CAJ | 학술논문

目的建立PKH26荧光标记肝纤维化大鼠骨髓内皮祖细胞(EPC)的方法,观察PKH26标记的EPC输注后在肝纤维化大鼠肝内的迁移和分化情况.方法分离和培养肝纤维化大鼠骨髓源性EPC,并在体外进行PKH26荧光标记,通过荧光共聚焦显微镜观察和采用流式细胞仪检测PKH26的标记率,并观察荧光标记对细胞生长状态的影响；将PKH26荧光标记的EPC经尾静脉输注入肝纤维化大鼠体内,观察其在大鼠肝内的迁移示踪情况,并用免疫荧光法检测内皮细胞标志物CD31和血管性假血友病因子(vWF)的表达.结果 PKH26标记的细胞呈红色荧光,标记率为96.65％；PKH26标记的EPC生长状态良好,与未标记的EPC相比,生长曲线基本一致；EPC迁移入肝后主要存在于沿纤维分布的血管内皮和肝小叶内的窦内皮,并表达内皮细胞特异性抗原CD31和vWF.结论 PKH26可在体外成功标记肝纤维化大鼠骨髓源性EPC,该细胞输注入肝纤维化大鼠体内后,可迁移至肝内血管周围,并向成熟的内皮细胞分化.
목적 건립PKH26형광표기간섬유화대서골수내피조세포(EPC)적방법,관찰PKH26표기적EPC수주후재간섬유화대서간내적천이화분화정황.방법 분리화배양간섬유화대서골수원성EPC,병재체외진행PKH26형광표기,통과형광공취초현미경관찰화채용류식세포의검측PKH26적표기솔,병관찰형광표기대세포생장상태적영향；장PKH26형광표기적EPC경미정맥수주입간섬유화대서체내,관찰기재대서간내적천이시종정황,병용면역형광법검측내피세포표지물CD31화혈관성가혈우병인자(vWF)적표체.결과 PKH26표기적세포정홍색형광,표기솔위96.65％；PKH26표기적EPC생장상태량호,여미표기적EPC상비,생장곡선기본일치；EPC천이입간후주요존재우연섬유분포적혈관내피화간소협내적두내피,병표체내피세포특이성항원CD31화vWF.결론 PKH26가재체외성공표기간섬유화대서골수원성EPC,해세포수주입간섬유화대서체내후,가천이지간내혈관주위,병향성숙적내피세포분화.
Objective To establish a method of PKH26 labeled bone marrow-derived endothelial progenitor cells (EPCs) into rats with liver fibrosis and observe cell immigration and differentiation in the liver after transplantation.Methods Bone marrow-derived EPCs were isolated and cultured from rats with liver fibrosis,and then labeled with PKH26 in vitro.Under the scanning confocal microscopy and flow cytometry,PKH26 fluorescent labeling rate and cell survival rate were measured.EPCs of PKH26 fluorescent labeling were transplanted into rats with liver fibrosis via the tail vein,and the migration situation was observed in the liver.Endothelial cell markers CD31 and von willebrand factor (vWF) were detected by using immunofluorescence.Results The PKH26-labeled EPCs appeared red fluorescence and the labeling rate was 96.65 ％.As compared with unlabeled cells,the labeled cells grew well,and had no significant changes in the growth curve.After transplantation into the liver of rats,the PKH26 labeled cells were mainly distributed in blood vessel endothelium along fibers and hepatic sinusoids in hepatic lobule.Endothelial cell-specific antigens such as CD31 and vWF could be detected along the vascular walls.Conclusion PKH26 could be used to label and track EPCs in the liver of rats with liver fibrosis in vitro.The PKH26-labeled cells may migrate to the surrounding of the hepatic vessels and differentiate into mature endothelial cells.