Interaction and internalization of gram-positive orthopaedic pathogens in MG-63 osteoblast-like cells and antibacterial activity of vancomycin-eluting polymethylmethacrylate

PURPOSE: Orthopaedic implant-associated infections (OIAIs) are frequently caused by Gram-positive bacteria capable of invading osteoblasts, promoting persistence and treatment failure. Local antibiotic delivery using polymethylmethacrylate (PMMA) with vancomycin may improve extracellular bacterial eradication but may not directly address intracellular bacterial reservoirs. This study evaluated the interaction and internalization of Gram-positive bacteria in MG-63 osteoblast-like cells and the antibacterial activity of vancomycin-eluted PMMA. METHODS: Adhesion, interaction, and internalization assays were performed using six previously sequenced clinical isolates of coagulase-negative staphylococci (CoNS) (Staphylococcus epidermidis, S. haemolyticus, S. capitis, S. warneri, and S. caprae) and Cutibacterium acnes. Intracellular localization was confirmed by transmission electron microscopy. Vancomycin-impregnated PMMA beads were added to infected cell cultures to assess antibiotic elution and antibacterial activity. Statistical analyses included ANOVA or Kruskal-Wallis tests, with paired t-test and Wilcoxon sensitivity analysis for biomaterial assays. RESULTS: All isolates adhered to, interacted with, and internalized into MG-63 cells. Staphylococcus warneri exhibited the greatest cellular interaction, whereas S. caprae showed the highest internalization index despite lower interaction. C. acnes also demonstrated efficient intracellular localization, confirmed by transmission electron microscopy. Vancomycin-eluted PMMA significantly reduced the bacterial inoculum of all CoNS isolates and C. acnes. CONCLUSIONS: CoNS and C. acnes can invade osteoblast-like cells, supporting their role in the persistence of orthopaedic implant-associated infections. Vancomycin-eluted PMMA demonstrated significant antibacterial activity, supporting local antibiotic delivery as a complementary strategy for managing osteoarticular infections.