COVID-19 and lung disease: developing therapy through immune cell re-programming (Ref: 3949)

SARS-CoV-2呼吸道感染可导致轻度或重度疾病（COVID-19）。除了男性和高血压、糖尿病等合并症外，衰老是一个关键的风险因素。同样的风险也是其他严重、无法治愈和致命的肺部疾病，如肺纤维化（PF）和慢性阻塞性肺疾病（COPD）的关键。新出现的数据表明，COVID-19可导致肺瘢痕（即PF），患有PF的患者因COVID-19而死亡率更高。肺泡性肺病已占英国总死亡人数的1%，而慢性阻塞性肺病是全球第三大杀手。我们最近使用英国生物银行（超过500,000参与者的遗传，生物标志物和健康数据资源）进行的分析揭示了PF和COPD与免疫细胞数量和端粒磨损的有趣关联（接受在柳叶刀呼吸医学上发表）；我们正在埃克塞特-布里斯托尔联合患者队列中跟踪这些发现。免疫细胞库也是COVID-19的主要研究重点。巨噬细胞在防御病原体方面发挥着关键作用，但也可能促进疾病的发展——尤其是通过炎症细胞因子的过度产生（在PF和COVID-19中都可见到）——这与线粒体功能有关。线粒体是细胞活动的关键调节器，充当病毒感染的细胞内信号“枢纽”，并通过炎性体介导细胞对炎症的反应；它们也是sars冠状病毒蛋白的靶标。有效的免疫力随着年龄的增长而下降，导致免疫衰老和慢性低度炎症。我们最近获得的两项MRC奖金（总计150万英镑）主要集中在COPD和PF的线粒体方面。我们有令人兴奋的数据表明，Whiteman教授开发的新药物可以产生生理剂量的线粒体靶向硫化氢（H2S），可以改善实验系统中的促炎和促纤维化反应。线粒体是一种新的控制先天免疫活性的药物靶点。Lindsay教授最近的工作进一步关注了长链非编码RNA （lncRNA）在炎症反应调节中的作用，而Scotton博士小组的初步数据表明，在小鼠模型中，人类过继转移可加剧纤维化疾病。我们的假设是，线粒体功能障碍可以通过我们的新型H2S药物来改变细胞代谢和转录组，减少超激活，促进组织修复。这些过程将与PF和COVID-19的功能破坏有关。有3个目的：1)利用RNA-Seq、生物信息学和标准体外试验（如细胞因子ELISA、流式细胞术、趋化和吞噬），表征PF患者与年龄匹配（60岁）和年轻对照的单核细胞源性巨噬细胞的lncRNA/mRNA转录组和表型：a)炎症刺激，包括热灭活的SARS-CoV-2病毒颗粒和Spike蛋白，b)巨噬细胞极化，如M（LPS）、M（IL4）。2)探讨H2S药物补充对LPS、IL-4或SARS-CoV-2/Spike蛋白刺激下巨噬细胞功能和lncRNA水平的功能影响。诱导多能干细胞衍生的肺泡也将用于增加与肺的相关性。3)在PF的实验体内模型中，确定年轻人和老年人过继转移到免疫功能低下的NOD/SCID小鼠后的纤维化潜力，以及我们的H2S药物的治疗效果。最终，我们希望将这些新药从实验室应用到临床。方法:RNASeq;中存在;ELISA;表型/功能流式细胞术；海马细胞外通量分析；PF实验小鼠模型；体内成像系统（IVIS）跟踪荧光标记测量肺胶原积累和离体微计算机断层扫描

SARS-CoV-2 infection of the respiratory tract can lead to mild or severe disease (COVID-19).Ageing is a crucial risk factor, alongside male sex and co-morbidities such as hypertension and diabetes. The same risks are key in other serious, incurable and fatal lung diseases such as pulmonary fibrosis (PF) and chronic obstructive pulmonary disease (COPD). Emerging data suggest that COVID-19 can lead to lung scarring (i.e. PF), and patients who have PF have greater mortality from COVID-19. PF already accounts for 1% of all UK deaths, while COPD is the 3rd biggest killer globally.Our recent analysis using the UK Biobank (a resource of genetic, biomarker and health data for over 500,000 participants) has revealed intriguing associations with immune cell numbers and also telomere attrition in PF and COPD (accepted for publication in the Lancet Respiratory Medicine); we are following up some of these findings in our joint Exeter-Bristol patient cohorts. Immune cell repertoires are also a major research focus in COVID-19.Macrophages play a key role in defence in against pathogens, but can also contribute to disease development - not least via the overproduction of inflammatory cytokines (seen in both PF and COVID-19) - and this is linked to mitochondrial function. Mitochondria are key regulators of cellular activity, act as intracellular signalling 'hubs' for viral infection, and mediate the cellular response to inflammation through the inflammasome; they are also targets for SARS-CoV proteins.Effective immunity deteriorates with age, leading to immunosenescence and chronic low-grade inflammation. Our two recent MRC awards (totalling >£1.5m) focus on mitochondria in the context of COPD and PF. We have exciting data showing that novel drugs developed by Prof Whiteman, which generate physiological doses of mitochondrial-targeted hydrogen sulfide (H2S), can ameliorate pro-inflammatory and pro-fibrotic responses in experimental systems. mitochondria are a novel drug target to control innate immune activity.Recent work from Prof Lindsay has further focused on the role of long non-coding RNA (lncRNA) in the regulation of inflammatory responses in while preliminary data from Dr Scotton's group has demonstrated that adoptive transfer of human can exacerbate fibrotic disease in murine models. Our hypothesis is that mitochondrial dysfunction in can be modified by our novel H2S drugs to alter cellular metabolism and transcriptome, reducing hyper-activation and facilitating tissue repair. These processes will be relevant for disrupted function in both PF and COVID-19. There are 3 Aims:1) Characterise the lncRNA/mRNA transcriptome and phenotype of monocyte-derived macrophages from patients with PF versus age-matched (60 year old) and young controls using RNA-Seq, bioinformatics, and standard in vitro assays (e.g. cytokine ELISA, flow cytometry, chemotaxis, and phagocytosis) during the response to: a) inflammatory stimuli, including heat-inactivated SARS-CoV-2 viral particles and Spike protein, and b) macrophage polarisation e.g. M(LPS), M(IL4).2) Interrogate the functional consequence of H2S drug supplementation on macrophage function and lncRNA levels in human stimulation with LPS, IL-4 or SARS-CoV-2/Spike protein. Induced pluripotent stem cell-derived alveolar will also be used to increase relevance to the lung.3) Determine the fibrogenic potential of young vs-old human following adoptive transfer into immunocompromised NOD/SCID mice in an experimental in vivo model of PF, and the therapeutic benefit of our H2S drugs. Ultimately, we would like to translate these novel drugs from bench to bedside. Methodology: RNASeq; qRT-PCR; ELISA; flow cytometry of phenotype/function; Seahorse extracellular flux analysis; experimental mouse model of PF; In Vivo Imaging System (IVIS) tracking of fluorescently-tagged measurement of lung collagen accumulation and ex vivo micro- Computed Tomography