Inhibition of PI3K promotes dilation of human small airways in a rho kinase‐dependent manner

Abstract

Background and PurposeAsthma manifests as a heterogeneous syndrome characterized by airway obstruction, inflammation and hyperresponsiveness (AHR). Although the molecular mechanisms remain unclear, activation of specific PI3K isoforms mediate inflammation and AHR. We aimed to determine whether inhibition of PI3Kδ evokes dilation of airways and to elucidate potential mechanisms.Experimental ApproachHuman precision cut lung slices from non‐asthma donors and primary human airway smooth muscle (HASM) cells from both non‐asthma and asthma donors were utilized. Phosphorylation of Akt, myosin phosphatase target subunit 1 (MYPT1) and myosin light chain (MLC) were assessed in HASM cells following either PI3K inhibitor or siRNA treatment. HASM relaxation was assessed by micro‐pattern deformation. Reversal of constriction of airways was assessed following stimulation with PI3K or ROCK inhibitors.Key ResultsSoluble inhibitors or PI3Kδ knockdown reversed carbachol‐induced constriction of human airways, relaxed agonist‐contracted HASM and inhibited pAkt, pMYPT1 and pMLC in HASM. Similarly, inhibition of Rho kinase also dilated human PCLS airways and suppressed pMYPT1 and pMLC. Baseline pMYPT1 was significantly elevated in HASM cells derived from asthma donors in comparison with non‐asthma donors. After desensitization of the β2‐adrenoceptors, a PI3Kδ inhibitor remained an effective dilator. In the presence of IL‐13, dilation by a β agonist, but not PI3K inhibitor, was attenuated.Conclusion and ImplicationsPI3Kδ inhibitors act as dilators of human small airways. Taken together, these findings provide alternative approaches to the clinical management of airway obstruction in asthma.

CF airway smooth muscle transcriptome reveals a role for PYK2

“…One of the small-molecule compounds with a high-connectivity score was GSK-1059615, a pan PI3K inhibitor. Inhibition of PI3K has been shown to reduce airway remodeling (26,27), reduce ovalbumin-induced airway hyperresponsiveness in mice (28), and reverse carbachol-induced constriction of human airways (18). The presence of this airway-modifying drug on our list of potential compounds supported our hypothesis that connectivity mapping could be used to identify therapeutics for airway smooth muscle dysfunction.…”

“…Using this analysis, we found 9 upstream regulators that were dysregulated due to CFTR loss (Supplemental Table 3). Interestingly, many of the proposed upstream regulators, such as phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K) and phosphodiesterase signaling, have been implicated in airway smooth muscle function and disease (18,19). Others, such as WNT and CREB, have been studied in CF (20)(21)(22).…”

Neuronal PAS Domain 2 (Npas2)‐Deficient Fibroblasts Accelerate Skin Wound Healing and Dermal Collagen Reconstruction

“…During migration, fibroblasts adhere to the extracellular matrix (ECM) through integrin molecules and generate a single‐cell traction force (Style et al, ). We used a recently developed single‐cell contraction assay that required cell adhesion to the FLECS printed with fibronectin (Koziol‐White et al, ; Pushkarsky et al, ). The FLECS assay showed that mouse dermal fibroblasts increased the cell contraction behavior by Npas2 KO mutation (Fig.…”

“…Single‐cell contraction was measured using fluorescently labeled elastomeric contractible surfaces (FLECS) (Forcyte Biotechnologies Inc., Los Angeles, CA) (Koziol‐White et al, ). FLECS plates with the soft silicone elastomer filmed bottom were micropatterned with fluorescent fibrinogen in uniform “X” shapes (70 μm diagonal by 10 μm thick).…”

Human Pulmonary 3D Models For Translational Research

“…Researchers thus concentrate their use of these systems on the investigation of rapid (seconds to hours) or short‐term (within days) responses. In asthma research, PCLS are used extensively for the investigation of new drugs or interventions interfering with bronchial constriction . Noteworthy, carefully done studies point to remarkable differences between mice and man with respect to the regulation of bronchial constriction .…”