Description of biomechanical properties of spheroids from human cells

球体是一种三维细胞培养物，由于其三维排列方式，比传统的二维细胞培养方式更能反映细胞在组织中的自然生理状况。因此，对于标准化的体外方法和个性化医疗的临床应用来说，它们都非常有趣。球体可以由一种细胞类型（单培养）或几种细胞类型（共培养）组成，通过直接细胞-细胞接触和细胞外基质（ECM）的蛋白质连接。每个球体的细胞数可以在很宽的范围内变化。由于球体是有生命的系统，球体内的细胞组织、细胞粘附蛋白钙粘蛋白和ecm蛋白的表达以及细胞骨架都不稳定，并且会随着时间（成熟）而改变，这强烈影响了它们的生物力学特性。这些老化过程在很大程度上是未知的，这使得很难产生具有明确生物力学特性的球体。本项目旨在研究球体的微观结构与生物力学特性之间的关系，并使用离散元法（DEM）建立数值模型来描述它们。为了实现这一目标，将系统地研究从正常人类真皮成纤维细胞（NHDF）中提取的球体的生物力学特性。为此，将在培养基中使用纳米压痕方法对球体和二维细胞层的机械性能进行广泛的实验研究。球体的微观结构将通过对球体的半薄和超薄切片的扫描电镜分析来确定。此外，实验还测定了细胞和分子生物学参数、钙粘蛋白、细胞骨架蛋白以及ECM蛋白的表达。这些数据将用于验证数值模型，这将允许对影响球体微观特性（微观结构，单细胞相互作用）和宏观生物力学特性之间关系的因素进行量化。对这些相关机制的基本理解将有助于在个性化医疗背景下生成具有定义属性的球体，这将显著提高应用的临床成功率。利用获得的生物力学特性和开发的DEM模型，可以预测不同应用（如组织工程、生物打印、球体工艺）中球体的工艺相关损伤，从而优化这些工艺。

Spheroids are 3D cell cultures, which reflect the natural physiological situation of cells in tissues better than traditional 2D cell culture approaches due to their 3D arrangement. Therefore, they are very interesting for standardized in vitro approaches but also for clinical applications in terms of a personalized medicine. Spheroids can consist of one cell type (mono-culture) or of several cell types (co-culture) connected by direct cell-cell contacts and proteins of the extracellular matrix (ECM). The number of cells per spheroid can be varied over a wide range. Since spheroids are living systems, cellular organization within the spheroids, expression of the cell adherence protein cadherin and of ECM-proteins as well as the cytoskeleton are not stable and will change over time (maturation), which strongly influence their biomechanical properties. These aging processes are largely unexplored, which makes it difficult to produce spheroids with defined biomechanical properties.This project aims to investigate the relationships between the microstructure and the biomechanical properties of spheroids over time and to describe them with a numerical model using the Discrete Element Method (DEM). To achieve this goal, the biomechanical properties of spheroids from Normal Human Dermal Fibroblasts (NHDF) will be systematically investigated. For this purpose, extensive experimental studies on the mechanical properties of spheroids as well as of 2D cell layers will be performed using the nanoindentation method in a culture medium. The microstructure of the spheroids will be determined by scanning electron microscopic analyses of semi- and ultra-thin sections of spheroids. Furthermore, the cell and molecular biological parameters, expression of cadherin, cytoskeletal proteins as well as proteins of the ECM are determined experimentally. These data will be used to validate the numerical models, which will allow a quantification of the factors influencing the relationships between the microscopic properties (microstructure, interactions of single cells) and macroscopic biomechanical properties of spheroids. A basic understanding of these relevant mechanisms will help to generate spheroids with defined properties for the respective requirements in the context of a personalized medicine, which will significantly improve the clinical success of an application. With the obtained biomechanical properties and developed DEM model, process-related damage to the spheroids in different applications (e.g. tissue engineering, bio-printing, spherox processes) can be predicted in order to optimize these processes.