The Role And Mechanism Of Transforming Growth Factor Beta 1 In Airway Hyperresponsiveness
Asthma, a chronic inflammatory airway disease characterized by airway hyperresponsiveness (AHR) and airway remodeling, affects 300 million people worldwide. Conventional asthma management includes the use of glucocorticoids and β2-agonist bronchodilators to combat inflammation and reverse airway narrowing. In severe asthma patients, whose asthma is poorly controlled with conventional therapies, frequent asthma exacerbations can lead to sustained AHR that remains insensitive to bronchodilator therapy. Therefore, it is essential to determine the mechanisms contributing to AHR and irreversible airflow obstruction in asthma to reduce patient morbidity and mortality. Transforming growth factor beta 1 (TGF-β1), a growth factor elevated in the airway of patients with asthma, perpetuates airway inflammation and airway remodeling in airway structural cells. The role of TGF-β1 in mediating AHR, however, remains unclear. In this dissertation, we demonstrate that TGF-β1 primes agonist-induced contractile responses and attenuates agonist-induced relaxation pathways in human airway smooth muscle (HASM), contributing to AHR and irreversible airflow obstruction in asthma. The dynamics of AHR and single-cell excitation-contraction coupling were measured using supravital microscopy, magnetic twisting cytometry, and biochemical assays in human precision-cut lung slices (hPCLS) and isolated primary HASM cells. We demonstrate that overnight TGF-β1 treatment augmented basal and agonist-induced shortening in hPCLS and isolated HASM cells. Interestingly, TGF-β1 increased HASM cell shortening and myosin light chain phosphorylation in a Smad3-dependent manner, with little effect on intracellular calcium levels. Additionally, we find that overnight TGF-β1 treatment decreased β2-agonist-induced relaxation responses in HASM cells via induction of phosphodiesterase 4 (PDE4), an enzyme that negatively regulates β2-agonist signaling pathways. Pharmacological and siRNA inhibition of PDE4 and Smad3, respectively, restored β2-agonist sensitivity in TGF-β1-treated HASM cells. Together, our data suggest that increased levels of TGF-β1 in the airway contribute to airway narrowing, asthma exacerbations, and bronchodilator resistance in asthma. Furthermore, our results establish TGF-β1 as a novel therapeutic target to decrease airway exacerbations in patients with severe and treatment-resistant asthma.