Ezigulini ezihlinzwa ngokufakelwa kwamathambo, izifo ezibangelwa amagciwane kanye nezimpendulo zamasosha omzimba ezibangelwa ukutheleleka bekulokhu kuyizingozi ezisongela ukuphila.Izinto ezivamile zebhayoloji zisengozini yokungcoliswa yizinto eziphilayo, okubangela ukuthi amagciwane ahlasele indawo elimele futhi abangele ukutheleleka kwangemva kokuhlinzwa.Ngakho-ke, kunesidingo esiphuthumayo sokuthuthukisa i-anti-infection kanye ne-immune yokuphunyuka kwezingubo zokufakelwa kwamathambo.Lapha, sithuthukise ubuchwepheshe obuthuthukisiwe bokuguqula indawo yokufakelwa kwamathambo ebizwa ngokuthi i-Lubricated Orthopedic Implant Surface (LOIS), egqugquzelwa ubushelelezi bezimbiza zezitshalo ze-pitcher.I-LOIS inoketshezi oluhlala isikhathi eside noluqinile oluxosha uketshezi oluhlukahlukene nezinto eziphilayo (okuhlanganisa amaseli, amaprotheni, i-calcium namagciwane).Ngaphezu kwalokho, siqinisekise ukuqina komshini ngokumelene nokuklwebheka namandla okulungisa ngokulingisa umonakalo ongenakugwenywa phakathi nokuhlinzwa kwe-in vitro.Imodeli yokuvuvukala komnkantsha wethambo logwaja yasetshenziswa ukutadisha kahle ukukala okulwa nebhayoloji kanye nekhono lokulwa nokutheleleka kwe-LOIS.Sibona ngeso lengqondo ukuthi i-LOIS, enezici zokunqanda i-biofouling kanye nokuqina kwemishini, iyisinyathelo esiya phambili ekuhlinzeni amathambo okungenazifo.
Namuhla, ngenxa yokuguga okuphelele, inani leziguli ezihlushwa izifo zamathambo (ezifana nokuphuka asebekhulile, izifo ezihlangene eziwohlokayo, kanye ne-osteoporosis) liye landa kakhulu (1, 2).Ngakho-ke, izikhungo zezokwelapha zibheka ukubaluleka okukhulu ekuhlinzweni kwamathambo, okuhlanganisa ukufakelwa kwamathambo kwezikulufu, amapuleti, izipikili namajoyinti okwenziwa (3, 4).Kodwa-ke, izimila zendabuko zamathambo kuye kwabikwa ukuthi zingakwazi ukunamathela kwebhaktheriya kanye nokwakheka kwe-biofilm, okungabangela ukutheleleka kwendawo yokuhlinzwa (SSI) ngemva kokuhlinzwa (5, 6).Uma i-biofilm yakhiwe ebusweni bokufakelwa kwamathambo, ukususwa kwe-biofilm kuba nzima kakhulu ngisho nokusetshenziswa kwemithamo emikhulu yemithi elwa namagciwane.Ngakho-ke, ngokuvamile kuholela ezifweni ezinzima zangemuva kokuhlinzwa (7, 8).Ngenxa yezinkinga ezingenhla, ukwelashwa kokufakelwa okuthelelekile kufanele kufake ukusebenza kabusha, kuhlanganise nokususwa kwazo zonke izimila kanye nezicubu ezizungezile;ngakho-ke, isiguli sizobhekana nobuhlungu obukhulu kanye nezingozi ezithile (9, 10).
Ukuxazulula ezinye zalezi zinkinga, izimila zamathambo ezi-eluting zenzelwe ukuvimbela ukutheleleka ngokuqeda amagciwane anamathele phezulu (11, 12).Nokho, isu lisabonisa imikhawulo eminingana.Kubikwe ukuthi ukufakwa isikhathi eside kwezimila ze-eluting zezidakamizwa kubangele ukulimala kwezicubu ezizungezile futhi kubangele ukuvuvukala, okungaholela ku-necrosis (13, 14).Ngaphezu kwalokho, izincibilikisi eziphilayo ezingaba khona ngemva kwenqubo yokukhiqiza izimila zamathambo ezi-eluting izidakamizwa, ezinqatshelwe ngokuphelele yi-US Food and Drug Administration, zidinga izinyathelo ezengeziwe zokuhlanzeka ukuze zihlangabezane namazinga azo (15).Ukufakwa kwezidakamizwa eluting kuyinselele ekukhishweni okulawulwayo kwezidakamizwa, futhi ngenxa yokulayishwa kwazo okulinganiselwe kwezidakamizwa, ukusetshenziswa kwesikhathi eside komuthi akunakwenzeka (16).
Elinye isu elijwayelekile ukumboza okokufakelwa nge-polymer evimbela ukungcola ukuze kuvinjwe izinto zebhayoloji namagciwane ukuthi anganamatheli phezulu (17).Isibonelo, ama-polymer e-zwitterionic adonse ukunaka ngenxa yezakhiwo zawo ezinganamatheli lapho ethintana namaprotheni e-plasma, amaseli, namagciwane.Kodwa-ke, inokulinganiselwa okuthile okuhlobene nokuzinza kwesikhathi eside kanye nokuqina kwemishini, okuvimbela ukusetshenziswa kwayo okungokoqobo ekufakweni kwamathambo, ikakhulukazi ngenxa yokukhuhla ngomshini phakathi nezinqubo zokuhlinzwa (18, 19).Ngaphezu kwalokho, ngenxa ye-biocompatibility yayo ephezulu, ukuntuleka kwesidingo sokuhlinzwa kokususwa, kanye nezakhiwo zokuhlanza indawo ngokugqwala, izimila zamathambo ezenziwe ngezinto ezibolayo zisetshenzisiwe (20, 21).Ngesikhathi sokugqwala, izibopho zamakhemikhali phakathi kwe-polymer matrix ziyaphulwa futhi zikhishwe phezulu, futhi abalandeli bahlanza indawo.Kodwa-ke, ukungcoliswa kwe-anti-biological ngokuhlanza indawo kusebenza ngempumelelo ngesikhathi esifushane.Ngaphezu kwalokho, izinto eziningi ezimuncayo ezihlanganisa i-poly(lactic acid-glycolic acid copolymer) (PLGA), i-polylactic acid (PLA) kanye nama-alloys asekelwe ku-magnesium zizobhekana nokuwohloka kwe-biodegradation nokungalingani emzimbeni, okuzothinta kabi ukuzinza kwemishini.(amashumi amabili nambili).Ngaphezu kwalokho, izingcezu zamapuleti ezinto eziphilayo zinikeza indawo yokunamathisela amagciwane, okwandisa amathuba okutheleleka ngokuhamba kwesikhathi.Le ngozi yokuwohloka kwemishini kanye nokutheleleka ikhawulela ukusetshenziswa okungokoqobo kokuhlinzwa kwepulasitiki (23).
Izindawo eziphakeme ze-Superhydrophobic (SHP) ezilingisa ukwakheka kwamaqabunga e-lotus seziphenduke isixazululo esingaba khona sezindawo ezilwa nokungcoliswa (24, 25).Lapho indawo ye-SHP icwiliswa oketshezini, amabhamuza omoya azovaleleka, ngaleyo ndlela enze amaphakethe omoya futhi avimbele ukunamathela kwebhaktheriya (26).Kodwa-ke, ucwaningo lwakamuva lubonise ukuthi indawo ye-SHP inokungalungi okuhlobene nokuqina kwemishini kanye nokuzinza kwesikhathi eside, okuvimbela ukusetshenziswa kwayo ekufakweni kwezokwelapha.Ngaphezu kwalokho, amaphakethe omoya azoncibilika futhi alahlekelwe izakhiwo zawo zokulwa nokungcolisa, ngaleyo ndlela kubangele ukunamathela okubanzi kwamagciwane ngenxa yendawo enkulu yendawo ye-SHP (27, 28).Muva nje, u-Aizenberg kanye nozakwabo bethule indlela emisha yokuhlanganisa i-anti-biofouling ebusweni ngokwakha indawo ebushelelezi egqugquzelwe isitshalo sembiza ye-Nepenthes (29, 30).Indawo ebushelelezi ikhombisa ukuzinza kwesikhathi eside ngaphansi kwezimo ze-hydraulic, ixosha uketshezi ngokwedlulele oketshezini lwebhayoloji, futhi inezinto zokuzilungisa.Kodwa-ke, ayikho indlela yokufaka i-coating ekufakweni kwezokwelapha okumile okuyinkimbinkimbi, futhi akuqinisekisiwe ukusekela inqubo yokuphulukisa yezicubu ezilimele ngemva kokufakwa.
Lapha, sethula i-lubricated orthopedic implant surface (LOIS), i-micro/nano-structured orthopedic implant surface futhi ehlanganiswe ngokuqinile nongqimba oluncane lwe-lubricant ukuvimbela ukuthi ingahlotshaniswa nokuhlinzwa kwepulasitiki Ukutheleleka kwamagciwane, okufana nokulungiswa kokuphuka.Ngenxa yokuthi isakhiwo se-fluorine-level micro/nano-level esisebenza kahle silungisa ngokuqinile okokugcoba esakhiweni, i-LOIS ethuthukisiwe ingaxosha ngokugcwele ukunamathela koketshezi oluhlukahlukene futhi igcine ukusebenza kokulwa nokungcolisa isikhathi eside.Izingubo ze-LOIS zingasetshenziswa ezintweni zomumo ohlukahlukene ezihloselwe ukuhlanganiswa kwamathambo.Izakhiwo ezinhle kakhulu zokulwa ne-biofouling ze-LOIS ngokumelene namagciwane e-biofilm [Pseudomonas aeruginosa kanye ne-Staphylococcus aureus (MRSA) engamelana ne-methicillin] nezinto zebhayoloji (amangqamuzana, amaprotheni kanye ne-calcium) kuqinisekisiwe ku-vitro.Izinga lokunamathela lokunamathela okubanzi ku-substrate lingaphansi kwe-1%.Ngaphezu kwalokho, ngisho nangemva kokucindezeleka komshini okufana nokuklwebheka kwendawo, ukuziphilisa okubangelwa ukugcoba okungena ngaphakathi kusiza ukugcina izakhiwo zayo zokulwa nokungcola.Imiphumela yokuhlolwa kokuqina komshini ibonisa ukuthi ngisho nangemva kokuguqulwa kwesakhiwo namakhemikhali, amandla aphelele ngeke ehliswe kakhulu.Ngaphezu kwalokho, ukuhlolwa kwe-in vitro okulingisa ukucindezelwa komshini endaweni yokuhlinzwa kwenziwa ukuze kuqinisekiswe ukuthi i-LOIS ingakwazi ukumelana nezingcindezi ezihlukahlukene zemishini ezenzeka phakathi nokuhlinzwa kwepulasitiki.Okokugcina, sisebenzise imodeli yokuphuka kwe-femoral esekwe ku-vivo, efakazele ukuthi i-LOIS inezinto eziphakeme zokulwa namagciwane kanye nokuhambelana kwe-biocompatibility.Imiphumela ye-radiological kanye ne-histological iqinisekise ukuthi ukuziphatha okuzinzile kwe-lubricant kanye nezakhiwo zokulwa ne-biofouling phakathi kwamaviki angu-4 ngemva kokufakelwa kungafinyelela ukusebenza okuphumelelayo kokulwa nokutheleleka kanye nokusebenza komzimba wokuphunyuka ngaphandle kokulibazisa inqubo yokuphulukisa amathambo.
