目的 探讨磁性纳米材料低分子量聚乙烯亚胺修饰的超顺磁性氧化铁(Alkyl-PEI2k/SPIO)标记成人外周血源性血管内皮祖细胞(EPCs)的标记效率及7.0 T MR体外成像效果.方法 密度梯度离心贴壁培养EPCs.Alkyl-PEI2k/SPIO(Fe终质量浓度7μg/ml)标记EPCs,普鲁士蓝染色检测标记率;以CCK-8实验、Transwell迁移实验及Matrigel成管实验分别检测标记后细胞的生长、增殖活性,迁移能力及体外成管能力;将Alkyl-PEI2k/SPIO标记的EPCs(0、5×10^3、1×10^4、5× 10^4、1×10^5、2.5×10^5、5×10^5、1× 10^6和1.5×10^6个)制成琼脂糖凝胶细胞模型进行7.0 T MR成像.对数据行两样本t检验和配对t检验.结果 EPCs标记率达100%,标记后EPCs的含铁量为(6.062±0.050) pg/细胞;各时间点标记和未标记细胞的吸光度间差异无统计学意义(均P>0.05);SPIO标记组[(80.6±8.0)个]和未标记组EPCs[(77.6±4.6)个]迁移细胞数差异无统计学意义(P>0.05);SPIO标记组成管数量[(9.0±1.0)个]和未标记组EPCs成管数量[(9.4±1.5)个]相当(P>0.05).EPCs琼脂糖凝胶模型MRI结果显示,T2、T2*图像信号强度和弛豫时间随细胞数目的增加而减低,不同细胞数凝胶模型T2和T2*间的信号强度、弛豫时间差异有统计学意义(P<0.01).结论 Alkyl-PEI2k/SPIO标记成人外周血源性EPCs的效率高,且对细胞活性无明显影响;高场强7.0T MR能够用于Alkyl-PEI2k/SPIO标记的EPCs体外成像.
Objective To assess the feasibility of labeling endothelial progenitor cells (EPCs) with N-Alkyl-polyethylenimine 2 kDa stabilized superparamagnetic iron oxide (Alkyl-PEI2k/SPIO) and imaging in vitro by a 7.0 T MR scanner.Methods EPCs were obtained from human peripheral blood by density gradient centrifugation and cultured adherently under given conditions.Alkyl-PEI2k/SPIO (7 μg/ml Fe) was used to label EPCs by incubation overnight,and the labeling efficiency was determined by Prussian blue staining.The viability and activity of labeled cells were evaluated using CCK-8,Transwell migration and tubulogenesis assays.Labeled EPCs agarose hydrogels with different number of cells (0,5×10^3,1×10^4,5×10^4,1×10^5,2.5×10^5,5×10^5,1×10^6 and 1.5×10^6) were prepared,and in vitro 7.0 T MR was performed.Signal intensity (SI) and relaxation time were measured.Data were analyzed by two-sample t test and paired t test.Results The labeling efficiency with Alkyl-PEI2k/SPIO at a 7 μg Fe/ml concentration was 100%.Quantitative analysis of cellular iron was (6.062±0.050) pg/cell.There was no statistical difference between the absorbance of labeled and unlabeled EPCs determined by CCK-8 assay (P〉0.05).Migration assay showed that the number of labeled migrated EPCs (80.6±8.0) was comparable to that of unlabeled EPCs (77.6±4.6; P〉0.05).Tubulogenesis assay suggested that the number of tube-like structures formed by labeled EPCs (9.0± 1.0) was not statistically different from that of the unlabeled EPCs (9.4± 1.5 ; P〉0.05).In vitro MRI showed that SI and relaxation time were reduced with the increased number of labeled EPCs.Both SI and relaxation time had significant differences between T2 and T2* (P〈0.01).Conclusions EPCs could be labeled with Alkyl-PEI2k/SPIO efficiently without significant side effect on cell activity.Magnetically labeled EPCs could be imaged in vitro by a 7.0 T MR scanner.