Human cultured epidermis accelerates wound healing regardless of its viability in a diabetic mouse model

Michiharu Sakamoto,Susumu Saito,Hiroki Yamanaka,Shuichi Ogino,Naoki Morimoto,Masukazu Inoie,Sunghee Lee,Feng Zhao,Itaru Tsuge,Yoshihiro Shimizu

doi:10.1371/journal.pone.0237985

Michiharu Sakamoto, Susumu Saito + Show 8 more

Open Access

https://doi.org/10.1371/journal.pone.0237985

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Journal: PLOS ONE	Publication Date: Aug 21, 2020
Citations: 7	License type: CC BY 4.0

Affiliation: Kyoto University, Shiga University of Medical Science

Abstract

Allogeneic cultured epidermis (allo-CE) is a cultured keratinocyte sheet manufactured from donor cells and promotes wound healing when used in deep dermal burns, donor sites, and chronic ulcers and serves as a wound dressing. Allo-CE is usually cryopreserved to be ready to use. However, the cryopreservation procedure will damage the cell viability, and the influence of Allo-CE, according to its viability or wound healing process, has not been evaluated sufficiently. In this study, we aimed to prove the influence of keratinocyte viability contained in allo-CEs on wound healing. We prepared CEs with Green’s method using keratinocytes obtained from a polydactyly patient and then prepared four kinds of CEs with different cell viabilities [fresh, cryopreserved, frozen, and FT (freeze and thaw)]. The cell viabilities of fresh, cryopreserved, frozen, and FT CEs were 95.7%, 59.9%, 16.7%, and 0.0%, respectively. The four CEs had homogeneous characteristics, except for small gaps found in the FT sheet by transmission electron microscopy observation. The four CEs were applied on the full-thickness skin defect of diabetic mice (BKS.Cg-Dock 7m +/+ Leprdb/Jcl), and the wound area and neoepithelium length were evaluated on days 4, 7, and 14. As a result, FT CEs without viable cells similarly promoted epithelialization on days 4 and 7 (p<0.05) and accelerated wound closure on day 7 (p<0.01) as fresh CEs compared with the control group. In conclusion, the promoting effect of allo-CE on wound healing does not depend on cell viability. Lyophilized CEs may be a suitable wound dressing with a long storage period at room temperature.

Highlights

Cultured epidermal autograft (CEA) is a cultured keratinocyte sheet manufactured from the patient’s skin biopsy and is effective in the treatment of severe burns [1,2]
Allogeneic cultured epidermis is a cultured keratinocyte sheet manufactured from donor cells; it can be initially prepared and stored to be applicable immediately after a burn injury
Allo-CE promotes wound healing when used in deep dermal burns [4,5,6,7,8], donor sites [9,10,11,12], and chronic ulcers [13,14,15] and serves as a wound dressing

Summary

Introduction

Cultured epidermal autograft (CEA) is a cultured keratinocyte sheet manufactured from the patient’s skin biopsy and is effective in the treatment of severe burns [1,2]. A delay of 3–4 weeks required for the generation of a CEA from the patient’s skin creates a serious clinical problem because it occurs during the life-threatening early phase in the treatment of severe. Human CE without viable cells accelerate wound healing did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The specific roles of these authors are articulated in the ‘author contributions’ section.’

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