In Vitro Studies Of The Antibacterial And Anti-inflammatory Effects Of Autogenous Oxygen Composite Hydrogels

Objectives:Periodontitis is an inflammatory disease of the periodontal tissues induced by plaque parasitic on the neck of the teeth,which is a common disease of the oral cavity.Periodontitis is currently the leading cause of premature tooth loss in adults and is a serious threat to human oral health.Currently,supragingival scaling,subgingival scraping and the application of antibiotics in periodontal pockets are the most common methods used in clinical practice to inhibit plaque and control periodontitis.However,basic periodontal treatment can only temporarily remove plaque,and with the passage of time,plaque will soon proliferate and multiply in the periodontal pockets again,and the long-term application of clinical antibacterial drugs increases the risk of drug-resistant strains and is usually accompanied by side effects such as gastrointestinal reactions.The plaque in periodontal pockets is mainly gram-negative anaerobic bacteria,which are sensitive to oxygen content,and increasing oxygen content can significantly inhibit the growth of anaerobic bacteria.In addition,increasing oxygen content can also promote bone tissue regeneration,which is beneficial to periodontal tissue regeneration.Moreover,this method can be applied for a long time without drug resistance and gastrointestinal reactions.To investigate the feasibility of oxygen therapy applied to the treatment of periodontitis in more depth,this study evaluated the relevant biological properties of oxygen-producing composite hydrogel in anti-inflammation by establishing an in vitro inflammation model,also tested the inhibitory effect of this self-oxygen-producing composite hydrogel on Fusobacterium nucleatum（F.nucleatum）to provide new ideas and methods for the treatment of periodontitis.Methods:In this study,the proportion screening and configuration of sodium alginate hydrogels have been completed,and sodium alginate hydrogels loaded with Ca O₂and ascorbic acid have been successfully generated.In this study,the cytotoxicity of the composite hydrogel was evaluated by detecting the cell proliferation activity by CCK-8 method;the inflammation model was established by inducing Raw264.7 cells with bacterial lipopolysaccharides（LPS）,and the nitric oxide content in the cell supernatant was detected by nitric oxide kit.The proliferation activity was measured using CCK-8 assay kit.The expression levels of TNF-α,l L-1βand l L-6 m RNA in Raw264.7 cells were detected by quantitative real-time fluorescence polymerization chain reaction（q-PCR）;the levels of inflammatory factors TNF-α,l L-1βand l L-6 in the supernatant of Raw264.7 cells were detected by enzyme-linked immunosorbent assay（ELISA）.In vitro,F.nucleatum was co-cultured with the composite hydrogel for 24 h.After taking the bacterial solution and coating the plate,colony counting was performed,while live-dead bacterial staining was done,and the number of live-dead bacteria was observed under laser confocal microscope to determine the inhibitory effect of the composite hydrogel on F.nucleatum.Results:1.The composite hydrogel is biocompatible.2.Successfully established an inflammatory model of Raw264.7 cells using LPS;and determined the optimal induction concentration of LPS to be 1μg/m L.3.Composite hydrogel down-regulates TNF-α,l L-1β,l L-6 m RNA expression in LPS-stimulated Raw264.7 cells at the transcriptional level.4.The composite hydrogel significantly reduces the release of inflammatory factors TNF-α,l L-1β,l L-6 in Raw264.7 cells stimulated by LPS.5.The composite hydrogel can significantly inhibit the growth of F.nucleatum.Conclusion:1.The composite hydrogel has good biocompatibility.2.The composite hydrogel can significantly inhibit the growth of periodontitis-causing bacteria F.nucleatum.3.The composite hydrogel can reduce the expression and release of cellular inflammatory factors in the inflammatory state,thus acting as a therapeutic agent for cells in the inflammatory state.