SLA表面处理TiCu合金的抗菌性能研究
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摘要
目的对TiCu合金进行SLA表面粗糙化处理,探究具有粗糙表面的TiCu合金的抗菌性能。方法 用喷砂+酸蚀(SLA)的方法对TiCu合金表面进行粗糙化处理(SLA-TiCu),并与经过相同处理的SLA-Ti进行对比研究。采用金黄色葡萄球菌作为实验菌株,通过平板菌落计数法计算杀菌率,采用活/死细菌染色实验观察表面细菌的粘附分布及生物被膜的形成情况,在透射电镜下观察细菌微观结构的变化,探究SLA-TiCu的抗菌作用及对细菌微观结构的影响。结果 SLA表面处理使TiCu合金具有微米-亚微米的复合双层孔洞结构表面,SLA-TiCu具有优异的抗菌性能,对表面粘附细菌的抗菌率为100%。染色结果表明,SLA-TiCu表面粘附的细菌中大量的是死细菌,没有形成生物被膜,而SLA-Ti表面已经形成细菌生物被膜。透射电镜观察表明,SLA-TiCu作用后的细菌出现了明显的细胞膜溶解现象,破损的细菌体内部出现透明和半透明的区域,细胞壁和细胞膜破损,细胞内的细胞质流出,在破损的细菌周围可以看到高电子密度的颗粒和沉淀。结论 SLA处理的TiCu合金具有优异的抗菌性能,更能有效发挥合金中Cu的杀菌作用,能够高效杀死表面粘附的细菌,从而抑制表面生物被膜的形成。因此,具有优异抗菌性能的SLA-TiCu具有极大潜力应用于牙种植体制造。
The work aims to explore the antibacterial performance of TiCu alloy with rough surface after SAL. The TiCu alloy was treated by sandblasting and acid etching(SLA) in order to acquire a rough surface and then compared with SLA-Ti for study. S. aureus was used as the experimental strains to calculate the sterilizing rate through the plate colony-counting method.Dead/live bacteria staining test was carried out to observe the bacteria adhesion distribution and formation of biofilm on the surface and the change of bacteria microstructure under TEM, in order to explore the antibacterial properties of SLA-TiCu and the effects on bacterial microstructure. The surface of SLA-TiCu had the micro-submicron dual-hole scale structure and excellent antibacterial properties, with an antibacterial ratio of 100% for surface adhesion bacteria. The staining result reflected that a large number of bacteria on the surface of SLA-TiCu were dead, and no biofilm was formed, but biofilm was observed on the surface of SLA-Ti. Observed under TEM, the bacteria on SLA-TiCu were found to have obvious cell membrane dissolution phenomenon and transparent and semitransparent areas appeared inside the damaged bacteria body. Cell walls and cell membranes were damaged, cytoplasm in cells flew out, and particles and precipitates with high electron density could be seen around the damaged bacteria. The TiCu alloy treated by SLA has excellent antibacterial properties, which can effectively play the antibacterial role of Cu ions and then effectively kill the bacteria adhering to the surface, thus inhibiting the formation of biofilms on the surface. Therefore, SLA-TiCu with outstanding antibacterial performance has great potential for application in dental implants.
The work aims to explore the antibacterial performance of TiCu alloy with rough surface after SAL. The TiCu alloy was treated by sandblasting and acid etching(SLA) in order to acquire a rough surface and then compared with SLA-Ti for study. S. aureus was used as the experimental strains to calculate the sterilizing rate through the plate colony-counting method.Dead/live bacteria staining test was carried out to observe the bacteria adhesion distribution and formation of biofilm on the surface and the change of bacteria microstructure under TEM, in order to explore the antibacterial properties of SLA-TiCu and the effects on bacterial microstructure. The surface of SLA-TiCu had the micro-submicron dual-hole scale structure and excellent antibacterial properties, with an antibacterial ratio of 100% for surface adhesion bacteria. The staining result reflected that a large number of bacteria on the surface of SLA-TiCu were dead, and no biofilm was formed, but biofilm was observed on the surface of SLA-Ti. Observed under TEM, the bacteria on SLA-TiCu were found to have obvious cell membrane dissolution phenomenon and transparent and semitransparent areas appeared inside the damaged bacteria body. Cell walls and cell membranes were damaged, cytoplasm in cells flew out, and particles and precipitates with high electron density could be seen around the damaged bacteria. The TiCu alloy treated by SLA has excellent antibacterial properties, which can effectively play the antibacterial role of Cu ions and then effectively kill the bacteria adhering to the surface, thus inhibiting the formation of biofilms on the surface. Therefore, SLA-TiCu with outstanding antibacterial performance has great potential for application in dental implants.
