新型氟化丝素 TiO2纳米复合材料支架外文文献

新型氟化丝素新型氟化丝素 TiO2TiO2纳米复合材料支架外文文献纳米复合材料支架外文文献 Contents listsavailable atScienceDirectMaterials ScienceReceived inrevised form26Augustxx Aepted1Septemberxx Corresponding author E mail address madaah sharif H R Madaah Hosseini Materials Sciencew1 w2andw3arethe weightof bottlefilled withethanol bottle containingethanol andscaf f old andbottle takenout ofethanol saturated scaf f old respectively eisthedensity ofethanol Fig 1 A FT IR spectra of a TiO2and b TiO2 F and B XRD patternsof a TiO2and b TiO2 F nanoparticles N Johari etal Materials ScienceTakaraBio INC Japan solution at37 C andpH7 4for periodsup to30days Each samplewas takenout rinsed and dried in5days intervals Thedried scaf f oldswere weightedand theweight lossduring degradationwascalculated usingeq 2 57 58 Three sampleswere usedfor eachcondition weight loss W W W1000t0 2 where W0and Wt aredry scaf f oldsweight atthe timevalues equalto0andt respectively For eachimmersed sample pH of the phosphatebuffered salinesolutionwas measuredby acalibrated pHmeter pH andion meterGLP22 Crison Spain every2days 2 8 In vitrobioactivity evaluationsThebioactivity behaviour of theSF TiO2 F scaf f oldswere in vestigated bysoaking in SBF where pHof SBFwas7 4at37 C Thestandard SBFsolutionwassynthesized aording to Kokubo s protocol 59 Prepared scaf f oldswere immersedin SBFat37 C for0 7 14 21and28days The scaf f oldswere washedwith distilledwater aftereachsoaking steptime and then dried Afterwards the concentrationsofcalcium andphosphorus ionsin SBFwere measured using ICP OES VISTA PRO Varian Co Australia Thefl uoriderelease aftereachsoaking stepand thevariations inpH weredetermined byan ion andpH meter GLP22 Crison Spain For eachcondition three sampleswereused The apatiteformation on the surface of thescaf f olds wasstudiedby SEMimages andits elementalanalysis wasevaluatedbyanenergy dispersive X ray spectroscopy EDX device TESCAN CzechRepublic In addition the phasestructure and functional groupsof theformedlayer on the surface of nanopositescaf f oldswerein vestigated usingX ray diff raction XRD patterns andFourier Trans form Infra Red FTIR Spectra of scaf f olds beforeand afterimmersion inSBFsolution 2 9 Cell culture2 9 1 Cell morphologyThehumanlikeSaOS 2osteoblast cell line Cell Bank PasteurInstitute ofIran was usedfor culturingin fl askscontaining Dulbeo smodif ied Eagle s medium DMEM Biowest France supplemented with10 fetal bovineserum FBS Biowest France 1 105cells wereculturedona96well platecontaining theSF TiO2 F scaf f oldsandcultured for1 3and5days inan incubatorhumidif iedwith5 CO2 95 air at37 C After3days ofculturing cells werefixed withglu taraldehyde 2 5 dehydrated withgraded ethanol 10 20 30 40 50 60 70 90and100vol anddried The morphologyof theat tached cellswas observedwith SEM 2 9 2 MTT assayAfterincubation ofSaOS 2cells for1 3and5days on SF TiO2 Fscaf f olds the cellswere washedwith DMEM The mediumwas replacedwith450 l DMEMand mixedwith50 l of5mg ml MTT After4hincubation at37 C the mediumwas removedfrom eachwell andref illed with0 1ml ofdimethyl sulphoxide DMSO Biowest France todissolve theformazan crystals The plateswere agitatedto pletelydissolveformazan crystals and theabsorbance at570nm wasmea suredusinga scanningmulti well spectrophotometer Anthos2020 Austria A blankOptical Density OD value wasderived fromeachsample reading Data werepresented asmean standard errorof themean Comparisons of the meanvalues wereperformedbyone wayanalysis ofvariance ANOVA using SPSS17 0statistical software 3 Results anddiscussion3 1 Phase