CAJ | 학술논문

背景:内皮细胞基础培养液丰要用于培养内皮祖细胞,作者所查用此液培养间充质干细胞的报道较少.目的:对以内皮细胞基础培养液为基础的不同方法培养脐血间充质干细胞的效果比较.设计、时间和单位:对比观察的细胞培养实验,于2005-09/2006-05在上海市新华医院市级重点实验室完成.材料:选取妊娠杂交犬8只,抽取脐血进行干细胞的分离和培养.内皮细胞基础培养液及内皮细胞培养基为美国Clonetics产品.鼠抗CDlla单克隆抗体,鼠抗CDllb单克隆抗体,鼠抗CD29单克隆抗体,鼠抗CD71单克隆抗体均为美国Antibody diagnosdca产品;鼠抗CD34单克隆抗体为美国Lab VisionCorporation产品.方法:收集的脐血干细胞分为A,B,C,D4组.A组用内皮细胞摹础培养液培养:B组用添加有微血管内皮细胞生长培养液的内皮细胞基础培养液在6孔板上培养.C组用添加有内皮细胞培养基的内皮细胞基础培养液在纤维连接蛋白包被的6孔板上培养.D组用添加有内皮细胞培养基的内皮细胞基础培养液在25cm2细胞培养瓶中培养.主要观察指标:观察各组细胞形态学变化和群体倍增数.应用免疫组化法检测细胞抗原CDlla、CDllb、CD34、CD29及CD71的表达.结果:各组均有成纤维细胞样细胞培养出.A组细胞形态不良,增殖缓慢;B组细胞增殖旺盛:C组细胞克隆不稳定,容易老化;同时出现另一种细胞克隆.D组细胞培养的结果与C组相似.免疫组化法检测抗原显示CD11(-),CD11b(-),CD34(-),CD29(+)及CD71(+).结论:在未经处理的6孔板上,用添加有内皮细胞培养基的内皮细胞基础培养液细胞培养液可以培养出生长良好、增殖旺盛的间充质干细胞.应用纤维连接蛋白和25cm2细胞培养瓶培养的效果不佳.
배경:내피세포기출배양액봉요용우배양내피조세포,작자소사용차액배양간충질간세포적보도교소.목적:대이내피세포기출배양액위기출적불동방법배양제혈간충질간세포적효과비교.설계、시간화단위:대비관찰적세포배양실험,우2005-09/2006-05재상해시신화의원시급중점실험실완성.재료:선취임신잡교견8지,추취제혈진행간세포적분리화배양.내피세포기출배양액급내피세포배양기위미국Clonetics산품.서항CDlla단극륭항체,서항CDllb단극륭항체,서항CD29단극륭항체,서항CD71단극륭항체균위미국Antibody diagnosdca산품;서항CD34단극륭항체위미국Lab VisionCorporation산품.방법:수집적제혈간세포분위A,B,C,D4조.A조용내피세포모출배양액배양:B조용첨가유미혈관내피세포생장배양액적내피세포기출배양액재6공판상배양.C조용첨가유내피세포배양기적내피세포기출배양액재섬유련접단백포피적6공판상배양.D조용첨가유내피세포배양기적내피세포기출배양액재25cm2세포배양병중배양.주요관찰지표:관찰각조세포형태학변화화군체배증수.응용면역조화법검측세포항원CDlla、CDllb、CD34、CD29급CD71적표체.결과:각조균유성섬유세포양세포배양출.A조세포형태불량,증식완만;B조세포증식왕성:C조세포극륭불은정,용역노화;동시출현령일충세포극륭.D조세포배양적결과여C조상사.면역조화법검측항원현시CD11(-),CD11b(-),CD34(-),CD29(+)급CD71(+).결론:재미경처리적6공판상,용첨가유내피세포배양기적내피세포기출배양액세포배양액가이배양출생장량호、증식왕성적간충질간세포.응용섬유련접단백화25cm2세포배양병배양적효과불가.
BACKGRoUND:Endothelial basal medium is mainly used to culture endothelial progenitor cells.Studies on mesenchymal stem cells (MSCs) cultured in this medium are few. OBJECTIVE:To compare the outcome of MSCs cultured in different mediums including endothelial basal medium. DESiGN.TIME AND SETTING:The control eell experiment was performed at the Muniopal Key Laboratory of Xinhua Hospital of Shanghai.China from September 2005 to May 2006. MATERIALS:Eight pregnant mongrel dogs were selected to obtain umbilical cord blood for isolation and culture of stem cells. Endothelial eell basal medium and endothelial cell medium were bought from Clonetics.USA.Mouse anti-CDlla monoclonal andbody,mouse anti-CDllb monoclonal antibody.mouse anti-CD29 monoclenal antibody and mouse anti-CD7l monoclonal antibody Were purchased from Antibody diagnostica,USA.Mouse anti-CD34 monoclonal antibody was obtained from Lab Vision Corporation.USA.METHODS:Umbilical cord blood stem cells were divided into four groups.Umbilical cord blood steTn cells in the group A were incubated in the endothellal basal medium.Umbilical cord blood steHl cells in the group B were incubated in the endothelial basal medium containing microvascular endothelial cells in a 6-well plate.Umbilical cord blood stem cells in the group C were incubated in the endothelial basal medium containing endotheliaI medium in a 6-well plate coated with fibronectin.Umbilical cord blood stelrn cells in the group D were incubated in the endothelial basal medium containing endothelial medium in a 25 cm2 ctdturing flask.MAIN OUTCOME MEASURES:Cell morphology and population doublings were observed.CDlla,CDllb,CD34,CD29 and CD71 expression was detected by immunohistochemistry. RESULTS:Fibroblast-like cells were measured in each group.The celIs grew badly in morphology and proliferated slowly in the group A,while cells proliferated rapidlyinthe group B.The cell clones were instable and inclined to aging in the group C,with a new cell clone was found.The cells in the group D were similar to those in the group C.Surface antigens detected by immunohistochemistry showed CDlla(-),CDllb(-),CD34(-),CD29(+)and CD7l(+). CONCLUSION:MSCs witll well growth and rapid proliferation can be detected in the endothelial basal medium containing endothellal medium in an intact 6-well plate.The outcome is bad in fibronectm-coated plate or 25 cm2 culture flask.