Effect of Puumala hantavirus infection on human umbilical vein endothelial cell hemostatic function: platelet interactions, increased tissue factor expression and fibrinolysis regulator release

Abstract

Puumala virus (PUUV) infection causes over 5000 cases of hemorrhagic fever in Europe annually and can influence the hemostatic balance extensively. Infection might lead to hemorrhage, while a recent study showed an increased risk of myocardial infarction during or shortly after PUUV infection. The mechanism by which this hantavirus influences the coagulation system remains unknown. Therefore we aimed to elucidate mechanisms explaining alterations seen in primary and secondary hemostasis during PUUV infection. By using low passage PUUV isolates to infect primary human umbilical vein endothelial cells (HUVECs) we were able to show alterations in the regulation of primary- and secondary hemostasis and in the release of fibrinolysis regulators. Our main finding was an activation of secondary hemostasis due to increased tissue factor (TF) expression leading to increased thrombin generation in a functional assay. Furthermore, we showed that during infection platelets adhered to HUVEC and subsequently specifically to PUUV virus particles. Infection of HUVEC with PUUV did not result in increased von Willebrand factor while they produced more plasminogen activator inhibitor type-1 (PAI-1) compared to controls. The PAI-1 produced in this model formed complexes with vitronectin. This is the first report that reveals a potential mechanism behind the pro-coagulant changes in PUUV patients, which could be the result of increased thrombin generation due to an increased TF expression on endothelial cells during infection. Furthermore, we provide insight into the contribution of endothelial cell responses regarding hemostasis in PUUV pathogenesis.

Thrombocytopenia associates with the severity of inflammation and variables reflecting capillary leakage in Puumala Hantavirus infection, an analysis of 546 Finnish patients

“…[33] Endothelial activation markers associate with thrombocytopenia [34] and pathogenic hantaviruses are known to direct the adherence of quiescent platelets through b3-integrin to the surface of infected endothelial cells [35] and specifically to PUUV particles. [36] The adherence of platelets may result in altered platelet activation and decreased circulating platelet count as well as a loss of vascular integrity. Increased thrombin formation and remarkable fibrinolysis, as well as decreased natural anticoagulants and altered platelet ligands reported in PUUV infected patients support the role of activated platelets and endothelium in the pathogenesis of hantavirus infection.…”

Influenza virus damages the alveolar barrier by disrupting epithelial cell tight junctions

“…Time to thrombin generation was measured essentially as described previously [29]. See the supplementary material for details.…”

The Association between Hantavirus Infection and Selenium Deficiency in Mainland China

“…To study the influence of selenium concentration on hantavirus replication in a well-established immune competent cell line of human origin we infected Human Umbilical Vein Endothelial Cells (HUVEC) with a low passage isolate of Puumala hantavirus (PUUV) with or without selenium supplementation. Much effort has been put in the isolation of PUUV within four passages over human cell lines to retain its virulence [ 29 ] ( Figure 4 shows the viral copy numbers in each replicate measured every 24 h up to 72 h post infection of HUVEC’s with a multiplicity of infection (MOI) of 0.05. Viral copy numbers significantly decreased after selenium supplementation of 200 ng/mL, suggesting a decrease in viral replication.…”