Preliminary report of novel technique for cryopreservation鈥擵acuum-assisted cryoprotectant infiltration

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• 作者：Kazutaka Soejima^a ; ^{soejima.kazutaka@nihon-u.ac.jp} ; Katsumi Shimoda^a ; Masaki Takeuchi^b ; Takashi Yamaki^b ; Taro Kono^b ; Hiroyuki Sakurai^b ; Hiroaki Nakazawa^a
• 关键词：Cryoprotective treatment ; Vacuum assistance ; Freezing damage ; Ice crystal formation
• 刊名：Cryobiology
• 出版年：2012
• 期刊代码：166_00112240
• 类别：bio
• 出版时间：August, 2012
• 卷：65
• 期：1
• 页码：21-26
• 文件大小：1581 K

摘要

To date, cryopreservation of large soft tissues has not been successfully achieved because of limitation of cryoprotective agent (CPA) infiltration into the tissue. This study aimed to investigate the effects of a vacuum on the tissue-infiltration of a CPA. An instant pickle-maker was modified for use as a vacuum apparatus, and glycerol was selected as the CPA. Twenty-six rats were used, and their thighs were divided into three treatment groups. Group 1: fresh control; Group 2: cryopreserved control, i.e., immersed in the CPA for 1 h under atmospheric pressure and cryopreserved; Group 3: vacuum-assisted CPA infiltration, i.e., immersed in the CPA under negative pressure (20, 40 and 60 cmHg, for durations of 10, 20 and 30 min at each) and cryopreserved. The Groups 2 and 3 specimens were thawed after 3 weeks of cryopreservation at 鈭?0 掳C and histologically examined, in comparison with Group 1. Skin: in Groups 2 and 3, the skin was well preserved. Muscle: in Group 2, both extracellular and intracellular ice crystal formation was widely distributed throughout the muscle tissue. In Group 3, under an adequate vacuum, the muscle tissue was well preserved, with no ice crystal formation. However, when the treatment was conducted under excessive vacuum conditions, the muscle tissue showed focal necrosis. Blood vessels: in Group 3, both the arteries and veins were well preserved up to the tunica intima. The method described in this paper may be a useful technique for achieving cryopreservation of large soft tissues.