Muscle-derived stem cells in tissue engineering:: defining cell properties suitable for construct design
Muscle-derived stem cells in tissue engineering:: defining cell properties suitable for construct design
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DOI：
10.14670/hh-20.891
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发表时间：
2005-07-01
影响因子：
2
通讯作者：
Bellón, JM
中科院分区：
文献类型：
--
作者：
Buján, J;Pascual, G;Bellón, JM
术语“构造”或“组织等同物”是指通过组织工程技术产生的新组织。构成结构的元素是支架，细胞在其上被“再造”，形成对某些细胞信号敏感的工程组织。细胞在支架上增殖和分化的能力是由固定、粘附、增殖和迁移等特性决定的。在最有希望用于设计结构的细胞类型中，组织居住干细胞或成体干细胞表现出两个主要特征：分化成许多细胞系的能力和自我更新的能力。这些特征使它们成为细胞替代疗法的良好候选者。在这里，我们报告了肌肉干细胞的鉴定、分离和培养，旨在建立理想的培养，以确定培养的细胞群何时能够表现出转移到支架上以获得特定结构的最佳特性。从新西兰白兔背肌中提取干细胞，体外培养5 ~ 14天。从同一实验动物身上获得的成纤维细胞作为对照。用光镜和扫描电镜观察干细胞。对于干细胞鉴定，我们使用了抗m-cadherin、抗cd34和抗myf -5抗体。使用的肌肉分化标志物为：抗波形蛋白。抗a-actin，抗desmin和抗myosin。Western blotting检测不同肌肉分化标志物及TGF β 1在细胞培养中的表达谱。通过监测氚化胸腺嘧啶掺入来测定增殖率。胸腺嘧啶掺入率在未分化细胞群体中明显高于从同一动物获得的对照成纤维细胞。在培养的前五天，除了m-cadherin， CD34和Myf-5外，大多数细胞对所有检测的标记物都是阴性的，尽管开始出现vimentin表达的离散迹象。培养14天后，成体干细胞显示vimentin（94.2%）和desmin（33.8%）的表达，而myosin（16.2%）和α -actin（8.3%）的标记较少。对照成纤维细胞表现出强烈的vimentin（99.3%）和α -actin（62.2%）标记，而不到2%的人群表达myosin（0.9%）和desmin（1.6%）。经过两周的培养，肌肉来源的干细胞在开始分化时表现出良好的增殖和粘附特性。这些条件似乎是获得所需构造的理想条件。
The terms construct or tissue equivalent refer to neotissue produced by tissue engineering techniques. The elements forming the construct are scaffolds on which cells are "recreated" to form an enginnered-tissue sensitive to certain cell signals. The ability of the cells to expand and differentiate on the scaffold is determined by properties such as fixation, adhesion, proliferation and migration. Among the cell types that seem to be most promising for designing constructs are tissue-residing, or adult, stem cells, which show two main features: a capacity to differentiate into many cell lineages and the power of self-renewal. These features make them good candidates for cell replacement therapies. Here, we report the identification, isolation and culture of muscle stem cells aimed at establishing the ideal culture in terms of defining when the cultured cell population would show optimal characteristics for transfer to the scaffold to obtain a particular construct. Stem cells harvested from the dorsal muscle of white New Zealand rabbits were cultured in vitro and characterized 5 to 14 days after the start of culture. Fibroblasts obtained from the same experimental animal served as controls. The stem cells were examined by light and scanning electron microscopy. For stem cell identification, we used the antibodies anti-m-cadherin, anti-CD34 and anti-Myf-5. The markers of muscle differentiation used were: anti-vimentin. anti-a-actin, anti-desmin and anti-myosin. The expression profiles of the different markers of muscle differentiation and TGF beta 1 in the cell cultures were confirmed by Western blotting. Proliferation rates were determined by monitoring tritiated thymidine incorporation.The thymidine incorporation rate was substantially higher for the population of undifferentiated cells than for control fibroblasts obtained from the same animal. During the first five days of culture, most cells were negative for all the markers examined, with the exception of m-cadherin, CD34 and Myf-5, although discrete signs of vimentin expression started to emerge. After 14 days of culture, the adult stem cells showed vimentin (94.2%) and desmin (33.8%) expression yet scarce labeling for myosin (16.2%) and alpha-actin (8.3%). Control fibroblasts showed intense labeling for vimentin (99.3%) and alpha-actin (62.2%), while less than 2% of the population expressed myosin (0.9%) and desmin (1.6%).After two weeks of culture, muscle-derived stem cells show good proliferative and adhesion properties as they initiate differentiation. These conditions seem ideal for obtaining the desired construct.