A Simple Method for Cryopreservation of Human Periodontal Ligament Cells in a Vitrification Solution Containing 20% Ethylene Glycol and 20% Dimethyl Sulfoxide

作者：Kim Yemi; Kim Myung Rae; Ahn Jung Hyuck; Kim Sun Jong^*

来源：Tissue Engineering and Regenerative Medicine, 2011, 8(4): 415-421.

• cryopreservation
• tooth banking
• vitrification
• periodontal ligament cells
• cell viability

Vitrification approaches for the cryopreservation of human periodontal ligament (PDL) cells have not yet been successfully standardized, although vitrification has been widely used in various cells and tissues. The aims of this study were to examine whether vitrification approaches could be used for the cryopreservation of human PDL cells and to establish the most suitable vitrification formula for tooth cryopreservation. Human PDL cells were cryopreserved with 4 solutions: VS1, 20% (v/v) ethylene glycol (EG) and 20% dimethyl sulfoxide (Me(2)SO); VS2, 40% EG; VS3, 20% (v/v) 1,2-propanediol (PROH) and 20% (v/v)Me(2)SO; and VS4, 40% PROH. All vitrification solutions contained 0.6 M sucrose and 18% (w/v) Ficoll 70 as non-permeating cryoprotectants. Cells were equilibrated in a 25% concentration of each vitrification solution for 3 minutes at room temperature and exposed to each full strength solution at 4 degrees C. The cells were then immediately transferred to liquid nitrogen and cryopreserved for 7 days. After stepwise warming with 0.5, 0.25 and 0.125 M sucrose and F-medium, cell viability was measured using trypan blue exclusion, MTT assay, Cell Counting Kit-8 assay and morphological appearance. Cell apoptosis was assessed using TUNEL assay and Annexin V assay. VS1 led to 93% viability which is significantly higher than other groups. Morphological appearance and the percentage of apoptotic cells were similar between the VS1 and control groups. These results indicate that the vitrification method may be applicable to the cryopreservation of PDL cells and that VS1 can be used in successful tooth cryopreservation.

• 出版日期2011-8