Umfanekiso 1A ubonisa umdwebo wohlelo lwe-LOIS ethuthukisiwe, etshalwe ngezakhiwo zesikali esincane/se-nano kumodeli yokuphuka kwefemoral onogwaja ukuze kuqinisekiswe izici zayo ezinhle kakhulu zokulwa nebhayoloji kanye nezindawo zokulwa nokutheleleka.Indlela ye-biomimetic iyenziwa ukulingisa indawo engaphezulu yesitsha sebhodwe lamanzi, kanye nokuvimbela ukungcoliswa kwezinto eziphilayo ngokuhlanganisa ungqimba lwe-lubricant ngaphakathi kwesakhiwo se-micro/nano esingaphezulu.Indawo ejovwe nge-lubricant inganciphisa ukuthintana phakathi kwezinto zebhayoloji nendawo engaphezulu.Ngakho-ke, ngenxa yokwakhiwa kwezibopho zamakhemikhali ezinzile endaweni, inokusebenza okuhle kakhulu kwe-antifouling nokuzinza kwesikhathi eside.Ngenxa yalokho, izakhiwo ze-anti-biofouling zendawo yokugcoba zivumela ukusetshenziswa okuhlukahlukene okusebenzayo ocwaningweni lwe-biomedical.Kodwa-ke, ucwaningo olubanzi mayelana nokuthi le ndawo ekhethekile isebenzisana kanjani emzimbeni ayikaqedwa.Ngokuqhathanisa i-LOIS nama-substrates anqunu ku-vitro esebenzisa i-albumin namabhaktheriya e-biofilm, ukunganamatheli kwe-LOIS kungaqinisekiswa (Umfanekiso 1B).Ukwengeza, ngokukhipha amaconsi amanzi ku-substrate engenalutho ethambekele kanye ne-LOIS substrate (Umfanekiso S1 kanye ne-Movie S1), ukusebenza kokungcoliswa kwezinto eziphilayo kungaboniswa.Njengoba kuboniswe esithombeni se-fluorescence microscope, i-substrate evulekile efakwe ekumisweni kwamaprotheni namagciwane ibonise inani elikhulu lezinto eziphilayo ezinamathela phezulu.Kodwa-ke, ngenxa yezakhiwo zayo ezinhle kakhulu zokulwa ne-biofouling, i-LOIS ayibonisi neze i-fluorescence.Ukuze kuqinisekiswe izakhiwo zayo zokulwa ne-biofouling kanye ne-anti-infection, i-LOIS yasetshenziswa ebusweni bokufakelwa kwamathambo ukuze kuhlanganiswe amathambo (amapuleti nezikulufu) futhi yafakwa kumodeli yokuphuka onogwaja.Ngaphambi kokufakelwa, ukufakelwa kwe-orthopedic enqunu kanye ne-LOIS kwafakwa ekumisweni kwebhaktheriya amahora angu-12.I-pre-incubation iqinisekisa ukuthi i-biofilm iyakhiwa ebusweni bokufakelwa obala ukuze kuqhathaniswe.Umfanekiso 1C ukhombisa isithombe sendawo yokuphuka emavikini ama-4 ngemuva kokufakwa.Kwesokunxele, unogwaja onesimila esingenalutho samathambo wabonisa izinga elinzima lokuvuvukala ngenxa yokwakheka kwe-biofilm ebusweni bokufakelwa.Umphumela ophambene wabonwa konogwaja abafakwe i-LOIS, okungukuthi, izicubu ezizungezile ze-LOIS azizange zibonise izimpawu zokutheleleka noma izimpawu zokuvuvukala.Ngaphezu kwalokho, isithombe sokubona esingakwesokunxele sibonisa indawo yokuhlinzwa kanogwaja ngokufakwa obala, okubonisa ukuthi akukho okunamathelayo okuningi okukhona ebusweni bokufakelwa okuveziwe okutholwe ebusweni be-LOIS.Lokhu kubonisa ukuthi i-LOIS inozinzo lwesikhathi eside futhi inamandla okugcina izinto zayo zokungcolisa izinto eziphilayo kanye nezokunqanda ukunamathela.
(A) Umdwebo wohlelo lwe-LOIS nokufakwa kwayo kumodeli yokuphuka kwefemu kanogwaja.(B) Isithombe se-Fluorescence microscopy sephrotheni nebhaktheriya biofilm endaweni engenalutho kanye ne-LOIS substrate.Amaviki angu-4 ngemva kokufakelwa, (C) isithombe sesithombe sesayithi lokuphuka kanye (D) nesithombe se-X-ray (sigqanyiswe unxande obomvu).Isithombe sihlonishwa: Kyomin Chae, Yonsei University.
Onogwaja abahlinzekiwe, abatshalwe ngendlela engalungile babonisa inqubo evamile yokuphulukisa amathambo ngaphandle kwanoma yiziphi izimpawu zokuvuvukala noma ukutheleleka.Ngakolunye uhlangothi, izimila ze-SHP ezifakwe ngaphambili ekumisweni kwebhaktheriya zibonisa ukuvuvukala okuhlobene nokutheleleka ezicutshini ezizungezile.Lokhu kungabangelwa ukungakwazi kwayo ukuvimbela ukunamathela kwebhaktheriya isikhathi eside (Umfanekiso S2).Ukuze kuqinisekiswe ukuthi i-LOIS ayithinti inqubo yokuphulukisa, kodwa ivimbela izifo ezingase zibe khona ezihlobene nokufakelwa, izithombe ze-X-ray ze-matrix enhle eveziwe kanye ne-LOIS endaweni yokuphuka kwaqhathaniswa (Umfanekiso we-1D).Isithombe se-X-ray sokufakelwa kahle okungenalutho sabonisa imigqa ephikelelayo ye-osteolysis, ebonisa ukuthi ithambo belingaphuluki ngokuphelele.Lokhu kusikisela ukuthi inqubo yokubuyisela amathambo ingase ibambezeleke kakhulu ngenxa yokuvuvukala okuhlobene nokutheleleka.Ngokuphambene nalokho, kwabonisa ukuthi onogwaja abafakwe nge-LOIS base bepholile futhi abazange babonise noma iyiphi indawo esobala yokuphuka.
Ukuze kuthuthukiswe izimila zezokwelapha ngokuzinza nokusebenza kwesikhathi eside (okuhlanganisa ukumelana ne-biofouling), imizamo eminingi yenziwe.Kodwa-ke, ukuba khona kwezinto ezihlukahlukene zebhayoloji kanye nokuguquguquka kokunamathela kwezicubu kukhawulela ukuthuthukiswa kwezindlela zabo ezinokwethenjelwa emtholampilo.Ukuze sinqobe lokhu kushiyeka, senze isakhiwo esinengqimba encane/ye-nano kanye nendawo eguquliwe ngamakhemikhali, ethuthukiswa ngenxa yamandla aphezulu e-capillary kanye nokuhambisana kwamakhemikhali ukuze kugcinwe okokugcoba okubushelelezi ngokwezinga elikhulu kakhulu.Umfanekiso 2A ukhombisa yonke inqubo yokukhiqiza ye-LOIS.Okokuqala, lungiselela insimbi engagqwali yezinga lezokwelapha (SS) 304 substrate.Okwesibili, ukwakheka kwe-micro/nano kwakhiwe ku-substrate ye-SS ngokufakwa kwamakhemikhali kusetshenziswa isisombululo se-hydrofluoric acid (HF).Ukuze ubuyisele ukumelana nokugqwala kwe-SS, isisombululo se-nitric acid (HNO3) (31) sisetshenziselwa ukucubungula i-substrate eqoshiwe.I-passivation ithuthukisa ukumelana nokugqwala kwe-substrate ye-SS futhi ibambezela ngokuphawulekayo inqubo yokugqwala enganciphisa ukusebenza kukonke kwe-LOIS.Bese, ngokwenza i-monolayer (SAM) ezihlanganisayo ene-1H, 1H, 2H, 2H-perfluorooctyltriethoxysilane (POTS), indawo engaphezulu iguqulwa ngamakhemikhali ukuze kuthuthukiswe ukusebenzisana kwamakhemikhali phakathi kwendawo kanye ne-Lubricant Affinity ebushelelezi.Ukuguqulwa kwendawo kunciphisa ngokuphawulekayo amandla angaphezulu kwendawo eyakhiwe ngesikali esincane/se-nano, ehambisana namandla angaphezulu kokokuthambisa okubushelelezi.Lokhu kuvumela i-lubricant ukuba imanziswe ngokuphelele, ngaleyo ndlela yakhe ungqimba oluzinzile lwe-lubricant ebusweni.Indawo eshintshiwe ibonisa i-hydrophobicity ethuthukisiwe.Imiphumela ibonisa ukuthi okokugcoba okushelelayo kubonisa ukuziphatha okuzinzile ku-LOIS ngenxa yokuhlangana kwamakhemikhali aphezulu namandla e-capillary okubangelwa ukwakheka kwe-micro/nano (32, 33).Izinguquko ze-optical ebusweni be-SS ngemva kokuguqulwa kwendawo kanye nomjovo we-lubricant zafundwa.Isakhiwo esinezingqimba ezincane/nano esakhiwe phezulu singabangela izinguquko ezibukwayo futhi sibe mnyama ebusweni.Lesi sigameko sibalulwe emthelela wokuhlakazeka kokukhanya othuthukisiwe endaweni embi, okwandisa ukubonakaliswa okusabalele okubangelwa indlela yokubamba ukukhanya (34).Ukwengeza, ngemva kokujova i-lubricant, i-LOIS iba mnyama.Isendlalelo sokugcoba sibangela ukukhanya okuncane ukuthi kubonakale kusuka ku-substrate, ngaleyo ndlela kube mnyama i-LOIS.Ukuze kuthuthukiswe i-microstructure/nanostructure ukukhombisa i-engeli encane yokushelela (i-SA) ukuze kuzuzwe ukusebenza kwe-anti-biofouling, i-electron microscopy (SEM) yokuskena kanye namapheya e-athomu asetshenziselwa ukwenza izikhathi ezihlukile zokuhlanganisa i-HF (0, 3)., 15 kanye nemizuzu engu-60) Phoqa Isibonakhulu (AFM) (Umfanekiso 2B).Izithombe ze-SEM ne-AFM zibonisa ukuthi ngemva kwesikhathi esifushane sokuqoshwa (imizuzu emi-3 yokuqopha), i-substrate engenalutho yenze ubuhwaqaqe be-nano-scale obungalingani.Ubulukhuni bendawo bushintsha ngesikhathi sokuqopha (Umfanekiso S3).Ijika elishintshashintshayo isikhathi libonisa ukuthi ubulukhuni obungaphezulu buyaqhubeka nokwanda futhi bufinyelele umvuthwandaba ngemizuzu engu-15 yokuqoshwa, bese kuba nokwehla okuncane kuphela kwevelu yokumahhadla kubonakala emizuzwini engama-30 yokuluma.Kuleli qophelo, i-nano-level roughness iyaqoshwa, kuyilapho i-micro-level roughness ikhula ngamandla, okwenza ukuguquka kushintshe kuzinze.Ngemva kokuqopha imizuzu engaphezu kwengu-30, ukwanda okwengeziwe kokuqina kubonakala, okuchazwa ngokuningiliziwe ngale ndlela elandelayo: I-SS yakhiwe ngensimbi, ehlanganiswe nezakhi ezihlanganisa insimbi, i-chromium, i-nickel, i-molybdenum nezinye izakhi eziningi.Phakathi kwalezi zakhi, i-iron, i-chromium ne-molybdenum idlala indima ebalulekile ekwakheni ukuhwaqeka kwesikali se-micron/nano-scale ku-SS ngokufakwa kwe-HF.Ezigabeni zokuqala zokugqwala, insimbi ne-chromium kugqwalile ngoba i-molybdenum inokumelana nokugqwala okuphezulu kune-molybdenum.Njengoba i-etching iqhubeka, isixazululo se-etching sifinyelela ekugcwaleni kwendawo, okwenza ama-fluoride nama-oxides abangelwa ukucwiliswa.I-fluoride ne-oxide ziyagqwala futhi ekugcineni ziphinde zibeke endaweni engaphezulu, kwakheka ukuhwashaza kwendawo ebangeni le-micron/nano (31).Lobu bunzima beleveli encane/ye-nano budlala indima ebalulekile ezicini zokuziphilisa ze-LOIS.Ingaphezulu lesilinganiso esikabili likhiqiza umphumela we-synergistic, okwandisa kakhulu amandla e-capillary.Lesi simo sivumela okokugcoba ukuthi kungene endaweni ngokuzinza futhi kube nomthelela ezimpahleni zokuziphilisa (35).Ukwakheka kwe-roughness kuncike esikhathini sokuqopha.Ngaphansi kwemizuzu eyi-10 yokushumeka, indawo engaphezulu iqukethe kuphela ukuhwaqeka kwesikali se-nano, okunganele ukubamba okokugcoba okwanele ukuze kube nokumelana ne-biofouling (36).Ngakolunye uhlangothi, uma isikhathi se-etching sidlula imizuzu engama-30, ukuhwalala kwe-nano-scale okwakhiwa ukumiswa kabusha kwe-iron ne-chromium kuzoshabalala, futhi kuphela ukuhwaqeka kwe-micro-scale okuzosala ngenxa ye-molybdenum.Indawo egxile kakhulu ayinakho ubulukhuni be-nano-scale futhi ilahlekelwa umphumela we-synergistic wezigaba ezimbili, okuphazamisa kabi izici zokuziphilisa ze-LOIS.Izilinganiso ze-SA zenziwa kuma-substrates anezikhathi ezihlukile zokufaka ukufakazela ukusebenza okuphikisana nokungcoliswa.Izinhlobo ezahlukene zoketshezi zakhethwa ngokusekelwe ku-viscosity namandla angaphezulu, okuhlanganisa amanzi e-deionized (DI), igazi, i-ethylene glycol (EG), i-ethanol (EtOH) ne-hexadecane (HD) (Figure S4).Iphethini ye-etching eshintshashintshayo ibonisa ukuthi oketshezini oluhlukahlukene olunamandla ahlukene angaphezulu nama-viscosities, i-SA ye-LOIS ngemva kwemizuzu engu-15 yokushumeka iphansi kakhulu.Ngakho-ke, i-LOIS ilungiselelwe ukuthi imile imizuzu engu-15 ukuze yakhe i-micron kanye ne-nano-scale roughness, elungele ukulondoloza ngempumelelo ukuqina kwe-lubricant kanye nezakhiwo ezinhle kakhulu zokulwa nokungcola.
