An Aligned-to-Random PLGA/Col1-PLGA/nHA Bilayer Electrospun Nanofiber Membrane Enhances Tendon-to-Bone Healing in a Murine Model

Background: The challenge of achieving effective tendon-to-bone healing remains a significant concern in sports medicine, necessitating further exploration. Biomimetic electrospun nanomaterials present promising avenues for improving this critical healing process. Purpose: To investigate the biological efficacy of a novel aligned-to-random PLGA/Col1-PLGA/nHA bilayer electrospun nanofiber membrane in facilitating tendon-to-bone healing. Study Design: Controlled laboratory study. Methods: The bilayer membrane’s composition, combining PLGA/Col1 for tendon attachment and PLGA/nHA for bone integration, was examined using scanning electron microscopy, Fourier transform infrared spectroscopy, and mechanical testing. Positioned between the Achilles tendon and bone, its design aimed for harmonious integration with both types of tissue. In vitro, biocompatibility, cell adhesion, and proliferation of the biomaterial were evaluated using live/dead staining and the CCK-8 assay. Collagen secretion and mineralization were measured for 2 cell types. In vivo, tendon-to-bone insertion samples harvested from mice were analyzed: micro–computed tomography assessed bone formation; histological staining evaluated chondrogenesis, tendinogenesis, and the 4-layer structure of the insertion; and biomechanical testing measured insertion strength. Real-time polymerase chain reaction identified genes involved in tendon-to-bone healing, and transcriptome analysis elucidated the underlying cellular and molecular mechanisms. Results: The optimal composition was determined as 10% 3:1 for aligned PLGA/Col1 and 9% 5:1 for PLGA/nHA. Coculture showed minimal cell death, firm cell adherence, and steady proliferation, with PLGA/Col1 enhancing collagen secretion. In vivo, the material promoted bone and cartilage formation and improved tendon-to-bone interface strength. Transcriptome analysis indicated links to TNF and NF-κB pathways and to genes IL-1β, ADAM8, and EGR2. Conclusion: The novel aligned-to-random PLGA/Col1-PLGA/nHA bilayer nanofiber membrane outperformed other materials in both in vitro and in vivo evaluations, significantly enhancing tendon-to-bone healing. It notably improved cartilage and bone formation, tendon maturation, and biomechanical strength at the surgical interface. These effects may be associated with the TNF and NF-κB pathways and with the genes IL-1β, ADAM8, and EGR2. Clinical Relevance: This study introduces a biomimetic nanofiber membrane enhancing tendon-to-bone healing, which is crucial for sports medicine. Its efficacy in improving healing outcomes, including bone and cartilage formation and biomechanical strength, could significantly lower failure rates in surgical procedures such as rotator cuff repair and anterior cruciate ligament reconstruction. This advancement offers promising implications for patient recovery and the effectiveness of surgical interventions in tendon-to-bone injuries.