Differentiation of Human Induced Pluripotent Stem Cells into Functional Enterocyte-like Cells Using a Simple Method
Differentiation of Human Induced Pluripotent Stem Cells into Functional Enterocyte-like Cells Using a Simple Method
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DOI：
10.2133/dmpk.dmpk-13-rg-005
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发表时间：
2014-02-25
影响因子：
2.1
通讯作者：
Matsunaga, Tamihide
中科院分区：
文献类型：
--
作者：
Iwao, Takahiro;Toyota, Masashi;Matsunaga, Tamihide
使用激活素A将人诱导多能干（iPS）细胞分化成内胚层，然后用成纤维细胞生长因子2（FGF 2）处理以分化成肠干细胞样细胞。这些未成熟的细胞，然后分化成肠上皮细胞样细胞使用表皮生长因子（EGF）在2%胎牛血清（FBS）。在分化的早期阶段，尾型同源异型盒2（CDX 2），一个主要的转录因子相关的肠发育和分化，和富含亮氨酸重复的G-蛋白偶联受体5（LGR 5），肠干细胞标记物的mRNA表达显着增加与FGF 2治疗。当细胞在含有EGF和低浓度FBS的培养基中培养时，肠上皮细胞的特异性标志物的mRNA表达，包括蔗糖酶-异麦芽糖酶、肠寡肽转运蛋白SLC 15 A1/肽转运蛋白1（PEPT 1）和主要代谢酶CYP 3A 4。此外，检测蔗糖酶-异麦芽糖酶蛋白表达和寡肽转运蛋白的荧光标记底物β-Ala-Lys-N-7-氨基-4-甲基香豆素-3-乙酸（β-Ala-Lys-AMCA）的摄取。这些结果证明了用于将人iPS细胞分化为功能性肠细胞样细胞的简单和直接的方法。
Human induced pluripotent stem (iPS) cells were differentiated into the endoderm using activin A and were then treated with fibroblast growth factor 2 (FGF2) for differentiation into intestinal stem cell-like cells. These immature cells were then differentiated into enterocyte-like cells using epidermal growth factor (EGF) in 2% fetal bovine serum (FBS). At the early stage of differentiation, mRNA expression of caudal type homeobox 2 (CDX2), a major transcription factor related to intestinal development and differentiation, and leucine-rich repeat-containing G-protein-coupled receptor 5 (LGR5), an intestinal stem cell marker, was markedly increased by treatment with FGF2. When cells were cultured in medium containing EGF and a low concentration of FBS, mRNAs of specific markers of intestinal epithelial cells, including sucrase-isomaltase, the intestinal oligopeptide transporter SLC15A1/peptide transporter 1 (PEPT1), and the major metabolizing enzyme CYP3A4, were expressed. In addition, sucrase-isomaltase protein expression and uptake of beta-Ala-Lys-N-7-amino-4-methylcoumarin-3-acetic acid (beta-Ala-Lys-AMCA), a fluorescence-labeled substrate of the oligopeptide transporter, were detected. These results demonstrate a simple and direct method for differentiating human iPS cells into functional enterocyte-like cells.