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首页> 外文期刊>Acta biomaterialia >DNA delivery in vitro via surface release from multilayer assemblies with poly(glycoamidoamine)s.
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DNA delivery in vitro via surface release from multilayer assemblies with poly(glycoamidoamine)s.

机译：通过从具有聚（糖胺基胺）的多层组件中释放表面来进行体外DNA递送。

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Localized controlled release of nucleic acid therapeutics could be an effective way to reduce the extracellular barriers associated with systemic delivery. Herein, we have used the layer-by-layer film deposition approach to construct ultrathin multilayer assemblies for in vitro controlled release of plasmid DNA (pDNA). Layer-by-layer assemblies containing alternate layers of cationic poly(l-tartaramidopentaethylenetetramine) (T4), and anionic pDNA were fabricated. The film thickness and the absorbance at 260 nm for different T4/pDNA multilayer assemblies were characterized by ellipsometry and UV-vis spectrophotometry, respectively. The results indicated an increased loading capacity of pDNA with respect to an increase in the number of T4/pDNA bilayers deposited. For the controlled-release studies we incubated the bilayers coated on quartz slides in phosphate-buffered saline (PBS) at 37 degrees C and collected the media at different incubation time points. The collected PBS samples were characterized for pDNA release by complexing solutions containing the released pDNA with Lipofectamine 2000 and following cellular pDNA uptake via flow cytometry and GFP gene expression assays with HeLa cells. The study showed that the multilayer films started to release pDNA after 1 day of incubation and increased after 7 days of incubation. Assays monitoring green fluorescent protein (GFP) expression in HeLa cells indicated that about 20% of the cells were positive for GFP expression at all sample time points up to 11 days. Although an increase in cells positive for Cy5-pDNA was found as the incubation time increased, the number of cells positive for GFP expression remained constant over the same time frame.
机译：核酸治疗剂的局部控制释放可能是减少与全身性输送相关的细胞外屏障的有效方法。在这里，我们已经使用了逐层膜沉积的方法来构建用于体外控制释放质粒DNA（pDNA）的超薄多层组件。制备了包含交替的阳离子聚（1-酒石酰胺戊四亚乙基四胺）（T4）和阴离子pDNA的逐层组件。通过椭圆偏振光度法和紫外可见分光光度法分别表征了不同的T4 / pDNA多层组件的膜厚度和在260 nm处的吸光度。结果表明，相对于沉积的T4 / pDNA双层数的增加，pDNA的负载能力增加。对于控释研究，我们在磷酸盐缓冲液（PBS）中于37℃孵育了涂在石英玻片上的双层，并在不同的孵育时间点收集了培养基。通过将含有释放的pDNA的溶液与Lipofectamine 2000络合，然后通过流式细胞术和HeLa细胞进行GFP基因表达分析来摄取细胞pDNA，来表征收集的PBS样品的pDNA释放。研究表明，多层膜在孵育1天后开始释放pDNA，在孵育7天后增加。监测HeLa细胞中绿色荧光蛋白（GFP）表达的分析表明，在长达11天的所有采样时间点，约有20％的细胞GFP表达呈阳性。尽管随着孵育时间的增加，发现Cy5-pDNA阳性的细胞增加，但在同一时间段内，GFP表达阳性的细胞数量保持恒定。

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