DataSheet1_Enteric Coronavirus Infection and Treatment Modeled With an Immunocompetent Human Intestine-On-A-Chip.PDF

Many patients infected with coronaviruses, such as SARS-CoV-2 and NL63 that use ACE2 receptors to infect cells, exhibit gastrointestinal symptoms and viral proteins are found in the human gastrointestinal tract, yet little is known about the inflammatory and pathological effects of coronavirus infection on the human intestine. Here, we used a human intestine-on-a-chip (Intestine Chip) microfluidic culture device lined by patient organoid-derived intestinal epithelium interfaced with human vascular endothelium to study host cellular and inflammatory responses to infection with NL63 coronavirus. These organoid-derived intestinal epithelial cells dramatically increased their ACE2 protein levels when cultured under flow in the presence of peristalsis-like mechanical deformations in the Intestine Chips compared to when cultured statically as organoids or in Transwell inserts. Infection of the intestinal epithelium with NL63 on-chip led to inflammation of the endothelium as demonstrated by loss of barrier function, increased cytokine production, and recruitment of circulating peripheral blood mononuclear cells (PBMCs). Treatment of NL63 infected chips with the approved protease inhibitor drug, nafamostat, inhibited viral entry and resulted in a reduction in both viral load and cytokine secretion, whereas remdesivir, one of the few drugs approved for COVID19 patients, was not found to be effective and it also was toxic to the endothelium. This model of intestinal infection was also used to test the effects of other drugs that have been proposed for potential repurposing against SARS-CoV-2. Taken together, these data suggest that the human Intestine Chip might be useful as a human preclinical model for studying coronavirus related pathology as well as for testing of potential anti-viral or anti-inflammatory therapeutics.

诸多感染冠状病毒的患者（如利用血管紧张素转换酶2（ACE2）受体侵染细胞的SARS-CoV-2与NL63）会表现出胃肠道症状，且人体胃肠道中可检测到病毒蛋白，但目前学界对冠状病毒侵染人体肠道所引发的炎症与病理效应仍知之甚少。本研究采用搭载患者类器官源肠道上皮、并与人体血管内皮细胞相接的微流控培养装置——人体肠道芯片（human intestine-on-a-chip，Intestine Chip），以探究宿主细胞及炎症应答针对NL63冠状病毒侵染的反应。与静态培养的类器官或Transwell小室培养的细胞相比，在肠道芯片中于流体环境及类似肠蠕动的机械形变条件下培养时，这类类器官源肠道上皮细胞的ACE2蛋白表达水平会显著升高。本研究中，肠道芯片内的肠道上皮被NL63侵染后，会引发内皮炎症，具体表现为屏障功能丧失、细胞因子生成增加以及循环外周血单个核细胞（peripheral blood mononuclear cells, PBMCs）的招募浸润。使用获批的蛋白酶抑制剂药物萘莫司他（nafamostat）处理NL63侵染的肠道芯片，可抑制病毒侵入，并同时降低病毒载量与细胞因子分泌；而瑞德西韦（remdesivir）——少数获批用于COVID-19患者的药物之一——不仅未展现出抗病毒效果，还会对血管内皮产生毒性。本研究还利用该肠道感染模型，测试了其他被提议用于重新适配治疗SARS-CoV-2感染的药物的效果。综合来看，上述研究结果表明，人体肠道芯片可作为研究冠状病毒相关病理的人类临床前模型，同时也可用于筛选潜在的抗病毒或抗炎治疗药物。