引文
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[13]REN L,MEMARZADEH K,ZHANG S,et al.A novel coping metal material CoCrCu alloy fabricated by selective laser melting with antimicrobial and antibiofilm properties[J].Materials science&engineering C:Materials for biological applications,2016,67:461-467.
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[22]ZHANG S,YANG C,REN G,et al.Study on behavior and mechanism of Cu2+ion release from Cu bearing antibacterial stainless steel[J].Materials technology,2015,30(B2):126-132.
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[24]LIU R,TANG Y,ZENG L,et al.In vitro and in vivo studies of anti-bacterial copper-bearing titanium alloy for dental application[J].Dental materials,2018,34(8):1112-1126.
[25]LIU R,MEMARZADEH K,CHANG B,et al.Antibacterial effect of copper-bearing titanium alloy(Ti-Cu)against Streptococcus mutans and Porphyromonas gingivalis[J].Scientific reports,2016,6:29985.
[26]MA Z,LI M,LIU R,et al.In vitro study on an antibacterial Ti-5Cu alloy for medical application[J].Journal of materials science materials in medicine,2016,27(5):91.
[27]SNT 2399-2010,抗菌金属材料评价方法[S].SNT 2399-2010,Evaluation method for antimicrobial metallic materials[S].
[28]ISO-10993-5,Biological evaluation of medical devicesPart 5:Tests in vitro forcytotoxicity:in vitro methods[S].
[29]HAN A,TSOI J K H,RODRIGUES F P,et al.Bacterial adhesion mechanisms on dental implant surfaces and the influencing factors[J].International journal of adhesion and adhesives,2016,69:58-71.
[30]JUNG W K,KOO H C,KIM K W,et al.Antibacterial activity and mechanism of action of the silver ion in Staphylococcus aureus and Escherichia coli[J].Applied and environmental microbiology,2008,74:2171-2178.
[2]HOLMBERG KV,ABDOLHOSSEINI M,LI Y,et al.Bioinspired stable antimicrobial peptide coatings for dental applications[J].Acta biomaterialia,2013,9(9):8224-8231.
[3]WANG X J,LIU H Y,REN X,et al.Effects of fluoride-ion-implanted titanium surface on the cytocompatibility in vitro and osseointegatation in vivo for dental implant applications[J].Colloids and surfaces B:Biointerfaces,2015,136:752-760.
[4]CHENG M,QIAO Y,WANG Q,et al.Calcium plasma implanted titanium surface with hierarchical microstructure for improving the bone formation[J].ACS applied materials&interfaces,2015,7(23):13053-13061.
[5]APARICIO C,PADROS A,GIL F J.In vivo evaluation of micro-rough and bioactive titanium dental implants using histometry and pull-out tests[J].Journal of the mechanical behavior of biomedical materials,2011,4(8):1672-1682.
[6]DAVIES J E,AJAMI E,MOINEDDIN R,et al.The roles of different scale ranges of surface implant topography on the stability of the bone/implant interface[J].Biomaterials,2013,34(14):3535-3546.
[7]ALBREKTSSON T,WENNERBERG A.Oral implant surfaces.Part 2.Review focusing on clinical knowledge of different surfaces[J].International journal of prosthodontics,2004,17(5):544-564.
[8]LAUER G,WIEDMANN-AL-AHMAD M,OTTEN J E,et al.The titanium surface texture effects adherence and growth of human gingival keratinocytes and human maxillar osteoblast-like cells in vitro[J].Biomaterials,2001,22(20):2799-2809.
[9]XIE Y,LI J,YU Z M,et al.Nano modified SLA process for titanium implants[J].Materials letters,2017,186:38-41.
[10]GITTENS RA,MCLACHLAN T,OLIVARES-NAVAR-RETE R,et al.The effects of combined micron-/submicronscale surface roughness and nanoscale features on cell proliferation and differentiation[J].Biomaterials,2011,32(13):3395-3403.
