Activated and non-activated PAMAM dendrimers for gene delivery in vitro and in vivo
Activated and non-activated PAMAM dendrimers for gene delivery in vitro and in vivo
复制标题
DOI：
10.1016/j.nano.2008.12.007
复制
发表时间：
2009-09-01
影响因子：
5.4
通讯作者：
Tros de ILarduya, Conchita
中科院分区：
文献类型：
--
作者：
Navarro, Gemma;Tros de ILarduya, Conchita
纳米技术虽然不是一个新概念，但在医学突破中已经变得越来越重要。像聚合物纳米颗粒这样的纳米系统的制备可以用于药物和基因输送。本研究对由第4代和第5代(G4、G5)聚酰胺胺(PAMAM)树枝状大分子和质粒DNA制备的树枝状纳米粒子进行了表征，并评价了它们在体内外对细胞的转染能力。此外，这些树枝状大分子在热处理后激活的有效性已经被测试，以试图提高比完整的树枝状大分子的转染活性。颗粒大小和Zeta电位随电荷比和聚合物生成的变化的测量表明，在不同电荷比下制备的未活化树枝状聚合物中，G4或G5聚合物的粒径没有显著差异。树枝状神经丛的Zeta电位是强正性的，差别很小。原子力显微镜图像显示，它们是球形的、个性化的、分布均匀的。与裸DNA相比，即使在60%胎牛血清存在的情况下，这些载体也能有效地保护DNA免受DNase I的攻击，并在体外提高了质粒介导的基因转移到肝(HepG2)和结肠癌(CT26)癌细胞的效率。在较高的电荷比下，表达增强，在电荷比为10：1(+/-)时达到最大值，并通过增加树枝状大分子的生成量来增强表达。最后，G4和G5树突状复合体在HepG2和CT26细胞中的转染活性通过树枝状大分子的激活而显著增强。在这方面，我们优化了激活时间，以获得最佳的基因表达水平。此外，静脉注射激活的G4和G5树枝状分子-DNA复合体在体内的基因转移效率也优于未激活的树枝状大分子-DNA复合体。综上所述，我们的结果表明，G4和G5 PAMAM树枝状大分子是一种有效的纳米系统，可以在体外将基因输送到结肠癌和肝癌细胞，也可以用于体内治疗应用。
Nanotechnology, though not a new concept, has gained importance in medical breakthroughs. The preparation of nanosystems like polymeric nanoparticles can be used for drug and gene delivery. In this study dendrimeric nanoparticles prepared with generations 4 and 5 (G4, G5) polyamidoamine (PAMAM) dendrimers and plasmid DNA were characterized and their ability to transfect cells in vitro and in vivo evaluated. Additionally, the efficacy of these dendrimers on activation after heat treatment has been tested to attempt an enhancement in transfection activity over that of intact dendrimers. Measurements of the particle size and zeta potential as a function of the charge ratio and the generation of the polymer reveal that no significant differences were obtained in size by using G4 or G5 polymers in nonactivated dendriplexes prepared at different charge ratios. The zeta potentials of the dendriplexes are strongly positive and differ only slightly. Atomic force microscopy images of complexes showed that they are spherical, individualized, and homogeneously distributed. These vectors were also highly effective in protecting DNA from attack by DNase I and increased the efficiency of plasmid-mediated gene transfer in vitro to liver (HepG2) and colon (CT26) cancer cells as compared with naked DNA, even in the presence of 60% fetal bovine serum. Expression is enhanced at higher charge ratios with maximal values obtained at a charge ratio of 10: 1 (+/-) and by increasing the dendrimer generation. Finally, the transfection activity of G4 and G5 dendriplexes was significantly enhanced in HepG2 and CT26 cells by activation of the dendrimers. In this respect we have optimized the time of activation to obtain the optimal levels of gene expression. Also, intravenously administered activated G4 and G5 dendrimer-DNA complexes are superior to the nonactivated ones in terms of gene transfer efficiency in vivo. In conclusion, our results showed that G4 and G5 PAMAM dendrimers are an effective nanosystem for gene delivery to colon and liver cancer cells in vitro, as well as for in vivo therapeutic applications.