[生物醫(yī)藥論文精品a]骨形成蛋白微球溫敏性凝膠復(fù)合系統(tǒng)的制備及釋藥特性研究

1骨形成蛋白微球溫敏性凝膠復(fù)合系統(tǒng)的制備及釋藥特性研究A1A0A2A3A4A5A6A4A7A8A4A9A10A11A12A13A14A15A16A17A18A19A20A21A22A23A24A25A17A18A25A26A27A28A28A29A28A30A31A32A33A34A35A36A37A38A39A40A41A42A43A44A45A46A47A48A49A50A51A52A53A54A55（RHBMP2）A56A57A58A59A60A61A62A63A25A64A65A66A67A68A69A56A57A40A67A68A69A56A57A70A71A72A73A74A75A76A77A78A61A79A80A58A22A81A72A82A83A84A85A86A87A88A89A88A87A90A91A92A93A23A94A95A96A97A98A91酸乙醇酸共聚物（PLGA）為材料A41A42A99A53A56A57A25A100A101A102A103A96A41A42RHBMP2A104A22A56A57A82A97A105A106A107A98A108A109A110A54A111A107A108A112A113A98A114A115MPEGPLGAA30A116A117A118A41A42A73A74A25A85A86A119A120A96A121A122A123A124A102A103A96A125A126A58A22A127A60A82A128A129微球呈現(xiàn)三相釋藥模式，而微球凝膠劑體外釋藥曲線符合HIGUCHI方程。采用PLGACOOH制備的RHBMP2微球，其突釋明顯小于以PLGA為材料制備的微球，增加內(nèi)水相與油相的體積比、在內(nèi)水相中加入氯化鈉均可以在一定程度上減小突釋。A128A130BMP微球溫敏性凝膠復(fù)合系統(tǒng)制備工藝，具有比RHBMP2微球劑更理想的體外釋藥特性。A131A132A133A131A132A133A134A134A46A47A48A49A50A51A52A53A54A55A135A98A91酸乙醇酸共聚物；A105A106A107A98A108A109A110A54A111A107A108A112A113A98A114A135A56A57A136A70A71A72A73A74A136A137A58PREPARATIONOFRECOMBINANTHUMANBONEMORPHOGENETICPROTEIN2LOADEDMICROSPHERES/THERMOSENSITIVEHYDROGELSCOMPLEXANDITSINVITRORELEASECHARACTERISTICFUXUDONG,LIUHONG，TANGREN,ZHANGMENGYOU（WUHANGENERALHOSPITALOFGUANZHOUCOMMAND,WUHAN430030,CHINA）ABSTRACTOBJECTIVETOEXAMINETHEEFFECTSOFFORMULATIONFACTORSONINVITRORELEASERATEOFRECOMBINANTHUMANBONEMORPHOGENETICPROTEIN2RHBMP2LOADEDMICROSPHERESANDCOMPAREDRUGRELEASEPROFILESOFMICROSPHERESANDMICROSPHERES/THERMOSENSITIVEHYDROGELSCOMPLEXMETHODSBLANKMICROSPHERESWEREPREPAREDWITHW/O/WEMULSIONSOLVENTEVAPORATIONMETHODRHBMP2LOADEDMICROSPHERESWASPREPAREDWITHABSORPTIONMETHODTHERMOSENSITIVEHYDROGELSWEREPREPAREDWITHMPEGPLGAENZYMELINKEDIMMUNOSORBENTASSAYWASUSEDTO1A138A139A140A141A142A143A144A145A146A147A148A149A150A151A152A153A154A154A155A156A156A157A154A158A159A157A160A161A162A163A164A165A142A166A167A168A169A170A169A171A172A173A174A175TEL/FAX02787649309EMAILFUXUDONG2005YAHOOCOMCN2DETERMINEDRUGCONTENTANDINVITRORELEASERATERESULTSTHEPROFILESOFRHBMP2MICROSPHERESWERECHARACTERIZEDBYTHREEPHASE,WHILETHEINVITRORELEASEPROFILEOFRHBMP2MICROSPHERES/THERMOSENSITIVEHYDROGELSCOMPLEXCONFORMEDTOHIGUCHIEQUATIONTHEAMOUNTOFRHBMP2RELEASEDFROMMICROSPHERESINTHEINITIALBURSTPHASEWASDECRESEDMARKEDLYWHENPLGACOOHWASUSEDTOPREPAREMICROSPHERESINSTEADOFPLGAINCREASINGTHEVOLUMERATIOOFINTERNALAQUEOUSANDADDINGNACLTOINTERNALAQUEOUSPHASECOULDREDUCEINITIALRELEASECONCLUSIONRHBMP2LOADEDMICROSPHERES/THERMOSENSITIVEHYDROGELSCOMPLEXEXERTEDADESIRABLERELEASECHARACTERISTICKEYWORDSRECOMBINANTHUMANBONEMORPHOGENETICPROTEIN2PLGAMPEGPLGATHERMOSENSITIVEHYDROGELSMICROSPHERE；INITIALBURSTRELEASE牙周病是世界衛(wèi)生組織列為重點(diǎn)防治的、人類普遍罹患的口腔疾病之一，是成年人失牙的主要原因，其治療的最終目標(biāo)是實(shí)現(xiàn)牙周組織包括牙槽骨、牙骨質(zhì)和牙周膜的完全功能性再生，而牙周組織再生的基礎(chǔ)是牙周膜細(xì)胞（PDLCS），但在牙周損傷部位PDLCS的來(lái)源非常有限，以致牙周組織很難達(dá)到有效地再生和重建。骨形成蛋白（BONEMORPHOGENETICPROTEIN,BMP）是目前發(fā)現(xiàn)的唯一能在體內(nèi)誘導(dǎo)骨與軟骨的細(xì)胞因子，業(yè)已證實(shí)BMP能夠促進(jìn)PDLCG86G3698G8554G2462牙周組織的G1474G38091，但是BMP在體內(nèi)G9213G5242和G18250的G1328G11004G991非常G4493G7143G1207G16886失G8975，限G2058G1114其發(fā)G4649和G5224G11004。G4559G6226G17878G4464的G14659G10301G17894G17877G13007G13491一G11464是BMP的G11752G12362G9921點(diǎn)之一，以生G10301G2499G19489G16311G7460G7021G2058G3803的G5506G10711G6122G13447G12871G12902G2058G2070G1328為BMP牙周G15967G4628部治療的G17745體G2475到G11752G12362G13785的G19750G11556，但是G17837G12193G13485G14659G13007G13491體G3818G18334G14659G7366G13459G5194G993是非常G10714G5831，G15932現(xiàn)G1998一G4462的G12373G18334G6122G993G16280G2029的G989G11468G18334G14659G8181G534723，而G1000G5506G12902G2058G2070G11004G1122牙周G15967G4628部G13485G14659G17836G4396在G1363G11004G993G1427G12573G13582點(diǎn)。G12520G13785以G13870G1095G18252G1069G18267G18252G1861G13870G10301為G7460G7021G2058G3803BMPG5506G10711，G17902G17819G3800G7053和G5049G14414的G1260G2282在一G4462G12255G5242G990G2499G19489G1314G12373G18334；G4570G2058G3803的G5506G10711G2010G6967在以G13870G1069G1120G18267G2345G11014G18286/G13870G1095G18252G1069G18267G18252G1861G13870G10301（MPEGPLGA）為G7460G7021G2058G3803的G1969G14026G1025，BMP在G5506G10711G1969G14026G3809G2524G13007G13491G1025的G18334G14659G17907G5242G5483到進(jìn)一G8505G6925G2904。MPEGPLGAG1867有G9213G6947性，一G4462G8999G5242的MPEGPLGAG8712G9354G9094在G4472G9213G991為G14270G11013流動(dòng)的G9094態(tài)G2499以G7053G1427醫(yī)生G11004G14659，在體G9213G991G2499發(fā)生G14026G1969而起到控G2058G14659G10301G18334放的G1328G11004。本文主要報(bào)道G1114G3800G7053和G2058G3803G5049G14414對(duì)BMPG5506G10711以G2462BMPG5506G10711G1969G14026G2070體G3818G18334G14659G7366G13459的影響。