The role of extracellular mechanics in skin tissue engineering and ageing

皮肤包含其驻留细胞的生物机械/化学和拓扑学上不同的小生境。表皮细胞位于坚硬（MPa）的胶原IV/VII和富含层粘连蛋白的细胞外基质（ECM）上，称为基底膜（BM），而真皮成纤维细胞位于相当柔软（0.1- 10 kPa）的胶原III/I和基于胶原蛋白/弹性蛋白的基质上。随时间的老化对皮肤生物力学有着深远的影响。由于ECM合成减少、胶原蛋白I/弹性蛋白断裂和重塑，皮肤逐渐失去其天然弹性并变得起皱和脆弱。胶原蛋白III/I比率在老化真皮中增加，而BM表现出明显的胶原蛋白VII损失和脆性。这些变化如何影响机械感测和老化皮肤细胞生理学知之甚少。然而，众所周知，衰老会导致细胞硬化，从而影响收缩性和流动性。然而，在组织工程中，皮肤组织的地形和时序机械可变性在很大程度上被忽视。标准细胞培养程序涉及在非生理硬（GPa）表面（即聚苯乙烯）上繁殖。然而，主要的细胞功能，如身份，增殖，信号传导，行为和架构是由细胞外环境的刚度。因此，目前的体外工程皮肤替代品不能准确代表天然皮肤并不奇怪。具体而言，塑料培养皿上的生长异常激活真皮成纤维细胞（例如纤维化），诱导应激信号传导途径，并改变细胞结构，同时它基本上限制了细胞增殖。因此，我们的假设是，仿生2D表面的使用和支持角质形成细胞和成纤维细胞机械传感的分子机制的研究将促进适当的全厚度3D年轻/老年皮肤模型的开发。开发这样的模型将有利于个人医疗保健和化妆品行业（目标验证，技术鉴定），也是研究体外皮肤老化机制的关键工具。

Skin contains bio-mechanically/-chemically and topologically distinct niches for its resident cells. Epidermal cells rest on a stiff (MPa) collagen IV/VII and laminin-rich extracellular matrix (ECM) termed the basement membrane (BM), while dermal fibroblasts reside on considerably softer (0.1-10kPa) collagen III/I and fibrillin/elastin-based matrixes. Chronological ageing has profound effects on skin biomechanics. Skin progressively loses its natural elasticity and becomes wrinkled and fragile due to reduced ECM synthesis, collagen I/elastin fragmentation and remodelling. The collagen III/I ratio increases in the aged dermis, while the BM exhibits marked collagen VII loss and fragility. How these changes affect mechano-sensing and aged skin cell physiology is poorly understood. However, it is well established that ageing yields cell stiffening, which affects contractility and mobility. The topographic and chronological skin tissue mechanical variability is however, largely ignored in tissue engineering. Standard cell culture procedures involve propagation on non-physiologically hard (GPa) surfaces (i.e. poly-styrene). Yet, major cellular functions such as identity, proliferation, signalling, behaviour and architecture are determined by the stiffness of the extracellular environment. It is thus not surprising that current in vitro-engineered skin-substitutes do not accurate represent native skin. Specifically, growth on plastic dishes abnormally activates dermal fibroblasts (e.g. fibrosis), induces stress signalling pathways, and alters cell architecture, while it substantially limits cell proliferation. Therefore our hypothesis is that the use of biomimetic 2D surfaces and the study of the molecular mechanisms that underpin keratinocyte and fibroblast mechano-sensing will facilitate the development of proper full thickness 3D young/aged skin models. Developing such models will be beneficial for the personal healthcare and cosmetics industry (target validation, technology identification) and a pivotal tool for the study of the mechanisms that underpin skin ageing in vitro.