Lynch J, Behan N, Birkinshaw Colin
Department of Materials Science, University of Limerick, Limerick, Ireland.
J Aerosol Med. 2007 Fall;20(3):257-68. doi: 10.1089/jam.2007.0605.
Pulmonary gene therapy has the potential to treat or cure respiratory diseases such as cystic fibrosis. Much work has focused on the delivery of genes to the lung using viral vectors with varying degrees of success. Viral vectors are problematic and undesirable for use in the lung because they can provoke an acute immune response. This study has focused on the characterization of nonviral, polymer-based gene vectors for use with nebulizers. Calf thymus DNA has been used as a model, and was complexed with each of the three polycations; 22 kDa linear polyethyleneimine, 25 kDa branched polyethyleneimine, and 29.5 kDa polylysine using water, glucose solution, and phosphate-buffered saline (PBS) as carrier liquids. Fourier transform infrared spectroscopy has shown that the DNA retains the B form during the complex formation. The complexes prepared at N:P ratios of 10, have been nebulized using a vibrating plate nebulizer and the particle size and Zeta potentials measured before and after nebulization. The particle size distributions of the DNA complexes prepared in water and glucose solution were unimodal before and after nebulization with a small increase in particle size following nebulization. Choice of complexing polymer is shown to have only a small effect on particle size with the dominant effect coming from the ionic character of the dispersion fluid. Complexes prepared in PBS, although originally unimodal, showed pronounced agglomeration on nebulization. With all polymers in water or glucose solution, the Zeta potential increases after nebulization, but with PBS as the carrier liquid the potential falls and is clearly associated with the observed agglomeration. Gel electrophoresis shows that the complexing polymers protect the DNA through the nebulization process in all cases.
肺部基因治疗有潜力治疗或治愈诸如囊性纤维化之类的呼吸系统疾病。许多工作聚焦于使用病毒载体将基因递送至肺部，且取得了不同程度的成功。病毒载体存在问题，并不适合在肺部使用，因为它们会引发急性免疫反应。本研究聚焦于用于雾化器的非病毒、基于聚合物的基因载体的特性研究。小牛胸腺DNA已被用作模型，并与三种聚阳离子中的每一种进行复合；使用水、葡萄糖溶液和磷酸盐缓冲盐水（PBS）作为载液，将22 kDa线性聚乙烯亚胺、25 kDa支化聚乙烯亚胺和29.5 kDa聚赖氨酸与小牛胸腺DNA进行复合。傅里叶变换红外光谱表明，在复合形成过程中DNA保留了B型结构。以N:P比为10制备的复合物，使用振动板雾化器进行雾化，并在雾化前后测量粒径和Zeta电位。在水和葡萄糖溶液中制备的DNA复合物的粒径分布在雾化前后均为单峰，雾化后粒径略有增加。结果表明，复合聚合物的选择对粒径的影响很小，主要影响来自分散液的离子特性。在PBS中制备的复合物，虽然最初是单峰的，但在雾化时显示出明显的团聚。对于在水或葡萄糖溶液中的所有聚合物，雾化后Zeta电位增加，但以PBS作为载液时，电位下降，并且明显与观察到的团聚有关。凝胶电泳表明，在所有情况下，复合聚合物在雾化过程中都能保护DNA。
