CAJ | 학술논문

目的观察骨髓间充质干细胞(MSC)在异位小肠移植大鼠体内的定植情况.方法骨髓细胞取自1月龄的雄性Lewis大鼠,通过密度梯度离心法结合贴壁法分离和培养骨髓MSC;通过流式细胞术对MSC表面抗原(CD29、CD90、CD34和CD45)进行鉴定;应用终浓度为5μmol/L的5(6)-羧基二乙酸荧光素琥珀酰亚胺酯(CFSE)对生长良好的第3代MSC进行标记.以成年雄性近交系F344大鼠作为供体、成年雄性Lewis大鼠作为受体,建立大鼠异位小肠移植模型.术后经阴茎背静脉将CFSE标记的MSC注入受体体内,术后第7天收集冰冻病理标本,在荧光显微镜下观察MSC的定位情况.结果成功分离培养出骨骼MSC,其表面抗原CD29、CD90、CD34和CD45的平均阳性表达率分别为96.48％、99.77％、2.41％和1.39％.CFSE能有效标记MSC,术后7d在移植小肠的间质、脾脏和胸腺可观察到大量绿色荧光,自体小肠组织仅见微量荧光,而心、肝和肺组织基本未见绿色荧光.结论骨骼MSC能成功定植于大鼠小肠移植术后的移植小肠中.
목적 관찰골수간충질간세포(MSC)재이위소장이식대서체내적정식정황.방법 골수세포취자1월령적웅성Lewis대서,통과밀도제도리심법결합첩벽법분리화배양골수MSC;통과류식세포술대MSC표면항원(CD29、CD90、CD34화CD45)진행감정;응용종농도위5μmol/L적5(6)-최기이을산형광소호박선아알지(CFSE)대생장량호적제3대MSC진행표기.이성년웅성근교계F344대서작위공체、성년웅성Lewis대서작위수체,건립대서이위소장이식모형.술후경음경배정맥장CFSE표기적MSC주입수체체내,술후제7천수집빙동병리표본,재형광현미경하관찰MSC적정위정황.결과 성공분리배양출골격MSC,기표면항원CD29、CD90、CD34화CD45적평균양성표체솔분별위96.48％、99.77％、2.41％화1.39％.CFSE능유효표기MSC,술후7d재이식소장적간질、비장화흉선가관찰도대량록색형광,자체소장조직부견미량형광,이심、간화폐조직기본미견록색형광.결론 골격MSC능성공정식우대서소장이식술후적이식소장중.
Objective To study the bone marrow mesenchymal stem cells (MSC) settled in rats after small intestinal transplantation.Methods Bone marrow MSCs were taken from 1-month male Lewis rats, isolated and cultured by density gradient centrifugation and differential adherent culture.The surface antigens (CD29, CD90, CD34 and CD45) of MSC were identified by flow cytometry.Final concentration of 5 μg/L CFSE was used to mark the third generation of MSCs.Adult male inbred line F344 rats were used as donor and adult male Lewis rats as acceptor.A heterotopic intestinal transplant rat model was established by F344 to Lewis.Labeled MSCs were injected into model rats through vena dorsalis penis after operation.Tissues at postoperative 7-day were collected for frozen pathology to reveal the location of transplanted MSCs under fluorescence microscope.Results MSCs were successfully isolated from rat bone marrow.The average positive expression rates of surface antigens CD29, CD90,CD34 and CD45 were 96.48％, 99.77％, 2.41％ and 1.39％ respectively.MSCs were successfully and effectively marked with CFSE.Seven days after operation, a large number of green fluorescence could be observed in transplanted intestine, spleen and thymus.Autograft intestinal tissues only showed trace fluorescence, and the heart, liver and lung tissue basically did not present the green fluorescence.Conclusion Bone marrow MSCs can settle in transplanted small intestine of rat.