Restoring immune-vascular axis integrity to alleviate acute lung injury in sepsis

项目总结/摘要 脓毒症引起的多器官功能障碍综合征，主要影响肺和肾， 全世界发病率和死亡率原因。急性肺损伤（ALI）或其最严重的形式，急性呼吸道 急性呼吸窘迫综合征（ARDS）是脓毒症最常见的并发症，目前尚无有效的治疗方法 除了支持性治疗因此，减轻脓毒症诱导的ALI的新的治疗策略是 尤其是在当前的SARS-CoV-2大流行面前。 脓毒症是由宿主对感染的反应失调引起的危及生命的器官功能障碍， 其中免疫细胞和内皮细胞都起关键作用。病原体诱导的炎症级联反应和 内皮通过自分泌或旁分泌环相互影响以增加血管通透性， 导致免疫失调，破坏免疫血管稳态。因此，治疗旨在 需要恢复免疫-血管稳态以减轻脓毒症诱导的并发症如ALI/ARDS。 神经纤毛蛋白-1（NRP 1）与血管通透性和炎症有关，可能被证明是一种可行的 通过恢复免疫血管完整性治疗脓毒症诱导的ALI的靶点。因此， 我们认为NRP 1在脓毒症诱导的ALI的免疫血管功能障碍中起关键作用， 靶向NRP 1以恢复免疫血管稳态是治疗这种疾病的可行治疗选择。 为了验证我们的假设，我们提出了三个具体目标。在目标1中，我们将分析ALI的影响- 内皮特异性NRP 1基因敲除小鼠肺内皮细胞相关细胞因子的体外研究 小鼠我们还将通过高剂量脂多糖（LPS）治疗或盲肠给药在这些小鼠中建立ALI 结扎和穿刺（CLP），以评估内皮NRP 1在脓毒症诱导的ALI严重程度中的作用， 体内血管通透性、白细胞滚动和外渗。目的2探讨共培养对细胞增殖的影响 从骨髓特异性和CD 4 + T细胞特异性NRP 1-T细胞分离的外周血单核细胞（PBMC）， 用野生型小鼠肺内皮细胞敲除小鼠。此外，我们将在这些NRP 1- 敲除小鼠模型，以研究NRP 1在这两种免疫细胞群体中的多效性作用， 脓毒症引起的ALI。目的3评价内皮细胞靶向脂质体的治疗效果 小分子NRP 1抑制剂（EG 00229）制剂，改善脓毒症诱导的ALI的严重程度。 我们预计，我们的建议将提供更深入的了解NRP 1在免疫中的作用- 败血症诱导的ALI中的血管功能障碍，并导致开发一种可行的治疗策略， 靶向NRP 1以克服其严重性。我们的建议的最终目的是使大量的 患有脓毒症引起的肺部并发症的患者，迄今为止还没有有效的治疗方法。

PROJECT SUMMARY/ABSTRACT Sepsis-induced multi organ dysfunction syndrome, which primarily affects the lungs and kidneys, is a major cause of morbidity and mortality worldwide. Acute lung injury (ALI) or its most severe form, acute respiratory distress syndrome (ARDS), are the most common complications of sepsis, for which no effective treatment except supportive care is available. Hence, novel therapeutic strategies for mitigating sepsis-induced ALI are desperately needed, more so in the face of the current SARS-CoV-2 pandemic. Sepsis is a life-threatening organ dysfunction caused by a dysregulated host response to an infection, where both immune cells and endothelial cells play critical roles. Pathogen-induced inflammatory cascade and the endothelium impact each other via autocrine or paracrine loops to increase vascular permeability and cause immune dysregulation to disrupt immune-vascular homeostasis. Hence, therapies aimed towards reinstating immune-vascular homeostasis are needed to alleviate sepsis-induced complications like ALI/ARDS. Neuropilin-1 (NRP1), implicated in vascular permeability and inflammation, may prove to be a viable target for treating sepsis-induced ALI by restoring immune-vascular integrity. Hence, the central hypothesis of our proposal is that NRP1 plays a critical role in the immune-vascular dysfunction in sepsis-induced ALI and targeting NRP1 for restoring the immune-vascular homeostasis is a viable therapeutic option for this disease. To validate our hypothesis, we propose three specific aims. In Aim 1, we will analyze the effect of ALI- relevant cytokines in vitro in mouse lung endothelial cells isolated from endothelial-specific NRP1 knockout mice. We will also develop ALI in these mice by high dose lipopolysaccharides (LPS) treatment or by cecal ligation and puncture (CLP) to evaluate the role of endothelial NRP1 in the severity of sepsis-induced ALI, in vivo vascular permeability, leukocyte rolling and extravasation. Aim 2 will investigate the effect of coculturing peripheral blood mononuclear cells (PBMCs) isolated from myeloid-specific and CD4+T-cell specific NRP1- knockout mice with wild-type mouse lung endothelial cells. Moreover, we will develop ALI in these NRP1- knockout mice models to investigate the pleotropic role of NRP1 in these two immune cell populations during sepsis-induced ALI. Aim 3 will evaluate the therapeutic efficacy of an endothelial cell (EC)-targeting liposomal formulation of a small-molecule NRP1 inhibitor (EG00229), in ameliorating the severity of sepsis-induced ALI. We anticipate that our proposal will provide a deeper insight into the role of NRP1 in the immune- vascular dysfunction in sepsis-induced ALI and lead to the development of a viable therapeutic strategy of targeting NRP1 to overcome its severity. The ultimate goal of our proposal is to benefit a large number of patients suffering from sepsis-induced lung complications for which there is no effective therapy till date.