(A) Umdwebo wohlelo wenqubo yokukhiqiza enezinyathelo ezine ye-LOIS.Isingeniso sibonisa i-SAM eyenziwe ku-substrate.(B) Izithombe ze-SEM ne-AFM, ezisetshenziselwa ukuthuthukisa ukwakheka kwe-micro/nano ye-substrate ngaphansi kwezikhathi zokuqopha ezihlukene.I-X-ray photoelectron spectroscopy (XPS) ye-spectra ye-(C) Cr2p kanye no-(D) F1s ngemva kokudlula kwendawo kanye ne-SAM coating.noma, iyunithi engafanele.(E) Izithombe ezimelele amaconsi amanzi kuma-substrates angenalutho, aqoshiwe, e-SHP kanye ne-LOIS.(F) I-engeli yokuxhumana (CA) kanye nesilinganiso se-SA soketshezi olunokungezwani okuhlukile kwendawo ku-SHP ne-LOIS.Idatha ivezwa njengencazelo ± SD.
Khona-ke, ukuze kuqinisekiswe ukuguqulwa kwezakhi zamakhemikhali ezingaphezulu, i-X-ray photoelectron spectroscopy (XPS) yasetshenziswa ukutadisha ushintsho ekubunjweni kwamakhemikhali endaweni engaphansi kwe-substrate ngemva kokumbozwa kwendawo ngayinye.Umfanekiso 2C ukhombisa imiphumela yesilinganiso ye-XPS yendawo enamathiselwe ye-HF kanye nendawo ephathwayo ye-HNO 3.Iziqongo ezimbili eziyinhloko ku-587.3 kanye no-577.7 eV zingabalulwa kubhondi ye-Cr-O ekhona kusendlalelo se-chromium oxide, okuwumehluko omkhulu endaweni eqoshiwe ye-HF.Lokhu kubangelwa ikakhulukazi ukusetshenziswa kwe-iron ne-chromium fluoride phezu kwe-HNO3.I-etching esekelwe ku-HNO3 ivumela i-chromium ukuthi yakhe ungqimba lwe-oxide edlulayo endaweni, okwenza i-SS egxilile imelane nokugqwala.Emdwebeni 2D, i-spectra ye-XPS yatholwa ukuze kuqinisekiswe ukuthi i-silane esekwe ku-fluorocarbon yakhiwe phezulu ngemva kokunamatheliswa kwe-SAM, okuxosha uketshezi oluphakeme kakhulu ngisho naku-EG, igazi kanye ne-EtOH.Ukufakwa kwe-SAM kuqedwa ngokusabela kwamaqembu asebenzayo we-silane anamaqembu e-hydroxyl akhiwe ukwelashwa kwe-plasma.Ngenxa yalokho, kwabonwa ukwanda okukhulu kweziqongo ze-CF2 ne-CF3.Amandla okubopha phakathi kuka-286 no-296 eV abonisa ukuthi ukuguqulwa kwamakhemikhali kuqedwe ngempumelelo yi-SAM coating.I-SHP ibonisa iziqongo ezinkulu ze-CF2 (290.1 ​​​​eV) kanye ne-CF3 (293.3 eV), ezibangelwa i-silane esekelwe ku-fluorocarbon eyakhiwe phezulu.Umfanekiso 2E ubonisa izithombe ezimele optical zezilinganiso ze-engeli yokuxhumana (CA) zamaqenjana ahlukene amanzi enziwe i-deionized axhumene ne-repe, eqoshiwe, i-SHP, ne-LOIS.Lezi zithombe zibonisa ukuthi indawo eqoshiwe iba yi-hydrophilic ngenxa yesakhiwo se-micro/nano esakhiwe yi-chemical etching ukuze amanzi akhishwe nge-deionized afakwe esakhiweni.Kodwa-ke, lapho i-substrate ihlanganiswe ne-SAM, i-substrate ibonisa ukuxoshwa kwamanzi okunamandla, ngakho-ke i-SHP engaphezulu iyakhiwa futhi indawo yokuxhumana phakathi kwamanzi nangaphezulu incane.Ekugcineni, ukwehla kwe-CA kwabonwa ku-LOIS, okungabalelwa ekungeneni kwe-lubricant ku-microstructure, ngaleyo ndlela kwandise indawo yokuxhumana.Ukuze kufakazelwe ukuthi indawo engaphezulu inezindawo ezinhle kakhulu zokuxosha uketshezi kanye nezindawo ezinganamatheli, i-LOIS yaqhathaniswa ne-SHP substrate ngokukala i-CA ne-SA kusetshenziswa uketshezi oluhlukahlukene (Umfanekiso 2F).Izinhlobo ezihlukahlukene zoketshezi zakhethwa ngokusekelwe ku-viscosity namandla angaphezulu, okuhlanganisa amanzi angcolile, igazi, i-EG, i-EtOH ne-HD (Umfanekiso S4).Imiphumela yokulinganisa ye-CA ibonisa ukuthi uma i-CA ijwayele ukuba HD, inani lokunciphisa le-CA, lapho i-CA inamandla aphansi kakhulu angaphezulu.Ngaphezu kwalokho, i-LOIS ye-CA iyonke iphansi.Nokho, ukulinganisa kwe-SA kukhombisa into ehluke ngokuphelele.Ngaphandle kwamanzi ane-ionized, lonke uketshezi lunamathela ku-substrate ye-SHP ngaphandle kokushelela.Ngakolunye uhlangothi, i-LOIS ikhombisa i-SA ephansi kakhulu, lapho lapho lonke uketshezi lutshekiswa nge-engeli engaphansi kuka-10° kuya ku-15°, lonke uketshezi luzosuka.Lokhu kukhombisa ngokuqinile ukuthi ukunganamatheli kwe-LOIS kungcono kunobuso be-SHP.Ngaphezu kwalokho, i-LOIS isetshenziselwa izinhlobo ezihlukahlukene zezinto zokwakha, kuhlanganise ne-titanium (Ti), i-polyphenylsulfone (PPSU), i-polyoxymethylene (POM), i-polyether ether ketone (PEEK) kanye nama-polymers e-bioabsorbable (PLGA), Ziyizinto ezisetshenziswayo zamathambo (Figure S5)).Izithombe ezilandelanayo zamaconsi ezintweni eziphathwe yi-LOIS zibonisa ukuthi izakhiwo zokulwa ne-biofouling ze-LOIS ziyafana kuwo wonke ama-substrates.Ngaphezu kwalokho, imiphumela yokulinganisa ye-CA ne-SA ibonisa ukuthi izakhiwo ezinganamatheli ze-LOIS zingasetshenziswa kwezinye izinto.
Ukuze kuqinisekiswe izici zokulwa nokungcoliswa kwe-LOIS, izinhlobo ezihlukahlukene zama-substrates (okuhlanganisa angenalutho, aqoshiwe, i-SHP ne-LOIS) zafakwa i-Pseudomonas aeruginosa ne-MRSA.Lawa mabhaktheriya amabili akhethwe njengama-bacteria asesibhedlela amele, angaholela ekwakhekeni kwama-biofilms, okuholela ku-SSI (37).Umfanekiso wesi-3 (A no-B) ubonisa izithombe zesibonakhulu se-fluorescence kanye nemiphumela yokulinganisa ye-coloni forming unit (CFU) yama-substrates afakwe ekumisweni kwamagciwane isikhathi esifushane (amahora angu-12) kanye nesikhathi eside (amahora angu-72), ngokulandelana.Esikhathini esifushane, amagciwane azokwakha amaqoqo futhi akhule ngosayizi, azimboze ngezinto ezifana namafinyila futhi avimbele ukususwa kwawo.Nokho, phakathi namahora angu-72 ekufukamela, amagciwane azovuthwa futhi kube lula ukuhlakazeka ukuze akhe amakoloni amaningi noma amaqoqo.Ngakho-ke, kungacatshangwa ukuthi ukufukamela amahora angu-72 kuyisikhathi eside futhi isikhathi esifanele sokufukamela sokwenza i-biofilm eqinile ebusweni (38).Esikhathini esifushane, indawo eqoshiwe kanye nengaphezulu le-SHP libonise ukunamathela kwebhaktheriya, okwehliswe cishe ngo-25% kuya ku-50% uma kuqhathaniswa ne-substrate engenalutho.Kodwa-ke, ngenxa yokusebenza kwayo okuhle kakhulu kwe-anti-biofouling nokuzinza, i-LOIS ayizange ibonise ukunamathela kwe-biofilm yebhaktheriya esikhathini esifushane nesikhathi eside.Umdwebo wohlelo (Umfanekiso 3C) uchaza incazelo yendlela yokungcolisa emelene nebhayoloji yesixazululo se-etching, i-SHP ne-LOIS.Ukucatshangwa ukuthi i-substrate ehlanganisiwe enezakhiwo ze-hydrophilic izoba nendawo enkulu kune-substrate engenalutho.Ngakho-ke, ukunamathela okwengeziwe kwebhaktheriya kuzokwenzeka ku-substrate eqoshiwe.Kodwa-ke, uma kuqhathaniswa ne-substrate engenalutho, i-substrate ehlanganisiwe ine-biofilm encane kakhulu eyakhelwe phezulu.Lokhu kungenxa yokuthi ama-molecule amanzi abophezela ngokuqinile endaweni ye-hydrophilic futhi asebenze njengesithambiso samanzi, ngaleyo ndlela aphazamise ukunamathela kwamabhaktheriya esikhathini esifushane (39).Nokho, ungqimba lwama-molecule amanzi luncane kakhulu futhi luncibilika ekumisweni kwamagciwane.Ngakho-ke, ungqimba lwama-molecule wamanzi luyanyamalala isikhathi eside, okuholela ekunamatheleni okubanzi kwebhaktheriya nokwanda.Ku-SHP, ngenxa yezindawo zayo zesikhashana ezingamanzisi, ukunamathela kwebhaktheriya kuvinjelwe.Ukunamathela kwebhaktheriya okuncishisiwe kungabangelwa amaphakethe omoya avaleleke esakhiweni esinezingqimba namandla angaphansi komhlaba, ngaleyo ndlela kuncishiswe ukuthintana phakathi kokumiswa kwebhaktheriya nendawo engaphezulu.Kodwa-ke, ukunamathela okubanzi kwebhaktheriya kwabonwa ku-SHP ngoba yalahlekelwa izakhiwo zayo zokulwa nokungcolisa isikhathi eside.Lokhu ikakhulukazi ngenxa yokunyamalala kwamaphakethe omoya ngenxa yokucindezela kwe-hydrostatic kanye nokuqedwa komoya emanzini.Lokhu kubangelwa ikakhulukazi ukunyamalala kwamaphakethe omoya ngenxa yokuhlakazeka kanye nesakhiwo esinezingqimba esinikeza indawo enkulu yokunamathela (27, 40).Ngokungafani nalawa ma-substrates amabili anomphumela obalulekile ekuzinzeni kwesikhathi eside, i-lubricant lubricant equkethwe ku-LOIS ijovwa esakhiweni se-micro/nano futhi ngeke zinyamalale ngisho nangesikhathi eside.Izinto zokugcoba ezigcwele izakhiwo ezincane / ze-nano zizinzile kakhulu futhi zikhangwa kakhulu ebusweni ngenxa yokuhlangana kwazo kwamakhemikhali aphezulu, ngaleyo ndlela kuvimbele ukunamathela kwamagciwane isikhathi eside.Umfanekiso we-S6 ubonisa isithombe se-confocal microscope esibukwayo se-lubricant-infusted substrate ecwiliswe ku-phosphate buffered saline (PBS).Izithombe eziqhubekayo zibonisa ukuthi ngisho nangemva kwamahora angu-120 okuzamazama kancane (120 rpm), isendlalelo se-lubricant ku-LOIS sihlala singashintshiwe, okubonisa ukuzinza kwesikhathi eside ngaphansi kwezimo zokugeleza.Lokhu kungenxa yokuhlobana kwamakhemikhali okuphezulu phakathi kwe-SAM esekelwe ku-fluorine kanye ne-perfluorocarbon-based lubricant, ukuze kwakhiwe ungqimba oluzinzile lwe-lubricant.Ngakho-ke, ukusebenza kwe-anti-fouling kuyagcinwa.Ngaphezu kwalokho, i-substrate ihlolwe ngokumelene namaprotheni amele (i-albumin ne-fibrinogen), ase-plasma, amangqamuzana ahlobene eduze nokusebenza komzimba (ama-macrophages nama-fibroblasts), nalawo ahlobene nokwakheka kwamathambo.Okuqukethwe kwe-calcium kuphezulu kakhulu.(Umfanekiso 3D, 1 kanye 2, kanye Figure S7) (41, 42).Ngaphezu kwalokho, izithombe ze-fluorescence microscope zokuhlolwa kokunamathela kwe-fibrinogen, i-albumin ne-calcium zibonise izici ezihlukene zokunamathela zeqembu ngalinye le-substrate (Umfanekiso S8).Ngesikhathi sokwakheka kwamathambo, izingqimba ezisanda kwakhiwa zamathambo ne-calcium zingase zizungeze ukufakelwa kwamathambo, okungenzi nje ukususwa kube nzima, kodwa kungase kubangele ukulimala okungalindelekile esigulini ngesikhathi sokususwa.Ngakho-ke, amazinga aphansi e-calcium efakwa kumapuleti amathambo nezikulufu anenzuzo ekuhlinzweni kwamathambo okudinga ukususwa kwezimila.Ngokusekelwe ekubalweni kwendawo enamathiselwe ngokusekelwe kubukhulu be-fluorescence nokubala kwamaseli, siqinisekisile ukuthi i-LOIS ibonisa izici ezinhle kakhulu zokulwa ne-biofouling yazo zonke izinto zebhayoloji uma ziqhathaniswa nezinye izingxenye ezincane.Ngokwemiphumela yokuhlolwa kwe-in vitro, i-anti-biological fouling i-LOIS ingasetshenziswa ekufakweni kwamathambo, okungakwazi ukuvimbela izifo ezibangelwa amagciwane e-biofilm, kodwa futhi kunciphise ukuvuvukala okubangelwa amasosha omzimba asebenzayo.
