Ventilation-Based Decellularization System of the Lung.

Abstract

The demand for donated organs greatly exceeds the availability. Alternatives to organ donation, such as laboratory-engineered organs, are therefore being developed. One approach is to decellularize the organ and reseed it with selected cells, ideally from the organ recipient. Organ decellularization has typically been attempted by the administration of detergents into vessels such as the portal vein in the liver. However, in the case of the lung, the airway provides another potential administration route, because it has a wide contact area between cells and detergents in the tracheal tree and alveoli. In the present study, we introduce a novel ventilation-based decellularization system for the lung and compare its efficacy to ordinary decellularization systems administering detergent through the pulmonary artery. Rat lungs were decellularized using 500 mL of 3-[(3-cholamidopropyl) dimethylammonio]-1-Propanesulfonate (CHAPS) decellularization solution administrated through the pulmonary artery (vessel group) or through the trachea (airway group). The vessel group was infused CHAPS solution using a gravitational pressure head of 20 cmH2O. The airway group was infused with the detergent using negative pressure and positive end-expiratory pressure, for a volume 10cc with each inspiration in a bioreactor. Pathological and immunohistochemical findings indicated that components of the extracellular matrix (ECM), including proteoglycans, elastic fibers, fibronectin, and laminin, were more decreased in the airway group than in the vessel group. Western blot analysis showed that MHC class I antigen and β-actin were not detected in both decellularized groups. A collagen assay showed that collagen was 70% preserved in both groups compared to native lung. Glycosaminoglycan (GAG) and DNA assays showed that GAG and DNA contents were strongly diminished in both decellularized groups, but those contents were smaller in the airway group than in the vessel group. Accordingly, the alveolar wall was thinner on electron microscopy, and DNA remnants were not observed in the airway group. Infusion of red blood cells indicated that capillary walls were preserved without blood leakage in both groups. In conclusion, we describe a novel approach for decellularization through the airway that represents a more stringent method for both DNA and ECM removal, with capillary wall preservation.