31材料和儀器骨形成蛋白（北京環(huán)亞泰克生G10301醫(yī)學(xué)技術(shù)有限公司）；G13870（G1095G18252G1069G18267G18252）G1861G13870G1030150/50（PLGA，G1095G18252G1069G18267G18252比例為5050，G2010子量為10000，山東省醫(yī)療器械G11752G12362所）；端羧基G13870G1095G18252羥基G1069G18252G1861G13870G10301（PLGACOOH,G1095G18252羥基G1069G18252比例為5050,G2010子量為10000，山東省醫(yī)療器械G11752G12362所），G13870G1069烯G18267（PVA124，日產(chǎn)進(jìn)口G2010裝，北京G2282學(xué)試G2070公司）；MPEGPLGALAGAEO的比例是821，G2010子量40005000，PEGG2010子量1000，濟(jì)南岱罡生G10301技術(shù)有限公司；BMP2ELISA試G2070盒（武漢博士德試G2070公司）。數(shù)顯G5347電動(dòng)攪拌器（G990海司樂(lè)儀器有限公司）；JY9202D超聲波細(xì)胞粉碎機(jī)（寧波新芝生G10301科技股份有限公司）；SK1快G17907混勻器（常州國(guó)華電器有限公司）；UV260型紫G3818G2010光光G5242計(jì)（日本島津公司）；SHZ82恒G9213振蕩器（常州國(guó)華電器有限公司）；BX50光學(xué)顯G5506鏡（日本OLYMPUS公司）；S450掃描電鏡（日本HITACHI公司）；ALPHA14冷凍干燥機(jī)（CHRIST,德國(guó)）。2實(shí)驗(yàn)方法21BMP微球的制備空白G5506G10711采G11004W/0/WG1095G2282G9354G2070揮發(fā)法G2058G3803，取G17878量G8712加入到2ML含有200MGPLGA的G1120氯G11014烷G1025，冰浴條G1226G991超聲G220G86，G9994G2530G4570其G1510入15MLG22G705的PVAG9354G9094G1025，冰浴條G1226G991G2500RPM攪拌10MING2530，G4570G17728G17907G19489G14279G2200RPM加入G225MLG8712，在G4472G9213G991G13499G13505攪拌4H，G9994G2530G17819G9400，G8939G9080，G11507空G1128G8699G2282G1120G11979干燥4G27H。BMPG5506G10711G17902G17819G2572G19480法G17745G14659，G457010MG空白G5506G10711混G5760在100L含50GML1的BMP25G705G10988G8700G18252G13543G1926G9094G1025，4G263G991G2572G1948024HG2530冷凍干燥。22BMP微球凝膠的制備G12228取MPEGPLGA4GG13634G112220MLG8712G1025，攪拌G2530G19757G13634G11224G263冰G12677G1025，G5194G7114G7114振蕩，G5465MPEGPLGAG1817G2010G9354G13972G2530，G2375G548320G705MPEGPLGAG8712G1969G14026。取10MGG17745G14659G5506G10711，加入25MLG1969G14026，在冰浴條G1226G991G1817G2010混勻，G2375G5483含BMP2GML1的G5506G10711G1969G14026。23BMP微球及微球凝膠劑體外釋放度的測(cè)定4G4570G17745有5G22037GBMP的G5506G10711G2010G2047混G7071在25ML的01MOLL1PH74的G11979G18252G11428G13543G1926G9094、10G705和20G705的MPEGPLGEG8712G9354G9094G1025，G12946G4506量取01MLG2010G2047G13634G11221ML的G121754G5527G12661G1025，加入1MLPH74的G11979G18252G11428G13543G1926G9094，G13634G1122恒G9213振蕩器G1025，振蕩G17907G5242為100RG22109MIN1、G9213G5242為（37G1021）G263，G2010G2047在05H、1、3、5、7、14、21和28D取G7691，2000RPMG12175G552710MING2530，G5335G6493G990G9177G3824，G8797G9108G10301G1025加入1MLPBS，超聲15MIN，2000RPMG12175G552710MING2530，取G990G9177G90940LMLG6365110比例G12244G18334G2530，采G11004G3853G5527法G18250G13864G1825G11135G2572G19480法G8991G4462G7691G2709G1025G8543G11053的G14659G10301，G17902G17819標(biāo)G1946G7366G13459計(jì)G12651BMP的含量，G16825G7053法的G7828G8991G14551G32726254000PG/ML，G8611份G7691G2709G5191G15904G8991G44622G8437。G18334放G5242為G18334放前G2530G13007G13491G1025的G14659G10301含量之G5058與G18334放前G13007G13491的G14659G10301含量的G11346G2010比。