Discussion
In this study, we measured temporal changes of PC20 in pediatric asthma patients with A(H1N1)pdm09 infection. PC20 was lowest at 1 month after discharge and significantly increased at 3 months after discharge. Furthermore, we comparatively evaluated AHR in a mouse model of asthma with either A(H1N1)pdm09 infection or seasonal H1N1 infection to investigate the pathophysiology of A(H1N1)pdm09-infected asthmatic children. Enhanced AHR was observed in asthmatic mice with A(H1N1)pdm09 infection, which peaked at 7 days post-infection and subsequently diminished at 10 days post-infection. These results were similar to those of asthmatic children. Histopathological analysis showed that the onset of lung inflammation in asthmatic mice with A(H1N1)pdm09 infection occurred earlier and was more prominent compared to that in mice with seasonal H1N1 infection; these effects peaked 7 days post-infection and diminished by 10 days post-infection, which was consistent with the observed changes in AHR. These data suggest that A(H1N1)pdm09 induces enhanced AHR complication as a severe phenotype of pneumonia in mice with asthma. The desaturation and enhanced AHR observed in asthmatic patients must be induced by the pulmonary inflammation during A(H1N1)pdm09 infection, but not seasonal H1N1.
The severity of AHR reflects the inflammatory state of the airways19. Several studies have reported that AHR can be enhanced by inflammatory and Th2 cytokines, such as tumour necrosis factor (TNF)-α, interleukin (IL)-6, and IL-1320-22. TNF-α secreted from airway macrophages or airway epithelial cells after respiratory virus infection increases levels of adhesion molecules, such as intercellular adhesion molecule-1 on epithelial cells, thereby inducing the recruitment of eosinophils and contributing to epithelial damage and AHR23-27. TNF-α is associated with wheezing in human infants20. IL-6 is secreted from epithelial cells in respiratory virus infection and induces airway inflammation and bronchospasms in patients with asthma and upper respiratory tract infections21, 24. Our previous study showed that IL-6 and TNF-α levels in the bronchoalveolar lavage fluid of A(H1N1)pdm09-infected mice were significantly higher than those in seasonal H1N1-infected mice within 3 days after infection14. Therefore, A(H1N1)pdm09 infection may enhance AHR by inducing the production of high levels of inflammatory cytokines during lung inflammation in asthmatic mice, which may also occur in human cases.
As explained above, AHR in A(H1N1)pdm09-infected children was alleviated by 3 months after discharge compared to findings at 1 month after discharge. This finding is supported by our data in A(H1N1)pdm09-infected mice showing that the enhanced AHR at 3 and 7 days post-infection decreased to the same level as in control mice at 10 days post-infection. Bozanich et al28 reported that increased AHR observed in seasonal H3N1 influenza-infected non-asthmatic wild-type mice at 4 days post-infection returned to control levels at 20 days post-infection, which is consistent with our findings. Together, these data indicate that it is pivotal to treat patients with severe asthma exacerbation in the acute phase of post-A(H1N1)pdm09 infection. Established treatments for rescuing acute severe asthma exacerbation complicated with severe pneumonia resulting from A(H1N1)pdm09 infection have not yet been developed. We are currently investigating approaches for treating acute severe asthma exacerbation occurring with A(H1N1)pdm09 infection.
There were some limitations to this study. First, we did not evaluate AHR, inflammatory or Th2 cytokines, or virus titres in the bronchoalveolar lavage fluid at the same time. Lung tissues were collected and used for pathological analysis after AHR evaluation, as we previously reported the cytokine profiles in the bronchoalveolar lavage fluid of A(H1N1)pdm09 mice13. Second, we could not evaluate AHRs of the pediatric participants before or during A(H1N1)pdm09 infection for ethical reasons to verify whether or not enhanced AHR during the infection were alleviated in the post-infection phase. Instead, we measured AHR at 1 and 3 months after discharge.
In conclusion, AHR was significantly enhanced in both pediatric asthma patients and asthmatic mice in acute phase of A(H1N1)pdm09 infection. Furthermore, A(H1N1)pdm09-infected asthma model mice showed more severe pulmonary inflammation in the acute phase post-infection, compared to that occurring in asthmatic mice with seasonal influenza infection. Enhanced AHR subsequently returned to normal levels with the amelioration of lung inflammation, suggesting that appropriate treatment during the acute phase after A(H1N1)pdm09 infection is essential for avoiding severe respiratory conditions.