acterizingwww

http://www.henhenlu.com/  时间:2021-03-05  阅读:()
Non-invasivecharacterizationofstructureandmorphologyofsilkbroinbiomaterialsusingnon-linearmicroscopyWilliamL.
Ricea,ShamarazFirdousb,SharadGuptaa,MartinHuntera,CherylW.
P.
Fooa,YongzhongWanga,HyeonJooKima,DavidL.
Kaplana,IreneGeorgakoudia,*aBiomedicalEngineeringDepartment,TuftsUniversity,4ColbyStreet,Room229,Medford,MA02155,USAbLaserLaboratory,DepartmentofPhysicsandAppliedMathematics(DPAM),PakistanInstituteofEngineeringandAppliedSciences(PIEAS),P.
O.
Nilore,Islamabad,PakistanReceived10October2007;accepted22December2007Availableonline21February2008AbstractDesigningbiomaterialscaffoldsremainsamajorchallengeintissueengineering.
Keytothischallengeisimprovedunderstandingoftherelationshipsbetweenthescaffoldpropertiesanditsdegradationkinetics,aswellasthecellinteractionsandthepromotionofnewmatrixdeposition.
Herewepresenttheuseofnon-linearspectroscopicimagingasanon-invasivemethodtocharacterizenotonlymorphological,butalsostructuralaspectsofsilkwormsilkbroin-basedbiomaterials,relyingentirelyonendogenousopticalcontrast.
Wedemonstratethattwophotonexciteduorescenceandsecondharmonicgenerationaresensitivetothehydration,overallbsheetcontentandmolecularorientationofthesample.
Thus,thefunctionalcontentandhighresolutionaffordedbythesenon-invasiveapproachesofferpromiseforidentifyingimportantconnectionsbetweenbiomaterialdesignandfunctionalengineeredtissuedevelopment.
Thestrategiesdescribedalsohavebroaderimplicationsforunderstandingandtrackingtheremodelingofdegradablebiomaterialsunderdynamicconditionsbothinvitroandinvivo.
2008ElsevierLtd.
Allrightsreserved.
Keywords:Silk;Fibroin;Twophotonexciteduorescence;Secondharmonicgeneration;Spectralanalysis;Non-invasivecharacterization1.
IntroductionDesignoptimizationofthebiomaterialsthatareusedasscaffoldsontowhichneworregeneratedtissuesareexpectedtogrowisoneofthemajorchallengesfacingtissueengineers.
Numerousaspectsofthebiomaterialsdesign,fromthemolec-ularstructureandorganizationtotheoverallmacro-architec-tureofthesesystemsareknowntoimpactsignicantlythedevelopmentofnewtissueinvitroandinvivo[1e4].
Scaf-foldsprovidestructuralsupportandimportantenvironmentalcuestocellsthatpopulatethem,thuscontrollingtoalargeextentcellularproliferation[1,5],differentiation[6e8],depo-sitionofnewstructuralproteins[9,10]andultimatelytheregenerationoffunctionaltissue[4,11,12].
Traditionally,methodssuchasNMR,FTIRandX-rayspectroscopyhavebeenusedtoassessbiomaterialsatthemolecularlevel[13e17],whileSEMandTEMhavebeeninvaluabletoolsforchar-acterizingthethree-dimensionalmorphologyofbiomaterialscaffolds[5,18e23].
Histologyandimmunostainingaswellasassaysfordeterminingtheexpressionlevelsofspecicpro-teinsareoftenusedtoassesshowscaffoldsinteractwithcellsastissuesdevelop[5,10,18,20,22e24].
Whilealloftheseap-proachesprovidesensitiveandspecicdata,theyareinvasive.
Asaresult,theyrevealinformationaboutasingletime-pointalongthedevelopmentofadynamicallychangingspecimen.
Thislimitationhindersfullcharacterizationandunderstandingoftherelationshipsthatexistbetweenthestructural,mechan-ical,architecturalandbiochemicalpropertiesofthescaffoldandthecorrespondingpropertiesofthedevelopingtissue.
Anumberofopticalmethodshavebeendevelopedtomon-itornon-invasivelydifferenttissuecomponentsinthecontextofdiseasediagnosisandmonitoring[25e29].
Suchmethods*Correspondingauthor.
Fax:16176273231.
E-mailaddress:irene.
georgakoudi@tufts.
edu(I.
Georgakoudi).
0142-9612/$-seefrontmatter2008ElsevierLtd.
Allrightsreserved.
doi:10.
1016/j.
biomaterials.
2007.
12.
049Availableonlineatwww.
sciencedirect.
comBiomaterials29(2008)2015e2024www.
elsevier.
com/locate/biomaterialshaveonlyrecentlystartedtobeexploitedastoolsforassessingdifferentpropertiesofthecellandmatrixcomponentsofengineeredtissues[30e33].
Thegoalofthisstudywastode-terminethetypeofmorphologicalandstructuralinformationthatcouldbeacquiredaboutsilk-basedbiomaterialscaffoldsusingspectraltwophotonexciteduorescence(TPEF)andsecondharmonicgeneration(SHG)imaging.
Thisinformationwillbeessentialindevelopinganopticalbiomarkertoolkitthatwillallowustomonitordynamicallyhowsuchscaffoldsinteractwithandaremodiedbycellsasengineeredtissuesgroweitherinvitroorinvivo.
Furthermore,theseopticaltoolkitscanalsobeextendedtootherbiomaterialmatricesasthesystemsaredevelopedandoptimized.
Silkisanaturalproteinpolymervaluedforitsbiocompat-ibility,lightweight,andstrength[34].
Processingmethodsforthispolymerarewellestablishedforcontrolofmorphology,mechanicalpropertiesandenvironmentalstability[35e38].
Duetotheseproperties,silkisanexcellentcandidateforgen-eratingbiomaterialscaffoldsforengineeredtissues.
TheBombyxmorisilkwormsilkprotein,broin,canbedescribedbytwostructuralmodels:SilkI,consistingoftypeIIbturn,randomcoildomains,andmixedstructuresincludingalphahelices,andSilkII,consistingmostlyofantiparallelbpleatedsheets[14,15].
