Development of a novel vitrification method for chondrocyte sheets

BMC Biotechnology

Hiroshi Nagashima²  Joji Mochida⁵  Munetaka Yokoyama⁵  Suong-Hyu Hyon³  Kazuaki Matsumura⁴  Michio Sato¹  Takahiro Kanai⁶  Mami Kokubo⁵  Hitomi Matsunari²  Masahito Watanabe²  Masato Sato⁵  Miki Maehara² 

[1]Laboratory of Microbial Genetics, School of Agriculture, Meiji University, 1-1-1 Higashimita, Tama, Kawasaki, Japan

[2]Meiji University International Institute for Bio-Resource Research (MUIIBR), 1-1-1 Higashimita, Tama, Kawasaki, Japan

[3]Center for Fiber and Textile Science, Kyoto Institute of Technology, Creation Core Kyoto Mikuruma 213, Kamigyo, Kyoto, Japan

[4]School of Materials Science, Japan Advanced Institute of Science and Technology, 1-1 Asahidai, Nomi, Ishikawa, Japan

[5]Department of Orthopaedic Surgery, Surgical Science, Tokai University School of Medicine, 143 Shimokasuya, Isehara, Kanagawa, Japan

[6]Laboratory of Developmental Engineering, School of Agriculture, Meiji University, 1-1-1 Higashimita, Tama, Kawasaki, Japan

关键词: Cartilage repair;    Cryopreservation;    Vitrification;    Chondrocyte sheet;    Cell sheet therapy;   

Others  :  1121367
DOI  :  10.1186/1472-6750-13-58

 received in 2013-04-17, accepted in 2013-07-22,  发布年份 2013

【 摘 要 】

Background

There is considerable interest in using cell sheets for the treatment of various lesions as part of regenerative medicine therapy. Cell sheets can be prepared in temperature-responsive culture dishes and applied to injured tissue. For example, cartilage-derived cell sheets are currently under preclinical testing for use in treatment of knee cartilage injuries. The additional use of cryopreservation technology could increase the range and practicality of cell sheet therapies. To date, however, cryopreservation of cell sheets has proved impractical.

Results

Here we have developed a novel and effective method for cryopreserving fragile chondrocyte sheets. We modified the vitrification method previously developed for cryopreservation of mammalian embryos to vitrify a cell sheet through use of a minimum volume of vitrification solution containing 20% dimethyl sulfoxide, 20% ethylene glycol, 0.5 M sucrose, and 10% carboxylated poly-L-lysine. The principal feature of our method is the coating of the cell sheet with a viscous vitrification solution containing permeable and non-permeable cryoprotectants prior to vitrification in liquid nitrogen vapor. This method prevented fracturing of the fragile cell sheet even after vitrification and rewarming. Both the macro- and microstructures of the vitrified cell sheets were maintained without damage or loss of major components. Cell survival in the vitrified sheets was comparable to that in non-vitrified samples.

Conclusions

We have shown here that it is feasible to vitrify chondrocyte cell sheets and that these sheets retain their normal characteristics upon thawing. The availability of a practical cryopreservation method should make a significant contribution to the effectiveness and range of applications of cell sheet therapy.

【 授权许可】

   
2013 Maehara et al.; licensee BioMed Central Ltd.

【 图 表 】

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