Schwarz L A, Johnson J L, Black M, Cheng S H, Hogan M E, Waldrep J C
Department of Molecular Physiology and Biophysics, Baylor College of Medicine, Houston, TX 77204, USA.
Hum Gene Ther. 1996 Apr 10;7(6):731-41. doi: 10.1089/hum.1996.7.6-731.
Aerosol delivery of gene therapy for treatment of lung diseases allows topical treatment of the airways with DNA concentrations not obtainable by systemic administration. We have investigated delivery of cationic liposomes complexed to plasmid DNA in a small particle aerosol. Plasmid cDNA-DMRIE/DOPE complexes were nebulized using either an Aerotech II or Puritan-Bennett 1600 (PB1600) nebulizer. Reservoir sampling showed that DNA-DMRIE/DOPE complexes were damaged to a significant degree during nebulization, such that activity of transfected gene was diminished. Of the nebulizers analyzed, DNA-DMRIE/DOPE complexes were more stable in the PB1600. The loss of effective transfection by DNA-DMRIE/DOPE, as detected by decreased reporter gene activity in A549 lung cells, was consistent with denaturation of the DMRIE/DOPE. In contrast, nebulized DNA-DOSPA/DOPE complexes retained complete ability to transfect. Adjustments to flow rate and reservoir volume of the PB1600 allowed a longer period of delivery of active DNA-DMRIE/DOPE particles. DNA-DMRIE/DOPE was radiolabeled with Technetium-99m (99mTc), nebulized, and the output captured in either an Andersen Sampler (AS) (Andersen, 1958) cascade impactor particle size analyzer or an all glass impinger. cDNA-cationic lipid complexes were detected in size ranges of 0.4-10 microns, with most particles found between 1-2 microns. Aerosol output was consistent from 0 to 5 min. These results show the feasibility of aerosol delivery of DNA-cationic lipids for the purposes of gene therapy to the lung.
通过气溶胶递送基因疗法来治疗肺部疾病，能够对气道进行局部治疗，且可达到全身给药无法实现的DNA浓度。我们研究了与质粒DNA复合的阳离子脂质体以小颗粒气溶胶形式的递送情况。使用Aerotech II或Puritan-Bennett 1600（PB1600）雾化器对质粒cDNA-DMRIE/DOPE复合物进行雾化。储液器采样显示，DNA-DMRIE/DOPE复合物在雾化过程中受到显著损伤，导致转染基因的活性降低。在所分析的雾化器中，DNA-DMRIE/DOPE复合物在PB1600中更稳定。如通过A549肺细胞中报告基因活性降低所检测到的，DNA-DMRIE/DOPE有效转染的丧失与DMRIE/DOPE的变性一致。相比之下，雾化的DNA-DOSPA/DOPE复合物保留了完全的转染能力。对PB1600的流速和储液器体积进行调整后，活性DNA-DMRIE/DOPE颗粒的递送时间得以延长。用锝-99m（99mTc）对DNA-DMRIE/DOPE进行放射性标记，然后雾化，并将输出物捕获在Andersen采样器（AS）（Andersen，1958年）级联冲击器粒度分析仪或全玻璃冲击器中。在0.4 - 10微米的尺寸范围内检测到cDNA - 阳离子脂质复合物，大多数颗粒在1 - 2微米之间。在0至5分钟内，气溶胶输出是一致的。这些结果表明，通过气溶胶递送DNA - 阳离子脂质用于肺部基因治疗是可行的。
