Maintenance, regeneration, and repair of skeletal systems: molecular regulation of autophagy in the joint

骨骼系统的维护、再生和修复：关节自噬的分子调节

• 批准号: BB/Y002504/1
• 负责人: Chrissy Hammond
• 金额: $ 78.38万
• 依托单位: University of Bristol
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2023
• 资助国家: 英国
• 起止时间: 2023 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FY002504%2F1
• 关键词: Maintenance; regeneration; repair; skeletal; systems

Our skeletons constantly adapt throughout life so they are strong enough to withstand our needs, while remaining relatively light to facilitate movement. This is achieved by balanced activity of bone building cells called osteoblasts, and bone resorbing cells called osteoclasts which remove old or poor quality bone. In ageing and in some genetic conditions, this balance can often be lost. If resorption exceeds the amount of bone being made it can lead to osteoporosis, where bones are less dense, more fragile and prone to fracture. In other cases, bone making can exceed resorption and this can lead to bone being made in places it normally wouldn't be found. This can include spurs on the edges of the bone that can impede joint movement in the hips, spine or knees, or mineralization of soft tissues like cartilage and ligaments, which normally help to facilitate smooth joint movement, leading to joint stiffness. However, we still don't fully understand how the cells of the skeletal system, the osteoblasts and osteoclasts of the bone and the chondrocytes that maintain joint cartilage, are affected by ageing. One hypothesis is that autophagy, which is a process by which cells break down and recycle parts of their own working machinery, becomes impaired in the ageing skeleton. This causes cells to become less healthy over time and less able to adapt to the needs to of the organism. Evidence for this idea comes from genetic studies which have linked some autophagy genes to osteoarthritis and osteoporosis, which suggests if autophagy is impaired it might lead to earlier onset of skeletal ageing. We want to use zebrafish as an animal model to test whether this is happening. Zebrafish are a model system which make bones in the same ways humans and other animals do, but are also translucent. This means that we can label the cells of the skeletal system with fluorescent markers and watch their behaviour in the living fish in response to injury, to ageing. We can also monitor the effects of the addition of drugs, such as steroids, or bisphosphonates which are also used clinically. Indeed, some of the drugs in the clinic were first identified using zebrafish. We have generated some lines of zebrafish in which some of the regulators of autophagy are switched off, and we have seen that these fish have abnormal skeletons, with changes both to cartilage and to bone. These changes become more severe as the fish age. We will use live imaging of fluorescently tagged cells in the living fish to monitor autophagy during skeletal development in normal and mutant fish, and to watch how skeletal cells behave in young and old fish in response to injury so that we can identify which cells might cause these differences. When we better understand which cells are causing these changes, we will test which proteins and processes in those cells are disrupted and see whether we can identify ways to change this, by working in tissue culture, in which cells are grown in petri dishes to allow us to study these processes in more detail.

我们的骨骼在整个生命过程中不断适应，因此它们足够坚固以满足我们的需求，同时保持相对较轻以便于运动。这是通过平衡称为成骨细胞的骨构建细胞和称为破骨细胞的骨再吸收细胞的活性来实现的，破骨细胞去除旧的或质量差的骨。在衰老和某些遗传条件下，这种平衡往往会失去。如果骨吸收超过了骨生成量，就会导致骨质疏松症，即骨密度降低，更脆弱，更容易骨折。在其他情况下，骨生成可能超过吸收，这可能导致骨生成在通常不会被发现的地方。这可能包括骨骼边缘的骨刺，这可能会阻碍髋关节，脊柱或膝盖的关节运动，或软骨和韧带等软组织的矿化，这些软组织通常有助于促进关节运动，导致关节僵硬。然而，我们仍然不完全了解骨骼系统的细胞，骨骼的成骨细胞和破骨细胞以及维持关节软骨的软骨细胞是如何受到衰老的影响的。一种假设是，自噬，这是一个过程，细胞分解和回收部分自己的工作机器，成为受损的老化骨骼。这导致细胞随着时间的推移变得不健康，并且不太能够适应生物体的需要。这一观点的证据来自遗传学研究，这些研究将一些自噬基因与骨关节炎和骨质疏松症联系起来，这表明如果自噬受损，可能会导致骨骼衰老的提前发生。我们想用斑马鱼作为动物模型来测试这种情况是否发生。 斑马鱼是一个模型系统，它以与人类和其他动物相同的方式制造骨骼，但也是半透明的。这意味着我们可以用荧光标记物标记骨骼系统的细胞，并观察它们在活鱼中对损伤和衰老的反应。我们还可以监测添加药物的效果，如类固醇或临床上使用的双膦酸盐。事实上，临床上的一些药物最初是用斑马鱼发现的。我们已经培养了一些斑马鱼品系，其中一些自噬调节因子被关闭，我们已经看到这些鱼有异常的骨骼，软骨和骨骼都发生了变化。随着鱼的年龄增长，这些变化变得更加严重。我们将使用活体荧光标记细胞的实时成像来监测正常和突变鱼骨骼发育过程中的自噬，并观察骨骼细胞在年轻和年老的鱼中对损伤的反应，以便我们可以识别哪些细胞可能导致这些差异。当我们更好地了解哪些细胞引起这些变化时，我们将测试这些细胞中的哪些蛋白质和过程被破坏，看看我们是否可以通过组织培养来确定改变这种情况的方法，其中细胞在培养皿中生长，使我们能够更详细地研究这些过程。