胍基和咪唑修饰的氨基聚甲基丙烯酸缩水甘油酯衍生物作为基因载体的构建及性能研究

发布时间：2018-12-20 15:03

【摘要】：近年来,拥有不同组分和生物性质的纳米载体已经广泛的应用于体外或体内的药物运输和基因传递。纳米载体家族包括聚合物纳米粒子、基于脂质体的载体、树枝状大分子、碳纳米管和金颗粒。在这些不同的体系中,聚合物载体有以下多种性质:合成简单、便宜、生物相容性好、生物可降解、无毒无免疫原性、水溶性好。另外,与阳离子脂质体相比,阳离子聚合物能够形成更加稳定的复合体,在细胞内转运过程中能够起到更好的保护作用。在实际应用方面,较高的转染效率和较低的细胞毒性对于基因载体系统至关重要。在过去的十年中,聚缩水甘油和其结构类似物在生物领域引起了人们极大的关注。在我们此次的研究中,首先通过原子自由基转移聚合(ATRP)反应合成了聚甲基丙烯酸缩水甘油酯(PGMA)。然后通过开环反应向侧链引入带有阳离子的乙二胺(EDA)得到EP。接下来将EP按顺序分别进行胍基功能化修饰得到GEP和希夫碱连接的咪唑功能化修饰得到IGEP。我们用核磁共振氢谱和傅里叶红外光谱对这几种合成的高分子的结构进行了表征。使用凝胶渗透色谱对高分子的分子量和分散性指数进行了测试,并对其进行了元素分析来确定高分子的氨基含量。接下来我们分别用3种高分子与DNA相复合,构建复合体,并对这些复合体作为基因载体的一些性能进行了测试。测试表明PGMA衍生物能够有效地与pDNA结合,形成最佳粒径为100nm的纳米粒子,表面电势为10-30mV,使用电子扫描显微镜对所形成的纳米粒子的形态进行了表征。在生物策略方面,我们选用了胍基。因为胍基能够促进载体与细胞膜的相互作用,所以能够提高细胞摄取效率,并且它还具有细胞核定位功能。希夫碱连接的咪唑有助于核内体逃逸和促进DNA释放,提高靶向细胞内的转染效率。值得注意的是,胍基和咪唑能够降低细胞毒性。因此,这些优点为构建能够应用于实际的、安全的、有效的基因载体提供了大量信息。
[Abstract]:In recent years, nano-carriers with different components and biological properties have been widely used in drug transport and gene transmission in vitro or in vivo. The nanocarrier family includes polymer nanoparticles, liposome-based carriers, dendritic macromolecules, carbon nanotubes and gold particles. In these different systems, the polymer carriers have the following properties: simple, inexpensive, biocompatible, biodegradable, non-toxic, non-immunogenicity and water-soluble. In addition, compared with cationic liposomes, cationic polymers can form more stable complexes and play a better protective role in intracellular transport. In practical application, higher transfection efficiency and lower cytotoxicity are very important for gene vector system. In the past decade, polyglycidyl and its structural analogues have attracted great attention in the biological field. In our study, we first synthesized polyglycidyl methacrylate (PGMA).) by atom free radical transfer polymerization (ATRP) reaction. Then EP. was obtained by introducing ethylenediamine (EDA) with cations into the side chain by ring-opening reaction. Next, EP was modified by guanidine functionalization in order to get GEP and Schiff base bound imidazole functionalized modification to obtain IGEP.. The structures of the synthesized polymers were characterized by NMR and Fourier transform infrared spectroscopy (FTIR). The molecular weight and dispersity index of the polymer were measured by gel permeation chromatography, and the amino content of the polymer was determined by elemental analysis. Then we used three kinds of polymers to combine with DNA to construct the complexes, and some properties of these complexes as gene vectors were tested. The results show that the PGMA derivatives can effectively combine with pDNA to form the best 100nm nanoparticles with a surface potential of 10-30mV. The morphology of the nanoparticles was characterized by electron scanning microscopy (SEM). In biological strategy, we chose guanidine. Because guanidinyl can promote the interaction between the carrier and cell membrane, it can improve the efficiency of cell uptake, and it also has the function of nuclear localization. Schiff alkali-linked imidazole can promote the release of DNA and enhance the transfection efficiency of target cells. It is worth noting that guanidine and imidazole can reduce cytotoxicity. Therefore, these advantages provide a large amount of information for the construction of practical, safe and effective gene vectors.
【学位授予单位】：天津理工大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R450

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