Thebsheetcontentandthealignmentofthesebsheetcrystals,alongwiththenon-crystallinedomainsoftheprotein,areimportantdeterminantsofthebulkmechanicalpropertiesanddegradationkineticsofbiomaterialsgeneratedfromsilk[14,36,37,39e41].
Mostofthebsheetcontentandorientationofthesecrystallinedomainsislostduringthepro-cessingofsilkbroinintoaqueoussolutions,asteprequiredfortheregenerationofnewbiomaterialscaffoldsfortissueculture[42].
Thebsheetcontentandorientationcanberecon-stitutedtodifferentextentsdependingonthemodeofmaterialpreparation[14,38,42].
Thus,thenon-invasive,opticalassess-mentofbsheetcontentandorientationofsilkbroinduringbiomaterialscaffoldformationwasoneofthespecicgoalsofthisstudy.
Linearopticalapproaches,suchasuorescenceandRamanspectroscopyhavebeenusedpreviouslytocharacterizemate-rialsmadefromsilkbroin[39,43,44].
However,therearenostudiestoourknowledgeonthenon-linearopticalpropertiesofsilk.
Non-linearopticalmethodssuchasTPEFandSHGofferadditionaladvantagesfornon-invasiveimaging,includingexcitationinthenearinfraredregionofthespectrum,wherescatteringistypicallylowerthaninthevisibleregion,andreducedphotobleaching[45].
InTPEFandSHG,twophotons,typicallyofthesameenergy,interactsimultaneouslywithamoleculeandyieldeitheruorescenceemission(TPEF)orscattering(SHG)ofasinglephoton.
InthecaseofSHG,thescatteredphotonhasthesameenergyasthecollectiveenergyofthetwoincidentphotons(i.
e.
thereisnonetenergylossandthewavelengthofthescatteredphotonisatexactlyhalfthewavelengthofeachoneoftheincidentphotons).
InthecaseofTPEF,thewavelengthoftheincidentphotonsisapproxi-matelytwiceaslongasthewavelengthofaphotonrequiredforlinearexcitation,whiletheuorescentlyemittedphotonshavenearlyidenticalspectralfeaturesasthoseresultingfromsinglephotonexcitation.
Becausetheprobabilityofsi-multaneousinteractionwithtwophotonsisordersofmagni-tudelowerthansinglephotoninteractions,TPEFandSHGprocessesrequirethepresenceofhighphotondensities.
Asaresult,theseeventsareconnedwithinasmallvolumeintheapexofafocusedconeoflightandautomaticallyyieldopticallysectioned,depth-resolvedimages.
Theconnementoftheopticaleffect,andtheuseoflowenergy(longerwave-length)photonsresultsinreducedthermalandphotodamagewithinandoutsidetheplaneoffocus[45].
Exploitingsuchprocessesinmicroscopicimagingplatformsallowsfrequentsampleassessmentoverlongperiodsoftime,withoutdamageorcontaminationfromelementsoutsidethetissuecultureenvironment.
2.
MaterialsandmethodsAllchemicalswereobtainedfromSigmaAldrich(St.
Louis,MO)unlessotherwisenoted.
2.
1.
PreparationofsilkbroinCocoonsofB.
moriwerekindlyprovidedbyM.
Tsukada(InstituteofSericulture,Tsukuba,Japan).
Thecocoonswereinitiallyprocessedtoremovetheglue-likesericinproteinsandtoextractthepuresilkbroincomponentaspreviouslydescribed[39].
Theresultingaqueoussilkbroinsolutionhadanapproximateconcentrationof8%(wt)silkbroin.
Forhexauoro-2-propanol(HFIP)silkbroinsolutions,aqueoussilkwaslyophilizedandredissolvedinHFIPresultingina6%(w/v)broinsolution.
2.
2.
PreparationoflmsandgelsSilkbroinlmsweremadebypipettingvolumesofaqueousorHFIPsilksolutionsonto35mmglassbottomdishes(MatTek,ActonMA)andallowingthesolventtoevaporate.
Gelsweremadefromaqueoussolutionsin35mmglassbottomdishesatsilkconcentrationsof8and4%with6%(vol)0.
1MHCl.
Thesilksolutionswereplacedonthedishesrstandtheacidwastitratedintothesolutionforanalvolumeof2.
5mL.
Disheswerethensealedwithparafnlmandplacedat37Cfor72h[42].
bsheetcontentwasinducedinsilkbroinlmsbyimmersioninmethanolfor30min.
Filmswerecom-pressedandstretchedbyplacinglmsinopposingclamps,applyingpressuretosecurethelmandthendrawingtheclampsapart.
2.
3.
PreparationofscaffoldsThree-dimensionalsilkscaffoldswerepreparedaspreviouslydescribed[39].
Thescaffoldswerecutintodiscs(5mmindiameterand3mminthick-ness)driedat60C,andautoclavedforfurtherexperiments.
Theporesizeofthescaffoldswas55030mm.
Humanmesenchymalstemcells(Cambrex,EastRutherfordNJ)wereseededontoanaqueoussilkscaffoldasdescribedbyKimetal.
[18].
Theseededscaffoldswereplacedonglassbottomdishesandimagedonday21.
2.
4.
TwophotonexciteduorescenceandsecondharmonicgenerationmicroscopyTPEFandSHGmicrographswereacquiredonaLeicaDMIRE2micro-scopewithaTCSSP2scanner(Wetzlar,Germany).
Thesystemwasequippedwitha10(NA0.
3)dry,20(NA0.
7)dryanda63(NA1.
2)waterimmer-sionobjective.
TheexcitationlightsourcewasaMaiTaitunable(710e920nm)titaniumsapphirelaseremitting100fspulsesat80MHz(SpectraPhysics,MountainViewCA).
Sampleswereplacedonculturedisheswithnumber1.
5coverglassbottoms(MATTEK,AshlandMA)andexcitedat2016W.
L.
Riceetal.
/Biomaterials29(2008)2015e2024DownloadEnglishVersion:https://daneshyari.
com/en/article/10486DownloadPersianVersion:https://daneshyari.
com/article/10486Daneshyari.
com