[11]BLATT S,PABST A M,SCHIEGNITZ E,et al.Early cell response of osteogenic cells on differently modified implant surfaces:Sequences of cell proliferation,adherence and differentiation[J].Journal of craniomaxillofacal surgery,2017,46(3):453-460.
[12]BURGHARDT I,LUTHEN F,PRINZ C,et al.A dual function of copper in designing regenerative implants[J].Biomaterials,2015,44(44):36-44.
[13]REN L,MEMARZADEH K,ZHANG S,et al.A novel coping metal material CoCrCu alloy fabricated by selective laser melting with antimicrobial and antibiofilm properties[J].Materials science&engineering C:Materials for biological applications,2016,67:461-467.
[14]ZHAO L,CHU P K,ZHANG Y,et al.Antibacterial coatings on titanium implants[J].Journal of biomedical materials research part B:Applied biomaterials,2009,91B(1):470-480.
[15]HAN A,LI X,HUANG B,et al.The effect of titanium implant surface modification on the dynamic process of initial microbial adhesion and biofilm formation[J].International journal of adhesion and adhesives,2016,69:125-132.
[16]HARDES J,AHRENS H,GEBERT C,et al.Lack of toxicological side-effects in silver-coated megaprostheses in humans[J].Biomaterials,2007,28(18):2869-2875.
[17]GOODMAN S B,YAO Z,KEENEY M,et al.The future of biologic coatings for orthopaedic implants[J].Biomaterials,2013,34:3174-3183.
[18]HARRIS L G,TOSATTI S,WIELAND M,et al.Staphylococcus aureus adhesion to titanium oxide surfaces coated with non-functionalized and peptide-functionalized poly(L-lysine)-grafted-poly(ethylene glycol)copolymers[J].Biomaterials,2004,25:4135-4148.
[19]ZHENG Y,LI J,LIU X,et al.Antimicrobial and osteogenic effect of Ag-implanted titanium with a nanostructured surface[J].International journal of nanomedicine,2012,7:875-884.
[20]ADIL M,SINGH K,VERMA P K,et al.Eugenol-induced suppression of biofilm-forming genes in Streptococcus mutans:An approach to inhibit biofilms[J].Journal of global antimicrobial resistance,2014,2(4):286-292.
[21]ZHANG E,LI F,WANG H,et al.A new antibacterial titanium-copper sintered alloy:preparation and antibacterial property[J].Materials science&engineering C:Materials for biological applications,2013,33(7):4280-4287.
[22]ZHANG S,YANG C,REN G,et al.Study on behavior and mechanism of Cu2+ion release from Cu bearing antibacterial stainless steel[J].Materials technology,2015,30(B2):126-132.
[23]REN L,NAN L,YANG K.Study of copper precipitation behavior in a Cu-bearing austenitic antibacterial stainless steel[J].Materials&design,2011,32(4):2374-2379.
[24]LIU R,TANG Y,ZENG L,et al.In vitro and in vivo studies of anti-bacterial copper-bearing titanium alloy for dental application[J].Dental materials,2018,34(8):1112-1126.
[25]LIU R,MEMARZADEH K,CHANG B,et al.Antibacterial effect of copper-bearing titanium alloy(Ti-Cu)against Streptococcus mutans and Porphyromonas gingivalis[J].Scientific reports,2016,6:29985.
[26]MA Z,LI M,LIU R,et al.In vitro study on an antibacterial Ti-5Cu alloy for medical application[J].Journal of materials science materials in medicine,2016,27(5):91.
[27]SNT 2399-2010,抗菌金属材料评价方法[S].SNT 2399-2010,Evaluation method for antimicrobial metallic materials[S].
[28]ISO-10993-5,Biological evaluation of medical devicesPart 5:Tests in vitro forcytotoxicity:in vitro methods[S].
[29]HAN A,TSOI J K H,RODRIGUES F P,et al.Bacterial adhesion mechanisms on dental implant surfaces and the influencing factors[J].International journal of adhesion and adhesives,2016,69:58-71.
[30]JUNG W K,KOO H C,KIM K W,et al.Antibacterial activity and mechanism of action of the silver ion in Staphylococcus aureus and Escherichia coli[J].Applied and environmental microbiology,2008,74:2171-2178.
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