Radioresistant Innate Immunity in SAVI Tissue-Specific Autoinflammation

cGas/STING胞质DNA传感途径检测微生物DNA，并在宿主防御中起关键作用。 越来越多的证据表明，在胞质溶胶中积累的自身DNA也参与了cGAS的刺激， 炎症此外，一系列导致STING组成性激活的功能获得性突变， 现在已经与一种称为SAVI的衰弱性自身炎症性疾病（刺痛相关性- 婴儿期发作的血管病变）。这些患者患有严重的血管炎性病变和进行性 间质性肺病（ILD），且经常死于与纤维化相关的难治性ILD。类似 对于与其他炎症性或纤维化肺病相关的肺部疾病， 这些患者的发病机制。因此，更好地了解这些机制， 为了开发更有效的治疗方法，需要条件。我们最近开发了一种小鼠模型 最常见的SAVI突变STINGV 154 M（VM），并表明这些小鼠重现了人类 根据各种标准，包括炎性/纤维化肺病的发展，可以评估疾病的严重程度。这些VM小鼠 提供了一个独特的实验工具，探索细胞类型和分子机制，负责 纤维化肺病的发生和发展。Rag 1缺陷完全从双肺拯救VM小鼠 炎症和肺纤维化，指出适应性免疫在VM ILD中的关键作用。STING的作用 已经集中在骨髓细胞和VM小鼠具有扩增和活化的骨髓/中性粒细胞 车厢然而，放射嵌合体研究已经确定了放射抗性细胞在肺纤维化中的关键作用 当用WT骨髓干细胞重建的致死性辐照VM小鼠发生极其严重的肺损伤时， 尽管这些嵌合体完全缺乏任何VM衍生的造血细胞，但它们仍然可以预防疾病。初步数据 提示淋巴管内皮细胞（LECs）是VM小鼠BALT形成的启动子。这个目标 本申请的目的是进一步探索直接或间接被辐射激活的抗辐射先天免疫细胞， 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 包括：（1）与内皮限制性Cre删除器杂交的新型VM条件KI系的体内分析 菌株和其他相关对照，（2）包括淋巴和淋巴细胞的基质群体的scRNAseq比较， 从WT和VM小鼠分离的血管内皮细胞;和（3）分析来自VM和WT的内皮细胞 通过VM突变或更常规的STING激动剂体外活化后的小鼠。目前迫切 需要为患有自身免疫性和自身炎性肺病的患者确定更好的治疗方法， 在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 相关的治疗目标。此外，这些研究为研究STING的影响提供了机会。 更普遍地，在基质细胞类型上的活化，作为内皮细胞中的STING活性的新兴研究领域， 细胞最近与严重COVID的发展有关19。

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.