Background: Each year, about 136,000 individuals worldwide are diagnosed with larynx carcinoma. Tissue engineered organs shows promising alternative to a conventional transplantation in both preclinical models and in clinical set up to address the issues of donor scarcity and graft rejection. Here we attempted to create a less immune, whole pig larynx scaffold comprising complete acellular and integrated muscle and cartilage of the larynx.

Methods: Pig larynxes (n=5) were decellularized with combination of perfusion-agitation and ultrasonication method, where ultrasonic energy used for mechanically loosen the collagenous matrix of laryngeal tissue and detergents such as Dimethyl Sulfoxide and Triton X-100 were used to wash away the cell debris from extracellular matrix (ECM). Decellularized larynxes were characterized by histology, immunohistochemistry, DNA quantification and growth factor analysis (Luminex technology). Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) were used to investigate the ultrastructural changes to the arrangement of ECM fibers in acellular larynxes after decellularization.

Results: Complete acellular laryngeal matrix preparation took 25 decellularization cycles; with well preserving muscles, cartilage, and blood vessels. Histological findings showed that the non cartilaginous part of decellularised pig larynges were devoid of cells. Also we were successful in making complete chondrocytes free cartilage in epiglottis and trachea. However very few cell debris was observed in the thyroid and cricoid cartilage. The DNA quantification analysis showed 99% reduction in DNA content in the decellularised larynges compared to the normal larynx. Luminex analysis showed presence of angiogenic growth factors in the acellular larynxes after decellularization method. SEM and TEM analysis confirmed the structural arrangements of ECM fibers in larynxes were well preserved after decellularization.

Conclusion: Our findings suggest that, porcine larynxes can be decellularized with detergent-ultrasonication method. This method preserves ECM proteins and angiogenic growth factors which can help in building the new tissue engineered larynx for in-vivo implantation.

« Back to 2015 American Transplant Congress