(A) Izithombe ze-Fluorescence microscope zeqembu ngalinye (zinqunu, ziqoshiwe, i-SHP ne-LOIS) ezifakwe ku-Pseudomonas aeruginosa kanye nokumiswa kwe-MRSA amahora angu-12 nangama-72.(B) Inombolo ye-CFU enamathelayo ye-Pseudomonas aeruginosa ne-MRSA ebusweni beqembu ngalinye.(C) Umdwebo wohlelo wendlela yokungcolisa i-anti-biological yesikhathi esifushane nesikhathi eside, i-SHP ne-LOIS.(D) (1) Inombolo yama-fibroblasts anamathele kumfanekiso we-substrate ngayinye kanye ne-fluorescence microscope yamaseli anamathela kokungenalutho kanye ne-LOIS.(2) Ukuhlolwa kokunamathela kwamaprotheni ahlobene nokuzivikela komzimba, i-albhamuin ne-calcium ehilelekile ekuphulukiseni amathambo (* P <0.05, ** P <0.01, *** P <0.001 kanye **** P <0.0001).ns, akubalulekile.
Endabeni yokucindezeleka okugxilile okungenakugwenywa, ukuqina komshini bekulokhu kuyinselele enkulu ekusetshenzisweni kwezindwangu ezivimbela ukungcola.Izindlela zendabuko zokulwa nokuthuthwa kwendle zisekelwe kuma-polymers ane-solubility ephansi yamanzi kanye ne-fragility.Ngakho-ke, ngokuvamile basengozini yokucindezeleka komshini emisebenzini ye-biomedical.Ngakho-ke, okokunamathela okuqinile okuqinile okuhlala kuyinselele ezisetshenziswayo ezifana nezimila zamathambo (43, 44).Umfanekiso we-4A (1) ubonisa izinhlobo ezimbili eziyinhloko zokucindezeleka ezisetshenziswa ekufakweni kwamathambo, okuhlanganisa ukunwaya (ukucindezeleka kwe-shear) nokucindezela ngesithombe se-optical sokufakelwa owonakele okukhiqizwa yi-forceps.Isibonelo, uma isikulufu siqiniswa nge-screwdriver, noma lapho udokotela ohlinzayo ebambe ipuleti lethambo aliqinise ngama-tweezers futhi esebenzisa amandla acindezelayo, i-plastic bone plate izolimala futhi iklwejwe kuzo zombili izikali ezinkulu ne-micro/nano (Umfanekiso 4A, 2) .Ukuze kuhlolwe ukuthi i-LOIS ekhiqiziwe ingakwazi yini ukumelana nale monakalo ngesikhathi sokuhlinzwa kwepulasitiki, i-nanoindentation yenziwa ukuze kuqhathaniswe ubulukhuni be-substrate engenalutho kanye ne-LOIS esikalini se-micro/nano ukuze kufundwe izakhiwo zemishini ze-micro/nano structure Impact (Umfanekiso 4B).Umdwebo wohlelo ubonisa ukuziphatha okuhlukile kwe-LOIS ngenxa yobukhona bezakhiwo ezincane/nano.Ijika le-force-displacement ladwetshwa ngokusekelwe emiphumeleni ye-nanoindentation (Umfanekiso 4C).Isithombe esiluhlaza okwesibhakabhaka simelela i-substrate engenalutho, ebonisa ukuwohloka okuncane kuphela, njengoba kubonwa ukujula okukhulu kokuhlehlisa okungu-0.26-μm.Ngakolunye uhlangothi, ukwanda kancane kancane kwamandla e-nanoindentation kanye nokufuduka okubonwa ku-LOIS (ijika elibomvu) kungase kubonise izimpawu zezakhiwo ezincishisiwe zemishini, okuholela ekujuleni kwe-nanoindentation kwe-1.61μm.Lokhu kungenxa yokuthi isakhiwo se-micro/nano esikhona ku-LOIS sinikeza isikhala sokuthuthuka esijulile se-tip ye-nanoindenter, ngakho-ke ukuguqulwa kwayo kukhulu kunaleyo ye-substrate engenalutho.U-Konsta-Gdoutos et al.(45) ikholelwa ukuthi ngenxa yokuba khona kwe-nanostructures, i-nanoindentation kanye ne-micro/nano roughness kuholela kumajika e-nanoindentation angajwayelekile.Indawo enomthunzi ihambisana nejika lokuguquguquka elingavamile elibalelwa ku-nanostructure, kuyilapho indawo engasikiwe ibhekwa njenge-microstructure.Lokhu kuguqulwa kungase kulimaze i-microstructure/nanostructure ye-lubricant yokubamba futhi kuthinte kabi ukusebenza kwayo kokulwa nokungcolisa.Ukuze kufundwe umthelela womonakalo ku-LOIS, umonakalo ongenakugwenywa ezakhiweni ezincane/nano waphindwaphindwa emzimbeni ngesikhathi sokuhlinzwa kwepulasitiki.Ngokusebenzisa ukuhlolwa kokunamathela kwegazi namaprotheni, ukuzinza kwezakhiwo ze-anti-biofouling ze-LOIS ngemva kwe-in vitro kunganqunywa (Figure 4D).Uchungechunge lwezithombe ezibonakalayo lubonisa umonakalo owenzekile eduze nezimbobo ze-substrate ngayinye.Ukuhlolwa kokunamathela kwegazi kwenziwa ukuze kuboniswe umphumela wokulimala kwemishini ku-anti-biofouling coating (Umfanekiso 4E).Njenge-SHP, izakhiwo zokulwa nokungcolisa ziyalahleka ngenxa yomonakalo, futhi i-LOIS ibonisa izinto ezinhle kakhulu zokulwa nokungcolisa ngokuxosha igazi.Lokhu kungenxa yokuthi, ngenxa yokuthi amandla angaphezulu aqhutshwa isenzo se-capillary esimboze indawo eyonakele, ukugeleza ku-lubricant ye-microstructured lubricant kubuyisela izakhiwo zokulwa nokungcolisa (35).Umkhuba ofanayo wabonwa ekuhlolweni kokunamathela amaprotheni kusetshenziswa i-albumin.Endaweni eyonakele, ukunamathela kwamaprotheni ebusweni be-SHP kubonakala kakhulu, futhi ngokulinganisa indawo yayo, ingalinganiswa njengengxenye yezinga lokunamathela le-substrate engenalutho.Ngakolunye uhlangothi, i-LOIS yagcina izakhiwo zayo zokulwa ne-biofouling ngaphandle kokubangela ukunamathela (Umfanekiso 4, F kanye no-G).Ngaphezu kwalokho, ingaphezulu lesikulufu livamise ukuba ngaphansi kwengcindezi eqinile yemishini, njengokubhoboza, ngakho-ke sifunde ikhono le-LOIS lokunamathela ukuze lihlale linjalo kusikulufa ku-vitro.Umfanekiso 4H ubonisa izithombe ezibonakalayo zezikulufu ezihlukene, okuhlanganisa okungenalutho, i-SHP ne-LOIS.Unxande obomvu umele indawo eqondiwe lapho kwenzeka khona ukucindezeleka okuqinile komshini ngesikhathi sokufakelwa kwethambo.Ngokufanayo nokuhlolwa kokunamathela kwamaprotheni epuleti, isibonakhulu se-fluorescence sisetshenziselwa ukwenza isithombe sokunamathela kwamaprotheni futhi kukalwe indawo yokumboza ukuze kufakazele ubuqotho be-LOIS yokunamathela, ngisho nangaphansi kokucindezeleka okunamandla kwemishini (Umfanekiso 4, I kanye no-J).Izikulufu ezilashwe nge-LOIS zibonisa ukusebenza okuhle kakhulu kokunqanda ukungcola, futhi cishe awekho amaprotheni anamathela phezulu.Ngakolunye uhlangothi, ukunamathela kwamaprotheni kwabonwa kuzikulufu ezingenalutho kanye nezikulufu ze-SHP, lapho ukumbozwa kwendawo kwezikulufu ze-SHP kwakuyingxenye eyodwa kwezintathu yalezo zezikulufu ezingenalutho.Ukwengeza, ukufakelwa kwamathambo okusetshenziselwa ukulungiswa kumele kube namandla ngomshini ukuze kumelane nokucindezeleka okufakwe endaweni yokuphuka, njengoba kuboniswe kuMfanekiso 4K.Ngakho-ke, ukuhlolwa kokugoba kwenziwa ukuze kutholwe umphumela wokuguqulwa kwamakhemikhali kuzakhiwo zemishini.Ukwengeza, lokhu kwenziwa ukuze kugcinwe ukucindezeleka okungaguquki okuvela ku-implant.Faka amandla emishini eqondile kuze kube yilapho isifakelo sigoqwe ngokugcwele futhi kutholakala ijika le-stress-strain (Umfanekiso 4L, 1).Izakhiwo ezimbili ezihlanganisa i-Young's modulus namandla aguquguqukayo ziqhathaniswe phakathi kwama-substrates angenalutho kanye ne-LOIS njengezinkomba zamandla azo emishini (Umfanekiso 4L, 2 kanye no-3).I-Young's modulus ikhombisa ikhono lempahla ukumelana nezinguquko zemishini.Imoduli Yentsha ye-substrate ngayinye ingu-41.48±1.01 kanye no-40.06±0.96 GPa, ngokulandelana;umehluko oboniwe ucishe ube ngu-3.4%.Ngaphezu kwalokho, kubikwa ukuthi amandla okugoba, anquma ukuqina kwento, yi-102.34±1.51 GPa ye-substrate engenalutho kanye ne-96.99±0.86 GPa ye-SHP.I-substrate engenalutho icishe ibe ngu-5.3% ngaphezulu.Ukuncipha kancane kwezici zokushintshwa kungase kubangelwe umphumela we-notch.Kumphumela we-notch, ubulukhuni be-micro/nano bungase busebenze njengesethi yezinotshi, okuholela ekugxilweni kwengcindezi yasendaweni futhi kuthinte izici zomshini zokufakelwa (46).Kodwa-ke, ngokusekelwe eqinisweni lokuthi ukuqina kwethambo le-cortical yomuntu kubikwa ukuthi kuphakathi kuka-7.4 no-31.6 GPa, futhi i-modulus ye-LOIS elinganiselwe idlula ithambo le-cortical yomuntu (47), i-LOIS yanele ukusekela ukuphuka kanye ne- izakhiwo zemishini zithinteka kancane ngokuguqulwa kwendawo.
