Kawashita Yujo, Fujioka Hikaru, Ohtsuru Akira, Kuroda Hiroaki, Eguchi Susumu, Kaneda Yasufumi, Yamashita Shunichi, Kanematsu Takashi
Department of Transplantation and Digestive Surgery, Graduate School of Biomedical Sciences, Nagasaki University School of Medicine, 1-7-1 Sakamoto, Nagasaki 852-8501, Japan.
J Surg Res. 2006 May;132(1):136-41. doi: 10.1016/j.jss.2005.09.027. Epub 2005 Dec 9.
Most virus mediated transfection systems are efficient; however, their highly immunogenic properties do tend to cause clinical problems. HVJ-liposome vector is a hybrid vector consisting of liposome and inactivated sendai virus (hemagglutinating virus of Japan HVJ), which has been reported to be have a low immunogenicity, while it can also be repeatedly administered. To enhance the transfection efficiency, especially in the liver, we investigated the efficacy of total vascular exclusion (TVE) during the portal vein injection (PVI) of the vectors.
beta-galactosidase and luciferase expression were used as reporter genes. Wistar rats were injected with HVJ-liposome through PVI without TVE (PVI group, n = 10) or PVI with TVE (PVI + TVE group, n = 10). The control rats were infused with equal volumes of saline through the portal vein (control group n = 12). The transfection efficiencies were assessed by beta-galactosidase staining and a luciferase assay. Biochemical and histological analyses were performed to evaluate the tissue toxicity after gene transfer.
The reporter genes expression in the liver dramatically increased after PVI + TVE in comparison to after PVI alone (1.2 x 10(5)versus 1.5 x 10(4) RLU/mg protein, P < 0.05 according to a luciferase assay). Notably, the extrahepatic "leaky" transgene expression could be minimized by PVI + TVE, whereas the general condition remained unchanged according to both the biochemical parameters and histological findings.
The present data indicate that PVI + TVE may thus facilitate the liver-specific gene delivery using the HVJ-liposome method and this modality might also be applicable to other gene transfer systems.
大多数病毒介导的转染系统效率很高；然而，其高度免疫原性往往会引发临床问题。HVJ-脂质体载体是一种由脂质体和灭活仙台病毒（日本血凝病毒HVJ）组成的杂交载体，据报道其免疫原性较低，而且还可重复给药。为提高转染效率，尤其是在肝脏中的转染效率，我们研究了在载体门静脉注射（PVI）过程中采用全血管阻断（TVE）的效果。
使用β-半乳糖苷酶和荧光素酶表达作为报告基因。将Wistar大鼠通过不进行TVE的PVI（PVI组，n = 10）或进行TVE的PVI（PVI + TVE组，n = 10）注射HVJ-脂质体。对照组大鼠通过门静脉输注等体积的生理盐水（对照组n = 12）。通过β-半乳糖苷酶染色和荧光素酶测定评估转染效率。进行生化和组织学分析以评估基因转移后的组织毒性。
与单独PVI后相比，PVI + TVE后肝脏中报告基因的表达显著增加（根据荧光素酶测定，分别为1.2×10⁵和1.5×10⁴RLU/mg蛋白，P < 0.05）。值得注意的是，PVI + TVE可将肝外“渗漏”的转基因表达降至最低，而根据生化参数和组织学结果，总体状况保持不变。
目前的数据表明，PVI + TVE可能因此有助于使用HVJ-脂质体方法进行肝脏特异性基因递送，并且这种方式也可能适用于其他基因转移系统。
