Institute for Innovative NanoBio Drug Discovery and Development, Graduate School of Pharmaceutical Sciences, Kyoto University, Kyoto, Japan.
PLoS One. 2012;7(7):e41319. doi: 10.1371/journal.pone.0041319. Epub 2012 Jul 23.
We have developed an in vivo transfection method for naked plasmid DNA (pDNA) and siRNA in mice by using a tissue suction device. The target tissue was suctioned by a device made of polydimethylsiloxane (PDMS) following the intravenous injection of naked pDNA or siRNA. Transfection of pDNA encoding luciferase was achieved by the suction of the kidney, liver, spleen, and heart, but not the duodenum, skeletal muscle, or stomach. Luciferase expression was specifically observed at the suctioned region of the tissue, and the highest luciferase expression was detected at the surface of the tissue (0.12±0.03 ng/mg protein in mice liver). Luciferase expression levels in the whole liver increased linearly with an increase in the number of times the liver was suctioned. Transfection of siRNA targeting glyceraldehyde 3-phosphate dehydrogenase (GAPDH) gene significantly suppressed the expression of GAPDH mRNA in the liver. Histological analysis shows that severe damage was not observed in the suctioned livers. Since the suction device can be mounted onto the head of the endoscope, this method is a minimally invasive. These results indicate that the in vivo transfection method developed in this study will be a viable approach for biological research and therapies using nucleic acids.
我们开发了一种用于在体转染裸质粒 DNA（pDNA）和 siRNA 的方法，使用一种组织抽吸装置。在静脉注射裸 pDNA 或 siRNA 后，用聚二甲基硅氧烷（PDMS）制成的装置抽吸目标组织。通过抽吸肾脏、肝脏、脾脏和心脏可以实现 pDNA 转染编码荧光素酶，但不能转染十二指肠、骨骼肌或胃。在组织的抽吸区域特异性观察到荧光素酶表达，并且在组织表面检测到最高的荧光素酶表达（小鼠肝脏中为 0.12±0.03ng/mg 蛋白）。随着肝脏抽吸次数的增加，肝脏中荧光素酶的表达水平呈线性增加。针对甘油醛 3-磷酸脱氢酶（GAPDH）基因的 siRNA 转染显著抑制了肝脏中 GAPDH mRNA 的表达。组织学分析表明，在抽吸的肝脏中未观察到严重损伤。由于抽吸装置可以安装在内窥镜的头部，因此该方法是一种微创方法。这些结果表明，本研究中开发的在体转染方法将成为使用核酸进行生物研究和治疗的可行方法。
