Cooperativity between β‐agonists and c‐Abl inhibitors in regulating airway smooth muscle relaxation

Abstract

Current therapeutic approaches to avoid or reverse bronchoconstriction rely primarily on β2 adrenoceptor agonists (β‐agonists) that regulate pharmacomechanical coupling/cross bridge cycling in airway smooth muscle (ASM). Targeting actin cytoskeleton polymerization in ASM represents an alternative means to regulate ASM contraction. Herein we report the cooperative effects of targeting these distinct pathways with β‐agonists and inhibitors of the mammalian Abelson tyrosine kinase (Abl1 or c‐Abl). The cooperative effect of β‐agonists (isoproterenol) and c‐Abl inhibitors (GNF‐5, or imatinib) on contractile agonist (methacholine, or histamine) ‐induced ASM contraction was assessed in cultured human ASM cells (using Fourier Transfer Traction Microscopy), in murine precision cut lung slices, and in vivo (flexiVent in mice). Regulation of intracellular signaling that regulates contraction (pMLC20, pMYPT1, pHSP20), and actin polymerization state (F:G actin ratio) were assessed in cultured primary human ASM cells. In each (cell, tissue, in vivo) model, c‐Abl inhibitors and β‐agonist exhibited additive effects in either preventing or reversing ASM contraction. Treatment of contracted ASM cells with c‐Abl inhibitors and β‐agonist cooperatively increased actin disassembly as evidenced by a significant reduction in the F:G actin ratio. Mechanistic studies indicated that the inhibition of pharmacomechanical coupling by β‐agonists is near optimal and is not increased by c‐Abl inhibitors, and the cooperative effect on ASM relaxation resides in further relaxation of ASM tension development caused by actin cytoskeleton depolymerization, which is regulated by both β‐agonists and c‐Abl inhibitors. Thus, targeting actin cytoskeleton polymerization represents an untapped therapeutic reserve for managing airway resistance.

The intermediate filament protein nestin serves as a molecular hub for smooth muscle cytoskeletal signaling

“…The role of nestin in smooth muscle contraction has not been previously investigated. We have established the technique to determine airway narrowing of PCLS [ 38 , 52 ]. We treated human PCLS with control or nestin siRNA.…”

Platelet-derived growth factor (PDGF)-BB regulates the airway tone via activation of MAP2K, thromboxane, actin polymerisation and Ca2+-sensitisation

“…Our results are supported by the facts that (1) actin polymerisation is of impact for the contractile effect of PDGF-BB in pulmonary veins (GP) [ 19 ] and (2) that actin polymerisation is of relevance for airway hyperresponsiveness [ 62 , 65 ]. Further, Nayak et al [ 66 ] recently demonstrated the additive effects of beta-agonists and Abl-inhibitors on the airway tone in murine PCLS and in mice with the flexiVent system [ 66 ].

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“…Our results are supported by those of Chopra et al [ 50 ] who proved a relaxant effect of the TKIs ST638, genistein and tyrphostin A47 in isolated bronchioles of rats. Recently, Nayak et al [ 66 ] showed in murine PCLS and in a murine in vivo model (flexiVent) that TKIs and β-agonist act synergistic within the context of bronchorelaxation. Beyond, the relaxant effects of several TKIs, e.g.…”

Use of human airway smooth muscle in vitro and ex vivo to investigate drugs for the treatment of chronic obstructive respiratory disorders

“…The day after, slices are washed and returned to the incubator for 1 h before experiments (Koziol‐White, 2022; Sanderson, 2011; Slack et al, 2013). The preparation of PCLS via inflation of warm agarose requires specific buffers such as the Hanks' balanced salt solution (HBSS) with Ca 2+ and Mg 2+ supplemented with HEPES and Dulbecco's modified Eagle medium (DMEM)/F‐12 with antibiotics (Bai et al, 2016; Nayak et al, 2021).…”