Gingival fibroblasts (GFs) can differentiate into osteoblast-like cells and induce osteoclast precursors to differentiate into osteoclasts. As it is unclear whether these two processes influence each other, we investigated how osteogenic differentiation of GFs affects their osteoclast-inducing capacity. To establish step-wise mineralization, GFs were cultured in four groups for 3 weeks, without or with osteogenic medium for the final 1, 2, or all 3 weeks. The mineralization was assessed by ALP activity, calcium concentration, scanning electron microscopy (SEM), Alizarin Red staining, and quantitative PCR (qPCR). To induce osteoclast differentiation, these cultures were then co-cultured for a further 3 weeks with peripheral blood mononuclear cells (PBMCs) containing osteoclast precursors. Osteoclast formation was assessed at different timepoints with qPCR, enzyme-linked immunosorbent assay (ELISA), TRAcP activity, and staining. ALP activity and calcium concentration increased significantly over time. As confirmed with the Alizarin Red staining, SEM images showed that the mineralization process occurred over time. Osteoclast numbers decreased in the GF cultures that had undergone osteogenesis. TNF-α secretion, a costimulatory molecule for osteoclast differentiation, was highest in the control group. GFs can differentiate into osteoblast-like cells and their degree of differentiation reduces their osteoclast-inducing capacity, indicating that, with appropriate stimulation, GFs could be used in regenerative periodontal treatments.

Bioactive glass (BAG) S53P4 is a clinically approved bone substitute with antibacterial, osteoconductive and osteostimulatory properties. Its antibacterial effect is associated with ion release, local pH elevation and osmolality, but the precise biochemical and biophysical mode-of-action is unclear. This study investigates the antibacterial mechanism of BAG S53P4 eluates. BAG eluates, collected at 2, 4, 8, and 24 h, eradicated Staphylococcus aureus. Elemental analysis revealed an early increase in concentrations of Si and Na, a later rise in Ca, depletion of P over time and rapid loss of Mg. Membrane disturbances occurred within 5 min, evident by permeability for SYTOX, aligning with time-kill kinetics for S. aureus and Bacillus subtilis. In B. subtilis, 2h-BAG-eluate induced rapid delocalization of marker proteins for cell division and DNA repair, signaling membrane potential collapse and nucleoid condensation. Transcriptomics revealed early transcription remodeling reflecting ionic and energetic imbalance, including disruption of central metabolism, redox homeostasis, and translational stability. Scanning electron microscopy revealed severe cell surface damage and particulate deposits on S. aureus. Transmission electron microscopy showed cell envelop disruptions and cytoplasmic leakage. Energy dispersive X-ray analysis identified Si on bacterial cell surface at 4 h and intracellular accumulation in punctured, empty cells at 24 h. Overall, BAG ionic dissolution products kill bacteria through a stepwise mechanism involving membrane damage, protein delocalization and metabolic impairment, accompanied by Si deposition on bacterial surfaces and loss of Mg. This finally leads to cell wall degradation, cytoplasmic content leakage and further Si deposition on the cells and inside cell ghosts.

Streptococcus suis is a largely neglected but emerging bacterial zoonotic pathogen of global concern for animal welfare, antibiotic resistance development, and human health. No effective vaccines are now available. Here, we identified and characterized the function and structure of two cell wall polysaccharide variants in pathogenic S. suis strains using genetic deletion and (heterologous) complementation, lectin staining, glycan composition analysis, and specialized NMR spectroscopy. Both glycan variants were anionic polymers that differed in the presence of glucose in the side chain as a result of allelic variation in a glycosyltransferase gene. Deletion of this variable glycosyltransferase revealed an identical glycan “core” and affected S. suis morphology and lysozyme resistance. Immunization of pigs with this core domain elicited antibodies that recognized antigenically diverse pathogenic S. suis strains and induced complement deposition on encapsulated pathogenic S. suis strains. This study provides valuable insights for developing next-generation glycoconjugate vaccines, whereby a single-glycan target could protect against the emerging zoonotic pathogen S. suis .