Ga majinyatan da ake yi wa tiyatar kafa kothopedic, cututtuka na ƙwayoyin cuta da ƙwayoyin cuta da ke haifar da rigakafi sun kasance haɗari masu haɗari.Kayayyakin halitta na al'ada suna da sauƙin kamuwa da gurɓataccen yanayi, wanda ke haifar da ƙwayoyin cuta su mamaye yankin da suka ji rauni kuma suna haifar da kamuwa da cuta bayan tiyata.Sabili da haka, akwai buƙatar gaggawa don haɓaka rigakafin kamuwa da cuta da suturar tserewa na rigakafi don ƙwanƙwasa orthopedic.Anan, mun ƙirƙira fasahar gyare-gyare na ci gaba don ƙwanƙwasa kasusuwa mai suna Lubricated Orthopedic Implant Surface (LOIS), wanda aka yi wahayi zuwa gare shi ta hanyar santsin saman tulun shukar tulu.LOIS yana da ɗorewa kuma mai ƙarfi mai hana ruwa zuwa nau'ikan ruwaye da abubuwan halitta (ciki har da sel, sunadarai, calcium da ƙwayoyin cuta).Bugu da kari, mun tabbatar da dorewar injina akan karce da kayyade karfi ta hanyar siffanta lalacewar da babu makawa yayin tiyatar in vitro.An yi amfani da ƙirar kasusuwan kasusuwa mai kumburi mai kumburi na mata don yin nazari sosai game da sikelin ƙwayoyin cuta da kuma rigakafin kamuwa da cuta na LOIS.Muna tunanin cewa LOIS, wanda ke da kaddarorin anti-biofouling da dorewar inji, mataki ne na ci gaba a cikin tiyatar ƙasusuwa marasa kamuwa da cuta.
A yau, saboda yawan tsufa, yawan marasa lafiya da ke fama da cututtuka na orthopedic (kamar tsofaffin tsofaffi, cututtuka na haɗin gwiwa, da osteoporosis) sun karu sosai (1, 2).Sabili da haka, cibiyoyin kiwon lafiya suna ba da mahimmanci ga aikin tiyata na kasusuwa, ciki har da gyare-gyaren kashin baya na sukurori, faranti, kusoshi da haɗin gwiwar wucin gadi (3, 4).Duk da haka, an ba da rahoton ƙwararrun ƙasusuwa na gargajiya suna da sauƙi ga mannewar ƙwayoyin cuta da kuma samuwar biofilm, wanda zai iya haifar da kamuwa da cutar ta wurin tiyata (SSI) bayan tiyata (5, 6).Da zarar an samar da biofilm a saman ƙwanƙwasa orthopedic, cirewar biofilm ya zama mai wuyar gaske har ma da amfani da manyan allurai na rigakafi.Saboda haka, yawanci yana haifar da cututtuka masu tsanani bayan tiyata (7, 8).Saboda matsalolin da ke sama, maganin cututtukan da suka kamu da cutar ya kamata ya haɗa da sake yin aiki, gami da cire duk abubuwan da aka sanyawa da nama da ke kewaye;saboda haka, mai haƙuri zai sha wahala mai tsanani da wasu haɗari (9, 10).
Don magance wasu daga cikin waɗannan matsalolin, an ƙera kayan dasa kothopedic da ke kawar da ƙwayoyi don hana kamuwa da cuta ta hanyar kawar da ƙwayoyin cuta da ke haɗe a saman (11, 12).Koyaya, dabarun har yanzu yana nuna iyakoki da yawa.An ba da rahoton cewa dasa shuki na dogon lokaci na ƙwayoyin cuta na ƙwayoyin cuta ya haifar da lalacewa ga kyallen da ke kewaye da su kuma ya haifar da kumburi, wanda zai iya haifar da necrosis (13, 14).Bugu da kari, abubuwan kaushi na kwayoyin halitta wadanda zasu iya wanzuwa bayan tsarin masana'antu na masana'anta na gyaran kasusuwa, wadanda Hukumar Abinci da Magunguna ta Amurka ta hana su, suna buƙatar ƙarin matakan tsarkakewa don saduwa da ƙa'idodinta (15).Abubuwan da ke kawar da ƙwayoyi suna ƙalubalanci don sarrafa magungunan ƙwayoyi, kuma saboda iyakancewar lodin ƙwayoyi, aikace-aikacen maganin na dogon lokaci ba zai yiwu ba (16).
Wata dabara ta gama gari ita ce a sa suturar da aka dasa tare da polymer antifouling don hana kwayoyin halitta da ƙwayoyin cuta daga mannewa saman (17).Misali, polymers na zwitterionic sun jawo hankali saboda abubuwan da ba su da mannewa lokacin da suke hulɗa da sunadaran plasma, sel, da ƙwayoyin cuta.Duk da haka, yana da wasu ƙayyadaddun ƙayyadaddun da suka danganci kwanciyar hankali na dogon lokaci da kuma dorewa na inji, wanda ke hana aikace-aikacensa na aiki a cikin kayan aiki na orthopedic, musamman saboda ƙwanƙwasa na inji a lokacin aikin tiyata (18, 19).Bugu da ƙari, saboda haɓakar haɓakaccen haɓakaccen haɓakaccen haɓaka, rashin buƙata don cire tiyata, da kayan tsaftacewa ta hanyar lalata, an yi amfani da kayan da aka yi da orthopedic da aka yi da kayan haɓaka (20, 21).A lokacin lalata, haɗin sinadarai tsakanin matrix polymer ya rushe kuma an cire su daga saman, kuma masu bin su suna tsaftace saman.Duk da haka, lalatawar ƙwayoyin cuta ta hanyar tsaftacewa na sama yana da tasiri a cikin ɗan gajeren lokaci.Bugu da kari, mafi absorbable kayan ciki har da poly (lactic acid-glycolic acid copolymer) (PLGA), polylactic acid (PLA) da magnesium na tushen gami za su sha m biodegradation da yashwa a cikin jiki, wanda zai barnatar da tasiri na inji kwanciyar hankali.(ashirin da biyu).Bugu da ƙari, gutsuttsuran farantin da za su iya ba da wuri don ƙwayoyin cuta su haɗa, wanda ke ƙara yiwuwar kamuwa da cuta a cikin dogon lokaci.Wannan haɗarin lalacewar injina da kamuwa da cuta yana iyakance aikace-aikacen aikin tiyata na filastik (23).
Fuskokin Superhydrophobic (SHP) waɗanda ke kwaikwayi tsarin tsarin ganyayen magarya sun zama mafita mai yuwuwa ga filaye masu ɓarna (24, 25).Lokacin da SHP surface aka nutsar a cikin ruwa, iska kumfa za a cikin tarko, game da samar da iska aljihu da kuma hana kwayan cuta adhesion (26).Duk da haka, binciken da aka yi kwanan nan ya nuna cewa SHP surface yana da rashin lahani da ke da alaka da ƙarfin injiniya da kwanciyar hankali na dogon lokaci, wanda ke hana aikace-aikacensa a cikin magungunan likita.Bugu da ƙari, aljihunan iska za su narke kuma su rasa kaddarorin su na ƙazanta, don haka yana haifar da mannewa na kwayan cuta saboda babban yanki na SHP (27, 28).Kwanan nan, Aizenberg da abokan aiki sun gabatar da wata sabuwar hanya ta hana ƙorafi ta hanyar haɓaka shimfidar wuri mai santsi wanda aka yi wahayi daga shukar Nepenthes pitter (29, 30).Filaye mai santsi yana nuna kwanciyar hankali na dogon lokaci a ƙarƙashin yanayi na ruwa, yana da matukar juriya ga ruwayen halittu, kuma yana da abubuwan gyara kai.Duk da haka, babu wata hanyar da za a yi amfani da sutura zuwa wani hadadden nau'i na likita, kuma ba a tabbatar da goyon bayan tsarin warkarwa na nama mai lalacewa bayan dasawa.
Anan, mun gabatar da lubricated orthopedic implant surface (LOIS), micro/nano-structured orthopedic implant surface da tam a haɗe tare da bakin ciki mai laushi Layer don hana shi daga hade da filastik tiyata Kwayoyin cututtuka, kamar karaya gyarawa.Saboda tsarin matakin micro/nano-mai aiki da fluorine yana tabbatar da mai mai akan tsarin, haɓakar LOIS na iya cika cikar mannewar ruwa iri-iri da kuma kula da aikin hana lalata na dogon lokaci.Ana iya amfani da suturar LOIS zuwa kayan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan nau'ikan) ana yin amfani da su ana amfani da su.Kyawawan kaddarorin anti-biofouling na LOIS akan ƙwayoyin cuta na biofilm [Pseudomonas aeruginosa da methicillin-resistant Staphylococcus aureus (MRSA)] da abubuwan halitta (kwayoyin halitta, sunadarai da calcium) an tabbatar dasu a cikin vitro.Adadin mannewa na mannewa mai yawa zuwa ga substrate bai wuce 1%.Bugu da ƙari, ko da bayan damuwa na inji irin su zazzage saman sama ya faru, warkar da kai da aka samu ta hanyar mai mai shiga yana taimakawa wajen kula da abubuwan da ba su da kyau.Sakamakon gwajin dorewa na inji ya nuna cewa ko da bayan gyare-gyaren tsari da sinadarai, jimlar ƙarfin ba zai ragu sosai ba.Bugu da ƙari, an gudanar da gwajin in vitro wanda ke kwatanta damuwa na inji a cikin aikin tiyata don tabbatar da cewa LOIS na iya jure wa matsalolin inji daban-daban da ke faruwa a lokacin aikin filastik.A ƙarshe, mun yi amfani da zomo na tushen a cikin vivo femoral fracture model, wanda ya tabbatar da cewa LOIS yana da mafi girman kaddarorin ƙwayoyin cuta da ƙwayoyin cuta.Sakamakon radiyo da na tarihi sun tabbatar da cewa bargaren halayen mai da kuma kaddarorin anti-biofouling a cikin makonni 4 bayan dasawa na iya samun ingantaccen rigakafin kamuwa da cuta da aikin tserewa na rigakafi ba tare da jinkirta tsarin warkar da kashi ba.
