Microfluidic photo-crosslinking fabrication of silk fibroin/hyaluronic acid composite microsphere-reinforced hydrogels.

Abstract

Objectives: To fabricate an injectable composite microsphere hydrogel reinforced with silk fibroin/hyaluronic acid microspheres, achieving synergistic enhance-ment of mechanical robustness and biofunctionality.

Methods: Methacrylated hyaluronic acid (HAMA) and thiolated silk fibroin (SF-GSH) precursors were synthesized. Monodis-perse microspheres (43~130 μm) generated via microfluidics were UV-crosslinked (420 nm) through thiol-ene click reaction. These microspheres were embedded in a HAMA/SF-GSH matrix to form photo-crosslinked composites. The physicochemical and biological properties of composite microsphere hydrogels were systematically characterized by ¹H-NMR, optical microscopy, rheometer, scanning electron microscopy, inverted confocal microscopy, rheology, X-ray energy dispersive spectrometer, compression testing, degrada-tion/swelling assays, calcein-AM/PI double staining, CCK-8 assay, etc.

Results: The grafting rates of methacrylated hyaluronic acid and thiolated silk fibroin was 48.03% and 17.99%, respectively. The diameter of microsphere hydrogel was evenly distributed in the range of 43~130 μm. The gelation time of the composite microsphere hydrogel system was 48~115s. The laser confocal imaging confirmed dynamic regulation charac-teristics of the composite microsphere hydrogel system. The compressive strength of the composite microsphere hydrogel reached 22.7 kPa, and maintained structural integrity at 40% strain after 20 compression cycles. The composite microsphere hydrogels exhibited differential deswelling behaviors in simulated physiological environments, and the reducing microsphere particle size could significantly enhance its stability under moist conditions. The degradation rate of the composite microsphere hydrogel was 49% after 200 h of degradation, and the water retention rate maintained at 49%~62% after 96 h of degradation. Biocompatibility assays confirmed >95% cell viability and unimpaired cell migration abilities.

Conclusions: The silk fibroin/hyaluronic acid composite microsphere hydrogel developed with microfluidic photo-crosslinking strategy in this study has charac-teristics of rapid fabrication, adjustable mechanical properties, enhanced environmental stability and excellent biocompatibility, its unique injectability and water retention provide a new material solution for tissue repair and regenerative medicine.