Infection of Endothelial and Mesothelial Cells with Andes Virus and NK Response

SCHIERLOH P.; MARTINEZ V; LANDONI V.I; FERNÁNDEZ G C; SASIAIN M.C.; PADULA P

Beijing

Conferencia; IX International Conference on HFRS HPS & Hantaviruses; 2013

SAI

In the early phase of the infection pathogenic hantaviruses escape induction of innate antiviral responses, specifically innate interferon system. The rol of NK cells in hantavirus infection remains to be investigated. Lung microvascular endothelium is considered to be the primary site for hantavirus replication. To determine whether two human pulmonary cell types support Andes virus (ANDV) replication and contribute to the earliest induction of the antiviral response, human microvascular pulmonary endothelial cell line HMEC-1 and cultured primary human mesothelial cells were mock infected or infected with ANDV. Infection of cells was confirmed by immunofluorescence and cell morphology was analyzed by confocal microscopy. Expression of cell surface markers related to NK cell activation was analyzed by flow cytometry. Viral RNA, infectious particles and cytokine and chemokine responses were quantified in culture media of both cell types. Infected cells were identified by the characteristically intra-cytoplasmatic pattern of staining for viral nucleoprotein (NP). The infection caused morphological changes in HMEC-1 and mesothelial cells. By day 4 post infection (p.i.) more than 20% of cells were positive for NP staining in HMEC-1 infected at a multiplicity of infection of 0.1. At the same day viral RNA in culture media increased drastically to reach 2, 5 x 109 S segment molecules per ml. The kinetics of infection in HMEC-1 was similar to that seen in Vero E6 cells, although in mesothelial cells the titer reached up to 4 days p.i. was lower. Expression of IL-1 and IL-8 and major histocompatibility complex class I (MHC-I), ICAM and CD80/CD86 surface markers were evaluated in each cell type. ANDV infection in HMEC-1and mesothelial cells significantly increased MHC-1 expression in cell surface and IL-8 secretion. However, ANDV seems to be unable to alter the pattern induced by some added inflammatory mediators such as TNF and IFN-g.