Hoto na 1A yana nuna zane-zane na LOIS da aka haɓaka, wanda aka dasa tare da ƙananan sikelin sikelin / nano-nano a cikin ƙirar zomo na mata don tabbatar da kyakkyawan yanayin lalata ƙwayoyin halitta da kaddarorin kamuwa da cuta.Ana aiwatar da hanyar biomimetic don kwaikwaya saman shukar tukunyar ruwa, da kuma hana ɓarkewar halitta ta hanyar haɗa Layer mai mai a cikin tsarin micro/nano na saman.Filayen da aka yi masa allura tare da mai mai na iya rage cudanya tsakanin abubuwan halitta da saman.Saboda haka, saboda samuwar barga sunadarai bond a saman, yana da kyakkyawan aikin antifouling da kwanciyar hankali na dogon lokaci.A sakamakon haka, Properties anti-biofouling na lubricating surface damar daban-daban m aikace-aikace a biomedical bincike.Duk da haka, bincike mai zurfi kan yadda wannan fili na musamman ke hulɗa a cikin jiki bai riga ya kammala ba.Ta hanyar kwatanta LOIS tare da tsirara tsirara a cikin vitro ta amfani da albumin da kwayoyin halitta na biofilm, ana iya tabbatar da rashin mannewa na LOIS (Hoto 1B).Bugu da kari, ta hanyar mirgine ɗigon ruwa a kan madaidaicin madaidaicin madauri da kuma ma'aunin LOIS (Hoto S1 da Fim ɗin S1), ana iya nuna aikin gurɓataccen yanayi.Kamar yadda aka nuna a hoton microscope mai kyalli, abin da aka fallasa da aka fallasa a cikin dakatarwar furotin da ƙwayoyin cuta ya nuna adadi mai yawa na kayan halitta masu mannewa saman.Koyaya, saboda kyawawan kaddarorin anti-biofouling, LOIS da kyar ke nuna wani haske.Don tabbatar da maganin ƙwayoyin cuta da ƙwayoyin cuta, an yi amfani da LOIS a saman kayan da aka yi da orthopedic don haɗin kashi (faranti da sukurori) kuma an sanya shi a cikin samfurin karayar zomo.Kafin a dasa shi, tsirara kothopedic implant da LOIS an sanya su a cikin dakatarwar kwayan cuta na sa'o'i 12.Pre-incubation yana tabbatar da cewa an samar da biofilm a kan farfajiyar da aka fallasa don kwatantawa.Hoto na 1C yana nuna hoton wurin karyewar makonni 4 bayan dasawa.A gefen hagu, wani zomo tare da daɗaɗɗen kashin baya ya nuna mummunan matakin kumburi saboda samuwar biofilm a saman da aka sanya.An lura da akasin sakamako a cikin zomaye da aka dasa tare da LOIS, wato, ƙwayoyin da ke kewaye da LOIS ba su nuna alamun kamuwa da cuta ko alamun kumburi ba.Bugu da ƙari, hoton hoton da ke gefen hagu yana nuna wurin aikin tiyata na zomo tare da dasa shuki, wanda ke nuna cewa ba a sami wasu nau'i-nau'i masu yawa da ke kan farfajiyar da aka fallasa a saman LOIS ba.Wannan yana nuna cewa LOIS yana da kwanciyar hankali na dogon lokaci kuma yana da ikon kula da abubuwan da ke lalata ƙwayoyin halitta da abubuwan da ke hana mannewa.
(A) Tsarin tsari na LOIS da dasa shi a cikin samfurin karyewar mata na zomo.(B) Hoton microscopy mai kyalli na furotin da kwayoyin halitta na kwayoyin halitta a kan danda ba a san shi ba da kuma wurin LOIS.Makonni 4 bayan dasawa, (C) hoton hoto na wurin karyewar da (D) hoton X-ray (wanda jan rectangle ya haskaka).Hoton hoto: Kyomin Chae, Jami'ar Yonsei.
Haifuwa, fallasa mara kyau dasa zomaye sun nuna tsarin warkar da kashi na yau da kullun ba tare da alamun kumburi ko kamuwa da cuta ba.A daya hannun, SHP implants pre-cubated a cikin wani kwayan cuta dakatar nuna kamuwa da cuta da alaka kumburi a kewaye da kyallen takarda.Ana iya danganta wannan ga rashin iya hana mannewar kwayan cuta na dogon lokaci (Hoto S2).Don tabbatar da cewa LOIS ba zai shafi tsarin warkaswa ba, amma yana hana yiwuwar cututtuka da suka shafi dasa, hotunan X-ray na matrix mai kyau da aka fallasa da LOIS a wurin da aka lalata (Figure 1D).Hoton X-ray na dasawa mai inganci ya nuna layin osteolysis na yau da kullun, yana nuna cewa kashi bai gama warkewa ba.Wannan yana nuna cewa tsarin dawo da kashi na iya jinkirtawa sosai saboda kumburi da ke da alaƙa da kamuwa da cuta.Akasin haka, ya nuna cewa zomaye da aka dasa tare da LOIS sun warke kuma basu nuna wani wurin karaya ba.
Don haɓaka kayan aikin likita tare da kwanciyar hankali na dogon lokaci da aiki (ciki har da juriya ga biofouling), an yi ƙoƙari da yawa.Duk da haka, kasancewar abubuwa daban-daban na ilimin halitta da haɓakar mannewar nama yana iyakance haɓaka hanyoyin dogaro da su na asibiti.Domin shawo kan waɗannan gazawar, mun ƙirƙiri wani tsari mai laushi na micro/nano da sinadarai gyare-gyare, wanda aka inganta saboda babban ƙarfin capillary da alaƙar sinadarai don kiyaye mafi kyawun mai mai zuwa mafi girma.Hoto 2A yana nuna gaba ɗaya tsarin masana'antu na LOIS.Na farko, shirya wani likita sa bakin karfe (SS) 304 substrate.Na biyu, an kafa tsarin micro/nano akan SS substrate ta hanyar sinadarai ta hanyar amfani da maganin hydrofluoric acid (HF).Don dawo da juriya na lalata na SS, ana amfani da maganin nitric acid (HNO3) (31) don aiwatar da ƙaƙƙarfan ɓangarorin.Passivation yana haɓaka juriya na lalata na SS kuma yana rage jinkirin tsarin lalata wanda zai iya rage ayyukan LOIS gabaɗaya.Sa'an nan kuma, ta hanyar samar da monolayer mai haɗa kai (SAM) tare da 1H, 1H, 2H, 2H-perfluorooctyltriethoxysilane (POTS), an gyare-gyaren saman ta hanyar sinadarai don inganta hulɗar sinadarai tsakanin farfajiya da santsi mai laushi.gyare-gyaren saman yana da mahimmanci yana rage ƙarfin daɗaɗɗen ƙirar micro/nano-sikelin da aka ƙera, wanda yayi daidai da ƙarfin saman mai mai santsi.Wannan yana ba da damar man shafawa gaba ɗaya ya jike, ta yadda zai samar da barga mai ƙoshin mai a saman.Fuskar da aka gyara tana nuna ingantaccen hydrophobicity.Sakamakon ya nuna cewa lubricant mai zamewa yana nuna halin kwanciyar hankali akan LOIS saboda babban alaƙar sinadarai da ƙarfin capillary wanda tsarin micro/nano ya haifar (32, 33).Canje-canje na gani a saman SS bayan gyaran fuska da allura mai mai.Tsarin sikirin micro/nano da aka kafa akan saman zai iya haifar da canje-canje na gani kuma ya duhunta saman.Ana danganta wannan al'amari ga ingantaccen tasirin watsawa na haske a kan ƙasa maras kyau, wanda ke ƙara haɓakar hangen nesa ta hanyar tsarin kama haske (34).Bugu da kari, bayan an yi wa mai mai, LOIS ya yi duhu.Layin mai mai yana haifar da ƙarancin haske da ke fitowa daga cikin ƙasa, don haka duhun LOIS.Domin inganta microstructure/nanostructure don nuna mafi ƙarancin kusurwar zamiya (SA) don cimma aikin anti-biofouling, ana amfani da microscopy na lantarki (SEM) da nau'ikan atomic don yin lokutan etching daban-daban na HF (0, 3)., Minti 15 da 60) Ƙarfin Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwararren Ƙwaƙwalwa 2B ) ya yi.Hotunan SEM da AFM sun nuna cewa bayan ɗan gajeren lokaci na etching (minti 3 na etching), ɓangarorin da ba su da tushe sun haifar da rashin daidaituwa na nano-sikelin.Ƙarfin saman yana canzawa tare da lokacin etching (Hoto S3).Matsakaicin sauye-sauyen lokaci yana nuna cewa ƙarancin saman yana ci gaba da karuwa kuma ya kai kololuwa a cikin mintuna 15 na etching, sannan kuma an sami raguwa kaɗan a ƙimar rashin ƙarfi a cikin mintuna 30 na etching.A wannan lokaci, ƙarancin matakin nano ya ɓace, yayin da ƙananan ƙananan matakan ke tasowa da ƙarfi, yana sa rashin ƙarfi ya canza mafi kwanciyar hankali.Bayan etching fiye da minti 30, ana ƙara haɓakar rashin ƙarfi, wanda aka yi bayani dalla-dalla kamar haka: SS ya ƙunshi ƙarfe, gami da abubuwan da suka haɗa da baƙin ƙarfe, chromium, nickel, molybdenum da sauran abubuwa masu yawa.Daga cikin waɗannan abubuwa, baƙin ƙarfe, chromium da molybdenum suna taka muhimmiyar rawa wajen samar da sikelin micron/nano akan SS ta HF etching.A farkon matakan lalata, baƙin ƙarfe da chromium sun fi lalacewa saboda molybdenum yana da juriya na lalata fiye da molybdenum.Yayin da etching ke ci gaba, maganin etching ya kai ga yawan kitse na gida, yana samar da fluorides da oxides da ke haifar da etching.Fluoride da oxide suna hazo kuma a ƙarshe suna sake ajiya akan saman, suna samar da rashin ƙarfi a cikin kewayon micron/nano (31).Wannan ƙaƙƙarfan matakin micro/nano yana taka muhimmiyar rawa a cikin abubuwan warkar da kai na LOIS.Fuskar sikelin dual yana haifar da tasirin aiki tare, yana ƙara ƙarfin ƙarfin capillary sosai.Wannan sabon abu yana ba da damar mai mai don kutsawa saman saman a tsaye kuma yana ba da gudummawa ga kaddarorin warkar da kai (35).Samuwar roughness ya dogara da lokacin etching.A ƙarƙashin mintuna 10 na etching, saman yana ƙunshe da ƙaƙƙarfan sikelin nano kawai, wanda bai isa ya riƙe isasshen mai don samun juriya na biofouling ba (36).A gefe guda, idan lokacin etching ya wuce minti 30, ƙarancin nano-sikelin da aka samu ta hanyar sake fasalin baƙin ƙarfe da chromium zai ɓace, kuma ƙananan sikelin kawai zai kasance saboda molybdenum.Wurin da ya wuce gona da iri ba shi da ƙaƙƙarfan ma'auni na nano kuma ya rasa tasirin haɗin gwiwa na ƙaƙƙarfan matakai biyu, wanda ya yi mummunan tasiri ga halayen warkarwa na LOIS.An yi ma'aunin SA akan ma'auni tare da lokuta daban-daban don tabbatar da aikin hana lalata.An zaɓi nau'ikan ruwa iri-iri dangane da danko da makamashi na sama, gami da ruwa (DI) ruwa, jini, ethylene glycol (EG), ethanol (EtOH) da hexadecane (HD) (Hoto S4).Tsarin etching na lokaci-lokaci yana nuna cewa ga ruwa daban-daban tare da kuzari daban-daban da kuma danko, SA na LOIS bayan mintuna 15 na etching shine mafi ƙanƙanta.Don haka, LOIS an inganta shi don tsarawa na mintina 15 don samar da micron da nano-sikelin roughness, wanda ya dace don kiyaye karko na mai mai da kyawawan kaddarorin hana lalata.
