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Introduction: Several scaffolds have been introduced for urethral tissue engineering. However, acellular human urethral scaffold harvested from deceased donors may provide significant advantages in comparison to synthetic, composite or other biological scaffolds. The aim of this study was to develop a protocol for decellularization of human urethra that preserves substantial components of extracellular matrix (ECM), which are essential for subsequent recellularization mimicking the natural environment of the original ECM.
Methodology: 12 human urethras were procured from deceased donors. Part of every procured urethra was used as a control sample for analysis. The protocol design was enzyme-detergent based, 0.25% trypsin, 1% Triton X-100 supplemented with enzymatic removal of DNA residues was used. Subsequently, the scaffold was washed in distilled water for 7 days. The efficiency of decellularization was determined using histology examination, immunohistochemical staining, electron microscopy and DNA quantification.
Results: Histology confirmed cell removal and preservation of urethral structure after decellularization. The preservation of collagen I, III, IV, elastin and fibronectin was confirmed by histologic examination and immunohistochemical staining. Scanning electron microscopy confirmed maintenance of ultrastructural architecture of ECM, ground substance and fibers. DNA content in decellularized urethra was significantly lower compared to the native sample and the criteria for decellularized tissue were met.
Conclusion: This study demonstrates the feasibility of an enzyme-detergent-based decellularization protocol for the removal of cellular components and maintenance of urethral ECM and its ultrastructure. Results provide solid ground for recellularization and urethral tissue engineering which will follow.
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