24微球粒徑及表面形態(tài)觀察G11004光學(xué)顯G5506鏡G2462掃描電鏡G16278G4531G5506G10711的G3818G16278形態(tài)，G8991G4462G993G4581G1122200G1022的G12902G5464，計(jì)G12651G12651術(shù)G5191G3355G12902G5464。3結(jié)果與討論31內(nèi)水相與油相的體積比對(duì)BMP微球初始釋藥行為的影響以PLGA50/50為G7460G7021，G13783G4531G5415內(nèi)G8712G11468與G8845G11468的體G12227比G2010G2047為15、110和120G7114G5506G10711的形態(tài)和G18334G14659G15904為。G3926G32821所G12046，G19555G11540內(nèi)G8712G11468與G8845G11468的體G12227比的G3698加，G5506G10711的G12902G5464G8821有G7138顯的G2476G2282（G3835G13434在20G794G220M），但是G5506G10711G15932G19766的G4392G8946G3698加5，G5506G10711在0G175HG794G22G71的G18334放G5242G19489G1314。G17837G2499能因?yàn)锽MP是G17902G17819G2572G19480G1328G11004G17745G14659的，G19555G11540內(nèi)G8712G11468體G12227G11468對(duì)G1955G4581，G5506G10711內(nèi)部和G15932G19766的G4392G8946G1955G4581（G16277G32822），G5506G10711與BMPG9354G9094G6521G16314G19766G12227G1955G4581，BMPG15999G2572G19480的量G1955G4581，G8821有G15999G2572G19480的BMP在G18334G14659G2033G7411很快G6205G6967到G18334放G1183質(zhì)，G5506G10711的G12373G18334G3698加。A176A177A176A178A176A179A176A180A176A181A182A183A184A185A186A184A187A188A189A190A191A192A193A194A195A196A197A198A199A200A201A199A202A200A203A197A198A199A200A201A199A202A200A202A204A197A198A199A200A201A199A202A200A205A204圖1內(nèi)水相與油相的體積比對(duì)BMP微球072H釋藥行為的影響5圖2內(nèi)水相與油相的體積比對(duì)BMP微球形態(tài)的影響A206A207A208A209A210A208A211A212A213A214A215A216A217A218A216A216A219A220A2162A21932內(nèi)水相中氯化鈉含量對(duì)BMP微球初始釋藥行為的影響G4570內(nèi)G8712G11468與G8845G11468的體G12227比G3278G4462在15，G1100410G705的氯G2282G19060G9354G9094G7379G1207G8712G1328為內(nèi)G8712G11468，G2499以發(fā)現(xiàn)G5506G10711G15932G19766的G4392G8946G7138顯G3698加（G3926G3282G22所G12046），因?yàn)镚19555G11540內(nèi)G8712G11468G9195G17891G2399G3698G20652，G3818G11468的G8712G7368G4493G7143進(jìn)入G5506G10711的內(nèi)部，G15932G19766的G4392G8946G7138顯G3698加（G16277G32824），和G256G22G171G257G20045G991G16278G4531到的G13479G7536G11468G1296G5506G10711在0G175HG794G22G71的G18334放G5242顯G14891G19489G1314。G17837G2499能G2528G7691是因?yàn)锽MP是G17902G17819G2572G19480G17745G14659，G5506G10711內(nèi)部和G15932G19766的G4392G8946為BMPG9354G9094的G9195G17891提供G1114很多的G17902道，有利G1122G14659G10301的G2572G19480。