The miR-183/96/182 Cluster in Pseudomonas aeruginosa-induced Keratitis

铜绿假单胞菌诱导的角膜炎中的 miR-183/96/182 簇

• 批准号: 10656958
• 负责人: SHUNBIN XU
• 金额: $ 42.88万
• 依托单位: WAYNE STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-04-01 至 2027-05-01
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10656958
• 关键词: Acceleration; Address; Afferent Neurons; Antibacterial Response; Antibiotic Resistance; Antibiotic Therapy; Apoptosis; Bacteria; Bacterial Antibiotic Resistance; Biogenesis; Biological; Blindness; C57BL/6 Mouse; Cells; Communicable Diseases; Cornea; Corneal Diseases; Data; Development; Disease; Disease Management; Disinhibition; Exhibits; Fluorescein; Gene Expression; Genes; Hour; Human; Immune; Immune response; In Vitro; Infection; Inflammation Mediators; Inflammatory; Inflammatory Response; Injections; Keratitis; Knock-out; Knockout Mice; Knowledge; Label; Lead; Lipopolysaccharides; Mediating; Metabolic; MicroRNAs; Molecular; Mus; Natural Immunity; Nerve; Neuroimmune; Neuropeptides; Neuropilins; Play; Production; Pseudomonas aeruginosa; Pseudomonas aeruginosa infection; Publishing; Regulation; Regulator Genes; Research; Resolution; Role; Sensory; Severities; Severity of illness; Structure of trigeminal ganglion; Substance P; TAC1 gene; TLR4 gene; TRPV1 gene; Testing; Tissues; Topical application; Untranslated RNA; Visual impairment; Wild Type Mouse; afferent nerve; alternative treatment; base; capsaicin receptor; conditional knockout; cytotoxic; density; formyl peptide; genome-wide analysis; human disease; in vivo; insight; knock-down; mouse model; nerve supply; neurite growth; neutrophil; novel; prevent; prophylactic; protective effect; receptor; resistant strain; therapeutic miRNA; therapeutic target; transcriptome sequencing

Pseudomonas aeruginosa (PA) keratitis is one of the most rapidly developing and destructive diseases of the cornea and a global cause of visual impairment and blindness. Emergence of antibiotic-resistant strains poses additional challenges for effective disease management. Development of alternative treatment is urgent. In this regard, microRNAs (miRNAs) are small, non-coding RNAs and important regulators of gene expression. miRNAs play critical roles in human diseases and are viable therapeutic targets. However, the roles of miRNAs in PA keratitis remain largely unexplored. Our long-term objectives are to uncover the molecular mechanisms of miRNAs in ocular infectious diseases, identify novel miRNA-based therapeutic targets and develop alternative treatment of these diseases. The proposed research will directly address this knowledge gap. It is built upon our recently published and strong preliminary data showing that application of anti-miRs targeting the miR-183/96/182 cluster (referred to as miR-183C from here on) and knockout of miR-183C in mice decreases corneal nerve density and neuropeptide production, while reducing the severity of PA keratitis; miR- 183C targets key genes regulating corneal sensory innervation, e.g., Nrp1, bacterium-induced sensory-neuron activation and neuropeptide production, e.g., Toll-Like Receptor (TLR)4, Formyl Protein Receptor (Fpr)1 and substance P (sP) precursor gene Tac1. These data lead us to the overarching hypothesis that, in addition to innate immunity, miR-183C modulates PA keratitis through its regulation of corneal sensory nerve function and neuroimmune interaction by targeting key genes involved in sensory innervation, PA-induced sensory-neuron activation and pro-inflammatory neuropeptide production. Three Aims are proposed in this application to test this hypothesis. In Aim 1, fluorescein amidites (FAM)-labeled anti-miR-183C will be applied to the cornea of wild-type (WT) mice to test the hypothesis that anti-miR-183C treatment upregulates Tac1, TLR4, Fpr1 and Nrp1 in corneal sensory nerves, resulting in increased pro-inflammatory neuropeptides (e.g., sP) and an early immune/inflammatory response (<24 hours post-infection) and an accelerated sensory-nerve reduction and decreased pro-inflammatory neuropeptides and immune/inflammatory response in a later stage during disease resolution. Aim 2 will test the hypothesis using a sensory neuron-specific miR-183C conditional knockout mouse model. Aim 3 will test the hypothesis that knockdown or knockout of miR-183C in vitro/ex vivo in both human and mouse sensory neurons parallels the in vivo data in that it will upregulate Tac1, TLR4 and Fpr1 to enhance PA-induced secretion of pro-inflammatory neuropeptides by sensory neurons, and increase Nrp1 expression to inhibit neurite growth. A genome-wide identification of miR-183C target genes in both human and mouse trigeminal ganglion sensory neurons will also be conducted by RNA seq. This study will provide novel biological insights into molecular bases of miR-183C's regulation of PA keratitis and other ocular infectious diseases. It will uncover mechanisms and reveal new targets for anti-miR therapy.

