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Pulmonary inflammatory diseases are among the top leading causes of death globally. The majority treatments for these diseases are aimed counteracting the symptoms rather than treating the underlying cause and many are not very effective. This is partly due to the poorly understood nature of the mechanisms underlying these diseases and lack of disease modifying strategies. The balance of inflammatory and suppressive cytokine is critical in controlling inflammatory responses, and the pro- and anti-inflammatory cytokines IL17A and IL10 has been implicated in numerous pulmonary inflammatory diseases. Hypersensitivity pneumonitis (HP) is a collection of lung diseases caused by repeated inhalation of environmental antigens leading to inflammation, tissue scarring, and some loss of lung function. Moreover, treatment options for HP are limited. Farmer’s lung disease (FLD), a type of HP caused by Saccharopolyspora rectivirgula (SR) was originally described as a TH1-mediated disease, but has been shown to depend on IL17A, a cytokine secreted predominantly by TH17 cells. The Tec kinase Itk plays a critical role in T cell activation and has been shown to be required for the production of IL17A. Furthermore, Foxp3+ T regulatory (Tregs) cells, suppressive producers of IL10, are negatively regulated by Itk. We have previously shown that Itk-/- mice do not develop allergic airway disease coupled with reduced production of IL17A in the lungs. The objective of this work is to investigate the source of IL17A and the role of the Tec kinase Itk in regulating development of IL17A dependent airway immune response in a murine model of FLD. We used exposure to SR of WT and Itk-/ -IL17A-GFP/Foxp3-RFP reporter mice, to examine the time course, source of IL17A, and requirement for Itk, during the development of lung inflammation, and how microbial diversity effects disease outcome; additionally, we explored the use of PEG-maleimide (PEG-MAL) hydrogels crosslinked with enzymatically degradable peptides as in situ crosslinkable depots that has the potential to manipulate inflammatory immune cells in vivo, for its potential to reduce inflammation in this model of HP. Such immunomodulatory approaches have been shown in cancer immunotherapy, but to date not in airway inflammation. We found that surprisingly, exposure to SR drives robust IL17A response in the lung initially by γδ T cells and then by CD4+ αβ T cells even in the absence of Itk, accompanied by the development of pulmonary inflammation. We also found that neutrophils were not a predominant source of IL17A as has been reported. SR also induced Itk independent development of a population of Foxp3+ Tregs cells that produce IL17A or IL10, with IL10 dominating early in the disease. RNA-sequencing and ATAC-sequencing was performed to determine the transcriptome and chromatin accessible status of SR-induced TH17 cells and T regulatory cells. Transcriptomic analysis of Itk independent TH17 cells induced by SR suggest that SR-induced inflammatory signals may override signals downstream of Itk to rescue TH17 responses. In addition, we explored the role of microbial diversity in the development of lung inflammation using SR induced HP or house dust mice (HDM) induced allergic airway disease (AAD). The microbiome plays an important role in the development of allergies and can modulate immune responses to diseases. We treated mice with antibiotic to reduce microbial diversity, or co-housing with pet store mice to increase microbial diversity, to investigate how varied microbial exposure affects inflammatory T cell responses in FLD or AAD. We found that reducing the microbiota decreased TH17 responses in FLD, while increasing the diversity of microbes led to a reduction in the inflammation in FLD and an increase in AAD. These studies suggest that TCR signaling through Itk differentially regulates the development of inflammatory TH17 cells and suppressive Tregs, dependent on the type of inflammation. Understanding how Itk modulates the development of TH17/Treg cytokine responses will allow us to better understand the precise role of Itk in the regulation of cytokines involved in in airway inflammation.

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260 pages


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Union Local


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August, Avery

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Singh, Ankur
Tait Wojno, Elia

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Biomedical and Biological Sciences

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Ph. D., Biomedical and Biological Sciences

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Doctor of Philosophy

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Government Document




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Attribution-NonCommercial-NoDerivatives 4.0 International


dissertation or thesis

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