This study found that when challenged intranasally with the same dose of 2009 H1N1 virus, older mice are less likely to die than younger mice. This fact that age is protective for pandemic H1N1 infection parallels the human data. Importantly, in this mouse model of pandemic H1N1 infection, all mice had not been exposed to any influenza viruses before the experiment; therefore, pre-existing anti-H1N1 or cross-reactive anti-influenza immunity is not a mechanism to be considered for age-related protection. On the other hand, immune responses to pathogen infection differ by age [20–22], and age is a factor to be considered in vaccination and therapy strategies.
Mechanistic examination by immunohistochemistry and pathology revealed that young mice are more sensitive to 2009 influenza A H1N1 virus infection than older mice, at least partially due to the greater sensitivity of their bronchi epithelial cells. In the study by Munster et al.  using an adult ferret model, the 2009 H1N1 virus was found to replicate efficiently in the lower respiratory tract, and viral antigens were found in epithelial cells of bronchioles and terminal bronchioles, alveolar macrophages, and also type I and type II alveolar epithelial cells. In contrast, seasonal influenza virus was only found in the epithelial cells of the upper respiratory tract, and this difference in upper versus lower respiratory tract infection pattern was proposed as why the novel H1N1 virus will cause a larger epidemic than the seasonal influenza virus. This result is also in agreement with studies by Itho et al. and Belser et al. [18, 23] that H1N1 viral antigens were detected in bronchiolar epithelia, desquamated cells in bronchiolar lumen, alveolar epithelium and histiocytes in thickening alveolus interstitial. In addition, our results from the murine model largely agree with the study in ferrets [16, 17] in that we also found most virus infection in the lower respiratory tract and of the cell types infected, only marked difference was that mouse alveolar epithelial cells are not sensitive to H1N1 virus infection. This difference in cell type sensitivity might be due to animal species difference and might account for the relative insensitivity of mice as a species to influenza infection . We suspect similar age-related sensitivity to H1N1 virus infection should also exist in ferrets and parallel study in ferrets should offer interesting insights to the age-distribution of the pandemic.
Related to immunohistochemical data, the virus lung titer assay affirmed more efficient virus proliferation in the younger mice in the first two to three days after infection. This higher virus titer might be explained by the more efficient infection at inoculation, as there will be more lung cells producing viruses, and it may or may not be due to more viruses made per mouse cell.
The iBALT, a tissue form that is not usually present in healthy animals [25, 26] has been demonstrated to form upon certain infections and its formation is a protective immune response in that it clears influenza viral infection with a better organized immune response than that mediated by primary lymphoid organs which might over-react and result in more severe lung pathology and even death [27, 28]. The role of innate response to influenza infection is complicated and can both help and inhibit viral clearance [29, 30]. An effective level is necessary for infection clearance [31–33] but overreaction has been blamed to be causing deaths in young people in the H5N1 epidemic [34, 35]. It has been reported that young mice have lower phagocytic functions and secret less TNF-alpha than adult mice [36, 37], suggesting young mice has lower immunity than adult mice and has less ability to organize an efficient response for clearing influenza virus. Our data suggest that the moderate inflammatory response in lung of adult and old mice, organized with the help to iBALT, might provided an efficient level of protective immune response to the infection at earlier time after virus infection, while the young mice lacked iBALT formation and had insufficient level of inflammatory response for viral clearance and control.
H1N1 virus induces apoptosis of infected cells, which in a degree is a resistance mechanism of the body to viral replication . In our study, at 3 days after virus infection, shedding of injured bronchial epithelial cells was observed, and some epithelium was seen in regeneration and repair stage. The regeneration of epithelium is slightly earlier in adult mice than in young mice (Figure 3D-F), suggesting adult mice have higher capacity in epithelium repair than young mice. The much more severe pathology observed in the lung of young mice as compared to adult and old mice appears to be explained by young mice's higher sensibility to infection, higher levels of virus replication and subsequent massive apoptosis and damage of their epithelium cells.
On the other hand, at the 10LD50 dose of H1N1 infection, we noted that the adult and older mice had much more severe damages in their spleen than the young mice, which might contribute to their eventual death. This data indicate that even the end result might be the same (although the older are more likely to survive a lower dose that's lethal to the young), young and older mice die from different pathological causes. If the same is true in humans, this serves a basis for adopting differential intervention strategies for different ages.
The murine influenza model has been widely used for studying influenza pathogenesis, viral therapy and vaccines  and the model has been used to study this 2009 H1N1 virus. Pathological findings in mice mimic that in the clinic . In addition, mice are the animal of choice for studying immune responses at different ages and development stages [41–43]. The current study found that 1) young mice are more susceptible to the 2009 pandemic influenza A H1N1 virus infection and are more likely to die from the infection than adult and old mice, which parallels the pandemic epidemiology in humans; 2) this age-related sensitivity to H1N1 infection might be explained by young mice's higher sensitivity of its bronchial and bronchiole epithelial cells to be infected, a weaker innate immune response of the young mice to control and clear infection, and therefore young mice are more likely to die from infection-caused massive apoptosis and damage of mouse bronchial epithelial cells and the lung structure and function. This is a first age-comparison study in animals for 2009 H1N1 infection, and by using an influenza-naive and immunocompetent mouse model it studied the age factor independently of two major influential factors of the pandemic. The study highlights the importance of age factor in the 2009 H1N1 virus infection and also suggests that animal model data with unspecified age or one age should be interpreted with caution. Despite the imperfections of the animal model, these observations might provide some mechanistic explanations for the age distribution in the 2009 human influenza pandemic.