载ANM33双靶标脂质超声微泡精准胞内释药可行性的体外研究  

An in vitro study on the feasibility of precise intracellular drug release from dual-targeted lipid ultrasound microbubbles loaded with ANM33

作　　者：刘芮宁 陈尚珂 拜合提亚·塔依尔 关丽娜[1,2] 穆玉明 Liu Ruining;Chen Shangke;Baihetiya·Tayier;Guan Lina;Mu Yuming(Department of Echocardiography,the First Affiliated Hospital of Xinjiang Medical University,Urumqi 830000,China;Xinjiang Key Laboratory of Ultrasound Medicine,Urumqi 830000,China)

机构地区：[1]新疆医科大学第一附属医院心脏超声诊断科,乌鲁木齐830000 [2]新疆超声医学重点实验室,乌鲁木齐830000

出　　处：《中华超声影像学杂志》2024年第7期617-625,共9页Chinese Journal of Ultrasonography

基　　金：国家自然科学基金(82060321);新疆维吾尔自治区自然科学基金(2022D01D67,2022D01C491);中国中央政府引导地方科技发展专项资金(ZYYD2023A02)。

摘　　要：目的制备载miR-33反义核苷酸(ANM33)的双靶标脂质超声微泡(HA-PANBs),体外评价其分阶段靶向泡沫细胞并实现细胞内精准释药的可行性。方法设计以纳米泡NBs为微泡核心,透明质酸(hyaluronic acid,HA)第一阶段靶向受损内皮细胞,适配子PM1第二阶段靶向泡沫细胞,ANM33为治疗因子的双靶标脂质超声微泡。完成脂质泡表征的同时检测其稳定性及体外造影能力;随后建立受损人脐静脉内皮细胞(HUVEC)和巨噬细胞(RAW264.7,MΦ)共培养模型,结合流式细胞术、油红O染色和小动物活体成像仪等评价HA-PANBs体外精准定位泡沫细胞并释放ANM33的能力。结果制备的双靶标脂质超声微泡HA-PANBs形态规则,稳定性好,粒径为(1357.53±140.20)nm,表面电位为(-5.61±0.73)mV,HA、PM1及ANM33均有效连接。在受损HUVEC/MΦ共培养模型中,HA-PANBs靶向下层泡沫细胞的效果最佳,且有效摄取量高达(80.65±2.12)%,较NBs组高56.74%;油红O染色显示HA-PANBs组泡沫细胞的胆固醇外排能力明显优于NBs组,差异有统计学意义[(629.80±21.99)a.u.比(1071.00±55.49)a.u.,P<0.05]。结论构建的双靶标脂质超声微泡HA-PANBs能精准靶向泡沫细胞并显著增强其胆固醇外排能力,为实现动脉粥样硬化的早期无创诊疗提供了新策略。Objective To prepare dual-targeted lipid ultrasound microbubbles loaded with ANM33(HA-PANBs)and evaluate its feasibility in targeting foam cells by stages and achieving precise intracellular drug release in vitro.Methods The dual-targeteded lipid ultrasound microbubbles were designed with nanobubbles(NBs)as the microbubble core,hyaluronic acid(HA)as the first-stage targeting ligand for damaged endothelial cells,aptamer PM1 as the second-stage targeting moiety for foam cells,and ANM33 as the therapeutic factor.Simultaneously with the characterization of the lipid bubbles,the stability and in vitro contrast-enhanced ultrasound imaging capability were detected.Then a co-culture model of damaged human umbilical vein endothelial cells(HUVEC)and macrophages(RAW264.7,MΦ)was established,combined with flow cytometry,oil red O staining and small animal in vivo imaging to evaluate the ability of HA-PANBs in targeting foam cells precisely and releasing ANM33.Results The HA-PANBs exhibited regular morphology and good structural stability,with a particle size of(1357.53±140.20)nm and a surface potential of(-5.61±0.73)mV.HA,PM1 and ANM33 were effectively connected.In the damaged HUVEC/MΦco-culture system,the HA-PANBs group demonstrated the best targeting effect on foam cells,with an effective uptake of(80.65±2.12)%,which was 56.74%higher than that of the NBs group.Oil red O staining revealed that the cholesterol efflux capacity of foam cells in the HA-PANBs group was significantly better than that in the NBs group,the results were statistically different[(629.80±21.99)a.u.vs(1071.00±55.49)a.u.,P<0.05].Conclusions The dual-targeted lipid ultrasound microbubbles(HA-PANBs)can precisely target foam cells and significantly enhance their cholesterol efflux,providing a new strategy for the early non-invasive diagnosis and treatment of atherosclerosis.

关 键 词：动脉粥样硬化 载ANM33双靶标脂质超声微泡 巨噬细胞 精准释药 

分 类 号：R943[医药卫生—药剂学]