Evaluation of changes in cartilage viability in detergent-treated tracheal grafts for immunosuppressant-free allotransplantation in dogs

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Abstract
OBJECTIVES: The first tissue-engineered clinical tracheal transplant prepared using the detergent-enzymatic method resulted in graft stenosis, possibly from detergent-enzymatic method-induced graft non-viability. We reported on the transplantation of de-epithelialized tracheal allografts while maintaining cartilage viability in dogs. No lethal stenosis occurred in allografts. Herein, on the basis of previous experimentation, we assessed cartilage viability in detergent-treated cartilages. METHODS: Six canine tracheal grafts were treated with detergent [1% t-octylphenoxypolyethoxyethanol (Triton X-100)] before transplantation. The histoarchitecture was evaluated, and the viable chondrocytes ratio was calculated. Glycosaminoglycan was detected using safranin-O staining. Collagen II was tested using immunohistochemistry. RESULTS: The epithelium was completely removed in 5 grafts. Compared with fresh tracheas, the viable chondrocyte ratio was significantly reduced in the de-epithelialized grafts (100 vs 54.70 ± 8.56%; P < 0.001). Image analysis revealed that the mean optical density of glycosaminoglycan (0.363 ± 0.027 vs 0.307 ± 0.012; P = 0.007) and collagen II (0.115 ± 0.013 vs 0.092 ± 0.011; P = 0.028) was decreased. The observation period ranged from 91 to 792 days. No stenosis occurred in 5 allografts; moderate stenosis developed in 1 allograft during the 4th week after surgery. The chondrocyte nuclei almost completely disappeared. Both glycosaminoglycan (0.307 ± 0.012 vs 0.164 ± 0.104; P = 0.044) and collagen II (0.092 ± 0.011 vs 0.068 ± 0.022; P = 0.022) were significantly degraded. CONCLUSIONS: This study demonstrated successful tracheal transplantation; about 50% of the viable chondrocytes were retained in the cartilage that could prevent development of a lethal stenosis in tracheal grafts.

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