The role of muscle-derived extracellular vesicles in adipocyte metabolism

胞外囊泡(EVS)是几乎所有类型的细胞都能释放的膜结合囊泡，潜在地 介导细胞间的通讯，从而在许多生理和病理过程中发挥重要作用 流程。这种能力依赖于电动汽车作为一种广泛的内源能源的运输工具的能力 货物分子，如RNA、蛋白质和脂类。EVS也被发现可以显示组织特异性 表面分子(如整合素和葡聚糖)介导的识别，使其成为有希望的药物 递送应用程序。然而，更好地了解电动汽车的组成和货物是必要的，以使 使用电动汽车作为运输工具。初步数据显示，在肥大刺激后，肌肉衍生 EVS(MEV)是专门输送到脂肪组织的。有趣的是，将MEV运送到脂肪组织 诱导高脂饮食小鼠脂肪分解和脂肪丢失。这一令人兴奋的初步数据可能代表着第一个 步入肥胖症治疗的发展阶段。此外，我们的初步数据显示， 二酰甘油(DAG)在MEV内，它可以在交付时介导蛋白激酶C(PKC)的激活和 诱导脂肪细胞代谢的变化。拟议的项目在我们初步数据的基础上进行了扩展，并基于 关于肥大刺激会改变MEV的脂质组成的总体假设，导致 脂肪细胞的代谢发生变化。更具体地说，假设肥厚性刺激将增加 MEV内的DAG浓度将通过激活PKC诱导脂肪细胞脂解，从而导致全身 新陈代谢的好处。为了确认我们的初步数据，我们将使用肌肉细胞(C2C12细胞)来 分离有或没有肥大刺激的MEV(用胰岛素样生长因子1治疗)，并进行 这些小泡的脂类分布。在建立了脂谱后，MEVS将被转移到脂肪细胞 (3T3L1细胞)或将尾静脉注射到肥胖小鼠体内。这个项目的目的是测试它的工作 通过追求三个具体目标来提出假设。具体目标1是确定脂类成分是否 骨骼肌胞外小泡对肥大刺激的反应是改变的；具体目标2是 确定MEVS是否通过激活PKC刺激脂肪细胞脂解；具体目标3是 确定细胞外小泡对肥大刺激的反应是否可以通过 增强肥胖小鼠的脂解作用。初步数据表明，MEVS对脂肪细胞有有益的影响 肥胖小鼠的代谢和诱导脂肪丢失。如果拟议的实验结果提供了证据 支持这一假设，未来的R01应用将寻求严格测试哪一类或哪些类的脂类可以 特异性地调节脂肪细胞代谢的变化。

Extracellular vesicles (EVs) are membrane-bound vesicles released by virtually all cell types, potentially mediating intercellular communication and thereby play essential roles in many physiological and pathological processes. This capacity relies on EVs’ ability to serve as delivery vehicles for a wide range of endogenous cargo molecules, such as RNAs, proteins, and lipids. EVs have also been found to display tissue-specific recognition mediated by surface molecules (such as integrins and glycans), making them promising for drug delivery applications. However, a better understanding of EV composition and cargo is necessary to enable the use of EVs as delivery vehicles. Preliminary data demonstrate that after a hypertrophic stimulus, muscle-derived EVs (mEVs) are specifically delivered to adipose tissue. Interestingly, the delivery of mEVs to adipose tissue induced lipolysis and fat loss in high-fat-diet-fed mice. This exciting preliminary data could represent the first steps into the development of therapy for obesity. Additionally, our preliminary data indicate high levels of diacylglycerol (DAG) within mEVs, which could, upon delivery, mediate protein kinase C (PKC) activation and induce changes in adipocyte metabolism. The proposed project expands upon our preliminary data and is based on the overarching hypothesis that a hypertrophic stimulus would change the lipid makeup of mEVs, leading to adipocyte metabolism changes. More specifically, it is hypothesized that a hypertrophic stimulus will increase DAG concentration within mEVs that will induce adipocyte lipolysis through PKC activation, resulting in wholebody metabolic benefits. In order to confirm our preliminary data, we will use muscle cells (C2C12 cells) to isolate mEVs with or without a hypertrophic stimulus (treatment with insulin-like growth factor 1) and perform a lipid profile of these vesicles. After establishing the lipid profile, mEVs will be transferred to adipocyte cells (3T3L1 cells) or will be tail vein injected into obese mice. The purpose of this project is to test this working hypothesis by pursuing three specific aims. Specific Aim 1 is to determine whether the lipid composition of skeletal muscle extracellular vesicles is altered in response to a hypertrophic stimulus; Specific Aim 2 is to determine whether mEVs stimulate adipocyte lipolysis through PKC activation; and Specific Aim 3 is to determine whether extracellular vesicles in response to a hypertrophic stimulus can induce fat loss through enhanced lipolysis in obese mice. Preliminary data suggest that mEVs provide beneficial effects on adipocyte metabolism and induce fat loss in obese mice. If the results of the proposed experiments provide evidence to support the hypothesis, a future R01 application will seek to rigorously test which class/classes of lipids can specifically mediate changes in adipocyte metabolism.