Cytotoxic Lung Endothelial Amyloids

项目摘要/摘要 肺内皮细胞屏障的完整性对于有效的气体交换和防止血源性 呼吸道病原体的传播。院内优势菌--铜绿假单胞菌 肺炎，通过诱导连接复合体破坏导致肺泡内皮细胞屏障 泛滥和低氧血症。在这里，我们建议研究一种以前未确定的、由以下定义的新的疾病过程 院内常见病原菌铜绿假单胞菌与宿主内皮细胞的相互作用铜绿假单胞菌 利用III型分泌系统(T3SS)将细胞毒性外切酶直接注入宿主细胞。胞外酶 Y(ExoY)是一种T3SS效应因子，由约90%的临床铜绿假单胞菌菌株表达，导致 胞内cGMP、cUMP和cAMP。CAMP的增加促进了内皮tau的磷酸化， 它从微管解离，随后微管崩溃，内皮间间隙形成，以及 渗透性增加。过度磷酸化的tau然后结合成细胞毒性tau低聚物，释放出来 进入细胞外环境。这些细胞毒性tau寡聚体是可穿透并自我传播的淀粉样蛋白。 CGMP和CUMP各自的作用目前尚不清楚，尽管初步数据证实了 ExoY产生的CUMP增加与Tau寡聚释放之间的时间和空间相关性 PMVEC。胞浆中的CUMP信号与胞浆tau的减少和胞浆tau的减少密切相关 伴随细胞外tau的增加。因此，ExoY煽动的CUMP增加在出口中有牵连 细胞毒性牛磺酸。此外，在神经退行性疾病的背景下，降解和再循环 自噬途径通常在病理上受到限制，从而防止过量或 调节失调的淀粉样蛋白。在自噬通量被抑制后，失调的淀粉样蛋白积累和 齐聚。新生的tau寡聚体的清除通常通过胞外体和/或分泌来促进。 出口。与我们的观察一致，ExoY最近被牵连到抑制先天的 通过抑制转化生长因子-β激活激酶1而产生的免疫反应 据报道会阻碍自噬。综上所述，PMVEC中TAK1的ExoY抑制可能有助于 自噬的抑制和感染诱导的tau蛋白的产生。因此，这项提案考验着 假设ExoY诱导的CUMP有助于抑制自噬通量，从而促进 产生和释放细胞毒性tau，可传递地破坏内皮细胞间连接并促进 烫发能力。 。

PROJECT SUMMARY/ABSTRACT Pulmonary endothelial barrier integrity is vital for efficient gas exchange and to prevent the hematogenous dissemination of respiratory pathogens. Pseudomonas aeruginosa, a predominant agent of nosocomial pneumonia, breaches the endothelial barrier by inducing junctional complex disruption leading to alveolar flooding and hypoxemia. Here, we propose to study a previously unidentified, novel disease process defined by the interaction of the common nosocomial pathogen, P. aeruginosa, with the host endothelium. P. aeruginosa utilizes a Type III Secretion System (T3SS) to inject cytotoxic exoenzymes directly into the host cell. Exoenzyme Y (ExoY), a T3SS effector expressed by ~90% of clinical P. aeruginosa strains, causes marked increases in cytosolic cGMP, cUMP, and cAMP. The increase in cAMP facilitates the phosphorylation of an endothelial tau, which dissociates from microtubules, followed by microtubule collapse, interendothelial gap formation, and increased permeability. Hyperphosphorylated tau then coalesces into cytotoxic tau oligomers that are released into the extracellular milieu. These cytotoxic tau oligomers are transmissibleand self-propagating amyloid prions. The respective roles of cGMP and cUMP are currently unknown, although preliminary data establishes both a temporal and spatial correlation between the increase in ExoY-generated cUMP and oligomeric tau release from PMVECs. The cUMP signal in the bulk cytosol closely coincides with both the reduction of cytosolic tau and the concomitant increase of extracellular tau. Thus, an ExoY-instigated increase in cUMP is implicated in the export of cytotoxic tau. Furthermore, within the context of neurodegenerative disease, the degradation and recycling pathway of autophagy is often pathologically constrained thereby preventing the breakdown of excess or dysregulated amyloids. Following the inhibition of autophagic flux, dysregulated amyloids accumulate and oligomerize. Clearance of nascent tau oligomers is often facilitated via exosomal and/or secretion mediated export. Concordant with our observations, ExoY has recently been implicated in the suppression of the innate immune response via the inhibition of transforming growth factor-β-activating kinase 1 (TAK1) which has also been reported to obstruct autophagy. Taken together, ExoY suppression of TAK1 in PMVECs may contribute to the inhibition of autophagy and the production of infection-induced tau prions. Therefore, this proposal tests the hypothesis that ExoY-induced cUMP contributes to the suppression of autophagic flux thereby promoting the generation and release of cytotoxic tau that transmissibly disrupts interendothelial junctions and promotes perm eability. .