Engineering skeletal myoblasts: roles of three-dimensional culture and electrical stimulation
Engineering skeletal myoblasts: roles of three-dimensional culture and electrical stimulation
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DOI：
10.1152/ajpheart.00610.2003
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发表时间：
2005-04-01
影响因子：
4.8
通讯作者：
Niklason, LE
中科院分区：
文献类型：
--
作者：
Pedrotty, DM;Koh, J;Niklason, LE
未成熟的骨骼肌细胞或成肌细胞已被用于细胞心肌成形术，试图通过将细胞注射到受损的心肌中来再生心肌组织。在一些研究中，成肌细胞植入部位内的肌肉组织在形态上可能与心肌相似。我们假设心脏环境的可识别方面可能有助于体内植入的成肌细胞的生长和发育。为了验证这一假设，我们设计了一种新型的体外系统，以模拟天然心肌的电和生化环境的某些方面。该系统使我们能够分离可能参与成肌细胞增殖和可塑性的三维（3- D）电信号和生化信号。成肌细胞生长在3- D聚乙醇酸网状支架在控制条件下，在cardiaclike电流通量的存在下，或在存在的培养基中，已被成熟的心肌细胞条件。DNA分析表明，在三维培养中，心肌样电流可引起成肌细胞数量的增加.通过增殖细胞核抗原和TdT介导的dUTP缺口末端标记确定，细胞数量的增加是由于细胞增殖的增加而不是凋亡的差异。心肌细胞条件培养液也显著增加成肌细胞增殖。通过免疫印迹法评价了调控沿着骨骼或心脏谱系分化的转录因子的表达。虽然这些测定是定性的，但没有观察到响应电流通量的分化状态沿沿着骨骼或心脏谱系的变化。此外，从这些实验中，条件培养基似乎没有改变骨骼肌成肌细胞的分化状态。因此，心脏环境似乎刺激骨骼肌成肌细胞的增殖，但不影响分化。
Immature skeletal muscle cells, or myoblasts, have been used in cellular cardiomyoplasty in attempts to regenerate cardiac muscle tissue by injection of cells into damaged myocardium. In some studies, muscle tissue within myoblast implant sites may be morphologically similar to cardiac muscle. We hypothesized that identifiable aspects of the cardiac milieu may contribute to growth and development of implanted myoblasts in vivo. To test this hypothesis, we designed a novel in vitro system to mimic some aspects of the electrical and biochemical environment of native myocardium. This system enabled us to separate the three- dimensional ( 3- D) electrical and biochemical signals that may be involved in myoblast proliferation and plasticity. Myoblasts were grown on 3- D polyglycolic acid mesh scaffolds under control conditions, in the presence of cardiaclike electrical current fluxes, or in the presence of culture medium that had been conditioned by mature cardiomyocytes. Cardiac- like electrical current fluxes caused increased myoblast number in 3- D culture, as determined by DNA assay. The increase in cell number was due to increased cellular proliferation and not differences in apoptosis, as determined by proliferating cell nuclear antigen and TdT- mediated dUTP nick- end labeling. Cardiomyocyte- conditioned medium also significantly increased myoblast proliferation. Expression of transcription factors governing differentiation along skeletal or cardiac lineages was evaluated by immunoblotting. Although these assays are qualitative, no changes in differentiation state along skeletal or cardiac lineages were observed in response to electrical current fluxes. Furthermore, from these experiments, conditioned medium did not appear to alter the differentiation state of skeletal myoblasts. Hence, cardiac milieu appears to stimulate proliferation but does not affect differentiation of skeletal myoblasts.