A novel supercritical CO2-based decellularization method for maintaining scaffold hydration and mechanical properties

Decellularized tissues are commonly utilized as tissue engineering scaffolds. Decellularization by extended exposure to aqueous detergents can damage the microstructure or deposit cytotoxic residue. Supercritical carbon dioxide (scCO(2)) has been proposed for decellularization, but reportedly causes dehydration and scaffold embrittlement. Presented herein is a novel decellularization method that preserves matrix hydration state and mechanical properties. Over 97% of the water in porcine aorta is maintained by presaturating scCO(2) with water; however, complete decellularization was not attained by any process utilizing only scCO(2). Instead, a novel hybrid method is presented that combines a brief (48 h) exposure of tissue to aqueous detergent, followed by washing with scCO(2) (1 h). The hybrid method fully decellularized the tissue, as confirmed by histology and DNA quantification (<0.04 lig DNA/mg tissue). This hybrid treatment was faster than the standard method (2 days compared to 4-7 days), while preserving tissue structure and mechanical properties.