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VWA proteins of Leptospira interrogans induce hemorrhage in leptospirosis by competitive inhibition of vWF/GPIb-mediated platelet aggregation.

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-Ib? (GPIb?) 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-GPIb?-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-GPIb?-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-GPIb?-IgG, rLep-vWA-I-IgG and rLep-vWA-II-IgG blocked this binding or inhibition. SPR and ITC revealed a tight interaction between Hu-GPIb? and rLep-vWA-I/rLep-vWA-II with KD values of 3.87?×?10-7-8.65?×?10-8?M. Hu-GPIb?-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-GPIb?-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.

SUBMITTER: Fang JQ

PROVIDER: S-EPMC6284457 | biostudies-literature | 2018 Nov

REPOSITORIES: biostudies-literature

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vWA proteins of Leptospira interrogans induce hemorrhage in leptospirosis by competitive inhibition of vWF/GPIb-mediated platelet aggregation.

Fang Jia-Qi JQ Imran Muhammad M Hu Wei-Lin WL Ojcius David M DM Li Yang Y Ge Yu-Mei YM Li Kai-Xuan KX Lin Xu'ai X Yan Jie J

EBioMedicine 20181015

<h4>Backgroud</h4>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.<h4>Methods</h4>Human (Hu)-platelet- and Hu-GP ...[more]

PMID: 30337247

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Project description:BackgroundLeptospirosis is caused by pathogenic spirochetes of the genus Leptospira spp. This zoonotic disease is distributed globally and affects domestic animals, including cattle. Leptospira interrogans serogroup Sejroe serovar Hardjo and Leptospira borgpetersenii serogroup Sejroe serovar Hardjo remain important species associated with this reproductive disease in livestock production. Previous studies on Brazilian livestock have reported that L. interrogans serovar Hardjo is the most prevalent leptospiral agent in this country and is related to clinical signs of leptospirosis, which lead to economic losses in production. Here, we described the isolation of three clinical strains (Norma, Lagoa and Bolivia) obtained from leptospirosis outbreaks that occurred in Minas Gerais state in 1994 and 2008.ResultsSerological and molecular typing using housekeeping (secY and 16SrRNA) and rfb locus (ORF22 and ORF36) genes were applied for the identification and comparative analysis of Leptospira spp. Our results identified the three isolates as L. interrogans serogroup Sejroe serovar Hardjo and confirmed the occurrence of this bacterial strain in Brazilian livestock. Genetic analysis using ORF22 and ORF36 grouped the Leptospira into serogroup Sejroe and subtype Hardjoprajitno. Genetic approaches were also applied to compare distinct serovars of L. interrogans strains by verifying the copy numbers of the IS1500 and IS1533 insertion sequences (ISs). The IS1500 copy number varied among the analyzed L. interrogans strains.ConclusionThis study provides evidence that L. interrogans serogroup Sejroe serovar Hardjo subtype Hardjoprajitno causes bovine leptospirosis in Brazilian production. The molecular results suggested that rfb locus (ORF22 and ORF36) could improve epidemiological studies by allowing the identification of Leptospira spp. at the serogroup level. Additionally, the IS1500 and IS1533 IS copy number analysis suggested distinct genomic features among closely related leptospiral strains.

Dataset Information

VWA proteins of Leptospira interrogans induce hemorrhage in leptospirosis by competitive inhibition of vWF/GPIb-mediated platelet aggregation.

Publications

vWA proteins of Leptospira interrogans induce hemorrhage in leptospirosis by competitive inhibition of vWF/GPIb-mediated platelet aggregation.

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