Alcohol and the alveolar epithelial barrier

项目摘要 新冠肺炎疫情代表着几代人以来发生的最严重的公共卫生危机。一位少校 新冠肺炎的死因是急性呼吸窘迫综合征。是否长期饮酒 障碍加剧新冠肺炎的严重程度尚不清楚，但考虑到已知的影响，很有可能 酒精对几个器官系统的影响，包括启动肺上皮屏障以增加细胞旁渗漏。在……里面 此外，众所周知，长期饮酒在以下情况下会加剧肺损伤的严重程度 再加上额外的侮辱，就是所谓的“二次打击”。值得注意的是，酒精肺综合征会增加 ARDS的发生率是非酒精性ICU患者的3-4倍。我们已经确定了财产 与病毒E蛋白有关的SARS-CoV-2病毒，使其易于进入较温和的感染阶段。这个 最初的SARS冠状病毒SARS-CoV-1有一个E蛋白，专门针对紧密连接蛋白PDZ 结合基序，因此很容易破坏受感染的上皮细胞的上皮屏障。相比之下，新冠肺炎 SARS-CoV-2病毒的E蛋白突变抑制了其与PDZ结合基序的结合能力，这有助于 保护受感染的上皮细胞屏障功能。然而，在酒精肺的背景下，SARS-CoV-2 综合征更有可能导致更严重的后果，因为上皮屏障已经作为一种 是长期酗酒的结果。原代培养的人细支气管上皮细胞来源的酒精和非酒精 酒精受试者将在体外感染SARS-CoV-2，我们将测量几个结果变量 与感染的严重程度有关。我们也有初步数据显示酗酒者的细胞 与非酒精细胞相比，基因表达和功能的标志性差异导致减少 屏障功能和细胞形态改变。这支持了一种假设，即长期接触酒精会导致 呼吸道上皮细胞基因组的持续性表观遗传学改变。在这份补充提案中，我们将 确定SARS-CoV-2对酗酒和非酗酒者宿主细胞表观基因组的影响， 专注于DNA甲基化。特别有兴趣的将是检查DNA甲基化诱导的影响 酒精和/或SARS-CoV-2感染对转录因子表达及下游靶点的影响 Nrf2和PU.1，已知因长期酒精暴露而受损，并在调节 上皮性抗病毒反应。我们还将确定显示差异模式的新宿主基因组基因座 与疾病严重程度相关的甲基化。我们预计识别差异甲基化的基因座 受长期酒精暴露和SARS-CoV-2感染的影响，将导致药物靶点有可能 降低酒精中毒患者新冠肺炎的严重程度。

Project Summary The COVID-19 pandemic represents the most significant public health crisis to occur in generations. A major cause of death due to COVID-19 is acute respiratory distress syndrome (ARDS). Whether chronic alcohol use disorder exacerbates the severity of COVID-19 is not known, but is highly likely given the known impact of alcohol on several organ systems including priming the lung epithelial barrier for increased paracellular leak. In addition, it is well established that chronic alcohol consumption exacerbates the severity of lung injury when combined with an additional insult, a so-called “second hit”. Of note, alcoholic lung syndrome increases the incidence of ARDS by 3-4 fold when compared with non-alcoholic ICU patients. We have identified properties of SARS-CoV-2 that predispose it towards a milder phase of infection, related to the virus E protein. The original SARS coronavirus, SARS-CoV-1, has an E protein that specifically targets tight junction protein PDZ binding motifs and thus readily disrupts the epithelial barrier of infected epithelia. By contrast, the COVID-19 virus SARS-CoV-2 has E protein mutations that inhibit its ability to bind to PDZ binding motifs which can help preserve infected epithelial cell barrier function. However, SARS-CoV-2 in the context of alcoholic lung syndrome is more likely to result in a more severe outcome, since the epithelial barrier is already impaired as a result of chronic alcohol abuse. Primary human bronchiolar epithelial cells derived from alcoholic and non- alcoholic subjects will be infected with SARS-CoV-2 in vitro and we will measure several outcome variables related to severity of infection. We also have preliminary data showing that cells from alcoholics retain signature differences in gene expression and function relative to non-alcoholic cells, leading to decreased barrier function and altered cell morphology. This supports a hypothesis that chronic alcohol exposure induces persistent, epigenetic changes to the airway epithelial cell genome. In this supplement proposal, we will determine the impact of SARS-CoV-2 on the epigenome of host cells from alcoholics and non-alcoholics, focusing on DNA methylation. Of particular interest will be to examine the impact of DNA methylation induced by alcohol and/or SARS-CoV-2 infection on the expression and downstream targets of the transcription factors Nrf2 and PU.1, known to be impaired by chronic alcohol exposure and that play key roles in regulating epithelial antiviral responses. We will also identify novel host genome loci showing patterns of differential methylation that correlate with disease severity. We anticipate that identifying differentially methylated loci impacted by chronic alcohol exposure and SARS-CoV-2 infection will lead to drug targets with the potential to reduce the severity of COVID-19 in alcoholic patients.