| BackgroundInfectious bone defect refers to the bone defect accompanied by infection or produced during the treatment of bone infection,which is complicated and complicated:To control bone infection and repair bone defects,new antibacterial bone tissue engineering materials have become a hot topic in clinical and basic research for the treatment of infected bone defects due to the deficiencies of autogenous bone allograft and drug resistance caused by antibiotic use.ObjectiveThe main purpose of this study was to construct CSH/β-TCP/BBR granular materials with antibacterial properties and bone-inducing activity,and systematically evaluate their biosafety,so as to provide a new method for the treatment of infectious bone defects,and provide scientific basis for preclinical application..MethodsIn the first part,the solution of berberine was prepared and the antibacterial effect of berberine on Staphylococcus aureus was measured by the diameter of bacteriostatic zone by AGAR drilling method.The minimum inhibitory concentration(MIC)and minimum antibacterial concentration(MBC)of berberine solution against Staphylococcus aureus were determined by microbroth dilution method.In the second part,CSH/β-TCP/BBR granules were co-cultured with rat bone marrow mesenchymal stem cells(BMSCs)CCK-8 method was used to test the effect of C SH/β-TCP/BBR granules on the vitality of rat BMSCs cells and the ALP activity of rat BMSCs cells.The third part established the infective bone defect model of SD rats(24 rats,divided into 4 groups,6 rats in each group)were implanted with scaffolds with different berberine concentrations(0,500μg/mL,750μg/mL,1000μg/mL)and the repair of infected bone defects was comprehensively evaluated by postoperative body temperature,body weight,intraoperative healing,blood routine and bacterial culture Micro-CT detection at 3 and 6 weeks after surgery:Results1.Berberine has good antibacterial activity against Staphylococcus aureus.The antibacterial zone range of berberine solution with concentration of 20mg/mL 100μl is 35.40±1.67mm,and the MBC and MIC values of berberine solution against Staphylococcus aureus are 62.5-125μg/mL.2.Gradient concentration of CSH/β-TCP/BBR granules has an effect on the growth and proliferation of rat mesenchymal stem cells.CSH/β-TCP/BBR granules with liquid berberine concentration higher than 1000μg/mL have obvious cytotoxicity.CSH/β-TCP/BBR granules with berberine concentration between 500 and 700μg/mL promoted cell proliferation and the expression level of osteogenic marker ALP was higher than that of the control group.3.At 3 and 6 weeks after operation,there was no significant difference in blood routine results among the four groups.Micro-CT and pathological results showed that the bone defect area of the experimental group was smaller than that of the blank control group,and the internal implants existed.The range of bone defects was reduced and new bone trabeculae were formed in the medium and low concentration groups,and the medium concentration group was more obvious.Six weeks after the operation,the bone marrow lavage fluid was cultured The antibacterial rate of the three experimental groups reached 100%,which was significantly different from that of the blank control group.ConclusionIn this study,calcium sulfate/calcium phosphate granules(CSH/β-TCP/BBR granules)containing Chinese medicine berberine were prepared by directly combining calcium sulfate and calcium phosphate solid phase powder with the liquid phase of Chinese medicine monomer berberine solution.The antibacterial activity,biosafety and bone inducability of the granules were detected and evaluated through in vitro experiments,combined with the experimental results of rats.It was found that the low and middle concentration group(liquid berberine concentration 750pg/mL)had good antibacterial activity and bone-inducing activity,while no obvious cytotoxicity was found in the concentration group,which provided a theoretical basis for the clinical application of biodegradable biomaterials containing TCM monomers. |
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