TNFα-induced airway smooth muscle cell proliferation depends on endothelin receptor signaling, GM-CSF and IL-6

Pathological proliferation of human airway smooth muscle cells (HASMCs) causes hyperplasia in chronic lung diseases. Signaling pathways that link airway inflammation to HASMC proliferation might provide therapeutic targets for the prevention of airway remodeling and chronic lung diseases. Endothelin-1 (ET-1) signals via endothelin-A- and B-receptors (ETAR, ETBR) to perpetuate HASMC-associated and TNF alpha-dependent inflammatory processes. Hypothesis: endothelin receptor antagonists (ERAs) suppress HASMC proliferation induced by inflammatory cytokines. HASMCs were stimulated ex vivo with cytokines in the presence or absence of ERAs (ETAR-specific/selective: BQ123, ambrisentan; ETBR-specific: BQ788; non-selective: bosentan, macitentan, ACT-132577) or cytokine-blocking antibodies. Cell counts, DNA-synthesis (BrdU-incorporation assay), cytokine production (ELISA) and ETBR expression (whole-genome microarray data, western blot) were analyzed. ET-1-induced HASMC proliferation and DNA synthesis were reduced by protein kinase inhibitors and ETAR-specific/selective ERAs but not by BQ788. TNF alpha-induced HASMC proliferation and DNA-synthesis were reduced by all ERAs. TNF alpha induced ET-1 and ETBR expression. TNF alpha- and ET-1-induced GM-CSF releases were both reduced by BQ123 and BQ788. TNF alpha- and ET-1-induced IL-6 releases were both reduced by BQ123 but not by BQ788. Combined but not single blockade of GM-CSF-receptor-alpha-chain and IL-6 reduced TNFa- and ET-1 induced HASMC proliferation and DNA-synthesis. Combined but not single treatment with GM-CSF and IL-6 induced HASMC proliferation and DNA-synthesis in the presence of ET-1. In conclusion, TNF alpha induces HASMC proliferation via ET-1/GM-CSF/IL-6. ETBR requires up-regulation by TNFa to mediate ET-1 effects on HASMC proliferation. This signaling cascade links airway inflammation to HASMC-associated remodeling processes and is sensitive to ERAs. Therefore, ERAs could prevent inflammation induced airway smooth muscle hyperplasia. (C) 2016 Elsevier Inc. All rights reserved.