Fig 5

Contribution of the HA multiple basic amino acid cleavage site to virus replication in human endothelial cells. Cells were infected with virus at an MOI of 0.01, and supernatants were collected for virus titration by plaque assay. (A) Immunofluorescence detection of nucleoprotein (NP) antigen in infected HMVEC-LBI. Cells seeded on chamber slides were infected with virus at an MOI of 1 and fixed for NP detection at 8 h p.i. (B) Virus replication in polarized bronchial epithelial cells (Calu-3). (C) Virus replication in HMVEC-LBI. Ten percent normal chicken egg allantoic fluid was added to the medium. For real-time PCR analysis, cells were infected with each virus at an MOI of 1 and RNA was collected at 24 h p.i. (D) Relative M1 gene levels in infected cells determined by real-time PCR. (E) Relative fold change of gene expression of type I interferon genes in infected cells.