Deparment of Drug Discovery Sciences, Boehringer Ingelheim Pharma GmbH & Co. KG, Biberach an der Riss, Germany.
Department of Non-clinical Drug Safety, Boehringer Ingelheim Pharma GmbH & Co. KG, Biberach an der Riss, Germany.
Am J Physiol Lung Cell Mol Physiol. 2022 Aug 1;323(2):L206-L218. doi: 10.1152/ajplung.00364.2021. Epub 2022 Jun 28.
Animal models are important to mimic certain pathways or biological aspects of human pathologies including acute and chronic pulmonary diseases. We developed a novel and flexible mouse model of acute epithelial lung injury based on adeno-associated virus (AAV) variant 6.2-mediated expression of the human diphtheria toxin receptor (DTR). Following intratracheal administration of diphtheria toxin (DT), a cell-specific death of bronchial and alveolar epithelial cells can be observed. In contrast to other lung injury models, the here described mouse model provides the possibility of targeted injury using specific tropisms of AAV vectors or cell-type-specific promotors to drive the human DTR expression. Also, generation of cell-specific mouse lines is not required. Detailed characterization of the AAV-DTR/DT mouse model including titration of viral genome (vg) load and administered DT amount revealed increasing cell numbers in bronchoalveolar lavage (BAL; macrophages, neutrophils, and unspecified cells) and elevation of degenerated cells and infiltrated leukocytes in lung tissue, dependent of vg load and DT dose. Cytokine levels in BAL fluid showed different patterns with higher vg load, e.g., IFNγ, TNFα, and IP10 increasing and IL-5 and IL-6 decreasing, whereas lung function was not affected. In addition, laser-capture microdissection (LCM)-based proteomics of bronchial epithelium and alveolar tissue revealed upregulated immune and inflammatory responses in all regions and extracellular matrix deposition in infiltrated alveoli. Overall, our novel AAV-DTR/DT model allows investigation of repair mechanisms following epithelial injury and resembles specific mechanistic aspects of acute and chronic pulmonary diseases.
动物模型对于模拟人类病理学的某些途径或生物学方面非常重要，包括急性和慢性肺部疾病。我们基于腺相关病毒（AAV）变体 6.2 介导的人白喉毒素受体（DTR）表达，开发了一种新型的、灵活的急性上皮性肺损伤小鼠模型。在气管内给予白喉毒素（DT）后，可以观察到支气管和肺泡上皮细胞的特异性死亡。与其他肺损伤模型相比，本文描述的小鼠模型提供了使用 AAV 载体的特异性嗜性或细胞类型特异性启动子来驱动人 DTR 表达进行靶向损伤的可能性。此外，不需要生成细胞特异性的小鼠系。对 AAV-DTR/DT 小鼠模型进行了详细的表征，包括病毒基因组（vg）载量和给予的 DT 量的滴定，结果显示，支气管肺泡灌洗液（BAL；巨噬细胞、中性粒细胞和未指定细胞）中的细胞数量增加，以及依赖于 vg 载量和 DT 剂量，肺组织中退化细胞和浸润的白细胞增多。BAL 液中的细胞因子水平显示出不同的模式，较高的 vg 载量，例如 IFNγ、TNFα 和 IP10 增加，IL-5 和 IL-6 减少，而肺功能不受影响。此外，支气管上皮和肺泡组织的激光捕获显微解剖（LCM）-基于蛋白质组学的研究显示，所有区域的免疫和炎症反应上调，以及浸润肺泡的细胞外基质沉积。总的来说，我们的新型 AAV-DTR/DT 模型允许研究上皮损伤后的修复机制，并类似于急性和慢性肺部疾病的特定机制方面。
