肽功能化载GOD智能响应型相变纳米粒用于乳腺癌超声诊疗的实验研究  

Peptide-functionalized glucose oxidase-loaded phase-transformation and smart responsive nanoparticles for ultrasound imaging and breast cancer treatment

作　　者：谢卓晏 罗远利 乔斌 李锦瑞 周志益 王志刚[2] 任建丽[1] Xie Zhuoyan;Luo Yuanli;Qiao Bin;Li Jinrui;Zhou Zhiyi;Wang Zhigang;Ren Jianli(Department of Ultrasonography,the Second Affiliated Hospital of Chongqing Medical University,Chongqing,400010,China;Chongqing Key Laboratory of Ultrasound Molecular Imaging,Institute of Ultrasound Imaging,Chongqing Medical University,Chongqing,400016,China;Department of Geriatrics,Chongqing People's Hospital,Chongqing 400013,China)

机构地区：[1]重庆医科大学附属第二医院超声科,400010 [2]重庆医科大学超声影像学研究所超声分子影像重庆市重点实验室,400016 [3]重庆市人民医院老年病科,400013

出　　处：《中华医学超声杂志（电子版）》2022年第8期837-846,共10页Chinese Journal of Medical Ultrasound(Electronic Edition)

基　　金：重庆市渝中区基础研究与前沿探索项目(20190112);国家自然科学基金面上项目(81873901);重庆市自然科学基金重点项目(cstc2019jcyj-zdxmX2020);重庆市中青年医学高端人才培养项目;宽仁骨干人才项目(KR2019G001)。

摘　　要：目的制备一种肿瘤归巢穿膜肽tLyp-1介导的载葡萄糖氧化酶(GOD)和全氟正戊烷(PFP)纳米粒(tLyp-1-GOD@PFP NPs),观察其基本表征、体内外超声成像以及对人乳腺癌MDAMB-231细胞的体外靶向及体内外抗肿瘤能力。方法采用乳化法制备tLyp-1-GOD@PFP NPs,观察其形态特点,检测其理化性质(粒径、电位、包封率、释药率);低强度聚焦超声(LIFU)辐照纳米粒后,光镜下观察其相变情况,并观察其体内外超声成像效果;激光共聚焦显微镜和流式细胞仪检测纳米粒对MDA-MB-231细胞的靶向能力;使用CCK-8法和流式法评估在LIFU辐照下该纳米粒对MDA-MB-231细胞的抗肿瘤能力;通过荷瘤裸鼠尾静脉注射纳米粒并观察肿瘤体积变化和裸鼠体重以探索其体内抑瘤效果。结果制备的纳米粒呈球形壳核结构,大小均一,其平均粒径为(227.4±12.5)nm,平均电位为(-16.5±2.7)mV,GOD的包封率为(12.93±0.46)%,载药率为(1.62±0.06)%,24 h释药率可达(51.73±3.22)%。光镜下相变和体外超声成像具有辐照时间依赖性。CCK-8和流式细胞仪检测结果证明tLyp-1-GOD@PFP NPs具有良好的生物安全性,并且在LIFU辐照后其抑瘤效果较好。同时,体内实验证实该纳米粒具有良好的超声造影能力和靶向能力,体内治疗结果也表明该纳米粒可有效抑制肿瘤增殖。结论本研究成功制备了tLyp-1介导的载GOD和PFP的相变型纳米粒,对MDA-MB-231细胞具有特异靶向能力,在LIFU辐照下,可实现体内外超声成像,同时产生良好的抗肿瘤效果。Objective To prepare tumor-homing tLyp-1-mediated nanoparticles loaded with glucose oxidase(GOD)and perfluoron-pentane(PFP)(TLyp-1-GOD@PFP NPs),to evaluate their physicochemical properties,ability of targeting,and antitumor efficacy in human breast cancer MDA-MB-231 cells under irradiation by low intensity focused ultrasound(LIFU),and to explore their use in ultrasound imaging in vitro and vivo.Methods TLyp-1-GOD@PFP NPs were prepared by a phacoemulsification method.Their characteristics including particle size,potential,drug release rate,and entrapment efficiency were detected.The phase-change of the nanoparticles irradiated by LIFU was observed under an optical microscope.Meanwhile,their use in ultrasound imaging in vitro and vivo was explored.The targeting efficacy of tLyp-1-GOD@PFP NPs in MDA-MB-231 cells was evaluated by laser confocal microscopy and flow cytometry.Besides,CCK-8 assay was applied to detect the cytotoxicity of the nanoparticles.Flow cytometry was also carried out to measure the percentage of apoptotic cells in all groups.In order to explore their antitumor effect in vivo,the nanoparticles were injected into the tail vein of nude mice bearing heterotopic xenografts of MDA-MB-231 cells and then,the changes of tumor volume and weight of nude mice were observed.Results TLyp-1-GOD@PFP NPs were successfully prepared,and showed spherical shell-core structure in shape and were uniform in size,with an average size of(227.4±12.5)nm,at an average potential of(-16.5±2.7)mV.The entrapment efficiency and the drug loading rate of GOD were(12.93±0.46)%and(1.62±0.06)%,respectively.The drug release rate under irradiation could reach(51.73±3.22)%at 24 h.Phase-change of the nanoparticles observed under microscope and ultrasound imaging in vitro were time-dependent.The results of CCK-8 assay and flow cytometry exhibited the excellent biosafety and proved that the tLyp-1-GOD@PFP NPs with LIFU enhanced the inhibition proliferation.In the meantime,the contrast-enhanced ultrasound ability and targeting ability o

关 键 词：肿瘤归巢穿膜肽 葡萄糖氧化酶 相变纳米粒 乳腺癌 超声诊疗 

分 类 号：R445.1[医药卫生—影像医学与核医学] R737.9[医药卫生—诊断学]