The Pseudomonas aeruginosa exoenzyme Y impairs endothelial cell proliferation and vascular repair following lung injury.
Exoenzyme Y (ExoY) is a Pseudomonas aeruginosa toxin that is introduced into host cells through the type 3 secretion system (T3SS). Once inside the host cell cytoplasm, ExoY generates cyclic nucleotides that cause tau phosphorylation and microtubule breakdown. Microtubule breakdown causes interendothelial cell gap formation and tissue edema. Although ExoY transiently induces interendothelial cell gap formation, it remains unclear whether ExoY prevents repair of the endothelial cell barrier. Here, we test the hypothesis that ExoY intoxication impairs recovery of the endothelial cell barrier following gap formation, decreasing migration, proliferation, and lung repair. Pulmonary microvascular endothelial cells (PMVECs) were infected with P. aeruginosa strains for 6 h, including one possessing an active ExoY (PA103 exoUexoT::Tc pUCPexoY; ExoY(+)), one with an inactive ExoY (PA103ΔexoUexoT::Tc pUCPexoY(K81M); ExoY(K81M)), and one that lacks PcrV required for a functional T3SS (ΔPcrV). ExoY(+) induced interendothelial cell gaps, whereas ExoY(K81M) and ΔPcrV did not promote gap formation. Following gap formation, bacteria were removed and endothelial cell repair was examined. PMVECs were unable to repair gaps even 3-5 days after infection. Serum-stimulated growth was greatly diminished following ExoY intoxication. Intratracheal inoculation of ExoY(+) and ExoY(K81M) caused severe pneumonia and acute lung injury. However, whereas the pulmonary endothelial cell barrier was functionally improved 1 wk following ExoY(K81M) infection, pulmonary endothelium was unable to restrict the hyperpermeability response to elevated hydrostatic pressure following ExoY(+) infection. In conclusion, ExoY is an edema factor that chronically impairs endothelial cell barrier integrity following lung injury.
Center for Lung Biology, University of South Alabama, Mobile, Alabama; tstevens@southalabama.edu.
Trevor C
TC
Stevens
Physician-Scientist Training Program, Department of Medicine, University of Texas-Southwestern Medical Center, Dallas, Texas; Division of Pulmonary and Critical Care, University of Texas-Southwestern Medical Center, Dallas, Texas;
Cristhiaan D
CD
Ochoa
Department of Pharmacology, University of South Alabama, Mobile, Alabama; Center for Lung Biology, University of South Alabama, Mobile, Alabama;
K Adam
KA
Morrow
Center for Lung Biology, University of South Alabama, Mobile, Alabama;
Matthew J
MJ
Robson
Herman B Wells Center for Pediatric Research, Department of Pediatrics, Indiana University, Indianapolis, Indiana;
Nutan
N
Prasain
Center for Lung Biology, University of South Alabama, Mobile, Alabama;
Chun
C
Zhou
Department of Pharmacology, University of South Alabama, Mobile, Alabama; Department of Medicine, University of South Alabama, Mobile, Alabama; Center for Lung Biology, University of South Alabama, Mobile, Alabama;
Diego F
DF
Alvarez
Department of Microbiology and Molecular Genetics, Medical College of Wisconsin, Milwaukee, Wisconsin; and Center for Infectious Disease Research, Medical College of Wisconsin, Milwaukee, Wisconsin.
Dara W
DW
Frank
Department of Cell Biology and Neuroscience, University of South Alabama, Mobile, Alabama; Center for Lung Biology, University of South Alabama, Mobile, Alabama;
Ron
R
Balczon
Department of Pharmacology, University of South Alabama, Mobile, Alabama; Department of Medicine, University of South Alabama, Mobile, Alabama; Center for Lung Biology, University of South Alabama, Mobile, Alabama;
Troy
T
Stevens
D000818:Animals; D001426:Bacterial Proteins / Q000502:physiology*; D049109:Cell Proliferation; D000242:Cyclic AMP / Q000378:metabolism; D004487:Edema / Q000276:immunology / Q000382:microbiology; D042783:Endothelial Cells / Q000276:immunology / Q000382:microbiology* / Q000502:physiology; D005964:Glucosyltransferases / Q000502:physiology*; D054884:Host-Pathogen Interactions; D008168:Lung / Q000098:blood supply / Q000276:immunology / Q000382:microbiology; D055370:Lung Injury / Q000276:immunology / Q000382:microbiology; D008297:Male; D055806:Microvessels / Q000473:pathology / Q000503:physiopathology; D018410:Pneumonia, Bacterial / Q000276:immunology* / Q000382:microbiology / Q000473:pathology; D011552:Pseudomonas Infections / Q000276:immunology* / Q000382:microbiology / Q000473:pathology; D011550:Pseudomonas aeruginosa / Q000201:enzymology* / Q000276:immunology / Q000502:physiology; D051381:Rats
D001426:Bacterial Proteins; D000242:Cyclic AMP; C111504:ExoY protein, bacteria; D005964:Glucosyltransferases
cyclase; microtubules; permeability; pulmonary edema; tau