Extra-pulmonary pathology caused by avian H5N1 and recombinant H1N1-1918 influenz

描述(由申请人提供)：目前禽流感在人类中的暴发强调了加强研究工作的必要性，以充分确定感染期间的发病机制。患者的临床过程和尸检病理表明，流感病毒存在于呼吸道以外的组织中，继发于呼吸道感染但仍未感染病毒的组织中，或两者兼而有之。动物研究还表明，禽流感病毒在肺外传播，其程度和涉及的器官似乎依赖于物种。评估非呼吸系统器官病理对细胞因子过度产生的确切贡献，这是人类疾病导致多器官衰竭的共同特征，对于成功的临床治疗至关重要，因为更好地理解导致脑病发展的因素也是常见的。不幸的是，感染后住院的时机，英勇的治疗努力的混乱效果，以及伦理考虑排除了对已经严重受损的人的主要器官活检，需要使用动物模型来研究肺部和其他可能受感染影响的主要器官的早期感染。在过去的3年里，我们成功地优化了人类流感感染的猕猴模型，用于基因组应用。为此，我们使用了一种轻度致病性的人类流感病毒(A/Texas/36/91)来仔细描述其临床病程、病理和对病毒mRNA、病毒蛋白或对活跃的病毒复制的转录调控。接下来，我们将用德克萨斯流感病毒感染食蟹猴，该病毒最多含有1918年大流行流感的三个基因(A/布雷维格使命/1/18)，据信来自一种禽类病毒，并感染高致病性禽流感(A/A/越南/1203/04)。除了前些年开展的工作外，我们还建议通过使用系统显微镜检查、流感抗原染色、免疫细胞原位表型和免疫组织化学染色等炎症和免疫过程的标记物，包括环氧合酶级联反应、巨噬细胞产生一氧化氮、低氧反应元件的诱导、血管生成等，对呼吸道以外的任何组织的病理进行深入表征。这一方法将通过对病变进行激光捕获显微解剖，然后对相应的mRNAs进行定量逆转录聚合酶链式反应来补充和证实。我们认为，结合几年来对这一模型的直接经验，以及对已知或可能对人类具有高毒力的相关流感病毒的使用，为阐明人类禽流感感染的这一重要方面提供了一个独特的机会。Bielefeldt Ohmann-R03叙述性禽流感在人类中的暴发已导致越来越高的发病率和死亡率。该病毒对人类宿主的进一步适应很可能导致一场世界性的流行病，如1918年或“西班牙”流感，据信是禽源性的，已导致全球2000万至4000万人死亡。感染禽流感引起的人类疾病不仅限于呼吸道，而且似乎影响到其他器官，导致感染者预后不良。我们建议对即将感染高致病性禽流感病毒和与1918年流感基因重组的人类病毒的非人类灵长类动物进行深入的病理学研究，以全面描述感染对呼吸道外组织的影响，并旨在为有助于将感染后的人类伤亡降至最低的知识体系做出贡献。

DESCRIPTION (provided by applicant): Ongoing outbreaks of avian influenza in humans stress the need for intensified research efforts to fully characterize the pathogenesis during infection. Clinical course in patients and post-mortem pathology have suggested either presence of influenza virus in tissues outside of the respiratory tract, disease secondary to respiratory infection but in tissues that remain virus-free, or both. Animal studies have also demonstrated extra- pulmonary spread of avian influenza viruses, to an extent and with organ-involvement that appear species- dependent. Assessing the exact contribution of non-respiratory organ pathology to cytokine overproduction, a common feature of the disease in humans leading to multi-organ failure, is critical to successful clinical management, as is a better understanding of the factors leading to development of encephalopathy, also commonly seen. Unfortunately, timing of hospitalization after infection, confounding effects of heroic therapeutic efforts, and ethical considerations precluding biopsy of major organs in already severely compromised individuals, necessitate the use of animal models to study early infection in lungs and other major organs likely affected by the infection. Over the past 3 years, we have successfully optimized a macaque model of human influenza infection for genomic applications. For this purpose, we have used a mildly pathogenic human influenza virus (A/Texas/36/91) to carefully characterize the clinical course, pathology, and transcriptional regulation in response to viral mRNA, viral proteins, or to active viral replication. Next, we will infect Cynomolgus macaques with Texas influenza virus containing up to three genes from the 1918 pandemic flu (A/Brevig Mission/1/18), believed to have originated from an avian virus, and with highly pathogenic avian influenza (A/A/Vietnam/1203/04). In addition to the work performed in previous years, we propose to do an in-depth characterization of any pathology in tissues other than the respiratory tract, through the use of systematic microscopic examination, influenza antigen staining, phenotyping of immune cells in situ, and immunohistochemistry staining for markers of inflammatory and immune processes, including the cyclooxygenase cascade, production of nitric oxide by macrophages, induction of hypoxia response elements, angiogenesis, and others. This approach will be supplemented and corroborated by laser capture microdissection of lesions followed by quantitative reverse transcription PCR for corresponding mRNAs. We believe that the combination of several years of direct experience with this model and the use of relevant influenza viruses, known to be or likely to be of high virulence in humans, provides a unique opportunity to elucidate this important aspect of avian influenza infection in humans. Bielefeldt Ohmann - R03 NARRATIVE Outbreaks of avian influenza in humans have caused increasingly high morbidity and mortality. Further adaptation of the virus to the human host is likely to result in a world-wide epidemic such as the 1918 or "Spanish" influenza, believed to have been of avian origin and to have caused 20 to 40 millions deaths worldwide. Human disease caused by infection with avian influenza is not limited to the respiratory tract, but appears to affect other organs in ways that result in poor prognosis in infected individuals. We propose to perform in-depth pathology work in non-human primates that are to be infected with a highly pathogenic avian virus and with a human virus recombined with genes from the 1918 influenza, in order to fully characterize the effect of infection on tissues outside of the respiratory tract and with the goal of contributing to the body of knowledge that will help minimize human casualties after infection.