HSV-1 Encoded MicroRNAs in the Pathogenesis and Treatment of Ocular Herpes

HSV-1 编码的 MicroRNA 在眼疱疹发病机制和治疗中的作用

• 批准号: 10569577
• 负责人: Afsar Raza Naqvi
• 金额: $ 41万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-03-01 至 2025-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10569577
• 关键词: Acyclovir; Affect; Aftercare; Animals; Antigen Presentation Pathway; Antigen-Presenting Cells; Antigens; Antiviral Agents; Antiviral Response; Attenuated; Autologous; B-Lymphocytes; Biological Process; Blindness; Blood; CD4 Positive T Lymphocytes; CD8B1 gene; Cell Line; Cell physiology; Cells; Code; Cornea; Data; Dendritic Cells; Development; Disease; Disease Progression; Disease model; Double Stranded DNA Virus; Drug resistance; Exhibits; Experimental Designs; Eye; Eye Infections; Eye diseases; Family; Flow Cytometry; Gene Expression; Genes; Genome; Goals; Herpes Labialis; Herpesviridae; Herpesviridae Infections; Herpesvirus 1; Herpetic Keratitis; Host Defense; Human; Immune; Immune Evasion; Immune response; Immune system; Impairment; Infection; Infiltration; Inflammation; Interferon Activation; Interferons; Keratitis; Kinetics; Knowledge; Lytic; Lytic Virus; Macrophage; Measures; Mediating; Messenger RNA; MicroRNAs; Morbidity - disease rate; Mus; Myeloid Cells; Natural Killer Cells; Oligonucleotides; Outcome; Pathogenesis; Pathology; Pathway interactions; Pattern; Persons; Production; Productivity; Proteins; RNA; Regulation; Reporting; Resolution; Role; Severities; Severity of illness; Sorting; Supporting Cell; Suspensions; Swab; T cell response; T-Cell Activation; T-Lymphocyte; Therapeutic; Tissues; Topical application; Transcript; Transfection; Tropism; United States; Untranslated RNA; Vaccines; Viral; Viral Antigens; Viral Genes; Viral Genome; Viral Load result; Viral Physiology; Virion; Virus; Virus Activation; Virus Replication; Work; analog; cytokine; cytotoxic; defense response; design; functional disability; immune cell infiltrate; immune function; immunoregulation; in vitro Assay; in vivo; inhibitor; insight; latent infection; lytic gene expression; member; mouse model; neutrophil; new therapeutic target; novel; pathogenic virus; prevent; response; seropositive; single-cell RNA sequencing; therapeutic target; translational study; uptake; virus host interaction

Herpes Simplex Virus-1 (HSV-1), a highly transmissible infection, is common and endemic throughout the world. Similar to other human herpesviruses (HHV), HSV-1 maintains lifelong latency inside host that requires immune evasion through various sophisticated mechanisms. Although HSV-1 is equipped with large repertoire (>80) of protein coding genes, the commonly accepted manifestations of viral gene expression during latency are the accumulation of a noncoding transcript and a set of microRNAs (miR). HSV-1 encoded viral miRNAs (v-miRs) are demonstrated to control expression of both viral and host transcripts and regulate viral tropism, lytic switching, immune subversion, etc. While multiple studies have examined HSV-1 profiles in various cell lines, key biological functions of these v-miRs remain unknown. Therefore, we propose to evaluate: (1) systematic expression dynamics of v-miRs during disease progression and reactivation, (2) comprehensive role in the pathogenesis by perturbing immune cells functions and (3) therapeutic targeting of v-miR by synthetic oligonucleotides to mitigate HSV-1-mediated ocular herpes. Using our established mouse model of ocular herpes, we will compare HSV-1 miRNA profiles in primary and reactivated mice corneal tissues and blood. Identifying positive and negative regulatory v-miRs can yield novel insights into host-virus interaction. The information gained will be used in designing therapeutic v-miR Inhibitors to silence candidate v-miRs functions. Effect of synthetic oligonucleotides (v-miR Inhibitors) targeting candidate v-miRs will be assessed on ocular disease progression in a mice model. V-miR inhibitors (alone or in combination) will be topically delivered and virus release, viral transcript/genome, and disease severity score will be measured. Next, we will evaluate how v-miRs can render host immune system dysfunctional, an integral feature required for HSV-1 persistence. Using v-miR inhibitors, we will assess whether immune infiltration and functions can be restored in vivo. Immune cell subsets will be comprehensively profiled in virus-infected animals treated with v-miR inhibitor using flow cytometry and single cell RNA sequencing. In addition, transcript expression profiles of genes related to antigen processing/presentation pathway, critical for potent antiviral response, will be quantified. This will identify the in vivo mechanisms through which v-miRs can facilitate immune evasion in ocular tissues. Next, we will dissect underlying mechanisms of v-miR-mediated dysregulation of antigen processing/presentation by macrophages and dendritic cells and activation of T cells. V-miR expressing myeloid cells will be assessed for uptake and processing of viral antigens and activation of autologous T helper (CD4+) and T cytotoxic (CD8+) cells. In addition, we will assess the impact of v-miRs on the polarization of CD4+ T cells. The data generated will provide significant information to existing knowledge gaps. Overall, the proposed translational study focuses on identifying the therapeutic and mechanistic aspect of v-miRs in ocular disease pathogenesis through modulation of immune cell responses.

