Introduction: The purpose of this study was to compare different decellularization protocols with the conventional detergent enzymatic method (DEM) using continuous agitation. Methods: The first experiment compared conventional DEM with sonication and lyophilization+freeze-thaw cycles. A second experiment was carried out to compare time-adjusted DEM (2-hour instead of 4-hour incubations with 4% deoxycholate) to decellularization in a bioreactor. Cellularity was determined by DNA-quantitation, H&E-staining and immunostaining for major histocompatibility complex-1 (MHC-1). Results: Compared to untreated trachea, DNA content significantly decreased after 2 cycles in all groups in the first experiment and dropped below the minimal criteria for efficient decellularization (<50 ng dsDNA/mg dry weight) after 4 cycles. However, nuclei were seen in the cartilage and MCH-1 staining was detected in some submucosal areas, indicating presence of chondrocytes and cellular residues that may render the scaffold immunogenic. In the second experiment DNA content significantly decreased after 1 cycle in both groups; however, even after 4 cycles, DNA content was above the minimal criteria for efficient decellularization. While collagen-levels remained stable, glycosaminoglycans diminished significantly after the initial cycles. Conclusions: Efficient decellularization can be achieved after only 4 cycles of DEM compared to the 17 cycles previously reported. The use of a bioreactor can preserve the integrity of the extracellular matrix.
ASJC Scopus subject areas
- Medicine (miscellaneous)
- Biomedical Engineering
Tracheal Decellularization using a Combination of Chemical, Physical and Bioreactor Methods. / Tchoukalova, Yourka D.; Hintze, Justin M.; Hayden, Richard E; Lott, David G.In: International Journal of Artificial Organs, 01.09.2017.
Research output: Contribution to journal › Article