于宿主生存。中枢神经系统的炎症的特征是将白细胞募集到大脑和 包括星形胶质细胞在内的常驻CNS细胞的激活。病毒期间星形胶质细胞的体内关键功能 脑炎的定义很差。 Toll像受体3（TLR3）是一种诱导干扰素的双链RNA传感器 这会感知病毒感染。尽管众所周知，星形胶质细胞表达功能性TLR3，但其在响应中的作用 对于中枢神经系统的病毒RNA感染仍然知之甚少。转录因子和 在病毒脑炎期间，microRNA及其上游模式识别受体尚未广泛 研究。 microRNA-155（miR-155）在TLR反应的微调中具有关键作用，并且施加了阳性 和炎症期间的负调节。我们和其他人已经将miR-155确定为最过分的 用巨噬细胞中的合成dsRNA激动剂聚（I：c）刺激TLR3后表示miRNA。在 CNS，miR-155在神经胶质细胞中也表达。但是，miR-155的CNS内在作用尚未定义 体内。转录（Stat）蛋白及其负调节剂的信号换能器和激活剂，即 细胞因子信号传导（SOCS）蛋白家族的抑制剂在调节许多细胞因子和 增长因素。最近，在miR-155的启动子中发现了两个STAT3结合位点。此外， miR-155靶向SOCS1和SOCS3抑制其对JAK/STAT信号的负面调节作用 路径。冠状病毒（COV）被包裹，阳性，单链，聚腺苷酸化的RNA病毒。 小鼠肝炎病毒JHM（MHV-JHM）是一种CNS循环菌株，可诱导与致命性脑炎有关 非保护，增强的巨噬细胞和中性粒细胞浸润，与恶化的趋化因子相关 和Th1响应，没有TH17参与。我们已经确定了TLR3依赖性miR-155 /STAT3调节 促进恶化，有害神经炎症和血液脑屏障（BBB）破坏的环路 对MHV-JHM致命感染的反应。有趣的是，Trif是唯一已知的TLR3适配器，TLR3-不需要 STAT3的依赖激活以及MiR-155的体外和体内诱导。我们已经确定了星形胶质细胞 随着驱动加剧的STAT3激活的细胞源，炎症单核细胞渗入大脑。我们的 调查结果促使我们研究以下假设：AIM 1 MHV-JHM在星形胶质细胞中感染和 巨噬细胞通过独立于TRIF的TLR3依赖性非统计轴诱导STAT3激活 受其适配器TRIF的负调节； AIM 2 STAT3和MIR-155的细胞类型特异性遗传消融 星形胶质细胞或髓样细胞中的表达将减少有害的神经炎症并增加宿主存活率 致命的MHV-JHM感染后； AIM 3 TLR3通过早期和持续而导致BBB中断 星形胶质细胞中miR-155表达的上调，这些表达向脑内皮细胞传递外泌体-MIR155 影响（在AIM 3中识别）AJP，TJP和/或MMP的表达。