Alesci Salvatore, Ramsey Walter J, Bornstein Stefan R, Chrousos George P, Hornsby Peter J, Benvenga Salvatore, Trimarchi Francesco, Ehrhart-Bornstein Monika
Pediatric and Reproductive Endocrinology Branch, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892-1583, USA.
Proc Natl Acad Sci U S A. 2002 May 28;99(11):7484-9. doi: 10.1073/pnas.062170099.
Recombinant adenoviral vectors are effective in transferring foreign genes to a variety of cells and tissue types, both in vitro and in vivo. However, during the gene transfer, they may alter the principal function and local environment of transfected cells. Increasing evidence exists for a selective adrenotropism of adenovirus during infections and gene transfer. Therefore, using bovine adrenocortical cells in primary culture, we analyzed the influence of different adenoviral deletion mutants on cell morphology and physiology. Transfection of cells with an E1/E3-deleted adenoviral vector, engineered to express a modified form of the Aequorea victoria green fluorescent protein, was highly efficient, as documented by fluorescent microscopy. Ultrastructural analysis, however, demonstrated nuclear fragmentation and mitochondrial alterations in addition to intranuclear viral particles. Basal secretion of 17-OH-progesterone, 11-deoxycortisol, and cortisol was significantly increased by E1/E3-deleted vectors; yet, the corticotropin-stimulated release of these steroids was decreased. Interestingly, neither purified viral capsids nor E3/E4-deleted adenoviral mutants altered basal and stimulated steroidogenesis of adrenocortical cells. An intact adrenal response is crucial for adaptation to stress and survival. Therefore, the implications of our findings need to be considered in patients with adenoviral infections and those undergoing clinical studies using adenoviral gene transfer. At the same time, the high level of transfection in adrenocortical cells might make appropriately modified adenoviral vectors suitable for gene therapy of adrenocortical carcinomas with poor prognosis.
重组腺病毒载体在体外和体内将外源基因转移到多种细胞和组织类型中均有效。然而,在基因转移过程中,它们可能会改变转染细胞的主要功能和局部环境。越来越多的证据表明,腺病毒在感染和基因转移过程中具有选择性嗜肾上腺性。因此,我们使用原代培养的牛肾上腺皮质细胞,分析了不同腺病毒缺失突变体对细胞形态和生理学的影响。用经工程改造以表达修饰形式的维多利亚多管水母绿色荧光蛋白的E1/E3缺失腺病毒载体转染细胞非常高效,荧光显微镜记录了这一点。然而,超微结构分析表明,除了核内病毒颗粒外,还存在核碎裂和线粒体改变。E1/E3缺失载体显著增加了17-羟孕酮、11-脱氧皮质醇和皮质醇的基础分泌;然而,促肾上腺皮质激素刺激的这些类固醇释放却减少了。有趣的是,纯化的病毒衣壳和E3/E4缺失腺病毒突变体均未改变肾上腺皮质细胞的基础和刺激的类固醇生成。完整的肾上腺反应对于适应应激和生存至关重要。因此,在腺病毒感染患者和接受腺病毒基因转移临床研究的患者中,需要考虑我们研究结果的影响。同时,肾上腺皮质细胞中的高转染水平可能使适当修饰的腺病毒载体适用于预后不良的肾上腺皮质癌的基因治疗。