Title: Lung localized protective responses to heterosubtypic influenza challenge
Description: Influenza A virus (IAV) is one of the most ubiquitous respiratory viruses in the world, causing significant disease burden in the United States and abroad. Current vaccination strategies that target the generation of humoral immunity offer limited heterosubtypic protection; T cells offer cross-strain protection and the promise of universal immunity against IAV. Local tissue immunity plays a key role in pathogen clearance and tissue protection, particularly in the form of tissue resident memory T cells (TRM), which are a non-circulating memory T cell subset that have been shown in a variety of tissue sites to be superior mediators of protection compared to circulating memory T cells. At the same time, T cell immunity has been associated with inflammatory processes that may also lead to lung immunopathology. How lung tissue localized T cell immunity mediates its protection during a recall response to IAV challenge is not well understood. Using the lymphocyte sequestering drug FTY720, we show that primary infection with H3N2 IAV strain X31 provides tissue localized heterosubtypic immunity independently of humoral immunity against an H1N1 PR8 IAV strain.

Within the lung resident niche, the recall response drives faster CD4+ and CD8+ T cell expansion compared to a primary infection. This rapid T cell expansion resulted from in situ TRM proliferation that was augmented by the migration of peripheral T cells. By tracking a naïve T cell population specific for the IAV strain used in secondary challenge, we demonstrate that influenza-specific T cells, including those specific for newly introduced antigens, migrate to the lung niche from the local mediastinal lymph node (medLN) where both CD4+ and CD8+ T cells experience enhanced priming and proliferation. We further show that primary infection fortifies the medLN with persistently increased numbers of T cells as well as both CD103+ and CD103- conventional dendritic cells (cDCs) that are transcriptionally similar to cDCs in an infection naïve mouse. By depleting Zbtb46+ cDCs, we determine that cDC fortification is a crucial mechanism for enhanced T cell priming and expansion in the medLN during a recall response.

We also found that lung localized CD4+ T cell responses exhibit significant immunomodulatory function. Polyclonal lung CD4+ TRM generated by influenza infection as well as lung OT-II TRM exhibit increased production of antiviral inflammatory cytokines in addition to enhanced IL-10 family cytokine production compared to splenic CD4+ effector memory T cells (TEM). During a heterosubtypic challenge, we further observed that lung niche non-TRM CD4+ T cells produce significantly more in situ IL-10 compared to a primary infection, which modulated airway IFN-ɣ and TNF-α production without any depreciation in viral clearance. Immunomodulatory characteristics of a recall response was reflected in lung tissue-wide transcriptional downregulation of innate responses such as type I IFN responses compared to a primary infection. This work demonstrates the dual antiviral and immunomodulatory protective role of enhanced tissue-localized T cell responses during the recall response to IAV challenge.