(A) Jadawalin tsari na tsarin samar da matakai huɗu na LOIS.Saitin yana nuna SAM da aka kafa akan ma'auni.(B) Hotunan SEM da AFM, waɗanda aka yi amfani da su don inganta tsarin micro/nano na substrate a ƙarƙashin lokuta daban-daban.X-ray photoelectron spectroscopy (XPS) spectra na (C) Cr2p da (D) F1s bayan wucewar sararin samaniya da shafi SAM.au, naúrar sabani.(E) Hotunan wakilci na ɗigon ruwa a kan danda, kwai, SHP da LOIS substrates.(F) kusurwar lamba (CA) da ma'aunin SA na ruwa tare da tashin hankali daban-daban akan SHP da LOIS.Ana bayyana bayanai azaman ma'anar ± SD.
Sa'an nan, domin tabbatar da canji a cikin sinadaran Properties na surface, X-ray photoelectron spectroscopy (XPS) da aka yi amfani da nazarin canji a cikin sinadaran abun da ke ciki na substrate surface bayan kowane surface shafi.Hoto 2C yana nuna sakamakon ma'aunin XPS na HF etched surface da kuma HNO 3 da aka yi magani.Babban kololuwa biyu a 587.3 da 577.7 eV ana iya danganta su da haɗin Cr-O da ke cikin Layer chromium oxide, wanda shine babban bambanci daga saman HF etched.Wannan ya faru ne saboda yawan amfani da ƙarfe da chromium fluoride a saman ta HNO3.Etching na tushen HNO3 yana ba da damar chromium don samar da Layer oxide mai wucewa a saman, wanda ke sa SS ɗin da aka ƙera ya sake juriya ga lalata.A cikin Hoto na 2D, an sami sifofin XPS don tabbatar da cewa silane na tushen fluorocarbon an kafa shi a saman bayan murfin SAM, wanda ke da matsanancin ƙarancin ruwa har ma ga EG, jini da EtOH.An kammala suturar SAM ta hanyar mayar da martani ga ƙungiyoyi masu aiki na silane tare da ƙungiyoyin hydroxyl waɗanda aka kafa ta hanyar jiyya na plasma.A sakamakon haka, an sami karuwa mai yawa a cikin CF2 da CF3 kololuwa.Ƙarfin dauri tsakanin 286 da 296 eV yana nuna cewa an sami nasarar kammala gyaran sinadarai ta hanyar shafan SAM.SHP yana nuna ƙananan kololuwar CF2 (290.1 ​​​​eV) da CF3 (293.3 eV), waɗanda ke haifar da silane na tushen fluorocarbon da aka kafa a saman.Hoto na 2E yana nuna ma'auni na ɗimbin hotuna na gani na wakilci na kusurwar lamba (CA) don ƙungiyoyi daban-daban na ruwan da aka lalatar da su a cikin hulɗa tare da danda, etched, SHP, da LOIS.Wadannan hotuna sun nuna cewa saman da aka kwaikwayi ya zama hydrophilic saboda tsarin micro/nano da aka samar ta hanyar sinadarai ta yadda za a tsotse ruwa a cikin tsarin.Duk da haka, lokacin da aka yi amfani da substrate tare da SAM, mai amfani yana nuna karfin ruwa mai karfi, don haka an kafa SHP surface kuma wurin da ke tsakanin ruwa da ƙasa yana da ƙananan.A ƙarshe, an sami raguwar CA a cikin LOIS, wanda za'a iya danganta shi da shigar da mai mai a cikin microstructure, ta haka yana ƙaruwa wurin hulɗa.Domin tabbatar da cewa saman yana da kyawawan ƙayyadaddun ƙayyadaddun ruwa da kaddarorin da ba a haɗa su ba, an kwatanta LOIS tare da ma'aunin SHP ta hanyar auna CA da SA ta amfani da ruwa daban-daban (Hoto 2F).An zaɓi nau'ikan ruwa iri-iri dangane da danko da kuzarin ƙasa, gami da ruwa mai tsafta, jini, EG, EtOH da HD (Hoto S4).Sakamakon ma'auni na CA ya nuna cewa lokacin da CA ke kula da HD, ƙimar ragewa na CA, inda CA ke da mafi ƙarancin makamashi.Bugu da ƙari, LOIS na gaba ɗaya CA yana da ƙasa.Koyaya, ma'aunin SA yana nuna wani al'amari na daban.Ban da ruwan ionized, duk ruwaye suna manne da ma'aunin SHP ba tare da zamewa ba.A gefe guda, LOIS yana nuna ƙarancin SA, inda duk ruwan ya karkata a kusurwa ƙasa da 10 ° zuwa 15 °, duk ruwan zai juye.Wannan yana nuna ƙarfi sosai cewa rashin mannewa na LOIS ya fi na SHP surface.Bugu da kari, LoIS coatings kuma ana amfani da daban-daban iri kayan, ciki har da titanium (Ti), polyphenylsulfone (PPSU), polyoxymethylene (POM), polyether ether ketone (PEEK) da kuma bioabsorbable polymers (PLGA), Su ne implantable orthopedic kayan (Figure). S5)).Hotunan jeri na ɗigogi a kan kayan da LOIS ke bi da su sun nuna cewa kaddarorin anti-biofouling na LOIS iri ɗaya ne akan duk abubuwan da ake amfani da su.Bugu da ƙari, sakamakon ma'auni na CA da SA ya nuna cewa za a iya amfani da abubuwan da ba su da mannewa na LOIS zuwa wasu kayan.
Domin tabbatar da kaddarorin hana lalata na LOIS, nau'ikan nau'ikan nau'ikan nau'ikan (ciki har da bare, etched, SHP da LOIS) an haɗa su tare da Pseudomonas aeruginosa da MRSA.An zaɓi waɗannan ƙwayoyin cuta guda biyu a matsayin wakili na ƙwayoyin cuta na asibiti, wanda zai iya haifar da samuwar biofilms, haifar da SSI (37).Hoto na 3 (A da B) yana nuna Hotunan kyakyawan kyakyawan kyalli da sakamakon ma'aunin ma'aunin ma'auni (CFU) na abubuwan da aka sanya a cikin dakatarwar kwayan cuta na ɗan gajeren lokaci (awanni 12) da na dogon lokaci (72 hours), bi da bi.A cikin kankanin lokaci kwayoyin cuta za su yi tari kuma su yi girma, suna rufe kansu da abubuwa masu kama da ƙora da hana cire su.Koyaya, a cikin sa'o'i 72, ƙwayoyin cuta za su girma kuma su zama masu sauƙin tarwatsewa don samar da ƙarin yankuna ko gungu.Sabili da haka, ana iya la'akari da cewa 72-hour incubation yana da dogon lokaci kuma shine lokacin da ya dace don samar da ingantaccen biofilm a saman (38).A cikin ɗan gajeren lokaci, ƙwanƙwasa da kuma saman SHP sun nuna alamar ƙwayar cuta, wanda aka rage da kusan 25% zuwa 50% idan aka kwatanta da simintin da ba a so.Koyaya, saboda kyakkyawan aikin anti-biofouling da kwanciyar hankali, LOIS bai nuna mannewar biofilm na kwayan cuta a cikin ɗan gajeren lokaci da dogon lokaci ba.Tsarin tsari (Hoto 3C) yana bayyana bayanin tsarin lalata ƙwayoyin cuta na maganin etching, SHP da LOIS.Zaton shi ne cewa ƙwanƙwasa da ke da abubuwan hydrophilic za su sami yanki mafi girma fiye da dandali.Sabili da haka, ƙarin mannewa na kwayan cuta zai faru a kan ƙwanƙwasa.Duk da haka, idan aka kwatanta da abin da ba a kwance ba, ƙwaƙƙwarar da aka yi da shi ba ta da ƙarancin biofilm da aka kafa a saman.Wannan shi ne saboda kwayoyin ruwa suna ɗaure da ƙarfi zuwa saman hydrophilic kuma suna aiki azaman mai mai don ruwa, don haka suna tsoma baki tare da manne kwayoyin cuta a cikin ɗan gajeren lokaci (39).Koyaya, Layer na kwayoyin ruwa yana da siriri sosai kuma yana narkewa a cikin dakatarwar kwayan cuta.Sabili da haka, layin kwayoyin ruwa yana ɓacewa na dogon lokaci, yana haifar da mannewa da yawa na ƙwayoyin cuta.Don SHP, saboda gajeriyar kaddarorin sa na ɗan gajeren lokaci, an hana mannewa na kwayan cuta.Ana iya danganta raguwar mannewa na kwayan cuta zuwa aljihun iska da aka makale a cikin tsari mai laushi da ƙananan makamashi na ƙasa, don haka rage hulɗa tsakanin dakatarwar kwayan cuta da saman.Koyaya, an lura da mannewa na kwayan cuta mai yawa a cikin SHP saboda ya rasa abubuwan da ya dade yana lalata su.Wannan ya faru ne saboda bacewar aljihunan iska saboda matsa lamba na ruwa da kuma narkar da iska a cikin ruwa.Wannan ya faru ne saboda bacewar aljihunan iska saboda rushewa da kuma tsarin da aka yi da shi wanda ke ba da wuri mafi girma don mannewa (27, 40).Ba kamar waɗannan nau'ikan nau'ikan guda biyu waɗanda ke da tasiri mai mahimmanci akan kwanciyar hankali na dogon lokaci, lubricating lubricant da ke cikin LOIS ana allura a cikin tsarin micro / nano kuma ba zai ɓace ba har ma a cikin dogon lokaci.Man shafawa da ke cike da ƙananan sifofi/nano suna da ƙarfi sosai kuma ana sha'awar su sosai a saman saboda yawan alaƙar sinadarai, don haka suna hana mannewar ƙwayoyin cuta na dogon lokaci.Hoto S6 yana nuna hoton microscope mai jujjuyawa na wani abu mai mai mai wanda aka nitse cikin salin buffered phosphate (PBS).Hotuna masu ci gaba suna nuna cewa ko da bayan sa'o'i 120 na ɗan girgiza (120 rpm), Layer na man shafawa akan LOIS ya kasance baya canzawa, yana nuna kwanciyar hankali na dogon lokaci a ƙarƙashin yanayin gudana.Wannan ya faru ne saboda ƙaƙƙarfan alaƙar sinadarai tsakanin rufin SAM na tushen fluorine da man shafawa na tushen perfluorocarbon, ta yadda za a iya samar da madaidaicin madaurin mai.Sabili da haka, ana kiyaye aikin hana lalata.Bugu da ƙari, an gwada substrate akan furotin na wakilci (albumin da fibrinogen), waɗanda ke cikin plasma, ƙwayoyin da ke da alaƙa da aikin rigakafi (macrophages da fibroblasts), da waɗanda ke da alaƙa da samuwar kashi.Abubuwan da ke cikin calcium yana da yawa sosai.(Hoto na 3D, 1 da 2, da Hoto S7) (41, 42).Bugu da ƙari, Hotunan microscope mai kyalli na gwajin mannewa don fibrinogen, albumin da calcium sun nuna halaye daban-daban na mannewa na kowane rukuni (Hoto S8).A lokacin samuwar kashi, sabbin kasusuwa da yadudduka na calcium na iya kewaye da kafawar kasusuwa, wanda ba wai kawai ya sa cirewa ke da wahala ba, amma kuma yana iya haifar da lahani maras tsammanin ga majiyyaci yayin aikin cirewa.Sabili da haka, ƙananan matakan alli a kan faranti na kasusuwa da sukurori suna da amfani ga aikin tiyata na orthopedic wanda ke buƙatar cirewa.Dangane da ƙididdigewa na yankin da aka haɗe dangane da ƙarfin hasken wuta da ƙididdigar tantanin halitta, mun tabbatar da cewa LOIS yana nuna kyawawan kaddarorin anti-biofouling ga duk abubuwan halitta idan aka kwatanta da sauran abubuwan.Dangane da sakamakon gwaje-gwajen in vitro, za a iya amfani da maganin hana ƙwayoyin cuta LOIS a cikin gyare-gyare na orthopedic, wanda ba zai iya hana cututtukan da kwayoyin halitta ke haifar da su ba, har ma da rage kumburi da tsarin garkuwar jiki ke haifarwa.
