超顺磁性氧化铁纳米粒子标记羊骨髓间充质干细胞的体外实验研究

目的：探究超顺磁性氧化铁纳米粒子体外标记羊骨髓间充质干细胞的最佳方案,进行磁共振扫描,观察与三维打印组织工程骨支架复合培养的效果。方法：取超顺磁性氧化铁纳米粒子与多聚左旋赖氨酸的复合物标记第3代羊骨髓间充干细胞,将两者（各mg/L）按配比分为4组,A组：25：0.75、B组：50：1.5、C组：100：3和D组：200：6,分别在4h、24h和48h检测细胞标记阳性率、存活率和增殖活力,选最佳方案组进行磁共振成像、观察细胞内外粒子分布并与支架复合培养。结果：A组、B组细胞标记24h和48h普鲁士蓝染色阳性率最高,均达100%,细胞外未见纳米粒子聚集块形成;A组细胞标记4h、24h细胞存活率最高,与对照组差异无统计学意义;4组纳米粒子标记不同时间均未影响细胞增殖活力;在3.0T磁共振成像下至少可检测到A组5×103个细胞;纳米粒子分布于胞浆内、可与支架有效粘附。结论：超顺磁性氧化铁纳米粒子与多聚左旋赖氨酸配比为25：0.75（mg/L）时,标记第3代羊骨髓间充质干细胞24h为最佳方案,可粘附于三维打印组织工程骨支架表面。

Experimental Study of Superparamagnetic Iron Oxide Nanoparticles Labeled Sheep Bone Marrow Mesenchymal Stem Cells in Vitro

Objective：To explore the optimal plan of superparamagnetic iron oxide nanoparticles labeled sheep bone marrow mesenchymal stem cells in vitro,the effect of taking MR imaging and the group of labeled cells compound cultivate with the 3D-printing of bone tissue engineering scaffold.Methods：We used the compound solution of nanoparticles and poly-L-lysine to label the third generation of the sheep bone marrow mesenchymal stem cells.According to different ratio of nanoparticles and poly-l-lysine,four groups were divided as follows,group A：25：0.75,group B：50：1.5,group C：100：3and group D：200：6.The cell labeling positive rate,the cell survival rate and cell proliferation activity were detected as time point 4h,24hand 48h.The best experimental group was imaged by MR imaging,intracellular and extracellular nanoparticles and scaffold compounded cultivate with the labeled cells were detected.Results：Group A and group B were labeled efficiently after 24hand 48h.The prussian blue staining positive rate was 100%.No visible of extracellular nanoparticles gathered.Group A had the highest survival rate after 4hand 24h.There was no significant difference with the control group.The labeling time of four groups did not affect cell proliferation activity.3.0T MR imaging could detect 5×103cells at least.Nanoparticles distributed in cytoplasm and the labeled cells could be seen adhere on the scaffold surface.Conclusion：The ratio of superparamagnetic iron oxide nanoparticles and poly-l-lysine as（25：0.75）μg/mL,labeled the third generation of the sheep bone marrow mesenchymal stem cells for 24hwas the optimal plan,and the labeled cells could adhere on the 3Dprinting of bone tissue engineering scaffold surface.