Purinergic Signaling in Varicella Zoster Virus Vasculopathy

水痘带状疱疹病毒血管病中的嘌呤能信号传导

基本信息

批准号：
9331756
负责人：
Maria Acena Nagel
金额：
$ 34.02万
依托单位：
UNIVERSITY OF COLORADO DENVER
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-09-01 至 2019-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9331756
关键词：
ADP Receptors Adenosine Alpha Cell Aneurysm Antiviral Agents Arteries Attenuated Binding Blood Vessels Brain hemorrhage Cadaver Cell surface Cells Central Nervous System Diseases Cerebrovascular system Chemosensitization DNA DNA Virus Infections Data Elderly Event Extracellular Matrix Fibroblasts Ganglia Germ Cells Herpes zoster disease Herpesviridae Herpesvirus 1 Herpesvirus Type 3 Human In Vitro Incidence Infection Integrins Ischemia Lead Measurement Measures Medial Mediating Membrane Microscopy Modeling Myofibroblast Pathologic Pathway interactions Phenotype Population Purinoceptor RNA Virus Infections Receptor Activation Risk Role Signal Pathway Signal Transduction Simplexvirus Smooth Muscle Smooth Muscle Myocytes Staining method Stains Stroke Structure Testing Time Tunica Adventitia Vascular remodeling Virus Virus Diseases Visual Zoster Vaccine cell motility cerebral artery cerebrovascular clopidogrel experimental study extracellular in vivo inhibitor/antagonist matrigel migration novel prevent public health relevance receptor stroke treatment tripolyphosphate varicella zoster virus vasculopathy viral DNA

项目摘要

DESCRIPTION (provided by applicant): Varicella zoster virus (VZV) is a ubiquitous, human alphaherpesvirus that can reactivate from ganglia, infect cerebral arteries, and cause stroke (VZV vasculopathy). Specifically, VZV-infected arteries have a thickened intima composed of myofibroblasts, likely originating from smooth muscle cells or adventitial fibroblasts, which can contribute to luminal occlusion and ischemia, and have loss of medial smooth muscle cells which can contribute to aneurysm formation and hemorrhagic stroke. Binding of extracellular adenosine 5' triphosphate (ATP) and its metabolites to specific purinergic receptors on the cell surface (purinergic signaling) has emerged as important in virus infection and pathological vascular remodeling. Our preliminary data show that: (1) compared to mock-infected cells, extracellular ATP is rapidly increased upon VZV infection of human cerebrovascular adventitial fibroblasts, which are the initial cells infected in arteries and which are the key regulators of vascular tone and structure, and (2) blockade of cellular ATP/ADP receptors, particularly with P2Y12 receptor inhibitor clopidogrel, attenuates virus infection. Thus, we hypothesize that specific purinergic receptor subtypes in cerebral vasculature mediate VZV infection of adventitial fibroblasts and also promote VZV-induced pathological cerebrovascular remodeling and stroke. Herein, we will identify the mechanism(s) by which activation of P2Y12 receptors promotes VZV infection in cerebrovascular adventitial fibroblasts (Aim 1). Since P2Y12 receptor activation has been shown to activate specific integrins downstream, including a3b2 and _b1, which can be used for virus entry, we will treat VZV-infected human cerebrovascular adventitial fibroblasts with select integrin inhibitors involved in the P2Y12 pathway, as well as in herpesvirus entry into cells. One and 3 days later, virus will be titered and viral DNA quantitated by PCR to identify integrin chains/heterodimers involved in VZV infection. After identification of these specific integrins, we will determine if specific integrin activation is downstream of P2Y12 receptor activation by examining possible potentiation effects of combined integrin/P2Y12 inhibitors on viral DNA and titers. We will also identify the specific purinergic receptor(s) that mediates VZV-induced cerebrovascular remodeling by measuring the migration of VZV or mock-infected human cerebrovascular adventitial fibroblasts and smooth muscle cells in vitro with and without specific purinergic receptor inhibitors using matrigel invasion chambers (Aim 2a). Finally, additional preliminary data showed that VZV-infection of cadaveric human cerebral arteries ex vivo mimics vessel wall changes seen in vivo in VZV vasculopathy (thickening of the intimal layer) which is absent in mock-infection - providing a model to identify specific purinergic receptors involved in VZV-induced remodeling within a whole artery (Aim 2b). Thus, we will VZV or mock-infect cadaveric human cerebral artery explants with and without specific purinergic receptor inhibitors and examine effects on vascular remodeling 2 and 4 weeks later by measuring intimal thickening.

描述（由适用提供）：水痘带状疱疹病毒（VZV）是一种无处不在的人α掌病毒，可以从神经节，感染大脑动脉重新激活并引起中风（VZV Vasculopopathy）。具体而言，VZV感染的动脉具有由肌成纤维细胞组成的增厚内膜，可能源自平滑肌细胞或晚期纤维细胞，可导致腔内闭塞和缺血，并导致培养基平滑肌细胞的丧失，从而对Aneurysm形成和血液造成的肌肉造成损失。细胞外腺苷5'三磷酸（ATP）及其代谢产物与细胞表面上特定的嘌呤能受体的结合（嘌呤能信号）在病毒感染和病理血管重塑中已经出现了很重要。我们的初步数据表明：（1）与模拟感染的细胞相比，在VZV感染人脑脑血管晚期成纤维细胞时，细胞外ATP迅速增加，这是动脉感染的初始细胞，它们是血管张力和结构的关键调节剂，以及（2）与细胞clop clop clop clop clop clop clop clop condifor intp py22 inf inf py2y12 inf inf2y12的调节器，病毒感染。这就是我们假设脑血管培养基VZV感染晚期成纤维细胞中的特定嘌呤能受体亚型，还促进了VZV诱导的病理脑血管重塑和中风。本文中，我们将确定P2Y12受体激活促进脑血管进步成纤维细胞中VZV感染的机制（AIM 1）。由于已显示P2Y12受体激活可以激活下游特定的整合素，包括A3B2和_b1，可用于病毒进入，我们将与参与P2Y12 Pathway的精选整合蛋白抑制剂以及Herspesvirus入口中的精选整合蛋白抑制剂一起治疗VZV感染的人口脑脑脑血压成纤维细胞。细胞。一日和3天后，PCR定量定量的病毒和病毒DNA，以鉴定与VZV感染有关的整合蛋白链/异二聚体。在鉴定出这些特定的整合素后，我们将通过检查联合素/P2Y12抑制剂对病毒DNA和滴度的联合素/P2Y12抑制剂的可能影响，确定特定的整联蛋白激活是否在P2Y12受体激活的下游。我们还将通过测量VZV或模拟感染的人口脑血管成纤维细胞和平滑肌细胞的迁移，并在使用基质受体抑制剂的情况下介导VZV或模拟感染的人口脑血管前成纤维细胞和平滑肌细胞，从而介导VZV诱导的脑血管重塑的特定特异性嘌呤能受体。最后，其他初步数据表明，尸体人体脑动脉的VZV感染在VZV VasculoPathy（内膜层的增厚）中在体内观察到的体内模拟船只壁变化，这些变化在模拟型中不存在 - 在VZV-RemodeLeled中鉴定了涉及vzv诱导的特定模型的模型，以识别特定的Purinerggic受体。这是，我们将有或没有特定的嘌呤能受体抑制剂进行VZV或模拟感染的尸体尸体脑动脉外植体，并通过测量亲膜增厚2和4周检查2和4周后对血管重塑的影响。