CHARACTERIZATION OF ENDOTHELIAL CELL INFECTION BY JUNIN VIRUS.

POZNER, ROBERTO G.; GOMEZ, RICARDO M.; LAZZARI, MARIA A.; LITWAK, SARA; MAUGERI, NORMA; SCHATTNER, MIRTA

Paris, Francia

Congreso; XIXth International Society on Thrombosis and Haemostasis Congress; 2001

International Society on Thrombosis and Haemostasis

Junin virus (JV), a member of the arenaviridae family, is the causative agent of Argentine Hemorrhagic Fever (AHF). The clinical manifestations of this disease include thrombocytopenia and several alterations in blood coagulation and the fibrinolytic system without an evident intravascular coagulation. We tested the hypothesis that endothelial cells (EC) play an important role in the defective hemostasis that occurs in AHF. EC derived from human umbilical vein (HUVECs) were infected with 103 PFU of JV. Seven days post infection, viral infectivity, 6-ceto-PGF1a (PGF1a), nitric oxide (NO) and von Willebrand Factor (vWF) in supernatants, and expression of VCAM-1 or ICAM-1 on cells were determined. Viral infectivity oscillated between 103 and 104 PFU/mL and no cytopathic effect was observed by phase contrast microscopy although expression of viral antigen was detected by flow cytometry and immunocytochemical procedure. Infected and non-infected cells reached confluent monolayer at the same time. Production of PGF1a (pg/mL, X±SEM, n=3) (ELISA) and NO (uM, n=9) (Griess reaction) was increased in infected cells compared to control samples (44±7 vs. 16±5 and 2.7±0.3 vs. 1.9±0.3 respectively, p<0.05). Integrin expression evaluated by flow cytometry showed that basal levels of both ICAM-1 and VCAM-1 (mean fluorescence intensity, n=6) augmented in infected cells (control: 35±8 and 13±3 vs. 64±18 and 22±2 p<0.05). Stimulation of normal HUVECs by TNF-a resulted in a 5 fold ICAM-1 increases while in JV-infected cells the increase was only of 2 fold. It was observed that intracellular content of vWF was diminished in permeabilized infected cells compared to non-infected (323±37 vs. 460±72, p<0.05, n=4). This reduction seemed to be related to a decrease in vWF synthesis since the amount of vWF release (ng/mL) in the virus infected culture medium (218±45) was lower (p<0.05 n=4) than in control samples (336±30). In addition, flow cytometry analysis of non-permeabilized infected HUVECs showed no vWF associated to the cell surface. These results suggest that endothelium effector responses are triggered after viral infection and it might have an important role in the pathogenesis of AHF.