Mechanisms of GalTKO Lung Xenograft Injury

描述：针对不同靶点(特别是Gal，3Gal)和补体的抗体是心、肺和其他器官超急性排斥反应的关键介质。然而，在猪到人和猪到非人的灵长类动物模型中，我们一直发现有效的补体调节和有效地清除抗猪抗体与异种肺移植的快速功能障碍有关。因此，除了引起其他器官超急性排斥反应的机制外，超急性肺排斥反应(HALR)也是由多种机制介导的。 这一模式得到了初步研究的加强，在体外灌流的三个Galt-KO猪肺平均保留其功能2小时(60、134和170分钟)，远远长于对照组(10分钟)。失败的原因是肺血管阻力突然、迅速增加。尽管在灌流10分钟和30分钟时获得的活检在组织学上并不显著，但在移植失败时观察到血管内血栓和毛细血管充血，尽管使用了高剂量的肝素抗凝。凝血途径激活(F1+2)延迟但未被阻止，血小板激活(凝血酶敏感蛋白释放)未减弱，灌流液中超过70%的血小板在开始灌流后几分钟内被隔离在肺内。补体在肺中的活化和沉积，虽然减少了，但并未被阻止。基于这些观察和我们先前的工作，证明凝血酶、血小板、补体和肺血管内巨噬细胞在Gal+肺超急性排斥反应中的关键作用，我们推测血管内凝血失调和补体残留激活是Galt KO猪肺移植急性损伤的主要原因。 为了验证这一假设，将在已建立的体外灌流模型中结合使用遗传学方法(GalTKO肺表达人类组织因子途径抑制物或衰变加速因子)和药理学方法(特定的血小板受体、凝血酶或补体抑制物)。在体外产生最佳肺功能的方法将在活体内通过维持生命的猪到狒狒肺异种移植模型进行验证。作为所提出的研究的结果，我们预计将第一次成功地预防超急性肺排斥反应，主要使用捐赠者指导的、基于机制的策略来始终如一地实现企鹅的猪肺的生命维持功能，从而使随后的免疫障碍能够在这个器官系统中得到解决。

DESCRIPTION: Antibodies directed at various targets (especially Gal al,3Gal) and complement are pivotal mediators of hyperacute rejection of the heart, lung, and other organs. However in pig-to-human and pig-to-non-human primate models, we have consistently found that potent complement regulation coupled with efficient removal of anti-pig antibody is associated with rapid dysfunction of lung xenografts. Thus hyperacute lung rejection (HALR) is mediated by mechanisms in addition to those that cause hyperacute rejection of other organs. This paradigm is reinforced by the preliminary studies, where three GalT-KO swine lungs perfused ex vivo retained their function for an average of 2 hours (60, 134, and 170 minutes), far longer than controls (<10 minutes). Failure was due to sudden, rapid increase in pulmonary vascular resistance. Although biopsies obtained at 10 and 30 minutes of perfusion were histologically unremarkable, intravascular thrombi and capillary congestion were observed at graft failure, despite anticoagulation with high-dose heparin. Coagulation pathway activation (Fl+2) was delayed but not prevented, platelet activation (thrombospondin release) was not attenuated, and over 70% of platelets in the perfusate were sequestered in the lung within minutes of initiating perfusion. Complement activation and deposition in the lung, while reduced, were not prevented. Based on these observations and our previous work demonstrating pivotal roles for thrombin, platelets, complement, and pulmonary intravascular macrophages in hyperacute rejection of Gal+ lungs, we hypothesize that dysregulated intravascular coagulation and residual complement activation are the principle cause of acute injury of GalT KO pig lung xenograft. To test this hypothesis, a combination of genetic (GalTKO lungs expressing human tissue factor pathway inhibitor or decay accelerating factor) and pharmacologic approaches (specific platelet receptor, thrombin, or complement inhibitors) will be used in an established ex vivo perfusion model. Approaches that yield optimal lung function ex vivo will then be validated in vivo in a life-supporting pig-to-baboon lung xenograft model. As a result of the studies proposed, we anticipate that hyperacute lung rejection will be successfully prevented for the first time using primarily donor-directed, mechanism-based strategies to consistently achieve life-supporting function of a pig lung in a baboon, allowing subsequent immunologic barriers to be addressed in this organ system.