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Liver- and Lobe-Specific Gene Transfer Following the Continuous Microinstillation of Plasmid DNA onto the Liver Surface in Mice: Effect of Instillation Speed

机译:连续微量滴注质粒DNa到小鼠肝脏表面后的肝脏和叶片特异性基因转移:滴注速度的影响

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摘要

Development of technology to deliver foreign gene(s) to a specific organ/tissue is one of the major challenges in gene therapy. Here, we show liver- and lobe-specific gene transfer following the continuous microinstillation of plasmid DNA (pDNA) onto the liver surface in mice. Naked pDNA was continuously instilled onto the right medial liver lobe using syringe pump in male ddY mice. Our previous studies showed liver- and lobe-selective gene expression after instillation of 30m l of pDNA solution onto the liver surface, but gene expression was also found in the other liver lobe, kidney and spleen. To improve target site selectivity of gene expression, the instillation volume was decreased; however, non-specific gene expression in the other liver lobe and diaphragm was still detected. To prevent immediate diffusion of the pDNA solution, we performed continuous microinstillation of pDNA using a syringe pump; as a result, target site selectivity was greatly improved. As for instillation speed, 5 min infusion was enough to prevent diffusion of pDNA solution. Furthermore, transfection efficiency in the target site was maintained when instillation speed was slowed. Wiping off residual pDNA solution from the applied liver lobe resulted in a further improvement in selectivity, suggesting not only immediate diffusion, but also gradual diffusion, are important factors for successful target site-specific gene transfer. Information in this study will be useful for further development of an effective gene delivery system targeted to a specific organ/tissue by use of other non-viral or viral vectors.
机译:将外源基因传递到特定器官/组织的技术的发展是基因治疗的主要挑战之一。在这里,我们显示了在小鼠的肝脏表面上连续微滴注质粒DNA(pDNA)后肝脏和肺叶特异性基因的转移。使用注射泵在雄性ddY小鼠中将裸露的pDNA连续滴注到右内侧肝叶。我们以前的研究表明,将30ml的pDNA溶液滴注到肝表面后,肝和叶的选择性基因表达,但在其他肝叶,肾脏和脾脏中也发现了基因表达。为了提高基因表达的靶位点选择性,减少了滴注量;然而,仍检测到其他肝叶和diaphragm肌中的非特异性基因表达。为了防止pDNA溶液立即扩散,我们使用注射泵对pDNA进行了连续微滴注。结果,目标位点的选择性大大提高。至于滴注速度,输注5分钟足以防止pDNA溶液扩散。此外,当滴注速度减慢时,在目标部位的转染效率得以维持。从应用的肝叶上清除残留的pDNA溶液可进一步提高选择性,这表明不仅立即扩散,而且逐渐扩散是成功靶向位点特异性基因转移的重要因素。这项研究中的信息将有助于通过使用其他非病毒或病毒载体进一步开发针对特定器官/组织的有效基因递送系统。

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