Active Extracellular Vesicles - defining a novel, extracellular metabolic compartment and its role in the control of inflammation.

炎症是免疫系统用来帮助抵御感染和修复创伤造成的损害的一种正常防御机制。然而，只有当反应被关闭，受感染或受伤的组织恢复正常时，炎症的好处才能实现。这个过程被称为“分解”，依赖于免疫细胞(已经完成了它们的保护工作)被移除。为了清除它们，这些不需要的细胞死亡，在这样做的同时，它们释放出小的膜袋(称为细胞外小泡：EV)。我们已经证明，这些EV是吸引“承担者”细胞(“巨噬细胞”)的关键，这些细胞通过吃掉濒临死亡的细胞来移除它们，至关重要的是，它们会产生额外的信号来驱动组织修复。相对来说，人们对这些EV的功能知之甚少。我们已经确定了一种促进巨噬细胞募集的分子，但这些EV促进巨噬细胞“修复”反应的方式尚不清楚。现在，我们的初步工作表明，这些EV携带一系列活性酶，这些酶将有助于产生可能控制炎症和修复反应的小分子脂分子。这里提出的工作将定义这些活性酶的作用，这些酶从死亡的细胞传递到EV的巨噬细胞。因此，这些“活性”EV代表了一种新的细胞外代谢活性隔间，能够将炎症控制信号传递给周围的细胞，如巨噬细胞。为什么这一点很重要？未能控制炎症反应可能会导致慢性炎症和许多与衰老相关的炎症性疾病。因此，通过了解促进炎症的细胞和促进分解和修复的细胞之间的通信机制，我们将更清楚地了解由炎症驱动的疾病(例如心血管疾病、癌症、神经退行性变)的可能治疗方法。此外，我们将从一系列其他已知有助于促进修复的细胞中研究EV。例如，一种名为“MSC”的干细胞在再生医学中大有希望，但最近从这些细胞中释放的EV被认为是活跃的，可以刺激“修复环境”。因此，我们将从MSC中对EV的炎症控制代谢活性进行详细的研究。这增加了我们能够确定EV修复活动所需的关键因素的可能性，从而为再生医学开辟了基于干细胞或无细胞的新疗法。为了验证我们的核心假设，我们将从一系列死亡和存活的细胞中分析EV的组成，以确定酶的存在、它们的底物和它们的产物。我们将在体外和体内测试EVS促进修复的能力，并抑制关键酶以评估这一活动的本质。此外，我们将寻求‘调整’或优化细胞培养条件，以最大限度地提高EV的产量和功能。这项工作将首次详细说明一个炎症控制、代谢活性的胞外隔间。它将在分子水平上定义死亡细胞如何与其他细胞沟通，以确保炎症得到控制。这一点很重要，因为对炎症的有效控制会导致疾病。因此，利用我们的工作将针对那些炎症有助于推动疾病的条件，并将使新的战略能够促进自我修复。

Inflammation is a normal defence mechanism used by the immune system to help protect from infection and to repair damage caused by trauma. However, the benefits of inflammation are only realised when the response is turned off and the infected or traumatised tissue is returned to normal. This process is known as 'resolution' and relies on immune cells (that have done their protective job) being removed. To allow their removal, these unwanted cells die and, as they do so, they release small membrane bags (called extracellular vesicles: EV). We have shown that these EV are key to attracting 'undertaker' cells ('macrophages') that remove the dying cells by eating them and, crucially, they produce additional signals that drive the tissue to repair.Relatively little is known of how these EV function. We have identified a molecule that promotes recruitment of macrophages but the manner in which these EV promote 'repair' responses in macrophages remains unknown. Now, our preliminary work has shown that these EV carry a family of active enzymes which will help to produce small lipid molecules that may control inflammation and repair responses. The work proposed here will define the action of these active enzymes that are delivered from dying cells to macrophages in EV. As such, these 'active' EV represent a novel extracellular metabolically-active compartment capable of transmitting inflammation-controlling signals to surrounding cells such as macrophages.Why is this important? A failure to control inflammatory responses can lead to chronic inflammation and many of the inflammatory diseases associated with ageing. So, by understanding the mechanisms of communication between cells promoting inflammation, and those cells promoting resolution and repair, we will have a clearer insight to possible therapeutic approaches for diseases that are driven by inflammation (e.g. cardiovascular disease, cancer, neurodegeneration). Furthermore, we will study EV from a range of other cells that are known to help promote repair. For example, a type of stem cell, an 'MSC', has held great promise for regenerative medicine but recently the EV released from these cells have been proposed to be active to stimulate a 'repair environment'. Consequently, we will study in detail the inflammation-controlling metabolic activity of EV from MSC. This raises the possibility that we may be able to define the crucial factors required for EV repair activity, opening up novel stem cell-based or cell-free therapies for regenerative medicine. In order to test our core hypothesis, we will analyse, from a range of dying and viable cells, EV composition for the presence of enzymes, their substrates and their products. We will test the EVs' ability to promote repair in vitro and in vivo and we will inhibit the key enzymes to assess the essential nature of this activity. Furthermore, we will seek to 'tune' or refine cell culture conditions to maximise EV production and function.This work will, for the first time, detail an inflammation-controlling, metabolically-active extracellular compartment. It will define, at a molecular level, how dying cells communicate with other cells to ensure inflammation is controlled. This is important because ineffective control of inflammation leads to disease. Thus, exploitation of our work will target those conditions where inflammation helps drive disease and will enable novel strategies to promote self-repair.