(A) Umdwebo wohlelo (1) wokucindezeleka komshini osetshenziswa ekufakweni kwamathambo ngesikhathi sokusebenza, kanye (2) nesithombe sokubonwayo sokufakelwa kwamathambo okonakele.(B) Umdwebo oyisikimu wokulinganisa izakhiwo ze-nano-mechanical nge-nanoindentation kanye ne-LOIS endaweni engenalutho.(C) Ijika le-Nanoindentation force-displacement yendawo engenalutho kanye ne-LOIS.(D) Ngemva kokuhlolwa kwe-in vitro, lingisa izithombe zamehlo zezinhlobo ezihlukene zamapuleti amathambo (indawo eyonakele igqanyiswe ngonxande obomvu) ukuze ulingise ukucindezelwa komshini okubangelwa phakathi nokusebenza.(E) Ukuhlolwa kokunamathela kwegazi kanye (F) nokuhlolwa kokunamathela kwamaprotheni eqembu lamapuleti amathambo alimele.(G) Linganisa ukumbozwa kwendawo kwephrotheni enamathela epuletini.(H) Izithombe zamehlo zezinhlobo ezahlukene zezikulufu zamathambo ngemva kokuhlolwa kwe-in vitro.(I) Ukuhlolwa kokunamathela kwamaprotheni ukuze kufundwe ubuqotho bezingubo ezihlukene.(J) Linganisa ukumbozwa kwendawo kwephrotheni enamathela kusikulufu.(K) Ukunyakaza kukanogwaja kuhloswe ngayo ukukhiqiza ingcindezi engaguquki ethanjeni eliphukile.(L) (1) Gobisa imiphumela yokuhlolwa nemifanekiso yokubona ngaphambi nangemuva kokugoba.Umehluko (2) kumoduli Yentsha kanye (3) namandla okugoba phakathi kokufakelwa okungenalutho kanye ne-SHP.Idatha ichazwa njengesho ukuthi ± SD (*P<0.05, **P<0.01, ***P<0.001 kanye ne-****P<0.0001).Isithombe sihlonishwa: Kyomin Chae, Yonsei University.
Ezimweni zomtholampilo, ukuthintana kwamagciwane okuningi ngezinto zebhayoloji nezindawo zamanxeba kuvela kuma-biofilms avuthiwe, avuthiwe (48).Ngakho-ke, i-US Centers for Disease Control and Prevention ilinganisela ukuthi i-65% yazo zonke izifo zabantu zihlobene nama-biofilms (49).Kulokhu, kuyadingeka ukuhlinzeka ngedizayini yokuhlola ye-vivo enikeza ukwakheka okungaguquki kwe-biofilm ebusweni bokufakelwa.Ngakho-ke, sakha imodeli yokuphuka kwefemu kanogwaja lapho izimila zamathambo zafakwa khona ngaphambili ekumisweni kwebhaktheriya bese zifakwa kumafemu onogwaja ukuze kufundwe izici zokulwa nokungcolisa ze-LOIS ku-vivo.Ngenxa yala maqiniso amathathu abalulekile alandelayo, izifo ezibangelwa amagciwane zibangelwa imikhuba yangaphambi kwesiko kunokujova ngokuqondile ukumiswa kwamagciwane: (i) Amasosha omzimba onogwaja anamandla ngokwemvelo kunawabantu;ngakho-ke, umjovo wokumiswa kwebhaktheriya kanye namabhaktheriya e-planktonic kungenzeka Awunawo umthelela ekwakhiweni kwama-biofilms.(Ii) Amagciwane e-Planktonic angenwa kalula yimithi elwa namagciwane, futhi ama-antibiotic avame ukusetshenziswa ngemva kokuhlinzwa;ekugcineni, (iii) ukumiswa kwamagciwane e-planktonic kungase kuhlanjululwe uketshezi lomzimba wesilwane (50).Ngokuhlakulela kusengaphambili ukufakelwa ekumisweni kwebhaktheriya ngaphambi kokufakwa, singacwaninga kahle imiphumela eyingozi yokutheleleka ngebhaktheriya nokusabela komzimba wangaphandle (FBR) enqubweni yokuphulukisa amathambo.Onogwaja banikelwa emavikini angu-4 ngemva kokufakwa, ngoba i-osseointegration ebalulekile enkambisweni yokuphulukisa amathambo izoqedwa phakathi kwamaviki angu-4.Ngemva kwalokho, ama-implants asusiwe konogwaja ukuze kwenziwe izifundo ezansi nomfula.Umfanekiso 5A ubonisa indlela yokwanda kwamabhaktheriya.Ukufakelwa kwamathambo okunegciwane kungeniswa emzimbeni.Njengomphumela we-pre-incubation ekumisweni kwebhaktheriya, onogwaja abayisithupha kwabayisithupha ababefakwe ngokufakwa benqunu batheleleka, kanti akekho unogwaja owafakelwe izimila ezilashwe yi-LOIS owatheleleka.Ukutheleleka kwamagciwane kuqhubeka ngezinyathelo ezintathu, okuhlanganisa ukukhula, ukuvuthwa kanye nokuhlakazeka (51).Okokuqala, amabhaktheriya anamathiselwe ayazalana futhi akhule phezulu, bese amabhaktheriya enza i-biofilm lapho ekhipha i-extracellular polymer (EPS), i-amyloid ne-extracellular DNA.I-Biofilm ayiphazamisi nje kuphela ukungena kwemithi elwa namagciwane, kodwa futhi ikhuthaza ukuqoqwa kwama-enzyme ehlisa isibulala-magciwane (njenge-β-lactamase) (52).Ekugcineni, i-biofilm isakaza amagciwane avuthiwe ezicutshini ezizungezile.Ngakho-ke, ukutheleleka kwenzeka.Ngaphezu kwalokho, lapho umzimba wangaphandle ungena emzimbeni, ukutheleleka okungabangela ukusabela okunamandla kwamasosha omzimba kungabangela ukuvuvukala okukhulu, ubuhlungu, nokunciphisa ukuzivikela komzimba.Umdwebo 5B unikeza ukubuka konke kwe-FBR okubangelwa ukufakwa kwe-orthopedic implant, kunokuba impendulo yokuzivikela komzimba ebangelwa ukutheleleka kwebhaktheriya.Amasosha omzimba abona ukufakwa okufakiwe njengomzimba wangaphandle, bese kubangela ukuthi amaseli nezicubu ziphendule ukuze zihlanganise umzimba wangaphandle (53).Ezinsukwini zokuqala ze-FBR, i-matrix yokuhlinzeka yakhiwa ebusweni bokufakelwa kwe-orthopedic, okwaholela ekukhangisweni kwe-fibrinogen.I-adsorbed fibrinogen bese yenza inethiwekhi ye-fibrin eminyene kakhulu, ekhuthaza ukunamathiselwa kwe-leukocyte (54).Uma inethiwekhi ye-fibrin yakhiwe, ukuvuvukala okunamandla kuzokwenzeka ngenxa yokungena kwama-neutrophils.Kulesi sinyathelo, izinhlobonhlobo ze-cytokines ezifana ne-tumor necrosis factor-α (TNF-α), i-interleukin-4 (IL-4) ne-IL-β zikhishwa, futhi ama-monocyte aqala ukungena endaweni yokufakelwa futhi ahlukanise amaseli amakhulu.Phage (41, 55, 56).Ukunciphisa i-FBR bekulokhu kuyinselele ngoba i-FBR eyeqile ingabangela ukuvuvukala okunamandla nokungapheli, okungaholela ezinkingeni ezibulalayo.Ukuze kuhlolwe umthelela wezifo ezibangelwa amagciwane ezicutshini ezizungeze ukufakelwa okungenalutho kanye ne-LOIS, kwasetshenziswa i-hematoxylin ne-eosin (H&E) kanye ne-Masson trichrome (MT) staining.Konogwaja abatshalwe ngama-substrates angenalutho, izifo ezinzima zamagciwane zaqhubeka, futhi amaslayidi ezicubu ze-H&E abonisa ngokucacile amathumba kanye ne-necrosis ebangelwa ukuvuvukala.Ngakolunye uhlangothi, indawo eqinile ye-anti-biofouling i-LOIS ivimbela ukunamathela kwamagciwane, ngakho ayibonisi zimpawu zokutheleleka futhi inciphisa ukuvuvukala (Umfanekiso 5C).Imiphumela ye-MT staining ibonise umkhuba ofanayo.Kodwa-ke, i-MT staining iphinde yabonisa i-edema konogwaja abafakwe nge-LOIS, okubonisa ukuthi ukululama kuzokwenzeka (Umfanekiso 5D).Ukuze kufundwe izinga lokuphendula komzimba, ukungcola kwe-immunohistochemical (IHC) kwenziwa kusetshenziswa i-cytokines TNF-α ne-IL-6 ehlobene nokusabela komzimba.Ukufakelwa kwe-negative enqunu okwakungavezwanga amagciwane kwaqhathaniswa ne-LOIS eyayichayeke kubhaktheriya kodwa ingenalo igciwane ukuze itadishe inqubo yokuphulukisa lapho kungekho ukutheleleka kwamagciwane.Umfanekiso 5E ubonisa isithombe sokubonwayo sesilayidi se-IHC esiveza i-TNF-α.Indawo ensundu imele ukuphendula kwamasosha omzimba, okubonisa ukuthi ukusabela komzimba ku-LOIS kuncishisiwe kancane.Ngaphezu kwalokho, inkulumo ye-IL-6 ku-LOIS yayingaphansi kakhulu kunenkulumo engalungile yokunqunu oyinyumba (Umfanekiso 5F).Inkulumo ye-cytokine yalinganiswa ngokulinganisa indawo ye-antibody staining ehambisana ne-cytokine (Umfanekiso 5G).Uma kuqhathaniswa nonogwaja abavezwe ekufakweni okungalungile, amazinga enkulumo onogwaja abafakwe nge-LOIS ayephansi, ebonisa umehluko ozwakalayo.Ukuncipha kwenkulumo ye-cytokine kukhombisa ukuthi izinto zesikhathi eside, ezizinzile zokulwa nokungcoliswa kwe-LOIS azihlobene nje kuphela nokuvinjwa kwezifo ezibangelwa amagciwane, kodwa futhi nokuncipha kwe-FBR, okubangelwa ama-macrophage anamathela ku-substrate (53), 57, 58).Ngakho-ke, ukusabela okuncishisiwe kokuzivikela komzimba ngenxa yezakhiwo zokuzivikela komzimba ze-LOIS kungase kuxazulule imiphumela emibi ngemva kokufakelwa, njengokuphendula ngokweqile komzimba ngemva kokuhlinzwa kwepulasitiki.
(A) Umdwebo oyisikimu wendlela yokwakheka kwe-biofilm nokusabalala endaweni yokufakelwa kwamathambo okuthelelekile.I-eDNA, i-extracellular DNA.(B) Umdwebo wohlelo lwempendulo yokuzivikela komzimba ngemva kokufakwa kwe-orthopedic implant.(C) I-H&E staining kanye (D) ne-MT staining yezicubu ezizungezile zokufakelwa kwe-orthopedic ene-positive engenalutho kanye ne-LOIS.I-IHC yama-cytokines ahlobene nokuzivikela komzimba (E) TNF-α kanye (F) IL-6 yizithombe ezingcolile zonogwaja abanqunu nabangenayo i-LOIS.(G) Ukulinganisa kwe-cytokine expression by area coverage measure (** P <0.01).
