CAJ | 학술논문

目的利用动力性三维细胞培养技术建立大鼠胰腺导管来源干细胞( PDSC)系.方法以大鼠胰腺组织为材料,采用V型胶原酶原位消化分离大鼠胰腺组织,通过不连续密度梯度离心使胰腺导管细胞与胰岛细胞初步分离,采用动力性三维培养技术进行培养,获得原代PDSC,并进行扩增培养、连续传代和纯化,建立PDSC系.对原代分离的PDSC进行鉴定,包括形态学的鉴定及表达特性的鉴定.结果通过胶原酶消化、不连续密度梯度离心、动力性三维细胞培养,可分离培养出大鼠PDSC,并经过动力性三维培养,可以建立大鼠PDSC系.形态学方面,PDSC呈单个核；生长行为方面,PDSC在三维载体上沿纤维贴壁生长；表达特性方面,通过流式细胞仪分选结果显示,PDSC高表达CD29、CD73,CD90、CD105,不表达CD14、CD19、CD34、CD45,证实P DSC为间充质干细胞.结论通过采用原位胶原酶消化法、不连续密度梯度离心、动力性三维细胞培养系统,可分离培养出大鼠PDSC,成功建立PDSC细胞系.
목적 이용동력성삼유세포배양기술건립대서이선도관래원간세포( PDSC)계.방법 이대서이선조직위재료,채용V형효원매원위소화분리대서이선조직,통과불련속밀도제도리심사이선도관세포여이도세포초보분리,채용동력성삼유배양기술진행배양,획득원대PDSC,병진행확증배양、련속전대화순화,건립PDSC계.대원대분리적PDSC진행감정,포괄형태학적감정급표체특성적감정.결과 통과효원매소화、불련속밀도제도리심、동력성삼유세포배양,가분리배양출대서PDSC,병경과동력성삼유배양,가이건립대서PDSC계.형태학방면,PDSC정단개핵；생장행위방면,PDSC재삼유재체상연섬유첩벽생장；표체특성방면,통과류식세포의분선결과현시,PDSC고표체CD29、CD73,CD90、CD105,불표체CD14、CD19、CD34、CD45,증실P DSC위간충질간세포.결론 통과채용원위효원매소화법、불련속밀도제도리심、동력성삼유세포배양계통,가분리배양출대서PDSC,성공건립PDSC세포계.
Objective To isolate,purify and identify pancreatic duct derived stem cells (PDSCs) from the pancreatic duct of rats,and culture in the three-dimension cell culture system.Methods Adult male Wistar rats underwent perfusion with collagenase V via the pancreatic duct,then the pancreas was surgically procured,digested,followed by discontinued density gradient centrifuge to isolate ductal tissue from islets.The acinar and ductal tissue was cultivated in serumcontaining medium in the three-dimension cell culture to obtain adherent cells,as PDSCs,which were expanded by consecutive passages.The morphology and characterization of PDSCs on phenotype were examined.Results PDSCs could be obtained through in situ collagenase V digestion,discontinued density gradient centrifuge,and culture in the three dimension cell culture system.Morphologically,PDSCs had remarkable size,most with one nucleus.PDSCs grew in many layers in three-dimension cell culture system.PDSCs was revealed to express CD29,CD73,CD90,CD105,but not CD14,CD19,CD34,CD45 by FACS,in agreement with MSCs.Conclusion PDSCs of rats could be obtained through in situ collagenase V digestion,discontinued density gradient centrifuge,and culture in the three-dimension cell culture system.PDSCs lines were successfully established.