Mind Your Memory: Modulation of Memory T Cell Generation During Influenza Infection

We have all heard of people complaining from their flu-like symptoms, but few people actually comprehend what influenza is. There are actually two main kinds of influenza viruses, Type A and B, that are responsible for the seasonal flu epidemics that happen each year. Viruses themselves are infectious agents that need host cells in order to reproduce. During a viral infection, cells called CD8+ T cells recognize antigens (that were originally part of a pathogen). This causes them to multiply into large numbers of CD8+ T cells that are specific for that antigen. These CD8+ T cells can help to eradicate the virus by secreting molecules that can kill infected target cells. Eventually these numbers of CD8+ T cells decrease and a small population of antigen-specific memory T cells can be left behind. These memory cells are a great asset to the immune system because if the same pathogen attacks the body for a second time and the same antigens are presented, the immune system can activate these cells to mount a stronger and faster response than what was observed upon the pathogen’s initial assault. The generation of memory T cells is also important for the generation of vaccines designed to prevent serious infection by a pathogen. Vaccines often contained a killed, inactivated, or weakened form of a pathogen so that a recipient’s body can recognize the pathogen’s antigens as foreign, combat them, and then generate memory CD8+ T cells to “remember” the antigens for future encounters so that they can be fought off more effectively.

It is important to note that vaccines are especially important for protection against the influenza virus because this virus mutates and changes over time. Initial exposure to an influenza virus may not confer protection upon a second exposure of this influenza virus because the second exposure may involve a different strain of influenza for which no CD8+ memory cells had been generated. This is also why it has been impossible to create a vaccine that grants a person lifelong immunity. Instead, annual vaccine programs are undertaken to generate memory CD8+ T cells against the strains of the virus that are most likely to cause illness.

The factors affecting the generation of memory CD8+ T cells, however, are not yet fully understood. A protein by the name of osteopontin seems to be involved in this process. Osteopontin has been involved in many different processes ranging from cancer (Chakraborty and Kundu 2008) to bone remodelling (Denhardt and Noda 1998). Recent research by Morimoto and colleagues (et al., 2011) has specifically focused on how osteopontin (Opn) affects the generation of memory CD8+ T cells.

To study the role of Opn, the researchers used a mouse model of the disease in which certain groups of mice were infected with an influenza virus (Type A). The first question that they addressed was whether the influenza virus infection led to the expression of Opn in the lungs. To do this, they used a technique called immunohistochemistry. Immunohistochemistry uses antibodies to detect specific proteins in tissue, and an antibody-antigen interaction can be visualized through color-producing reactions. This technique revealed that levels of Opn were significantly increased in the lungs (which are the main site of influenza infection) in infected mice in comparison to the levels observed prior to infection.

Given that influenza infections have the potential to cause serious disease and damage in the lungs, which is associated with the presence of inflammatory cells, the researchers investigated whether a deficiency in Opn could affect the magnitude of lung inflammation. Inflammation is a local response at a site of infection caused by the influx of innate leukocytes, which are cells that attempt to combat infection through general recognition mechanisms. Previous research had shown that Opn can attract inflammatory cells to itself (Morimoto et al., 2004). In this study, the researchers used both wild-type (WT) Opn mice that express normal amounts of Opn and Opn knock-out (KO) mice that had been genetically engineered so that they could not express Opn. These mice were given a dose of the influenza virus which was not fatal but would lead to a severe illness. Findings using a technique called flow cytometry (used for counting and categorizing cells) revealed that the number of inflammatory cells in the lung fluid of Opn WT and KO mice were similar. Therefore, Opn actually did not affect the severity of lung inflammation even though its expression was induced in the lungs after influenza infection. The researchers also showed that Opn deficiency did not affect the generation of virus-specific CD8+ T cells, which are involved in killing virus-infected cells.