DiyVM:499元/月香港沙田服务器,L5630*2/16G内存/120G SSD硬盘/5M CN2线路

DiyVM是一家成立于2009年的国人主机商,提供的产品包括VPS主机、独立服务器租用等,产品数据中心包括中国香港、日本大阪和美国洛杉矶等,其中VPS主机基于XEN架构,支持异地备份与自定义镜像,VPS和独立服务器均可提供内网IP功能。商家VPS主机均2GB内存起步,三个地区机房可选,使用优惠码后每月69元起;独立服务器开设在香港沙田电信机房,CN2线路,自动化开通上架,最低499元/月起。下面以...

ZoeCloud:香港BGP云服务器,1GB内存/20GB SSD空间/2TB流量/500Mbps/KVM,32元/月

zoecloud怎么样?zoecloud是一家国人商家,5月成立,暂时主要提供香港BGP KVM VPS,线路为AS41378,并有首发永久8折优惠:HKBGP20OFF。目前,解锁香港区 Netflix、Youtube Premium ,但不保证一直解锁,谢绝以不是原生 IP 理由退款。不保证中国大陆连接速度,建议移动中转使用,配合广州移动食用效果更佳。点击进入:zoecloud官方网站地址zo...

舍利云30元/月起;美国CERA云服务器,原生ip,低至28元/月起

目前舍利云服务器的主要特色是适合seo和建站,性价比方面非常不错,舍利云的产品以BGP线路速度优质稳定而著称,对于产品的线路和带宽有着极其严格的讲究,这主要表现在其对母鸡的超售有严格的管控,与此同时舍利云也尽心尽力为用户提供完美服务。目前,香港cn2云服务器,5M/10M带宽,价格低至30元/月,可试用1天;;美国cera云服务器,原生ip,低至28元/月起。一、香港CN2云服务器香港CN2精品线...

http://www.henhenlu.com/为你推荐
广东GDP破10万亿中国GDP10万亿,广东3万亿多。占了中国三分之一的经纪。如果,我是说如果。广东独立了。中国会有什李子柒年入1.6亿新晋网红李子柒是不是背后有团队是摆拍、炒作为的是人气、流量?曲妙玲张婉悠香艳版《白蛇传》是电影还是写真集?冯媛甑夏如芝是康熙来了的第几期?www.idanmu.com新开奇迹SF|再创发布网|奇迹SF|奇迹mu|网通奇迹|电信奇迹|www.zhiboba.com登录哪个网站可以看nba当天的直播 是直播baqizi.cc誰知道,最近有什麼好看的電視劇m.yushuwu.org花样滑冰名将YU NA KIM的资料谁有?www.toutoulu.com安装好派克滤芯后要检查其是否漏气www.175qq.com这表情是什么?
美国网站空间 韩国vps 域名服务器的作用 已备案域名出售 securitycenter 网站保姆 个人免费空间 100x100头像 天互数据 韩国名字大全 免费dns解析 香港新世界中心 闪讯官网 smtp虚拟服务器 photobucket 双线空间 工信部icp备案查询 域名转入 测试网速命令 中国联通宽带测试 更多