The shortest route to the brain is through the olfactory system. This consists of olfactory neurons (OSNs) embedded in epithelial cells. OSNs extend their axons into the lamina propria (LP) where they are wrapped by olfactory glia (OECs) and fibroblasts forming the olfactory nerve connecting to the brain. Highly pathogenic strains of influenza virus have been reported to target cells within the olfactory system disseminating into the CNS. While some strains of influenza are restricted within the olfactory system following experimental intranasal infection. However, exact mechanism of virus spread or lack thereof through the olfactory system is not well understood.
Here we infect murine olfactory epithelial cultures (containing OSNs and epithelial cells), olfactory LP cultures (containing OECs and fibroblasts) and forebrain (Fb) cultures (containing neurons and astrocytes) with a highly pathogenic strain of Influenza A virus (H5N1/VN1203/2004) and a low pathogenic strain- pandemic (pm) (H1N1/CA/07/2009). We then performed multicycle growth kinetic studies and confocal microscopy investigating influenza infection through the olfactory system and compared this to CNS host cell response.
Viral growth kinetic studies revealed strain specific differences, with robust replication seen for H5N1 compared to pmH1N1. We also observed cell specific differences. While pmH1N1 appeared to replicate successfully within Fb cultures this was restricted within olfactory cultures. On the other hand, both olfactory and forebrain neuron cultures were conducive for H5N1 replication, . Using confocal microscopy, we were able to identify unique viral infection trends. H5N1 infection within olfactory epithelial cultures initial infection was observed mainly within non-neuronal (presumably epithelial cells) cells which appeared to spread to OSNs in later stages of infection (48 – 72 h). On the other hand, H5N1 infections was predominantly observed within OECs in the LP cultures with most infection resolved by 72 h. Furthermore, infection with H5N1 produced change in OEC morphology with increased cell area and tunnelling nanotubes (TNTs) seen between adjacent cells with influenza nucleoprotein within these structures. Similar changes were also observed in astrocytes, indicating potential viral manipulation for cell-cell transmission within glial cells.
Further molecular and microfluidic studies will be performed to investigate these differences in infection.