PRELIMINARY STUDY ON IN VITRO TENDON ENGINEE-RING USING TENOCYTES AND POLYGLYCOLIC ACIDS

Objective To investigate the feasibility of tendon engineering in vitro using tenocyws and polyglycolic acids （ PGA ）. Methods Tenocytes were isolated by tissue explant method and expanded in vitro. Cells of the second passage were collected and seeded onto PGA scaffolds made from PGA unwoven fibers at the density of 20 × 10^6 cells/ml. At 1 week postseeding ,the constructs were divided into three groups as follows： cell-scaffold constructs under constant tension generated by a U-shaped spring as the experimental group （ n = 5 ）, cell-scaffold constructs under no tension as control group 1 （ n = 4 ）, cell-free scaffolds under constant tension as control group 2 （n =3）. Samples were harvested at 2, 4 and 6 weeks for histological and immunohistochemical （ IHC ） examinations. Transmission electron microscopy （TEM） and mechanical test were performed to evaluate the constructs of 6 weeks. Results At 2 weeks, the constructs were mainly composed of undegraded PGA fibers. Gross and histological examination revealed no difference between the groups. At 4 weeks, neo-tendon was visible through gross observation in experimental group and control group 1. Histology and immunohistochemistry revealed the formation of collagen fibers. While in control group 2, PGA fibers were mostly degraded. At 6 weeks, the constructs were much thinner in experimental group than those in control group 1 （ 1.44 ± 0.13mm vs 2.55 ± 0. 18mm in diameter ）. TEM showed periodical strata of collagen fibers in the constructs from experimental group and control group 1. However, histology in experimental group revealed longitudinal alignment of collagen fibers, which more resembled natural tendon than neotendon formed in control group 1. Besides, the maximum load to failure（ Newton/mm^2 ） was greater in experimental group than that in control group 1 （1. 107 ±0. 327 vs 0. 294 ± 0. 138, P 〈0.05）. Conclusion It＇ s possible to engineer tendon substitutes in vitro. Cyclic strain generated by a bioreactor may be the optimal mechanical stimulation and is currently under investigation.

Engineering an extracellular matrix-functionalized,load-bearing tendon substitute for effective repair of large-to-massive tendon defects

A significant clinical challenge in large-to-massive rotator cuff tendon injuries is the need for sustaining high mechanical demands despite limited tissue regeneration,which often results in clinical repair failure with high retear rates and long-term functional deficiencies.To address this,an innovative tendon substitute named“BioTenoForce”is engineered,which uses(i)tendon extracellular matrix(tECM)’s rich biocomplexity for tendon-specific regeneration and(ii)a mechanically robust,slow degradation polyurethane elastomer to mimic native tendon’s physical attributes for sustaining long-term shoulder movement.Comprehensive assessments revealed outstanding performance of BioTenoForce,characterized by robust core-shell interfacial bonding,human rotator cuff tendon-like mechanical properties,excellent suture retention,biocompatibility,and tendon differentiation of human adipose-derived stem cells.Importantly,BioTenoForce,when used as an interpositional tendon substitute,demonstrated successful integration with regenerative tissue,exhibiting remarkable efficacy in repairing large-to-massive tendon injuries in two animal models.Noteworthy outcomes include durable repair and sustained functionality with no observed breakage/rupture,accelerated recovery of rat gait performance,and>1 cm rabbit tendon regeneration with native tendon-like biomechanical attributes.The regenerated tissues showed tendon-like,wavy,aligned matrix structure,which starkly contrasts with the typical disorganized scar tissue observed after tendon injury,and was strongly correlated with tissue stiffness.Our simple yet versatile approach offers a dual-pronged,broadly applicable strategy that overcomes the limitations of poor regeneration and stringent biomechanical requirements,particularly essential for substantial defects in tendon and other load-bearing tissues.

Facile and rapid fabrication of a novel 3D-printable,visible light-crosslinkable and bioactive polythiourethane for large-to-massive rotator cuff tendon repair

Facile and rapid 3D fabrication of strong,bioactive materials can address challenges that impede repair of large-to-massive rotator cuff tears including personalized grafts,limited mechanical support,and inadequate tissue regeneration.Herein,we developed a facile and rapid methodology that generates visible light-crosslinkable polythiourethane(PHT)pre-polymer resin(~30 min at room temperature),yielding 3D-printable scaffolds with tendon-like mechanical attributes capable of delivering tenogenic bioactive factors.Ex vivo characterization confirmed successful fabrication,robust human supraspinatus tendon(SST)-like tensile properties(strength:23 MPa,modulus:459 MPa,at least 10,000 physiological loading cycles without failure),excellent suture retention(8.62-fold lower than acellular dermal matrix(ADM)-based clinical graft),slow degradation,and controlled release of fibroblast growth factor-2(FGF-2)and transforming growth factor-β3(TGF-β3).In vitro studies showed cytocompatibility and growth factor-mediated tenogenic-like differentiation of mesenchymal stem cells.In vivo studies demonstrated biocompatibility(3-week mouse subcutaneous implantation)and ability of growth factor-containing scaffolds to notably regenerate at least 1-cm of tendon with native-like biomechanical attributes as uninjured shoulder(8-week,large-to-massive 1-cm gap rabbit rotator cuff injury).This study demonstrates use of a 3D-printable,strong,and bioactive material to provide mechanical support and pro-regenerative cues for challenging injuries such as large-to-massive rotator cuff tears.