Mesenchymal stem cell-derived extracellular vesicles as a cell-free therapy for ARDS.

急性呼吸窘迫综合征（ARDS）是一种严重的临床综合征，影响20%的重症监护危重患者。不幸的是，这些人中约有30-50%将死于这种疾病，幸存者的生活质量可能因随之而来的慢性肺部问题而严重降低。目前还没有有效的治疗方法，因此迫切需要新药。ARDS的特点是肺部过度和失调的炎症，导致免疫细胞不适当的浸润，肺部充满水，无法维持其主要功能-呼吸。为了呼吸，这些病人需要机械通气。近年来，细胞疗法越来越受到临床医生和基础科学家的关注。其中最有希望的细胞治疗候选者是被称为间充质干细胞（MSCs）的细胞，它代表了一种成体干细胞（可以很容易地从成年患者或健康志愿者的组织中分离出来），因此不存在伦理问题。MSCs在实验室条件下易于繁殖和操作，并具有产生多种具有免疫调节特性的介质的能力。然而，对MSC给药的长期影响（如肿瘤的发展）的安全性以及后勤挑战的担忧可能会限制其成功的临床转化。越来越多的证据表明，间充质干细胞主要通过分泌不同的生物活性介质发挥作用。因此，研究人员越来越多地考虑MSC细胞产品（它不能复制，因此形成肿瘤）作为全细胞治疗的替代方案。在这个项目中，我们将研究由间充质干细胞释放的小膜包裹的“细胞外囊泡”的治疗潜力。这些囊泡代表了间充质干细胞分泌组的关键成分，并且相对容易通过高速离心从间充质干细胞条件培养基中分离出来。之前，我们和其他人已经证明，这些囊泡能够在各种ARDS临床前模型中重现MSCs的治疗效果。此外，在我们最近的工作中，我们发现这些囊泡含有负责维持细胞生物能量的细胞器（线粒体）。这是一个非常令人兴奋的发现，因为众所周知，线粒体功能在急性炎症条件下受损，如ARDS。在本项目中，我们将使用ARDS的小动物模型来研究这些囊泡的治疗潜力，优化最佳给药途径和策略，了解它们在组织中的生物分布，并研究功能活跃的线粒体对治疗效果是否重要，以及哪些组织是ev中转移的线粒体的主要受体。这些研究将进一步补充EV线粒体转移到体外原代人肺细胞的研究。这些数据是监管机构（如英国药品和保健产品监管机构和欧洲药品管理局）对新疗法临床转化的基本要求。该项目的主要预期成果将是建立一个明确的临床开发策略，并了解MSC衍生的细胞外囊泡的具体作用机制（基于线粒体转移）。这些数据将立即为进一步在大型ARDS动物模型中进行临床前研究提供信息，以获得有关MSCs ev的安全性、有效性和作用机制的必要数据。小型和大型动物模型的总体数据将为ARDS患者的MSC细胞外囊泡的早期临床试验提供信息。

Acute Respiratory Distress Syndrome (ARDS) is a severe clinical syndrom that affects 20% of all critically ill patients in intensive care. Unfortunately, around 30-50% of these people will die from the condition and the quality of life of the survivers can be seriously reduced due to the consequent chronic lung problems.There is no effective treatments for this condition at this time and therefore new medicines are urgently needed. ARDS is characterised by an excessive and disregulated inflammation in the lung that leads to inappropriate infiltration of immune cells, lungs fill with water and fail to maintain its main function - breathing. In order to breath these patients require mechanical ventilation.Recently more and more clinicians and basic scientists attention is drawn to cell-based therapies. One of the most promising candidate for a cell based therapy are cells termed Mesenchymal Stem Cells (MSCs), which represent one kind of adult stem cells (can be easily isolated from tissues of adult patients or healthy volunteers), because of this, there is no ethical issues. MSCs could easily be propagated and manipulated under laboratory conditions and have capacity to produce multiple mediators with immuno-modulatory properties. However, concerns regarding safety of long-term effects of MSC administration (e.g. development of tumors) as well as logistical challenges may limit their successful clinical translation. Accumulating evidence now suggests that MSCs act primarily through secretion of different biologically active mediators. Therefore researchers are increasingly considering MSC cell products (which can not replicate and therefore form tumors) as an alternative to the whole cell therapy.In this project we will investigate the therapeutic potential of small membrane wrapped "exracellular vesicles" released by MSCs. These vesicles represent the key component of MSCs secretome and are relatively easy to isolate from MSC conditioned medium by high speed centrifugation. Previously we and others had demonstrated that these vesicles were capable to recapitulate therapeutic effects of MSCs in various pre-clinical models of ARDS. Furthemore, in our recent work we had found that these vesicles contain organelles responsible for maintaining cell bioenergetics (mitochondria). This is very exciting finding as it is known that mitochondrial function is impaired in acute inflammatory conditions such as ARDS.In this project we will use small animal model of ARDS to investigate the therapeutic potential of these vesicles, optimise best route and strategy of administration, gain isights into their bio-distribution in the tissues and also investigate whether or not functionally active mitochondria are important for the therapeutic effect and which tissues are major recipients of the mitochondria transferred in the EVs. These studies will be further complemented by the investigation of the EV mitochndrial transfer to primary human lung cells in vitro. These data are an essential requirement by regulatory bodies such as UK Medicines and Healthcare Products Regulatory Agency and European Medicines Agency for clincal translation of new therapies. The principle anticipated output of this project will be the establishment of a defined strategy for clinical development and knowledge of the specific mechanism of action (based on mitochondrial transfer) of the MSC derived extracellular vesicles. More immediately these data will inform further pre-clinical studies in larger animal models of ARDS required to obtain neccessary data on safety, efficacy and mechanism of action of MSCs EVs. Collectively data in small and large animal models will inform early phase clinical trials for MSC extracellular vesicles in patients with ARDS.