Balancing TGFbeta/BMP in pulmonary arterial hypertension

Abstract

Introduction: Hemodynamic alterations contribute to disease progression in Pulmonary Arterial Hypertension (PAH), a deadly lung disease associated with mutations in the Transofrming Growth Factor beta (TGFb) –Superfamily. Mutations of the bone morphogenetic protein receptor 2 (BMPR2) and increased expression of TGFb support the idea for a central role of TGFb/BMP imbalance in PAH. Therefore we investigated whether shear stress induces dysfunctional TGFb/BMP responses in pulmonary microvascular endothelial cells (MVEC) from PAH patients. Methods: Control MVECs from cancer lobectomies were compared to PAH MVECs isolated from autopsy and transplantation biopsies. Confluent cells were stimulated with TGFb1 or BMP9 or subjected to shear stress, and downstream phosphorylation of Smad2/3 and Smad1/5 was quantified. Reversal of shear stress induced responses was tested by addition of TGFb neutralizing antibody 1D11 and the selective inhibitor of the BMP type I receptor kinases LDN-193189. Results: Both control and PAH MVECs showed an increased phosphorylation of Smad2/3 upon TGFb1 and shear stress stimulation, which was blocked by 1D11. Suprisingly, BMPR2 mediated responses appeared to be intact and led to increased phosphorylation of Smad1/5 when exposed to BMP9 and shear stress, which was inhibited in presence of LDN-193189. Conclusions: Under the in vitro conditions used, both TGFb and BMP mediated signaling is intact in PAH MVECs. Upon exposure to shear stress or ligand stimulation, control and PAH MVECs showed no differences in downstream signaling of the TGFb/BMP pathway. The effective inhibition of Smad phosphorylation during shear stress shows the importance of TGFb ligands and BMPRI with regards to shear adaptation in MVECs.