铜绿假单胞菌 (PA) 角膜炎是人类发展最快、最具破坏性的疾病之一。 角膜和视力障碍和失明的全球原因。抗生素耐药菌株的出现造成 有效疾病管理面临的额外挑战。开发替代疗法刻不容缓。 在这方面，microRNA (miRNA) 是小型非编码 RNA，也是基因表达的重要调节因子。 miRNA 在人类疾病中发挥着关键作用，并且是可行的治疗靶点。然而，miRNA 的作用 PA 角膜炎的发病机制在很大程度上仍未被探索。我们的长期目标是揭示分子机制 眼部感染性疾病中的 miRNA 的研究，确定基于 miRNA 的新治疗靶点并开发 这些疾病的替代治疗。拟议的研究将直接解决这一知识差距。这是 基于我们最近发布的强有力的初步数据，表明抗 miR 靶向的应用 miR-183/96/182簇（以下简称miR-183C）和小鼠中miR-183C的敲除 降低角膜神经密度和神经肽的产生，同时减轻 PA 角膜炎的严重程度； miR- 183C 靶向调节角膜感觉神经支配的关键基因，例如 Nrp1、细菌诱导的感觉神经元 激活和神经肽生成，例如 Toll 样受体 (TLR)4、甲酰蛋白受体 (Fpr)1 和 P物质（sP）前体基因Tac1。这些数据使我们得出一个总体假设：除了 先天免疫，miR-183C 通过调节角膜感觉神经功能来调节 PA 角膜炎 通过靶向参与感觉神经支配、PA 诱导的感觉神经元的关键基因来实现神经免疫相互作用 激活和促炎神经肽的产生。本申请提出了三个目标来测试 这个假设。在目标 1 中，荧光素酰胺 (FAM) 标记的抗 miR-183C 将应用于患者的角膜。 野生型 (WT) 小鼠来检验抗 miR-183C 治疗上调 Tac1、TLR4、Fpr1 和 角膜感觉神经中的 Nrp1，导致促炎神经肽（例如 sP）增加，并导致早期 免疫/炎症反应（感染后<24小时）和加速的感觉神经减少和 疾病后期促炎神经肽和免疫/炎症反应减少 解决。目标 2 将使用感觉神经元特异性 miR-183C 条件敲除来检验假设 鼠标模型。目标 3 将检验以下假设：在体外/离体中敲低或敲除 miR-183C 人类和小鼠感觉神经元与体内数据相似，因为它会上调 Tac1、TLR4 和 Fpr1 增强 PA 诱导的感觉神经元促炎神经肽的分泌，并增加 Nrp1 表达抑制神经突生长。人类和人类中 miR-183C 靶基因的全基因组鉴定 小鼠三叉神经节感觉神经元也将通过 RNA seq 进行。 这项研究将为 miR-183C 调节 PA 角膜炎的分子基础提供新的生物学见解 以及其他眼部感染性疾病。它将揭示抗 miR 治疗的机制并揭示新的靶点。

项目成果

Mutation Screening in the miR-183/96/182 Cluster in Patients With Inherited Retinal Dystrophy.

• DOI: 10.3389/fcell.2020.619641
• 发表时间: 2020
• 期刊: Frontiers in cell and developmental biology
• 影响因子: 5.5
• 作者: Xu S;Coku A;Muraleedharan CK;Harajli A;Mishulin E;Dahabra C;Choi J;Garcia WJ;Webb K;Birch D;Goetz K;Li W
• 通讯作者: Li W

The MicroRNA-183-96-182 Cluster Promotes T Helper 17 Cell Pathogenicity by Negatively Regulating Transcription Factor Foxo1 Expression.

• DOI: 10.1016/j.immuni.2016.05.015
• 发表时间: 2016-06-21
• 期刊: Immunity
• 影响因子: 32.4
• 作者: Ichiyama K;Gonzalez-Martin A;Kim BS;Jin HY;Jin W;Xu W;Sabouri-Ghomi M;Xu S;Zheng P;Xiao C;Dong C
• 通讯作者: Dong C