外磁场下磁性纳米C3转移酶载体在大鼠损伤脊髓中的分布

Target distribution of magnetic nano C3-transferase carrier under external magnetic field in injured spinal cord in rats

目的:观察施加外磁场后磁性纳米C3转移酶药物载体(简称载药微球)在大鼠损伤脊髓局部的分布情况,并观察不同时间作用的外磁场对该载体分布的影响。方法:构建载药微球,检测其粒径、Zeta电位,透射电镜观察形态、测定磁场顺应性及药物释放;MTT法测定其细胞毒性;观察体外培养条件下细胞的摄取情况。82只大鼠建立T10损伤模型(NYU法),随机分为5组:A组,经尾静脉注射异硫氰酸荧光素组,20只;B组,经尾静脉注射载药微球组,20只;C组,经尾静脉注射载药微球+外磁场15min组,20只;D组,经尾静脉注射载药微球+外磁场30min组,12只;E组,经尾静脉注射载药微球+外磁场1h组,10只。A、B、C组各取10只大鼠,经尾静脉注射1h后,取肝、肾、脾及T10为中心的脊髓组织做冰冻切片并观察载药微球在其中的分布;5组各10只大鼠经尾静脉注射1h后取T10为中心4cm脊髓组织,火焰原子吸收分光光度法测定铁含量;D组取2只大鼠,电镜观察载药微球在脊髓组织中分布。结果:载药微球分散性好,载药微球磁化强度饱和度值为63.5emu/g,载药微球缓慢释放药物且释药时间超过9d,载药微球与细胞共培养,细胞平均存活率为78.10%,与细胞共培养5s后能够在细胞内达到良好聚集效果。C组脊髓损伤中心荧光强度高于A、B组。C组脊髓铁元素含量高于A、B组,D组测定铁含量高于C组(P<0.05),E组测定铁含量高于C组(P<0.05),D组测定铁含量与E组无统计学意义(P>0.05)。电镜观察载药微球聚集在损伤中心,可进入神经细胞胞体内。结论:磁性纳米C3转移酶药物载体可靶向聚集在损伤区局部,载药微球可进入脊髓损伤区神经细胞内;损伤后施加外磁场30min,载药微球可达到最佳聚集效果。

Objectives： To observe the target distribution of magnetic nano C3-transferase carrier under external magnetic field in injured spinal cord and explore this time-related effect.Methods： Drug carrier microsphere was constructed;the particle size,zeta potential,magnetic properties,drug release were assessed;the morphology was observed by transmission electron microscopy;the in vitro cytotoxicity was tested either;and the cellular uptake was analyzed using fluorescence microscopy.Eight-two SCI rats were divided into 5 groups randomly： Group A（n=20）,fluorescein isothiocyanate group（FITC） administered by caudal vein,20 rats;Group B（n=20）,drug carrier microsphere administered by caudal vein;Group C（n=20）,drug carrier microsphere administered by caudal vein and external magnetic 15min;Group D（n=12）,drug carrier microsphere administered by caudal vein and external magnetic 30min;Group E（n=10）,drug carrier microsphere administered by caudal vein and external magnetic 1h.10 rats in Group A,B and C respectively were selected,and the tissues including liver,kindey,spleen and T10 spinal cord were harvested 1h after administration by caudal vein,then the liposome distribution in these tissues was observed under fluorescence microscopy.10 rats in Group A,B,C,D and E respectively were sacrificed and the T10 spinal cord was harvested 1h after administration by caudal vein,and the iron contents were assayed by flame atomic absorption spectrophotometry.2 rats in Group D sacrificed and the T10 spinal cord was harvested 1h after administration by caudal vein,and the distribution of drug carrier microsphere was observed under electron microscope.Results： Drug carrier microspheres showed good dispersion,with the saturation value of magnetization of 63.5emu g,and the carrier could maintain drug realease in vitro until day 9;when co-culturing with MCF-7 cells,the average cell survival rate was 78.10%;FITC fluorescence density of Group C in injured center was more than group A and B.The iron contents in spinal cord of Group C showed significant difference than Group A and B（P〈0.05）,and those in group E were more than group C（P〈0.05）,while those in group D showed no difference than group C.The carriers were noted gathering in the center of injury and into neurocyte under electron microscopy.Conclusions： Magnetic nano C3-transferase drug carriers can accumulate into injured site of spine cord and enter neurocyte;adding external magnetic field of 30min after SCI can achieve optimal effect for aggregation.