Experimental Study Of Bone Cement Loaded With Nano-silver On The Prophylaxis Of Infection After Joint Replacement

BackgroundInfection after joint replacement is a catastrophic complication. Many measures havebeen taken to prevent infection. Among them, antibiotics impregnated in bone cement is aneffective one, but at the same time it has given rise to an increasing number ofmicroorganisms resistant to antibiotics. Antibiotics in the bone cement can not play apreventive role for these microorganisms. There is a need to find a new antimicrobialagent.Silver has a strong antimicrobial potential, which has been used since the ancient times.Now it is used in different areas of medicine, such as cardiovascular implants, contactlenses, catheter and wound care. Nanotechnology changes silver into the nano-size andthus it has a better contact with the microorganisms, owing to high surface-area-to-volumeratio..Nano-silver has proved to have good antibacterial and antiviral properties.The gold standard for fixation of artificial joints is bone cement. If nanoparticles silveradded to the bone cement has antibacterial properties, it will provide a preliminary basisfor prevention arthroplasty infections and improvement of success rate for infectedarthroplasty.Part OneObjective: To detect wether the addition of a small dose of.nano-silver to bone cementwill have adversely effect on the mechanical properties of bone cement.Methods: Nano-silver powder and bone cement polymer powder were mixed, then thebone cement monomer was added and stirred under vacuum.0.1%,0.5%,1.0%nano-silverbone cement were measured as study groups, while polymethylmetacrylate(PMMA) ascontrol group. Compressive properties, bending strength and fatigue properties were testedfor every group. Results: There was no significant difference between0.1%,0.5%,1.0%nano-silver bonecement and ordinary bone cemen when mechanical properties such as compressiveproperties, bending strength and fatigue properties were compared (P>0.05).Conclusion: The addition of a dose less than1.0%nano-silver to bone cement had noadverse effects.on the mechanical properties of bone cement. Part TwoObjective: To assess whether nano-silver bone cement have antimicrobial activity andcytotoxicity.in vitro.Methods: Experiments were divided into five groups: ordinary bone cement was used asnegative control group and2%gentamicin bone cement was used as positive control group,0.1%,0.5%,1%nano-silver bone cement were studied as experimental group. Viable cellscount of in vitro bactericidal test was tested to measure the antibacterial activity of everygroup of bone cement to Staphylococcus epidermidis and methicillin-resistantStaphylococcus epidermidis. In vitro cytotoxicity testing was measured by release oflactate dehydrogenase and protein content of the incubated mouse fibroblasts. Experimentsfor in vitro cytotoxicity testing were divided into three groups: extraction medium after1%nano-silver bone incubated was studied as experimental group,while1%toxic TritonX-100as postive control group and non-toxic cell culture medium as a negative controlgroup.Results:1%nano-silver bone cement and gentamicin bone cement completely inhibittedthe proliferation of Staphylococcus epidermidis, showing strong bactericidal effect.1%nano silver bone cement completely inhibitted the proliferation of methicillin-resistantStaphylococcus epidermidis (MRSE), while2%gentamicin bone cement had no inhibitoryeffect on MRSE. There was no significant difference in LDH release of1%nano-silverbone cement when compared with that of non-toxic control group (P＞0.05). There wassignificant difference in LDH release of1%nano-silver bone cement when compared withthat of toxic Triton X-100(P＜0.01). There was no significant difference in the amount of total protein between extraction liquid from nano-silver bone cement and that of non-toxiccell culture medium (P＞0.05). There was significant difference in the amount of totalprotein between extraction liquid from nano-silver bone cement and that of toxic TritonX-100(P＜0.01). There was significant difference in the amount of total protein betweennon-toxic cell culture medium and that of toxic Triton X-100(P＜0.01).Conclusion:1%nano-silver bone cement had strong antibacterial activity againstStaphylococcus epidermidis and MRSE. Gentamicin bone cement had strong antibacterialactivity against Staphylococcus epidermidis, but had no effect on MRSE.1%nano-silverbone cement was free of in vitro cytotoxicity. Part ThreeObjective: To evaluate the effect of1%nano-silver bone cement on the formation ofbiofilm and preventing infection.Methods: Implant of a Kirschne about3cm long in concave of rabbit femoral condylesimulated arthroplasty, with1mm expsosed in articular space. The Kirschne was fixed withordinary bone cement. After skin closure,0.5ml different concentrations of Staphylococcusepidermidis were injected into knee. Erythrocyte sedimentation rate and C-reactive proteinwere measured preoperatively and postoperatively. Tissues in the knee were cultured forbacterial two weeks postoperatively. Then appropriate concentration of Staphylococcusepidermidis was choosed to establish infection model.36rabbits were randomly divided into three groups, each group consisted of12rabbits:ordinary bone cement was treated as a negative control group,2%gentamicin bone cementwas treated as a positive control group, and1%nano-silver bone cement was treated asexperimental group. According to group, different bone cement was used to fixe prosthesis.Then0.5ml suitable concentration of Staphylococcus epidermidis was injected into knee.Erythrocyte sedimentation rate and C-reactive protein were measured regularly aftersurgery. Biofilm on the surface of prosthesis was detected by scanning electron microscope(SEM) and tissue in the knee were cultured for bacterial two weekspostoperatively.Results:0.5ml1×10~6and1×10~8S. epidermidis could successfully lead to infection and1×10~6was choosed for next step experiment. In experiment of preventing infection,erythrocyte sedimentation rate and C-reactive protein was significantly lower innano-silver bone cement group and gentamicin bone cement group than that of ordinarybone cement group (P <0.01). There was no significant difference between nano-silverbone cement group and gentamicin bone group in erythrocyte sedimentation rate andC-reactive protein (P>0.05). Positive rate of bacterial culture for ordinary bone cementgroup, gentamicin bone cement group and nano-silver bone cement group was10~0%(12/12),0%(0/12) and0%(0/12) respectively; there was significant difference betweennano-silver bone cement group and ordinary bone cement group (P <0.01), there wassignificant difference between gentamicin bone cement group and ordinary bone cementgroup (P <0.01), no significant difference existed betwwen nano-silver bone cement groupand gentamicin bone cement group (P>0.05). SEM showed that bacteria on the surface ofthe prosthesis gathered and clusters，forming biofilm in ordinary bone cement group;andthat the bacteria isolated, no cluster formation in nano-silver group and gentamicin group.Conclusion:1%nano-silver bone cement inhibitted biofilm formation and preventedinfection in knee replacement of rabbit.