Poxvirus adverse effects on cardiac cells and the heart

描述（申请人提供）：为了应对生物恐怖分子可能在美国释放天花病毒的担忧，政府实施了多阶段疫苗接种计划。然而，接种疫苗后观察到的心脏不良事件导致疾病控制中心建议将有心脏病风险的人排除在疫苗接种计划之外。值得注意的是，美国最近有18名天花疫苗接种者被诊断出心肌炎。尽管心肌炎和天花疫苗接种之间最近的时间关联以及类似的历史关联，但没有研究痘病毒对心脏细胞的影响，也没有开发任何动物模型来研究痘病毒诱导的心肌炎。对于其他病毒，心肌炎可以反映病毒对心肌细胞的直接致细胞病变作用，或者可以由免疫细胞、细胞因子或其他可溶性因子介导。痘病毒可以直接致细胞病变，但也编码许多细胞因子应答的调节剂。牛痘病毒含有影响免疫反应的已知正痘病毒辅助基因的完整补充，而DryVax（目前的牛痘病毒疫苗株）编码较少的辅助基因，而改良的安卡拉牛痘病毒（MVA，在人类细胞中有复制缺陷）含有更少的辅助基因。我们的初步数据表明，牛痘病毒和DryVax复制小鼠原代心肌细胞培养物（PCMCs），而MVA没有。此外，牛痘、DryVax和MVA对这些培养物均具有致细胞病变性，尽管致细胞病变性的类型是病毒株特异性的。我们推测痘病毒对心肌细胞具有病毒株特异性的直接或间接的不良影响，导致心肌细胞和心脏的损伤。在特定目标1中，我们将确定这三种痘病毒（牛痘、DryVax和MVA）对小鼠原代心肌细胞培养物（PCMCs）和对照培养物的影响，分析病毒感染和基因表达，以及细胞致病性的诱导。在具体目标2中，我们将确定细胞因子和痘病毒感染对PCMC和对照培养物的细胞致病性之间的关联。在具体目标3中，我们将确定这些痘病毒是否在各种小鼠品系中诱导心肌炎。R21申请中提出的研究将为未来的深入研究提供基础，以改进当前和未来的天花疫苗。

DESCRIPTION (provided by applicant): In response to concerns that bioterrorists may release smallpox virus in the United States, the government has implemented a multi-stage vaccination program. However, the observation of cardiac adverse events following vaccination has resulted in the Centers for Disease Control recommending that people at risk for heart disease should be excluded from the vaccination program. Notably, myocarditis has been diagnosed in 18 recent smallpox vaccinees in the USA. Despite this recent temporal association between myocarditis and smallpox vaccination as well as a similar historic association, there have been no studies of the effects of poxviruses on cardiac cells, or the development of any animal models to study poxvirus-induced myocarditis. For other viruses, myocarditis can reflect a direct cytopathogenic effect of the virus on cardiac myocytes, or can be mediated by immune cells, cytokines, or other soluble factors. Poxviruses can be directly cytopathogenic, but also encode a number of modulators of cytokine responses. Cowpox virus contains the full complement of known orthopoxvirus accessory genes that affect immune responses, while DryVax (the current vaccine strain of vaccinia virus) encodes fewer accessory genes, and modified vaccinia Ankara virus (MVA, which is replication-defective in human cells) contains even fewer accessory genes. Our preliminary data demonstrate that cowpox virus and DryVax replicate in murine primary cardiac myocyte cultures (PCMCs) whereas MVA does not. Moreover, cowpox, DryVax, and MVA are each cytopathogenic to these cultures, though the type of cytopathogenicity is virus strain-specific. We hypothesize that poxviruses have virus strain-specific adverse effects on cardiac cells, either directly or indirectly, resulting in damage to cardiac myocytes and the heart. In Specific Aim 1, we will determine the effect of these three poxviruses (cowpox, DryVax, and MVA) on murine primary cardiac myocyte cultures (PCMCs) and control cultures, analyzing viral infection and gene expression, and induction of cytopathogenicity. In Specific Aim 2, we will determine the association between cytokines and poxvirus infection on cytopathogenicity to PCMCs and control cultures. In Specific Aim 3, we will determine whether these poxviruses induce myocarditis in a variety of mouse strains. Studies proposed in this R21 application will provide the foundation for future in-depth studies with the goal of improving current and future smallpox vaccines.