The evaluation of periodontal ligament cells of rat teeth after low-temperature preservation under high pressure

Jin-Ho Chung; Jin Kim; Seong-Ho Choi; Eui-Seong Kim; Jiyong Park; Seung-Jong Lee

doi:10.5395/JKACD.2010.35.4.285

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Chung, Kim, Choi, Kim, Park, and Lee: The evaluation of periodontal ligament cells of rat teeth after low-temperature preservation under high pressure

Basic Research

Journal of Korean Academy of Conservative Dentistry

Journal of Korean Academy of Conservative Dentistry 2010; 35(4): 285-294.

Published online: 31 July 2010

DOI: https://doi.org/10.5395/JKACD.2010.35.4.285

The evaluation of periodontal ligament cells of rat teeth after low-temperature preservation under high pressure

Jin-Ho Chung¹, Jin Kim², Seong-Ho Choi³, Eui-Seong Kim¹, Jiyong Park⁴, Seung-Jong Lee¹

¹Department of Conservative Dentistry, College of Dentistry, Yonsei University, Seoul, Korea.

²Department of Oral Pathology, College of Dentistry, Yonsei University, Seoul, Korea.

³Department of Periodontology, College of Dentistry, Yonsei University, Seoul, Korea.

⁴Department of Biotechnology, College of Life Science and Biothchnology, Yonsei University, Seoul, Korea.

Corresponding Author: Seung-Jong Lee. Department of Conservative Dentistry, Yonsei University School of Dentistry 134, Shinchon-dong, Sudaemoon-gu, Seoul, 120-752, Korea. Tel: +82-2-2228-3148 Fax: +82-2-313-7575, sjlee@yuhs.ac

Received 29 May 2010 Revised 19 June 2010 Accepted 3 July 2010

Abstract

The purpose of this study was to evaluate the viability of periodontal ligament cells of rat teeth after low-temperature preservation under high pressure by means of MTT assay, WST-1 assay. 12 teeth of Sprague-Dawley white female rats of 4 week-old were used for each group.

Both side of the first and second maxillary molars were extracted as atraumatically as possible under tiletamine anesthesia. The experimental groups were group 1 (Immediate extraction), group 2 (Slow freezing under pressure of 3 MPa), group 3 (Slow freezing under pressure of 2 MPa), group 4 (Slow freezing under no additional pressure), group 5 (Rapid freezing in liquid nitrogen under pressure of 2 MPa), group 6 (Rapid freezing in liquid nitrogen under no additional pressure), group 7 (low-temperature preservation at 0℃ under pressure of 2 MPa), group 8 (low-temperature preservation at 0℃ under no additional pressure), group 9 (low-temperature preservation at -5℃ under pressure of 90 MPa). F-medium and 10% DMSO were used as preservation medium and cryo-protectant. For cryo-preservation groups, thawing was performed in 37℃ water bath, then MTT assay, WST-1 assay were processed. One way ANOVA and Tukey HSD method were performed at the 95% level of confidence. The values of optical density obtained by MTT assay and WST-1 were divided by the values of eosin staining for tissue volume standardization.

In both MTT and WST-1 assay, group 7 (0℃/2 MPa) showed higher viability of periodontal ligament cells than other group (2-6, 8) and this was statistically significant (p < 0.05), but showed lower viability than group 1, immediate extraction group (no statistical significance).

By the results of this study, low-temperature preservation at 0℃ under pressure of 2 MPa suggest the possibility for long term preservation of teeth.

Keywords: Periodontal ligament cell, MTT, WST-1, Viability, Low temperature preservation under pressure, Rapid freezing, Cryopreservation

Figures and Tables

Figure 1

Schematic diagram of program freezer with pressure vessel.

a. Oxygen container: 2 or 3 MPa of pressure, b. Program freezer, c. Pressure bottle, d. 2 ml Cryotube: 1 ml F medium + 10% DMSO, e. Tooth treated 10% DMSO, f. Pressure valve, g. Thermometer.

Table 1

Experimental Groups

Table 2

The averages and standard deviations of optical density of MTT

a,b,c,d,e,f: Different superscript letters means statistical difference (p < 0.05).

Table 3

The averages and standard deviations of optical density of WST-1

a,b,c,d,e,f: Different superscript letters means statistical difference (p < 0.05).

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