Revealing the Auxetic Behavior of Biomimetic Multimaterial and Region-Specific Nanofibrous Fascicle-Inspired Scaffolds via Synchrotron Multiscale Digital Volume Correlation: Innovative Building Blocks for the Enthesis Regeneration

Enthesis lesions are one of the prevalent causes of injuries in the tendon tissue. The gradient of mineralization, extracellular matrix organization and auxetic mechanical properties, make enthesis regeneration challenging. Innovative electrospun fascicle-inspired nanofibrous poly(L-lactic)acid/collagen type I blend scaffolds are developed. Specifically, a mineralized fibrocartilage-inspired region (with/without nano-mineralization with hydroxyapatite), where random and aligned nanofibers coexist, is connected to a tendon-like region made of aligned nanofibers, through a conical non-mineralized fibrocartilage-inspired junction. Scanning electron microscopy and synchrotron X-ray nano-tomography show the morphological biomimicry of scaffolds with the natural tendon fascicles. Cultures of human mesenchymal stromal cell spheroids confirm a balanced expression of both tendon, cartilage, and bone markers on the non-mineralized scaffolds compared with the mineralized ones. Mechanical tests, at different physiological strain-rates, reveal a biomimetic mechanical behavior of scaffolds and the ability of junctions to tune the mechanics of their surrounding sites. Multiscale synchrotron in situ tensile tests, coupled with digital volume correlation, elucidate the full-field strain distribution of scaffolds from the structural down to the nanofiber level, highlighting the auxetic mechanical behavior of junctions typical of the natural enthesis. The findings and cutting-edge investigations of this study suggest the suitability of these enthesis-inspired fascicles as innovative scaffolds for enhanced enthesis regeneration.