(A) Hotunan microscope na Fluorescence na kowane rukuni (tsirara, etched, SHP da LOIS) da aka sanya su a cikin Pseudomonas aeruginosa da dakatarwar MRSA na awanni 12 da 72.(B) Yawan masu bin CFU na Pseudomonas aeruginosa da MRSA a saman kowane rukuni.(C) Tsarin tsari na tsarin lalata ƙwayoyin cuta na gajeren lokaci da na dogon lokaci, SHP da LOIS.(D) (1) Adadin fibroblasts da ke manne da kowane nau'i da hotuna masu kyalli na sel da ke manne da danda da LOIS.(2) Gwajin adhesion na sunadaran da ke da alaƙa da rigakafi, albumin da calcium da ke cikin tsarin warkar da kashi (* P <0.05, ** P <0.01, *** P <0.001 da **** P <0.0001).ns, ba mahimmanci.
A cikin yanayin matsalolin da ba za a iya kaucewa ba, ƙarfin injin ya kasance koyaushe babban ƙalubale don aikace-aikacen suturar lalata.Hanyoyin gel na anti-najasa na al'ada sun dogara ne akan polymers tare da ƙarancin ruwa da rashin ƙarfi.Sabili da haka, yawanci suna da sauƙi ga damuwa na inji a aikace-aikacen ilimin halitta.Sabili da haka, kayan kwalliyar antifouling masu ɗorewa na injiniya sun kasance ƙalubale ga aikace-aikace kamar su ƙwanƙwasa orthopedic (43, 44).Hoto na 4A (1) yana nuna nau'ikan nau'ikan nau'ikan damuwa guda biyu da ake amfani da su a cikin ƙwanƙwasa orthopedic, ciki har da ƙwanƙwasa (danniya mai ƙarfi) da matsawa tare da hoton gani na lalacewa da aka samu ta hanyar tilastawa.Misali, lokacin da aka ɗora dunƙule tare da screwdriver, ko kuma lokacin da likitan fiɗa ya riƙe farantin kasusuwa tare da tweezers kuma yana amfani da ƙarfi mai ƙarfi, farantin kashin filastik zai lalace kuma a toshe shi akan ma'aunin macro da micro/nano (Hoto 4A, 2) .Don gwada ko ƙera LOIS zai iya jure wa waɗannan lalacewa yayin aikin tiyata na filastik, an yi nanoindentation don kwatanta taurin dandali da kuma LOIS akan sikelin micro/nano don yin nazarin kaddarorin inji na tasirin tsarin micro/nano (Hoto). 4B).Zane-zane yana nuna nau'ikan nakasar LOIS saboda kasancewar ƙananan sifofi/nano.An zana lanƙwasa-ƙarfi dangane da sakamakon nanoindentation (Hoto 4C).Hoton shudin shudi yana wakiltar ƙashin ƙasa, wanda ke nuna ɗan lahani kaɗan kawai, kamar yadda mafi girman zurfin shigar 0.26-μm ya gani.A gefe guda, haɓakar sannu-sannu a cikin ƙarfin nanoindentation da ƙaura da aka lura a LOIS (jan lanƙwasa) na iya nuna alamun raguwar kayan aikin injiniya, wanda ya haifar da zurfin nanoindentation na 1.61μm.Wannan shi ne saboda tsarin micro/nano da ke cikin LOIS yana ba da sararin ci gaba mai zurfi don tip na nanoindenter, don haka nakasar ta ya fi na abin da ba shi da tushe.Konsta-Gdoutos et al.(45) ya yi imanin cewa saboda kasancewar nanostructures, nanoindentation da micro/nano roughness suna haifar da ƙananan nanoindentation marasa daidaituwa.Wurin da aka shaded yayi daidai da madaidaicin nakasar nakasar da aka danganta ga nanostructure, yayin da yankin da ba shi da inuwa ana danganta shi da microstructure.Wannan nakasawa na iya lalata microstructure/nanostructure na mai mai riko da mummunan tasiri akan aikin sa na hana lalata.Domin yin nazarin tasirin lalacewa akan LOIS, lalacewar da babu makawa ga sifofin micro/nano an kwaikwayi su a cikin jiki yayin tiyatar filastik.Ta hanyar yin amfani da gwaje-gwajen adhesion na jini da furotin, ana iya ƙayyade kwanciyar hankali na anti-biofouling Properties na LOIS bayan in vitro (Figure 4D).Jerin hotuna na gani yana nuna barnar da ta faru a kusa da ramukan kowane yanki.An yi gwajin mannewa na jini don nuna tasirin lalacewar injiniya a kan murfin anti-biofouling (Hoto 4E).Kamar SHP, abubuwan da aka lalata sun ɓace saboda lalacewa, kuma LOIS yana nuna kyawawan abubuwan da ba su da kyau ta hanyar kawar da jini.Wannan shi ne saboda, saboda makamashin saman yana motsa shi ta hanyar aikin capillary wanda ke rufe yankin da ya lalace, yawo a cikin man shafawa na microstructured yana mayar da kaddarorin kariya (35).An lura da irin wannan yanayin a gwajin mannewar furotin ta amfani da albumin.A cikin yankin da aka lalace, an lura da mannewar sunadaran a saman SHP, kuma ta hanyar auna ɗaukar hoto, ana iya ƙididdige shi a matsayin rabin matakin mannewa na abin da ba shi da tushe.A gefe guda, LOIS ya kiyaye kaddarorin anti-biofouling ba tare da haifar da mannewa ba (Hoto 4, F da G).Bugu da kari, saman dunƙule sau da yawa yana fuskantar matsalolin injiniya mai ƙarfi, kamar hakowa, don haka mun yi nazarin ikon murfin LOIS don ci gaba da kasancewa a kan dunƙule in vitro.Hoto 4H yana nuna hotunan gani na sukurori daban-daban, gami da danda, SHP da LOIS.Jan rectangle yana wakiltar yankin da aka yi niyya inda tsananin damuwa na inji ke faruwa yayin dashen kashi.Hakazalika da gwajin mannewa na furotin na farantin, ana amfani da microscope mai kyalli don hoton mannewar furotin da kuma auna wurin ɗaukar hoto don tabbatar da amincin murfin LOIS, har ma a ƙarƙashin ƙarfin ƙarfin injiniya (Hoto 4, I da J).Screws ɗin da aka yi wa LOIS suna nuna kyakkyawan aikin hana lalata, kuma kusan babu furotin da ke manne da saman.A gefe guda, an lura da mannewar sunadaran a cikin screws da SHP screws, inda yanki na yanki na SHP screws ya kasance kashi ɗaya bisa uku na screws.Bugu da ƙari, ƙwanƙwasa orthopedic da aka yi amfani da shi don gyaran gyare-gyare dole ne ya kasance da ƙarfi da ƙarfi don tsayayya da damuwa da ake amfani da shi a wurin da aka karya, kamar yadda aka nuna a cikin Hoto 4K.Sabili da haka, an yi gwajin lanƙwasawa don sanin tasirin gyare-gyaren sinadarai a kan kaddarorin inji.Bugu da ƙari, ana yin wannan don kula da ƙayyadaddun damuwa daga shigarwa.Aiwatar da ƙarfin injina a tsaye har sai an gama naɗewa daɗaɗɗen kuma an sami madaurin damuwa (Hoto 4L, 1).An kwatanta kaddarori guda biyu da suka haɗa da modules ɗin matasa da ƙarfin sassauƙa tsakanin tsiran alade da LOIS a matsayin masu nuna ƙarfin injin su (Hoto 4L, 2 da 3).Matsakaicin matashi yana nuna ikon abu don jure canje-canjen inji.Matsakaicin Matasa na kowane yanki shine 41.48± 1.01 da 40.06±0.96 GPA, bi da bi;Bambancin da aka lura shine kusan 3.4%.Bugu da ƙari, an bayar da rahoton cewa ƙarfin lanƙwasawa, wanda ke ƙayyade taurin kayan, shine 102.34 ± 1.51 GPa don ƙananan ƙananan da 96.99 ± 0.86 GPa don SHP.Matsakaicin dandali yana da kusan 5.3% mafi girma.Ƙananan raguwa a cikin kayan aikin injiniya na iya haifar da sakamako mai daraja.A cikin sakamako mai daraja, ƙananan micro/nano roughness na iya yin aiki azaman saiti na ƙima, yana haifar da ƙaddamar da damuwa na gida kuma yana shafar kayan aikin injin da aka saka (46).Duk da haka, dangane da gaskiyar cewa an ba da rahoton taurin kashin cortical ɗan adam yana tsakanin 7.4 da 31.6 GPa, kuma ma'aunin LOIS da aka auna ya wuce na kashin cortical na ɗan adam (47), LOIS ya isa ya goyi bayan karyewar da gabaɗayan sa. na inji kaddarorin suna kadan tasiri a kan surface gyara.
