制备用于间充质干细胞成像的荧光成像/磁共振成像/正电子发射断层扫描三模态纳米探针

Preparation of a Fluorescence Imaging/Magnetic Resonance Imaging/Positron Emission Tomography Trimodal Nanoprobe for Mesenchymal Stem Cell Imaging

为了在间充质干细胞移植的各个阶段更好地获取移植干细胞的命运信息,制备了一种能同时实现荧光成像(FI)、磁共振成像(MRI)和正电子发射断层扫描(PET)的影像探针。首先制备了一种新型两亲性嵌段共聚物,第1嵌段为叔丁氧羰基保护的2-((叔丁氧羰基)氨基)丙烯酸乙酯的均聚物,第2嵌段为短链聚乙二醇丙烯酸酯和五氟苯酚丙烯酸酯的无规共聚物,再通过氨解和酰胺化反应为聚合物修饰上双功能螯合剂DOTA和近红外染料Cy5.5,通过核磁共振和凝胶渗透色谱对聚合物结构及相对分子质量进行了表征和分析。聚合物在水中自组装并螯合Gd^(3+)得到1 mg/mL的纳米探针Gd-DOTA-Cy5.5-NP,动态光散射和透射电镜观察到探针为平均粒径30 nm的球形颗粒,并用电感耦合等离子体光谱法测得Gd^(3+)浓度为0.084 mmol/L。将纳米探针与间充质干细胞共孵育,通过Cell Counting Kit-8试剂验证1 mg/mL以下的Gd-DOTA-Cy5.5-NP相对细胞毒性较低,同时激光共聚焦显微镜在675 nm激发波长下观测细胞出现红色荧光,利用3.0 T磁共振成像系统测得探针纵向弛豫率为46.969 mmol/(L·s)。最后Gd-DOTA-Cy5.5-NP螯合^(68)Ga^(3+),得到三模态纳米探针^(68)Ga/Gd-DOTA-Cy5.5-NP,伽马计数仪测得间充质干细胞对该探针的摄取在90 min达到饱和。

To track transplanted mesenchymal stem cells,a nanoprobe that can simultaneously realize fluorescence imaging(FI),magnetic resonance imaging(MRI),and positron emission tomography(PET)was prepared.The synthesis started from an amphiphilic diblock copolymer with the first block of poly-2-((tert-butoxycarbonyl)amino)ethyl acrylate homopolymer,and the second block of a random copolymer of poly(ethylene glycol)acrylate and pentafluorophenyl acrylate.The obtained copolymer was modified with bifunctional chelator DOTA and near-infrared dye Cy5.5 by ammoniation and amination reactions.Then the copolymers self-assembled in water and chelated ^(68)Ga^(3+)/Gd^(3+)to form trimodal ^(68)Ga/Gd-DOTA-Cy5.5-NP nanoprobes of 30 nm in diameter.In cytotoxicity experiments,^(68)Ga/Gd-DOTA-Cy5.5-NP under 1 mg/mL shows low toxicity to mesenchymal stem cells.Meanwhile,the nanoprobes exhibit a significant red fluorescence in 675 nm excitation,a notable high longitudinal relaxivity(r11)46.969 mmol/(L·s)in magnetic resonance imaging(MRI),and reach saturated uptake within 90 min in positron emission tomography(PET).