Endothelial Cell Permeability during Hantavirus Infection Involves Factor XII-Dependent Increased Activation of the Kallikrein-Kinin System

Abstract

Hemorrhagic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome (HPS) are diseases caused by hantavirus infections and are characterized by vascular leakage due to alterations of the endothelial barrier. Hantavirus-infected endothelial cells (EC) display no overt cytopathology; consequently, pathogenesis models have focused either on the influx of immune cells and release of cytokines or on increased degradation of the adherens junction protein, vascular endothelial (VE)-cadherin, due to hantavirus-mediated hypersensitization of EC to vascular endothelial growth factor (VEGF). To examine endothelial leakage in a relevant in vitro system, we co-cultured endothelial and vascular smooth muscle cells (vSMC) to generate capillary blood vessel-like structures. In contrast to results obtained in monolayers of cultured EC, we found that despite viral replication in both cell types as well as the presence of VEGF, infected in vitro vessels neither lost integrity nor displayed evidence of VE-cadherin degradation. Here, we present evidence for a novel mechanism of hantavirus-induced vascular leakage involving activation of the plasma kallikrein-kinin system (KKS). We show that incubation of factor XII (FXII), prekallikrein (PK), and high molecular weight kininogen (HK) plasma proteins with hantavirus-infected EC results in increased cleavage of HK, higher enzymatic activities of FXIIa/kallikrein (KAL) and increased liberation of bradykinin (BK). Measuring cell permeability in real-time using electric cell-substrate impedance sensing (ECIS), we identified dramatic increases in endothelial cell permeability after KKS activation and liberation of BK. Furthermore, the alterations in permeability could be prevented using inhibitors that directly block BK binding, the activity of FXIIa, or the activity of KAL. Lastly, FXII binding and autoactivation is increased on the surface of hantavirus-infected EC. These data are the first to demonstrate KKS activation during hantavirus infection and could have profound implications for treatment of hantavirus infections.

Hypoxia Induces Permeability and Giant Cell Responses of Andes Virus-Infected Pulmonary Endothelial Cells by Activating the mTOR-S6K Signaling Pathway

“…One study suggests that hantavirusinfected ECs transiently degraded VE-cadherin at a single specific time point (98). Consistent with our findings, another study of infected HUVECs demonstrated no change in VE-cadherin degradation in cells grown persistently in VEGF but didn't assess VEcadherin internalization or VEGF responsiveness (75). Interestingly, this paper demonstrates a role for kallikrein-directed bradykinin responses to increase the permeability of hantavirusinfected ECs (75).…”

“…HPS patients often seek medical attention as a result of the sudden onset of respiratory distress resulting from pulmonary edema that accumulates by as much as 1.0 liter per hour (2,(11)(12)(13). Potential pathogenic mechanisms accounting for the high rate of pulmonary fluid accumulation have yet to be demonstrated (13,18,22,41,47,48,(72)(73)(74)(75)(76) but appear be a consequence of the noncytolytic hantavirus infection of endothelial cells. Although, MECs and LECs are not permeabilized by hantavirus infection alone (47,(49)(50)(51)(52), hantavirus infection of the endothelium provides a means for the virus to alter EC responses that normally regulate capillary leakage and pulmonary fluid clearance.…”

“…Although, MECs and LECs are not permeabilized by hantavirus infection alone (47,(49)(50)(51)(52), hantavirus infection of the endothelium provides a means for the virus to alter EC responses that normally regulate capillary leakage and pulmonary fluid clearance. In vitro data indicate that ANDV infection of human pulmonary MECs and LECs alters VEGF A-mTOR signaling responses within ECs (40,41,47,(49)(50)(51)(52)65), and recent studies suggest that bradykinin may contribute to hantavirus-directed vascular permeability (75,76).…”

“…also participate in hypoxia-induced pulmonary edema during hantavirus infection that include responses of T cells (105), induced cytokines (106), and recently reported findings of kalikrein-directed bradykinin permeability responses (75,76) that may also enhance VEGFR2 signaling responses (82,83,85,99). Explanations for acute thrombocytopenia during hantavirus infections have yet to be defined.…”

Coagulation Factor XII –A Key Pro-Inflammatory and Pro-Coagulant Protein

“…This suggests that FXII can be activated following local activation of endothelial cells. This is consistent with a previous study in which FXII was activated following Hantavirus-induced hypersensilisation of endothelial cells to VEGF [15]. In another in vitro study, the inflammatory cytokine BK stimulated cultured endothelial cells to secrete plasminogen, prostacyclin, thromboxane A 2 and NO, thereby regulating platelet function and promote fibrinolysis [16].…”