Radioresistant Innate Immunity in SAVI Tissue-Specific Autoinflammation

CGAS/STING胞质DNA传感通路检测微生物DNA，在宿主防御中发挥关键作用。 越来越多的证据表明，积累在细胞质中的自身DNA也参与cGAs的激发 发炎。此外，导致STIN结构性激活的一系列功能获得突变 现在都与一种称为SAVI(刺痛相关的)的衰弱自体炎症疾病有关 血管病变，起病于婴儿期)。这些患者患有严重的脉管炎损害和进展性 间质性肺疾病(ILD)，并经常屈从于与纤维化相关的治疗耐药的ILD。类似 对于与其他炎症性或纤维性肺部疾病相关的肺部疾病，人们对 这些患者的致病机制。因此，更好地了解导致这些问题的机制 为了开发更有效的治疗方法，需要有适当的条件。我们最近开发了一种小鼠模型 对于最常见的SAVI突变STINGV154M(Vm)，并表明这些小鼠重现了人类 疾病由多种标准决定，包括发展为炎症性/纤维性肺病。这些虚拟小鼠 为探索细胞类型和分子机制提供了一种独特的实验工具 纤维性肺病的发生和发展。Rag1缺陷完全拯救双肺VM小鼠 炎症和肺纤维化提示获得性免疫在VM ILD中起重要作用。刺痛的作用 一直专注于髓系细胞和VM小鼠具有扩大和激活的髓系/中性粒细胞 车厢。然而，辐射嵌合体研究已经确定了辐射抵抗细胞在肺纤维化中的关键作用。 用WT骨髓干细胞重建的VM小鼠在受到致死照射后出现极严重的肺损伤 即使这些嵌合体完全缺乏任何VM来源的造血细胞。初步数据 提示淋巴管内皮细胞(LECs)是Vm小鼠BALT形成的始动细胞。这样做的目的是 应用是进一步探索辐射抵抗的先天性免疫细胞直接或间接激活 Vm突变并确定这些细胞促进肺部炎症的机制。我们的方法将 涉及：(1)一种新的VM条件性KI细胞系与内皮限制性Cre缺失器交叉的体内分析 菌株和其他相关对照，(2)间质种群的scRNAseq比较，包括淋巴和 WT和Vm小鼠血管内皮细胞分离；(3)Vm和WT小鼠血管内皮细胞分析 由Vm突变或更传统的刺痛激动剂体外激活的小鼠。有一件急事 需要为患有自身免疫性和自体炎症性肺疾病的患者确定更好的治疗方法 本申请中提出的研究应该提供关键的见解，使我们能够设计出最 相关治疗靶点。此外，这些研究为研究刺痛的影响提供了机会。 更广泛地说，对基质细胞类型的激活是一个新兴的研究领域，如内皮细胞中的刺痛活性 细胞最近被认为与严重的COVID19的发展有关。

The cGas/STING cytosolic DNA sensing pathway detects microbial DNA and plays a critical role in host defense. Growing evidence indicates that self-DNA that accumulates in the cytosol also engages cGAS to incite inflammation. In addition, a series of gain-of-function mutations that result in constitutive activation of STING have now been associated with a debilitating auto-inflammatory disease known as SAVI (STING-Associated- Vasculopathy with onset in Infancy). These patients suffer from severe vasculitic lesions and progressive interstitial lung disease (ILD), and frequently succumb to treatment-resistant ILD-associated with fibrosis. Similar to the lung disorders associated with other inflammatory or fibrotic lung diseases, very little is known about the pathogenic mechanisms in these patients. Thus, a better understanding of the mechanisms responsible for these conditions is needed in order to develop more effective therapies. We have recently developed a murine model for the most common SAVI mutation, STINGV154M (VM), and shown that these mice recapitulate the human disease by a variety of criteria, including the development of inflammatory/fibrotic lung disease. These VM mice provide a unique experimental tool for exploring the cell types and molecular mechanisms responsible for the initiation and progression of fibrotic lung disease. Rag1-deficiency completely rescues VM mice from both lung inflammation and lung fibrosis pointing to a critical role of the adaptive immunity in VM ILD. The role of STING has been focused on myeloid cells and VM mice have an expanded and activated myeloid/neutrophil compartment. However, radiation chimera studies have identified a key role for radioresistant cells in lung fibrosis as lethally irradiated VM mice reconstituted with WT bone marrow stem cells develop extremely severe lung disease even though these chimeras completely lack any VM-derived hematopoietic cells. Preliminary data implicate lymphatic endothelial cells (LECs) as initiators of BALT formation in VM mice. The goal of this application is to further explore the radioresistant innate immune cells that are directly or indirectly activated by the VM mutation and define the mechanisms by which these cells promote lung inflammation. Our approach will involve: (1) the in vivo analysis of a novel VM conditional KI line crossed to endothelial-restricted Cre deleter strains and other relevant controls, (2) scRNAseq comparison of stromal populations including lymphatic and vascular endothelial cells isolated from WT and VM mice; and (3) analysis of endothelial cells from VM and WT mice following in vitro activation by the VM mutation or more conventional STING agonists. There is an urgent need to identify better therapies for patients afflicted with autoimmune and autoinflammatory lung disorders and the studies proposed in this application should provide critical insights that will enable us to design the most relevant therapeutic targets. Further, these studies provide an opportunity to study the impact of STING activation on stromal cell types more generally, an emerging area of research as STING activity in endothelial cells has recently been linked to the development of severe COVID19.