The researchers also investigated whether Opn could affect the generation of CD8+ memory precursor T cells. After a viral or bacterial infection, virus-specific CD8+ T cells can expand their numbers and turn into either memory precursor effector cells or terminal effector CD8+ T cells. The former group of precursor cells then has the ability to become long-lived memory cells, but the latter group of terminal cells does not. Opn deficiency in Opn KO mice favored the generation of these memory precursor effector CD8+ T cells (rather than terminal cells) in their spleens, when compared to the Opn wild-type mice. This was observed at day 9 post-infection, at the peak of the influenza response. At 60 days after the initial infection, the Opn deficient mice also had higher numbers of memory CD8+ T cells in their spleens when compared to normal mice, indicating the prolonged lifespan of these memory cells.

The next aim of this study was to identify which cells were producing the Opn. Cells were collected from Opn wild-type mice spleens at 9 days post infection. Surprisingly the CD8+ effector T cells did not produce the Opn even after they were stimulated by their corresponding antigen. Instead, dendritic cells (DCs) isolated from the spleen at 7 days post infection produced the Opn. Dendritic cells are immune cells who main function is to process antigens (such as from an influenza virus) and present it on their surfaces to other cells of the immune system. Thus, Opn made by DCs seems to regulate the generation of memory precursor effector CD8+ T cells.

Morimoto and his colleagues then wanted to know if the CD8+ T cells made in the Opn deficient mice were still functional and had the ability to lyse cells (and thus destroy them). Target cells which presented the influenza virus were labelled with a fluorescent dye and injected into either Opn wild-type or Opn knockout mice. Opn wild-type and Opn deficient mice that had never been exposed to influenza before did not eliminate these cells. On the other hand, Opn wild-type and Opn KO mice that were primed by a prior exposure to influenza were able to completely eliminate these cells. During a secondary exposure to a higher dose of influenza, both Opn wild-type and KO mice that had been primed were also able to clear the virus. The Opn KO mice, however, had lower level of virus circulating in their lungs during the early stage of the memory cell response as a result of their increased numbers of memory CD8+ T cells. This demonstrated that the memory CD8+ T cells were not impaired by an Opn deficiency and could still be activated upon a second assault by a pathogen.

This research revealed how a deficiency of osteopontin, which can be produced by dendritic cells, leads to a persistent and increased number of memory CD8+ T cells in mice after an influenza virus infection. Nevertheless, this study could be extended further. Given that dendritic cells and not CD8+ T cells appear to be produce osteopontin, a relevant future direction for these researchers is to investigate the mechanisms that regulate the production of osteopontin. This knowledge might lead them to identify inhibitors of osteopontin production in order to increase the generation of memory CD8+ T cells. Also, as the researchers themselves mentioned, it would be interesting to identify which other cell types could be contributing to the production of osteopontin.

These findings presented in this study are of significance because the establishment of memory CD8+ T cells is important for combating a subsequent assault by the same pathogen and for the creation of effective vaccines. Once more data is gathered about the factors that regulate osteopontin production, perhaps a manipulation of these factors will be incorporated into vaccine designs. After all, disease prevention is a key aspect of public health; it is far better to prevent a disease rather than to treat it.

Primary Reference:

Morimoto, J., Sato, K., Nakayama, Y., Kimura, C., Kajino, K., Matsui, Y., . . . Uede, T. (2011). Osteopontin modulates the generation of memory CD8+ T cells during influenza virus infection. Journal of Immunology (Baltimore, Md.: 1950), 187(11), 5671-5683. doi:10.4049/jimmunol.1101825

Additional References:

Chakraborty, G., S. Jain, and G. C. Kundu. 2008. Osteopontin promotes vascular endothelial growth factor-dependent breast tumor growth and angiogenesis via autocrine and paracrine mechanisms. Cancer Res. 68: 152–161.

Denhardt, D. T., and M. Noda. 1998. Osteopontin expression and function: role in bone remodeling. J. Cell. Biochem. 72(Suppl.): 92–102.

Morimoto, J., M. Inobe, C. Kimura, S. Kon, H. Diao, M. Aoki, T. Miyazaki, D. T. Denhardt, S. Rittling, and T. Uede. 2004. Osteopontin affects the persistence of b-glucan-induced hepatic granuloma formation and tissue injury through two distinct mechanisms. Int. Immunol. 16: 477–488.