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: 10391770
• 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/10391770
• 关键词: Acyclovir; Address; 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; CD8-Positive T-Lymphocytes; Cell Line; Cell physiology; Cells; Code; Cornea; Data; Dendritic Cells; Dendritic cell activation; Development; Disease; Disease Progression; Disease model; Double Stranded DNA Virus; Drug resistance; Exhibits; Experimental Designs; Eye; Eye Infections; 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; Measures; Mediating; Messenger RNA; MicroRNAs; Morbidity - disease rate; Mus; Myeloid Cells; Natural Killer Cells; Oligonucleotides; Outcome; Pathogenesis; Pathology; Pathway interactions; Pattern; Persons; Production; Proteins; RNA; Regulation; Reporting; Resolution; Role; Severities; Severity of illness; Sorting - Cell Movement; Supporting Cell; Suspensions; Swab; T cell response; T-Cell Activation; T-Lymphocyte; Therapeutic; Tissues; Topical application; Transcript; Tropism; Tumor-infiltrating immune cells; United States; Untranslated RNA; Vaccines; Viral; Viral Antigens; Viral Genes; Viral Genome; Viral Load result; Virion; Virus; Virus Activation; Virus Replication; Work; analog; cytokine; cytotoxic; defense response; design; functional disability; immune function; immunoregulation; in vitro Assay; in vivo; inhibitor; insight; latent infection; lytic gene expression; macrophage; 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.

1 型单纯疱疹病毒 (HSV-1) 是一种高度传播的感染，在全世界都很常见和流行。相似的 与其他人类疱疹病毒 (HHV) 不同，HSV-1 在宿主体内维持终生潜伏期，需要通过以下方式进行免疫逃避： 各种复杂的机制。尽管 HSV-1 配备了大量 (>80) 的蛋白质编码基因，但 潜伏期病毒基因表达的普遍接受的表现是非编码基因的积累 转录本和一组 microRNA (miR)。 HSV-1 编码的病毒 miRNA (v-miR) 被证明可以控制 病毒和宿主转录本并调节病毒向性、裂解转换、免疫颠覆等。多项研究 虽然已经检查了各种细胞系中的 HSV-1 谱，但这些 v-miR 的关键生物学功能仍然未知。所以， 我们建议评估：（1）疾病进展和重新激活期间 v-miR 的系统表达动态，（2） 通过扰乱免疫细胞功能在发病机制中发挥综合作用，以及（3）通过 v-miR 进行治疗靶向 合成寡核苷酸可减轻 HSV-1 介导的眼部疱疹。使用我们建立的小鼠眼模型 疱疹，我们将比较原代小鼠和再激活小鼠角膜组织和血液中的 HSV-1 miRNA 谱。识别 正向和负向调节 v-miR 可以对宿主-病毒相互作用产生新的见解。获得的信息将 可用于设计治疗性 v-miR 抑制剂以沉默候选 v-miR 功能。合成效果 将评估针对候选 v-miR 的寡核苷酸（v-miR 抑制剂）对小鼠眼部疾病进展的影响 模型。 V-miR抑制剂（单独或组合）将被局部递送并释放病毒、病毒转录物/基因组、 并将测量疾病严重程度评分。接下来，我们将评估v-miRs如何渲染宿主免疫系统 功能失调，这是 HSV-1 持续存在所需的一个不可或缺的特征。使用 v-miR 抑制剂，我们将评估是否免疫 在体内可恢复浸润和功能。将全面分析病毒感染的免疫细胞亚群 使用流式细胞术和单细胞 RNA 测序使用 v-miR 抑制剂治疗动物。另外，转录 与抗原加工/呈递途径相关的基因的表达谱，对于有效的抗病毒反应至关重要， 将被量化。这将确定 v-miRs 促进眼部免疫逃避的体内机制。 组织。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/呈递以及 T 细胞的激活。表达 V-miR 的骨髓细胞将 评估病毒抗原的摄取和加工以及自体 T 辅助细胞 (CD4+) 和 T 细胞毒性的激活 (CD8+) 细胞。此外，我们将评估 v-miR 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，拟议的转化研究侧重于 通过调节 v-miRs 在眼部疾病发病机制中的治疗和机制方面 免疫细胞反应。