Abstract 11784: Reduced BMPR2 Increases GM-CSF mRNA Translation by Inhibiting eIF2α Mediated Stress Granule Formation and Propensity to Pulmonary Vascular Disease
Background: Inflammation in human and experimental pulmonary artery hypertension (PAH) is associated with reduced BMPR2. We reported an increase in the pro-inflammatory cytokine GMCSF in lung and pulmonary arteries (PA) from patients with idiopathic PAH. Reducing BMPR2 by siRNA in cultured human PA endothelial cells (EC) enhanced TNF-α induced-GMCSF mRNA translation by prolonging p38 phosphorylation. Translation of cytokine mRNAs is normally repressed when phosphorylation of eIF2 recruits mRNAs and RNA binding proteins such as HuR and TIA-1 to stress granules (SGs). We therefore hypothesized that reduced BMPR2 via p-p38 enhances GM-CSF mRNA translation by dis-assembly of SGs and that GM-CSF worsens PVD.
Methods: In TNF-α stimulated PAEC, we determined, by western immunoblot, the effects of loss of BMPR2, SB202190 (p38 inhibitor) and salubrinal, [inihibitor of protein phosphatase 1(PP1)] on phosphorylation of eIF2. Dis-assembly of SGs was analyzed by immunofluorescent staining for HuR and TIA-1. To establish the clinical significance of our observations, we assessed the distribution of GM-CSF receptor (R)α (+) cells in the PAs of IPAH patients by immunohistochemistry and microfluidic single cell transcriptional analysis. We also administered murine GM-CSF to mice exposed to chronic hypoxia for three weeks to determine whether this increased PVD.
Results: TNF-α mediated eIF2α phosphorylation and SG assembly was reversed by dropping BMPR2 levels in human PA ECs The decline in phospho-eIF2α required p-p38 activation of PPI phosphatase. In the IPAH lung, GM-CSFRα co-distributed with endothelium and macrophages in diseased PAs. Flow cytometory of freshly isolated lung CD31 (+) cells revealed significant expansion of GM-CSFRα (+) cells from IPAH lungs (P<0.05). Single cell transcriptional analysis demonstrated two subpopulations one associated with EC the other with monocyte/macrophage genes. Exogeneous GM-CSF exacerbated hypoxia-induced PH in mice, related to more muscularized PAs and perivascular macrophages.
Conclusions: Loss of BMPR-II causes prolonged p-p38 activity that is required for GADD34/PPI complex to dis-assemble SGs and enhance GM-CSF mRNA translation. GM-CSF contributes to PVD by recruiting inflammatory cells.
- © 2011 by American Heart Association, Inc.