From ageing to space travel: Developing an organotypic model of skeletal tissue disuse for understanding degeneration in altered environments

骨组织在维持钙平衡和器官功能方面的作用是必不可少的。钙作为一种通用货币，对大多数生理过程都是必不可少的，包括细胞通讯、肌肉收缩、血液凝结和神经功能。骨骼组织的大量丢失发生在几种临床情况下，包括废用性骨质疏松症、衰老、脊髓损伤、在微重力下的静止和失重，所有这些都以承重区域的骨量迅速和显著丢失为特征，包括腿部、脊柱和髋部。这增加了骨折的风险，损害了愈合过程，给医疗保健系统和与干预相关的成本带来了巨大的负担。虽然众所周知，骨量随着负荷的减少而成比例减少，但管理它的细胞过程需要进一步了解。人们普遍认为，骨吸收增加，骨沉积减少，然而，还没有强有力的方法来研究骨重建的不平衡。目前已经有成熟的动物模型来研究肌肉骨骼的废弃和骨丢失，这些动物模型可以通过手术切除参与骨骼代谢的腺体、使用毒素固定、手术切除神经、肌腱或脊髓，或者通过尾部悬吊方法促进后肢卸载，从而导致动物骨量减少。其中一些过程对动物非常有害，另一些过程干扰了骨骼动态平衡的生物化学。此外，这些结果并不能完全代表人类的情况，因为这两个物种之间以及同一实验动物的不同品系之间的骨重建过程存在差异。本工作的目的是建立一种能够改进和减少用于了解肌肉骨骼退化的动物数量的模型，并提供一种研究器皿中骨丢失的方法。这项工作将产生一种与生理相关的模型，在该模型中，人类来源的特殊骨细胞以及这些细胞类型的组合将在人类衍生的生物支架内培养。细胞将使用几个旋转培养生物反应器进行机械卸载，这些生物反应器可以使用不断旋转的容器保持细胞持续悬浮，从而模拟失重状态。该平台将与一系列来自人体组织的活性基质结合使用，例如纤维蛋白(血栓样)，它们是可降解的，在病理上具有代表性。这些细胞将在形态上适应为悬浮培养的球体，并可以提供一定程度的支持，同时允许骨细胞用胶原基质取代它们，大量矿化这个模板并将自己埋入其中，如基础工作所示。这个模型将有助于研究早期的骨丢失过程，这对于了解废用病理学是必不可少的，可以提供细胞毒性、遗传毒性和不相容的化合物的第一阶段消除步骤，从而使更少的有害物质被用于体内测试，并将允许测试许多有前景的药物和潜在的治疗方法。它还将帮助研究人员在广泛的骨骼条件下开发治疗方法，不仅与废用性骨质疏松症有关，而且还与过度的骨骼研究、骨癌、炎症降解和多系统研究有关。

The role of bone tissue in maintaining calcium balance and organ function is essential. Calcium acts as a universal currency and is essential for most physiological processes, including cellular communication, muscle contraction, blood clotting and nerve function. A substantial loss of skeletal tissue takes place in several clinical contexts, including disuse osteoporosis, ageing, spinal cord injury, immobilisation and weightlessness in microgravity, all characterised by rapid and significant loss in bone mass in the load-bearing regions, including lower limbs, spine and hip. This increases the risk in fractures and impairs the healing process, placing a significant burden on the healthcare system and the costs associated with interventions.While bone mass is known to decrease proportionally with reduced loading, the cellular processes governing it require further understanding. It is widely accepted that bone resorption is increased and bone deposition is decreased, however, there is no robust way of studying the imbalance in bone remodelling.There are well-established animal models for studying musculoskeletal disuse and bone loss, which cause a reduction in bone mass in animals either through surgical removal of glands involved in bone metabolism, immobilisation using toxins, surgical resection of nerves, tendons or the spinal cord, or a tail suspension method facilitating hindlimb unloading. Some of these processes are very detrimental for the animals and others interfere with the biochemistry of skeletal homeostasis. Moreover, the results are not entirely representative of the human conditions, as differences exist in the bone remodelling process between the two species and between strains of the same laboratory animal.The aim of this work is to produce a model that can refine and reduce the number of animals used for understanding musculoskeletal degeneration and to provide a method to study bone loss in a dish. The work will generate a physiologically-relevant model, in which specialised bone cells of human origin as well as combinations of these cell types will be cultured inside human-derived biological scaffolds. Cells will be provided with mechanical unloading using several rotary culture bioreactors that can keep cells in a constant suspension using constantly rotating vessels, thus simulating a weightless state. This platform will be used in combination with a range of active matrices derived from human tissue, such as fibrin (blood clot-like), which are degradable and pathologically representative. These will be morphologically adapted into spheroids for suspended culture and can provide a degree of support while allowing bone cells to replace them with collagenous matrix, heavily mineralise this template and bury themselves inside it, as shown by foundation work. This model will help in studying early bone loss processes which are essential for understanding disuse pathology, can provide a first-stage elimination step of cytotoxic, genotoxic and incompatible compounds leading to less harmful agents being progressed for in vivo testing, and will allow the testing of numerous promising drugs and potential therapeutics. It will also help researchers developing treatments in a wide range of skeletal conditions, not only relevant to disuse osteoporosis, but also in excessive bone research, bone cancers, inflammatory degradation and multi-systemic research.