内皮祖细胞靶向归巢的双模态成像活体示踪研究  

Dual-modality imaging in vivo tracking for endothelial progenitor cells homing

作　　者：陈龙 董兵 刘晓玲 何江[2,3] 余冰波 罗丽芳 夏文豪 Chen Long;Dong Bing;Liu Xiaolin;He Jiang;Yu Bingbo;Luo Lifang;Xia Wenhao(The International Medical Department of Shenzhen Hospital,Southern Medical University,Shenzhen 510086,China;Department of Hypertension and Vascular Disease,the First Affiliated Hospital of Sun Yat-Sen University,Guangzhou 510080,China;Key Laboratory on Assisted Circulation Ministry of Health,Guangzhou 510080,China;Department of Dermatology,the First Affiliated Hospital of Guangdong Pharmaceutical University,Guangzhou 510080,China)

机构地区：[1]南方医科大学深圳医院国际医学部,深圳510086 [2]中山大学附属第一医院高血压血管病科,深圳510080 [3]国家卫生健康委员会辅助循环重点实验室,深圳510080 [4]广东药科大学附属第一医院皮肤性病科,深圳510080

出　　处：《中华细胞与干细胞杂志（电子版）》2021年第6期351-357,共7页Chinese Journal of Cell and Stem Cell（Electronic Edition）

基　　金：国家自然科学基金(81671379,81670226);广东省自然科学基金(2020A1515011264,2017A030313823)。

摘　　要：目的整合磁共振成像和光学成像的优势,采用双模态成像活体动态观察内皮祖细胞(EPCs)移植后靶向归巢到血管内皮损伤局部的生物学过程。方法利用化学法合成负载磁共振显影剂SPIO和近红外染料Cy7.5的PEG-PEI纳米载体建立裸大鼠颈动脉内膜损伤模型,随机分为空载体对照组(PEG-PEI组)和实验组(PEG-PEI-SPIO组),体外分别通过动态光散射实验检测其纳米粒子的粒径、电位,CCK-8实验和Annexin V流式实验检测其细胞毒性;体内借助磁共振成像和活体荧光成像动态观察大鼠EPCs尾静脉移植后靶向归巢到血管内皮损伤局部的过程,两组间比较采用t检验。结果动态光散射实验表明:当N/P=12时,PEG-PEI-SPIO纳米粒子与质粒复合后的粒径为(129.61±1.05)nm,表面电位为2.41 mV;电镜检测结果发现:复合后的纳米粒子呈现形态一致的圆形,SPIO位于复合体系中心,复合体系的粒径在(130.06±1.21)nm;CCK-8实验和Annexin V流式实验表明:当N/P=12时,PEG-PEI组与PEG-PEI-SPIO组纳米载体的增殖毒性和凋亡毒性差异无统计学意义;普鲁士蓝染色显示:PEG-PEI-SPIO纳米粒子可被EPCs吞至细胞质中。活体荧光显像结果显示:Cy7.5标记的EPCs能够在移植后2 d内归巢到血管内皮损伤处(高荧光信号强度);磁共振显像结果显示:SPIO标记的EPCs移植后2 d内到达损伤部位血管内皮下(MRI T_(2)强度下降)。结论纳米载体PEG-PEI-SPIO具有很好的质粒复合功能、稳定的形态学特征、低细胞毒性以及良好的活体可视化功能,可有效整合磁共振成像和光学成像在空间分辨率、敏感性、特异性以及定量分析等各方面的优势,活体示踪EPCs在体移植后的靶向归巢过程,对活体评价干细胞移植疗效,优化移植策略提供客观指标和理论指导。Objective Integrated magnetic resonance and optical imaging to create dynamically dual-modality imaging to observe the biological process of endothelial progenitor cells(EPCs)homing to the local vascular endothelial injury after transplanted.Methods Chemically synthesized PEG-PEI nanocarriers loaded with magnetic resonance imaging agent SPIO and near-infrared dye Cy7.5 were randomly divided into an empty carrier control group(PEG-PEI group)and an experimental group(PEG-PEI-SPIO group);The particle size and potential of the nanoparticles were measured by dynamic light scattering experiment in vitro,CCK-8 experiment and Annexin V flow cytometry;A nude rat carotid artery intima injury model was established,and the tail vein of EPCs was dynamically observed by magnetic resonance imaging and intravital fluorescence imaging After transplantation,the process of homing to the local vascular endothelial injury is targeted.t test was used for comparison between two groups.Results Dynamic light scattering experiments show that the particle size of PEG-PEI-SPIO nanoparticles is 129.6 nm,and the surface potential is 2.4 mV when N/P=12;electron microscopy Results show that the composite nanoparticles has circles shape,SPIO is located in the center of the composite system,and the particle size of the composite system is about 130 nm;CCK-8 experiment and Annexin V flow cytometry experiment show that when N/P=12,compared to the nanocarrier PEG-PEI control group,the proliferation toxicity and apoptosis toxicity of PEG-PEI-SPIO nanocarriers has no statistic differences;Prussian blue staining shows that PEG-PEI-SPIO nanoparticles can be endothelialized into the cytoplasm.In vivo fluorescence imaging Results showed that Cy7.5-labeled EPCs home to the vascular endothelial injury within 2 days after transplantation(increased fluorescence signal intensity);MRI Results showed that SPIO-labeled EPCs reached the sub-endothelium injury location within 2 days after transplantation(the intensity of MRI T_(2) decreased).Conclusion PEG-PEI nano

关 键 词：内皮祖细胞 血管损伤 双模态成像 活体示踪 

分 类 号：R457.7[医药卫生—治疗学] R-332[医药卫生—临床医学]