I-biocompatibility ye-LOIS kanye nomthelela wayo enqubweni yokuphulukisa amathambo kwahlolwa ku-vivo kusetshenziswa i-diagnostic imaging [i-x-ray ne-micro-computed tomography (CT)] kanye ne-osteoclast IHC.Umfanekiso we-6A ubonisa inqubo yokuphulukisa amathambo ehlanganisa izigaba ezintathu ezihlukene: ukuvuvukala, ukulungisa, nokulungisa kabusha.Uma kwenzeka ukuphuka, amangqamuzana okuvuvukala kanye nama-fibroblasts azongena ethanjeni eliphukile futhi aqale ukukhula abe yizicubu ze-vascular.Phakathi nesigaba sokulungisa, i-ingrowth yezicubu ze-vascular isakazeka eduze nendawo yokuphuka.Izicubu ze-Vascular zinikeza izakhi zokwakhiwa kwethambo elisha, elibizwa ngokuthi i-callus.Isigaba sokugcina senqubo yokuphulukisa amathambo yisigaba sokulungiswa kabusha, lapho ubukhulu be-callus buncishiswa ubukhulu bethambo elivamile ngosizo lokwanda kwezinga lama-osteoclasts acushiwe (59).Ukwakhiwa kabusha kwe-Three-dimensional (3D) yendawo yokuphuka kwenziwa kusetshenziswa ama-micro-CT scans ukuze kubonwe umehluko wezinga lokubunjwa kwe-callus eqenjini ngalinye.Bheka isigaba esiphambanayo se-femur ukuze ubone ukushuba kocingo oluzungeze ithambo eliphukile (Umfanekiso 6, B no-C).Ama-X-ray nawo asetshenziselwa ukuhlola izindawo zokuphuka zawo wonke amaqembu isonto ngalinye ukuze kugcinwe izinqubo ezihlukene zokuvuselela amathambo eqenjini ngalinye (Umfanekiso S9).I-Callus namathambo avuthiwe aboniswa eluhlaza okwesibhakabhaka / eluhlaza kanye nezinyo lendlovu, ngokulandelana.Izicubu eziningi ezithambile zihlungwa ngomkhawulo omisiwe.I-Nude positive kanye ne-SHP iqinisekise ukwakheka kwenani elincane le-callus endaweni yokuphuka.Ngakolunye uhlangothi, i-negative eveziwe ye-LOIS kanye nesayithi lokuphuka lizungezwe i-callus ewugqinsi.Izithombe ze-Micro-CT zibonise ukuthi ukwakheka kwe-callus kwavinjelwa ukutheleleka kwebhaktheriya nokuvuvukala okuhlobene nokutheleleka.Lokhu kungenxa yokuthi amasosha omzimba abeka phambili ukuphulukiswa kokulimala kwe-septic okubangelwa ukuvuvukala okuhlobene nokutheleleka, kunokululama kwamathambo (60).I-IHC kanye ne-Tartrate-resistant Acid Phosphatase (TRAP) staining yenziwa ukuze kubhekwe umsebenzi we-osteoclast kanye ne-bone resorption (Figure 6D) (61).Ama-osteoclast ambalwa kuphela acushiwe afakwe onsomi atholwe ezintweni ezinhle ezinqunu kanye ne-SHP.Ngakolunye uhlangothi, ama-osteoclast amaningi acushiwe abonwa eduze kwamathambo anqunu navuthiwe e-LOIS.Lesi senzakalo sibonisa ukuthi phambi kwama-osteoclasts, i-callus ezungeze indawo yokuphuka ibhekene nenqubo yokuvuselela enobudlova (62).Ivolumu yethambo nendawo ye-osteoclast ye-osteoclast ye-callus ikalwa ukuze kuqhathaniswe izinga lokwakheka kwe-callus ezungeze indawo yokuphuka kuwo wonke amaqembu, ukuze kulinganiswe imiphumela ye-micro-CT scan kanye ne-IHC (Umfanekiso 6E, 1 kanye no-2).Njengoba kulindelekile, ama-negative anqunu kanye nokwakheka kwe-callus ku-LOIS kwakuphakeme kakhulu kunamanye amaqembu, okubonisa ukuthi ukuguqulwa kwamathambo okuhle kwenzeka (63).Umfanekiso S10 ubonisa isithombe sokubonwayo sesayithi lokuhlinza, umphumela obala we-MT wethishu eqoqwe eduze kwesikulufu, kanye nomphumela wamabala we-TRAP ogqamisa isixhumi esibonakalayo se-screw-bone.Ku-substrate engenalutho, ukwakheka kwe-callus eqinile kanye ne-fibrosis kwabonwa, kuyilapho ukufakelwa kwe-LOIS-treatment kubonisa indawo engenakulinganiswa.Ngokufanayo, uma kuqhathaniswa nezinto ezingezinhle ezinqunu, i-fibrosis ephansi yabonwa onogwaja abafakwe nge-LOIS, njengoba kuboniswa imicibisholo emhlophe.Ukwengeza, i-edema eqinile (umcibisholo oluhlaza okwesibhakabhaka) ingabangelwa izakhiwo zokubalekela i-LOIS, ngaleyo ndlela yehlise ukuvuvukala okukhulu.I-non-stick surface ezungeze i-implant kanye ne-fibrosis encishisiwe iphakamisa ukuthi inqubo yokususa ilula, okuvame ukuholela kwezinye iziphuka noma ukuvuvukala.Inqubo yokuphola kwethambo ngemva kokukhishwa kwesikulufu yahlolwa umsebenzi we-osteoclast kusixhumi esibonakalayo se-screw-bone.Kokubili ithambo elingenalutho kanye nesixhumi esibonakalayo sokufakelwa kwe-LOIS kwamunca amazinga afanayo ama-osteoclasts ukuze kuqhutshekwe nokuphulukiswa kwamathambo, okubonisa ukuthi ukunamathela kwe-LOIS akunawo umthelela omubi ekwelapheni kwamathambo noma ekuphenduleni kwamasosha omzimba.Ukuze kuqinisekiswe ukuthi ukuguqulwa kwendawo okwenziwa ku-LOIS akuphazamisi inqubo yokuphulukisa amathambo, ukuhlolwa kwe-X-ray kwasetshenziswa ukuze kuqhathaniswe ukuphulukiswa kwamathambo onogwaja nama-ion angalungile avelayo kanye namaviki angu-6 okufakelwa kwe-LOIS (Umfanekiso 6F).Imiphumela yabonisa ukuthi uma kuqhathaniswa neqembu elinqunu elingenalo igciwane, i-LOIS ibonise izinga elifanayo lokuphulukiswa kwamathambo, futhi kwakungekho zimpawu ezicacile zokuphuka (umugqa oqhubekayo we-osteolysis) kuwo womabili amaqembu.
(A) Umdwebo wohlelo lwenqubo yokuphulukisa amathambo ngemva kokuphuka.(B) Umehluko kwizinga lokubunjwa kwekholi leqembu ngalinye elingaphezulu kanye (C) nesithombe esiphambanayo sesayithi lokuphuka.(D) I-TRAP staining ukuze ubone ngeso lengqondo umsebenzi we-osteoclast kanye ne-bone resorption.Ngokusekelwe kumsebenzi we-TRAP, ukwakheka kwe-callus yangaphandle yethambo le-cortical kwahlaziywa ngobuningi yi-(E) (1) micro-CT kanye (2) nomsebenzi we-osteoclast.(F) Emavikini angu-6 ngemva kokufakelwa, izithombe ze-X-ray zethambo eliphukile lenegethivu eveziwe (eqokonyiswe unxande odayishiwe obomvu) kanye ne-LOIS (ezigqanyiswe unxande odayishiwe oluhlaza okwesibhakabhaka).Ukuhlaziywa kwezibalo kwenziwe ngokuhlaziywa kwendlela eyodwa yokuhluka (ANOVA).*P <0.05.** P <0.01.
Ngamafuphi, i-LOIS inikeza uhlobo olusha lwesu lokutheleleka nge-antibacterial kanye ne-immune yokuphunyuka kokufakwa kwamathambo.Izimila ezivamile zamathambo ezine-SHP functionalization zibonisa izici zesikhashana ezinqanda i-biofouling, kodwa azikwazi ukugcina izakhiwo zazo isikhathi eside.I-superhydrophobicity ye-substrate ibamba amabhamuza omoya phakathi kwebhaktheriya ne-substrate, ngaleyo ndlela yakhe amaphakethe omoya, ngaleyo ndlela ivimbele ukutheleleka kwamagciwane.Nokho, ngenxa yokusatshalaliswa komoya, la maphakethe omoya asuswa kalula.Ngakolunye uhlangothi, i-LOIS ifakazele kahle ikhono layo lokuvimbela izifo ezihlobene ne-biofilm.Ngakho-ke, ngenxa yezakhiwo zokulwa nokwenqatshwa kwesendlalelo se-lubricant esifakwe endaweni yesakhiwo se-micro/nano egqinsiwe, ukuvuvukala okuhlobene nokutheleleka kungavinjelwa.Izindlela ezihlukahlukene zokuhlukanisa izinhlamvu ezihlanganisa i-SEM, i-AFM, i-XPS ne-CA izilinganiso zisetshenziselwa ukuthuthukisa izimo zokukhiqiza ze-LOIS.Ngaphezu kwalokho, i-LOIS ingasetshenziswa ezintweni ezihlukahlukene zebhayoloji ezivame ukusetshenziswa emishinini yokulungisa amathambo, njenge-PLGA, Ti, PE, POM kanye ne-PPSU.Khona-ke, i-LOIS yahlolwa ku-vitro ukuze ifakazele izakhiwo zayo zokulwa ne-biofouling ngokumelene namabhaktheriya nezinto eziphilayo ezihlobene nokusabela komzimba.Imiphumela ikhombisa ukuthi inemiphumela emihle kakhulu ye-antibacterial kanye ne-anti-biofouling uma iqhathaniswa nokufakwa okungenalutho.Ngaphezu kwalokho, i-LOIS ibonisa amandla emishini ngisho nangemva kokusebenzisa ukucindezeleka kwemishini, okungenakugwemeka ekuhlinzeni kwepulasitiki.Ngenxa yezakhiwo zokuziphilisa ze-lubricant ebusweni besakhiwo se-micro/nano, i-LOIS igcine ngempumelelo izakhiwo zayo zokungcolisa i-anti-biological.Ukuze kufundwe i-biocompatibility kanye nezakhiwo zokulwa namagciwane ze-LOIS ku-vivo, i-LOIS yafakwa ku-rabbit femur amaviki angu-4.Akukho ukutheleleka ngebhaktheriya okubonwe konogwaja abafakwe i-LOIS.Ngaphezu kwalokho, ukusetshenziswa kwe-IHC kubonise izinga elincishisiwe lokuphendula kwamasosha omzimba wendawo, okubonisa ukuthi i-LOIS ayivimbeli inqubo yokuphulukisa amathambo.I-LOIS ibonisa izinto ezinhle kakhulu zokulwa namagciwane kanye nokuvinjelwa kokuzivikela komzimba, futhi ifakazelwe ukuthi ivimbela ngempumelelo ukwakheka kwe-biofilm ngaphambi nangesikhathi sokuhlinzwa kwamathambo, ikakhulukazi ukuhlanganiswa kwamathambo.Ngokusebenzisa imodeli yokuvuvukala komnkantsha wethambo logwaja, umthelela wezifo ezihlobene ne-biofilm enqubweni yokuphulukisa amathambo ebangelwa ukufakwa kwangaphambili okufakwe ngaphambili kwacwaningwa ngokujulile.Njengocwaningo lwesikhathi esizayo, imodeli entsha ye-vivo iyadingeka ukuze kufundwe izifo ezingase zibe khona ngemva kokufakwa ukuze kuqondwe ngokugcwele futhi kuvinjwe izifo ezihlobene ne-biofilm phakathi nayo yonke inqubo yokuphulukisa.Ukwengeza, i-osteoinduction iseyinselele engaxazululiwe ekuhlanganisweni ne-LOIS.Ucwaningo olwengeziwe luyadingeka ukuze kuhlanganiswe ukunamathela okukhethiwe kwamaseli e-osteoinductive noma umuthi ovuselelayo ne-LOIS ukuze unqobe inselele.Sekukonke, i-LOIS imele ukufakwa okuthembisayo kwamathambo okunokuqina kwemishini kanye nezakhiwo ezinhle kakhulu zokulwa ne-biofouling, ezinganciphisa i-SSI kanye nemiphumela emibi yokuzivikela komzimba.
