Targeting Ischemia/Reperfusion Stress to Inhibit Cytomegalovirus Reactivation After Lung Transplant

针对肺移植后缺血/再灌注应激抑制巨细胞病毒再激活

• 批准号: 10453246
• 负责人: Zheng J Zhang
• 金额: $ 24万
• 依托单位: NORTHWESTERN UNIVERSITY 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: 2022-02-25 至 2024-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10453246
• 关键词: Address; Alveolar Macrophages; Animal Model; Binding; Birds; Cells; Cellular Stress; Chemicals; Clinical; Complication; Cytomegalovirus; DNA biosynthesis; Data; Day Surgery; Disease; Donor person; Early Promoters; Enzymes; Epigenetic Process; Foundations; Future; Gene Expression; Genetic Transcription; Goals; Hour; Human; Immediate-Early Genes; Immune; Immunosuppression; In Vitro; Inflammation; Inflammatory; Infusion procedures; Inositol; Interleukin-1 Receptors; Ischemia; Kidney; Kidney Transplantation; Knockout Mice; Left; Lung; Lung Transplantation; Mediating; Molecular; Murid herpesvirus 1; Mus; Myelogenous; NF-kappa B; Opportunistic Infections; Organ; Organ Donor; Organ Preservation; Organ Transplantation; Pathway interactions; Perioperative; Peripheral; Primary Infection; Procedures; Prophylactic treatment; RNA analysis; Regimen; Reperfusion Injury; Reperfusion Therapy; Role; Salivary Glands; Signal Pathway; Signal Transduction; Signaling Molecule; Site; Stress; System; Testing; Therapeutic; Time; Tissues; Toll-like receptors; Transcription Factor AP-1; Transcriptional Activation; Transplant Recipients; Transplantation; Transplantation Surgery; Viral; Viral Load result; Virus; Virus Replication; allograft rejection; cell type; clinical translation; clinically relevant; high risk; implantation; inhibitor; innovation; mRNA Expression; macrophage; mouse model; nanocarrier; novel; novel therapeutics; post-transplant; pre-clinical; preconditioning; prevent; reactivation from latency; response; sensor; seropositive; therapeutic evaluation; therapeutically effective; transcription factor; transcriptome; viral DNA

PROJECT SUMMARY Cytomegalovirus (CMV) is the most common opportunistic infection in lung transplant recipients, with the highest risk seen in seronegative recipients (R-) of seropositive organ donors (D+). While anti-rejection immunosuppression (IS) has been implicated as a primary cause of CMV reactivation, our studies have newly suggested otherwise. We have recently shown that CMV reactivation occurs independent of IS and seemingly independent of the allo-immune response following a D+/R- kidney transplant in mice. Treatment with a regimen of clinically relevant IS, while failing to induce CMV reactivation in latently infected mice (no transplant), does permit subsequent viral replication and dissemination following transplant, despite inhibition of the host rejection response and inflammation. These findings suggest that ischemia/reperfusion (I/R) stress inherent in a transplant procedure due to organ preservation and revascularization is necessary and sufficient to induce reactivation. However, molecular mechanisms underlying I/R stress mediated transcriptional reactivation from latency in the grafts remain elusive. The goal for this study is to identify key molecular pathways responsible for I/R stress- mediated CMV reactivation and to develop effective therapeutics to prevent both I/R injury and CMV reactivation following lung transplant. In our preliminary studies using a mouse model of syngeneic D+/R- lung transplant, we observed that expression of MCMV immediate early (IE) genes is induced within 48 hours of transplant surgery, followed by increased viral DNA replication in the lung grafts and dissemination to peripheral organs by day 14 post-transplant. We revealed that pre-transplant depletion of donor alveolar macrophages (AMs) decreased viral load in transplanted lungs and salivary glands, suggesting that AMs are primary sites of MCMV latency and reactivation, consistent with the findings from HCMV studies. We have further showed that Myeloid differentiation primary response 88 (Myd88) was critical in mediating transplant induced I/R injury, likely via activation of transcription factors (TFs) NF-kB, AP-1 and XBP-1. These TFs are shown to bind to MIEP and promote MCMV/HCMV reactivation. Thus, we hypothesize that I/R stress mediates Myd88 dependent activation of TFs in viral harboring donor lung cells (e.g. AMs); leading to reactivation of latent CMV in the transplanted lungs. Thus, tissue/cell specific inhibition of Myd88 pathways in latently infected lung donors may prevent viral reactivation. We will use a preclinical mouse model of lung transplantation to test our hypotheses. In Aim 1, we will 1) investigate how donor macrophage-derived Myd88 regulate expression of viral IE genes and viral DNA replication by utilizing genetically modified time-specific myeloid Myd88 knockout mice and 2) delineate impact of Myd88 on I/R stress mediated signaling pathways in the lung cells using single cells RNA analysis. In Aim 2, we will determine the therapeutic potentials of tissue/cell-specific targeting of Myd88 in mitigating I/R stress and MCMV reactivation and dissemination leveraging our novel nanocarrier delivery system. Data generated from this study will provide proof of concept for developing novel therapeutics for further clinical translation.

