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- Title
Development and In Vitro/In Vivo Comparative Characterization of Cryopreserved and Decellularized Tracheal Grafts.
- Authors
Stocco, Elena; Barbon, Silvia; Mammana, Marco; Trojan, Diletta; Bianchin, Alice; Favaretto, Francesca; Contran, Martina; Zambello, Giovanni; Vogliardi, Andrea; Confalonieri, Marta; Todros, Silvia; Pavan, Piero G.; Romanato, Filippo; Conconi, Maria Teresa; Macchi, Veronica; De Caro, Raffaele; Rea, Federico; Porzionato, Andrea
- Abstract
Tracheal reconstruction represents a challenge when primary anastomosis is not feasible. Within this scenario, the study aim was to develop a new pig-derived decellularized trachea (DecellT) to be compared with the cryopreserved counterpart (CryoT) for a close predictive analysis. Tracheal segments underwent decellularization by a physical + enzymatic + chemical method (12 cycles); in parallel, cryopreserved samples were also prepared. Once decellularized (histology/DNA quantification), the two groups were characterized for Alpha-Gal epitopes/structural proteins (immunohistochemistry/histology/biochemical assays/second harmonic generation microscopy)/ultrastructure (Scanning Electron Microscopy (SEM))/mechanical behaviour. Cytotoxicity absence was assessed in vitro (extract-test assay/direct seeding, HM1SV40 cell line) while biocompatibility was verified in BALB/c mice, followed by histological/immunohistochemical analyses and SEM (14 days). Decellularization effectively removed Alpha-Gal epitopes; cartilage histoarchitecture was retained in both groups, showing chondrocytes only in the CryoT. Cryopreservation maintained few respiratory epithelium sparse cilia, not detectable in DecellT. Focusing on ECM, preserved structural/ultrastructural organization and collagen content were observed in the cartilage of both; conversely, the GAGs were significantly reduced in DecellT, as confirmed by mechanical study results. No cytotoxicity was highlighted by CryoT/DecellT in vitro, as they were also corroborated by a biocompatibility assay. Despite some limitations (cells presence/GAGs reduction), CryoT/DecellT are both appealing options, which warrant further investigation in comparative in vivo studies.
- Subjects
SECOND harmonic generation; TRACHEA; IMMUNOHISTOCHEMISTRY; CYTOSKELETAL proteins; HISTOLOGY; FROZEN semen; TRACHEAL cartilage; FROZEN human embryos
- Publication
Cells (2073-4409), 2023, Vol 12, Issue 6, p888
- ISSN
2073-4409
- Publication type
Article
- DOI
10.3390/cells12060888