Geza i-substrate engu-15mm x 15mm x 1mm 304 SS (Dong Kang M-Tech Co., Korea) nge-acetone, i-EtOH kanye namanzi e-DI imizuzu engu-15 ukuze ususe ukungcola.Ukuze kwakheke ukwakheka kwezinga le-micro/nano-level ngaphezulu, i-substrate ehlanzekile icwiliswa kusixazululo se-HF esingu-48% kuya ku-51% (DUKSAN Corp., South Korea) ku-50°C.Isikhathi sokufaka sihluka kusuka kumaminithi angu-0 kuye kwangu-60.Khona-ke, i-substrate ehlanganisiwe yahlanzwa ngamanzi e-deionized futhi yafakwa ku-65% yesisombululo se-HNO3 (Korea DUKSAN Corp.) ku-50 ° C imizuzu engu-30 ukuze kwakhe ungqimba lwe-chromium oxide passivation ebusweni.Ngemuva kokudlula, i-substrate igezwa ngamanzi angcolile futhi yomiswe ukuze ithole i-substrate enesakhiwo esinezingqimba.Okulandelayo, i-substrate yavezwa ku-oxygen plasma (100 W, 3 amaminithi), futhi yacwiliswa ngokushesha kusixazululo esingu-8.88 mM POTS (Sigma-Aldrich, Germany) ku-toluene ekamelweni lokushisa amahora angu-12.Khona-ke, i-substrate eboshwe nge-POTS yahlanzwa nge-EtOH, futhi yafakwa ku-150 ° C amahora angu-2 ukuze kutholwe i-POTS SAM eminyene.Ngemuva kokugcoba kwe-SAM, ungqimba lokuthambisa lwakhiwe ku-substrate ngokusebenzisa i-lubricant ye-perfluoropolyether (Krytox 101; DuPont, USA) enomthamo wokulayisha we-20 μm / cm 2. Ngaphambi kokusetshenziswa, hlunga i-lubricant ngesihlungi se-micron esingu-0.2.Susa i-lubricant eyeqile ngokutshekisa nge-engeli engu-45° imizuzu engu-15.Inqubo efanayo yokukhiqiza yasetshenziselwa izimila zamathambo ezenziwe nge-304 SS (ipuleti lokukhiya nesikulufa sokukhiya esikhombe; i-Dong Kang M-Tech Co., Korea).Wonke ama-orthopedic implants aklanyelwe ukulingana nejometri ye-rabbit femur.
I-morphology yangaphezulu ye-substrate kanye nezimila ze-orthopedic yahlolwa yi-SEM ephumayo yasendle (Hlola i-F50, FEI, USA) kanye ne-AFM (XE-100, Park Systems, South Korea).Ubukhulu obungaphezulu (Ra, Rq) bulinganiswa ngokuphindaphinda indawo engu-20 μm ngo-20 μm (n=4).Isistimu ye-XPS (PHI 5000 VersaProbe, ULVAC PHI, Japan) efakwe umthombo we-X-ray we-Al Kα enosayizi wamabala ongu-100μm2 yasetshenziselwa ukuhlaziya ukwakheka kwamakhemikhali angaphezulu.Isistimu yokulinganisa ye-CA efakwe ikhamera yokuthwebula isithombe eguqukayo (SmartDrop, FEMTOBIOMED, ​​​​South Korea) yasetshenziswa ukukala uketshezi lwe-CA ne-SA.Esilinganisweni ngasinye, 6 kuya ku-10 μl wamaconsi (amanzi akhishiwe, igazi lehhashi, EG, 30% ethanol, kanye ne-HD) abekwa phezulu ukuze kukalwe i-CA.Lapho i-angle yokuthambekela ye-substrate ikhuphuka ngesivinini esingu-2 °/s (n = 4), i-SA ilinganiswa lapho i-droplet iwa.
I-Pseudomonas aeruginosa [Iqoqo Le-American Type Culture (ATCC) 27853] ne-MRSA (ATCC 25923) zathengwa kwa-ATCC (eManassas, eVirginia, eU.SA), futhi isiko lesitoko lagcinwa ku -80°C.Ngaphambi kokusetshenziswa, isiko eliqandisiwe lalifukanyelwa kumhluzi kabhontshisi wesoya we-trypsin-thawed (i-Komed, Korea) ku-37°C amahora angu-18 bese idluliswa kabili ukuze isebenze.Ngemva kokufukamela, isiko laligxiliswe ku-10,000 rpm imizuzu engu-10 ku-4 ° C futhi ligezwe kabili ngesisombululo se-PBS (pH 7.3).Isiko le-centrifuged libe selifakwa ngaphansi kwe-blood agar plates (BAP).I-MRSA kanye ne-Pseudomonas aeruginosa zilungiswe ubusuku bonke futhi zakhuliswa kumhluzi we-Luria-Bertani.Ukugxiliswa kwe-Pseudomonas aeruginosa ne-MRSA ku-inoculum kwanqunywa ngokwesilinganiso yi-CFU yokumiswa ekuhlanjululweni kwe-serial ku-agar.Bese, lungisa ukugxila kwebhaktheriya ku-0.5 McFarland ejwayelekile, okulingana no-108 CFU/ml.Bese unciphisa ukumiswa kwebhaktheriya okusebenzayo izikhathi eziyi-100 kuya ku-106 CFU/ml.Ukuhlola izici zokunamathela kwe-antibacterial, i-substrate yafakwa inzalo ku-121 ° C imizuzu engu-15 ngaphambi kokusetshenziswa.I-substrate yabe idluliselwa ku-25 ml yokumiswa kwebhaktheriya futhi yafakwa ku-37 ° C ngokuqhaqhazela okunamandla (200 rpm) amahora angu-12 nangama-72.Ngemva kokufukamela, i-substrate ngayinye yakhishwa ku-incubator futhi yagezwa izikhathi ezi-3 nge-PBS ukuze kukhishwe noma imaphi amagciwane antantayo phezulu.Ukuze ubuke i-biofilm ku-substrate, i-biofilm yayihlanganiswe ne-methanol futhi ingcoliswe nge-1 ml ye-crimidine orange imizuzu engu-2.Kwabe sekusetshenziswa isibonakhulu se-fluorescence (BX51TR, Olympus, Japan) ukuze kuthathwe izithombe ze-biofilm enamabala.Ukuze kulinganiswe i-biofilm ku-substrate, amaseli anamathiselwe ahlukaniswa ne-substrate ngendlela ye-bead vortex, eyayibhekwa njengendlela efanelekile kakhulu yokukhipha amabhaktheriya anamathiselwe (n = 4).Usebenzisa ama-forcep oyinyumba, susa i-substrate endaweni yokukhula bese ucindezela ipuleti lomthombo ukuze ususe uketshezi oluningi.Amaseli anamathiselwe ngokuxekethile akhishwe ngokuwashwa kabili nge-PBS eyinyumba.I-substrate ngayinye yabe idluliselwa eshubhuni lokuhlola eliyinyumba eliqukethe u-9 ml we-0.1% protein ept saline (PSW) kanye no-2 g wobuhlalu bengilazi obuyinyumba obungu-20 kuya ku-25 (ububanzi obungu-0.4 kuya ku-0.5 mm).Yabe isivalwa amaminithi angu-3 ukuze kukhishwe amaseli kusampula.Ngemuva kokuvuthwa, ukumiswa kwahlanjululwa ngokuphindwe ka-10 ngo-0.1% we-PSW, bese kuthi u-0.1 ml we-dilution ngayinye ifakwe ku-BAP.Ngemva kwamahora angu-24 ekufukamela ku-37°C, i-CFU yabalwa mathupha.
Kumaseli, igundane fibroblasts NIH/3T3 (CRL-1658; American ATCC) kanye negundane macrophages RAW 264.7 (TIB-71; American ATCC) asetshenziswa.Sebenzisa u-Dulbecco's modified Eagle medium (DMEM; LM001-05, Welgene, Korea) ukuze wenze ama-fibroblasts egundane amasiko futhi wengeze nge-10% ye-calf serum (S103-01, Welgene) kanye no-1% penicillin-streptomycin (PS ; LS202-02, Welgene (Welgene) Sebenzisa i-DMEM kuma-macrophage egundane amasiko, ahlanganiswe ne-fetus serum (S001-01, Welgene) kanye no-1% PS Beka i-substrate epuleti le-cell culture elinemithombo eyisithupha, Futhi ugobe amaseli kumaseli angu-105/cm2. Amaseli afakwe ngobusuku ku-37 ° C kanye no-5% we-CO2 Ukuze kufakwe i-cell staining, amaseli ahlanganiswe ne-4% ye-paraformaldehyde imizuzu engu-20 futhi afakwe ku-0.5% ye-Triton X Incubate imizuzu engu-5 ku-100 ku-37°C imizuzu engu-30 Ngemva kwenqubo yokufukamela, sebenzisa i-substrate ene-4′,6-diamino-2-phenylindole (H -1200, Vector Laboratories, UK) i-VECTASHIELD fixation medium (n = 4 ngeseli). , i-fluorescein, i-fluorescein isothiocyanate-albumin (A9771, Sigma-Aldrich, Germany) kanye ne-plasma yomuntu I-Alexa Fluor 488-conjugated fibrinogen (F13191, Invitrogen, USA) yachithwa ku-PBS (10 mM, pH 7.4).Ukugxila kwe-albumin ne-fibrinogen kwakuyi-1 ne-150 μg/ml, ngokulandelana.Ngemuva kwe-substrate Ngaphambi kokucwiliswa kwikhambi le-protein, lihlanza nge-PBS ukuze libuyisele amanzi ebusweni.Bese ucwilisa wonke ama-substrates epuleti elinemithombo eyisithupha eliqukethe isisombululo seprotheni bese ufukamela ku-37°C imizuzu engama-30 kanye nengama-90.Ngemva kokufukamela, I-substrate yabe isikhishwa esixazululweni samaprotheni, igezwe ngobumnene nge-PBS izikhathi ezingu-3, ​​futhi ilungiswe nge-4% ye-paraformaldehyde (n = 4 yeprotheni ngayinye).Okwe-calcium, i-sodium chloride (0.21 M) ne-potassium phosphate (3.77 mM) ) Yancibilika emanzini angcolile.I-pH yesisombululo yalungiswa yaba ngu-2.0 ngokwengeza isixazululo se-hydrochloride (1M).Kwabe sekuthi i-calcium chloride (5.62 mM) incibilike esixazululo.Ngokungeza i-1M tris(hydroxymethyl)-amino I-Methane ilungisa i-pH yesixazululo ibe ngu-7.4.Gxilisa wonke ama-substrates epuleti elinemithombo eyisithupha eligcwele isixazululo se-calcium phosphate engu-1.5× bese ususa esixazululweni ngemva kwemizuzu engama-30.Ukuze wenze amabala, 2 g Alizarin Red S (CI 58005) Hlanganisa no-100 ml wamanzi akhishwe.Bese, sebenzisa u-10% we-ammonium hydroxide ukuze ulungise i-pH ibe ngu-4. Daya i-substrate nge-Alizarin Red solution imizuzu engu-5, bese uxubha udayi oweqile bese uvala.Ngemuva kwenqubo yokugubha, susa i-substrate.Izinto ezibonakalayo ziphelelwe amanzi, bese zicwiliswa ku-acetone imizuzu engu-5, bese zicwiliswa kusisombululo se-acetone-xylene (1: 1) imizuzu engu-5, futhi ekugcineni zigezwe nge-xylene (n = 4).Kusetshenziswa isibonakhulu se-fluorescence (i-Axio Imager) enamalensi angu-×10 kanye no-×20..I-A2m, i-Zeiss, iJalimane) izithombe zonke izingxenye ezingaphansi.I-ImageJ/FIJI (https://imagej.nih.gov/ij/) isetshenziselwe ukulinganisa idatha yokunamathela yezinto eziphilayo eqenjini ngalinye lezindawo ezine ezihlukene zokucabanga.Guqula zonke izithombe zibe izithombe kanambambili ezinemibundu engaguquki ukuze kuqhathaniswe ne-substrate.
I-Zeiss LSM 700 confocal microscope yasetshenziswa ukuqapha ukuzinza kwesendlalelo sokuthambisa ku-PBS kumodi yokubonisa.Isampula yengilazi ehlanganiswe ne-SAM esekelwe ku-fluorine enesendlalelo sokuthambisa esijovwe yacwiliswa kusixazululo se-PBS, futhi yahlolwa kusetshenziswa i-orbital shaker (SHO-1D; Daihan Scientific, South Korea) ngaphansi kwezimo zokuzamazama okuncane (120 rpm).Bese uthatha isampula bese uqapha ukulahleka kokugcoba ngokulinganisa ukulahleka kokukhanya okubonisiwe.Ukuze uthole izithombe ze-fluorescence kumodi yokubonisa, isampula sivezwa kulaser engu-633 nm bese iqoqwa, ngoba ukukhanya kuzovela kusampula.Amasampula akalwe ngezikhawu zesikhathi ezingu-0, 30, 60, namahora angu-120.
