文献：Echogenic PEGylated PEI-Loaded Microbubble As Efficient Gene Delivery System

文献链接：https://www.semanticscholar.org/paper/Echogenic-PEGylated-PEI-Loaded-Microbubble-As-Gene-Liufu-Li/c9d0e576c0bdc2f62da99a5c4c6fd5a0ed6e4e8a

作者：Chun Liufu,Yue Li,Jiawei Tu,Hui Zhang, Jinsui Yu,Yi Wang,Pintong Huang, Zhiyi Chen

相关产品：

DSPE-PEG2000-Biotin  二硬脂酰磷脂酰乙醇胺 - 聚乙二醇 2000 - 生物素

Biotin-PEG(2k)-SS-NHS  生物素 - 聚乙二醇 2000- 二硫键 - 活性酯

原文摘要：Background: Cancer stem cells (CSCs) are responsible for cancer therapeuticresistance and metastasis. To date, in addition to surgery, chemotherapy, and radiotherapy, gene delivery has emerged as a potential therapeutic modality for ovarian cancer. Efficient and safe targeted gene delivery is complicated due to the tumor heterogeneity barrier. Ultrasound (US)-stimulated microbubbles (MBs) have demonstrated a method of enabling non-invasive targeted gene delivery.

Purpose: The purpose of our study was to show the utility of poly(ethylene glycol)-SSpolyethylenimine-loaded microbubbles (PSP@MB) as an ultrasound theranostic and redoxresponsive agent in a gene delivery system. Patients and methods: PSP nanoparticles were conjugated to the MB surface through biotin–avidin linkage, increasing the gene-loading efficiency of MB. The significant increase in the release of genes from the PSP@MB complexes was achieved upon ultrasound exposure. The positive surface charge in PSP@MB can condense the plasmid through electrostatic interactions; agarose-gel electrophoresis further confirmed the ability of PSP@MB to condense plasmids. The morphology, particle sizes and zeta potential of PSP@MB were characterized by transmission electron microscopy and dynamic light scattering. Results: Laser confocal microscopy showed that the combination of ultrasound with PSP@MB could promote the cellular uptake of plasmids. Plasmids which encode enhanced green fluorescence protein (EGFP) reporter genes or luciferase reporter genes were delivered to CSCs in vitro and to subcutaneous xenografts in vivo via the combination of ultrasound with PSP@MB. Gene transfection efficiency was evaluated by fluorescence microscopy and In Vivo Imaging Systems. This study demonstrated that the combination of ultrasound with PSP@MB can remarkably promote gene delivery to solid tumors as well as diminishing the toxicity towards normal tissues in vivo. The combination of PSP@MB and the use of ultrasound can efficiently enhance accumulation, extravasation and penetration into solid tumors.

Conclusion: Taken together, our study showed that this novel PSP@MB and ultrasoundmediated gene delivery system could efficiently target CSCs.

DSPE-PEG2000-Biotin结合了 PEG 的水溶性和生物相容性、DSPE 的细胞膜亲和性以及生物素的靶向功能，可作为纳米化合物载体。能与化合物结合，实现化合物的靶向传递，提高效果并减少副作用。Biotin-PEG(2k)-SS-NHS可用于构建具有刺激响应性的化合物递送系统。在正常生理环境下，二硫键保持稳定，确保化合物在运输过程中的稳定性；当到达特定的还原环境时，二硫键断裂，实现化合物的靶向释放。PSP纳米颗粒通过生物素-亲和素连接偶联到MB表面，提高了MB的基因装载效率。其具体应用如下：

图为：PSP@MB的构造示意图

Biotin-PEG(2k)-SS-NHS在PSP NPs合成与表征中的应用：

生物素化的PSP是由PEI和Biotin-PEG(2k)-SS-NHS通过酰胺键构建的。通过PSP在D2O中的1 H NMR谱，证实了PSP的成功合成。在D2O中PSP的1 H NMR谱中发现了“a”中的PEG和“b”中的PEI的信号。动态光散射（DLS）测量结果表明，PSP NPs的平均直径由于聚乙二醇化作用，有轻微的正zeta电位。为了评价还原敏感性，将PSP NPs与 GSH孵育。GSH处理后，PSP NPs的尺寸增加，表面电荷增加。

DSPE-PEG-Biotin在MB和PSP@MB合成与表征中的应用：

为了将生物素化的PSP NPs装载生物素化的脂质MBs，使用了生物素-亲和素法。为此，DSPE-PEG-Biotin功能化的脂质被纳入MB壳层中。用薄膜水化法合成了生物素化的MBs。FITC-链霉亲和素与PSP NPs和MBs偶联。荧光显微镜证实了PSP负载的MBs的成功连接。DLS测量结果显示，MB和PSP@MB的透射电镜（TEM）图像显示与DLS的粒径相似。

图为：PSP、MB和PSP@MB的表征

结论：Biotin-PEG (2k)-SS-NHS 中的 NHS（N - 羟基琥珀酰亚胺）基团是一种高活性的酯类基团，可以与含有氨基的分子发生酰胺化反应。在 PSP NPs 的合成过程中，如果纳米颗粒表面或相关分子中存在氨基，那么该试剂可以通过 NHS 基团与之反应，从而将Biotin-PEG (2k)-SS部分连接到纳米颗粒上。通过测量含有 Biotin-PEG (2k)-SS-NHS 修饰的 PSP NPs 的粒径、多分散性指数和表面电位等参数，可以了解纳米颗粒的大小分布和表面电荷情况。这些参数对于评估纳米颗粒的稳定性、生物分布和细胞摄取等性能非常重要。在 MB 的合成过程中，DSPE-PEG-Biotin 可以作为一种表面修饰剂，通过与微泡的磷脂膜相互作用，将生物素和聚乙二醇（PEG）部分连接到微泡表面。含有生物素标记的 MB 和 PSP@MB 可以用于体外细胞实验和体内动物实验，评估微泡的生物学性能。