靶向载长春新碱超声微泡的制备及体外寻靶实验研究  

Preparation and in Vitro Targeting Experiment of Vincristine-Loaded Ultrasound Microbubbles

作　　者：杨红 陈秋宏 黄娟 郭婧仪 王昱媛 吴奇 YANG Hong;CHEN Qiuhong;HUANG Juan;GUO Jingyi;WANG Yuyuan;WU Qi(Department of Cardiovascular Medicine,the Second Affiliated Hospital of Chengdu Medical College/Nuclear Industry 416 Hospital,Chengdu 610057,China;Chengdu Medical College,Chengdu 610500,China)

机构地区：[1]成都医学院第二附属医院核工业四一六医院心血管内科,成都610057 [2]成都医学院,成都610500

出　　处：《转化医学杂志》2024年第2期169-176,共8页Translational Medicine Journal

基　　金：四川省中医药管理局科研项目(2023MS177);成都市科技厅技术创新研发项目(2022-YF05-01459-SN);成都市医学重点专科(CDS2022Z076)。

摘　　要：目的 制备载长春新碱(VCR)且靶向结合外周神经髓鞘细胞的超声脂质微泡,观察其体外寻靶情况。方法 采用薄膜水化-机械振荡法制作载药超声微泡(VCR-MBs),生物素-亲和素桥接法制作靶向载药超声微泡(VCR-TMBs),观察VCR-TMBs形态、粒径、稳定性、体外显影、包封率及靶向性等理化性质。流式细胞仪检测VCR-TMBs对细胞的损伤及凋亡作用,体外通过VCR-TMBs联合超声(US)对大鼠肾动脉处理,HE染色观察脱髓鞘改变。结果 MBs、VCR-MBs及VCR-TMBs光镜下均成圆形,分布均匀。MBs、VCR-MBs及VCR-TMBs粒径分别为(3.22±1.07)、(3.48±1.06)及(3.22±0.79)μm,差异无统计学意义(P>0.05);与MBs、VCR-MBs相比,VCR-TMBs保存3 d时浓度明显下降(P<0.05);采用高效液相色谱法测定VCR-TMBs的包封率及载药率分别为(82.90±9.98)%、(2.07±0.25)%;免疫荧光法测定VCR-TMBs可靶向聚集在Schwann细胞膜上;流式细胞仪检测示VCR-TMBs+US促细胞凋亡作用优于单纯VCR+US及VCR-MBs+US(P<0.05);组织HE染色示VCR-TMBs联合US可有效使体外大鼠肾动脉神经脱髓鞘及变性坏死。结论 成功制备VCR-TMBs,其可靶向识别Schwann细胞,促进细胞凋亡,使大鼠肾动脉神经脱髓鞘改变,可为后期体内VCR-TMBs实现肾动脉周围神经去交感化做前期准备。Objective To prepare vincristine(VCR)-loaded lipid ultrasound(US)microbubbles(VCR-MBs)targeting to peripheral nerve myelin cells,and to observe the in vitro targeting experiment.Methods VCR-MBs were prepared by thin film hydration and mechanical oscillation method,and targeted VCR-MBs(VCR-TMBs)was prepared by biotin-avidin bridging method.The morphology,particle size,stability,in vitro development,encapsulation rate and targeting properties of VCR-TMBs were observed.The effect of VCR-TMBs on cell injury and apoptosis was detected by flow cytometry.The rat renal artery was treated by VCR-TMBs combined with US in vitro,and demyelination was observed by HE staining.Results MBs,VCR-MBs and VCR-TMBs were all circular and evenly distributed under light microscope.The particle sizes of MBs,VCR-MBs and VCR-TMBs were(3.22±1.07),(3.48±1.06)and(3.22±0.79)μm,respectively,with no significant difference(P>0.05).Compared with MBs and VCR-MBs,the concentration of VCR-TMBs decreased significantly after 3 days of storage(P<0.05).The encapsulation rate and drug loading rate of VCR-TMBs determined by high-performance liquid chromatography were(82.90±9.98)%and(2.07±0.25)%,respectively.VCR-TMBs was detected by immunofluorescence and could be targeted to accumulate on Schwann cell membrane.Flow cytometry showed that VCR-TMBS+US had better apoptotic effect than VCR+US and VCR-MBS+US(P<0.05).HE staining showed that VCR-TMBs+US could effectively demyelinate and denaturate the renal artery nerve in vitro.Conclusion The successful preparation of VCR-TMBs can target Schwann cells,promote cell apoptosis and demyelinate renal artery nerves of rats,which can be a preliminary preparation for the later realization of renal artery nerve dessympathization by VCR-TMBs in vivo.

关 键 词：长春新碱 靶向载药超声微泡 靶向识别 SCHWANN细胞 肾去交感化 色谱法 高压液相 细胞凋亡 

分 类 号：R9-33[医药卫生—药学]