超顺磁性氧化铁和DAPI双标记骨髓间充质干细胞

背景：骨髓间充质干细胞目前已经成为重要的组织工程种子细胞,而若想深入研究其在体内外增殖、分化的规律,首先需要解决如何能高效、安全地标记骨髓间充质干细胞。 目的：观察超顺磁性氧化铁及DAPI对大鼠骨髓间充质干细胞的双标记效果及其对细胞存活、增殖以及凋亡的影响。 方法：分离培养大鼠骨髓间充质干细胞,用超顺磁性氧化铁及DAPI双标记后分别用普鲁士蓝染色法和激光共聚焦显微镜观察其铁标记率及荧光标记率。用锥虫蓝检测细胞活力;MTT法检测标记干细胞增殖力;Calcein-AM/PI以及AO/PI双染细胞,检测细胞存活率以及凋亡率。 结果与结论：超顺磁性氧化铁及DAPI在体外双标记大鼠骨髓间充质干细胞效率高,可达100%;锥虫蓝染色显示标记细胞的存活率为97%;MTT法检测发现双标记干细胞组的活力以及增殖力与未标记干细胞组相比差异均无显著性意义(P > 0.05);Calcein-AM/PI染色,未标记细胞及双标记细胞的存活率分别为96%和95%;AO/PI染色,未标记及双标记细胞的凋亡率均为1%。提示超顺磁性氧化铁和DAPI双标记大鼠骨髓间充质干细胞后,对于干细胞的存活、增殖以及凋亡无影响。

Superparamagnetic iron oxide and 4',6-diamidino-2-phenylindole double-labeled bone marrow derived mesenchymal stem cells

BACKGROUND:Now, bone marrow mesenchymal stem cells (BMSCs) have become important seed cells of tissue engineering. It is necessary to solve the problem that how to label BMSCs efficiently and safely for further study on BMSCs proliferation and differentiation in vitro and in vivo. OBJECTIVE:To investigate the effects of superparamagnetic iron oxide (SPIO) and 4',6-diamidino-2-phenylindole (DAPI) double-labeling on BMSCs, and to study whether there is any effect on cell viability, proliferation and apoptosis. METHODS:BMSCs were isolated and double-labeled with SPIO and DAPI. The labeling rate was identified by Prussian blue staining and fluorescence microscope, then cell vitality was detected by Trypan blue staining and proliferation activity was measured by MTT assay. Cell survival rate and apoptosis rate were observed by using Calceein AM/PI and AO/PI staining. RESULTS AND CONCLUSION:The stem cell double-labeling rate by SPIO and DAPI were nearly 100%. The cell survival rate with Trypan blue staining was 97%. MTT method detected that there was no significant difference for proliferation activity between double-labeled and unlabeled stem cells (P > 0.05). Calcein-AM/PI staining detected that the survival rate of double-labeled and unlabeled stem cells were 95% and 96%. AO/PI staining detected that the apoptosis rate of double-labeled and unlabeled stem cells were both 1%. These suggested that there is no effect on survival, proliferation and apoptosis of BMSCs after SPIO and DAPI double-labeling.