(A) Jadawalin tsari na (1) damuwa na inji da aka yi amfani da shi a cikin kashin baya yayin aiki, da (2) hoton gani na lalacewa na kashin baya.(B) Tsarin tsari na ma'aunin kayan aikin nano-kanikanci ta hanyar nanoindentation da LOIS akan saman da ba kowa.(C) Nanoindentation ƙarfi-maurayi kwana na danda surface da LOIS.(D) Bayan gwaje-gwajen in vitro, kwaikwayi hotunan gani na nau'ikan faranti na orthopedic daban-daban (yankin da aka lalace yana haskakawa tare da jajayen rectangle) don kwaikwayi damuwa na inji da aka haifar yayin aikin.(E) Gwajin mannewar jini da (F) gwajin mannewar furotin na rukunin farantin kasusuwa da suka lalace.(G) Auna kewayon yanki na furotin da ke manne da farantin.(H) Hotunan gani na nau'ikan screws daban-daban na orthopedic bayan gwajin in vitro.(I) Gwajin mannewa sunadaran don nazarin amincin sutura daban-daban.(J) Auna kewayon yanki na furotin da ke manne da dunƙule.(K) An yi nufin motsi na zomo don haifar da ƙayyadaddun damuwa akan kashin da ya karye.(L) (1) Lanƙwasa sakamakon gwaji da hotunan gani kafin da bayan lankwasawa.Bambanci a cikin (2) Matsalolin Matasa da (3) Ƙarfin lankwasawa tsakanin danda babu da SHP.Ana bayyana bayanai azaman ma'ana ± SD (* P <0.05, ** P <0.01, *** P <0.001 da **** P <0.0001).Hoton hoto: Kyomin Chae, Jami'ar Yonsei.
A cikin yanayin asibiti, yawancin hulɗar ƙwayoyin cuta tare da kayan halitta da wuraren rauni suna zuwa daga balagagge, balagagge biofilms (48).Saboda haka, Cibiyar Kula da Cututtuka da Cututtuka ta Amurka ta kiyasta cewa 65% na duk cututtukan ɗan adam suna da alaƙa da biofilms (49).A wannan yanayin, yana da mahimmanci don samar da ƙirar gwaji na in vivo wanda ke ba da daidaiton samuwar biofilm akan saman dasa.Sabili da haka, mun haɓaka samfurin ɓarkewar mata na zomo wanda aka riga an riga an shigar da ƙwanƙwasa orthopedic a cikin dakatarwar kwayan cuta sannan kuma an dasa shi a cikin mata na zomo don yin nazarin abubuwan da ba su da kyau na LOIS a vivo.Saboda muhimman abubuwa guda uku masu zuwa, al’ada ta riga-kafi ne ke jawo kamuwa da cutar bakteriya maimakon alluran dakatarwar da kwayoyin cuta ke yi: (i) Tsarin garkuwar jikin zomayen a dabi’ance ya fi na mutane karfi;sabili da haka, allurar dakatarwar kwayan cuta da ƙwayoyin cuta na planktonic yana yiwuwa Ba shi da wani tasiri akan samuwar biofilms.(Ii) Kwayoyin Planktonic sun fi kamuwa da maganin rigakafi, kuma yawanci ana amfani da maganin rigakafi bayan tiyata;A ƙarshe, (iii) dakatarwar ƙwayoyin cuta na planktonic na iya narkewa da ruwan jikin dabba (50).Ta hanyar al'ada da shuka a cikin dakatarwar kwayan cuta kafin a dasa, za mu iya yin nazari sosai game da illar kamuwa da cutar kwayan cuta da kuma amsawar jikin waje (FBR) akan tsarin warkar da kashi.An yi hadaya da zomaye 4 makonni bayan dasa, saboda osseointegration da muhimmanci ga kashi waraka tsari za a kammala a cikin 4 makonni.Sa'an nan kuma, an cire kayan da aka saka daga zomaye don nazarin ƙasa.Hoto na 5A yana nuna tsarin yaduwar kwayoyin cuta.An shigar da dasa kothopedic mai cutar a cikin jiki.Sakamakon riga-kafin da aka yi a cikin dakatarwar ƙwayoyin cuta, shida daga cikin zomaye shida da aka dasa tare da tsirara sun kamu da cutar, yayin da babu ɗaya daga cikin zomayen da aka dasa da naman da aka yi wa LOIS ɗin da ya kamu da cutar.Kwayoyin cututtuka suna ci gaba a matakai uku, ciki har da girma, balagagge da watsawa (51).Na farko, ƙwayoyin cuta da aka haɗe suna haifuwa kuma suna girma a saman, sannan ƙwayoyin cuta suna yin biofilm lokacin da suka fitar da polymer na waje (EPS), amyloid da DNA na extracellular.Biofilm ba wai kawai yana tsoma baki tare da shigar da maganin rigakafi ba, har ma yana haɓaka tarin ƙwayoyin cuta masu lalata enzymes (kamar β-lactamase) (52).A ƙarshe, biofilm yana yada balagaggun ƙwayoyin cuta a cikin kyallen da ke kewaye.Saboda haka, kamuwa da cuta yana faruwa.Bugu da ƙari, lokacin da jikin waje ya shiga cikin jiki, kamuwa da cuta wanda zai iya haifar da amsawar rigakafi mai karfi zai iya haifar da kumburi mai tsanani, zafi, da rage rigakafi.Hoto na 5B yana ba da bayyani na FBR da aka haifar ta hanyar shigar da kashin baya, maimakon amsawar rigakafi ta hanyar kamuwa da cuta.Tsarin garkuwar jiki yana gane abin da aka saka a matsayin wani baƙon jiki, sannan yana haifar da sel da kyallen takarda don ɗaukar jikin baƙon (53).A farkon kwanakin FBR, an samar da matrix na samar da kayan aiki a saman kayan da aka sanya na orthopedic, wanda ya haifar da adsorption na fibrinogen.Fibrinogen da aka tallata sannan ya samar da hanyar sadarwa mai yawa na fibrin, wanda ke haɓaka haɗewar leukocytes (54).Da zarar an kafa cibiyar sadarwar fibrin, kumburi mai tsanani zai faru saboda kutsawar neutrophils.A cikin wannan mataki, an saki nau'in cytokines iri-iri irin su tumor necrosis factor-α (TNF-α), interleukin-4 (IL-4) da IL-β, kuma monocytes sun fara shiga cikin wurin dasa shuki kuma sun bambanta cikin manyan sel.Shafi (41, 55, 56).Rage FBR ya kasance ƙalubale koyaushe saboda yawan FBR na iya haifar da kumburi mai tsanani da na yau da kullun, wanda zai iya haifar da rikice-rikice masu mutuwa.Don tantance tasirin cututtukan ƙwayoyin cuta a cikin kyallen da ke kewaye da dandali da LOIS, an yi amfani da tabo na hematoxylin da eosin (H&E) da Masson trichrome (MT).Don zomaye da aka dasa tare da ƙananan ƙwayoyin cuta, cututtukan ƙwayoyin cuta masu tsanani sun ci gaba, kuma zane-zane na H&E sun nuna a fili abscesses da necrosis wanda kumburi ya haifar.A gefe guda, maɗaukakiyar ƙaƙƙarfan anti-biofouling surface LOIS yana hana ƙwayar ƙwayar cuta, don haka ba ya nuna alamun kamuwa da cuta kuma yana rage kumburi (Figure 5C).Sakamakon MT tabo ya nuna irin wannan yanayin.Duk da haka, MT tabo kuma ya nuna edema a cikin zomaye da aka dasa tare da LOIS, yana nuna cewa farfadowa yana gab da faruwa (Hoto 5D).Don yin nazarin digiri na amsawar rigakafi, an yi amfani da maganin rigakafi na immunohistochemical (IHC) ta amfani da cytokines TNF-a da IL-6 da suka danganci amsawar rigakafi.An kwatanta tsiraici mara kyau wanda ba a fallasa ga ƙwayoyin cuta da LOIS wanda aka fallasa ga ƙwayoyin cuta amma bai kamu da cutar ba don nazarin tsarin warkarwa a cikin rashin kamuwa da ƙwayoyin cuta.Hoto 5E yana nuna hoton gani na nunin IHC wanda ke bayyana TNF-a.Yankin launin ruwan kasa yana wakiltar amsawar rigakafi, yana nuna cewa amsawar rigakafi a LOIS ya dan ragu.Bugu da ƙari, furucin IL-6 a cikin LOIS ya kasance ƙasa da ƙarancin magana mara kyau na tsirara (Hoto 5F).An ƙididdige bayanin cytokine ta hanyar auna yankin tabon antibody daidai da cytokine (Hoto 5G).Idan aka kwatanta da zomaye da aka fallasa su zuwa ƙananan ƙwayoyin cuta, matakan maganganu na zomaye da aka dasa tare da LOIS sun kasance ƙananan, suna nuna bambanci mai ma'ana.Ragewa a cikin maganganun cytokine yana nuna cewa dogon lokaci, ƙayyadaddun ƙayyadaddun abubuwan lalata na LOIS ba kawai suna da alaƙa da hana cututtukan ƙwayoyin cuta ba, har ma da raguwar FBR, wanda ke haifar da macrophages manne ga substrate (53, 57, 58 ).Sabili da haka, rage yawan amsawar rigakafi saboda kaddarorin ƙauracewa na LOIS na iya magance abubuwan da ke faruwa bayan dasawa, irin su wuce kima na rigakafi bayan tiyatar filastik.
(A) Zane-zane na tsarin samar da biofilm da yaduwa a saman wani ƙwayar ƙwayar cuta mai cutarwa.eDNA, DNA extracellular.(B) Tsarin tsari na amsawar rigakafi bayan shigar da kashin baya.(C) H&E tabo da kuma (D) MT tabo na kewayen kyallen jikin bangon kasusuwa tare da tabbataccen inganci da LOIS.IHC na cytokines masu alaƙa da rigakafi (E) TNF-a da (F) IL-6 sune hotunan tsirara-mara kyau da zomaye da aka dasa na LOIS.(G) Ƙididdigar maganganun cytokine ta ma'aunin ɗaukar hoto (** P <0.01).
