Department of Orthodontics and Craniofacial Developmental Biology, Hiroshima University, Graduate School of Biomedical Science, 1-2-3 Kasumi, Minami-ku, Hiroshima 734-8553, Japan.
Cryobiology. 2011 Jun;62(3):181-7. doi: 10.1016/j.cryobiol.2011.03.001. Epub 2011 Mar 21.
The purpose of this study was to evaluate the effects of long-term cryopreservation on the isolated human periodontal ligament cells (PDL) and pulp tissues. In the first part of study, 10 freshly extracted teeth were selected and divided into two groups. In the cryopreserved group, the teeth were frozen for 5 years using a programmed freezer combined with a magnetic field, known as Cells Alive System "CAS". As for the control group, freshly extracted teeth were used. In each group, extracted PDL tissues were cultured and gene expression and protein concentration of collagen type I, alkaline-phosphatase (ALP) and vascular endothelial growth factor (VEGF) was compared between the two groups. In the second part, pulp tissues were obtained from 10 mature and immature third molars which were freshly extracted or cryopreserved for three months. Expression of VEGF and nerve growth factor (NGF) mRNAs and the protein concentration in the supernatant were investigated. Results indicated that long-term cryopreservation with the use of CAS freezer cannot affect the growth rate and characteristics of PDL cells. There was no significant difference in VEGF expression and VEGF and NGF protein concentration of pulp cells derived from cryopreserved teeth with immature apex and control group with mature root formation. Finally, proper PDL regeneration and appropriate apexogenesis after transplanting magnetically cryopreserved immature tooth was clinically confirmed. These findings demonstrate that teeth banking with the use of magnetic field programmed freezer can be available for future autotransplantation as a treatment modality for replacing missing teeth.
本研究旨在评估长期冷冻保存对分离的人牙周韧带细胞（PDL）和牙髓组织的影响。在研究的第一部分，选择了 10 颗新鲜拔出的牙齿，并将其分为两组。在冷冻保存组中，使用程序冷冻机结合磁场（称为“细胞活系统”CAS）将牙齿冷冻保存 5 年。对照组则使用新鲜拔出的牙齿。在每组中，培养提取的 PDL 组织，并比较两组之间胶原 I 型、碱性磷酸酶（ALP）和血管内皮生长因子（VEGF）的基因表达和蛋白浓度。在第二部分，从 10 颗成熟和未成熟的第三磨牙中获得牙髓组织，这些牙齿是新鲜拔出或冷冻保存三个月。研究了 VEGF 和神经生长因子（NGF）mRNA 的表达以及上清液中的蛋白浓度。结果表明，使用 CAS 冷冻机进行长期冷冻保存不会影响 PDL 细胞的生长速度和特性。未成熟根尖冷冻保存牙和成熟根形成对照组牙髓细胞的 VEGF 表达以及 VEGF 和 NGF 蛋白浓度没有显著差异。最后，临床证实了移植磁性冷冻保存的未成熟牙后适当的 PDL 再生和适当的根尖形成。这些发现表明，使用磁场程控冷冻机进行牙齿储存可用于未来的自体移植，作为替代缺失牙齿的治疗方法。
