Department of Clinical Pharmacy, Graduate School of Pharmaceutical Sciences, Nagoya City University.
Drug Metab Pharmacokinet. 2014;29(1):44-51. doi: 10.2133/dmpk.dmpk-13-rg-005. Epub 2013 Jul 2.
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 β-Ala-Lys-N-7-amino-4-methylcoumarin-3-acetic acid (β-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.
人诱导多能干细胞（iPS）经激活素 A 分化为内胚层，然后用成纤维细胞生长因子 2（FGF2）处理分化为肠干细胞样细胞。这些未成熟的细胞然后在 2%胎牛血清（FBS）中使用表皮生长因子（EGF）分化为肠上皮样细胞。在分化的早期阶段，通过用 FGF2 处理，与肠发育和分化相关的主要转录因子尾型同源盒 2（CDX2）和肠干细胞标志物富含亮氨酸重复的 G 蛋白偶联受体 5（LGR5）的 mRNA 表达明显增加。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 mRNAs，包括蔗糖酶-异麦芽糖酶、肠寡肽转运体 SLC15A1/肽转运体 1（PEPT1）和主要代谢酶 CYP3A4，表达。此外，检测到蔗糖酶-异麦芽糖酶蛋白表达和β-Ala-Lys-N-7-氨基-4-甲基香豆素-3-乙酸（β-Ala-Lys-AMCA）的摄取，β-Ala-Lys-AMCA 是寡肽转运体的荧光标记底物。这些结果表明了一种将人 iPS 细胞分化为功能性肠上皮样细胞的简单而直接的方法。
