Unique Differentiation Profile of Mouse Embryonic Stem Cells in Rotary and Stirred Tank Bioreactors
Unique Differentiation Profile of Mouse Embryonic Stem Cells in Rotary and Stirred Tank Bioreactors
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DOI：
10.1089/ten.tea.2010.0166
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发表时间：
2010-11-01
影响因子：
4.1
通讯作者：
Roy, Krishnendu
中科院分区：
文献类型：
--
作者：
Fridley, Krista M.;Fernandez, Irina;Roy, Krishnendu
胚胎干（ES）细胞来源的谱系特异性干细胞，例如造血干细胞，可以为可移植细胞提供潜在的无限来源，特别是对于基于细胞的疗法。然而，必须开发可重复的方法来最大化和扩大ES细胞分化，以产生临床相关数量的治疗性细胞。基于生物反应器的动态培养条件适合于大规模细胞生产，但很少有研究评估各种生物反应器类型和培养参数如何影响ES细胞分化，特别是造血。我们的研究结果表明，细胞接种密度和生物反应器的速度显着影响拟胚体的形成和随后的造血干细胞和祖细胞在搅拌罐（转瓶）和旋转微重力（Synthecon（TM））型生物反应器的一代。一般而言，在两个生物反应器中产生高百分比的造血干细胞和祖细胞，尤其是在高细胞密度下。此外，Synthecon生物反应器产生了更多的sca-1（+）祖细胞，转瓶产生了更多的c-Kit（+）祖细胞，证明了它们独特的分化特征。参与多能性，胚层形成和造血分化的基因的cDNA微阵列分析表明，在分化的第7天，来自两个生物反应器的胚状体由胚胎发育的所有三个胚层组成。然而，在两个生物反应器中观察到独特的基因表达谱;例如，在Synthecon培养物中特异性造血基因的表达比在转瓶中显著上调。我们的结论是，生物反应器类型和培养参数可用于控制ES细胞分化，增强独特的祖细胞群，并为大规模生产可移植治疗性细胞提供手段。
Embryonic stem (ES)-cell-derived lineage-specific stem cells, for example, hematopoietic stem cells, could provide a potentially unlimited source for transplantable cells, especially for cell-based therapies. However, reproducible methods must be developed to maximize and scale-up ES cell differentiation to produce clinically relevant numbers of therapeutic cells. Bioreactor-based dynamic culture conditions are amenable to large-scale cell production, but few studies have evaluated how various bioreactor types and culture parameters influence ES cell differentiation, especially hematopoiesis. Our results indicate that cell seeding density and bioreactor speed significantly affect embryoid body formation and subsequent generation of hematopoietic stem and progenitor cells in both stirred tank (spinner flask) and rotary microgravity (Synthecon (TM)) type bioreactors. In general, high percentages of hematopoietic stem and progenitor cells were generated in both bioreactors, especially at high cell densities. In addition, Synthecon bioreactors produced more sca-1(+) progenitors and spinner flasks generated more c-Kit(+) progenitors, demonstrating their unique differentiation profiles. cDNA microarray analysis of genes involved in pluripotency, germ layer formation, and hematopoietic differentiation showed that on day 7 of differentiation, embryoid bodies from both bioreactors consisted of all three germ layers of embryonic development. However, unique gene expression profiles were observed in the two bioreactors; for example, expression of specific hematopoietic genes were significantly more upregulated in the Synthecon cultures than in spinner flasks. We conclude that bioreactor type and culture parameters can be used to control ES cell differentiation, enhance unique progenitor cell populations, and provide means for large-scale production of transplantable therapeutic cells.