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Mater Sci Eng C Mater Biol Appl. 2019 Nov;104:109841. doi: 10.1016/j.msec.2019.109841. Epub 2019 Jun 6.

Enhanced decellularization technique of porcine dermal ECM for tissue engineering applications.

Author information

1
Department of Regenerative Medicine, College of Medicine, Soonchunhyang University, Cheonan, South Korea.
2
Institute of Tissue Regeneration, College of Medicine, Soonchunhyang University, Cheonan, South Korea.
3
Department of Regenerative Medicine, College of Medicine, Soonchunhyang University, Cheonan, South Korea; Institute of Tissue Regeneration, College of Medicine, Soonchunhyang University, Cheonan, South Korea. Electronic address: lbt@sch.ac.kr.

Abstract

Effective removal of cellular components while retaining extracellular matrix (ECM) proteins is the ultimate goal of decellularization. The aim of this study is to produce a decellularized ECM with highly preserved ECM proteins and to determine the effect of isopropanol as a decellularization solvent on the characteristics of the decellularized porcine skin. Two different protocols were used for porcine skin decellularization. Protocol 1 consisted of Triton-X and sodium dodecyl sulfate (SDS) in water while protocol 2 consisted of Triton-X and SDS in 70% isopropanol. After decellularization, DNA components decreased significantly in protocol 2 with lower amount of lipid content and higher ECM proteins such as collagen (92.91 ± 9.02 μg/mg sample), α-elastin (142.32 ± 6.74 μg/mg sample) and sulfated glycosaminoglycan (sGAG; 7.44 ± 1.30 μg/mg sample) compared with protocol 1 ECM. Higher amount of vascular endothelial growth factor (VEGF; 11.26 ± 0.44 pg/mg sample) content was quantified in protocol 2 compared with protocol 1 while higher trace amount of bone morphogenic protein 2 (BMP-2; 0.28 ± 0.04 pg/mg sample) was also observed in protocol 2 compared with protocol 1. Protocol 2 ECM did not significantly affect the cell viability and exhibited no cytotoxicity when exposed to three different cell lines: L929 fibroblast cells, MC3T3-E1 pre-osteoblast cells, and rat mesenchymal stem cells (BMSC). Subcutaneous implantation after 7 and 21 days revealed higher cell infiltration in protocol 2 ECM and enhanced neovascularization. Isopropanol/surfactants proved to be effective in cell and lipid removal during decellularization while preserving the higher amount of ECM proteins.

KEYWORDS:

Decellularization; Extracellular matrix; Isopropanol; Porcine skin

PMID:
31499993
DOI:
10.1016/j.msec.2019.109841

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