An yi nazarin yanayin yanayin LOIS da tasirinsa akan tsarin warkarwa na kashi a cikin vivo ta amfani da hoton bincike [x-ray da micro-computed tomography (CT)] da osteoclast IHC.Hoto 6A yana nuna tsarin warkar da kashi wanda ya ƙunshi matakai daban-daban guda uku: kumburi, gyare-gyare, da gyare-gyare.Lokacin da raunin ya faru, ƙwayoyin kumburi da fibroblasts za su shiga cikin kashin da ya karye kuma su fara girma a cikin nama na jijiyoyin jini.A lokacin gyaran gyare-gyare, ƙaddamar da ƙwayar ƙwayar ƙwayar ƙwayar ƙwayar ƙwayar ƙwayar ƙwayar ƙwayar ƙwayar ƙwayar ƙwayar ƙwayar ƙwayar ƙwayar ƙwayar ƙwayar ƙwayar ƙwayar cuta.Nama na jijiyoyi yana ba da abinci mai gina jiki don samuwar sabon kashi, wanda ake kira callus.Mataki na ƙarshe na tsarin warkaswa na kashi shine mataki na gyare-gyare, wanda girman girman kiran ya rage zuwa girman kashi na al'ada tare da taimakon karuwa a matakin osteoclasts da aka kunna (59).An yi gyare-gyaren nau'i uku (3D) na wurin raguwa ta amfani da micro-CT scans don lura da bambance-bambance a cikin matakin ƙirƙira kira a kowace ƙungiya.Kula da sashin giciye na femur don lura da kauri na kiran da ke kewaye da kashin da ya karye (Hoto 6, B da C).An kuma yi amfani da haskoki na X don bincika wuraren ɓarke ​​​​na duk ƙungiyoyi a kowane mako don lura da tsarin gyaran kashi daban-daban a cikin kowane rukuni (Figure S9).Ana nuna Callus da manyan ƙasusuwa a cikin shuɗi/kore da hauren giwa, bi da bi.Yawancin kyallen takarda masu laushi ana tace su tare da saiti.Tsirara tabbatacce kuma SHP sun tabbatar da samuwar ƙaramin adadin kira a kusa da wurin karyewar.A gefe guda kuma, ƙarancin LOIS da aka fallasa da kuma wurin karyewa suna kewaye da lokacin kira mai kauri.Hotunan Micro-CT sun nuna cewa samuwar callus ya hana kamuwa da cuta da kamuwa da cuta.Wannan shi ne saboda tsarin rigakafi ya ba da fifiko ga warkar da raunin da ya faru ta hanyar kamuwa da cututtuka, maimakon farfadowa na kashi (60).IHC da Tartrate-resistant Acid Phosphatase (TRAP) tabo an yi su don lura da aikin osteoclast da resorption kashi (Figure 6D) (61).'Yan kaɗan da aka kunna osteoclasts masu launin shuɗi ne kawai aka samo su a cikin tabbataccen tsirara da SHP.A gefe guda, an lura da yawancin osteoclasts da aka kunna kusa da tsirara tabbatacce kuma ƙasusuwan da balagagge na LOIS.Wannan al'amari yana nuna cewa a gaban osteoclasts, callus a kusa da wurin karyewar yana fuskantar wani tashin hankali na gyaran fuska (62).An auna ƙarar kashi da yanki na osteoclast na callus don kwatanta matakin ƙirar ƙira a kusa da wurin fashe a duk ƙungiyoyi, don ƙididdige sakamakon micro-CT da sakamakon IHC (Hoto 6E, 1 da 2).Kamar yadda aka sa ran, tsirara tsirara da kuma kiran kira a cikin LOIS sun kasance mafi girma fiye da sauran kungiyoyi, yana nuna cewa ingantaccen gyaran kashi ya faru (63).Hoto S10 yana nuna hoton gani na wurin aikin tiyata, sakamakon MT ɗin tabo na nama da aka tattara a kusa da dunƙule, da kuma sakamakon tabo na TRAP yana ba da haske game da ƙirar dunƙule-kashi.A cikin dandali, an lura da samuwar callus da fibrosis mai ƙarfi, yayin da aka yi wa LOIS magani ya nuna wani wuri mara kyau.Hakazalika, idan aka kwatanta da tsirara, an lura da ƙananan fibrosis a cikin zomaye da aka dasa tare da LOIS, kamar yadda fararen kiban suka nuna.Bugu da ƙari, ƙaƙƙarfan edema (kibiya mai launin shuɗi) za a iya danganta shi da kaddarorin gujewa na LOIS, don haka rage kumburi mai tsanani.Wurin da ba shi da tsayi a kusa da dasawa da rage fibrosis yana nuna cewa tsarin cirewa ya fi sauƙi, wanda yawanci yakan haifar da wasu karaya ko kumburi.An kimanta tsarin warkar da kashi bayan cire dunƙule ta hanyar aikin osteoclast a madaidaicin dunƙule-kashi.Dukansu ƙashi maras tushe da ƙirar ƙirar LOIS sun shafe irin wannan matakan osteoclasts don ƙara warkar da kashi, yana nuna cewa murfin LOIS ba shi da wani mummunan tasiri akan warkar da kashi ko amsawar rigakafi.Don tabbatar da cewa gyaran gyare-gyaren da aka yi a kan LOIS baya tsoma baki tare da tsarin warkar da kashi, an yi amfani da gwajin X-ray don kwatanta maganin kasusuwa na zomaye tare da ions mara kyau da aka fallasa da 6 makonni na shigar da LOIS (Figure 6F).Sakamakon ya nuna cewa idan aka kwatanta da ƙungiyar tsirara marasa lafiya, LOIS ya nuna irin nau'in warkarwa na kashi, kuma babu alamun alamun karaya (layin osteolysis na ci gaba) a cikin ƙungiyoyin biyu.
(A) Tsarin tsari na tsarin warkar da kashi bayan karaya.(B) Bambanci a cikin digiri na samuwar callus na kowane rukunin saman da (C) hoton giciye na wurin fashe.(D) Tsuntsun TRAP don ganin aikin osteoclast da resorption na kashi.Dangane da ayyukan TRAP, an ƙididdige samuwar kiran waje na kasusuwa na cortical da yawa ta hanyar (E) (1) micro-CT da (2) ayyukan osteoclast.(F) Makonni 6 bayan dasawa, Hotunan X-ray na kashin da ya karye na mara kyau da aka fallasa (wanda aka yi hasashe ta jajayen dashed rectangle) da LOIS (haske da shudin dashed rectangle).An yi nazarin ƙididdiga ta hanyar bincike na bambance-bambancen hanya ɗaya (ANOVA).P <0.05.** P <0.01.
A taƙaice, LOIS yana ba da sabon nau'in dabarun kamuwa da cuta na ƙwayoyin cuta da kuma kariya ta kariya don shigar da kasusuwa.Abubuwan da aka kafa na kasusuwa na al'ada tare da aikin SHP suna nuna kaddarorin anti-biofouling na gajeren lokaci, amma ba za su iya kula da kadarorin su na dogon lokaci ba.A superhydrophobicity na substrate tarko iska kumfa tsakanin kwayoyin da substrate, game da shi samar da iska aljihu, game da shi hana kamuwa da cuta.Koyaya, saboda yaduwar iska, ana cire waɗannan aljihunan iska cikin sauƙi.A gefe guda, LOIS ya tabbatar da ikonsa na hana cututtuka masu alaka da biofilm.Sabili da haka, saboda kaddarorin anti-ƙin yarda na Layer mai mai wanda aka allura a cikin shimfidar micro/nano tsarin shimfidar wuri, ana iya hana kumburin kamuwa da cuta.Hanyoyi daban-daban da suka haɗa da ma'aunin SEM, AFM, XPS da CA ana amfani da su don inganta yanayin masana'antar LOIS.Bugu da kari, LOIS kuma za a iya amfani da daban-daban na halitta kayan da aka saba amfani da orthopedic gyara kayan aiki, kamar PLGA, Ti, PE, POM da PPSU.Bayan haka, an gwada LOIS a cikin vitro don tabbatar da kaddarorin anti-biofouling akan kwayoyin cuta da abubuwan halitta masu alaƙa da amsawar rigakafi.Sakamakon ya nuna cewa yana da kyakkyawan sakamako na antibacterial da anti-biofouling idan aka kwatanta da dandali.Bugu da ƙari, LOIS yana nuna ƙarfin injin ko da bayan yin amfani da damuwa na inji, wanda ba zai yuwu a cikin aikin filastik ba.Saboda kaddarorin warkar da kai na mai mai a saman tsarin micro/nano, LOIS ya sami nasarar kiyaye kaddarorin sa na lalata ƙwayoyin halitta.Domin yin nazarin abubuwan da suka dace da ƙwayoyin cuta da ƙwayoyin cuta na LOIS a cikin vivo, an dasa LOIS cikin femur zomo na makonni 4.Ba a ga kamuwa da cutar kwayan cuta a cikin zomaye da aka dasa da LOIS ba.Bugu da ƙari, yin amfani da IHC ya nuna raguwar matakin amsawar rigakafi na gida, yana nuna cewa LOIS ba ya hana tsarin warkar da kashi.LOIS yana nuna kyawawan kaddarorin kawar da ƙwayoyin cuta da rigakafin rigakafi, kuma an tabbatar da su yadda ya kamata don hana samuwar biofilm kafin da kuma lokacin tiyatar orthopedic, musamman don haɗin kashi.Ta yin amfani da ƙirar ƙirar kashi mai narkewa mai narkewa, sakamakon cututtukan bioflm-da ke da alaƙa da warkar da kashin kashi wanda aka sanyaya shi da aka yi nazari sosai.A matsayin nazari na gaba, ana buƙatar sabon samfurin in vivo don nazarin yiwuwar cututtuka bayan dasawa don fahimta sosai da kuma hana cututtukan da ke da alaka da kwayoyin halitta a duk tsarin warkarwa.Bugu da ƙari, osteoinduction har yanzu ƙalubalen da ba a warware shi ba ne a cikin haɗin kai tare da LOIS.Ana buƙatar ƙarin bincike don haɗa zaɓin mannewa na ƙwayoyin osteoinductive ko maganin farfadowa tare da LOIS don shawo kan ƙalubalen.Gabaɗaya, LOIS yana wakiltar ƙwaƙƙwaran ƙwayar ƙwayar ƙwayar ƙwayar cuta tare da ƙarfin injina da kyawawan kaddarorin anti-biofouling, wanda zai iya rage SSI da illolin rigakafi.
