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Cardiac differentiation of induced pluripotent stem cells on elastin-like protein-based hydrogels presenting a single-cell adhesion sequence
Polymer Journal ( IF 2.8 ) Pub Date : 2018-08-03 , DOI: 10.1038/s41428-018-0110-2
Yusuke Kambe , Takayuki Tokushige , Atsushi Mahara , Yasuhiko Iwasaki , Tetsuji Yamaoka

AbstractSubstrate-dependent cardiac differentiation of induced pluripotent stem cells (iPSCs) has been studied on various extracellular matrix (ECM)-derived substrates, such as collagen type I (Col-I). However, ECM-derived substrates have multiple cell-adhesive amino acid sequences and stimulate various signaling pathways in cells, making it difficult to clarify the mechanism of substrate-dependent stem cell differentiation. A substrate presenting one of these sequences is a powerful tool for elucidating the mechanism. We designed elastin-like proteins (ELPs) composed of repetitive VPGIG sequences with or without the RGD cell adhesion motif (ELP-RGD/ELP-Ctrl) and used a chemical crosslinker to generate hydrogels. By adjusting the ELP and crosslinker concentrations, we obtained ELP-Ctrl and ELP-RGD hydrogels with a Young’s modulus of 0.3 kPa. The ELP-Ctrl and ELP-RGD gels were used as a substrate for the cardiac differentiation of cultured murine iPSCs. Cells on the ELP-RGD gel showed four times higher gene expression of the contractile protein troponin T type 2 than those on a Col-I gel, which is an effective substrate for iPSC cardiac differentiation. The ELP-RGD gel might stimulate integrin-derived signaling pathways in the cells to promote cardiac differentiation. This study showed the potential of ELP hydrogels for studying substrate-dependent iPSC cardiac differentiation by enabling the control of cell-adhesive sequence presentation.We designed elastin-like proteins (ELPs) composed of repetitive VPGIG sequences with or without the RGD cell adhesion motif (ELP-RGD/ELP-Ctrl) and used tetrakis(hydroxymethyl)phosphonium chloride (THPC) as a chemical crosslinker to generate hydrogels. The ELP-RGD and ELP-Ctrl gels were used as substrates for cardiac differentiation culture of murine induced pluripotent stem cells (iPSC). Cells on the ELP-RGD gel showed four times higher expression of the contractile protein gene, troponin T type 2 (TnT2), than those on a collagen type I gel, which is an effective substrate for iPSC cardiac differentiation.

中文翻译:

在呈现单细胞粘附序列的基于弹性蛋白的水凝胶上诱导多能干细胞的心脏分化

摘要已经在各种细胞外基质 (ECM) 衍生的基质上研究了诱导多能干细胞 (iPSC) 的基质依赖性心脏分化,例如 I 型胶原蛋白 (Col-I)。然而,ECM 来源的底物具有多个细胞粘附氨基酸序列并刺激细胞内的各种信号通路,使得难以阐明底物依赖性干细胞分化的机制。呈现这些序列之一的底物是阐明机制的有力工具。我们设计了由重复 VPGIG 序列组成的类弹性蛋白 (ELP),带有或不带有 RGD 细胞粘附基序 (ELP-RGD/ELP-Ctrl),并使用化学交联剂生成水凝胶。通过调整 ELP 和交联剂浓度,我们获得了杨氏模量为 0.3 kPa 的 ELP-Ctrl 和 ELP-RGD 水凝胶。ELP-Ctrl 和 ELP-RGD 凝胶用作培养小鼠 iPSC 心脏分化的基质。ELP-RGD 凝胶上的细胞显示收缩蛋白肌钙蛋白 T 2 型的基因表达比 Col-I 凝胶上的细胞高四倍,Col-I 凝胶是 iPSC 心脏分化的有效底物。ELP-RGD 凝胶可能会刺激细胞中整合素衍生的信号通路,以促进心脏分化。这项研究显示了 ELP 水凝胶通过控制细胞粘附序列呈现来研究底物依赖性 iPSC 心脏分化的潜力。我们设计了由重复 VPGIG 序列组成的弹性蛋白样蛋白 (ELP),有或没有 RGD 细胞粘附基序。 ELP-RGD/ELP-Ctrl)并使用四(羟甲基)氯化鏻(THPC)作为化学交联剂来生成水凝胶。ELP-RGD 和 ELP-Ctrl 凝胶用作小鼠诱导多能干细胞 (iPSC) 心脏分化培养的底物。ELP-RGD 凝胶上的细胞显示收缩蛋白基因 2 型肌钙蛋白 T (TnT2) 的表达比 I 型胶原凝胶上的细胞高四倍,胶原蛋白是 iPSC 心脏分化的有效底物。
更新日期:2018-08-03
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