Figure 2.

ICAM-1/2^-/- mice infected with PR8 exhibit a similar disease progression and viral clearance compared to WT mice. (A) Pulmonary imaging of infected mice using three-dimensional (3D) light sheet fluorescence microscopy (LSM). WT mice were I.T. instilled with 3x10³ PFU of PR8-mCherry influenza virus. Four days post infection (dpi), mice were sacrificed, lungs were cleared and individual lobes were imaged using an UltraMicroscope II (LaVision BioTec). The distribution of the PR8-mCherry influenza virus was mainly detected in the bronchi and bronchioles. Scale bar represents 500 μm. n=2. (B) Kaplan-Meier survival curves of WT (left panel) and ICAM-1/2^-/- (right panel) mice infected I.T. or I.N. with the indicated doses of PR8 influenza (15 PFU n=5, 155 PFU n=4, 3x10³ PFU n=3 (I.T.), 30 PFU n=4 (I.N.)) were assessed in at least two independent experiments. (C) Experimental design and timeline of primary and secondary infections. (D) Body weight change in either WT or ICAM-1/2^-/- mice initially infected I.N. with 30 PFU (n=13 for each group, top panel) or I.T. with 15 PFU (n=8 for WT and n=6 for ICAM-1/2^-/-, bottom panel) (1ry), in at least two independent experiments. For a secondary challenge, both groups were challenged I.T. with a lethal dose (3x10³ PFU) of PR8 influenza on day 40 (2ry). (E) Experimental design and timeline of viral titers in lungs. Mice were infected I.N. with 30 PFU of PR8 influenza, lungs were harvested on day 4 and 12 post infection and the virus lung titer was determined for both mice groups using RT-qPCR. I.N. PBS administration was performed in control experiments. Red dotted line represents the detection limit. (D) Statistical analysis was performed using a one-way ANOVA test comparing % body weight loss between groups. (E) Statistical significance was determined by two-tailed, unpaired Student’s t tests. ****P < 0.0001, ns, not significant. The error bars indicate the SEM of each measurement.