Choice of Feeders Is Important When First Establishing iPSCs Derived From Primarily Cultured Human Deciduous Tooth Dental Pulp Cells.
Choice of Feeders Is Important When First Establishing iPSCs Derived From Primarily Cultured Human Deciduous Tooth Dental Pulp Cells.
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DOI：
10.3727/215517915x689038
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发表时间：
2015-12
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通讯作者：
I. Saitoh;E. Inada;Y. Iwase;H. Noguchi;Tomoya Murakami;Miki Soda;Naoko Kubota;H. Hasegawa;Eri Akasaka;Y. Matsumoto;K. Oka;Y. Yamasaki;H. Hayasaki;Masahiro Sato
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作者：
I. Saitoh;E. Inada;Y. Iwase;H. Noguchi;Tomoya Murakami;Miki Soda;Naoko Kubota;H. Hasegawa;Eri Akasaka;Y. Matsumoto;K. Oka;Y. Yamasaki;H. Hayasaki;Masahiro Sato
饲养细胞通常需要维持胚胎干细胞（ESC）/诱导多能干细胞（iPSC）。小鼠胚胎成纤维细胞（MEFs）和STO小鼠基质细胞系是应用最广泛的饲养细胞。本研究的目的是确定哪些细胞适合从人乳牙牙髓细胞（HDDPC）中建立iPSCs。用一种新的电穿孔方法将含有人OCT 3/4、SOX 2/KLF 4或LMYC/LIN 28和pmaxGFP的三种质粒共转染原代培养的HDDPC，然后在ESC合格培养基中培养15天。将新生集落重新接种到丝裂霉素C处理的MEF或STO细胞上。将集落连续传代最多26代。在此期间，评估集落形态以确定细胞是否表现出ESC样形态和碱性磷酸酶活性以评估细胞重编程的状态。维持在MEF上的HDDPC成功地重编程为iPSC，而维持在STO细胞上的HDDPC则没有。一旦建立，将iPSC维持在STO细胞上而不丧失多能性。我们的结果表明，MEFs是比STO细胞更好的饲养细胞，用于建立iPSC。饲养层的选择是实现iPSC高效生成的关键因素。
Feeder cells are generally required to maintain embryonic stem cells (ESCs)/induced pluripotent stem cells (iPSCs). Mouse embryonic fibroblasts (MEFs) isolated from fetuses and STO mouse stromal cell line are the most widely used feeder cells. The aim of this study was to determine which cells are suitable for establishing iPSCs from human deciduous tooth dental pulp cells (HDDPCs). Primary cultures of HDDPCs were cotransfected with three plasmids containing human OCT3/4, SOX2/KLF4, or LMYC/LIN28 and pmaxGFP by using a novel electroporation method, and then cultured in an ESC qualified medium for 15 days. Emerging colonies were reseeded onto mitomycin C-treated MEFs or STO cells. The colonies were serially passaged for up to 26 passages. During this period, colony morphology was assessed to determine whether cells exhibited ESC-like morphology and alkaline phosphatase activity to evaluate the state of cellular reprogramming. HDDPCs maintained on MEFs were successfully reprogrammed into iPSCs, whereas those maintained on STO cells were not. Once established, the iPSCs were maintained on STO cells without loss of pluripotency. Our results indicate that MEFs are better feeder cells than STO cells for establishing iPSCs. Feeder choice is a key factor enabling efficient generation of iPSCs.