Human Intestinal Enteroids as Ex Vivo Models of Human Rotavirus Infection

项目总结 轮状病毒(RV)是5岁以下儿童中威胁生命的腹泻疾病的主要原因，以及 每年导致近50万人死亡，尽管使用了两种获得许可的轮状病毒疫苗。虽然房车 疫苗是可用的，但在最需要它们的国家，它们并不能以最佳方式发挥作用。人类 轮状病毒(HRV)在培养的细胞中生长不良，不会感染只对动物敏感的小鼠 轮状病毒感染。我们不了解HRV感染是如何导致危及生命的疾病的，并质疑 现有动物模型的知识完全适用于HRV。我们率先建立了人类 肠类肠道(HIEs)作为研究宿主-病原体相互作用的新型体外模型“迷你肠道”系统 和病理生理学。HIE是自组织的、三维生理活性的多细胞培养物， 模仿肠道的有序结构。我们已经发现HRV毒株在HIE中复制得非常好 模仿在人体内观察到的宿主迁移性，从而建立一个研究HRV感染的新模型 和发病机制。我们建议进行研究，直接检查病理生理学和遗传学机制。 控制HRV感染和生长。最近发现，HRV毒株使用的细胞受体是 组织血液组抗原(HBGA)导致了对宿主遗传影响的思维范式的转变 为轮状病毒的人畜传播提供了一个新的视角。独特的区别在于 HBGA的表达可能解释了非洲异常HRV毒株的高流行率和低疫苗 在不同的种族人群中看到的效果。我们假设肠道类的HRV感染是遗传的。 限制性和允许性感染导致新的细胞内和细胞外固有信号和反应 心率变异性发病机制和病理生理学的基础途径。此外，HRV感染或 疫苗接种可以通过有益的微生物进行调节。我们建议在目标1中进行研究，以确定 HIE基因控制的易感性调节因子(HBGA)和独特的先天反应调节HRV 感染和病理生理学(核心B和C)。目标2的研究将确立共生或共生的影响 病原体对HRV或疫苗感染的影响。这些研究将阐明菌株的分子基础- 在不同种族的儿童中观察到的特定宿主范围限制，并与理解相关 疫苗特性。此外，对含有人类共生体的改良HIE培养物的评估， 新的定制生物材料平台中的益生菌或其他病原体(具有核心B、C和项目2) 项目3)将有助于更好地理解上皮细胞对病毒感染的反应是如何调节的。 总的来说，这项工作将对轮状病毒的病理生理学和随后的 触发免疫和疾病的人类关键适应性反应。

PROJECT SUMMARY Rotavirus (RV) is the leading cause of life-threatening diarrheal disease in children under the age of 5, and results in nearly half a million deaths annually, despite the use of two licensed RV vaccines. Although RV vaccines are available, they do not work optimally in countries where they are needed most. Human rotaviruses (HRVs) grow poorly in cultured cells and do not infect mice, which are only susceptible to animal rotavirus infection. We do not understand how HRV infections cause life-threatening disease and question if knowledge from existing animal models is fully applicable to HRVs. We pioneered establishing human intestinal enteroids (HIEs) as novel ex vivo model “mini-gut” systems to examine host-pathogen interactions and pathophysiology. HIEs are self-organizing, 3-dimensional physiologically active, multicellular cultures that mimic the ordered structure of the intestine. We have found that HRV strains replicate extremely well in HIEs mimicking the host permissiveness observed in people, thus establishing a new model to study HRV infections and pathogenesis. We propose studies to directly examine mechanisms of pathophysiology and genetic control of HRV infection and growth. The recent discovery that cellular receptors used by HRV strains are histoblood group antigens (HBGAs) resulted in a paradigm shift in thinking about the host's genetic influence on RV infection and provides a new perspective on zoonotic transmission of rotaviruses. Unique differences in HBGA expression may explain the higher prevalence of unusual HRV strains in Africa and lower vaccine efficacy seen in distinct ethnic populations. We hypothesize that HRV infection of enteroids is genetically restricted and permissive infection leads to novel intracellular and extracellular innate signaling and response pathways that underlie HRV pathogenesis and pathophysiology. In addition, the outcome of HRV infection or vaccination can be modulated by beneficial microbes. We propose studies in Aim 1 to determine whether genetically-controlled modulators of susceptibility (HBGAs) and distinct innate responses in HIEs regulate HRV infection and pathophysiology (with Cores B and C). Studies in Aim 2 will establish the effect of commensal or pathogenic organisms on HRV or vaccine infection. These studies will elucidate the molecular basis for strain- specific host range restriction observed in children of different ethnic groups and be relevant to understanding vaccine properties. Furthermore, evaluation of modified HIE cultures that house human commensals, probiotics or other pathogens (with Cores B, C and Project 2) in new tailored biomaterial platforms (with Project 3) will lead to a better understanding of how epithelial responses to a viral infection are modulated. Collectively, this work will provide a new understanding of rotavirus pathophysiology and the subsequent critical adaptive responses in humans that trigger immunity and disease.