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ExoU-induced procoagulant activity in Pseudomonas aeruginosa-infected airway cells

Departamento de Microbiologia, Imunologia e Parasitologia, Faculdade de Ciências Médicas, Universidade do Estado do Rio de Janeiro, Rio de Janeiro, Brazil.

CORRESPONDENCE: M. C. Plotkowski, Departamento de Microbiologia, Imunologia e Parasitologia, Faculdade de Ciências Médicas UERJ, Av. 28 de Setembro, 87 fundos, 3° andar. 20 551-030, Rio de Janeiro, Brazil. Fax: 55 2125876476. E-mail: crisplot{at}yahoo.com.br

Keywords: ExoU, lung injury, procoagulant activity, Pseudomonas aeruginosa, sepsis, tissue factor

Received: June 7, 2008
Accepted July 19, 2008

The present study addressed the question whether ExoU, a Pseudomonas aeruginosa toxin with phospholipase A2 (PLA2) activity, may induce airway epithelial cells to overexpress tissue factor (TF) and exhibit a procoagulant phenotype.

Cells from the human bronchial epithelial BEAS-2B line were infected with an ExoU-producing P. aeruginosa strain, pre-treated or not with the cytosolic PLA2 inhibitor methylarachidonyl fluorophosphate (MAFP), or with two ExoU-deficient mutants. Control noninfected and infected cells were assessed for the expression of: 1) TF mRNA by RT-PCR; 2) cell-associated TF by enzyme immunoassay and flow cytometry; 3) procoagulant activity by a colorimetric assay; and 4) microparticle-associated TF by flow cytometry. An enzyme immunoassay was also used to assess cell-associated TF in lung extracts from mice infected intratracheally with ExoU-producing and -deficient bacteria.

Cells infected with the wild-type bacteria had higher levels of TF mRNA, cell-associated TF expression, procoagulant activity and released microparticle-associated TF than cells infected with the mutants. Bacterial treatment with MAFP significantly reduced the expression of TF by infected cells. Lung samples from mice infected with the wild-type bacteria exhibited higher levels of cell-associated TF and procoagulant activity.

The present results demonstrate that ExoU may contribute to the pathogenesis of lung injury by inducing a tissue factor-dependent procoagulant activity in airway epithelial cells.