PCI-enhanced adenoviral transduction employs the known uptake mechanism of adenoviral particles
PCI-enhanced adenoviral transduction employs the known uptake mechanism of adenoviral particles
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DOI：
10.1038/sj.cgt.7700808
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发表时间：
2005-05-01
影响因子：
6.4
通讯作者：
Mælandsmo, GM
中科院分区：
文献类型：
--
作者：
Engesæter, BO;Bonsted, A;Mælandsmo, GM
如何将治疗基因高效、特异地递送到靶组织中，是进一步发展体内基因治疗的一个重要问题。本研究将物理靶向技术光化学内化（PCI）与腺病毒联合应用。与单独使用腺病毒相比，PCI和腺病毒转导的结合已被证明是有利的，本研究的目的是验证腺病毒受体的作用，并确定光化学处理细胞中腺病毒颗粒使用的摄取途径。所有检测的细胞系在pci处理后都显示出增强的转导效率，与常规感染的细胞相比，转基因表达最多增加13倍。阻断CAR诱导完全抑制pci增强的转基因表达。然而，光化学处理成功地提高了重靶向病毒AdRGD-GFP的转导效率，这也表明病毒与car的相互作用对于获得对腺病毒转导的光化学效应并不重要。阻断α (V)-整合素可显著降低光化学处理细胞的基因表达。在感染后将表达阴性突变动力蛋白的HeLa细胞进行光照处理，基因转移量没有显著增加，而在表达野生型动力蛋白的HeLa细胞中，基因转移量增加了7倍。此外，氯丙嗪以剂量依赖的方式抑制光化学转导，而Filipin III对基因转移没有影响。总之，这些数据表明腺病毒受体结合是重要的，网格蛋白介导的内吞作用是光化学处理细胞中腺病毒颗粒的主要摄取机制。
The development of methods for efficient and specific delivery of therapeutic genes into target tissues is an important issue for further development of in vivo gene therapy. In the present study, the physical targeting technique, photochemical internalization (PCI), has been used together with adenovirus. The combination of PCI and adenoviral transduction has previously been shown to be favorable compared to adenovirus used alone, and the aim of this study was to verify the role of the adenoviral receptors and identify the uptake pathway used by adenoviral particles in photochemically treated cells. All examined cell lines showed augmented transduction efficiency after PCI-treatment, with a maximum of 13-fold increase in transgene expression compared to conventionally infected cells. Blocking of CAR induced a complete inhibition of PCI-enhanced transgene expression. However, photochemical treatment managed to enhance the transduction efficiency of the retargeted virus AdRGD-GFP showing also that the virus-CAR interaction is not vital for obtaining a photochemical effect on adenoviral transduction. Blocking the alpha(V)-integrins reduced the gene expression significantly in photochemically treated cells. Subjecting HeLa cells expressing negative mutant-dynamin to light treatment after infection gave no significant increase in gene transfer, while the gene transfer were enhanced seven-fold in cells with wild-type dynamin. Furthermore, chlorpromazine inhibited photochemical transduction in a dose-dependent manner, whereas Filipin III had no effect on the gene transfer. In summary, the data presented imply that adenoviral receptor binding is important and clathrin-mediated endocytosis is the predominant uptake mechanism for adenoviral particles in photochemically treated cells.