Role of Macrophages in CBD mediated attenuation of SEB-induced ARDS

项目摘要/摘要 这种名为SARS-CoV-2的病毒已经引发了新冠肺炎，并夺走了超过24万美国人和 全球有1270,000人。严重病例可导致急性呼吸窘迫 综合征(ARDS)，脓毒症，可因肺部炎症和破坏而致命 上皮细胞和内皮细胞衬里。了解这些疾病背后的机制对于 制定有效的预防和治疗策略。葡萄球菌肠毒素B(SEB)诱导 ARDS模拟重症患者出现的细胞因子风暴、脓毒症和多器官衰竭 新冠肺炎。已经证明，与尖峰相关的超抗原结构和序列 SARS-CoV-2的蛋白质与SEB相似。这种SEB诱导的ARDS模型也导致了 不同遗传背景的小鼠的疾病严重程度的不同表现，COVID- 19个在人类身上。用SEB处理C3H/HeJ小鼠，其存活率降至0%。在我们的研究中， 我们发现，在SEB治疗后给予大麻二醇(CBD)，可导致100%的存活率 无限期的。比较幼稚和幼稚肺的全单细胞测序数据的初步评估 SEB诱导的ARDS小鼠显示中性粒细胞增加，炎性 巨噬细胞和促炎细胞因子(IL-1β和肿瘤坏死因子-α)以及肺上皮细胞的丢失 细胞。探讨CBD治疗改善炎症反应的机制 响应，进行microRNA表达分析，显示表达显著降低 SEB治疗组miR-124-3p的表达与肿瘤坏死因子-α和IL-1β的上调有直接关系 表达以及巨噬细胞激活基因CEBP。我们假设CBD会减弱SEB- 肺浸润性细胞miRNA异常诱导ARDS，尤其是诱导miR-124-3p 它下调CEBP的表达，导致巨噬细胞的激活减少。目标1将 阐明驻留巨噬细胞和单核细胞来源的巨噬细胞在疾病中的作用以及CBD对 那些亚群。目标2将阐明CBD是否影响CEBP的表达，以及 MIR-124-3P无疾病表现。目标3将确定表观遗传因素调节 MiR-124-3p的表达。本研究将探索CBD作为ARDS和/或 败血症不仅是由SEB引起的，还包括其他病原体，如SARS-CoV-2。K99R00将提供 与职业发展和培训有关的机会--组学方法和生物统计学。 总的来说，我的导师、顾问委员会、顾问和研究环境 南卡罗来纳大学将帮助我成功地过渡到一名独立调查员。

PROJECT SUMMARY/ABSTRACT The virus, SARS-CoV-2 has caused COVID-19 and claimed the lives of over 240,000 Americans and 1,270,000 people worldwide. Severe cases of the disease leads to Acute Respiratory Distress Syndrome (ARDS), sepsis and can be fatal due to pulmonary inflammation and destruction of the epithelial and endothelial cell lining. Understanding the mechanisms behind these diseases is vital to develop effective preventive and therapeutic strategies. Staphylococcus enterotoxin B (SEB)-induced ARDS mimics the cytokine storm, sepsis and multiple organ failure presented in patients with severe COVID-19. It has been shown that the superantigen structure and sequence associated with the spike protein of the SARS-CoV-2 is similar to that of SEB. This SEB-induced ARDS model also results in various presentations of severity of illness in mice of different genetic backgrounds, as does COVID- 19 in humans. When C3H/HeJ mice are treated with SEB, their survival rate drops to 0%. In our study, we found that Cannabidiol (CBD) administration following SEB treatment, led to 100% survival indefinitely. Initial evaluation of whole single cell sequencing data comparing lungs from naïve with SEB-induced ARDS mice illustrated that there was an increase in neutrophils, inflammatory macrophages and pro-inflammatory cytokines (IL-1β and TNF-α) as well as a loss in lung epithelial cells. To characterize the mechanism by which CBD treatment led to amelioration of the inflammatory response, microRNA expression analysis was done that showed a significant decrease in expression of miR-124-3p in SEB-treated group which is directly associated with upregulation of TNF-α and IL-1β expression as well as macrophage activation gene, Cebp. We hypothesized that CBD attenuates SEB- induced ARDS by miRNA dysregulation in lung-infiltrating cells, specifically by inducing miR-124-3p which downregulates Cebp expression resulting in reduced activation of macrophages. Aim 1 will elucidate the role of resident and monocyte-derived macrophages in disease and the effect CBD on those subpopulations. Aim 2 will elucidate whether CBD affects Cebp expression and the effects that miR-124-3p has on manifestation of disease. Aim 3 will determine the epigenetic factors regulating expression of miR-124-3p. This study will explore CBD as a potential therapeutic for ARDS and/or sepsis induced not only by SEB but other pathogens such as SARS-CoV-2. The K99R00 will provide opportunities associated with Career Development and training in –omics approaches and biostatistics. Taken together, my mentors, advisory committee, consultant, and research environment at the University of South Carolina will nurture my successful transition to an Independent Investigator.