Conventional thinking is that premature infants are predisposed to more severe and frequent infections, such as influenza virus and respiratory syncytial virus (RSV) infections, due to the immaturity of their immune systems. However, recent data suggest that impaired responses to infection persist beyond infancy (19
), raising questions about whether and how early-life hyperoxia affects the immune system. To date, there have been no clinical or animal studies assessing antiviral CD8+
T cell function in subjects that received neonatal oxygen supplementation. The findings in the current study indicate that the detrimental effects of neonatal oxygen supplementation on host responses to infection are unlikely due to intrinsic defects in the ability to mount an effective CD8+
T cell response to the infection. This is evident from the equivalent numbers of virus-specific CD8+
T cells, T cell functional readouts, and viral clearance kinetics for the two groups in the study. Combined with previously published data showing normal influenza virus-specific serum antibody titers in mice exposed to 100% oxygen as neonates (31
) and with influenza immunization studies of premature infants demonstrating antibody responses comparable to those of full-term controls (11
), our findings support the idea that neonatal hyperoxia does not adversely affect adaptive immunity. In other words, premature infants are not predisposed to more frequent or severe respiratory infections simply due to the immaturity of the immune system.
In the absence of intrinsic deficits in key adaptive responses to infection, it is unclear which mechanisms are responsible for the poorer survival of adult animals that received supplemental oxygen for a few days after birth. One possibility is that early-life exposure to a high oxygen level reprograms innate host responses to infection. This could include lung epithelial and endothelial cells. Indeed, it has long been known that exposure of newborn rodents to high oxygen levels causes alveolar simplification and defects in vascular development, which are attributed in part to loss of vascular endothelial growth factor signaling and circulating endothelial cell precursors (7
). Furthermore, studies of mice suggest that supplemental oxygen disrupts the frequency of type I and type II epithelial cells within the developed lung (47
). These changes in lung cell programming and frequency may contribute to the increased inflammation observed (31
) or adversely influence tissue repair mechanisms.
A surprising finding from the current study was the contribution of memory CD8+
T cells to the development of infection-induced fibrosis in mice exposed to 100% oxygen as neonates. Very few investigations have evaluated how memory CD8+
T cells affect fibrosis development in infectious models. Previous studies have suggested that fibrosis associated with chronic hepatitis is promoted by nonspecific activation of memory CD8+
T cells (28
). For a bleomycin-induced fibrosis model, it was hypothesized that recently activated memory T cells may contribute to lung pathology via interaction with inflammatory dendritic cells (DCs) (1
). The potential role of CD8+
T cells in these models is believed to be as IFN-γ-mediated activators of inflammatory macrophages/dendritic cells, which generate fibrosis-inducing molecules such as tumor necrosis factor alpha (TNF-α), transforming growth factor beta (TGF-β), interleukin-1β (IL-1β), and IL-13 (44
). Whether immune activation of these molecules alone or in the context of severe epithelial injury drives fibrosis remains to be determined. While the issue of antigen-specific activation versus nonspecific activation may explain why pathology was improved in our infectious model compared to the aforementioned studies, it is possible that the altered kinetics of immune activation is responsible for the improved outcome. It is well known that the presence and early recruitment of memory T cells to the lung confer accelerated pathogen clearance and improved survival (16
). In addition to reducing the duration of infection, recent studies are starting to address how the presence of memory T cells affects the nature and magnitude of the innate immune response (24
). The altered kinetics of proinflammatory cytokines may alter the activation state or reduce the recruitment of innate immune cells that contribute to the development of fibrosis in mice exposed to 100% oxygen as neonates.
In addition to reduced lung pathology, the presence of memory T cells also improved the survival of adult mice that had received supplemental oxygen as neonates. While it was expected that having virus-specific memory would improve host resistance, the fact that generating memory T cells restored survival to that observed in controls exposed to room air, even at virus doses that induce 70% lethality, was surprising. This suggests that whatever benefit is conferred by the memory T cells is independent of infection severity. As with the effects on fibrosis, the presence of T cell memory likely influenced the kinetics of the innate immune response and accelerated viral clearance. A similar phenotype has been observed for influenza virus-infected obese mice, where pathology in obese mice was enhanced independent of viral clearance (30
). Despite the ability of obese mice to effectively clear the infection, survival was improved in these mice with the addition of antiviral medication that reduced the duration of infection. These data support the idea that reducing the duration of infection may keep neonates exposed to 100% oxygen from developing persistent, pathological inflammation and may improve survival.
Considering the fragility of infants who require supplemental oxygen and their sensitivity to respiratory infections, they are prime candidates for receiving the seasonal influenza vaccine. The recent emergence of the 2009 pandemic influenza (H1N1) and avian influenza (H5N1) viruses has triggered a renewed interest in elements that confer immune protection across multiple strains of influenza virus. Multiple groups have reported that seasonal influenza vaccination can generate CD8+
T cells that cross-react against pandemic and avian influenza virus antigens (26
). In a mouse model, adoptive transfer of influenza virus H3N2-specific T cells can confer protection against the 2009 H1N1 influenza virus (17
). These studies predict that cross-protective memory T cells generated by the seasonal influenza vaccine may reduce the susceptibility of infants treated with supplemental oxygen and/or diagnosed with BPD to related influenza virus infections. Considering the relatively small number of influenza virus-specific cells that were adoptively transferred into the mice in our study—equivalent to the number of cells found in the lungs of i.p. challenged mice ()—protection may not require a large expansion of memory T cells by the vaccine. If this proves true for humans, it will mean that many current vaccine formulations might be adequate to prime the immune response of infants requiring supplemental oxygen at birth, better protecting them from respiratory infection later in life. Since there is no current RSV vaccine and many infants become infected with viruses before vaccination, the use of antiviral drugs may be beneficial to shorten the duration of infection and reduce the risk of developing chronic inflammation. Further studies with children who received the seasonal influenza vaccine are needed to ascertain whether the incidences of influenza virus infection and/or hospitalization are different between children born at term and premature infants requiring supplemental oxygenation.
In conclusion, the current findings demonstrate for the first time that neonatal hyperoxia confers susceptibility to various subtypes of influenza virus but does not impair the CD8+ T cell response. Conversely, the presence of memory CD8+ T cells protects oxygen-exposed neonates from enhanced pathology and mortality associated with infection. These results suggest that premature infants who require supplemental oxygen therapy would benefit from vaccination strategies that promote T cell immunity against respiratory viral infections as well as from early antiviral treatments that reduce the duration of infection. A better understanding of how oxygen supplementation of neonates contributes to postinfection pathology will be helpful in developing appropriate vaccines and/or drugs to protect this fragile population.