Wanke 15mm x 15mm x 1mm 304 SS substrate (Dong Kang M-Tech Co., Korea) a cikin acetone, EtOH da DI ruwa na mintuna 15 don cire gurɓatattun abubuwa.Domin samar da tsarin micro/nano-matakin a saman, ana tsoma shi a cikin 48% zuwa 51% HF bayani (DUKSAN Corp., Koriya ta Kudu) a 50 ° C.Lokacin etching ya bambanta daga minti 0 zuwa 60.Sa'an nan, da etched substrate aka tsabtace da deionized ruwa da kuma sanya a cikin 65% HNO3 (Korea DUKSAN Corp.) bayani a 50 ° C na 30 minutes don samar da wani chromium oxide passivation Layer a saman.Bayan wucewar, ana wanke ma'aunin da ruwa da aka bushe kuma a bushe don samun abin da ke da tsari mai laushi.Bayan haka, an fallasa ma'adinin zuwa plasma oxygen (100 W, 3 minutes), kuma nan da nan an nutsar da shi a cikin wani bayani na 8.88 mM POTS (Sigma-Aldrich, Jamus) a cikin toluene a dakin da zafin jiki na 12 hours.Sa'an nan, da substrate mai rufi da tukwane aka tsabtace da EtOH, da kuma annealed a 150 ° C na 2 hours don samun m POTS SAM.Bayan shafewar SAM, an kafa wani nau'i mai laushi a kan ma'auni ta hanyar yin amfani da lubricant perfluoropolyether (Krytox 101; DuPont, Amurka) tare da nauyin nauyin 20 μm / cm 2. Kafin amfani, tace mai mai ta hanyar 0.2 micron tace.Cire yawan mai ta hanyar karkata a kusurwa 45° na mintuna 15.An yi amfani da hanyar masana'anta iri ɗaya don gyare-gyaren orthopedic da aka yi da 304 SS (kulle farantin karfe da makullin kulle cortical; Dong Kang M-Tech Co., Koriya).An ƙera duk abubuwan da aka sanyawa orthopedic don dacewa da lissafi na femur zomo.
An duba yanayin yanayin da ake ciki na substrate da orthopedic implants ta hanyar fitar da filin SEM (Duba F50, FEI, Amurka) da AFM (XE-100, Park Systems, Koriya ta Kudu).Ana auna tarkacen saman (Ra, Rq) ta hanyar ninka yanki na 20 μm ta 20 μm (n=4).An yi amfani da tsarin XPS (PHI 5000 VersaProbe, ULVAC PHI, Japan) wanda aka sanye shi da tushen X-ray na Al Kα tare da girman tabo na 100μm2 don nazarin abubuwan sinadaran saman.An yi amfani da tsarin ma'aunin CA sanye take da kyamarar ɗaukar hoto mai ƙarfi (SmartDrop, FEMTOBIOMED, ​​Koriya ta Kudu) don auna ruwa CA da SA.Ga kowane ma'auni, 6 zuwa 10 μl na droplets (ruwan da aka lalata, jinin doki, EG, 30% ethanol, da HD) ana sanya su a kan saman don auna CA.Lokacin da kusurwar karkata na substrate ya karu a gudun 2°/s (n = 4), ana auna SA lokacin da ɗigon ruwa ya faɗi.
Pseudomonas aeruginosa [Tsarin Al'adu na Amurka (ATCC) 27853] da MRSA (ATCC 25923) an saya su daga ATCC (Manassas, Virginia, Amurka), kuma an kiyaye al'adun jari a -80 ° C.Kafin amfani, an sanya al'adun daskararre a cikin broth waken soya mai narkar da trypsin (Komed, Koriya) a 37 ° C na awanni 18 sannan a canza shi sau biyu don kunna shi.Bayan shiryawa, al'adun sun kasance a tsakiya a 10,000 rpm na minti 10 a 4 ° C kuma an wanke su sau biyu tare da bayani na PBS (pH 7.3).Sannan al'adar da aka kafa ta tana ƙarƙashin al'adar akan faranti na agar jini (BAP).MRSA da Pseudomonas aeruginosa an shirya su cikin dare kuma an tsara su a cikin ruwan Luria-Bertani.Matsakaicin Pseudomonas aeruginosa da MRSA a cikin inoculum an ƙididdige su ta hanyar CFU na dakatarwa a cikin serial dilutions akan agar.Sa'an nan, daidaita ƙwayar ƙwayar cuta zuwa 0.5 McFarland misali, wanda yayi daidai da 108 CFU/ml.Sa'an nan kuma tsoma dakatarwar kwayar cutar sau 100 zuwa 106 CFU/ml.Don gwada kaddarorin mannewa na ƙwayoyin cuta, an haifuwa substrate a 121 ° C na mintuna 15 kafin amfani.Daga nan aka canja wurin substrate zuwa 25 ml na dakatarwar kwayan cuta kuma an sanya shi a 37 ° C tare da girgiza mai ƙarfi (200 rpm) na awanni 12 da 72.Bayan shiryawa, an cire kowace ƙasa daga cikin incubator kuma an wanke sau 3 tare da PBS don cire duk wani ƙwayar cuta da ke iyo a saman.Domin lura da biofilm a kan substrate, an gyara biofilm tare da methanol kuma an lalata shi da 1 ml na crimidine orange na minti 2.Sa'an nan kuma an yi amfani da microscope mai haske (BX51TR, Olympus, Japan) don ɗaukar hotuna na tabo na biofilm.Don ƙididdige biofilm akan substrate, ƙwayoyin da aka haɗe sun rabu da su ta hanyar hanyar vortex na bead, wanda aka ɗauka shine hanya mafi dacewa don cire ƙwayoyin cuta (n = 4).Yin amfani da bakararre bakararre, cire madaidaicin daga matsakaicin girma kuma danna farantin rijiyar don cire ruwa mai yawa.An cire ƙwayoyin da aka haɗe da su ta hanyar wankewa sau biyu tare da PBS mara kyau.Daga nan sai aka canjawa kowane sifari zuwa bututun gwajin da bakararre mai dauke da 9 ml na 0.1% protein ept saline (PSW) da 2 g na beads na gilashin bakararre 20 zuwa 25 (0.4 zuwa 0.5 mm a diamita).Daga nan sai aka juyar da shi na tsawon mintuna 3 don cire sel daga samfurin.Bayan vortexing, an narkar da dakatarwar a jere sau 10 tare da 0.1% PSW, sa'an nan kuma an shafe 0.1 ml na kowane dilution akan BAP.Bayan sa'o'i 24 na shiryawa a 37 ° C, an ƙidaya CFU da hannu.
Don sel, an yi amfani da fibroblasts na linzamin kwamfuta NIH/3T3 (CRL-1658; American ATCC) da linzamin kwamfuta macrophages RAW 264.7 (TIB-71; American ATCC).Yi amfani da Dulbecco's modified Eagle matsakaici (DMEM; LM001-05, Welgene, Korea) zuwa al'ada linzamin kwamfuta fibroblasts da kari tare da 10% maraƙi serum (S103-01, Welgene) da 1% penicillin-streptomycin (PS; LS202-02, Welgene (Welgene) ). An shigar da kwayoyin halitta a cikin dare a 37 ° C da 5% CO2 Don lalata tantanin halitta, an gyara sel tare da 4% paraformaldehyde na minti 20 kuma an sanya su a cikin 0.5% Triton X Incubate na minti 5 a cikin -100 a 37 ° C na minti 30 Bayan tsarin shiryawa, yi amfani da substrate tare da 4′,6-diamino-2-phenylindole (H -1200, Vector Laboratories, UK) VECTASHIELD matsakaicin daidaitawa (n = 4 da tantanin halitta). , fluorescein, fluorescein isothiocyanate-albumin (A9771, Sigma-Aldrich, Jamus) da kuma plasma na mutum An narkar da Alexa Fluor 488-conjugated fibrinogen (F13191, Invitrogen, Amurka) a cikin PBS (10 mM, pH 7.4).Yawan adadin albumin da fibrinogen sun kasance 1 da 150 μg/ml, bi da bi.Bayan substrate Kafin a nutse cikin maganin furotin, kurkura su da PBS don sake sanya ruwa a saman.Sa'an nan kuma a nutsar da duk abubuwan da ake amfani da su a cikin farantin rijiyar shida mai dauke da maganin furotin kuma a sanya shi a 37 ° C na minti 30 da 90.Bayan shiryawa, an cire substrate daga maganin furotin, a wanke a hankali tare da PBS sau 3, kuma an gyara shi tare da 4% paraformaldehyde (n = 4 ga kowane furotin).Don alli, sodium chloride (0.21 M) da potassium phosphate (3.77 mM) An narkar da su a cikin ruwa mai narkewa.An daidaita pH na maganin zuwa 2.0 ta hanyar ƙara maganin hydrochloride (1M).Sannan an narkar da calcium chloride (5.62 mM) a cikin maganin.Ta ƙara 1M tris (hydroxymethyl) -amino Methane yana daidaita pH na maganin zuwa 7.4.A nutsar da duk abubuwan da ake amfani da su a cikin farantin rijiyar shida mai cike da 1.5 × calcium phosphate bayani kuma cire daga maganin bayan minti 30.Don tabo, 2 g Alizarin Red S (CI 58005) Mix tare da 100 ml na ruwa mai narkewa.Sa'an nan, yi amfani da 10% ammonium hydroxide don daidaita pH zuwa 4. Rina da substrate da Alizarin Red bayani na 5 minutes, sa'an nan kuma girgiza da wuce haddi rini da goge.Bayan aiwatar da girgiza, cire substrate.Abun yana bushewa, sa'an nan kuma a nutsar da shi a cikin acetone na minti 5, sa'an nan kuma a nutsar da shi a cikin maganin acetone-xylene (1: 1) na minti 5, kuma a karshe an wanke shi da xylene (n = 4).Ana amfani da microscope na Fluorescence (Axio Imager) tare da ×10 da ×20 tabarau na haƙiƙa..A2m, Zeiss, Jamus) hotuna duk abubuwan da aka gyara.An yi amfani da HotunaJ/FIJI (https://imagej.nih.gov/ij/) don ƙididdige bayanan mannewa na abubuwan halitta akan kowane rukuni na wurare huɗu daban-daban na hoto.Mayar da duk hotuna zuwa hotunan binaryar tare da kafaffen ƙofa don kwatanta ƙasa.
An yi amfani da na'urar microscope na Zeiss LSM 700 don saka idanu da kwanciyar hankali na Layer mai mai a cikin PBS a cikin yanayin tunani.Samfurin gilashin SAM mai rufi na tushen fluorine tare da allurar mai mai allura an nutsar da shi a cikin maganin PBS, kuma an gwada shi ta amfani da shaker orbital (SHO-1D; Daihan Scientific, Koriya ta Kudu) a ƙarƙashin yanayin girgiza mai laushi (120 rpm).Sannan ɗauki samfurin kuma saka idanu akan asarar mai ta hanyar auna asarar haske mai haske.Don samun hotunan kyalli a cikin yanayin tunani, ana nuna samfurin zuwa Laser na 633 nm sannan kuma a tattara shi, saboda hasken zai nuna baya daga samfurin.An auna samfuran a cikin tazarar lokaci na 0, 30, 60, da 120 hours.
