Title Page Endothelin Receptor B Protects Granulocyte Macrophage Colony Stimulating Factor mRNA from Degradation

Evidence lacks on the differential effects of the two therapeutic concepts of endothelin receptor antagonists (ERA): the blockade of only the endothelin-receptorA (ETAR) (selective antagonism) versus both ETAR and ETBR (dual blockade). Ambrisentan, a selective ERA, and bosentan, a dual blocker, are both available for therapy. We hypothesized that there are differences in the potential of ERAs to ameliorate inflammatory processes in human airway smooth muscle cells (HASMCs) and aimed to unravel underlying mechanisms. We used HASMC culture, ELISA and qRTPCR. TNFα induced transcription and expression of CXCL2, CXCL3, GM-CSF and MMP12 in HASMCs. In concentration-response experiments, bosentan led to a significantly greater reduction of GM-CSF and MMP12 protein release than ambrisentan while there was no significant difference in their effect on GM-CSF and MMP12 mRNA. Both ERA reduced CXCL3 protein and mRNA equally but had no effect on CXCL2. Blocking mitogen-activated protein kinases revealed that both ETAR and ETBR signal through p38, but ETBR also signals through ERK-1/-2 to induce GMCSF expression. In the presence of the transcription inhibitor actinomycin D, bosentan but not ambrisentan reduced GM-CSF but not MMP12 and CXCL3 mRNA. In conclusion, blockade of each endothelin receptor subtype reduces GM-CSF transcription but blocking ETBR additionally protects GM-CSF mRNA from degradation via ERK-1/-2. Accordingly, blocking both ETAR and ETBR leads to a stronger reduction of TNFα induced GM-CSF protein expression. This mechanism might be specific for GM-CSF. Our data stress the anti-inflammatory potential of ERA and warrants further investigation of their utility in chronic inflammatory airway diseases. This article has not been copyedited and formatted. The final version may differ from this version. JPET Fast Forward. Published on April 8, 2015 as DOI: 10.1124/jpet.114.215822 at A PE T Jornals on D ecem er 3, 2017 jpet.asjournals.org D ow nladed from