Gioventù Silvia, Andriolo Gabriella, Bonino Ferruccio, Frasca Stefania, Lazzari Lorenza, Montelatici Elisa, Santoro Franco, Rebulla Paolo
Department of Surgical Sciences, Reconstruction and Diagnostics, University of Milan, Faculty of Dentistry, IRCCS Fondazione Ca' Granda Ospedale Maggiore Policlinico, via Francesco Sforza 35, 20122 Milano, Italy.
Transfus Apher Sci. 2012 Oct;47(2):199-206. doi: 10.1016/j.transci.2012.06.005. Epub 2012 Jul 11.
Dental pulp stem cells (DPSC), a cell type of mesenchymal origin showing high proliferation and plasticity, are an emerging source of adult stem cells offering interesting features in view of potential applications in regenerative medicine. These features prompted us to develop a new method to cryopreserve DPSC inside a whole tooth, thus avoiding the need to purify the cells before cryopreservation and reducing the initial costs and workload of tooth banking. In this study we cryopreserved 4 human deciduous whole teeth after digging micro-channels into the tooth with an Nd:YAG laser beam (laser piercing) to allow the cryopreservative to reach the dental pulp and preserve the cells at -80°C. Then, we isolated, expanded and characterized in vitro the stem cells after tooth thawing and mechanical fracture. In parallel, we characterized cells extracted from 2 teeth cryopreserved without laser piercing and from 4 non cryopreserved, non laser pierced, freshly fractured teeth. Our data demonstrate that DPSC isolated from laser pierced cryopreserved teeth show mesenchymal stem cells morphology, immunophenotype, viability and proliferation rate similar to those of cells isolated from fresh, non cryopreserved teeth, whereas significant loss of cell viability and proliferation rate was shown by cells isolated from teeth cryopreserved without laser piercing. These data support the use of this method for prospective whole tooth banking.
牙髓干细胞（DPSC）是一种间充质来源的细胞类型，具有高增殖能力和可塑性，鉴于其在再生医学中的潜在应用，它是一种新兴的成体干细胞来源。这些特性促使我们开发一种新方法，将DPSC冷冻保存在完整牙齿内部，从而避免在冷冻保存前对细胞进行纯化，并降低牙齿库的初始成本和工作量。在本研究中，我们用Nd:YAG激光束在牙齿上钻出微通道（激光穿孔），使冷冻保护剂能够到达牙髓，并在-80°C下保存细胞，之后对4颗人类乳牙进行了冷冻保存。然后，在牙齿解冻和机械破碎后，我们对干细胞进行了体外分离、扩增和表征。同时，我们对从2颗未经激光穿孔冷冻保存的牙齿以及4颗未冷冻保存、未激光穿孔、新鲜破碎的牙齿中提取的细胞进行了表征。我们的数据表明，从激光穿孔冷冻保存的牙齿中分离出的DPSC表现出与从新鲜的、未冷冻保存的牙齿中分离出的细胞相似的间充质干细胞形态、免疫表型、活力和增殖率，而从未经激光穿孔冷冻保存的牙齿中分离出的细胞则表现出明显的细胞活力和增殖率损失。这些数据支持将该方法用于前瞻性的全牙库保存。
