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. 2024 Jun;25(2):685-695.
doi: 10.1007/s10561-024-10131-6. Epub 2024 Feb 21.

Development and validation of cryopreserved or freeze-dried decellularized human dermis for transplantation

Affiliations

Development and validation of cryopreserved or freeze-dried decellularized human dermis for transplantation

Giulia Montagner et al. Cell Tissue Bank. 2024 Jun.

Abstract

For decades, dermal tissue grafts have been used in various regenerative, reconstructive, and augmentative procedures across the body. To eliminate antigenicity and immunogenic response while still preserving the individual components and collective structural integrity of the extracellular matrix (ECM), dermis can be decellularized. Acellular dermal matrix (ADM) products like such are produced to accurately serve diverse clinical purposes. The aim of the present study is to evaluate the efficacy of a novel decellularization protocol of the human dermis, which eliminates residual human genetic material without compromising the biomechanical integrity and collagenous content of the tissue. Moreover, a freeze-drying protocol was validated. The results showed that though our decellularization protocol, human dermis can be decellularized obtaining a biocompatible matrix. The procedure is completely realized in GMP aseptic condition, avoiding tissue terminal sterilization.

Keywords: Acellular dermal matrix; Allograft; Decellularization; Decellularized dermis; Tissue bank.

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Conflict of interest statement

The authors have no competing interests to declare that are relevant to the content of this article.

Figures

Fig. 1
Fig. 1
Uniaxial tensile tests setup (a). Mechanical parameters extracted from a typical stress–strain curve (b)
Fig. 2
Fig. 2
Haematoxylin and eosin staining of representative histological sections of non-decellularized dermis (A) and decellularized dermis (B), 10 × magnification. Absence of nuclei is demonstrated in decellularized samples. Extracellular matrix is similar between non-decellularized dermis (A) and decellularized dermis (B). Nuclear fluorescence staining with Hoechst in representative samples of non-decellularized dermis (C) and decellularized dermis (D). Nuclei are absent in decellularized sample. 20 × magnification
Fig. 3
Fig. 3
DNA content in four batches of dermis (native and decellularized) obtained from four different donors. The residual DNA in decellularized dermis was reduced to less than 10% with respect to the DNA quantity of native dermis; the reduction depicted is statistically significant (p < 0.05)
Fig. 4
Fig. 4
Electrophoresis analysis of nuclei acids. The image demonstrates the presence of DNA in non-decellularized samples (2, 4, 6) and the absence of nucleic acids fragments in decellularized samples (3, 5, 7). Samples were compared to a marker (1)
Fig. 5
Fig. 5
Representative figures of contact cytotoxicity test. Fibroblasts BJ CRL-2522 were grown in absence of cytotoxic agent (a), in presence of a cytotoxic agent (b, cyanoacrylate glue), in presence of non-decellularized dermis (c) or decellularized dermis (d). Cells were visualized with Giemsa stain at light microscopy and 10 × magnification
Fig. 6
Fig. 6
Viability of fibroblasts BJ CRL-2522 cultured in preconditioned medium with 0,1 g/ml and 0,2 g/ml of native dermis or decellularized dermis. Three batches of both native and decellularized samples obtained from three different donors were tested. No cytotoxicity occured, since any reduction of cell viability by more of 30% was observed
Fig. 7
Fig. 7
Mechanical tests results: ultimate tensile strength (a), elongation at break (b), linear elastic modulus (c), and toe elastic modulus (d). Asterisks above bars indicate statistically significant differences (p < 0.5)

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