Designing a novel vacuum aspiration system to decellularize large-size enthesis with preservation of physicochemical and biological properties

BACKGROUND: Functional and rapid enthesis regeneration remains a challenge after arthroscopic rotator cuff (RC) repair. Tissue-engineering a large-size biomimetic scaffold may be an adjuvant strategy to improve this clinical dilemma. Herein, we developed an optimized protocol to decellularize large-size enthesis as scaffolds for augmenting RC tear. METHODS: A novel vacuum aspiration system (VAS) was set up, which can provide a negative pressure to suck out cellular substances from tissue blocks without using chemical detergents. Large-size enthesis tissue specimens were harvested from canine infraspinatus tendon (IT) insertion, and then decellularized with an optimized protocol [freeze-thaw processing followed by nuclease digestion and phosphate buffer saline (PBS) rinsing in the custom-designed VAS], or a conventional protocol (freeze-thaw processing followed by nuclease digestion and PBS rinsing), thus fabricating two kinds of acellular enthesis matrix (AEM), namely C-AEM and O-AEM. After that, the C-AEM and O-AEM were comparatively evaluated from the aspect of their physicochemical and biological properties. RESULTS: Physiochemically, the O-AEM preserved the morphologies, ingredients, and tensile properties much better than the C-AEM. Biologically, in vitro studies demonstrated that both C-AEM and O-AEM show no cytotoxicity and low immunogenicity, which could promote stem cells attachment and proliferation. Interestingly, O-AEM showed better region-specific inducibility on the interacted stem cell down osteogenic, chondrogenic and tenogenic lineages compared with C-AEM. Additionally, using a canine IT repair model, the injured enthesis patched with O-AEM showed a significant improvement compared with the injured enthesis patched with C-AEM or direct suture histologically. CONCLUSIONS: The proposed VAS may help us fabricate large-size AEM with good physicochemical and biological properties, and this AEM may have potential clinical applications in patching large/massive RC tear.