A221A222A221A223A221A224A221A225A221A226A221A227A221A221A228A226A223A225A229A223A230A231A232A233A234A235A236A237A238A239A240A241A242A243A244A245A246A222A221A247A248A249A250圖3內(nèi)水相中氯化鈉含量對(duì)BMP微球072H釋藥行為的影響10MA10MB10MC6圖4內(nèi)水相與油相的體積比對(duì)BMP微球形態(tài)的影響A251A252A253A254A255A1A0A2A3A4A14A252A253A254A255A5A12A6A24A2A3A433PLGA及PLGACOOH對(duì)BMP微球釋放度的影響G3926G32825所G12046，采G11004PLGACOOHG2058G3803的G5506G10711在0G175HG794G22G71的G18334放G5242G7138顯G1314G1122PLGAG5506G10711，G17837是因?yàn)锽MP是一G12193堿性蛋白（G12573電點(diǎn)在9G170左右），G2499與PLGACOOHG2010子鏈G1025的羧基G2499以發(fā)生G12175子G11468互G1328G11004而G1363BMPG2572G19480量G3698加6，因此PLGACOOHG5506G10711G1025未G2572G19480的BMPG1955G4581，G12373G18334G7138顯G19489G1314。A7A8A7A9A7A10A7A11A7A13A7A15A7A7A16A13A9A11A17A9A18A19A20A21A22A23A25A26A27A28A29A30A31A32A29A30A31A32A33A34A35A35A36圖5PLGA種類對(duì)BMP微球072H釋藥行為的影響34MPEGPLGA濃度對(duì)微球凝膠劑中BMP釋放度的影響G3926G32826所G12046，BMPG5506G10711G15932現(xiàn)G1998G989G11468G18334G14659G8181G534770G7941D的快G17907G18334放G7411。G17837是G11013G1122G15932G19766未G2572G19480G14659G10301快G17907G9354G16311所導(dǎo)致的，G13434有30G705左右的G14659G10301G18334放；第1DG7945D的G7114滯G7411G14659G10301G18334放G17907G5242非常G13543慢，G17837是因?yàn)镚2572G19480G1122G5506G10711G15932G19766的G14659G10301已經(jīng)G18334放完畢，而PLGA的G19489G16311G17907G5242較慢，G2010子量為121KDA的BMP很難從G5506G10711G1025G6205G696720MA20MB7G1998來(lái)；第5DG79414DG2530G18334G14659G17907G5242G7138顯加快，是G11013G1122G19555G11540PLGA逐漸G19489G16311成G8712G9354性小G2010子，G5506G10711內(nèi)部和G15932G19766的G4392隙G3698G3835，為G14659G10301的G6205G6967提供G1114G7368多G17902道。BMPG5506G10711G1969G14026G2070G8821有G15932現(xiàn)G1998G7138顯的G989G11468G18334G14659G8181G5347，體G3818G7366G13459符G2524HIGUICHIG7053G12255，6631654215TQR09884A37其G12373G18334G7138顯G1314G1122G5506G10711G2070，而在1G7945D的G18334G14659G17907G5242又略快G1122G5506G10711G2070，G2499能是混G5760在G1969G14026G1025的G5506G10711就像是包G990G1114一層衣，G20652粘G5242的G1969G14026層，G19489G1314G1114G5506G10711G990未G15999G2572G19480的BMP向G1183質(zhì)G6205G6967的G17907G5242。BMPG5506G10711G1969G14026G2070在5G79414D也G8821有像G5506G10711G2070那G7691呈現(xiàn)快G17907G18334放的特征，G17837是因?yàn)镚1969G14026層阻止G1114G8712G2010的G9195G17891，PLGAG19489G16311G17907G5242G1955慢所致。A38A39A38A40A38A41A38A42A38A43A38A380481216202428A44A45A46A47A48A49A50A51A52A53A54A55A54A55A56A57圖6MPEGPLGA濃度對(duì)BMP微球釋藥曲線的影響4結(jié)論本文比較G1114BMPG5506G10711和BMPG5506G10711G1969G14026G2070的體G3818G18334G14659特征，G2528G7114G17836G13783G4531G1114G3800G7053和G2058G3803G5049G14414對(duì)BMPG17745G14659G5506G10711G18334放G5242的影響。G11752G12362G15932G7138BMPG5506G10711呈現(xiàn)G989G11468G18334G14659G8181G5347，而B(niǎo)MPG5506G10711G1969G14026G2070體G3818G18334G14659G7366G13459