Pseudomonas aeruginosa uses a type III secretion system to introduce the adenylyl and guanylyl cyclase exotoxin Y (ExoY) into the cytoplasm of endothelial cells. ExoY induces Tau hyperphosphorylation and insolubility, microtubule breakdown, barrier disruption and edema, although the mechanism(s) responsible for microtubule breakdown remain poorly understood. Here we investigated both microtubule behavior and centrosome activity to test the hypothesis that ExoY disrupts microtubule dynamics. Fluorescence microscopy determined that infected pulmonary microvascular endothelial cells contained fewer microtubules than control cells, and further studies demonstrated that the microtubule-associated protein Tau was hyperphosphorylated following infection and dissociated from microtubules. Disassembly/reassembly studies determined that microtubule assembly was disrupted in infected cells, with no detectable effects on either microtubule disassembly or microtubule nucleation by centrosomes. This effect of ExoY on microtubules was abolished when the cAMP-dependent kinase phosphorylation site (Ser-214) on Tau was mutated to a non-phosphorylatable form. These studies identify Tau in microvascular endothelial cells as the target of ExoY in control of microtubule architecture following pulmonary infection by Pseudomonas aeruginosa and demonstrate that phosphorylation of tau following infection decreases microtubule assembly.

Pneumonia causes short‐ and long‐term cognitive dysfunction in a high proportion of patients, although the mechanism(s) responsible for this effect are unknown. Here, we tested the hypothesis that pneumonia‐elicited cytotoxic amyloid and tau variants: (1) are present in the circulation during infection; (2) lead to impairment of long‐term potentiation; and, (3) inhibit long‐term potentiation dependent upon tau. Cytotoxic amyloid and tau species were recovered from the blood and the hippocampus following pneumonia, and they were present in the extracorporeal membrane oxygenation oxygenators of patients with pneumonia, especially in those who died. Introduction of immunopurified blood‐borne amyloid and tau into either the airways or the blood of uninfected animals acutely and chronically impaired hippocampal information processing. In contrast, the infection did not impair long‐term potentiation in tau knockout mice and the amyloid‐ and tau‐dependent disruption in hippocampal signaling was less severe in tau knockout mice. Moreover, the infection did not elicit cytotoxic amyloid and tau variants in tau knockout mice. Therefore, pneumonia initiates a tauopathy that contributes to cognitive dysfunction.

Lung endothelia in the arteries, capillaries, and veins are heterogeneous in structure and function. Lung capillaries in particular represent a unique vascular niche, with a thin yet highly restrictive alveolar-capillary barrier that optimizes gas exchange. Capillary endothelium surveys the blood while simultaneously interpreting cues initiated within the alveolus and communicated via immediately adjacent type I and type II epithelial cells, fibroblasts, and pericytes. This cell-cell communication is necessary to coordinate the immune response to lower respiratory tract infection. Recent discoveries identify an important role for the microtubule-associated protein tau that is expressed in lung capillary endothelia in the host-pathogen interaction. This endothelial tau stabilizes microtubules necessary for barrier integrity, yet infection drives production of cytotoxic tau variants that are released into the airways and circulation, where they contribute to end-organ dysfunction. Similarly, beta-amyloid is produced during infection. Beta-amyloid has antimicrobial activity, but during infection it can acquire cytotoxic activity that is deleterious to the host. The production and function of these cytotoxic tau and amyloid variants are the subject of this review. Lung-derived cytotoxic tau and amyloid variants are a recently discovered mechanism of end-organ dysfunction, including neurocognitive dysfunction, during and in the aftermath of infection.

Patients who recover from pneumonia subsequently have elevated rates of death after hospital discharge as a result of secondary organ damage, the causes of which are unknown. We used the bacterium Pseudomonas aeruginosa , a common cause of hospital‐acquired pneumonia, as a model for investigating this phenomenon. We show that infection of pulmonary endothelial cells by P. aeruginosa induces production and release of a cytotoxic amyloid molecule with prion characteristics, including resistance to various nucleases and proteases. This cytotoxin was self‐propagating, was neutralized by anti‐amyloid Abs, and induced death of endothelial cells and neurons. Moreover, the cytotoxin induced edema in isolated lungs. Endothelial cells and isolated lungs were protected from cytotoxin‐induced death by stimulation of signal transduction pathways that are linked to prion protein. Analysis of bronchoalveolar lavage fluid collected from human patients with P. aeruginosa pneumonia demonstrated cytotoxic activity, and lavage fluid contained amyloid molecules, including oligomeric t and Ab. Demonstration of long‐lived cytotoxic agents after Pseudomonas infection may establish a molecular link to the high rates of death as a result of end‐organ damage in the months after recovery from pneumonia, and modulation of signal transduction pathways that have been linked to prion protein may provide a mechanism for intervention.—Balczon, R., Morrow, K. A., Zhou, C., Edmonds, B., Alexeyev, M., Pittet, J.‐F., Wagener, B. M., Moser, S. A., Leavesley, S., Zha, X., Frank, D. W., Stevens, T. Pseudomonas aeruginosa infection liberates transmissible, cytotoxic prion amyloids. FASEB J. 31, 2785–2796 (2017). www.fasebj.org

Patients who recover from hospital‐acquired pneumonia exhibit a high incidence of end‐organ dysfunction following hospital discharge, including cognitive deficits. We have previously demonstrated that pneumonia induces the production and release of cytotoxic oligomeric tau from pulmonary endothelial cells, and these tau oligomers can enter the circulation and may be a cause of long‐term morbidities. Endothelial‐derived oligomeric tau is hyperphosphorylated during infection. The purpose of these studies was to determine whether Ser‐214 phosphorylation of tau is a necessary stimulus for generation of cytotoxic tau variants. The results of these studies demonstrate that Ser‐214 phosphorylation is critical for the cytotoxic properties of infection‐induced oligomeric tau. In the lung, Ser‐214 phosphorylated tau contributes to disruption of the alveolar–capillary barrier, resulting in increased permeability. However, in the brain, both the Ser‐214 phosphorylated tau and the mutant Ser‐214‐Ala tau, which cannot be phosphorylated, disrupted hippocampal long‐term potentiation suggesting that inhibition of long‐term potentiation was relatively insensitive to the phosphorylation status of Ser‐214. Nonetheless, phosphorylation of tau is essential to its cytotoxicity since global dephosphorylation of the infection‐induced cytotoxic tau variants rescued long‐term potentiation. Collectively, these data demonstrate that multiple forms of oligomeric tau are generated during infectious pneumonia, with different forms of oligomeric tau being responsible for dysfunction of distinct end‐organs during pneumonia.

Patients who survive pneumonia have elevated death rates in the months following hospital discharge. It has been hypothesized that infection of pulmonary tissue during pneumonia results in the production of long-lived cytotoxins that can lead to subsequent end organ failure. We have developed in vitro assays to test the hypothesis that cytotoxins are produced during pulmonary infection. Isolated rat pulmonary endothelial cells and the bacterium Pseudomonas aeruginosa are used as model systems, and the production of cytoxins following infection of the endothelial cells by the bacteria is demonstrated using cell culture followed by direct quantitation using lactate dehydrogenase assays and a novel microscopic method utilizing ImageJ technology. The amyloid nature of these cytotoxins was demonstrated by thioflavin T binding assays and by immunoblotting and immunodepletion using A11 anti-amyloid antibody. Further analyses using immunoblotting demonstrated that oligomeric tau and Aβ were produced and released by endothelial cells following infection by P. aeruginosa. These methods should be readily adaptable to analyses of human clinical samples.