Investigating the link between pericyte dysfunction and loss of glucose homeostasis in COVID-19

摘要 冠状病毒病2019年(新冠肺炎)是一种全球性的医疗危机，在美国每1人死亡约1人 等一下。它是由严重急性呼吸综合征冠状病毒-2(SARS-CoV-2)引起的。一边呼吸一边呼吸 衰竭仍然是新冠肺炎患者最常见的死亡原因，严重的表现随处可见 多个器官，包括胰腺。新冠肺炎和胰腺之间存在着错综复杂的关系 胰腺炎和糖尿病等疾病。了解导致糖尿病的病理生理学机制 感染患者的胰腺功能障碍和葡萄糖不耐受是最重要的。发病机制 糖尿病的发生与胰岛功能障碍密切相关。有很多研究 表明SARS-CoV-2病毒直接攻击血管系统。然而，胰岛微血管 功能障碍在新冠肺炎患者中引发血糖内稳态的丧失尚未被探讨。这个项目 重点关注血管周细胞，因为我们公布的初步数据表明：(A)人类中的周细胞 胰岛对血管紧张素II非常敏感，并且表达SARS-CoV-2的关键宿主细胞受体ACE2； (B)周细胞可感染SARS-CoV-2假进入病毒；及。(C)与SARS-CoV-2孵育。 SPEKE重组蛋白可提高周细胞基础钙水平。因此，我们假设SARS-CoV-2 感染胰腺周细胞，干扰其收缩特性，损害其功能。周细胞 功能障碍会减少局部血流量，导致组织缺氧和炎症，并损害 内分泌细胞活动。我们将使用来自人类和仓鼠的活胰腺切片来验证这一假设 鉴于它们的血管紧张素转换酶2蛋白序列高度同源性，它们是适合于新冠肺炎研究的动物模型。 这一假设将在两个方面得到验证：1)确定周细胞是否是SARS-CoV-2在 检测SARS-CoV-2对胰岛周细胞功能和微血管反应的影响 体外和体内。在目标1中，我们将寻找SARS-CoV-2病毒颗粒，描述微血管病变 新冠肺炎患者胰腺周细胞表型。我们会感染活的胰腺切片 SARS-CoV-2假侵入和活病毒，并确定周细胞是否允许感染。在目标2中，我们 将通过与ACE2的交互来评估操纵ACE2表达和活动的功能后果 SARS-CoV-2刺突蛋白对活体胰腺切片中周细胞钙反应和血管运动的影响 通过在仓鼠胰腺的导管内注射假进入病毒而获得活体。我们将纵向跟踪 对糖代谢的影响，以及局部缺氧、炎症、内分泌细胞质量和 功能。这项研究有可能确定胰腺内分泌功能障碍的机制。如果 假设是正确的，周细胞是这种冠状病毒的细胞靶标。他们的功能障碍可以解释 为什么新冠肺炎与血糖动态平衡的丧失和其他胰腺疾病有关。我们期待我们的 对新冠肺炎相关糖尿病的研究，以揭示该疾病自然病史的新机制。

ABSTRACT The coronavirus disease 2019 (COVID-19) is a global healthcare crisis that in the USA kills about 1 person every minute. It is caused by the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2). While respiratory failure is still the most common cause of mortality in COVID-19 patients, serious manifestations are seen across multiple organs, including the pancreas. There is an intricate relationship between COVID-19 and pancreatic diseases such as pancreatitis and diabetes. Understanding the pathophysiological mechanisms that lead to pancreatic dysfunction and glucose intolerance in infected patients is of utmost importance. The pathogenesis of diabetes is intimately associated with the dysfunction of the pancreatic islet. There are numerous studies indicating that SARS-CoV-2 directly attacks the vascular system. However, the possibility that islet microvascular dysfunction triggers loss of glucose homeostasis in COVID-19 patients has not been explored. This project focuses on the vascular pericyte because our published and preliminary data show that (a) pericytes in human islets are very responsive to angiotensin II and express the key host cellular receptor of SARS-CoV-2, ACE2; (b) pericytes can be infected with SARS-CoV-2 pseudo-entry viruses; and (c) incubation with a SARS-CoV-2 Spike recombinant protein increases pericyte basal Ca2+ levels. We therefore hypothesize that SARS-CoV-2 infects pancreas pericytes, interfering with their contractile properties and impairing their function. Pericyte dysfunction will decrease local blood flow, leading to tissue hypoxia and inflammation and compromising endocrine cell activity. We will test this hypothesis using living pancreas slices from humans and hamsters which are a suitable animal model for COVID-19 research given the high homology of their ACE2 protein sequences. This hypothesis will be tested in two Aims: 1) determine if pericytes are cellular targets of SARS-CoV-2 in the pancreas, and (2) examine the effects of SARS-CoV-2 on islet pericyte function and microvascular responses ex vivo and in vivo. In Aim 1, we will search for SARS-CoV-2 viral particles, characterize microvascular lesions and the pericyte phenotype in the pancreas of COVID-19 patients. We will infect living pancreas slices with SARS-CoV-2 pseudo-entry and live viruses and determine if pericytes are permissive for infection. In Aim 2, we will assess the functional consequences of manipulating ACE2 expression and activity through interactions with SARS-CoV-2 spike protein on pericyte Ca2+ responses and vasomotion ex vivo in living pancreas slices, and in vivo by intraductal injections of pseudo-entry viruses in the hamster pancreas. We will follow longitudinally the effects on glucose metabolism, as well as changes in local hypoxia, inflammation, endocrine cell mass and function. This research has the potential to identify a mechanism of dysfunction in the endocrine pancreas. If the hypothesis is correct, pericytes are cellular targets of this coronavirus. Their functional impairment could explain why COVID-19 is linked to a loss of glucose homeostasis and other pancreatic disorders. We anticipate our study of COVID-19–related diabetes to uncover novel mechanisms of the natural history of this disease.