Isolation of dental pulp stem cells from iPS cells induced by human deciduous tooth dental pulp cells using genetic engineering procedure

利用基因工程程序从人乳牙牙髓细胞诱导的 iPS 细胞中分离牙髓干细胞

• 批准号: 23792439
• 负责人: SAITOH Issei
• 金额: $ 2.75万
• 依托单位: Niigata University (2012)Kagoshima University (2011)
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Young Scientists (B)
• 财政年份: 2011
• 资助国家: 日本
• 起止时间: 2011 至 2012
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-23792439/
• 关键词: 小児歯科学; iPS細胞; 乳歯; 遺伝子工学; 歯髄幹細胞

First, we generated a set of gene-engineered STO feeder cells that confer to resistance to several commercially available drugs. The STO cells were transfected with pcBIH (carrying bleomycin resistance gene (ble) and hygromycin B phosphotransferase gene (Hyg), pcBIP (carrying ble and puromycin resistance gene (puro)) or pcBSN (carrying ble and neomycin resistance gene (neo)). The resulting stably transfectants (termed SHB for pcBIH, SPB for pcBIP and SNB for pcBSN) exhibited bleomycin/Hygromycin, bleomycin/puromycin or puromycin/neomycin, as expected. The morphology of these cells passaged over 18 generations was indistinguishable from that of parental STO cells. Of isolated clones successfully supported the growth of mouse ES cells in an undifferentiated state, when co-culture was performed. Next, we examined the possibility that STO cells could phagocytose Streptococcus mutans (a bacteria causing tooth decay), which always contaminates cultures of primarily isolated human deciduous dental pulp cells (HDDPCs). There was no bacterial contamination in the cultures containing STO cells supplemented with mitomycin C (MMC).HDDPCs can be reprogrammed using 4 reprogramming factors, and dental-pulp-stem-cell(DPSC) specific promoter was transfected to HDDPC-derived iPS. For teratoma formation,recombinant HDDPC-iPS cells were injected subcutaneously into immune compromised mice. The teratoma was dispersed into primary culture, and we tried to select DPSC under the basic medium supplemented neomycin. Unfortunately, we were unable to gain DPSC because of the uncertainty of promoter expression and differentiation in vivo. We will review our transfection procedure and try to use the materials like scaffold in the future.

首先，我们生成了一组基因工程的Sto馈线细胞，这些细胞允许对几种市售药物的抗性。 The STO cells were transfected with pcBIH (carrying bleomycin resistance gene (ble) and hygromycin B phosphotransferase gene (Hyg), pcBIP (carrying ble and puromycin resistance gene (puro)) or pcBSN (carrying ble and neomycin resistance gene (neo)). The resulting stably transfectants (termed SHB for pcBIH, SPB for PCBIP和PCBN的SNB表现出博来霉素/湿霉素，如预期的，这些细胞的形态与隔离的clone coult coult coult coult coult coult coult coult coult coult coult coult coult posploy。 STO细胞可以吞噬链球菌突变（导致牙齿衰减的细菌），这总是会污染孤立的人类落叶牙髓细胞（HDDPCS）的培养物（HDDPCS）。将特定启动子转染到HDDPC衍生的IPS。对于畸胎瘤的形成，将重组HDDPC-IPS细胞皮下注射到免疫损害的小鼠中。畸胎瘤分散到原发性培养物中，我们试图在补充新霉素的基本培养基下选择DPSC。不幸的是，由于启动子表达和体内分化的不确定性，我们无法获得DPSC。我们将审查转染程序，并尝试在将来使用脚手架之类的材料。

项目成果

小児歯科学

儿科牙科

• 发表时间: 2011
• 作者: 早崎治明，大島邦子，長谷川大子，齊藤一誠，山崎要一;稲田絵美，齊藤一誠，早崎治明，徳冨順子，佐藤秀夫，武元嘉彦，乃村俊樹，糀谷 淳，齊藤陽子，椙山加綱，山崎要一;窪田直子，深水 篤，齊藤一誠，岩崎智憲，長谷川大子，伊藤千晶，村上大輔，糀谷 淳，齊藤陽子，遠矢明菜，椙山加綱，山崎要一;有村栄次郎，余 永，稲田絵美，齊藤一誠，下田平貴子，福重雅美，北上真由美，山崎要一;早崎治明，山田千晶，大島邦子，稲田絵美，齊藤一誠，山崎要一;齊藤一誠;齊藤一誠;山崎要一,岩崎智憲,齊藤一誠
• 通讯作者: 山崎要一,岩崎智憲,齊藤一誠

Choice of feeders is important for the preparation of iPS cells from primarily cultured human deciduous tooth dental pulp cells

饲养层的选择对于从原代培养的人乳牙牙髓细胞制备 iPS 细胞很重要

• 发表时间: 2013
• 作者: Iwase Y;Saitoh I;Okamoto A;Nakakura-Ohshima K;Inada E;Yamada C;Takemoto Y;Yamasaki Y;Hayasaki H;村上智哉,齊藤一誠,稲田絵美,岩瀬陽子,長谷川大子,窪田直子,松本祐子,大島邦子,岡暁子,山崎要一,早崎治明
• 通讯作者: 村上智哉,齊藤一誠,稲田絵美,岩瀬陽子,長谷川大子,窪田直子,松本祐子,大島邦子,岡暁子,山崎要一,早崎治明

初代ヒト乳歯歯髄細胞における幹細胞特異的遺伝子の探索

在原代人乳牙牙髓细胞中寻找干细胞特异性基因

• 发表时间: 2012
• 作者: Y Nakamura;A Hatakeyama;C Kanda;M Inoue;乃村俊樹,齊藤一誠,稲田絵美,長谷川大子,松本裕子,窪田直子,山崎要一
• 通讯作者: 乃村俊樹,齊藤一誠,稲田絵美,長谷川大子,松本裕子,窪田直子,山崎要一

口腔組織からのiPS細胞の作製とその歯科領域への利用(小児歯科の観点から)

口腔组织iPS细胞的产生及其在牙科中的应用（从儿科牙科的角度）

• 发表时间: 2012
• 作者: 中村由紀;畠山文;井上誠;矢作理花;齊藤一誠
• 通讯作者: 齊藤一誠

新潟大学小児歯科学分野

新泻大学儿童牙科系