Mueller Christian, Torrez Daniel, Braag Sofia, Martino Ashley, Clarke Tracy, Campbell-Thompson Martha, Flotte Terence R
Department of Pediatrics and Powell Gene Therapy Center, College of Medicine, University of Florida, Gainesville, FL, USA.
J Gene Med. 2008 Jan;10(1):51-60. doi: 10.1002/jgm.1119.
Recently, we have developed a model of airway inflammation in a CFTR knockout mouse utilizing Aspergillus fumigatus crude protein extract (Af-cpe) to mimic allergic bronchopulmonary aspergillosis (ABPA) 1, an unusual IgE-mediated hypersensitivity syndrome seen in up to 15% of cystic fibrosis (CF) patients and rarely elsewhere. We hypothesized that replacement of CFTR via targeted gene delivery to airway epithelium would correct aberrant epithelial cytokine signaling and ameliorate the ABPA phenotype in CFTR-deficient (CFTR 489X - /-, FABP-hCFTR + / +) mice. CFTR knockout mice underwent intra-tracheal (IT) delivery of recombinant adeno-associated virus serotype 5 (rAAV5Delta-264CFTR) or rAAV5-GFP at 2.58 x 10(12) viral genomes/mouse. All mice were then sensitized with two serial injections (200 microg) of crude Af antigen via the intra-peritoneal (IP) route. Untreated mice were sensitized without virus exposure. Challenges were performed 2 weeks after final sensitization, using a 0.25% solution containing Aspergillus fumigatus crude protein extract delivered by inhalation on three consecutive days. The rAAV5Delta-264CFTR-treated mice had lower total serum IgE levels (172513 ng/ml +/- 1312) than rAAV5-GFP controls (26 892 ng/ml +/- 3715) (p = 0.037) and non-treated, sensitized controls (24 816 +/- 4219 ng/ml). Serum IgG1 levels also were lower in mice receiving the CFTR vector. Interestingly, splenocytes from rAAV5Delta-264CFTR-treated mice secreted less IL-13, INFg, TNFa, RANTES and GM-CSF after ConA stimulation. Gene therapy with rAAV5Delta-264CFTR attenuated the hyper-IgE response in this reproducible CF mouse model of ABPA, with systemic effects also evident in the cytokine response of stimulated splenocytes.
最近，我们利用烟曲霉粗蛋白提取物（Af-cpe）建立了一种CFTR基因敲除小鼠气道炎症模型，以模拟变应性支气管肺曲霉病（ABPA），这是一种不常见的IgE介导的超敏综合征，在高达15%的囊性纤维化（CF）患者中可见，在其他地方很少见。我们假设，通过将CFTR靶向递送至气道上皮来替代CFTR，将纠正异常的上皮细胞因子信号传导，并改善CFTR缺陷（CFTR 489X - / -，FABP-hCFTR + / +）小鼠的ABPA表型。CFTR基因敲除小鼠经气管内（IT）注射2.58×10¹²病毒基因组/小鼠的重组腺相关病毒5型（rAAV5Delta-264CFTR）或rAAV5-GFP。然后所有小鼠通过腹腔内（IP）途径用两次连续注射（200μg）的粗Af抗原进行致敏。未处理的小鼠在无病毒暴露的情况下进行致敏。在最后一次致敏后2周进行激发，连续三天通过吸入给予含0.25%烟曲霉粗蛋白提取物的溶液。接受rAAV5Delta-264CFTR治疗的小鼠总血清IgE水平（172513 ng/ml±1312）低于rAAV5-GFP对照组（26892 ng/ml±3715）（p = 0.037）和未处理的致敏对照组（24816±4219 ng/ml）。接受CFTR载体的小鼠血清IgG1水平也较低。有趣的是，来自接受rAAV5Delta-264CFTR治疗的小鼠的脾细胞在刀豆蛋白A刺激后分泌的IL-13、INFg、TNFa、RANTES和GM-CSF较少。在这个可重复的CF小鼠ABPA模型中，用rAAV5Delta-264CFTR进行基因治疗减弱了高IgE反应，全身效应在刺激的脾细胞的细胞因子反应中也很明显。