单纯疱疹病毒1型(HSV-1)是一种高度传染性的传染病，在世界各地普遍存在，并在世界范围内流行。类似 对于其他人类疱疹病毒(HHV)，HSV-1在宿主内保持终身潜伏，需要通过免疫逃避 各种复杂的机械装置。尽管HSV-1配备了大量蛋白质编码基因(&gt；80)，但 在潜伏期间病毒基因表达的普遍表现是非编码基因的积累 转录本和一组microRNAs(MiR)。HSV-1编码的病毒miRNAs(v-miRs)被证明控制 病毒和宿主转录并调节病毒嗜性、裂解转换、免疫颠覆等，而多项研究 已经研究了不同细胞系中的HSV-1图谱，但这些v-miR的关键生物学功能尚不清楚。因此， 我们建议评估：(1)v-miRs在疾病进展和再激活过程中的系统表达动态，(2) 干扰免疫细胞功能在发病机制中的综合作用和(3)v-miR靶向治疗 合成寡核苷酸用于缓解HSV-1介导的眼部疱疹。使用我们建立的眼部小鼠模型 对于疱疹病毒，我们将比较原始和重新激活的小鼠角膜组织和血液中HSV-1的miRNA图谱。识别 正向和负向调节v-miRs可以对宿主与病毒的相互作用产生新的见解。所获得的信息将 用于设计治疗性v-miR抑制剂以沉默候选v-miRs功能。人工合成的效果 靶向候选v-miR的寡核苷酸(v-miR抑制剂)将在小鼠的眼部疾病进展中进行评估 模特。V-miR抑制剂(单独或联合使用)将局部给药，病毒释放，病毒转录本/基因组， 并测量疾病严重程度评分。接下来，我们将评估v-miRs如何使宿主免疫系统 功能障碍，这是HSV-1持续存在所需的一个完整特征。使用v-miR抑制剂，我们将评估免疫 在体内可以恢复渗透和功能。免疫细胞亚群将在感染病毒的患者中进行全面分析 应用流式细胞术和单细胞RNA测序技术对v-miR抑制剂处理的动物进行检测。此外，文字记录 与抗原处理/提呈途径有关的基因的表达谱，这些基因对有效的抗病毒反应至关重要， 将会被量化。这将确定v-miRs促进眼部免疫逃避的体内机制。 纸巾。接下来，我们将剖析v-miR介导的抗原失调的潜在机制。 巨噬细胞和树突状细胞的处理/提呈和T细胞的激活。表达V-miR的髓系细胞将 评估病毒抗原的摄取和处理以及自体T辅助细胞(CD4+)和T细胞毒的激活 (CD8+)细胞。此外，我们还将评估v-miRs对CD4+T细胞极化的影响。生成的数据将 为现有的知识差距提供重要信息。总体而言，拟议的翻译研究侧重于 通过调控v-miRs在眼病发病机制中的作用 免疫细胞反应。