Quantitative and mechanism-based investigation of post-nuclear delivery events between adenovirus and lipoplex

被引：102

作者：

Hama, Susumu

Akita, Hidetaka

Iida, Shinya

Mizuguchi, Hiroyuki

Harashima, Hideyoshi ^{[1
]}

机构：

[1] Hokkaido Univ, Fac Pharmaceut Sci, Sapporo, Hokkaido 0600812, Japan

[2] Natl Inst Biomed Innovat, Lab Gene Transfer & Regulat, Osaka 5670085, Japan

来源：

NUCLEIC ACIDS RESEARCH | 2007年 / 35卷 / 05期

基金：

日本学术振兴会;

关键词：

D O I：

10.1093/nar/gkl1165

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

Quantitative and mechanism-based information on differences in transfection efficiency between viral and non-viral vectors would be highly useful for improving the effectiveness of non-viral vectors. A previous quantitative comparison of intracellular trafficking between adenovirus and LipofectAMINE PLUS (LFN) revealed that the three orders of magnitude lower transfection efficiency of LFN was dominantly rate limited by the post-nuclear delivery process. In the present study, the contribution of transcription and translation processes to the overall differences in the transgene expression efficiency of nucleus-delivered DNA was independently evaluated by quantifying mRNA. As a result, transcription efficiency (E-transcript) of LFN, denoted as transgene expression divided by the amount of nuclear pDNA was about 16 times less than that for adenovirus. Furthermore, translation efficiency (E-translate), denoted as transfection activity divided by mRNA expression was approximately 460 times less in LFN. Imaging of the decondensed form of DNA by in situ hybridization revealed that poor decondensation efficiency of LFN is involved in the inferior E-transcript- Moreover, the inferior translation efficiency (E-translate) of LFN was mainly due to electrostatic interactions between LFN and mRNA. Collectively, an improvement in nuclear decondensation and the diminution of the interaction between vector and mRNA is essential for the development of new generations of non-viral vectors.

引用

页码：1533 / 1543

页数：11

共 42 条

[1] Evaluation of the nuclear delivery and intra-nuclear transcription of plasmid DNA condensed with μ (mu) and NLS-μ by cytoplasmic and nuclear microinjection:: a comparative study with poly-L-lysine [J].