Targeted infection of endothelial cells by avian influenza virus A/FPV/Rostock/34 (H7N1) in chicken embryos

The tissue tropism and spread of infection of the highly pathogenic avian i nfluenza virus A/FPV/Rostock/34 (H7N1) (FPV) were analyzed in 11-day-old ch icken embryos, As shown by in situ hybridization, the virus caused generali zed infection that was strictly confined to endothelial cells in all organs , Studies with reassortants of FPV and the apathogenic avian strain A/chick /Germany/N/49 (H10N7) revealed that endotheliotropism was linked to FPV hem agglutinin (EW). To further analyze the factors determining endotheliotropi sm, the HA-activating protease furin was cloned from chicken tissue. Ubiqui tous expression of furin and other proprotein convertases in the chick embr yo indicated that proteolytic activation of HA was not responsible for rest riction of infection to the endothelium. To determine the expression of vir us receptors in embryonic tissues, histochemical analysis of alpha 2,3- and alpha 2,6-linked neuraminic acid was carried out by Lectin-binding assays. These receptors were found on endothelial tells and on several epithelial cells, but not on tissues surrounding endothelia. Finally, we analyzed the polarity of virus maturation in endothelial cells, Studies on cultured huma n endothelial cells employing confocal laser scanning microscopy revealed t hat EIA is specifically targeted to the apical surface of these cells, and electron microscopy of embryonic tissues showed that virus maturation occur s also at the luminar side. Taken together, these observations indicate tha t endotheliotropism of FPV in the chicken embryo is determined, on one hand , by the high cleavability of HA, which mediates virus entry into the vascu lar system, and, on the other hand, by restricted receptor expression and p olar budding, which prevent spread of infection into tissues surrounding en dothelia.