Defining the mechanisms of MSC extracellular vesicle modulation of microglia metabolism and bioenergetics in traumatic brain injury recovery

项目摘要 神经炎症在创伤性脑损伤（TBI）的发生和发展中起着关键作用; 然而，大多数疗法无法解决多方面的问题。TBI后，小胶质细胞 被激活并产生破坏血液的炎性细胞因子（包括TNF-α和IL-6） 脑屏障（BBB），紧密连接，并导致周围免疫细胞的浸润，如 中性粒细胞、单核细胞和T细胞。确定新的免疫调节策略， 小胶质细胞和促进神经保护环境对于治疗这种毁灭性疾病至关重要。 间充质基质细胞（MSC）是一种有前途的治疗再生医学应用， 它们的免疫调节功能，这是由分泌的细胞外囊泡（MSC-EV）介导的 具有不同的表面组成和囊内货物。我们的团队展示了 MSC-EV调节参与神经炎症的细胞类型（如小胶质细胞、T细胞和 周细胞MSC-EV是TBI的有希望的治疗剂，因为它们可以i）具有可比的 免疫调节功能，ii）穿过BBB，和iii）解决安全性问题 与MSC递送相关的疾病（即肿瘤发生和血栓形成）。此外，瞄准能力 （由表面信号介导）和MSC-EV货物可以通过引发（预处理） MSC与不同的微环境线索，如细胞因子干扰素-γ和肿瘤 坏死因子α然而，在MSC-EV的作用和机制方面存在知识空白 在TBI的背景下，小胶质细胞的调制，以及这种效果是否可以通过引发增强。 拟议的工作旨在阐明MSC-EV调节小胶质细胞的机制， 特别关注MSC-EV线粒体转移。我们的中心假设是，MSC-EV产生了 将具有更大的功能，通过调节小胶质细胞向 更多的神经保护性表型（例如，炎性细胞因子和反应性细胞因子的产生减少） 氧物种），并且这种作用是由MSC-EV衍生的体外线粒体转移介导的， in vivo.我们将从以下几个方面来验证这一假说：1）确定线粒体的机制， 从MSC-EV转移对小胶质细胞代谢的影响，以及2）评估MSC-EV在 猪TBI模型。成功完成拟议的工作将创造一个新颖的，可调的方法 通过更好地了解MSC-EV，针对大脑的免疫系统和治疗TBI 行动机制。

PROJECT SUMMARY Neuroinflammation plays a critical role in both the onset and progression of traumatic brain injury (TBI); however, most therapies are unable to address the multifaceted aspects. Following TBI, microglia become activated and produce inflammatory cytokines (including TNF-α and IL-6) that damage blood brain barrier (BBB), tight junctions, and lead to infiltration of peripheral immune cells such as neutrophils, monocytes, and T cells. Identification of new immunomodulatory strategies that target microglia and promote a neuroprotective environment is critical for treating this devastating disease. Mesenchymal stromal cells (MSCs) are a promising therapy for regenerative medicine applications due to their immunomodulatory function, which is mediated by secreted extracellular vesicles (MSC-EVs) that possess distinct surface composition and intravesicular cargo. Our group has demonstrated the ability of MSC-EVs to modulate cell-types involved in neuroinflammation such as microglia, T cells and pericytes. MSC-EVs are a promising therapeutic for TBI because they can i) have comparable immunomodulatory function to parent MSCs, ii) cross the BBB, and iii) address safety concerns associated with MSC delivery (i.e. tumorigenesis and thrombosis). Further, the targeting capabilities (mediated by surface signals) and MSC-EV cargo can be engineered through priming (preconditioning) of MSCs with different microenvironmental cues such as cytokines Interferon-gamma and Tumor necrosis factor alpha. However, there is a gap in knowledge over the role and mechanisms of MSC-EV modulation of microglia in the context of TBI and whether this effect can be enhanced through priming. The proposed work seeks to elucidate the mechanisms by which MSC-EVs modulate microglia with a specific focus on MSC-EV mitochondrial transfer. Our central hypothesis is that MSC-EVs produced from cytokine-primed MSCs will have greater functionality through modulation of microglia towards a more neuroprotective phenotype (e.g. reduced production of inflammatory cytokines and reactive oxygen species) and that this effect is mediated by MSC-EV derived mitochondrial transfer in vitro and in vivo. We will test this hypothesis in the following aims: 1) Define the mechanisms of mitochondrial transfer from MSC-EV on microglia metabolism, and 2) Assess MSC-EV therapeutic efficacy in a porcine TBI model. Successful completion of the proposed work will create a novel, tunable approach for targeting the brain’s immune system and treating TBI through a better understanding of MSC-EV mechanisms of action.