PKH26荧光素标记脂肪间充质干细胞的生物学特性的研究

In vitro study of biological characteristics of adipose tissue derived mesenchymal stem cells after PKH26-labeled

目的研究PKH26荧光素染料对脂肪间充质干细胞(ADSCs)生物学特性的影响。方法分离培养新生SD大鼠的ADSCs,通过流式细胞仪、成脂、成骨、成软骨诱导鉴定ADSCs的免疫表型和多向分化的能力。用细胞膜荧光素染料PKH26对第二代脂肪干细胞进行标记,通过倒置相差显微镜、荧光显微镜以及CCK-8法评估标记后细胞的形态、荧光强度以及增殖活性的变化。结果成功分离和培养ADSCs,其纯度达95%以上,并成功将ADSCs诱导成脂肪、骨、软骨;PKH26标记24 h后,在荧光显微镜下可见细胞膜上均匀分布的强红色荧光,细胞的形态以及增殖活性与未标记的细胞无明显差异。结论 PKH26荧光素染料标记后对ADSCs的形态及增殖活性无明显影响,是一种理想的细胞标志物,能够很好的应用于示踪间充质干细胞的迁移转归及干细胞标记等方面的研究。

Objective To investigate the effects of PKH26 fluorescent labeling on the biological characteristics of adipose derived mesenchymal stem cells（ ADSCs）. Methods The ADSCs were isolated from neonatal SD rats,the immunophenotype of ADSCs were identified by flow cytometry and the multilineage differentiation ability of ADSCs was detected by adipogenic,osteogenic and chondrogenic differentiation. The cell membrane of the second generation ADSCs was labeled with PKH26,and then the morphology,fluorescence intensity and proliferation activity of the cells were evaluated by inverted phase contrast microscopy,fluorescence microscopy and CCK-8. Results The ADSCs with purity above 95% were successfully isolated from neonatal SD rats,and were successfully induced into fat,bone and cartilage. After labeled with PKH26 for 24 h,the red fluorescence was uniformly distributed on the cell membrane under fluorescence microscope. The morphology and proliferation of the cells were not significantly different from those of unlabeled cells. Conclusion PKH26 labeling had no significant effect on cell morphology and proliferation activity,PKH26 is an ideal cell marker,which can be used to study the migration and prognosis of mesenchymal stem cells.