Sufficient therapeutic effect of cryopreserved frozen adipose-derived regenerative cells on burn wounds

Yasuhiko Kaita; Takehiko Tarui; Hideaki Yoshino; Takeaki Matsuda; Yoshihiro Yamaguchi; Takatoshi Nakagawa; Michio Asahi; Masaaki Ii

doi:10.1016/j.reth.2019.01.001

Sufficient therapeutic effect of cryopreserved frozen adipose-derived regenerative cells on burn wounds

Regen Ther. 2019 Feb 1:10:92-103. doi: 10.1016/j.reth.2019.01.001. eCollection 2019 Jun.

Authors

Yasuhiko Kaita¹, Takehiko Tarui¹, Hideaki Yoshino², Takeaki Matsuda¹, Yoshihiro Yamaguchi¹, Takatoshi Nakagawa³, Michio Asahi³, Masaaki Ii⁴

Affiliations

¹ Department of Trauma and Critical Care Medicine, Kyorin University School of Medicine, Tokyo, Japan.
² Division of Cardiology, Second Department of Internal Medicine, Kyorin University, School of Medicine, Tokyo, Japan.
³ Department of Pharmacology, Faculty of Medicine, Osaka Medical College, Osaka, Japan.
⁴ Division of Research Animal Laboratory and Translational Medicine, Research and Development Center, Osaka Medical College, Osaka, Japan.

Abstract

Introduction: The purpose of this study was to evaluate whether cryopreserved (frozen) adipose-derived regenerative cells (ADRCs) have a therapeutic effect on burn wound healing as well as freshly isolated (fresh) ADRCs.

Methods: Full thickness burns were created on dorsum of nude mice and burn wound was excised. The wound was covered by artificial dermis with; (i) fresh ADRCs, (ii) frozen ADRCs, and (iii) PBS (control). The assessment for wound healing was performed by morphological, histopathological and immunohistochemical analyses.

Results: In vivo analyses exhibited the significant therapeutic effect of frozen ADRCs on burn wound healing up to the similar or higher level of fresh ADRCs. There were significant differences of wound closure, epithelized tissue thickness, and neovascularization between the treatment groups and control group. Although there was no significant difference of therapeutic efficacy between fresh ADRC group and frozen ADRC group, frozen ADRCs improved burn wound healing process in dermal regeneration with increased great type I collagen synthesis compared with fresh ADRCs.

Conclusions: These findings indicate that frozen ADRCs allow us to apply not only quickly but also for multiple times, and the cryopreserved ADRCs could therefore be useful for the treatment of burn wounds in clinical settings.

Keywords: ADRCs, adipose-derived regenerative cells; AdSCs, adipose-derived stem cells; Burn; CM, conditioned medium; DAPI, 4′,6-diamidino-2-phenylindole; DMEM-F12, Dulbecco's modified Eagle's medium supplemented with F12; EGF, Epidermal growth factor; FBS, Fetal bovine serum; FGF2, Fibroblast growth factor 2; GAPDH, Glyceraldehyde 3-phosphate dehydrogenase; HGF, Hepatocyte growth factor; IGF-1, Insulin-like growth factor-1; ILB4, Isolectin B4; MSC, Mesenchymal stem cell; NHDF, Normal human dermal fibroblast; NHEK, Normal human epithelial keratinocyte; PBS, Phosphate-buffered saline; PVDF, Poly vinylidene di-fluoride; SDS-PAGE, Sodium dodecyl sulfate-polyacrylamide gel electrophoresis; SVF, Stromal vascular fraction; Stem cells; VEGF, Vascular endothelial growth factor; Wound healing; qRT-PCR, Quantitative real-time reverse transcription polymerase chain reaction.