Liu Qiang, Zaiss Anne K, Colarusso Pina, Patel Kamala, Haljan Gregory, Wickham Thomas J, Muruve Daniel A
Department of Medicine, University of Calgary, Calgary, Alberta, T2N 4N1 Canada.
Hum Gene Ther. 2003 May 1;14(7):627-43. doi: 10.1089/104303403321618146.
Adenovirus (Ad) vectors can produce inflammatory responses at high doses. Intravenous administration of an Ad vector expressing green fluorescent protein (AdGFP) to naive mice induced a biphasic pattern of liver cytokine/chemokine gene expression over 7 days. Tumor necrosis factor alpha (TNF-alpha), macrophage inflammatory protein 2 (MIP-2), and interferon gamma-inducible protein 10 (IP-10) genes were upregulated, with two distinct peaks of mRNA expression occurring at 6 hr and 5 days. The administration of transcription-defective AdGFP particles induced the early but not the late peak of chemokine/cytokine gene expression, confirming that Ad vector-induced inflammation is capsid dependent in the early phase and transcription dependent in the late phase. To determine the role of adenoviral capsid motifs in the early phase, capsid-modified Ad vectors were employed. The intravenous administration of the RGD-deleted Ad vector AdL.PB*, the fiber mutant AdL.F*, or the double mutant AdL.FPB induced similar levels of cytokine/chemokine expression compared with the wild-type vector AdLuc. Kupffer cell blockade significantly reduced liver TNF-alpha, MIP-2, and IP-10 gene expression and liver inflammation after the administration of AdL.PB* or AdL.FPB. Fluorescence microscopy of AdLuc- and AdL.PB*-transduced liver at 1 hr revealed localization of Ad vectors to liver sinusoids in Kupffer cell-depleted mice. AdL.PB* induced less E-selectin and VCAM-1 gene expression in liver, confirming reduced endothelial activation in mice receiving RGD-deleted Ad vectors. In vitro studies of endothelial cells demonstrated reduced transduction and endothelial activation by AdL.PB* compared with AdLuc. These results demonstrate that adenovirus capsid RGD motifs are required for efficient transduction and endothelial cell activation. Altering vector tropism represents a feasible strategy to modulate the innate response to Ad vectors in nontargeted tissues.
腺病毒（Ad）载体在高剂量时可引发炎症反应。向未接触过抗原的小鼠静脉注射表达绿色荧光蛋白的腺病毒载体（AdGFP），在7天内诱导出肝脏细胞因子/趋化因子基因表达的双相模式。肿瘤坏死因子α（TNF-α）、巨噬细胞炎性蛋白2（MIP-2）和干扰素γ诱导蛋白10（IP-10）基因上调，mRNA表达出现两个不同的峰值，分别在6小时和5天时出现。给予转录缺陷型AdGFP颗粒可诱导趋化因子/细胞因子基因表达的早期峰值，但不能诱导晚期峰值，证实腺病毒载体诱导的炎症在早期阶段依赖衣壳，在晚期阶段依赖转录。为了确定腺病毒衣壳基序在早期阶段的作用，采用了衣壳修饰的腺病毒载体。与野生型载体AdLuc相比，静脉注射缺失RGD的腺病毒载体AdL.PB*、纤维突变体AdL.F或双突变体AdL.FPB诱导的细胞因子/趋化因子表达水平相似。给予AdL.PB或AdL.FPB后，库普弗细胞阻断显著降低了肝脏TNF-α、MIP-2和IP-10基因表达以及肝脏炎症。在1小时时对AdLuc和AdL.PB转导的肝脏进行荧光显微镜检查发现，在库普弗细胞缺失的小鼠中，腺病毒载体定位于肝血窦。AdL.PB在肝脏中诱导的E-选择素和血管细胞黏附分子-1（VCAM-1）基因表达较少，证实接受缺失RGD的腺病毒载体的小鼠内皮细胞活化减少。内皮细胞的体外研究表明，与AdLuc相比，AdL.PB*的转导和内皮细胞活化减少。这些结果表明，腺病毒衣壳RGD基序是有效转导和内皮细胞活化所必需的。改变载体嗜性是调节非靶向组织对腺病毒载体固有反应的一种可行策略。
