Investigation of anthrax lethal and edema toxins pulmonary effects in an isolated perfused lung model

炭疽杆菌（B.炭疽病）仍然是发达国家的健康威胁。致死毒素和水肿毒素（分别为LT和ET）均参与炭疽感染时器官损伤和致死的发病机制。了解毒素致病作用的机制对于改善这种致命感染的结果将是重要的。体外研究结果确实表明，每种毒素都能引起内皮损伤和血管完整性丧失以及通透性增加。虽然从未在肺中进行过试验，但毒素增加的内皮渗透性可能导致液体外渗、氧转移和肺顺应性降低以及肺血管阻力增加。为了研究这些可能性，本研究将采用离体灌流大鼠肺模型来检查LT或ET是否确实引起肺内皮损伤和肺血管通透性增加。该离体模型将允许直接测量随时间变化的肺重量，这是计算肺渗透系数所需的。确定毒素是否改变肺通透性将提高我们对炭疽相关肺损伤的发病机制和临床管理的理解。 这项研究将分两部分进行。在第一部分（第1部分）中，将开发分离的灌注大鼠肺模型，并使用从健康大鼠分离的肺测试模型测量内皮渗透性、氧合、顺应性和血管阻力变化的能力。作为阳性对照，第1部分将包括用血栓烷A2（一种已知可增加肺内皮通透性的介质）激发的肺实验。在第二部分（第2部分）中，将用导入隔离肺灌注回路的LT或ET激发肺，并测量肺功能。本研究这一部分的一些实验还将包括使用Raxibacumab（一种抑制宿主细胞摄取LT和ET的单克隆抗体）和Adefovir（一种ET的细胞内抑制剂），以进一步探索这些毒素致病作用的机制。第2部分中测试的功能参数将包括渗透性、氧合、顺应性和血管阻力。第2部分的一些肺部将被送往组织学和电子显微镜检查。

Bacillus anthracis (B. anthracis or anthrax) remains a health threat for the developed world. Both lethal and edema toxin (LT and ET respectively) contribute to the pathogenesis of organ injury and lethality during anthrax infection. Understanding the mechanisms underlying the toxins pathogenic effects will be important for improving the outcome with this lethal infection. In vitro findings do suggest that each toxin can produce endothelial injury and loss of vascular integrity and increased permeability. Although never tested in the lung, increased endothelial permeability with the toxins could result in extravasation of fluid, reductions in oxygen transfer and lung compliance, and increased pulmonary vascular resistance. To investigate these possibilities, the present study will employ an isolated perfused rat lung model to examine whether LT or ET does cause pulmonary endothelial injury and increased pulmonary vascular permeability. This ex vivo model will allow a direct measure of changing lung weight over time, which is required to calculate a lung permeability coefficient. Determining whether either toxin alters lung permeability will improve our understanding of the pathogenesis and management of anthrax associated lung injury clinically. This study will be done in two parts. In the first part (Part 1), an isolated perfused rat lung model will be developed and the models ability to measure changes in endothelial permeability, oxygenation, compliance and vascular resistance tested with lungs isolated from healthy rats. As a positive control, Part 1 will include experiments in lungs challenged with thromboxane A2, a mediator known to increase lung endothelial permeability. In the second part (Part 2), lungs will be challenged with LT or ET introduced into the isolated lungs perfusion circuit and lung function will be measured. Some experiments in this part of the study will also include the use of Raxibacumab, a monoclonal antibody that inhibits host cell uptake of LT and ET, and adefovir, an intracellular inhibitor of ET, to further explore mechanisms underlying these toxins pathogenic effects. The functional parameters tested in Part 2 will include permeability, oxygenation, compliance and vascular resistance. Some lungs from Part 2 will be sent for histology and examination by electron microscopy.