structureandfunctionalgroups analysesFig 1 A shows theresults ofFT IR analysisfor TiO2and TiO2 Fnanoparticles In the TiO2spectrum Fig 1 A a the absorption linecorresponding toTieOeTi ispresented at476cm 1and therespectiveOeH absorptionbands hasappeared at1639cm 1and3430cm 1 While the characteristicTieOeTi bandin the TiO2eF spectrum Fig 1 A b is atthe samewave number the OeHbands haveex perienced ashift andappeared at1631cm 1and3418cm 1 Thisshift isattributed to the structuralchanges ourredbyfl uorineaddi tion Itisbelieved thatsome ofOH anionson theTiO2surface arebeingreplaced by Fluorine anionswhich areresponsiblefor the structurealterationon the surface of fluoridate TiO2nanoparticles In addition the OHabsorptionlineintensity isreduced due tothedecrease numberofOH bandson thesurface of TiO2 However TieOeTi absorptionFig 2 A FT IR spectraof a pure SF b SF 5TiO2 F c SF 10TiO2 F d SF 15TiO2 F and e SF 20TiO2 F and B XRD patternsof a TiO2nanoparticles b SF 5TiO2 F c SF 10TiO2 F d SF 15TiO2 F and e SF 20TiO2 F scaf f olds N Johari etal Materials Science Engineering C82 2018 265 276267band is not shifted Thus the crystalstructureof TiO2has remainedthesame and TiO2 x Fx poundwas notformed This justifi cationisconf i rmedby XRD patternspresented in Fig 1 B Comparing theXRDpatterns ofTiO2 Fig 1 B a and TiO2 F Fig 1 B b reveals that noadditional peaksor peakshift areintroduced afterfluoridation The FTIR spectraof SF TiO2 F nanopositescaf f olds withdif ferent amounts ofTiO2 F areshown in Fig 2 A amide groupsof Pro tein arenormally appearedat diff erentvibration peakranges Thecharacteristic vibrationbands ofproteins areobserved at1650 1630cm 1for amideI C O stretching 1540 1520cm 1forFig 3 SEM Micrographs ofA pure SF B SF 5TiO2 F C SF 10TiO2 F D SF 15TiO2 F and E SF 20TiO2 F scaf f olds Table1Porosity percentand pore size of pure SF and SF TiO2 F nanopositescaf f olds Valuesare mean SD n 3 Sample Porosity Pore size m SF88 2460 80SF 5TiO2 F85 5405 50SF 10TiO2 F82 2350 70SF 15TiO2 F80 5320 55SF 20TiO2 F78 3270 80N Johari etal Materials Science Engineering C82 2018 265 276268amide II secondary NeHbending and1270 1230cm 1for amideIII CeN andNeH functionalgroups 60 X ray diff ractionpatterns ofSF TiO2 F nanopositescaf f oldsareillustrated in Fig 2 B Thecharacteristic peaksof SF 60 andTiO2 aording toXRD JCPDS dataf ile No 04 0477 peaks canbe recognized The intensityofTiO2peaksincreased by TiO2content in the nanopositescaff olds 61 The lackofany additionalpeaks provesthatnochemical reactionhas ourredbetweenTiO2 Fand SF 62 3 2 Morphology studiesFig 3depicts theSEM micrographsof theSF TiO2 F scaff oldsbyvarious amountsofTiO2 F nanoparticles Micrographsof the producedscaff olds with0 5 10 15and20wt are presentedin Figs 3 A 3 B 3 C 3 D and3 E Respectively The openpore structures of SF TiO2 F scaff oldsare observablein Fig 3 High levelsof interconnectedporosity aswell asan appropriateporesize arerequired toof fer sufficient spacefortissuegrowth andtoincrease thevolume ofthe invasion of surroundingtissue inbio patible andbiodegradable scaff olds 63 64 Sinsawat etal 65 have demonstratedthat poresize between50and150 m couldcauseosteoid growthand poreslarger than150 m promotedproliferation ofcells vascular ingrowth and internalmineralized boneformation Table1describes the porosity percentageand poresize ofall SF TiO2 F scaff olds The producednanopositescaff olds had openmacroporesand theporesizeis inthe rangeof200 500 m In addi tion from Table1 it isobvious thatan increaseinthe amount ofTio2 Fleads toa decreaseintheporesizeofthescaff