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《大鼠骨髓間充質(zhì)干細胞的培養(yǎng)鑒定以及向神經(jīng)干細胞分化的實驗研究》由會員上傳分享，免費在線閱讀，更多相關內(nèi)容在工程資料-天天文庫。
1、大鼠骨髓間充質(zhì)干細胞的培養(yǎng)鑒定以及向神經(jīng)干細胞分化的實驗研究蒲曉姝，果磊重慶醫(yī)科大學附屬第一醫(yī)院燒傷整形外科，重慶400016【摘要】目的體外培養(yǎng)并鑒定大鼠骨髓間充質(zhì)干細胞（BMSCs），研究其生物學特性，初步探索酸性成纖維細胞生長因子（aFGF）誘導其向神經(jīng)干細胞（NSCs）分化的可行性。方法全骨髓貼壁培養(yǎng)法分離培養(yǎng)大鼠BMSCs，倒置相差顯微鏡觀察其形態(tài)特征，MTT法繪制生長曲線，流式細胞儀（FCM）測定細胞周期并鑒定細胞表面標志物。用80ng/mlaFGF的無血清DMEM/F12培養(yǎng)液誘導大鼠BMSCs向NSCs分化，鏡下觀察細胞形態(tài)特征，免
2、疫熒光染色方法檢測神經(jīng)巢蛋白（nestin）、神經(jīng)元特異性烯醇化酶（NSE）的表達。結果大鼠BMSCs貼壁生長，呈梭形，多角形。第1、3、5代BMSCs的生長曲線均呈S形，活性無明顯差異。細胞周期顯示94.34%BMSCs處于G0/G1期，有較強的分裂增殖能力。FCM檢測CD29、CD54、CD90表達陽性，CD45表達陰性。誘導6h后細胞的胞體收縮成橢圓形或球形并伸出細長突起，免疫熒光染色顯示nestin表達陽性，繼續(xù)誘導發(fā)現(xiàn)雙極和多極細胞增多，誘導6d后相互連接成網(wǎng)狀，成典型的神經(jīng)元樣細胞，NSE表達陽性。結論全骨髓貼壁培養(yǎng)法能培養(yǎng)出高純度的B
3、MSCs，細胞生長穩(wěn)定、增殖快、可多次傳代，具有干細胞特性。經(jīng)aFGF誘導后具有向NSCs分化的潛能，NSCs能進一步分化為神經(jīng)元樣細胞?！娟P鍵詞】骨髓間充質(zhì)干細胞；全骨髓貼壁培養(yǎng)法；誘導分化；神經(jīng)干細胞；酸性成纖維細胞生長因子TheexperimentalstudyoncultivationandidentificationofratbonemarrowmesenchymalstemcellsandtheirdifferentiationintoneuralstemcellsPuXiaoshu,GuoLei.DepartmentofBurnand
4、PlasticSurgery,theFirstAffiliatedHospitalofChongqingMedicalUniversity,Chongqing400016,China.［Abstract］ObjectiveTocultureinvitroandidentifyratbonemarrowmesenchymalstemcells(BMSCs),studytheirbiologicalcharacteristics,andinitiallytentativelyexplorethefeasibilityofacidicfibroblast
5、?growthfactor?(aFGF)inducingBMSCstodifferentiateintoneuralstemcells(NSCs).MethodsRatBMSCswereisolatedandculturedbythewholebonemarrowadherentculturemethod.ThemorphologicalcharacteristicsofBMSCswereobservedundertheinvertedphasecontrastmicroscopeandthecellgrowthcurvesweredrawnbyM
6、TTmethod.Thecellcycleandsurfacemarkersweredetectedwithflowcytometry(FCM).RatBMSCswereinducedtodifferentiateintoNSCsby80ng/mlaFGFofDMEM/F12medium.ThecellmorphologicalchangeswerecontinuouslyobservedafterusingaFGF.Immunofluorescencestainingwasusedtodetecttheexpressionsofnestinand
7、neuronspecificenolase(NSE).ResultsRatBMSCsshowedadherentgrowthandspindle-shapedorpolygonalappearance.The?growthcurve?of?the1st,3rd,5th-generation?BMSCs?showedS-shapeandnosignificantdifferencein?activity.FCMshowed?that?therewere94.34%?ofBMSCs?in?G0/G1phaseofthecellcycle,havings
8、trong?proliferation?ability.FCMdetected?thatCD29,CD54andCD90?
