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1、中國組織工程研究第16卷第14期2012–04–01出版ChineseJournalofTissueEngineeringResearchApril1,2012Vol.16,No.14綿羊羊膜上皮細胞分離培養(yǎng)及鑒定*☆朱雪敏,李海軍,米焱,楊銀鳳,關(guān)洪敏,曹貴方Isolation,cultureandidentiticationofOvisariesamnionepithelialcellsZhuXue-min,LiHai-jun,MiYan,YangYin-feng,GuanHong-min,CaoGui-f
2、angAbstractBACKGROUND:Theamnionepithelialcells(AECs)withstemcellspropertieshavebeensuccessfullyobtainedafterisolating,digestingandculturinghumanandratamnionandAECshavethepotentialtodifferentiateintocellsoftheendoderm(liver,lung,epithelium),mesoderm(bone,fat)
3、,andectoderm(neuralcells),OBJECTIVE:ToobtaintheOvisariesAECsanddetectthestemcellcharacteristics.METHODS:Ovisariesamniontissuewaspeeledoffbymechanicalwayandthelowvelocity-trypLE(pancreaticenzymereplacement)digestionmethodwasusedtoisolateandculturetheOvisaries
4、amnion,andtheOvisariesAECswereobtained.RESULTSANDCONCLUSION:TheimmunofluorescenceobservationshowedthatthelabeledproteinofAECssuchasOct-4,SSEA-1,SSEA-3,SSEA-4,TRA-1-60,TRA-1-81couldbeexpressedonOvisariesAECs.RT-PCRexperimentsconfirmedthatthegenesofOct-4,Sox-2
5、andRex-1wereexpressedandNanogwasnotexpressed.TheresultsindicatethattheOvisariesAECswithstemcellspropertieshavebeenachievedsuccessfully.ZhuXM,LiHJ,MiY,YangYF,GuanHM,CaoGF.Isolation,cultureandidentiticationofOvisariesamnionepithelialcells.ZhongguoZuzhiGongchen
6、gYanjiu.2012;16(14):2559-2562.[http://www.crter.cnhttp://en.zglckf.com]摘要背景:目前有研究發(fā)現(xiàn)人羊膜和大鼠羊膜分離消化培養(yǎng)后,可成功獲得具有干細胞特性的人羊膜上皮細胞和鼠羊膜上皮細胞,在體外一些外源因子作用下具有向三胚層細胞分化的潛能。目的:獲取綿羊羊膜上皮細胞,并對其干細胞特性進行鑒定。方法:采用機械方法剝離綿羊羊膜組織,運用低速旋轉(zhuǎn)TrypLE(胰酶替代物)消化法對綿羊羊膜進行分離培養(yǎng)獲得綿羊羊膜上皮細胞。結(jié)果與結(jié)論:細胞免疫熒光檢測結(jié)
7、果表明綿羊羊膜上皮細胞表達Oct-4、SSEA-1、SSEA-3、SSEA-4、TRA-1-60、TRA-1-81等胚胎干細胞標記蛋白;RT-PCR技術(shù)鑒定結(jié)果顯示綿羊羊膜上皮細胞的Oct-4、Sox-2、Rex-1全能性相關(guān)基因表達,而Nanog基因不表達。檢測結(jié)果提示實驗成功獲得具有干細胞特性的綿羊羊膜上皮細胞。關(guān)鍵詞:綿羊羊膜上皮細胞;分離;培養(yǎng);細胞免疫熒光;RT-PCR縮略語注釋:AECs:amnionepithelialcells,羊膜上皮細胞doi:10.3969/j.issn.1673-8225
8、.2012.14.018朱雪敏,李海軍,米焱,楊銀鳳,關(guān)洪敏,曹貴方.綿羊羊膜上皮細胞分離培養(yǎng)鑒定[J].中國組織工程研究,2012,16(14):2559-2562.[http://www.crter.orghttp://cn.zglckf.com]LaboratoryofAnimalEmbryoandDevelopmentalBiology,InnerMongoliaAgricu