Inhibition of the prolyl isomerase Pin1 improves endothelial function and attenuates vascular remodelling in pulmonary hypertension by inhibiting TGF-beta signalling

Pulmonary arterial hypertension (PAH) is a devastating disease, characterized by obstructive pulmonary vascular remodelling ultimately leading to right ventricular (RV) failure and death. Disturbed transforming growth factor-beta (TGF-beta)/bone morphogenetic protein (BMP) signalling, endothelial cell dysfunction, increased proliferation of smooth muscle cells and fibroblasts, and inflammation contribute to this abnormal remodelling. Peptidyl-prolyl isomerase Pin1 has been identified as a critical driver of proliferation and inflammation in vascular cells, but its role in the disturbed TGF-beta/BMP signalling, endothelial cell dysfunction, and vascular remodelling in PAH is unknown. Here, we report that Pin1 expression is increased in cultured pulmonary microvascular endothelial cells (MVECs) and lung tissue of PAH patients. Pin1 inhibitor, juglone significantly decreased TGF-beta signalling, increased BMP signalling, normalized their hyper-proliferative, and inflammatory phenotype. Juglone treatment reversed vascular remodelling through reducing TGF-beta signalling in monocrotaline + shunt-PAH rat model. Juglone treatment decreased Fulton index, but did not affect or harm cardiac function and remodelling in rats with RV pressure load induced by pulmonary artery banding. Our study demonstrates that inhibition of Pin1 reversed the PAH phenotype in PAH MVECs in vitro and in PAH rats in vivo, potentially through modulation of TGF-beta/BMP signalling pathways. Selective inhibition of Pin1 could be a novel therapeutic option for the treatment of PAH.Cancer Signaling networks and Molecular Therapeutic

Volume load-induced right ventricular failure in rats is not associated with myocardial fibrosis

Background Right ventricular (RV) function and failure are key determinants of morbidity and mortality in various cardiovascular diseases. Myocardial fibrosis is regarded as a contributing factor to heart failure, but its importance in RV failure has been challenged. This study aims to assess whether myocardial fibrosis drives the transition from compensated to decompensated volume load-induced RV dysfunction.MethodsWistar rats were subjected to aorto-caval shunt (ACS, n = 23) or sham (control, n = 15) surgery, and sacrificed after 1 month, 3 months, or 6 months. Echocardiography, RV pressure-volume analysis, assessment of gene expression and cardiac histology were performed.ResultsAt 6 months, 6/8 ACS-rats (75%) showed clinical signs of RV failure (pleural effusion, ascites and/or liver edema), whereas at 1 month and 3 months, no signs of RV failure had developed yet. Cardiac output has increased two- to threefold and biventricular dilatation occurred, while LV ejection fraction gradually decreased. At 1 month and 3 months, RV end-systolic elastance (Ees) remained unaltered, but at 6 months, RV Ees had decreased substantially. In the RV, no oxidative stress, inflammation, pro-fibrotic signaling (TGF beta 1 and pSMAD2/3), or fibrosis were present at any time point.ConclusionsIn the ACS rat model, long-term volume load was initially well tolerated at 1 month and 3 months, but induced overt clinical signs of end-stage RV failure at 6 months. However, no myocardial fibrosis or increased pro-fibrotic signaling had developed. These findings indicate that myocardial fibrosis is not involved in the transition from compensated to decompensated RV dysfunction in this model.Therapeutic cell differentiatio

Egr-1 and its effectors in flow-associated pulmonary arterial hypertension.

Background: In children with congenital heart disease (CHD), pulmonary arterial hypertension (PAH) is a serious cause for morbidity and mortality. PAH is a vasoproliferative disease in which the formation of irreversible neointimal lesions leads to progressive obstruction of the pulmonary arterioles. Increased pulmonary blood flow is a known trigger for this disease. In a model of experimental flow-induced PAH, Egr-1 was identified as a flow-specific trigger. It is expressed progressively throughout disease development. In our rat model for flow-PAH, Egr-1 inhibition using ED5 has attenuated neointimal formation. The spatiotemporal expression of Egr-1 in human, pediatric disease is unknown. The objective of this study is to elucidate the pathways through which Egr-1 mediates disease progression in our rat model and to determine Egr-1 expression in both early- and end-stage pediatric PAH. Methods: We used rats in which flow-PAH was created through monocrotaline injection and an aorto-caval shunt. In addition rats were treated with either saline, scrambled ED5 or ED5. Rats were sacrificed 1 week (early-stage) and 3 weeks (end-stage) after surgery. Using immunohistochemistry (IHC), we performed a proliferation and apoptosis assay and assessed the spatiotemporal expression of 5 possible PAH and Egr-1 associated proteins: tissue factor (TF), platelet-derived growth factor-B (PDGF-B), transforming growth factor-β1 (Tgf-β1), interleukin-6 (IL-6) and p53. Human Egr-1 expression was assessed in 29 children with either early- or end-stage CHD-PAH by IHC analysis of lung biopsies taken prior to correction of their cardiac anomaly. Egr-1 expression was then related to patient characteristics and hemodynamic measurements. Results: Main findings are that patterns of proliferation and apoptosis both peak in early-stage PAH. Egr-1 inhibition causes a decrease in proliferation in early-stage PAH and an increase in apoptosis in end-stage PAH. PDGF-B, Tgf-β1, IL-6 and p53 expression increase both in early and end-stage PAH and their expression decreases on Egr-1 inhibition. TF was only up regulated in end-stage PAH and down regulated by Egr-1 inhibition in that stage. In human end-stage PAH, Egr-1 expression is increased more than in early-stage disease. Egr-1 expression did not correlate to age, sex, shunt characteristics or any of the hemodynamic variables. Conclusions: In this study of experimental PAH, Egr-1 inhibition changed vascular remodeling by reducing hyper proliferation and allowing normal apoptosis. PDGF-B and IL-6 may be important effectors of Egr-1, that induce proliferation in both early and end-stage PAH, while TF might be an Egr-1 effector only in end-stage disease. The Tgf-β1 response is possibly not initiated but maintained by Egr-1. p53 might be an Egr-1 effector with a protective factor in PAH development. This renders more insight in the function of Egr-1 in the pathobiology of flow-associated PAH and might provide a starting point for a more specifically directed approach for therapeutic intervention. Furthermore, we confirmed the presence of Egr-1 in pediatric early-stage CHD-PAH and demonstrated a difference with expression in end-stage disease. The factors that might cause this difference remain unclear.

Reply to Piquereau and Perros and to Pullamsetti and de Jesus Perez

Therapeutic cell differentiatio