Production of arrays of cardiac and skeletal muscle myofibers by micropatterning techniques on a soft substrate
Production of arrays of cardiac and skeletal muscle myofibers by micropatterning techniques on a soft substrate
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DOI：
10.1007/s10544-008-9245-9
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发表时间：
2009-04-01
影响因子：
2.8
通讯作者：
Elvassore, Nicola
中科院分区：
文献类型：
--
作者：
Cimetta, Elisa;Pizzato, Sara;Elvassore, Nicola
微图案化和微制造技术已被广泛用于在细胞表面图案化细胞，并更深入地了解细胞生物学中的许多过程，如细胞黏附和与周围环境的相互作用。这项研究的目的是开发一种简单和通用的技术，利用软基质上的微图案技术体外生产功能性心肌和骨骼肌肌纤维阵列。心肌细胞用于产生定向的心肌纤维，而小鼠肌肉卫星细胞用于产生分化的平行肌管。我们在光聚合到功能化玻片上的软聚丙烯酰胺水凝胶上进行了细胞外基质蛋白的微接触打印。我们的方法被证明是简单、可重复和有效的，在获得心肌细胞和卫星细胞在有图案的软水凝胶表面上的高度选择性黏附方面。培养2天后，心肌细胞形成排列整齐的心肌纤维，能够表现出同步的收缩活动。我们首次证明，在软水凝胶基质上培养的小鼠卫星细胞在培养7天后融合并形成排列的肌管。免疫荧光分析证实细胞表型、分化标志物和肌瘤组织表达正确。这些结果是在来自野生型和MDX小鼠卫星细胞的肌管中获得的，这两种细胞都是研究肌肉营养不良的研究模型。这些心肌和骨骼肌肌纤维阵列可用作单一纤维水平的药理筛选试验或生物学研究的体外模型。
Micropatterning and microfabrication techniques have been widely used to pattern cells on surfaces and to have a deeper insight into many processes in cell biology such as cell adhesion and interactions with the surrounding environment. The aim of this study was the development of an easy and versatile technique for the in vitro production of arrays of functional cardiac and skeletal muscle myofibers using micropatterning techniques on soft substrates. Cardiomyocytes were used for the production of oriented cardiac myofibers whereas mouse muscle satellite cells for that of differentiated parallel myotubes. We performed micro-contact printing of extracellular matrix proteins on soft polyacrylamide-based hydrogels photopolymerized onto functionalized glass slides. Our methods proved to be simple, repeatable and effective in obtaining an extremely selective adhesion of both cardiomyocytes and satellite cells onto patterned soft hydrogel surfaces. Cardiomyocytes resulted in aligned cardiac myofibers able to exhibit a synchronous contractile activity after 2 days of culture. We demonstrated for the first time that murine satellite cells, cultured on a soft hydrogel substrate, fuse and form aligned myotubes after 7 days of culture. Immunofluorescence analyses confirmed correct expression of cell phenotype, differentiation markers and sarcomeric organization. These results were obtained in myotubes derived from satellite cells from both wild type and MDX mice which are research models for the study of muscle dystrophy. These arrays of both cardiac and skeletal muscle myofibers could be used as in vitro models for pharmacological screening tests or biological studies at the single fiber level.