There is growing interest in the functional roles of the extracellular matrix (ECM) in regulating the fate of pluripotent stem cells (PSCs). An artificially bioengineered ECM provides an excellent model for studying the molecular mechanisms underlying self‐renewal and differentiation of PSCs, without multiple unknown and variable factors associated with natural substrates. Here, we have engineered multifunctional fusion proteins that are based on peptides from laminin, including p20, RGD, and elastin‐like polypeptide (ELP), where laminin peptides work as cell adhesion molecules (CAMs) and ELP to promote anchorage. The functionality of these chimeric proteins, referred to as ERE‐p20 and E‐p20, was assessed by determining their ability to immobilize cells on a hydrophobic polystyrene surface, improve mouse induced pluripotent stem cells (miPSCs) attachment, and promote miPSC differentiation to neural progenitors. ERE‐p20 and E‐p20 proteins showed hydrophobic binding saturation to the polystyrene plates around 500 nM (2.39 μg/cm²) and 750 nM (2.27 μg/cm²) protein concentrations, respectively. The apparent maximum cell binding to ERE‐p20 and E‐p20 was approximately 81% and 73%, respectively, relative to gelatin. For neural precursors, neurite outgrowth was enhanced by the presence of RGD and p20 peptides. The expression levels of neuronal marker protein MAP2 were upregulated approximately 2.5‐fold and threefold by ERE‐p20 and E‐p20, respectively, relative to laminin. Overall, we have shown that elastin‐mimetic fusion proteins consisting of p20 with and without RGD peptides are able to induce neuronal differentiation. In conclusion, our newly designed bioengineered fusion proteins allow preparation of specific bioactive matrices or coating/scaffold for miPSCs differentiation.

中文翻译：

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构建具有层粘连蛋白衍生短肽的多功能融合蛋白以促进小鼠诱导多能干细胞的神经分化。

人们对细胞外基质 (ECM) 在调节多能干细胞 (PSC) 命运方面的功能作用越来越感兴趣。人工生物工程 ECM 为研究 PSC 自我更新和分化的分子机制提供了一个很好的模型，没有与天然底物相关的多种未知和可变因素。在这里，我们设计了基于层粘连蛋白肽的多功能融合蛋白，包括 p20、RGD 和弹性蛋白样多肽 (ELP)，其中层粘连蛋白肽作为细胞粘附分子 (CAMs) 和 ELP 来促进锚定。这些嵌合蛋白（称为 ERE-p20 和 E-p20）的功能是通过确定它们将细胞固定在疏水聚苯乙烯表面上的能力来评估的，改善小鼠诱导的多能干细胞 (miPSC) 附着，并促进 miPSC 向神经祖细胞分化。ERE-p20 和 E-p20 蛋白对聚苯乙烯板显示疏水结合饱和度约为 500 nM (2.39 μg/cm² ) 和 750 nM (2.27 μg/cm ² ) 蛋白质浓度，分别。相对于明胶，与 ERE-p20 和 E-p20 的表观最大细胞结合率分别约为 81% 和 73%。对于神经前体，RGD 和 p20 肽的存在增强了神经突的生长。相对于层粘连蛋白，ERE-p20 和 E-p20 分别将神经元标记蛋白 MAP2 的表达水平上调了约 2.5 倍和 3 倍。总的来说，我们已经表明，由带有和不带有 RGD 肽的 p20 组成的弹性蛋白模拟融合蛋白能够诱导神经元分化。总之，我们新设计的生物工程融合蛋白允许制备特定的生物活性基质或涂层/支架，用于 miPSCs 分化。