Fabrication and Characterization of Decellularized Periodontal Ligament Cell Sheet Constructs

Decellularized tissue engineered constructs have the potential to promote regeneration by providing a biomimetic extracellular matrix that directs tissue specific regeneration when implanted in situ. Recently, the use of cell sheets has shown promising results in promoting periodontal regeneration. Here, we describe the fabrication of decellularized periodontal cell sheets with intact extracellular matrix structural and biological properties. Melt electro-spun polycaprolactone (PCL) scaffolds are used as a carrier for the inherently fragile cell sheets, in order to provide support during the processes of decellularization. An optimized decellularization method is outlined using perfusion with a combination of NH₄OH and Triton X-100 together with a DNase treatment step for DNA removal. The maintenance of extracellular matrix structural and biological integrity is important, and here, we describe the assessment of these properties using immunostaining for extracellular matrix proteins and ELISA for growth factor quantification.