Regulatory T Cell Response During Influenza Infection and Vaccination In The Ferret
Regulatory T cells (Tregs) suppress effector immune responses and have been implicated in promoting chronic viral infections. Their role during influenza infection and vaccination, however, is still unclear. Influenza is a major public health concern, claiming over 49,000 lives annually in the U.S. alone. Vaccination is the best approach for preventing disease but frequent mutations of immunogenic epitopes requires a new vaccine to be formulated and administered annually. This poses a challenge for vaccine manufacturing and may strain patient compliance. A universal influenza vaccine, which targets the highly conserved extracellular domain of the influenza matrix protein 2 (M2e), may circumvent this problem by generating cross-protective immunity. In this study, we tested the efficacy of the M2e universal vaccine in the ferret, and determined whether vaccination induces a Treg response after influenza infection. We found that vaccination promotes the development of M2e specific IgM and IgG antibodies after boosting. Upon challenge with A/Memphis H1N1, vaccinated ferrets exhibited a lower body temperature and reduced virus titer compared to non-vaccinated animals. Together these findings suggest that the M2e vaccine protects ferrets against influenza infection. In order to determine whether Tregs increase after vaccination in ferrets, we had to first clone and characterize genes involved with Treg phenotype and function including CD25, Foxp3, and IL-10. The reciprocal nature between Tregs and Th17 cells and their involvement during influenza infection prompted us to also clone ferret IL-17F. Using these sequences, we designed a qRT-PCR array to measure the expression of Foxp3, IL-10, and IL-17F in ferret tissue. We also identified cross-reactive antibodies against ferret CD8, CD25, and Foxp3 for use in FACS, western blot, and ICC. Using these tools, we found that vaccination significantly increased the expression of Foxp3 in the spleen. An increased percentage of Foxp3+ lymphocytes was detected in both the PBMCs and splenocytes of immunized animals. In contrast, IL-10 and IL-17F expression decreased significantly in both immunized and non-immunized ferrets compared with naïve animals. These studies suggest that the M2e influenza vaccine induces a regulatory T cell response in ferrets and protects against influenza infection.