Establishing AMP-activated protein kinase as a regulator of adipose stem cell plasticity and function in health and disease

为了生长、复制和发挥功能，细胞必须感知并对环境做出反应。以三磷酸腺苷(ATP)形式存在的能量是所有细胞过程所必需的，包括构建蛋白质、复制DNA、修复细胞膜和抗击感染。为了维持最佳的ATP浓度，多蛋白质系统持续监测细胞内的核苷酸水平，在ATP充足时推动生长和增殖，在ATP匮乏时停止能量消耗并启动分解代谢。其中一个网络是由AMP激活的蛋白激酶(AMPK)控制的，AMPK是一种几乎在所有生物体中都存在的酶，负责直接感知细胞的能量水平。能量应激下AMPK的激活会导致能量储存的崩溃，以及产生大部分ATP的受损或功能失常的线粒体的移除(有丝分裂)。AMPK的激活还会增加新的、健康的线粒体的产生(生物发生)，导致高效的ATP产生，并鼓励细胞摄取葡萄糖，以恢复和维持ATP。因此，AMPK提供了一个有吸引力的药物靶点，用于治疗许多能量稳态改变突出的疾病。干细胞存在于大多数组织中，负责组织的维护和损伤后的修复。代谢疾病，如肥胖、糖尿病和心血管疾病，通常在能量平衡失调时发生，仅靠改变生活方式很难逆转。在新陈代谢应激条件下，如炎症、缺氧和细胞衰竭，这些干细胞不会做出适当的反应，损害变得不可逆转。衰老也是如此，达到自我更新极限的干细胞不再能够修复组织。白色脂肪组织是一种专门的器官，主要负责荷尔蒙的产生和脂肪的安全储存，通常与代谢性疾病有关。白色脂肪组织过度膨胀会导致脂肪细胞死亡、全身炎症和胰岛素抵抗，这些都是代谢综合征的特征。脂肪干细胞(ADSCs)具有多潜能干细胞的许多特征，能够形成包括软骨、骨、骨骼和心肌在内的多种细胞类型。可获得性和可塑性的结合使ADSCs在再生和恢复性医学的靶向干细胞治疗中具有吸引力。我们之前的工作发现了一种新的脂肪细胞群体，我们称之为SMART细胞，它是通过在肥胖小鼠模型中通过基因激活AMPK产生的。这些细胞具有高线粒体含量，增加了有丝分裂能力，并表达了肌肉中通常存在的蛋白质，使它们能够产生热量，保护小鼠免受饮食诱导的肥胖和相关器官损伤。由于SMART细胞在培养中受到刺激时会表达这些肌肉特异性蛋白，因此它们可能对治疗代谢和年龄相关疾病有用。然而，人们对这些细胞的起源知之甚少，也不知道为什么当AMPK激活时，它们会发育而不是正常的白色脂肪细胞。我们的目的是了解AMPK在正常人类脂肪组织发育中的作用，以及干细胞分化关键点的代谢变化对细胞命运和功能的影响。利用人类永生化脂肪细胞来源的干细胞(h-iADSCs)和新鲜的人类ADSCs，我们的目标是确定AMPK在脂肪干细胞维持、细胞谱系规范以及成熟脂肪细胞所产生的功能中的作用。这项工作还将通过分离代谢性疾病患者的ADSCs来探索代谢不稳定对脂肪细胞谱系和功能的影响。为了确定AMPK激活的治疗价值，我们将AMPK激活剂C455处理的患者来源的ADSC移植到小鼠体内，在那里我们将评估它们形成完整脂肪组织的能力，以及它们对全身能量动态平衡的影响。

To grow, replicate and function, cells must sense and respond to their environment. Energy, in the form of adenosine triphosphate (ATP) is required for all cellular processes; to build proteins, replicate DNA, repair membranes and fight infections. To maintain optimal ATP concentrations, multi-protein systems continuously monitor nucleotide levels within the cell, driving growth and proliferation when ATP is plentiful, and by halting energy expenditure and initiating catabolism when ATP is scarce. One such network is governed by AMP-activated protein kinase (AMPK), an enzyme found in almost all organisms, responsible for directly sensing cellular energy levels. Activation of AMPK under energy stress drives the breakdown of energy stores and the removal of damaged or malfunctioning mitochondria (mitophagy), where the majority of ATP is generated. AMPK activation also increases the production of new, healthy mitochondria (biogenesis), leading to efficient ATP production, and encourages the uptake of glucose by the cell, to restore and maintain ATP. AMPK therefore presents an attractive drug target for the treatment of many diseases in which alterations in energy homeostasis feature prominently. Stem cells reside in most tissues, and are responsible for tissue maintenance and repair after damage. Metabolic diseases, such as obesity, diabetes and cardiovascular disease, often occur when energy balance is dysregulated, and are difficult to reverse with lifestyle change alone. Under conditions of metabolic stress, such as inflammation, hypoxia and cell exhaustion, these stem cells do not respond appropriately and the damage becomes irreversible. This process is also true of aging, when stem cells that reach their limits of self-renewal are no longer capable of tissue repair.White adipose tissue, a specialised organ primarily responsible for hormone production and the safe storage of lipids, is often implicated in metabolic disease. Excessive expansion of white adipose tissue leads to adipocyte death, systemic inflammation and insulin resistance, hallmarks of metabolic syndrome. Adipose-derived stem cells (ADSCs) share many features of pluripotent stem cells, capable of forming a diverse range of cell types, including cartilage, bone, skeletal and cardiac muscle. The combination of accessibility and plasticity make ADSCs attractive for targeted stem cell therapies in regenerative and restorative medicine. Our previous work identified a novel population of adipocytes, which we termed SMART cells, generated by genetically activating AMPK in a mouse model of obesity. These cells have a high mitochondrial content, increased mitophagy and express proteins usually found in muscle that allow them to generate heat, protecting the mice from diet-induced obesity and associated organ damage. As SMART cells express these muscle-specific proteins when stimulated in culture, they may be useful for the treatment of metabolic and age-related disease. However, little is known about the origin of these cells, or why they develop instead of normal white adipocytes when AMPK is active. Our aim is to understand the role of AMPK in the development of normal human adipose tissue, and the consequence of metabolic alterations at key points in stem cell differentiation on cell fate and function. Using human immortalised adipocyte derived stem cells (h-iADSCs), and fresh human ADSCs, we aim to establish the role of AMPK in adipose stem cell maintenance, in the specification of cell lineage, and in the resulting function of the mature adipocyte. The work will also explore the effect of metabolic instability on adipocyte lineage and function by isolating ADSCs from patients with metabolic disease. To determine the therapeutic value of AMPK activation, we will transplant patient-derived ADSCs, treated with AMPK activator C455, into mice, where we will assess their ability to form whole adipose tissue, and their effect on whole-body energy homeostasis.