ABSTRACT
BACKGROUND: Suboptimal healing of the tendon-bone interface remains an unsolved problem. The authors hypothesized that (1) platelet-rich plasma and prolonged in vitro incubation will produce interface scaffolds with greater reseeding of viable adipose-derived stem cells; and (2) when implanted with extracellular matrix hydrogel, constructs will display superior in vivo strength repair and biocompatibility. METHODS: Achilles-calcaneal composite tendon-bone interface scaffold grafts were harvested from 30 Wistar rats. After physicochemical decellularization and lyophilization, scaffolds were revitalized in rat plasma or 100% activated rat platelet-rich plasma and reseeded with viable adipose-derived stem cells. For part 2 of the study, 90 Sprague-Dawley rats underwent reconstruction with one of five decellularized, lyophilized scaffold revitalization/reseeding conditions: (1) phosphate-buffered saline; (2) lyophilized, 100% activated platelet-rich plasma; (3) platelet-rich plasma and extracellular matrix hydrogel; (4) platelet-rich plasma and 14-day reseeding with ASC-luc2-eGFP cells; and (5) plasma, reseeding, and hydrogel. RESULTS: In part 1, platelet-rich plasma-revitalized grafts demonstrated greater live viable adipose-derived stem cell loads at 3, 7, and 14 days and total adipose-derived stem cell loads at 7 and 14 days with visibly greater live surface cellularity, layering, migration, and penetration. In part 2, bioluminescence imaging confirmed cell viability to day 22 after implantation. Biomechanical strength testing demonstrated a significant increase in ultimate failure load for reseeded groups compared with all other groups at week 2, whereas only reseeded grafts with hydrogel remained significantly stronger at weeks 4 and 8. Histologic examination demonstrated most increased tendinous cellular invasion and fibrocartilage repopulation at 8 weeks in the reseeded group with hydrogel. Masson trichrome staining demonstrated persistence of the scaffold structure at week 8 and blinded ImageJ analysis demonstrated significantly more type III collagen in the reseeded/hydrogel group at 2, 4, and 8 weeks. CONCLUSIONS: Decellularized lyophilized allogeneic tendon-bone interface scaffolds can be optimized by revitalization in platelet-rich plasma, reseeding with viable adipose-derived stem cells, and supplemented by an extracellular matrix tendon hydrogel at the time of implantation. When this is done, they display greater repair strength and biocompatibility.
