双效阳离子纳米递送体系的免疫治疗策略用于牙周炎辅助治疗的研究OA

Objective To investigate the effects of a dual-functional cationic nucleic acid nano delivery system(DCNA)loaded with small interfering RNA targeting Bruton's tyrosine kinase(siBTK)on the immune-inflammatory microenvironment of periodontitis by regulating BTK expression.Methods DCNAsiBTK was prepared using a double-emulsion solvent evaporation method,and its physicochemical properties and sustained-release per-formance were characterized.Cytotoxicity was evaluated using the cell counting kit-8(CCK-8)assay.Antibacte-rial activity against periodontal pathogens was assessed by colony-forming assays.BTK mRNA and protein expres-sion levels were determined by reverse transcription quantitative real-time polymerase chain reaction(RT-qPCR)and Western blot,respectively.Enzyme-linked immunosorbent assay(ELISA)was performed to quantify the lev-els of inflammatory cytokines,including IL-1β,IL-6,and TNF-α.A murine periodontitis model was established and treated with DCNAsiBTK.Therapeutic efficacy and biosafety were evaluated using quantitative polymerase chain reaction(qPCR),micro-computed tomography(micro-CT),hematoxylin-eosin(HE)staining,and immunohisto-chemistry(IHC).Results Stable DCNAsiBTK nanoparticles with an average particle size of(168.8±2.2)nm and a zeta potential of(17.9±0.8)mV were successfully prepared.Transmission electron microscopy(TEM)revealed uniformly spherical morphology with favorable sustained-release properties.CCK-8 assays demonstrated no signifi-cant cytotoxicity of DCNAsiBTK within the concentration range of 0-100 nmol/L.In vitro studies showed that DCNAsi-BTK effectively inhibited the growth of Porphyromonas gingivalis(Pg),Aggregatibacter actinomycetemcomitans(Aa),and Fusobacterium nucleatum(Fn),indicating broad-spectrum antibacterial activity.RT-qPCR and West-ern blot analyses demonstrated that DCNAsiBTK downregulated BTK mRNA and protein expression in a dose-dependent manner.ELISA results showed that DCNAsiBTK significantly reduced the secretion of IL-1β,IL-6,and TNF-α,thereby attenuating inflammatory responses.In vivo experiments further demonstrated that DCNAsiBTK sup-pressed anaerobic bacterial proliferation within periodontal pockets,alleviated alveolar bone resorption and peri-odontal inflammation,and caused no obvious pathological damage in major organs,including heart,liver,spleen,lungs,and kidneys.Conclusion DCNAsiBTK efficiently encapsulated and released siBTK,and exhibited dual ac-tivities including antibacterial and anti-inflammatory.By suppressing BTK expression,DCNAsiBTK effectively at-tenuated the progression of periodontitis,providing a promising novel strategy for adjunctive periodontal therapy.