The utilization of human-induced pluripotent stem cells (hiPSCs) in cell therapy has a tremendous potential but faces many practical challenges, including costs associated with cell culture media and growth factors. There is an immediate need to establish an optimized culture platform to direct the differentiation of hiPSCs into germ layers in a defined nutritional microenvironment to generate cost-effective and robust therapeutics. The aim of this study was to identify the optimal nutritional environment by mimicking the in vivo concentrations of three key factors (glucose, pyruvate, and oxygen) during the spontaneous differentiation of hiPSCs derived from cord blood, which greatly differ from the in vitro expansion and differentiation scenarios. Moreover, we hypothesized that the high glucose, pyruvate, and oxygen concentrations found in typical growth media could inhibit the differentiation of certain lineages. A design of experiments was used to investigate the interaction between these three variables during the spontaneous differentiation of hiPSCs. We found that lower oxygen and glucose concentrations enhance the expression of mesodermal (Brachyury, KIF1A) and ectodermal (Nestin, β-Tubulin) markers. Our findings present a novel approach for efficient directed differentiation of hiPSCs through the manipulation of media components while simultaneously avoiding the usage of growth factors thus reducing costs.

中文翻译：

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使用实验设计优化人诱导多能干细胞分化的营养环境——概念验证

在细胞治疗中利用人类诱导的多能干细胞 (hiPSC) 具有巨大的潜力，但面临许多实际挑战，包括与细胞培养基和生长因子相关的成本。迫切需要建立一个优化的培养平台，在特定的营养微环境中指导 hiPSCs 分化为胚层，以产生具有成本效益和稳健的治疗方法。本研究的目的是通过模拟脐带血来源的 hiPSCs 自发分化过程中三个关键因素（葡萄糖、丙酮酸和氧气）的体内浓度来确定最佳营养环境，这与体外扩增和差异化场景。此外，我们假设高葡萄糖、丙酮酸、在典型的生长培养基中发现的氧浓度可以抑制某些谱系的分化。使用实验设计来研究这三个变量在 hiPSC 自发分化过程中的相互作用。我们发现较低的氧气和葡萄糖浓度会增强中胚层（Brachyury，KIF1A）和外胚层（Nestin，β-Tubulin）标记物的表达。我们的研究结果提出了一种通过操纵培养基成分有效定向分化 hiPSC 的新方法，同时避免使用生长因子，从而降低成本。我们发现较低的氧气和葡萄糖浓度会增强中胚层（Brachyury，KIF1A）和外胚层（Nestin，β-Tubulin）标记物的表达。我们的研究结果提出了一种通过操纵培养基成分有效定向分化 hiPSC 的新方法，同时避免使用生长因子，从而降低成本。我们发现较低的氧气和葡萄糖浓度会增强中胚层（Brachyury，KIF1A）和外胚层（Nestin，β-Tubulin）标记物的表达。我们的研究结果提出了一种通过操纵培养基成分有效定向分化 hiPSC 的新方法，同时避免使用生长因子，从而降低成本。