The Fragility of Cryopreserved Insulin-producing Cells Differentiated from Adipose-tissue-derived Stem Cells

• Tokuda, Kazunori

• Ikemoto, Tetsuya

• Saito, Yu

• Miyazaki, Katsuki

• Yamashita, Shoko

• Yamada, Shinichiro

• Imura, Satoru

• Morine, Yuji

• Shimada, Mitsuo

Abstract

The aim of our study is to determine whether insulin-producing cells (IPCs) differentiated from adipose-tissue-derived stem cells (ADSCs) can be cryopreserved. Human ADSCs were differentiated into IPCs using our two-step protocol encompassing a three-dimensional culture and xenoantigen-free method. Thereafter, IPCs were frozen using three different methods. First, IPCs were immediately frozen at −80°C (−80°C group). Second, IPCs were initially placed into a Bicell freezing container before freezing at −80°C (BICELL group). Third, a vitrification method for oocytes and embryos was used (CRYOTOP group). Cell counting kit-8 (CCK-8) assay showed that cell viability was decreased in all groups after cryopreservation (P < 0.01). Corroboratively, the amount of adenosine triphosphate was markedly decreased after cryopreservation in all groups (P < 0.01). Immunofluorescence staining showed a reduced positive staining area for insulin in all cryopreservation groups. Furthermore, 4′,6-diamidino-2-phenylindole and merged immunofluorescence images showed that cryopreserved cells appeared to be randomly reduced in the −80°C group and CRYOTOP group, while only the central region was visibly reduced in the BICELL group. Using immunohistochemical staining, IPCs after cryopreservation were shown to be positive for cleaved caspase-3 antibody in all groups. Finally, insulin secretion following glucose stimulation was significantly reduced in IPCs from all groups after cryopreservation (P < 0.01). In conclusion, IPCs may be too fragile for cryopreservation with accomplished methods and further investigations for a suitable preservation method are required.

Journal

• Cell Transplantation

  Cell Transplantation 29 096368972095479-, 2020-09-02

  SAGE Publications

Keywords

• insulin-producing cell
• cryopreservation
• adipose-derived stem cell
• regenerative medicine

Details

• CRID: 1050571859101284352
• NII Article ID: 120007175421
• NII Book ID: AA1084767X
• ISSN: 09636897
• ISSN: 15553892
• DOI: 10.1177/0963689720954798
• Web Site: http://repo.lib.tokushima-u.ac.jp/116513
• Web Site: http://journals.sagepub.com/doi/pdf/10.1177/0963689720954798
• Web Site: http://journals.sagepub.com/doi/full-xml/10.1177/0963689720954798
• Text Lang: en
• Article Type: journal article
• Data Source:
  • IRDB
  • Crossref
  • CiNii Articles
  • KAKEN