Friday, September 22, 2023
9/22/2023
Eosinophilia is a characteristic feature of allergic airway inflammation (AAI) including asthma. Eosinophils (Eos) are one of the most pro-inflammatory granulocytes that contribute to pathogenesis and exacerbation of AAI. There is also increasing evidence suggesting that defects in the immune response and barrier properties of airway epithelial cells (AEC), which serve as the first line of defense against allergens, contributes to the development/exacerbation of AAI including asthma. Allergic asthma is a heterogeneous disease with multiple phenotypes that might reflect distinct types of inflammation; thus identifying and understanding cellular mechanisms that drive the inflammation underlying these phenotypes is essential to the development of improved/tailored treatments. The objective of this proposal is to understand the mechanism by which Tropomyosin receptor kinase (Trk) A, a member of the Trk family of receptor tyrosine kinases and the high- affinity receptor for nerve growth factor (NGF), regulates AAI. Although growing evidence suggests that signaling via TrkA is involved in airway inflammatory diseases, due to the lack of specific methods to interrogate the NGF/TrkA signaling pathway (TrkA gene knock-down in mice is lethal and TrkA inhibitors have limited drug specificity), the mechanism by which TrkA regulates allergic asthma remains poorly understood. We show for the first time that TrkA in Eos and AEC can be activated by ligands other than NGF (i.e., CCL11 [eotaxin-1] in Eos and the fungal allergen Alternaria alternata in AEC) and that TrkA can regulate important cellular functions (migration, tight junction formation) in these cells. In addition, using a genetic-chemical approach to specifically control the TrkA kinase activity in vitro and in vivo (TrkA knock-in [TrkA-KI] mice), we provide unequivocal evidence for an important role of TrkA signaling in AAI. The TrkA-KI mouse strain offers the unique advantage of selectively inhibiting TrkA signaling using a PP1 derivative (1NM-PP1) compared to using pharmacological inhibitors in a wild-type setting. Preliminary studies with A. alternata-challenged TrkA-KI mice indicate that blockade of TrkA signaling leads to a dramatic inhibition of airway eosinophilia and prevention of allergen-induced loss of the barrier-forming protein occludin in AEC. In Aim 1 of this application, we will determine the mechanism by which TrkA is activated in Eos by CCL11 and how TrkA signaling regulates Eos trafficking and migration in vitro and in vivo within inflamed blood vessels using 2-color intravital microscopy. Findings will be correlated to in vivo studies examining the role of TrkA in promoting AAI/asthma in acute and chronic models using TrkA-KI mice. In Aim 2, we will determine the mechanism by which TrkA is activated in AEC by A. alternata and how activation of TrkA regulates AEC function (immune response) and barrier integrity (tight junction protein expression). These explorative studies will reveal novel cell type-specific functions regulated by TrkA signaling in Eos and AEC, two cell types that play significant roles in the pathogenesis of allergic asthma, and provide the basis for the development of novel and effective therapeutics.
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