Defining the diversity of phenotypic respecification using multiple cell lines and reprogramming regimens.
Defining the diversity of phenotypic respecification using multiple cell lines and reprogramming regimens.
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DOI：
10.1089/scd.2013.0040
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发表时间：
2013-10
影响因子：
4
通讯作者：
Bradly Alicea;S. Murthy;S. Keaton;P. Cobbett;J. Cibelli;S. Suhr
中科院分区：
文献类型：
--
作者：
Bradly Alicea;S. Murthy;S. Keaton;P. Cobbett;J. Cibelli;S. Suhr
为了更好地了解细胞重编程的变异基础，我们进行了两个主要目标的实验：第一，在考虑技术变量后，确定类似类型的细胞系之间重编程效率的差异程度，第二，比较多个相似细胞系转换为两种不同的重编程方案-诱导神经元和诱导骨骼肌-的效率。使用两种重新编程方案，可以确定转换后的细胞是可能来自通常容易重新编程的不同亚群，还是来自具有重新指定给定表型的独立能力的细胞。我们的结果表明，当考虑到重编程方案的技术成分时，重编程效率在给定的原代成纤维细胞系中是可重现的，但在不同系之间差异很大。各行之间重新编程效率的差异足以解释已公布的结果中的一些差异。我们还发现，转换为一种表型的效率并不能预测重新编程为另一种表型的可能性，这表明重新编程的能力并不是来自于对细胞身份通常具有“弱控制”的特定亚群。我们的发现表明，为了准确评估直接重编程过程的效率，可能需要对来自几个来源的多个细胞系进行平行测试，而测试更多的成纤维细胞系--甚至是来源相似的细胞系--可能是提高重编程效率的最直接手段。
To better understand the basis of variation in cellular reprogramming, we performed experiments with two primary objectives: first, to determine the degree of difference, if any, in reprogramming efficiency among cells lines of a similar type after accounting for technical variables, and second, to compare the efficiency of conversion of multiple similar cell lines to two separate reprogramming regimens-induced neurons and induced skeletal muscle. Using two reprogramming regimens, it could be determined whether converted cells are likely derived from a distinct subpopulation that is generally susceptible to reprogramming or are derived from cells with an independent capacity for respecification to a given phenotype. Our results indicated that when technical components of the reprogramming regimen were accounted for, reprogramming efficiency was reproducible within a given primary fibroblast line but varied dramatically between lines. The disparity in reprogramming efficiency between lines was of sufficient magnitude to account for some discrepancies in published results. We also found that the efficiency of conversion to one phenotype was not predictive of reprogramming to the alternate phenotype, suggesting that the capacity for reprogramming does not arise from a specific subpopulation with a generally "weak grip" on cellular identity. Our findings suggest that parallel testing of multiple cell lines from several sources may be needed to accurately assess the efficiency of direct reprogramming procedures, and that testing a larger number of fibroblast lines--even lines with similar origins--is likely the most direct means of improving reprogramming efficiency.