Introduction and Objectives 5%–10% of asthmatic patients display relative insensitivity to the therapeutic benefits corticosteroids and are termed severe asthmatics. This has been attributed, in severe asthma (SA) airway smooth muscle cells (ASMCs), to defective corticosteroid-induced nuclear translocation of the glucocorticoid receptor (GR). Importins, including importin-7, mediate GR nuclear translocation. Studies report increased importin-7-GR interaction in response to corticosteroids in ASMCs. Long-acting beta agonists (LABAs) augment corticosteroid-induced GR nuclear translocation in COPD macrophages. The primary aim of this project was to determine whether the LABA salmeterol could reverse impaired corticosteroid (dexamethasone)-induced GR nuclear translocation in severe asthmatic ASMCs through an importin-7-dependent mechanism.

Methods ASMCs derived from bronchial biopsies from patients with non-severe asthma (NSA) (n=4) and SA (n=4) were cultured and treated with dexamethasone (10–8 M) in the absence or presence of salmeterol (10–10 M or 10–8 M). 30 min post-stimulation, nuclear and cytoplasmic protein fractions underwent western blot analysis to determine levels of importin-7 and GR. RNA extraction was conducted at 4 hours post-stimulation followed by RT-qPCR to investigate the levels of the GR-activated genes, mitogen activated protein kinase phosphatase-1 (MKP-1) and glucocorticoid leucine zipper mRNA levels (GILZ).

Results The Results show a reduced dexamethasone-induced GR nuclear translocation in SA when compared to NSA ASMCs, which was increased with co-treatment with dexamethasone and salmeterol. Co-treatment with dexamethasone and salmeterol also increased GR-activated gene expression leading to an increase in the expression of MKP-1 in SA ASMCs compared to dexamethasone alone. Nuclear importin-7 levels in NSA and SA ASMCs were not increased by co-treatment compared to dexamethasone alone.

Conclusion We have demonstrated impaired dexamethasone-induced GR nuclear translocation in SA ASMCs, which can be reversed by the addition of the LABA salmeterol in combination with dexamethasone. No differences were observed in importin-7 nuclear localisation after co-treatment compared to dexamethasone alone in SA ASMCs. This could suggest that importin-7 is either rapidly recycled back into the cytoplasm after entry into the nucleus, or that the effect of LABAs are not mediated through Importin-7 in severe asthmatic patients. Further real-time experiments would elucidate the role of importin-7 in LABA mediated GR nuclear translocation.