Development and Manufacturing of Scaffold-less Constructs for Tendon/Ligament Repair.

Soft tissues, such as rotator cuff tendons and the anterior cruciate ligament (ACL), integrate with the subchondral bone through a complex multi-tissue interface that functions to minimize the formation of stress concentrations and enable the efficient transfer of load between tendon or ligament and bone. Current rotator cuff tendon and ACL repair techniques, requiring the reattachment of the tendon/ligament to its original bony footprint, fail to regenerate this interface. Instead, the repaired insertion site transitions from tendon/ligament to bone through a disorganized, fibrovascular scar tissue with weak mechanical properties, leaving it prone to failure and compromising long-term clinical outcomes. To improve tendon-bone integration following rotator cuff repairs, the objective of this thesis was to utilize a scaffold-less tissue engineered construct to promote the regeneration of the tendon-bone interface and develop a reproducible, automated manufacturing system to facilitate the advancement of the construct towards clinical use. Matrix organization and mechanical properties of the regenerated enthesis were evaluated in both acute (immediate repair) and chronic (repair 4 weeks post injury) supraspinatus tear rat models. Utilization of tissue-engineered constructs resulted in superior enthesis regeneration compared to current mechanical fixation techniques. Next, to enhance the reproducibility and uniformity of existing multi-phasic scaffold-less construct fabrication methodologies, protocol standards and a novel delamination system were developed and later extrapolated for use with human derived constructs. The novel construct fabrication methods yielded an increased number of engineered constructs of consistent size and mechanical properties. Temporal gene expression confirmed the commitment of human derived constructs toward tendon and bone-like tissues. Lastly, to facilitate the eventual large-scale commercial production of our multiphasic tissues, a novel semi-closed bioreactor system was developed and validated. The use of the bioreactor successfully facilitated the co-culture and integration of two distinct tissue types in a single chamber without any direct user manipulation. The findings described in this thesis will lead to the development of a new soft-tissue-to-bone repair strategy to improve functional tendon/ligament repair outcomes and provide the framework for expediting the clinical and commercial translation of our tissue engineering technologies.