Fig. 4: Influenza-infected neonates have greater oxidative stress imbalance in response to IFNβ.

Three-day-old neonatal and 8-week-old adult WT mice were intranasally infected with PR8 influenza A virus. Age-matched uninfected mice were used as controls. Mice were harvested 2-days post-infection and Type II epithelial cells were isolated. The TIIEC were treated with an antioxidant N-acetylcysteine (NAC), IFNβ, or tert-Butyl hydroperoxide (TBHP), or media (Untreated) ex vivo for an hour. NAC treated wells were used as background staining controls. The CellInsight CX7 high content screening platform was used to quantify CellROX intensity at the individual cell level. a Representative images at 10X magnification from IFNβ-treated uninfected and infected neonates and IFNβ-treated infected adults are depicted. The TIIEC were co-stained with DAPI (blue), WGA (red) and CellROX (green). b The percentage of CellROX positive cells relative to total cells in each treatment group is indicated. c Fold change of CellROX intensity relative to age-matched uninfected animals. n = 9 in neonates, n = 3 in adults, 3 independent experiments. To confirm the role of IFNβ in the neonatal program of oxidative stress imbalance during influenza virus infection, IFNαβR^−/− and WT neonates were infected as above and TIIEC were harvested 2-days post-infection. d Relative fluorescence units of average CellROX intensity by flow cytometry in IFNβ-treated WT (white bar) and IFNαβR^−/− neonates (black bar) is shown. Statistical differences between groups were assessed using Student’s T test was used when comparing 2 groups, where denoted *p < 0.05, **p < 0.01.