Ukuze kutholwe umthelela wenqubo yokuguqulwa kwendawo ezimpahleni ze-nanomechanical zokufakelwa kwamathambo, i-nanoindenter (TI 950 TriboIndenter, Hysitron, USA) efakwe ithiphu ledayimane le-Berkovich elinezinhlangothi ezintathu elimise okwephiramidi lasetshenziswa ukukala i-nanoindenedione.Umthwalo ophakeme ngu-10 mN futhi indawo ingu-100μmx 100μm.Kuzo zonke izilinganiso, isikhathi sokulayisha nesokukhipha singamasekhondi ayi-10, futhi isikhathi sokubamba ngaphansi komthwalo omkhulu wokuhlehlisa singamasekhondi ama-2.Thatha izilinganiso ezindaweni ezinhlanu ezahlukene bese uthatha isilinganiso.Ukuze kuhlolwe ukusebenza kwamandla okusebenza ngaphansi komthwalo, ukuhlolwa kokugoba kwamaphuzu amathathu okuphambanayo kwenziwa kusetshenziswa umshini wokuhlola wendawo yonke (Instron 5966, Instron, USA).I-substrate icindezelwe ngesilinganiso esingaguquki esingu-10 N/s ngomthwalo owandayo.Uhlelo lwesoftware lweBluehill Universal (n = 3) lusetshenziswe ukubala imodulus eguquguqukayo kanye nengcindezi enkulu yokucindezela.
Ukuze kulingise inqubo yokusebenza kanye nokulimala komshini okuhlobene okudalwe phakathi nokusebenza, inqubo yokusebenza yenziwa nge-vitro.Ama-femur aqoqwa konogwaja abamhlophe baseNew Zealand ababulawa.I-femur yahlanzwa futhi yalungiswa ku-4% ye-paraformaldehyde isonto elingu-1.Njengoba kuchazwe endleleni yokuhlola isilwane, i-femur engaguquki yahlinzwa.Ngemuva kokuhlinzwa, ukufakelwa kwe-orthopedic kwacwiliswa egazini (igazi lehhashi, i-KISAN, Korea) nge-10 s ukuze kuqinisekiswe ukuthi ukunamathela kwegazi kwenzeka ngemva kokulimala komshini (n = 3).
Isamba sabesilisa abangama-24 onogwaja abamhlophe base-New Zealand (isisindo esingu-3.0 kuye ku-3.5kg, isilinganiso sobudala bezinyanga eziyisi-6) bahlukaniswa ngokungahleliwe baba ngamaqembu amane: abanqunu abanqunu, abanqunu, i-SHP kanye ne-LOIS.Zonke izinqubo ezibandakanya izilwane zenziwa ngokuhambisana nezindinganiso zokuziphatha zeKomidi Lokunakekelwa Kwezilwane Lesikhungo Sesikhungo (I-IACUC igunyaziwe, i-KOREA-2017-0159).I-orthopedic implant iqukethe ipuleti lokukhiya elinezimbobo ezinhlanu (ubude obungu-41 mm, ububanzi obungu-7 mm nobukhulu obungu-2 mm) kanye nezikulufu zokuvala i-cortical (ubude obungu-12 mm, ububanzi obungu-2.7 mm) ukuze kulungiswe ukuphuka.Ngaphandle kwalawo mapuleti nezikulufu ezisetshenziswa eqenjini elingenalutho-negative, wonke amapuleti nezikulufu zafakwa ku-MRSA ukumiswa (106 CFU/ml) amahora angu-12.Iqembu elinqunu-negative (n=6) laphathwa ngezifakelo zangaphandle ezinqunu ngaphandle kokuchayeka ekumisweni kwebhaktheriya, njengokulawula okungekuhle kokutheleleka.Iqembu elingenalutho (n = 6) laphathwa ngokufakelwa kwendawo engenalutho evezwe kubhaktheriya njengokulawula okuhle kokutheleleka.Iqembu le-SHP (n = 6) lelashwe ngezimila ze-SHP ezivezwe ngamagciwane.Ekugcineni, iqembu le-LOIS laphathwa ngokufakwa kwe-LOIS okuvezwe ngamagciwane (n = 6).Zonke izilwane zigcinwa ekhejini, futhi kunikezwa ukudla okuningi namanzi.Ngaphambi kokuhlinzwa, onogwaja babezila amahora angu-12.Izilwane zaxoshwa nge-anesthetized ngomjovo we-intramuscular we-xylazine (5mg / kg) kanye nomjovo we-intravenous we-paclitaxel (3mg / kg) wokungeniswa.Ngemva kwalokho, hambisa i-2% isoflurane kanye ne-50% kuya ku-70% umoya-mpilo wezokwelapha (izinga lokugeleza 2 L/min) ngohlelo lokuphefumula ukuze kugcinwe i-anesthesia.Ifakwe ngendlela eqondile ku-lateral femur.Ngemuva kokususwa kwezinwele kanye nokukhishwa kwe-povidone-iodine esikhumbeni, ukusika okungaba ngu-6 cm ubude kwenziwa ngaphandle kwefemur emaphakathi kwesokunxele.Ngokuvula igebe phakathi kwemisipha ehlanganisa i-femur, i-femur ivezwa ngokugcwele.Beka ipuleti ngaphambi kwe-femoral shaft bese uyilungisa ngezikulufo ezine.Ngemuva kokulungiswa, sebenzisa i-saw blade (ubukhulu obuyi-1 mm) ukuze udale ukuphuka ngokuzenzakalelayo endaweni ephakathi kwembobo yesibili kanye nomgodi wesine.Ekupheleni kokuhlinzwa, isilonda sasigezwa nge-saline futhi sivalwe ngemishudo.Unogwaja ngamunye wajovwa ngaphansi kwesikhumba nge-enrofloxacin (5 mg/kg) exutshwe ingxenye eyodwa kwezintathu ku-saline.I-X-ray ye-postoperative ye-femur yathathwa kuzo zonke izilwane (0, 7, 14, 21, 28, kanye nezinsuku ezingu-42) ukuze kuqinisekiswe i-osteotomy yethambo.Ngemuva kokubulawa izinzwa ezijulile, zonke izilwane zabulawa ngomthambo i-KCl (2 mmol/kg) ezinsukwini ezingama-28 nezingu-42.Ngemva kokubulawa, i-femur iskenwa yi-micro-CT ukuze ibone futhi iqhathanise inqubo yokuphulukisa amathambo kanye nokwakheka kwamathambo amasha phakathi kwamaqembu amane.
Ngemva kokubulawa, izicubu ezithambile ezazithintana ngokuqondile nezitho zamathambo zaqoqwa.Ithishu yalungiswa ku-10% ye-neutral buffered formalin ngobusuku obubodwa bese iphelelwa amanzi ku-EtOH.Izicubu eziphelelwe amanzi zafakwa kupharafini futhi zahlukaniswa ngogqinsi lwama-40 μm kusetshenziswa i-microtome (400CS; EXAKT, Germany).Ukuze ubone ngeso lengqondo ukutheleleka, ukugcotshwa kwe-H&E kanye ne-MT staining kwenziwa.Ukuze uhlole impendulo yomsingathi, izicubu ezihlukaniswe ngezigaba zafakwa anti-TNF-α primary antibody (AB6671, Abcam, USA) kanye ne-rabbit anti-IL-6 (AB6672; Abcam, USA), bese zelashwa nge-horseradish.I-oxidase.Faka isistimu yokwenza amabala ye-avidin-biotin complex (ABC) ezigabeni ngokuya ngemiyalelo yomkhiqizi.Ukuze kubonakale njengomkhiqizo wokusabela onsundu, i-3,3-diaminobenzidine isetshenziswe kuzo zonke izingxenye.Isithwebuli samaslayidi sedijithali (i-Pannoramic 250 Flash III, 3DHISTECH, Hungary) sasetshenziswa ukuze kubonakale zonke izingcezu ngeso lengqondo, futhi okungenani ama-substrates amane eqenjini ngalinye ahlaziywa isofthiwe ye-ImageJ.
Izithombe ze-X-ray zathathwa kuzo zonke izilwane ngemva kokuhlinzwa futhi masonto onke ukuze kuqashwe ukuphulukiswa kokuphuka (n=6 iqembu ngalinye).Ngemva kokubulawa, i-high-resolution micro-CT yasetshenziselwa ukubala ukwakheka kwe-callus ezungeze i-femur ngemva kokuphulukiswa.I-femur etholakele yahlanzwa, yafakwa ku-4% ye-paraformaldehyde izinsuku ezingu-3, ​​futhi yaphelelwa amanzi ngo-75% we-ethanol.Amathambo aphelelwe amanzi abe esethwetshulwa kusetshenziswa i-micro-CT (SkyScan 1173, Brooke Micro-CT, Kandy, Belgium) ukuze kukhiqizwe izithombe ze-voxel ze-3D (amaphikseli angu-2240×2240) wesampula yethambo.Sebenzisa isihlungi esingu-1.0 mm Al ukuze unciphise umsindo wesignali futhi usebenzise ukulungiswa okuphezulu kuzo zonke izikena (E = 133 kVp, I = 60 μA, isikhathi sokuhlanganisa = 500 ms).Isofthiwe ye-Nrecon (inguqulo engu-1.6.9.8, i-Bruker microCT, i-Kontich, e-Belgium) yasetshenziselwa ukukhiqiza ivolumu ye-3D yesampula eskeniwe kusukela ku-2D etholiwe ye-lateral projection.Ukuze kuhlaziywe, isithombe esakhiwe kabusha se-3D sihlukaniswe ngamakhyubhu angu-10mm×10mm×10mm ngokuya ngendawo yokuphuka.Bala i-callus ngaphandle kwethambo le-cortical.I-DataViewer (inguqulo 1.5.1.2; isofthiwe ye-Bruker microCT, Kontich, Belgium) yasetshenziselwa ukuqondisa kabusha ngedijithali umthamo wamathambo askeniwe, kanye nesofthiwe ye-CT-Analyzer (inguqulo 1.14.4.1; Bruker microCT, Kontich, Belgium) ukuze kuhlaziywe.Ama-coefficients ahlobene wokumuncwa kwe-x-ray ethanjeni elivuthiwe kanye ne-callus ahlukaniswa ngokuminyana kwawo, bese ivolumu ye-callus ibalwa (n = 4).Ukuze kuqinisekiswe ukuthi i-biocompatibility ye-LOIS ayilibazisi inqubo yokuphulukisa amathambo, ukuhlaziya okwengeziwe kwe-X-ray kanye ne-micro-CT kwenziwa onogwaja ababili: amaqembu anqunu-negative kanye ne-LOIS.Womabili amaqembu abulawa ngesonto lesi-6.
Ama-femurs avela ezilwaneni ezihlatshelwe aqoqwa futhi afakwa ku-4% we-paraformaldehyde izinsuku ezi-3.I-orthopedic implant ikhishwa ngokucophelela ku-femur.I-femur yaqedwa izinsuku ezingama-21 kusetshenziswa i-0.5 M EDTA (EC-900, National Diagnostics Corporation).Khona-ke i-femur esusiwe yacwiliswa ku-EtOH ukuze iphelelwe amanzi.I-femur ephelelwe amanzi emzimbeni ikhishwe ku-xylene futhi ifakwe kupharafini.Ngemva kwalokho isampula yasikwa nge-automatic rotary microtome (Leica RM2255, Leica Biosystems, Germany) enogqinsi luka-3 μm.Ngokugcotshwa kwe-TRAP (F6760, Sigma-Aldrich, Germany), amasampuli ahlukaniswe akhishwa ipharafini, afakwa kabusha emanzini futhi afakwa ku-reagent ye-TRAP ku-37°C ihora elingu-1.Izithombe zitholwe kusetshenziswa isithwebuli samaslayidi (i-Pannoramic 250 Flash III, 3DHISTECH, Hungary) futhi salinganiswa ngokulinganisa ukumbozwa kwendawo yendawo enamabala.Esivivinyweni ngasinye, okungenani ama-substrates amane eqenjini ngalinye ahlaziywa isofthiwe ye-ImageJ.
Ukuhlaziywa kokubaluleka kwezibalo kwenziwa kusetshenziswa i-GraphPad Prism (GraphPad Software Inc., USA).Ukuhlolwa kwe-t okungabhanqiwe nokuhlaziywa kwendlela eyodwa yokuhluka (ANOVA) kwasetshenziswa ukuze kuhlolwe umehluko phakathi kwamaqembu okuhlola.Izinga lokubaluleka liboniswa emfanekisweni ngale ndlela elandelayo: *P<0.05, **P<0.01, ***P<0.001 kanye ****P<0.0001;NS, awukho umehluko obalulekile.
Ukuze uthole izinto ezengeziwe zalesi sihloko, sicela ubheke ku-http://advances.sciencemag.org/cgi/content/full/6/44/eabb0025/DC1
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