olds which isconf irmedby SEM micrographs Fig 3 3 3 Mechanical propertiesFig 4 A and4 B show theresults ofthe pressive strength andmodulus of preparedporous SF TiO2 F scaff olds withdifferent amountsof TiO2 F respectively The pressive strengthofporous SF TiO2 Fscaf f olds increases byTiO2 F nanoparticlesweight percent Fig 4 A The maximumpressivestrengthwas achievedby SF 15TiO2 Fnanoposite scaff old It hasbeen reportedthat the mechanicalproperties ofscaff oldsareimproved byenhancing nanoparticlesto apolymer matrix 66 Furthermore increasing theamount ofporositydecreases the pressivestrengthofscaff olds 55 In addition Fig 4 B shows thatthe pressive modulus ofthescaff oldsalso in creases by theTiO2 F amount The pressivemodulus ofbiode gradable polymerposites reinforcedby HAor otherceramic par ticles hasbeen improvedduetothe presence ofarigid andinorganicparticulate phaseinapolymermatrix 66 67 Herein thepressivestrength andmodulusof SF TiO2 F nano positescaff olds havebeen improvedby increasing theTiO2 F na noparticles contentup to15wt Indeed themechanicalproperties ofSF 20TiO2 F scaffolddecreased duetotheaggregation ofTiO2 F na noparticles Fig 5illustrates TEM images of SF 15TiO2 Fand SF 20TiO2 F scaff olds It canbe seenthat TiO2 F nanoparticleshave beenuniformlydispersed inthe polymericSF matrixof SF 15TiO2 F nano positescaffolds Fig 5 A However intheSF 20TiO2 F nano positescaffold TiO2 F nanoparticlesdid nothave auniform dis persion in SFmatrix Fig 5 B that is the highTiO2 F nanoparticlescontentmade themsignif i cantly aggregate Indeed it isnot possibletoprepare theSF 20TiO2 F nanopositescaffold withuniform disper sionofTiO2 F nanoparticles Therefore this positionisnotpre ferred asa boissue engineeredscaffold Fig 4 TEMimagesofA SF 15TiO2 F and B SF 20TiO2 F Fig 5 A pressivestrength and B pressivemodulusof SF TiO2 F scaffolds N Johari etal Materials Science Engineering C82 2018 265 2762693 4 Biodegradation evaluationFig 6 A shows theweight lossof SF TiO2 F scaffoldsat diff erentincubationtimes The weight loss hasincreased withthe incubationperiodandthe amountsofTiO2 F After30days ofincubation theweight lossof SF TiO2 F scaffolds have changedaround2 5 in na noposites containing0 5 10 15and20wt ofTiO2 F nano particles Studies demonstratedthatthe gradual degradationcouldmatch thegrowth rateof newbone Indeed thegradualdegradation ofthescaffold providesthe structuralsupport fornewly grownbone 68 69 The weightofpureSF scaffoldshas notaltered toa largeextentduring thewhole incubationperiod Studies havedemonstrated thatmore pacted structureshave a reduced degradation 57 70 Ac cordingtoTable1 by increasing theamountsofTiO2 F nanoparticlesinscaf folds theporositypercentageofscaffolds isincreased andtheirstructures wouldbe lesspact Therefore the degradationrates ofnanopositescaffoldshaveincreased by increasing theconcentra tionofTiO2 F nanoparticles In fact morepactstructuresof SF TiO2 F nanopositescaffoldswithhigh SFconcentration hadare duced dissolution The pHpattern changeofthe PBS solutioncon taining SF TiO2 F nanopositescaffoldsareshown inFig 6 B ThepH ofthePBSsolution hasdecreased fromthe initialvalue of7 4in allnanopositescontaining0 5 10 15and20wt TiO2 F afterthef irst weekof soaking Acidif i cationofthe mediumis duetothe presenceof acidicproducts ofthe polymerdegradation 71 The degradationproductsof Fibroinare aminoacids suchas Glycineand Alanine 72 The acidityofthesolution increasesbyincreasingtheTiO2 F nano particles contentin scaffolds since thescaffolds degradationrate in