Enteric coronavirus infection and treatment modeled with an immunocompetent human intestine-on-a-chip

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 (PMBCs). 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. 4 uninfected organ-on-chips (2 donors; 2 chips/donor) and 2 infected chips (1 donor). Replicate Seq-Well arrays of all chips aside from 2 uninfected chips (1 array only).

众多感染冠状病毒的患者——例如借助血管紧张素转换酶2（ACE2）受体感染宿主细胞的严重急性呼吸综合征冠状病毒2（SARS-CoV-2）与NL63冠状病毒——均会表现出胃肠道症状，且人体胃肠道中可检测到病毒蛋白；但目前学界对于冠状病毒感染引发的人体肠道炎症与病理损伤机制仍知之甚少。本研究采用由患者类器官来源肠上皮与人体血管内皮共同构建的人体肠道芯片（Intestine Chip）微流控培养装置，探究NL63冠状病毒感染后的宿主细胞应答与炎症反应。相较于静态培养的类器官或Transwell插入式培养体系，在肠道芯片中于流体剪切力与蠕动样机械形变条件下培养的类器官来源肠上皮细胞，其ACE2蛋白表达水平会显著升高。在芯片模型中用NL63感染肠上皮细胞，可引发内皮炎症，具体表现为屏障功能受损、细胞因子生成增加以及循环外周血单核细胞（PMBCs）的招募浸润。使用获批蛋白酶抑制剂萘莫司他（nafamostat）处理感染NL63的芯片，可抑制病毒入侵，并同时降低病毒载量与细胞因子分泌水平；而获批用于新型冠状病毒肺炎（COVID-19）患者的少数药物之一瑞德西韦（remdesivir）不仅未展现出抗病毒活性，还会对血管内皮产生毒性作用。本研究构建的肠道感染模型还被用于测试其他拟通过药物重定位开发以对抗SARS-CoV-2的药物的效应。综上，上述实验数据表明，人体肠道芯片可作为人体临床前模型，用于研究冠状病毒相关病理机制，以及筛选潜在的抗病毒或抗炎治疗药物。本研究共设置4个未感染器官芯片样本（2名供体，每名供体2个芯片）以及2个感染芯片样本（1名供体）。除2个未感染芯片外，其余所有芯片均进行了重复Seq-Well阵列检测（仅1组阵列）。