大鼠骨髓间充质干细胞磁标记及MR成像研究

Magnetic labeling and in vitro MR imaging of rat bone marrow mesenchymal stem cells

目的应用菲立磁-多聚左旋赖氨酸复合物标记大鼠骨髓间充质干细胞,探讨MR成像显示磁标记干细胞的可行性。方法制备菲立磁-多聚左旋赖氨酸复合物。分离培养Wistar大鼠骨髓间充质干细胞,以菲立磁-多聚左旋赖氨酸复合物标记干细胞。分别于标记后24h及1、2、3周行普鲁士蓝染色观察细胞内铁,台盼蓝排除试验检测细胞活力。应用1.5TMR仪,以SE序列T1WI、T2WI和梯度回波(GRE)序列T2WI行磁标记干细胞成像。结果普鲁士蓝染色显示细胞质内大量铁颗粒存在,标记率100%;随细胞分裂增殖,细胞内铁颗粒逐渐减少。干细胞磁标记后24h及1、2、3周的台盼蓝拒染率分别为91.00%、93.00%、91.75%和92.50%,与未标记细胞相比较差异无统计学意义(P>0.05)。103、104、105个磁标记干细胞T2WI信号降低分别为63.75%、82.31%、91.92%,T2WI信号降低分别为68.24%、83.01%、93.94%。105个干细胞磁标记后24h及1、2、3周T2WI信号降低分别为93.75%、75.92%、41.75%、8.83%。结论应用菲立磁-多聚左旋赖氨酸复合物标记大鼠间充质干细胞安全、有效;T2WI对磁标记干细胞的显示最敏感;MR信号改变与干细胞数目及分裂增殖状态相关。

Objective To label rat bone marrow mesenchymal stem cells with feridex combined with poly-l-lysine （ PLL）, and to determine the feasibility of detection of magnetically labeled stem cells with MR imaging. Methods Feridex were incubated with PLL for 1 hour to obtain a complex of feridex-PLL. Mesenchymal stem cells isolated from the bone marrows of Wistar rats were cultured and expanded. By the 42 passage, cells were co-incubated overnight with the feridex-PLL complex. Prussian blue staining for demonstrating intracytoplastic nanoparticles and trypan-blue exclusion test for cell viability were performed respectively at 24 h,1 w,2 w,3 w after labeling. MR imaging of cell suspensions was performed by using T1WI, T2WI and T2 ＊ WI sequences at a clinical 1.5 T MR system. Results Numerous intracytoplastic iron particles were stained with Prussian blue. With division of stem cells, the stained particles were seen decreased gradually. Trypan blue exclusion test at 24 h, 1 w, 2 w and 3 w showed that the viability of the labeled cells was 91.00%, 93.00%, 91.75%, and 92. 50%, not significantly different with that of nonlabeled cells（P 〉 0.05）. For 10^3, 10^4 and 10^5 cells, T2 signal intensity decreased by 63.75%, 82. 31% and 91.92% respectively, T2 ＊ signal intensity decreased by 68. 24%, 83.01%, and 93.94% respectively. For l0s labeled cells, T2＊ signal intensity decreased by 93.75%, 75.92%, 41.75% and 8. 83% respectively at 24 h, 1 w, 2 w and 3 w after labeling. Conclusion Magnetic labeling of rat bone marrow stem cells with feridex-PLL complex is feasible, efficient and safe. T2 ＊ WI is the most sensitive sequence to detect the labeled cells. The degree of T2 signal decreasing may be related to the cell count and division phase.