creasesbyincreasingtheamountsofTiO2 F Fig 6 A Fig 7 A and7 B shows theSEM micrographsof SF 15TiO2 Fnanoposite scaffolds prior tothe immersion in PBSand after30days respectively Fig 7 B shows aslight degradationinSF TiO2 Fnanoposite scaffold and confi rmstheweightloss presentedinFig 6 A 3 5 In vitrobioactivity behaviourThein vitrobioactivityof SF TiO2 F nanopositescaffolds wasevaluatedby theformation of apatite on their surfacesduring immer sionin SBF undernormal physiologicalconditions at pH 7 4and37 C By incubatingfor0 7 14 21and28days in SBF as canbe seeninFig 8 a signif i cantnumber ofspherical particleshasbeenformed onthesurfaces ofthe samples after21days The particlesformation wasinvestigatedbyXRDand EDXanalyses Fig 9 A shows theXRD patternof SF TiO2 F nanopositescaffold after21days incubationin SBF As shown inFig 9 A the hydroxyapatite HA peaks arepresented at2 32 33and34 and theyareinagreement withJCPDSdatafileNo 03 0747which isrepresentative ofHA presenceon theSF TiO2 Fscaf folds surface The EDX analysis ofthese sphericalparticles afterFig 6 A weightlosschanges and B pH changesof PBSsolution usedfor immersionofSF TiO2 F nanopositeand pureSF scaffoldfor variousperiods Fig 7 SEM micrographsof SF TiO2 F nanopositescaffolds A before and B 30days afterimmersion inPBS N Johari etal Materials Science Engineering C82 2018 265 276270three weeksof immersionin SBFis showninFig 9 B High calciumandphosphorous peaksare detectedby EDXanalysis The Ca P ratioisabout1 64 which isclose toHA This Ca P ratioreveals theformationof Calcium def icient hydroxyapatite CDHA on thesurfaceofSF TiO2 F nanopositescaffolds 68 Typical FTIRspectra whichare showninFig 10independently confirmthe HAformation Fig 11 A and11 B show thechanges inCa and P concentrationafter immersing inSBF The obtainedresults revealedthat Caand Pconcentration in SBFmedium ofSF TiO2 F scaffoldsare decreasedat soakingperiod due tothepresence ofOeH groupson TiO2nanoparticles surfaces OeHgroups arethe activesites forapatite nucleation since theyare prooabsorb CaandPand formingapatite 73 74 The variationof fl uorineionconcentration releasedfromtheSF TiO2 F scaffolds incubated inSBFup to14days isshowninFig 11 C The higherbioactivityofSF TiO2 F nanopositescaffoldswithrespect toSF TiO2is evidencedbythe SEMmicrographs showninFig 12 Up to21days this behaviourisFig 8 SEMmicrographsofSF TiO2 F nanopo sitescaffoldsafter A 0 B 7 C 14 D 21 and E 28days incubationin SBF N Johari etal Materials Science Engineering C82 2018 265 276271mainly causedbytheformation ofspherical apatiteparticles on thesurface ofSF TiO2 F nanopositescaffold whichhave coveredmostof thesurface Fig 12 A However exceeding21days ofimmersinginSBF theamountofapatiteonthesurfaceofSF TiO2nan opositescaf fold isnot high Fig 12 B whereas ourprevious work 54 re vealedthatmost ofthesurfaceofSF TiO2nanopositescaffoldwascovered byapatite after28days 3 6 BiopatibilityFor cellularresponse evaluationoftheSF TiO2 F scaffolds theSaOS 2osteoblast like cellswere seededon all samples Fig 13depictsthe morphologiesofthe cell grownontheSF TiO2 F scaffoldsbearing0 5and15wt TiO2 F nanoparticlesafter beingcultured for3days The cellswere well attached astheTiO2 F nanoparticlesincreased suggesting theproper osteoconductingcharacteristics oftheSF TiO2 Fnanoposite scaffolds This behaviourforthecase ofSF 15TiO2 Fnanoposite scaffoldcanbeobserved inFig 13 C Dehydrogenase ofmitochondrial inliving cellsleads toreduce theyellowMTT toa purpleformazan dye Therefore it canbeusedtoestimate thecell viability Fig 14shows theabsorbance values