A novel bioreactor for the generation of highly aligned 3D skeletal muscle-like constructs through orientation of fibrin via application of static strain
A novel bioreactor for the generation of highly aligned 3D skeletal muscle-like constructs through orientation of fibrin via application of static strain
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DOI：
10.1016/j.actbio.2015.06.033
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发表时间：
2015-09-15
影响因子：
9.7
通讯作者：
Fuchs, Christiane
中科院分区：
文献类型：
--
作者：
Heher, Philipp;Maleiner, Babette;Fuchs, Christiane
生成功能性仿生骨骼肌结构体仍然是骨骼肌组织工程的基本挑战之一。随着结构强烈地决定了功能的概念，无数的细胞类型、支架材料和刺激策略已经被结合起来。为了进一步优化肌肉工程结构，我们开发了一种新的生物反应器系统（MagneTissue），用于快速工程的骨骼肌样结构，目的是在结构、基因表达谱和成熟度方面与天然肌肉相似。纤维蛋白是一种天然的水凝胶，作为细胞外基质，嵌入成肌细胞通过磁力传输受到机械刺激。我们确定静态机械应变是细胞排列的触发因素，同时支架的取向也成为高度组织的纤维蛋白原纤维。这最终产生的肌管具有更成熟的表型，在肌瘤图案，直径和长度方面。在分子水平上，肌源性基因表达程序的快速进展是显而易见的，当施加应变时，肌源性测定标记MyoD和Myogenin以及Ca2+依赖性收缩结构标记TnnT1显着上调。MagneTissue生物反应器系统的主要优点是，所产生的张力并不完全依赖于细胞自身对支架锚定的反应，而是能够使结构体单独调节应变方案。在未来的工作中，这将允许在组织工程构建物的成熟过程中应用不同应变制度的机械刺激，并阐明机械转导在肌肉形成中的作用。组织工程骨骼肌结构的机械刺激是增加组织功能的一种很有前途的方法。我们开发了一种新型的基于生物反应器的3D培养系统，使用户可以应用不同的应变机制，如静态、循环或斜坡应变，以嵌入在纤维蛋白支架中的肌原性前体细胞。施加静态机械应变导致纤维蛋白原纤维沿应变轴排列，并伴随高度排列的肌管形成。此外，肌生成基因的表达模式遵循体内观察到的时间进展，当施加静态应变时，更彻底地诱导肌生成程序。最终，本研究中使用的应变方案导致更高程度的肌肉成熟，通过增强的肌体图案和增加的肌管直径和长度来证明。引入的生物反应器系统为肌肉组织工程提供了新的可能性，因为更长的培养周期和不同的菌株应用将产生具有改进功能和仿生学特征的组织工程类肌肉结构。(C) 2015材料学报Elsevier Ltd.出版。版权所有。
The generation of functional biomimetic skeletal muscle constructs is still one of the fundamental challenges in skeletal muscle tissue engineering. With the notion that structure strongly dictates functional capabilities, a myriad of cell types, scaffold materials and stimulation strategies have been combined. To further optimize muscle engineered constructs, we have developed a novel bioreactor system (MagneTissue) for rapid engineering of skeletal muscle-like constructs with the aim to resemble native muscle in terms of structure, gene expression profile and maturity. Myoblasts embedded in fibrin, a natural hydrogel that serves as extracellular matrix, are subjected to mechanical stimulation via magnetic force transmission. We identify static mechanical strain as a trigger for cellular alignment concomitant with the orientation of the scaffold into highly organized fibrin fibrils. This ultimately yields myotubes with a more mature phenotype in terms of sarcomeric patterning, diameter and length. On the molecular level, a faster progression of the myogenic gene expression program is evident as myogenic determination markers MyoD and Myogenin as well as the Ca2+ dependent contractile structural marker TnnT1 are significantly upregulated when strain is applied. The major advantage of the MagneTissue bioreactor system is that the generated tension is not exclusively relying on the strain generated by the cells themselves in response to scaffold anchoring but its ability to subject the constructs to individually adjustable strain protocols. In future work, this will allow applying mechanical stimulation with different strain regimes in the maturation process of tissue engineered constructs and elucidating the role of mechanotransduction in myogenesis.Statement of SignificanceMechanical stimulation of tissue engineered skeletal muscle constructs is a promising approach to increase tissue functionality. We have developed a novel bioreactor-based 3D culture system, giving the user the possibility to apply different strain regimes like static, cyclic or ramp strain to myogenic precursor cells embedded in a fibrin scaffold. Application of static mechanical strain leads to alignment of fibrin fibrils along the axis of strain and concomitantly to highly aligned myotube formation. Additionally, the pattern of myogenic gene expression follows the temporal progression observed in vivo with a more thorough induction of the myogenic program when static strain is applied. Ultimately, the strain protocol used in this study results in a higher degree of muscle maturity demonstrated by enhanced sarcomeric patterning and increased myotube diameter and length. The introduced bioreactor system enables new possibilities in muscle tissue engineering as longer cultivation periods and different strain applications will yield tissue engineered muscle-like constructs with improved characteristics in regard to functionality and biomimicry. (C) 2015 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.