Chronic use of glucocorticoids (GCs), which are among the most frequently prescribed drugs, has been associated with various side effects including cardiovascular complications such as systemic hypertension and cardiac dysfunction. However, its effects on the pulmonary vascular bed are less clearly described. Elevated levels of reactive oxygen species (ROS) have been associated with a variety of disorders of the systemic and pulmonary vasculature, although the role of ROS in GCs signaling is controversial.

We therefore evaluated the impact of the synthetic GC dexamethasone (DEX) on vascular ROS signalling in vitro and in vivo.

Treatment with DEX induced a rapid but sustained elevation of ROS production in a dose dependent manner in endothelial cells as well as in explanted murine systemic and pulmonary vessels.

Depletion of the NADPH oxidase subunits NOX2 or NOX4 by RNAi decreased ROS levels in DEX treated cells and vessels. In addition, DEX increased NOX2 activity, as well as NOX mRNA and protein levels.

Since we previously could show that ROS derived from NADPH oxidases can activate hypoxia inducible transcription factors (HIFs) we further determined the impact of DEX on HIF activity. We found that treatment with DEX not only enhanced HIF activity but also increased the levels of HIF-1α, and that the depletion of NOX2 or NOX4 prevented these DEX-induced responses. Since the HIF system has been described to be highly involved in controlling angiogenesis we evaluated DEX effects on endothelial proliferative responses. Treatment with DEX enhanced endothelial cell proliferation and tube formation in an in vitro angiogenesis assay, and these effects were dependent on ROS derived from NOX2 and NOX4.

Subsequent in vivo experiments showed that chronic treatment of mice with low doses of DEX induced pulmonary vascular remodelling and increased the size of the right ventricle. When mice with a deficiency of HIF-1alpha in the endothelium were treated with DEX, induction of pulmonary vascular remodelling and right ventricular hypertrophy was substantially diminished.

Taken together, our data show that treatment with DEX increases endothelial cell ROS production via NOX2 and NOX4, which leads to activation of the HIF system and a subsequent angiogenic response. This pathway also seems to be important for in vivo effects of chronic DEX treatment resulting in pulmonary vascular remodelling and right ventricular hypertrophy.