Long Term Culture of Human Kidney Proximal Tubule Epithelial Cells Maintains Lineage Functions and Serves as anEx vivoModel for Coronavirus Associated Kidney Injury
Long Term Culture of Human Kidney Proximal Tubule Epithelial Cells Maintains Lineage Functions and Serves as anEx vivoModel for Coronavirus Associated Kidney Injury
复制标题
人肾近端小管上皮细胞的长期培养维持谱系功能并作为冠状病毒相关肾损伤的离体模型
DOI：
10.1007/s12250-020-00253-y
复制
发表时间：
2020-06-29
影响因子：
5.5
通讯作者：
Li, Hui
中科院分区：
文献类型：
--
作者：
Xia, Siyu;Wu, Ming;Li, Hui
SARS-CoV-2如何导致严重的多器官衰竭的机制在很大程度上是未知的。急性肾损伤（阿基）是重症COVID-19患者常见的器官损伤之一。已有研究表明，人肾小管细胞可能是SARS-CoV-2潜在的靶细胞。传统的癌细胞系或永生化细胞系在遗传和表型上与宿主细胞不同。动物模型被广泛应用，但由于种属嗜性，往往不能反映生理和致病状态。对于用于疾病建模的正常人上皮细胞存在未满足的需求。在这项研究中，我们成功地建立了正常人肾近端小管上皮细胞（KPTECs）的长期培养在2D和3D培养系统中使用条件重编程（CR）和类器官技术。这些细胞具有分化和修复DNA损伤的能力，并且不显示转化性质。重要的是，CR KPTEC通过表达特异性转运蛋白（SLC 34 A3和cubilin）维持谱系功能。它们还表达SARS-CoV和SARS-CoV-2的受体血管紧张素转换酶2（ACE 2）。相反，癌细胞系不表达内源性SLC 34 A3、cubilin和ACE 2。非常有趣的是，ACE 2表达在3D类器官培养中比在2D CR培养条件下高约两倍。假病毒体实验证明SARS-CoV S蛋白能够携带荧光素酶报告基因进入CR细胞。这种集成的2D CR和3D类器官培养物提供了一个生理学的活体模型来研究肾功能，肾细胞对病毒的先天免疫反应，以及药物发现和安全性评价的新平台。
The mechanism of how SARS-CoV-2 causes severe multi-organ failure is largely unknown. Acute kidney injury (AKI) is one of the frequent organ damage in severe COVID-19 patients. Previous studies have shown that human renal tubule cells could be the potential host cells targeted by SARS-CoV-2. Traditional cancer cell lines or immortalized cell lines are genetically and phenotypically different from host cells. Animal models are widely used, but often fail to reflect a physiological and pathogenic status because of species tropisms. There is an unmet need for normal human epithelial cells for disease modeling. In this study, we successfully established long term cultures of normal human kidney proximal tubule epithelial cells (KPTECs) in 2D and 3D culture systems using conditional reprogramming (CR) and organoids techniques. These cells had the ability to differentiate and repair DNA damage, and showed no transforming property. Importantly, the CR KPTECs maintained lineage function with expression of specific transporters (SLC34A3 and cubilin). They also expressed angiotensin-converting enzyme 2 (ACE2), a receptor for SARS-CoV and SARS-CoV-2. In contrast, cancer cell line did not express endogenous SLC34A3, cubilin and ACE2. Very interestingly, ACE2 expression was around twofold higher in 3D organoids culture compared to that in 2D CR culture condition. Pseudovirion assays demonstrated that SARS-CoV spike (S) protein was able to enter CR cells with luciferase reporter. This integrated 2D CR and 3D organoid cultures provide a physiologicalex vivomodel to study kidney functions, innate immune response of kidney cells to viruses, and a novel platform for drug discovery and safety evaluation.