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vWA proteins of Leptospira interrogans induce hemorrhage in leptospirosis by competitive inhibition of vWF/GPIb-mediated platelet aggregation
- Source :
- EBioMedicine
- Publication Year :
- 2018
- Publisher :
- Elsevier BV, 2018.
-
Abstract
- Backgroud Leptospira interrogans is the major causative agent of leptospirosis, a worldwide zoonotic disease. Hemorrhage is a typical pathological feature of leptospirosis. Binding of von Willebrand factor (vWF) to platelet glycoprotein-Ibα (GPIbα) is a crucial step in initiation of platelet aggregation. The products of L. interrogans vwa-I and vwa-II genes contain vWF-A domains, but their ability to induce hemorrhage has not been determined. Methods Human (Hu)-platelet- and Hu-GPIbα-binding abilities of the recombinant proteins expressed by L. interrogans strain Lai vwa-I and vwa-II genes (rLep-vWA-I and rLep-vWA-II) were detected by flowcytometry, surface plasmon resonance (SPR) and isothermal titration calorimetry (ITC). Hu-platelet aggregation and its signaling kinases and active components were detected by lumiaggregometry, Western analysis, spectrophotometry and confocal microscopy. Hu-GPIbα-binding sites in rLep-vWA-I and rLep-vWA-II were identified by SPR/ITC measurements. Findings Both rLep-vWA-I and rLep-vWA-II were able to bind to Hu-platelets and inhibit rHu-vWF/ristocetin-induced Hu-platelet aggregation, but Hu-GPIbα-IgG, rLep-vWA-I-IgG and rLep-vWA-II-IgG blocked this binding or inhibition. SPR and ITC revealed a tight interaction between Hu-GPIbα and rLep-vWA-I/rLep-vWA-II with KD values of 3.87 × 10−7-8.65 × 10−8 M. Hu-GPIbα-binding of rL-vWA-I/rL-vWA-II neither activated the PI3K/AKT-ERK and PLC/PKC kinases nor affected the NO, cGMP, ADP, Ca2+ and TXA2 levels in Hu-platelets. G13/R36/G47 in Lep-vWA-I and G76/Q126 in Lep-vWA-II were confirmed as the Hu-GPIbα-binding sites. Injection of rLep-vWA-I or rLep-vWA-II in mice resulted in diffuse pulmonary and focal renal hemorrhage but this hemorrhage was blocked by rLep-vWA-I-IgG or rLep-vWA-II-IgG. Interpretation The products of L. interrogans vwa-I and vwa-II genes induce hemorrhage by competitive inhibition of vWF-mediated Hu-platelet aggregation.
- Subjects :
- 0301 basic medicine
Research paper
Clinical Sciences
030106 microbiology
Renal Hemorrhage
Competitive inhibition
Hemorrhage
von Willebrand factor
General Biochemistry, Genetics and Molecular Biology
Hemontage
law.invention
Mice
03 medical and health sciences
Bacterial Proteins
Von Willebrand factor
law
Human Umbilical Vein Endothelial Cells
Animals
Humans
Leptospirosis
Platelet aggregation
Platelet
Protein kinase C
vwa-I and vwa-II genes
biology
Chemistry
Kinase
Platelet Glycoprotein GPIb-IX Complex
General Medicine
biology.organism_classification
Molecular biology
Good Health and Well Being
030104 developmental biology
Public Health and Health Services
biology.protein
Recombinant DNA
Female
Leptospira interrogans
Biotechnology
Subjects
Details
- ISSN :
- 23523964
- Volume :
- 37
- Database :
- OpenAIRE
- Journal :
- EBioMedicine
- Accession number :
- edsair.doi.dedup.....7f499ced1c66abc238f58dbfc81db904