| Summary: | 博士 === 臺北醫學大學 === 藥學研究所 === 95 === Modifications of both carriers and host barriers have been investigated for efficient inhalation gene delivery to lung. Here we used a biocompatible non-ionic poly(ethyleneoxide) -poly(propyleneoxide)-poly(ethyleneoxide) (PEO-PPO-PEO) polymeric micelles (PM) as a carrier and combined it with ethanol (10-40%) to enhance tissue penetration of delivered DNA. The inhalation delivery with six 100 μg doses of pCMV-Lac Z with PM to nude mice in two days at 8 hr interval could enhance the gene expression in the lung around bronchioles (38%), trachea (32%), stomach (22%) and duodenum (26%) areas under only with 10% ethanol. Using intra-, intercellular penetration markers (estradiol and mannitol), 10% ethanol also increased the apparent intercellular permeability (Pmannitol) by 54% in stomach and by 41% in intestine, and increased the apparent intracellular permeability (Pestradiol) by 42% in stomach and by 141% in intestine at 48 hr after the first dosage of delivery. Also delivery of DNA encoding a functional human cystic fibrosis transmembrane protein (CFTR) using the same inhalation delivery method co-formulated with 10% ethanol, an increased expression of CFTR in lung was detected by immunostaining. Ethanol effect on biophysical alterations in protein and lipids of GI tissues was evaluated using attenuated total reflection/Fourier transform infrared (ATR/FTIR) spectroscopy. The IR spectra of both tissues showed that β–sheet/α-helix ratios of protein compositions in the secondary structure-dependent amide I band (1600-1700 cm-1) increased linearly with increasing concentrations of ethanol as well as tissue intercellular permeation. The decrease of absorbance in C-H stretching band, an indictor of lipid extraction was also correlated with intercellular permeation, but not consisted with intracellular permeation with increasing ethanol concentration. Using the P=O- (1237 cm-1) stretching band, an indicator of hydration state in tissues, both tissues only pretreated with 20%-40% ethanol was significantly shifted to lower wave number in IR spectra. Also the significant decrease of hydration level (HL) % in both tissues only with pretreated of 20%-40% ethanol was observed. Thus, we concluded that 10% ethanol co-formulated with the PM system could enhance inhaled gene delivery to airway and GI tract due to the intracellular transport of GI tissues increased. The dehydration ability of ethanol at higher concentration (20%-40%) may overshadow intracellular permeation of tissues with decreasing intracellular permeation.
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