Vacuolisation of human microvascular endothelial cells by enterohaemorrhagic Escherichia coli

Martina Bielaszewska; Andreas Bauwens; Lilo Greune; Björn Kemper; Ulrich Dobrindt; Joyce M. Geelen; Kwang S. Kim; M. Alexander Schmidt; Helge Karch

doi:10.1160/TH09-07-0499

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 2009; 102(06): 1080-1092
DOI: 10.1160/TH09-07-0499

Theme Issue Article

Schattauer GmbH

Vacuolisation of human microvascular endothelial cells by enterohaemorrhagic Escherichia coli

Authors

Martina Bielaszewska^*

¹Institute of Hygiene, University of Münster, Münster, Germany
Andreas Bauwens^*

¹Institute of Hygiene, University of Münster, Münster, Germany
Lilo Greune

²Institute of Infectiology, Center for Molecular Biology of Inflammation (ZMBE), University of Münster, Münster, Germany
Björn Kemper

³Center for Biomedical Optics and Photonics, University of Münster, Münster, Germany
Ulrich Dobrindt

⁴Institute for Molecular Infection Biology, University of Würzburg, Würzburg, Germany
Joyce M. Geelen

⁵Department of Pediatric Nephrology, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands
Kwang S. Kim

⁶Division of Pediatric Infectious Diseases, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA
M. Alexander Schmidt

²Institute of Infectiology, Center for Molecular Biology of Inflammation (ZMBE), University of Münster, Münster, Germany
Helge Karch

¹Institute of Hygiene, University of Münster, Münster, Germany

Abstract

Summary

Enterohaemorrhagic Escherichia coli (EHEC) cause haemolytic uraemic syndrome (HUS),a thrombotic microangiopathy resulting from endothelial injury in the renal glomeruli and other organs. EHEC virulence factors that damage the microvascular endothelium play therefore major roles in the pathogenesis of HUS.We identified an EHEC strain that vacuolates and kills primary human glomerular microvascular endothelial cells (GMVECs) and a human brain microvascular endothelial cell (HBMEC) line. Because the vacuolating effect closely resembles those elicited on other cells by the vacuolating cytotoxin of Helicobacter pylori (VacA), we designated the factor responsible for this effect EHEC vacuolating cytotoxin (EHEC-Vac). EHEC-Vac (a secreted non-serine protease protein) binds to HBMECs rapidly and irreversibly, vacuolates within 30 min after exposure and the effect is maximally apparent at 48 h. Despite the vacuolisation, HBMECs survive for several days before they undergo necrosis. Electron and immunofluorescence microscopy demonstrate that the vacuoles induced by EHEC-Vac originate from lysosomes.Accordingly, they stain with neutral red indicating an acidic microenvironment. Similar to VacA, the EHEC-Vac-mediated vacuolisation is both prevented and reverted by the vacuolar proton pump inhibitor bafilomycin A1, suggesting a similar mechanism of vacuole formation by these toxins. Despite the similarity of phenotypes elicited by EHEC-Vac and VacA, genomic DNA from the EHEC-Vac-producing strain failed to hybridise to a vacA probe, as well as to probes derived from presently known E. coli vacuolating toxins.Through its microvascular endothelium-injuring potential combined with the ability to induce interleukin 6 release from these cells EHEC-Vac might contribute to the pathogenesis of HUS.

Keywords

Enterohaemorrhagic E. coli - haemolytic uraemic syndrome - vacuolating cytotoxin - EHEC-Vac - microvascular endothelial injury

Full Text

References

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