Experimental Study On Synergistic Treatment Of Periodontitis By Photothermal/Photodynamic/Chemodynamic Therapy Of Bismuth Sulfide-Anchored Cu-TCPP Nanosheets

Background:Periodontitis is a common oral inflammatory disease,and humans have paid a heavy price for the treatment of this disease over the long history.Periodontal tissue is the supporting tissue of teeth,anatomically,it mainly includes alveolar bone,periodontal ligament,and gingival tissue.The main pathogenic factor of periodontitis is dental plaque,which is a soft,unmineralized bacterial colony wrapped in a matrix that adheres to each other or to the surface of the tooth or restoration and cannot be washed away by water.It can also be thought of as a bacterial biofilm that cannot be washed away by water.Under the continuous stimulation of these bacteria and their metabolites,the infection of the periodontal tissue leads to the destruction of periodontal fibers and the absorption of alveolar bone,which eventually leads to the loss of teeth.Therefore,the effective pathways of removing dental plaque are particularly essential for inhibiting the occurrence and development of periodontitis.At present,the control of dental plaque is mainly through physical means and medical treatment.Physical means include supragingival and subgingival scaling.For patients with severe symptoms,in addition to the above physical means,antibiotics are also needed.However,due to the abuse of antibiotics,bacterial resistance is becoming more serious and could not be ignored,these problems force us to adopt new therapeutic strategies which can effectively kill bacteria,inhibit the further development of periodontitis,and avoid the side effects of treatment.Objective:Bismuth sulfide（Bi₂S₃）nanoparticles were uniformly distributed on the surface of Cu-TCPP nanosheets to form Bi₂S₃/Cu-TCPP nanocomposites.The ability of this nano-heterogeneous structure to ablate biofilm and the treatment of periodontitis are studied through in vitro and in vivo experiments.Methods:1.Synthesis of Cu-TCPP nanosheets and Bi₂S₃/Cu-TCPP nanocomposites by hydrothermal method.2.Characterize the nanocomposites by transmission electron microscopy（TEM）,ultraviolet spectroscopy,X-ray diffraction（XRD）,X-ray photoelectron spectroscopy（XPS）,photoluminescence（PL）spectroscopy,Zeta potential,Electron Paramagnetic Resonance（EPR）and other means.3.To study the effects of photothermal therapy（PTT）,photodynamic therapy（PDT）,and Chemodynamic therapy（CDT）of different nanomaterials in vitro.4.Through electrochemical experiments,the mechanism of Z-scheme heterostructure enhancing PDT performance was explored.5.The biological safety and cytotoxicity of Bi₂S₃/Cu-TCPP were evaluated by in vitro experiments（CCK-8 and cell staining）,in vivo experiments（H&E staining of important tissues and organs）,and hemolytic experiments.6.The bactericidal ability of Bi₂S₃/Cu-TCPP on Porphyromonas gingivalis（P.gingivalis）and Fusobacterium nucleatum（F.nucleatum）was explored by single biofilm or mixed biofilm live/dead staining,colony forming unit（CFU）counting,SEM,and polymerase chain reaction（Polymerase Chain Reaction,PCR）.7.Evaluate the therapeutic effect of Bi₂S₃/Cu-TCPP on periodontitis by in vivo experiments in rats（～180g,male）.Results:1.Bi₂S₃ nanoparticles are uniformly distributed on Cu-TCPP nanosheets,XPS and related characterizations show that there is a strong interaction between them.2.It is confirmed by in vitro experiments that 2-Bi₂S₃/Cu-TCPP has better PDT/PTT/CDT performance under the irradiation of 635 nm laser,and these three functions are mutually promoted,which proves that the load of Bi₂S₃ is greatly improved the function of Cu-TCPP.3.The electrochemical results show that this composite material does have a Z-type heterostructure and can significantly promote the separation and transport of photogenerated carriers,and further amplify the effect of PDT.4.Through biocompatibility tests in vitro and in vivo,the truth that Bi₂S₃/Cu-TCPP has excellent biocompatibility and low cytotoxicity was confirmed.5.According to the results of the antibacterial experiment,the ratio of viable bacteria and the thickness of the biofilms in the Bi₂S₃/Cu-TCPP experimental group were significantly reduced in the live/dead staining,and the CFU was reduced by3-log of magnitude,proving that the synergistic effect of PDT/PTT/CDT has the good bactericidal ability.6.In vivo studies have shown that Bi₂S₃/Cu-TCPP can significantly reduce the degree of alveolar bone resorption and inhibit the further development of inflammation.Conclusions:In this study,Bi₂S₃/Cu-TCPP nanocomposites were designed with the synergistic effect of PDT/PTT/CDT under 635 nm laser irradiation to eliminate biofilms and alleviate inflammation in the model of periodontal disease.In contrast with Cu-TCPP nanosheets,Bi₂S₃/Cu-TCPP nanocomposites accelerated photoinduced electron–hole separation and enhanced generation of ROS under 635 nm laser irradiation.The results of electrochemical experiments proved the rationality of the Z-Scheme heterostructure and revealed the mechanism of enhanced photodynamic activity by charge transfer and energy evolution pathways.Meanwhile,the photothermal effect of Bi₂S₃ nanoparticles further amplified CDT by accelerating the release of Cu²⁺and reaction rate.This synergistic effect of PDT/PTT/CDT showed an excellent ability to eradicate single-species or multi-species biofilms.These nanocomposites had a superior bactericidal effect in the treatment of periodontitis,which means that they could play an important role in the treatment of other diseases,especially tumors.