Therapy of acute radiation syndrome and its complications by mesenchymal stromal cells conditioned with Toll-like receptor 9 agonists

项目摘要 仍然迫切需要制定医疗对策来治疗急性和迟发性辐射损伤 这可能是由大规模的放射性核事件造成的。不可预测的性质，以及复杂的 核爆炸造成的伤害使其成为一个具有挑战性的世界卫生问题。如果事件确实发生， 幸存者最关心的是辐射和损伤等机会性感染的发展 损害免疫系统。由于保护潜在的高数量的治疗选择有限， 个人可能会受到不同的辐射，新的和创新的战略是必要的，以促进 放射性损伤的治疗。基于细胞的治疗在许多疾病过程中显示出益处 与损伤和免疫功能障碍有关。间充质基质细胞（MSC）是理想的候选人， 细胞治疗，因为我们已经表明，他们改善生存鼠败血症通过调节免疫 反应和促进炎症消退。MSC的预处理可以提高其在肿瘤中的功效。 氧化炎性微环境，它们在辐射损伤后被施用到其中。我们已经表明 Toll样受体9激动剂（CpG-ODN）可能是预处理MSC的最佳候选物。我们 发现特异性CpG-ODN序列保护小鼠免受损伤后感染和辐射后败血症。 因此，该提议的首要目标是开发具有长CpG的MSC和CpG预处理的MSC。 使用CpG-MSC作为细胞治疗剂以降低辐射暴露的死亡率和发病率的长期目标 受伤我们推测，通过CpG-ODN刺激预处理MSC将增强其有益的 对抗微生物免疫功能和造血系统恢复的影响，以改善辐射损伤， 复苏为了验证我们的假设，我们提出了三个目标。在目标1中，我们将系统地测试和开发MSC 和CpG-ODN预处理的MSC作为放射诱导的损伤和感染的治疗在临床前 观察存活率、细菌计数和免疫细胞恢复的动物模型。初步数据显示， 与对照组相比，给予CpG条件MSC的小鼠在辐射和感染后的死亡率较低。 未处理和MSC处理的小鼠。在目的2中，我们将探索CpG-ODN刺激对细胞增殖的生物学效应。 MSC表型、功能、归巢损伤、对氧化应激的反应以及它们与 照射和未照射的免疫细胞。我们还将使用RNAseq来分析MSC的CpG-ODN应答， 识别机械目标在目标3中，我们将使用项目过程中获得的数据来确定 介导CpG-ODN调节的MSC的有益活性的机制和途径。为此，我们 还将确定用MSC和CpG MSC治疗是否会影响骨髓间充质干细胞延迟效应的发展。 辐射这项研究具有重要意义，因为从这项研究中获得的知识将为以下方面提供临床前数据： 将CpG预处理的MSC转化为治疗各种损伤的有效医学对策 可能发生在放射性核事件之后。

Project Summary There remains an urgent need to develop medical countermeasures to treat acute and delayed radiation injuries that could result from a large-scale, radio-nuclear event. The unpredictable nature, as well as the complexity of the injuries from a nuclear blast makes this a challenging world health problem. If an event does occur, an immediate concern for survivors will be the development of opportunistic infections as radiation and injury damage the immune system. As there are limited treatment options for protecting the potentially high numbers of individuals that might be exposed to varying radiation, new and innovative strategies are necessary to advance the treatment of radiation injury. Cellular based therapy has shown benefit in numerous disease processes associated with injury and immune dysfunction. Mesenchymal stromal cells (MSCs) are ideal candidates for a cellular therapeutic as we have shown that they improve survival in murine sepsis by modulating the immune response and promoting resolution of inflammation. Pre-conditioning of MSCs may improve their efficacy in the oxidative inflammatory microenvironment into which they are administered after radiation injury. We have shown that the Toll-like receptor 9 agonist (CpG-ODN) may be an optimal candidate for pre-conditioning of MSCs. We found that a specific CpG-ODN sequence protected mice from post-injury infection and sepsis post-radiation. Thus, the overarching aim of this proposal is to develop MSCs and CpG pre-conditioned MSCs with the long- term goal of using CpG-MSCs as a cellular therapeutic to reduce the mortality and morbidity of radiation exposure injuries. We hypothesize that pre-conditioning MSCs by CpG-ODN stimulation will enhance their beneficial effects on anti-microbial immune function and hematopoietic system recovery to improve radiation injury and recovery. To test our hypothesis, we propose three aims. In Aim 1 we will systematically test and develop MSCs and CpG-ODN pre-conditioned MSCs as a treatment for radiation-induced injury and infection in a pre-clinical animal model looking at survival, bacterial counts, and immune cell recovery. Our preliminary data show that mice given CpG conditioned MSCs had a lower mortality rate following radiation and infection compared with untreated and MSC-treated mice. In Aim 2 we will explore the biological effects of CpG-ODN stimulation on MSC phenotype, function, homing to injury, response to oxidative stress, as well as their interaction with irradiated and non-irradiated immune cells. We will also use RNAseq to profile the CpG-ODN response of MSCs and identify mechanistic targets. In Aim 3 we will use data gained during the course of this project to identify mechanisms and pathways that mediate the beneficial activity of CpG-ODN conditioned MSCs. In this aim we will also determine whether treatment with MSCs and CpG MSCs impacts development of delayed effects of radiation. This research has significance, as knowledge gained from this study will provide pre-clinical data for translating CpG pre-conditioned MSCs as effective medical countermeasures for treating the variety of injuries that could occur following a radio-nuclear event.