Department of Cardiac and Thoracic Surgery, Heart Center of the University, University of Cologne, Cologne, Germany.
Center for Molecular Medicine, University of Cologne, Cologne, Germany.
J Thorac Cardiovasc Surg. 2012 Nov;144(5):1176-1184.e1. doi: 10.1016/j.jtcvs.2012.07.036. Epub 2012 Sep 11.
The effect of mechanical preconditioning on skeletal myoblasts in engineered tissue constructs was investigated to resolve issues associated with conduction block between skeletal myoblast cells and cardiomyocytes.
Murine skeletal myoblasts were used to generate engineered tissue constructs with or without application of mechanical strain. After in vitro myotube formation, engineered tissue constructs were co-cultured for 6 days with viable embryonic heart slices. With the use of sharp electrodes, electrical coupling between engineered tissue constructs and embryonic heart slices was assessed in the presence or absence of pharmacologic agents.
The isolation and expansion procedure for skeletal myoblasts resulted in high yields of homogeneously desmin-positive (97.1% ± 0.1%) cells. Mechanical strain was exerted on myotubes within engineered tissue constructs during gelation of the matrix, generating preconditioned engineered tissue constructs. Electrical coupling between preconditioned engineered tissue constructs and embryonic heart slices was observed; however, no coupling was apparent when engineered tissue constructs were not subjected to mechanical strain. Coupling of cells from engineered tissue constructs to cells in embryonic heart slices showed slower conduction velocities than myocardial cells with the embryonic heart slices (preconditioned engineered tissue constructs vs embryonic heart slices: 0.04 ± 0.02 ms vs 0.10 ± 0.05 ms, P = .011), lower maximum stimulation frequencies (preconditioned engineered tissue constructs vs embryonic heart slices: 4.82 ± 1.42 Hz vs 10.58 ± 1.56 Hz; P = .0009), and higher sensitivities to the gap junction inhibitor (preconditioned engineered tissue constructs vs embryonic heart slices: 0.22 ± 0.07 mmol/L vs 0.93 ± 0.15 mmol/L; P = .0004).
We have generated skeletal myoblast-based transplantable grafts that electrically couple to myocardium.
研究机械预处理对工程化组织构建体中的骨骼肌成肌细胞的影响，以解决骨骼肌成肌细胞与心肌细胞之间电传导阻滞的问题。
使用鼠源性骨骼肌成肌细胞生成具有或不具有机械应变施加的工程化组织构建体。在体外肌管形成后，将工程化组织构建体与活的胚胎心脏切片共培养 6 天。使用锋利的电极，在存在或不存在药物的情况下评估工程化组织构建体与胚胎心脏切片之间的电耦合。
骨骼肌成肌细胞的分离和扩增过程产生了高产量的均一性肌联蛋白阳性（97.1%±0.1%）细胞。在基质凝胶化过程中对工程化组织构建体中的肌管施加机械应变，生成预处理的工程化组织构建体。观察到预处理的工程化组织构建体与胚胎心脏切片之间的电耦合；然而，当工程化组织构建体未受到机械应变时，没有观察到耦合。来自工程化组织构建体的细胞与胚胎心脏切片中的细胞的耦合显示出比胚胎心脏切片中的心肌细胞更慢的传导速度（预处理的工程化组织构建体与胚胎心脏切片：0.04±0.02 ms 与 0.10±0.05 ms，P=0.011），更低的最大刺激频率（预处理的工程化组织构建体与胚胎心脏切片：4.82±1.42 Hz 与 10.58±1.56 Hz；P=0.0009），以及对缝隙连接抑制剂更高的敏感性（预处理的工程化组织构建体与胚胎心脏切片：0.22±0.07 mmol/L 与 0.93±0.15 mmol/L；P=0.0004）。
我们已经生成了可电耦合到心肌的基于骨骼肌成肌细胞的可移植移植物。
