Immobilized lysozyme-based multifunctional nanofiber membrane incorporating α-ketoglutarate modulates the...

Osteomyelitis represents a major challenge in bone regeneration due to persistent bacterial infection, chronic inflammation, excessive osteoclastogenesis, and impaired osteogenesis. Here, an immobilized lysozyme-based multifunctional nanofiber membrane (PGHpAL) was developed as an in vitro proof-of concept platform for modulating the osteomyelitis-associated bone microenvironment. The membrane was fabricated by incorporating α-ketoglutarate (AKG) into a polycaprolactone/gelatin/nanohydroxyapatite electrospun matrix, followed by surface immobilization of lysozyme. The physicochemical characterization confirmed the successful fabrication and functional integration of the membrane. In addition, lysozyme immobilization significantly reduced AKG release compared with the non-coated membrane. Furthermore, PGHpAL exhibited excellent cytocompatibility and strong antibacterial activity against E. coli (94.12%) and S. aureus (85.44%). Moreover, relative to the LPS group, the nanofiber membrane decreased Tnfa and Il6 expressions to approximately 0.64- and 0.56-fold, respectively, while increasing Il10 and Cd206 expressions to 1.89- and 4.3-fold, respectively, consistent with a shift toward an anti-inflammatory and pro-healing macrophage response. This response was associated with attenuation of JNK and p38 MAPK activation and suppression of osteoclastogenesis. PGHpAL also promoted osteogenic differentiation and mineralization under physiological and inflammatory conditions, with marked enhancement of mineralized matrix formation under LPS-induced inflammatory stress. Collectively, these in vitro findings demonstrate the ability of PGHpAL to concurrently regulate antibacterial, inflammatory, osteoclastogenic, and osteogenic responses associated with the osteomyelitis bone microenvironment.