项目摘要 巨细胞病毒（CMV）是肺移植受者中最常见的机会感染， 血清阳性器官供体（D+）的血清神经受体（R-）中看到的最高风险。而反拒绝 免疫抑制（IS）被认为是CMV重新激活的主要原因，我们的研究具有新的 建议其他建议。我们最近表明，CMV重新激活是独立于IS，看似 在小鼠中进行D+/R肾移植后，独立于同种免疫反应。用疗法治疗 临床上相关的是，同时未能诱导潜在感染的小鼠（无移植）的CMV重新激活 尽管抑制了宿主排斥反应，但随后允许随后的病毒复制和传播 反应和炎症。这些发现表明移植中固有的缺血/再灌注（I/R）应力 由于器官保存和血运重建而导致的程序是必要的，足以诱导重新激活。 但是，I/R应力介导的转录重新激活的分子机制是从潜伏期中的转录重新激活 移植物仍然难以捉摸。这项研究的目的是确定负责I/R应力的关键分子途径 - 介导的CMV重新激活并开发有效的治疗剂，以防止I/R损伤和CMV重新激活 肺移植后。在我们的初步研究中，使用同步D+/r肺移植的小鼠模型 我们观察到MCMV的表达早期（IE）基因在移植后的48小时内诱导 手术，然后在肺移植物中增加病毒DNA复制，并通过 第14天移植。我们揭示了供体肺泡巨噬细胞（AMS）的移植前耗竭 移植肺和唾液腺中病毒载量降低，表明AM是MCMV的主要部位 潜伏期和重新激活，与HCMV研究的发现一致。我们进一步表明髓样 分化主要响应88（MYD88）对于介导移植引起的I/R损伤至关重要，可能是通过 转录因子（TFS）NF-KB，AP-1和XBP-1的激活。这些TF显示与MIEP结合，并且 促进MCMV/HCMV重新激活。因此，我们假设I/R应力介导MYD88依赖性激活 病毒携带供体肺细胞的TF（例如AMS）；导致在移植中的潜在CMV重新激活 肺。因此，在潜在感染的肺供体中对MyD88途径的组织/细胞特异性抑制可能会预防病毒 重新激活。我们将使用肺移植的临床前小鼠模型来检验我们的假设。在AIM 1中，我们 将1）研究供体巨噬细胞衍生的MyD88如何调节病毒IE基因和病毒DNA的表达 通过利用转基因特异性的髓样MyD88敲除小鼠的复制和2）描述撞击 使用单细胞RNA分析，在肺细胞中I/R应力介导的信号通路上的MyD88的of。在AIM 2中， 我们将确定MyD88的组织/细胞特异性靶向I/R应力和 MCMV重新激活和传播利用了我们的新型纳米载体输送系统。生成的数据 这项研究将为开发新的治疗剂提供进一步临床翻译的概念证明。