磁性荧光双标人骨髓间充质干细胞归巢至大鼠急性损伤肝组织示踪显像

发布时间：2018-10-09 08:58

【摘要】：目的观察磁粒子和荧光染料双标人骨髓间充质干细胞归巢至大鼠急性损伤肝组织的示踪。方法以超顺磁性氧化铁(SPIO)和氯甲基苯甲酰氨(CM-Dil)标记永生化人骨髓间充质干细胞(UE7T-13)。普鲁士蓝染色检测细胞内铁,荧光显微镜和流式细胞仪检测CM-Dil标记阳性率。建立12只急性肝损伤大鼠模型,分为实验组(n=6)和对照组(n=6),将标记细胞经尾静脉移植入实验组大鼠,未标记细胞移植入对照组大鼠。分别于移植前3h、移植后3h、3天、5天、7天应用MR T2*W、T2*map对大鼠活体成像,获得肝脏R2*值。并与肝脏组织切片普鲁士蓝染色和CM-Dil荧光表达情况对照。结果双标细胞的SPIO-PLL标记率接近100%,流式细胞仪检测CM-Dil标记阳性率达99.97%。实验组细胞移植后3天、5天肝脏R2*值均较移植前明显升高(t=7.282、7.608,P均0.01),对照组细胞移植后各时间点与移植前比较,R2*值差异均无统计学意义(F=0.99,P0.05)。普鲁士蓝阳性细胞主要分布于肝小叶中央静脉周围病变区,荧光显微镜下观察红色荧光阳性细胞与普鲁士蓝阳性细胞分布基本一致。结论 SPIO和CM-Dil可有效标记人永生化骨髓间充质干细胞(UE7T-13),不影响细胞增殖能力,临床应用型1.5T MR可对磁粒子标记的UE7T-13进行肝归巢活体示踪,CM-Dil有助于证实存活干细胞的肝内定植。
[Abstract]:Objective to observe the homing of human bone marrow mesenchymal stem cells (BMSCs) with magnetic particles and fluorescent dyes to acute liver injury in rats. Methods immortalized human bone marrow mesenchymal stem cells (UE7T-13) were labeled with superparamagnetic ferric oxide (SPIO) and chloromethylbenzoyl amine (CM-Dil). Intracellular iron was detected by Prussian blue staining, CM-Dil positive rate was detected by fluorescence microscope and flow cytometry. Twelve rat models of acute liver injury were established and divided into two groups: experimental group (n = 6) and control group (n = 6). The labeled cells were transplanted through the tail vein into the experimental group and the unlabeled cells were transplanted into the control group. Rat liver R2 * value was obtained by using MR T2WT2map 3 hours before transplantation and 3 days after transplantation and 5 days and 7 days after transplantation. The results were compared with Prussian blue staining and CM-Dil fluorescence expression in liver tissue sections. Results the SPIO-PLL labeling rate of double labeled cells was close to 100 and the positive rate of CM-Dil labeling was 99.97 by flow cytometry. The R2 * value of liver in the experimental group was significantly higher than that before transplantation on the 3rd day and 5th day after transplantation (t = 7.282n 7.608, P < 0.01), but there was no significant difference between the control group and the control group at all time points and before transplantation (P 0.05). Prussian blue positive cells were mainly located in the lesion area around the central vein of hepatic lobule. The distribution of red fluorescent positive cells was basically consistent with that of Prussian blue positive cells under fluorescence microscope. Conclusion SPIO and CM-Dil can effectively label human immortalized bone marrow mesenchymal stem cells (UE7T-13) and have no effect on cell proliferation. Clinical application of 1.5T MR can be used to trace live liver homing UE7T-13 labeled by magnetic particles. CM-Dil is helpful to confirm the intrahepatic colonization of viable stem cells.
【作者单位】：广州市第一人民医院放射科;中山大学附属第三医院放射科;中山大学介入放射学研究所;
【基金】：国家自然科学基金(81271621;81070349;81172193) 广东省科技攻关项目(2010B031600204)
【分类号】：R575;R445.2

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