Impact of Human Lung CD8+ Tissue Resident Memory T Cell and Epithelial Crosstalk in Antiviral Immunity Restricted; Files Only
Mattingly, Cameron (Fall 2025)
Abstract
Tissue-resident memory CD8+ T cells (TRM) are essential components of barrier immunity, where their rapid response capabilities can limit viral replication at mucosal surfaces such as the respiratory tract. While murine models have demonstrated the protective role of respiratory TRM in reducing viral burden and immunopathology following heterosubtypic influenza infection, the effector functions and protective mechanisms of human lung TRM remain poorly defined.
To address this gap, we investigated the functionality of antigen-specific CD8+ T cells from non-transplantable, healthy human lungs to determine how virus-specific lung CD8+ TRM impact the local cellular environment following antigen recognition. Upon antigen encounter, lung TRM initiated a robust IFNg-driven response that activated both local innate immune cells and the airway epithelium. Upon further investigation into the epithelial cell response, we identified that basal epithelial cells exhibited the strongest response to IFNg activation, which correlated with elevated baseline expression of IFNgR. Using Air-Liquid Interface (ALI) cultures, we modeled the effects of IFNg-mediated epithelial cell activation on influenza replication and discovered that rapid IFNg signaling was necessary and sufficient to induce antiviral programs in stratified epithelium and protect against infection. Taken together our findings describe how TRM induce a crucial early resistance program in lung epithelium to restrict respiratory virus propagation.
To better understand the mechanisms underlying this rapid cytokine response by TRM , we used a mouse model to compare phenotypically distinct TRM subsets throughout respiratory tract. In exploring the functional differences among phenotypically distinct CD8+ TRM subsets—CD69 single-positive (SP) and CD69+CD103+ double-positive (DP), we found that DP TRM were enriched in the upper respiratory tract and exhibited intrinsically superior effector capacity, characterized by increased speed and production of IFNg production. Mechanistically, this heightened responsiveness was not driven by transcriptional activity or mRNA stability, but instead by greater stores of pre-formed IFNG mRNA, enabling rapid cytokine output upon antigen stimulation.
Together, this dissertation defines how human lung CD8+ TRM orchestrate early antiviral immunity through rapid cytokine production and epithelial activation. By extending mechanisms previously described in murine models into the human lung and further exploring subset-specific TRM functional heterogeneity in mice, this work underscores the importance of TRM derived IFNg and –epithelial crosstalk in early viral control. These findings support ongoing efforts to develop vaccines that generate lung CD8+ TRM and motivate continued investigation into the mechanisms of T cell–mediated protection in both human and experimental systems.
Table of Contents
Table of Contents
CHAPTER I: INTRODUCTION ........................................................................................................................ 1
INFLUENZA ......................................................................................................................................................... 1
Global Health Burden .................................................................................................................................... 1 Virology.......................................................................................................................................................... 2
Vaccines ......................................................................................................................................................... 4
AIRWAY EPITHELIUM........................................................................................................................................ 5
IMMUNE RESPONSE TO INFLUENZA .................................................................................................................. 8
Innate Immune Response ............................................................................................................................... 8
Initiation of adaptive response....................................................................................................................... 9
T Cell Response............................................................................................................................................ 13
CD8 + T Cell Memory................................................................................................................................... 14
CD8+ TISSUE RESIDENT MEMORY T CELLS ................................................................................................... 15
CD8+ TRM Surface Marker Expression ........................................................................................................ 15
Establishment and Maintenance of CD8+ TRM ............................................................................................. 17
Function of CD8+ TRM .................................................................................................................................. 19
Duration of CD8+ TRM.................................................................................................................................. 20
TECHNIQUES FOR STUDYING CD8+ TRM IN THE RESPIRATORY TRACT ......................................................... 21
Methods in Mice........................................................................................................................................... 21
Methods in Humans ..................................................................................................................................... 22
CHAPTER II: HUMAN LUNG CD8+ TRM DERIVED IFNg ORCHESTRATES SUBSET-SPECIFIC ANTIVIRAL PROGRAMMING IN AIRWAY EPITHELIAL CELLS .............................. 24
ABSTRACT ........................................................................................................................................................ 24
INTRODUCTION ................................................................................................................................................ 24
MATERIALS & METHODS ................................................................................................................................ 27
RESULTS ........................................................................................................................................................... 34
DISCUSSION ...................................................................................................................................................... 45
Limitations ................................................................................................................................................... 50
MAIN FIGURES ................................................................................................................................................. 52
Figure 1........................................................................................................................................................ 52
Figure 2........................................................................................................................................................ 54
Figure 3........................................................................................................................................................ 56
Figure 4........................................................................................................................................................ 59
Figure 5........................................................................................................................................................ 61
Figure 6........................................................................................................................................................ 63
Figure 7........................................................................................................................................................ 66
SUPPLEMENTAL INFORMATION ...................................................................................................................... 68
Supplemental Table 1................................................................................................................................... 68
Supplemental Table 2................................................................................................................................... 69
Supplemental Figure 1 ................................................................................................................................. 70
Supplemental Figure 2 ................................................................................................................................. 72
Supplemental Figure 3 ................................................................................................................................. 73
Supplemental Figure 4 ................................................................................................................................. 75
Supplemental Figure 5 ................................................................................................................................. 77
Supplemental Figure 6 ................................................................................................................................. 78
Supplemental Figure 7 ................................................................................................................................. 80
CHAPTER III: FUNCTIONAL HETEROGENEITY OF CD8+ TISSUE RESIDENT MEMORY T CELLS THROUGHOUT THE RESPIRATORY TRACT .......................................................... 81
ABSTRACT ........................................................................................................................................................ 81
INTRODUCTION ................................................................................................................................................ 82
MATERIALS & METHODS ................................................................................................................................ 84
RESULTS ........................................................................................................................................................... 86
DISCUSSION ...................................................................................................................................................... 90
MAIN FIGURES ................................................................................................................................................. 94
Figure 1........................................................................................................................................................ 94
Figure 2........................................................................................................................................................ 96
Figure 3........................................................................................................................................................ 98
Figure 4........................................................................................................................................................ 99
CHAPTER IV: DISCUSSION......................................................................................................................... 101 REFERENCES.................................................................................................................................................. 112
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