Tomasch Janine, Maleiner Babette, Heher Philipp, Rufin Manuel, Andriotis Orestis G, Thurner Philipp J, Redl Heinz, Fuchs Christiane, Teuschl-Woller Andreas H
Department Life Science Engineering, University of Applied Sciences Technikum Wien, Vienna, Austria.
The Austrian Cluster for Tissue Regeneration, Vienna, Austria.
Front Bioeng Biotechnol. 2022 May 20;10:836520. doi: 10.3389/fbioe.2022.836520. eCollection 2022.
Fibrin hydrogels have proven highly suitable scaffold materials for skeletal muscle tissue engineering in the past. Certain parameters of those types of scaffolds, however, greatly affect cellular mechanobiology and therefore the myogenic outcome. The aim of this study was to identify the influence of apparent elastic properties of fibrin scaffolds in 2D and 3D on myoblasts and evaluate if those effects differ between murine and human cells. Therefore, myoblasts were cultured on fibrin-coated multiwell plates ("2D") or embedded in fibrin hydrogels ("3D") with different elastic moduli. Firstly, we established an almost linear correlation between hydrogels' fibrinogen concentrations and apparent elastic moduli in the range of 7.5 mg/ml to 30 mg/ml fibrinogen (corresponds to a range of 7.7-30.9 kPa). The effects of fibrin hydrogel elastic modulus on myoblast proliferation changed depending on culture type (2D vs 3D) with an inhibitory effect at higher fibrinogen concentrations in 3D gels and in 2D. The opposite effect was evident in differentiating myoblasts as shown by gene expression analysis of myogenesis marker genes and altered myotube morphology. Furthermore, culture in a 3D environment slowed down proliferation compared to 2D, with a significantly more pronounced effect on human myoblasts. Differentiation potential was also substantially impaired upon incorporation into 3D gels in human, but not in murine, myoblasts. With this study, we gained further insight in the influence of apparent elastic modulus and culture type on cellular behavior and myogenic outcome of skeletal muscle tissue engineering approaches. Furthermore, the results highlight the need to adapt parameters of 3D culture setups established for murine cells when applied to human cells.
过去，纤维蛋白水凝胶已被证明是用于骨骼肌组织工程的高度合适的支架材料。然而，这些类型支架的某些参数会极大地影响细胞力学生物学，进而影响成肌结果。本研究的目的是确定二维和三维纤维蛋白支架的表观弹性特性对成肌细胞的影响，并评估这些影响在小鼠和人类细胞之间是否存在差异。因此，将成肌细胞培养在涂有纤维蛋白的多孔板上（“二维”）或嵌入具有不同弹性模量的纤维蛋白水凝胶中（“三维”）。首先，我们在7.5毫克/毫升至30毫克/毫升纤维蛋白原浓度范围内（相当于7.7 - 30.9千帕的范围）建立了水凝胶的纤维蛋白原浓度与表观弹性模量之间几乎呈线性的相关性。纤维蛋白水凝胶弹性模量对成肌细胞增殖的影响因培养类型（二维与三维）而异，在三维凝胶和二维中，较高纤维蛋白原浓度时具有抑制作用。如成肌标记基因的基因表达分析和改变的肌管形态所示，在分化的成肌细胞中则表现出相反的效果。此外，与二维培养相比，三维环境中的培养减缓了增殖，对人类成肌细胞的影响更为明显。在人类成肌细胞中，掺入三维凝胶后分化潜能也受到显著损害，但在小鼠成肌细胞中则没有。通过这项研究，我们进一步深入了解了表观弹性模量和培养类型对骨骼肌组织工程方法中细胞行为和成肌结果的影响。此外，结果强调了将为小鼠细胞建立的三维培养设置参数应用于人类细胞时需要进行调整。
