Supercritical Carbon Dioxide Assisted Decellularization for Regenerative Medicine

In living tissues, the extracellular matrix (ECM) provides biochemical and structural cues to regulate cell phenotype and function.  A prominent strategy of regenerative medicine is to decellularize the ECM (rid the ECM of its native cells and DNA while maintaining its biochemical and structural properties) and repopulate it with stem cells to engineer functional tissues and organs.  Traditional methods of decellularization employ surfactants such as sodium dodecyl sulfate.  While these chemicals effectively decellularize the ECM, they are damaging to its structural proteins.  Supercritical carbon dioxide (scCO₂), a non-toxic, non-deforming solvent, is commonly used in extraction applications, making it a potential aid for decellularization.  We explored decellularization of confluent human dermal fibroblast cells using scCO₂ by investigating the effects of the scCO₂ pressure, scCO₂ saturation time, isopropyl alcohol addition, releasing patterns, and sonication.  It was found that overnight storage at 4°C, a longer saturation time, and sonication improved the decellularization efficiency.  The time- and cost-effective nature of this procedure has the potential to allow for mass-production of engineered tissues to be used in regenerative medicine.