Don sanin tasirin tsarin gyare-gyaren saman akan abubuwan da ke cikin nanoindenter (TI 950 TriboIndenter, Hysitron, Amurka) an yi amfani da tip lu'u-lu'u mai siffar pyramid mai gefe uku don auna nanoindenedione.Mafi girman nauyin shine 10mN kuma yankin shine 100μmx 100μm.Don duk ma'auni, lokacin lodi da saukewa shine s 10, kuma lokacin riƙewa a ƙarƙashin ƙyalli mafi girma shine 2 s.Ɗauki ma'auni daga wurare daban-daban guda biyar kuma ɗauki matsakaicin matsakaici.Don kimanta aikin ƙarfin injina ƙarƙashin kaya, an yi gwajin lanƙwasawa mai lamba uku ta amfani da injin gwaji na duniya (Instron 5966, Instron, Amurka).Ana matsawa da ma'auni a matsakaicin ƙimar 10 N / s tare da ƙarin kaya.An yi amfani da shirin software na Bluehill Universal (n = 3) don ƙididdige ma'auni mai sauƙi da matsakaicin matsawa.
Don yin kwatankwacin tsarin aiki da lalacewar injin da ke da alaƙa da aka haifar yayin aikin, an aiwatar da aikin a cikin vitro.An tattara matan ne daga fararen zomayen New Zealand da aka kashe.An tsaftace femur kuma an gyara shi a cikin 4% paraformaldehyde na mako 1.Kamar yadda aka bayyana a cikin hanyar gwajin dabba, an yi wa kafaffen femur tiyata.Bayan aikin, an yi amfani da ƙwayar orthopedic a cikin jini (jinin doki, KISAN, Koriya) don 10 s don tabbatar da ko haɗin jini ya faru bayan an yi amfani da raunin injiniya (n = 3).
Jimlar 24 na maza na New Zealand fararen zomaye (nauyin 3.0 zuwa 3.5kg, matsakaicin shekaru 6 watanni) an raba su ba tare da izini ba zuwa kungiyoyi hudu: tsiraici mara kyau, tsirara tsirara, SHP da LOIS.Dukkan hanyoyin da suka shafi dabbobi an yi su daidai da ka'idodin da'a na Cibiyar Kula da Dabbobi da Amfani (IACUC ta amince, KOREA-2017-0159).Ƙaƙwalwar ƙwayar ƙwayar cuta ta ƙunshi farantin kulle tare da ramuka biyar (tsawon 41 mm, nisa 7 mm da kauri 2 mm) da kuma makullin kulle cortical (tsawon 12 mm, diamita 2.7 mm) don gyaran karaya.Ban da waɗancan faranti da sukurori da aka yi amfani da su a cikin rukunin mara kyau, duk faranti da sukurori an sanya su a cikin dakatarwar MRSA (106 CFU/ml) na awanni 12.Ƙungiyar tsirara (n=6) an bi da ita tare da tsirara tsirara ba tare da fallasa ga dakatarwar kwayan cuta ba, a matsayin mummunan iko ga kamuwa da cuta.Ƙungiya mai kyau (n = 6) an bi da ita tare da dasa shuki da aka fallasa ga ƙwayoyin cuta a matsayin ingantaccen iko don kamuwa da cuta.Ƙungiyar SHP (n = 6) an bi da ita tare da shigar da SHP da aka fallasa ta kwayoyin cuta.A ƙarshe, ƙungiyar LOIS an bi da su tare da ƙwayoyin LOIS da aka fallasa da ƙwayoyin cuta (n = 6).Ana ajiye dukan dabbobi a cikin keji, kuma ana ba da abinci da ruwa da yawa.Kafin aikin, an yi azumin zomaye na sa'o'i 12.An kwantar da dabbobin ta hanyar allurar intramuscular na xylazine (5mg/kg) da kuma allurar paclitaxel (3mg/kg) na ciki don shigar da su.Bayan haka, isar da 2% isoflurane da 50% zuwa 70% oxygen oxygen (yawan kwarara 2 L / min) ta hanyar tsarin numfashi don kula da maganin sa barci.An dasa shi ta hanyar kai tsaye zuwa ga femur na gefe.Bayan cire gashi da kuma lalata fata na povidone-iodine, an yi wani yanki mai tsayi kimanin 6 cm a waje na tsakiyar femur na hagu.Ta hanyar buɗe rata tsakanin tsokoki da ke rufe femur, femur yana bayyana cikakke.Sanya farantin a gaban shingen femoral kuma gyara shi tare da sukurori hudu.Bayan gyarawa, yi amfani da igiyar gani (kauri 1 mm) don ƙirƙirar karaya ta hanyar wucin gadi a cikin yanki tsakanin rami na biyu da rami na huɗu.A karshen aikin, an wanke raunin da gishiri kuma an rufe shi da sutures.An yi wa kowane zomo allurar subcutaneously tare da enrofloxacin (5 mg/kg) an diluted daya bisa uku a cikin saline.An dauki X-ray bayan tiyata na femur a cikin dukkan dabbobi (0, 7, 14, 21, 28, da 42 days) don tabbatar da osteotomy na kashi.Bayan maganin sa barci mai zurfi, KCl (2 mmol/kg) na cikin jini ya kashe duk dabbobi a cikin kwanaki 28 da 42.Bayan aiwatar da kisa, an duba femur ta hanyar micro-CT don lura da kwatanta tsarin warkar da kashi da sabon kasusuwa tsakanin kungiyoyi hudu.
Bayan aiwatar da kisa, an tattara nama mai laushi waɗanda ke cikin hulɗar kai tsaye tare da ƙwanƙwasa orthopedic.An gyara nama a cikin 10% tsaka tsaki buffered formalin na dare sannan ya bushe a cikin EtOH.An saka naman da aka bushe a cikin paraffin kuma an raba shi a kauri na 40 μm ta amfani da microtome (400CS; EXAKT, Jamus).Domin ganin kamuwa da cutar, an yi tabon H&E da MT.Don duba amsawar mai watsa shiri, an haɗa nau'in nau'in nama tare da zomo anti-TNF-α farko antibody (AB6671, Abcam, Amurka) da zomo anti-IL-6 (AB6672; Abcam, Amurka), sa'an nan kuma bi da tare da horseradish.Oxidase.Aiwatar da tsarin tabo na avidin-biotin (ABC) zuwa sassan bisa ga umarnin masana'anta.Domin ya bayyana azaman samfurin amsawar launin ruwan kasa, 3,3-diaminobenzidine an yi amfani dashi a duk sassa.An yi amfani da na'urar daukar hotan takardu ta dijital (Pannoramic 250 Flash III, 3DHISTECH, Hungary) don hange dukkan yanki, kuma software na ImageJ ta tantance aƙalla sassa huɗu a cikin kowace ƙungiya.
An ɗauki hotunan X-ray a cikin duk dabbobi bayan tiyata da kowane mako don saka idanu akan warkar da karaya (n=6 kowace rukuni).Bayan aiwatar da kisa, an yi amfani da ƙananan micro-CT don ƙididdige samuwar kira a kusa da femur bayan warkarwa.An tsabtace femur da aka samu, an gyara shi a cikin 4% paraformaldehyde na kwanaki 3, kuma ya bushe a cikin 75% ethanol.Sannan an duba ƙasusuwan da ba su da ruwa ta hanyar amfani da micro-CT (SkyScan 1173, Brooke Micro-CT, Kandy, Belgium) don samar da hotunan voxel na 3D (2240 ​​× 2240 pixels) na samfurin kashi.Yi amfani da 1.0 mm Al tace don rage hayaniyar sigina kuma amfani da babban ƙuduri ga duk sikanin (E = 133 kVp, I = 60 μA, lokacin haɗin kai = 500 ms).An yi amfani da software na Nrecon (version 1.6.9.8, Bruker microCT, Kontich, Belgium) don samar da ƙarar 3D na samfurin da aka bincika daga tsinkayar 2D da aka samu.Don bincike, hoton da aka sake ginawa na 3D yana kasu kashi 10mm × 10mm × 10mm cubes bisa ga wurin karyewa.Yi lissafin kira a waje da kashin cortical.DataViewer (version 1.5.1.2; Bruker microCT, Kontich, Belgium) software da aka yi amfani da lambobi a tura da scanning girma na kashi, da kuma CT-Analyzer (version 1.14.4.1; Bruker microCT, Kontich, Belgium) software da aka yi amfani da bincike.Ƙwayoyin shayarwar x-ray na dangi a cikin balagagge kashi da callus ana bambanta su da yawa, sa'an nan kuma an ƙididdige ƙarar kira (n = 4).Don tabbatar da cewa daidaituwa na LOIS ba ya jinkirta tsarin warkar da kashi, an yi ƙarin nazarin X-ray da micro-CT a cikin zomaye guda biyu: tsirara-rauni da ƙungiyoyin LOIS.An kashe kungiyoyin biyu ne a mako na 6.
An tattara femurs daga dabbobin hadaya kuma an gyara su a cikin 4% paraformaldehyde na kwanaki 3.Sa'an nan kuma an cire kasusuwa a hankali daga femur.An lalata mata na tsawon kwanaki 21 ta hanyar amfani da 0.5 M EDTA (EC-900, National Diagnostics Corporation).Daga nan sai aka nitse femur a cikin EtOH don ya bushe.An cire femur da ba shi da ruwa a cikin xylene kuma an saka shi a cikin paraffin.Sa'an nan samfurin da aka yanka tare da atomatik rotary microtome (Leica RM2255, Leica Biosystems, Jamus) tare da kauri na 3 μm.Don tabon TRAP (F6760, Sigma-Aldrich, Jamus), samfuran sassan da aka raba an cire su, an sake su kuma an sanya su a cikin TRAP reagent a 37 ° C na awa 1.An samo hotuna ta amfani da na'urar daukar hotan takardu (Pannoramic 250 Flash III, 3DHISTECH, Hungary) kuma an ƙididdige su ta hanyar auna yanayin yanki na tabo.A cikin kowane gwaji, aƙalla nau'i-nau'i huɗu a cikin kowace ƙungiya an bincika ta software na ImageJ.
An yi nazarin mahimmancin ƙididdiga ta amfani da GraphPad Prism (GraphPad Software Inc., Amurka).An yi amfani da gwajin t-test da ba a haɗa su ba da kuma nazarin bambance-bambancen hanya ɗaya (ANOVA) don gwada bambance-bambance tsakanin ƙungiyoyin kimantawa.An nuna matakin mahimmanci a cikin adadi kamar haka: * P <0.05, ** P <0.01, *** P<0.001 da ****P <0.0001;NS, babu bambanci mai mahimmanci.
Don ƙarin kayan don wannan labarin, da fatan za a duba http://advances.sciencemag.org/cgi/content/full/6/44/eabb0025/DC1
Wannan labarin buɗaɗɗen shiga ne wanda aka rarraba ƙarƙashin sharuɗɗan Lasisin Halayen Haɗin Kai-Ba na Kasuwanci ba, wanda ke ba da damar amfani, rarrabawa da haifuwa a kowane matsakaici, muddin amfani ba don riba ta kasuwanci bane kuma jigo shine asalin. aiki yayi daidai .Magana.
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