Wang Yue, Li Xiaoli, Liu Lanlan, Liu Bingruo, Wang Feng, Chen Changsheng
Department of Ultrasound, Peking University Shenzhen Hospital, Shenzhen, 518035 People's Republic of China.
Key Laboratory of Biomedical Materials and Implant Devices, Research Institute of Tsinghua University in Shenzhen, Nanshan Hi-new Technology and Industry Park, Shenzhen, 518057 Guangzhou People's Republic of China.
Cell Mol Bioeng. 2019 Sep 4;13(1):99-112. doi: 10.1007/s12195-019-00597-w. eCollection 2020 Feb.
Ultrasound-targeted microbubble destruction (UTMD) has been shown a promising approach for target-specific gene delivery and treatment of many diseases in the past decade. To improve the therapeutic potential of UTMD, the gene carrier of microbubbles should possess adequate DNA condensation capability and (or) specific cell or tissue selectivity. The tissue-targeted and ultrasound-targeted cationic microbubbles were developed to meet gene therapy.
A tissue-targeted stearic acid-inserted cationic microbubbles (SCMBs) were prepared for ultrasound-targeted gene delivery. Branched PEI was modified with stearic acid and further mixed with 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) and biot-1,2-distearoyl-sn-glycero-3-phosphoethanolamine--[methoxy (polyethylene glycol)-2000] (ammonium salt) (Biot-DSPE-PEG2000), intercellular adhesion molecule-1 (ICAM-1) antibody and plasmid DNA to prepare cationic microbubbles through ultrasonic hydration. The ICAM-1 antibody and plasmid DNA were expected to assemble to the surface of SCMBs biotin-avidin interaction and electrostatic interaction, respectively.
It was found that the SCMBs had higher zeta potential compared with neutral microbubbles (NMBs) and cationic microbubbles (CMBs). In contrast, DNA incorporated SCMBs4 showed negative potential, exhibiting good DNA-binding capacity. Confocal images showed that the HeLa cells were attached around by the SCMBs4 from the view of green fluorescence of fluorescein isothiocyanate-loaded IgG which conjugated to ICAM-1 antibody on their surface. After ultrasound treatment, HeLa cells treated with SCMBs exhibited slightly stronger red fluorescence under confocal laser scanning microscope, indicating a synergistic promotion for transfection efficiency.
This tissue- and ultrasound-targeted cationic microbubble demonstrated here showed a promising strategy for improving gene therapy in the future.
在过去十年中，超声靶向微泡破坏（UTMD）已被证明是一种用于靶向特定基因传递和治疗多种疾病的有前景的方法。为了提高UTMD的治疗潜力，微泡的基因载体应具备足够的DNA凝聚能力和（或）特定的细胞或组织选择性。为满足基因治疗的需求，研发了组织靶向和超声靶向阳离子微泡。
制备了一种组织靶向的硬脂酸插入型阳离子微泡（SCMBs）用于超声靶向基因传递。用硬脂酸修饰支链PEI，然后将其与1,2-二硬脂酰-sn-甘油-3-磷酸胆碱（DSPC）、生物素-1,2-二硬脂酰-sn-甘油-3-磷酸乙醇胺-[甲氧基（聚乙二醇）-2000]（铵盐）（Biot-DSPE-PEG2000）、细胞间黏附分子-1（ICAM-1）抗体和质粒DNA混合，通过超声水化制备阳离子微泡。预期ICAM-1抗体和质粒DNA分别通过生物素-抗生物素蛋白相互作用和静电相互作用组装到SCMBs表面。
发现SCMBs与中性微泡（NMBs）和阳离子微泡（CMBs）相比具有更高的ζ电位。相比之下，掺入DNA的SCMBs4显示出负电位，具有良好的DNA结合能力。共聚焦图像显示，从表面与ICAM-1抗体偶联的异硫氰酸荧光素标记的IgG的绿色荧光来看，HeLa细胞被SCMBs4围绕附着。超声处理后，在共聚焦激光扫描显微镜下，用SCMBs处理的HeLa细胞显示出稍强的红色荧光，表明对转染效率有协同促进作用。
本文展示的这种组织和超声靶向阳离子微泡为未来改善基因治疗提供了一种有前景的策略。