ofpurple formazanat570nm forSF TiO2 F nanopositescaffolds withdifferentamountsofTiO2 F nanoparticles i e 0 5 10 15and20wt after1 3and5days cellculturing The cell viability onthesurfaceofall preparedsamples wassignifi cantly differedfrom thatofthetissueculture plate The sameresults wereobtained forallsamplesafter1 3and5days The absorbancevaluesofcells culturedonSF 15TiO2 Fnanoposite scaffoldswere noticeablyhigher thanthose culturedonother samples Aording toourpreviouswork 54 SF TiO2nanopositescaf foldshadno significanteff ect onthecell attachment andviability ofSaOS 2osteoblast cell line Here the effectofSF TiO2and SF TiO2 Fnanoposite scaffoldson cellattachment andviability ofhuman likeosteoblastSaOS 2celllinehasbeenpared Fig 15shows theFig 9 A XRD pattern and B EDXanalysisofSF TiO2 F na nopositescaffold after21days incubationinSBF Fig 10 FTIRspectraofSF TiO2 F nanopositescaffolda before immersingand b after21days incubationinSBF N Johari etal Materials Science Engineering C82 2018 265 276272pression ofcell viabilityofSF TiO2andSF TiO2 F nanopositescaffolds In addition Fig 16exhibits thecellattachmentofSF TiO2 Fig 16 A andSF TiO2 F Fig 16 B scaffolds The obtainedresultsrevealed thatthepresenceoffl uorideincreases thecellviability Fig 15 and attachment Fig 16 The improvementoftheosteoblastresponses dueto fl uorideaddition waspreviously reportedby Tiaineal 13 In fact Fluoride ionsby excitingosteogenic cells aeleratesbone regeneration 4 ConclusionsIn thisstudy novel SF TiO2 F nanopositescaffoldswerepre pared throughphase separationmethod containingdifferentamountsofTiO2 F nanoparticles 0 5 10 15 and20wt TiO2nanoparticleswere modifiedbyFluorine ions The obtainedresults demonstratedthatthe mechanicalbehaviourofscaffoldshavebeenimprovedbyin creasingtheTiO2 F nanoparticlesup to15wt However a detri mentaleffectwasobservedbyafurtherincreaseintheTiO2 F content Theporosityofscaffoldswas decreasedwith TiO2 F content The pre senceoffluorineionsalso improvedthe bioactivityand biopat ibility ofSF TiO2 F nanopositescaffolds Furthermore thecellat tachmentandviability werealso significantlyimproved In addition introducing fluoridetothenanoposite resultedin improvingtheosteoblast responses Evaluating themechanicalproperties bioactivitybehaviour andbiopatibility ofSF TiO2 F nanopositescaffoldsrevealed thatSF 15TiO2 F nanopositescaffold hadthe optimizedposition Fig 11 A Calcium B Phosphorous and C Fluorine ionconcentration changesafter immersinginSBF N Johari etal Materials Science Engineering C82 2018 265 276273Fig 12 A SF TiO2 F nanopositescaffoldafter21days and B SF TiO2nanopositescaffoldafter28days immersioninSBF Fig 13 SaOS 2celllinemorphologies after3days culturing ontheSF TiO2 F scaffoldsbearing A 0 B 5 and C 15wt TiO2 F nanoparticles Fig 14 SaOS 2celllineviability after1 3and5days cellculturingonSF TiO2 F nanopositescaffolds n 3 and p 0 05 N Johari etal Materials Science Engineering C82 2018 265 276274AcknowledgementsThe authorsare gratefulforthesupports byIran NationalScienceFoundation INSF under thegrant number94003968and thankIranSilkworm ResearchCenter ISRC and MissNaf ise Taromi References 1 M J Olszta X Cheng S S Jee R Kumar Y Y Kim M J Kaufman E P Douglas L B Gower Bone structureand formation a newperspective Mater Sci Eng R Rep 58 xx 77 116 2 R Murugan S Ramakrishna Development ofnanoposites forbone grafting Compos Sci Technol 65 xx 2385 2406 3 J R Porter T T Ruckh K C Popat Bone tissueengineering a reviewin bonebiomimeticsand drugdelivery strategies Biotechnol Prog 25 xx 1539 1560 4 J O Hollinger T A Ei