Application And Mechanism Study Of Multifunctional Titanium Implants Augmented By Two-dimensional Niobium Carbide Nanosheets For The Treatment Of Implant Infections

Purpose: Implant infection is a catastrophic complication after orthopaedic implant surgery.Bacterial biofilm and tissue repair disorder caused by excessive inflammatory response at the infection site are two major reasons for the refractory characteristics of orthopaedic implant infection.Therefore,it is of great significance to design orthopedic implants that can simultaneously remove biofilm and inhibit excessive inflammatory response.Methods: Nb2C was first synthesized by etching and exfoliation of Nb2 AlC bulks,then deposited on the surface of aminated titanium by electrostatic adsorption,thus obtaining two-dimensional Nb2C nanosheets modified titanium implants(Nb2C@TP).Then we characterized Nb2C@TP by SEM,Raman,XPS and other detection methods.The antibacterial effect of Nb2C@TP was determined by dilution plate counting,SEM,CLSM and other detection studies,and the antibacterial mechanism of Nb2C@TP was explored by RNA-seq and qPCR.The in vitro photothermal antibacterial effect of Nb2C@TP was evaluated by TEM and flow cytometry.A mouse subcutaneous implant infection model was constructed,and the in vivo antibacterial effect of Nb2C@TP was verified by plate spreading,SEM,fluorescent bacterial construction,Giemsa staining and other methods.The in vivo anti-inflammatory effects of Nb2C@TPbased photothermal therapy were evaluated by F4/80 immunohistochemical staining,immunofluorescence staining of IL-6 and TNF-α,and quantitative analysis of MDA and SOD content in tissues.DCFHDA probe was used to detect the regulation of Nb2C@TP on ROS generation in macrophages stimulated by LPS.The ability of Nb2C@TP to promote the repair of infected tissue was assessed by Masson staining,immunofluorescence staining for CD31 and VEGF.Results: Two-dimensional nanosheets modified titanium implant(Nb2C@TP)was successfully constructed,with excellent biocompatibility and photothermal conversion efficiency in vitro and in vivo.In vitro experiments verified that Nb2C@TP has excellent anti-biofilm effect and moderate planktonic bacteria killing ability.RNA-seq studies showed that Nb2C@TP can interfere with bacterial energy metabolism pathways for direct bacterial killing,and interfere with accessory gene regulator(Agr)in the QS signaling pathway to inhibit biofilm formation and promote the dissociation of as-formed biofilm.In vitro photothermal experiments demonstrated that Nb2C@TP could increase the sensitivity of bacteria to thermal killing,and Nb2C@TP based photothermal therapy could further amplify the antibacterial efficacy of Nb2C@TP implant.In vivo experiments proved that Nb2C@TP equips with excellent antibacterial and anti-inflammatory effect.Besides,Nb2C@TP can promote tissue repair process in vivo,verifying its multimodal functions in treating implant infections.Conclusions: Herein,a Nb2C@TP implant with practical multimodal antiinfection functions was developed.Such multiple modes are capable of destroying biofilms for direct bacteria elimination through down-regulating bacterial energy metabolism pathway,suppressing biofilm formation and enhancing as-formed biofilm detachment via activating Agr.Another intriguing feature of this nanomedicine is the sensitization ability toward bacteria via photothermal transduction,which reduces the temperature necessary for the bacteria eradication and mitigate the possible normal tissue damage.Moreover,the Nb2C@TP medical implant is able to alleviate proinflammatory responses by scavenging excessive ROS in infectious microenvironment(IME),benefiting angiogenesis and tissue remodeling.