In the Absence of Effector Proteins, the Pseudomonas aeruginosa Type Three Secretion System Needle Tip Complex Contributes to Lung Injury and Systemic Inflammatory Responses

Abstract

Herein we describe a pathogenic role for the Pseudomonas aeruginosa type three secretion system (T3SS) needle tip complex protein, PcrV, in causing lung endothelial injury. We first established a model in which P. aeruginosa wild type strain PA103 caused pneumonia-induced sepsis and distal organ dysfunction. Interestingly, a PA103 derivative strain lacking its two known secreted effectors, ExoU and ExoT [denoted PA103 (ΔU/ΔT)], also caused sepsis and modest distal organ injury whereas an isogenic PA103 strain lacking the T3SS needle tip complex assembly protein [denoted PA103 (ΔPcrV)] did not. PA103 (ΔU/ΔT) infection caused neutrophil influx into the lung parenchyma, lung endothelial injury, and distal organ injury (reminiscent of sepsis). In contrast, PA103 (ΔPcrV) infection caused nominal neutrophil infiltration and lung endothelial injury, but no distal organ injury. We further examined pathogenic mechanisms of the T3SS needle tip complex using cultured rat pulmonary microvascular endothelial cells (PMVECs) and revealed a two-phase, temporal nature of infection. At 5-hours post-inoculation (early phase infection), PA103 (ΔU/ΔT) elicited PMVEC barrier disruption via perturbation of the actin cytoskeleton and did so in a cell death-independent manner. Conversely, PA103 (ΔPcrV) infection did not elicit early phase PMVEC barrier disruption. At 24-hours post-inoculation (late phase infection), PA103 (ΔU/ΔT) induced PMVEC damage and death that displayed an apoptotic component. Although PA103 (ΔPcrV) infection induced late phase PMVEC damage and death, it did so to an attenuated extent. The PA103 (ΔU/ΔT) and PA103 (ΔPcrV) mutants grew at similar rates and were able to adhere equally to PMVECs post-inoculation indicating that the observed differences in damage and barrier disruption are likely attributable to T3SS needle tip complex-mediated pathogenic differences post host cell attachment. Together, these infection data suggest that the T3SS needle tip complex and/or another undefined secreted effector(s) are important determinants of P. aeruginosa pneumonia-induced lung endothelial barrier disruption.

Achromobacter Respiratory Infections

“…Such a system is classically found in Bordetella, Yersinia, Vibrio, and Salmonella species, as well as in Pseudomonas aeruginosa. In fact, the needle tip complex of the latter organism's T3SS has recently been shown to induce alveolar injury and an inflammatory response, even in the absence of effector proteins (44).…”

Mitochondrial DNA Damage Initiates Acute Lung Injury and Multi-Organ System Failure Evoked in Rats by Intra-Tracheal Pseudomonas Aeruginosa

“…It remains unknown, however, whether enhanced mtDNA repair forestalls progression of ALI to MOSF. Accordingly, here we took advantage of a rat model of PA103-induced lung endothelial injury and distant organ dysfunction recently described by Audia and coworkers (14) to test the hypothesis that pharmacologic enhancement of mtDNA repair prevents and reverses bacteria-related ALI, MOSF, and mortality.…”

Dephosphorylation of Y685-VE-Cadherin Involved in Pulmonary Microvascular Endothelial Barrier Injury Induced by Angiotensin II

“…The mixture was incubated at room temperature for 30 min. Fluoromount-G (Southern Biotech, category number 0100-01, Birmingham, USA) was used to block the coverslip [22]. Finally, the images were observed using a BX53 fluorescence microscope (Olympus, Tokyo, Japan).…”