Elevated Cytokines, Thrombin and PAI-1 in Severe HCPS Patients Due to Sin Nombre Virus

Abstract

Sin Nombre Hantavirus (SNV, Bunyaviridae Hantavirus) is a Category A pathogen that causes Hantavirus Cardiopulmonary Syndrome (HCPS) with case fatality ratios generally ranging from 30% to 50%. HCPS is characterized by vascular leakage due to dysregulation of the endothelial barrier function. The loss of vascular integrity results in non-cardiogenic pulmonary edema, shock, multi-organ failure and death. Using Electric Cell-substrate Impedance Sensing (ECIS) measurements, we found that plasma samples drawn from University of New Mexico Hospital patients with serologically-confirmed HCPS, induce loss of cell-cell adhesion in confluent epithelial and endothelial cell monolayers grown in ECIS cultureware. We show that the loss of cell-cell adhesion is sensitive to both thrombin and plasmin inhibitors in mild cases, and to thrombin only inhibition in severe cases, suggesting an increasing prothrombotic state with disease severity. A proteomic profile (2D gel electrophoresis and mass spectrometry) of HCPS plasma samples in our cohort revealed robust antifibrinolytic activity among terminal case patients. The prothrombotic activity is highlighted by acute ≥30 to >100 fold increases in active plasminogen activator inhibitor (PAI-1) which, preceded death of the subjects within 48 h. Taken together, this suggests that PAI-1 might be a response to the severe pathology as it is expected to reduce plasmin activity and possibly thrombin activity in the terminal patients.

RIG-I-like receptor activation drives type I IFN and antiviral signaling to limit Hantaan orthohantavirus replication

“…Although, the vascular endothelium is the primary cellular target of hantavirus infection in both humans and in reservoir hosts, infection drives very different outcomes of acute pathogenesis in humans while persistent, nonpathogenic infection occurs in reservoir hosts [10]. Hantavirus disease in humans begins with intense muscle pain, fever, and nausea, accompanied by elevated levels of the proinflammatory cytokines IL-1β, TNFα, and IL-6 and others [2,[11][12][13]. Vascular leakage, either in the lung microvasculature or kidneys, is the hallmark of disease and can be linked to lethal disease.…”

Electric Cell-Substrate Impedance Sensing To Monitor Viral Growth and Study Cellular Responses to Infection with Alphaherpesviruses in Real Time

“…Thus far, ECIS has been used primarily to study viral growth and cellular responses to infection with RNA viruses, such as respiratory syncytial virus, influenza A virus (IAV), and Sin Nombre virus (7–9). Additionally, one study described the use of ECIS to study the effects of transient overexpression of Kaposi’s sarcoma-associated herpesvirus proteins MIR-1 and MIR-2 on the attachment, spreading, and junction formation of immortalized dermal microvascular endothelial cells (10).…”

Small-Volume Flow Cytometry-Based Multiplex Analysis of the Activity of Small GTPases

“…These signals ultimately combine to induce profound changes in vascular endothelial cells, including increased endothelial monolayer permeability [ 44 , 47 , 48 ]. High concentrations of thrombin expressed in the circulation of HCPS subjects significantly contribute to loss of cell barrier function in endothelial cells [ 18 ]. Argatroban, an orthosteric inhibitor of thrombin ( K i ~ 10 −8 M), can be used to block thrombin activity [ 49 ].…”

“…The first example measures the signaling cascade of GTPases that are activated to allow β 3 integrin-mediated cellular entry of Sin Nombre virus (SNV) in the course of a productive infection [ 16 , 17 ]. The second example measures GTPase activity downstream of signaling of protease-activated receptors (PARs), after exposure to thrombin found in the plasma samples drawn from patients with hantavirus cardiopulmonary syndrome (HCPS) [ 18 ]. The third example measures GTPase activation due to bacterial factors [ 12 , 19 ] present in a plasma sample from a septic patient.…”