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Demonstration of transdifferentiation of other tissue stem cells into dental pulp stem cells by a pulp biomarker

通过牙髓生物标志物证明其他组织干细胞转分化为牙髓干细胞

基本信息

批准号：
20390504
负责人：
NAKASHIMA Misako
金额：
$ 12.48万
依托单位：
National Institute for Longevity Sciences,NCGG
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2008
资助国家：
日本
起止时间：
2008 至 2010
项目状态：
已结题

项目摘要

The present study aimed at identification of a specific biomarker for dental pulp stem cells by systemic protein analysis by differential proteomic expression profiles of dental pulp stem cells and other tissue stem cells. As a result, a new method should be established to induce the dental pulp stem cells from other stem cells and obtain a new cell source with safety and no invasive method. First of all, dental pulp cells were isolated from the extracted tooth pulp after getting the patient's agreement, and CD105^+ cells were isolated by flow cytometry. Moreover, bone marrow CD105^+ cells and amnion CD105+ cells were isolated from a human bone marrow cells and amnion mesenchymal stem cells. After the protein was extracted from these three types of cells respectively, patterns were compared after silver staining in the two-dimensional electrophoretic gels. Nine protein spots were detected only in pulp CD105^+ cells. The proteins were identified by nano liquid chromatography-mass spectrometry (nano-LC/MS) and database searching. They were excised, and the reduction alkylation and the tryptic digestion were done. FK506 binding protein and unnamed protein product of the intermediate filament, were detected. One of the proteins identified with high confidence in pulp CD105+ cells was vimentin. Expression of vimentin mRNA was highest in pulp tissue compared with other variety of tissues, 18 human main tissues and 2 embryonic tissues. Next, interstitial pulp cells were strongly immunostained and the periodontal ligament cells were weakly immunostained with vimentin. All the regenerated tissues after transplantation of the dental pulp stem cells into root canal were similarly immunostained with vimentin. Thus, vimentin can serve as an useful marker for pulp regeneration.

本研究旨在通过系统蛋白质组学分析牙髓干细胞和其他组织干细胞的差异表达谱来鉴定牙髓干细胞的特异性生物标志物。因此，需要建立一种新的方法，从其他干细胞中诱导出牙髓干细胞，获得一种安全、无创的新的细胞来源。首先，在征得患者同意后，从拔出的牙髓中分离牙髓细胞，并通过流式细胞术分离CD 105 ^+细胞。此外，从人骨髓细胞和羊膜间充质干细胞分离骨髓CD 105+细胞和羊膜CD 105+细胞。分别从这三种细胞中提取蛋白质后，在双向电泳凝胶中进行银染色，比较其图谱。9个蛋白质点仅在牙髓CD 105 ^+细胞中检测到。通过纳米液相色谱-质谱（nano-LC/MS）和数据库检索鉴定蛋白质。将它们切除，并进行还原烷基化和胰蛋白酶消化。检测到FK 506结合蛋白和中间丝的未命名蛋白产物。在牙髓CD 105+细胞中以高置信度鉴定的蛋白质之一是波形蛋白。波形蛋白mRNA在牙髓组织中的表达量最高，其次是18种人体主要组织和2种胚胎组织。接下来，间质牙髓细胞被强烈免疫染色，牙周膜细胞被弱免疫染色与波形蛋白。所有牙髓干细胞移植后的再生组织均进行波形蛋白免疫组化染色。因此，波形蛋白可以作为牙髓再生的有用标记物。