Pulmonary arterial hypertension (PAH) is a devastating disease, characterized by obstructive pulmonary vascular remodelling ultimately leading to right ventricular (RV) failure and death. Disturbed... Show morePulmonary 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. Show less
14q32 microRNAs are known to play a role in various forms of vascular remodelling. This thesis elucidated that snoRNAs of the 14q32 locus are also involved in vascular remodelling processes. The... Show more14q32 microRNAs are known to play a role in various forms of vascular remodelling. This thesis elucidated that snoRNAs of the 14q32 locus are also involved in vascular remodelling processes. The expression of both noncoding RNA types in the human vasculature has been found to be vascular location and vessel type specific and are therefore promising targets for future implementation in clinical practice.The second part of this thesis focuses on three different types of 14q32 microRNA expression regulation in order to affect various vascular remodelling processes. 14q32 DNA methylation, myostatin and CIRBP were tested for their effect on 14q32 microRNA expression and the (subsequent) effect on vein graft disease and tissue ischemia, restenosis and angiogenesis, respectively. DNA methylation is not correlated with 14q32 microRNA expression, but directly interacts with vascular remodelling process status. Myostatin negatively affects 14q32 microRNA expression in vascular smooth muscle cells, but not in inflammatory cells involved in restenosis. Due to this latter finding, overall restenosis was not inhibited by myostatin. Inhibition of CIRBP inhibited 14q32 microRNA expression post-transcriptionally and therefore increased in vitro angiogenesis. These promising findings provide novel indirect regulators of vascular remodelling processes and future research will elucidate the potential for clinical application. Show less
Aims Pulmonary arterial hypertension (PAH) is associated with increased levels of circulating growth factors and corresponding receptors such as platelet derived growth factor, fibroblast growth... Show moreAims Pulmonary arterial hypertension (PAH) is associated with increased levels of circulating growth factors and corresponding receptors such as platelet derived growth factor, fibroblast growth factor and vascular endothelial growth factor. Nintedanib, a tyrosine kinase inhibitor targeting primarily these receptors, is approved for the treatment of patients with idiopathic pulmonary fibrosis. Our objective was to examine the effect of nintedanib on proliferation of human pulmonary microvascular endothelial cells (MVEC) and assess its effects in rats with advanced experimental pulmonary hypertension (PH).Methods and results Proliferation was assessed in control and PAH MVEC exposed to nintedanib. PH was induced in rats by subcutaneous injection of Sugen (SU5416) and subsequent exposure to 10% hypoxia for 4weeks (SuHx model). Four weeks after re-exposure to normoxia, nintedanib was administered once daily for 3 weeks. Effects of the treatment were assessed with echocardiography, right heart catheterization, and histological analysis of the heart and lungs. Changes in extracellular matrix production was assessed in human cardiac fibroblasts stimulated with nintedanib. Decreased proliferation with nintedanib was observed in control MVEC, but not in PAH patient derived MVEC. Nintedanib treatment did not affect right ventricular (RV) systolic pressure or total pulmonary resistance index in SuHx rats and had no effects on pulmonary vascular remodelling. However, despite unaltered pressure overload, the right ventricle showed less dilatation and decreased fibrosis, hypertrophy, and collagen type III with nintedanib treatment. This could be explained by less fibronectin production by cardiac fibroblasts exposed to nintedanib.Conclusion Nintedanib inhibits proliferation of pulmonary MVECs from controls, but not from PAH patients. While in rats with experimental PH nintedanib has no effects on the pulmonary vascular pathology, it has favourable effects on RV remodelling. Show less
Cardiovascular disease (CVD) is the collective term for diseases that involve the heart or circulation and CVDs are a major cause of mortality and morbidity worldwide. The aim of thesis was to... Show moreCardiovascular disease (CVD) is the collective term for diseases that involve the heart or circulation and CVDs are a major cause of mortality and morbidity worldwide. The aim of thesis was to investigate the role of inflammation in CVD related cardiac and vascular remodelling, which may lead to potential therapeutic agents. We investigated the therapeutic potential of antibodies directed against phosphorylcholine (PC), an endogenous ligand capable of triggering the innate immune system, which is expressed by apoptotic cells and oxidized LDL, in mouse models for myocardial infarction (MI). We found that treatment with anti-PC antibodies reduces adverse cardiac remodelling after both permanent MI as myocardial ischemia reperfusion (MI-R) injury. Furthermore, we found that treatment with annexin A5 also reduces adverse cardiac remodelling after MI-R injury. Interestingly, both anti-PC as annexin A5 treatment reduced the post MI inflammatory response. Next, we investigated the role of PCAF, an inflammatory related epigenetic factor, in vascular remodelling. We found that PCAF deficiency and treatment with a PCAF inhibitor reduces adverse vascular remodelling. Finally, we investigated the role of microRNAs, small RNA molecules that can affect expression of many different gene simultaneously, in vascular remodelling. We show that inhibition of microRNA-495 reduces adverse vascular remodelling. Show less
This thesis describes the role of 14q32 microRNAs in vascular remodelling. The 14q32 microRNA cluster contains 54 microRNAs in humans and is highly conserved in mammals. In part I of this thesis,... Show moreThis thesis describes the role of 14q32 microRNAs in vascular remodelling. The 14q32 microRNA cluster contains 54 microRNAs in humans and is highly conserved in mammals. In part I of this thesis, we describe the role of 14q32 microRNAs in several processes of vascular remodelling. We have shown that inhibition of several 14q32 microRNAs, miR-329, miR-494 and miR-495, results in increased neovascularisation after hindlimb ischemia in mice. In addition, inhibition of the same microRNAs reduced atherosclerotic plaque formation and restenosis in experimental mouse models under hypercholesterolemic conditions. In part II of this thesis, we zoom in to the post-transcriptional regulation of 14q32 microRNAs through RNA binding proteins. The third and last part of this thesis studies the expression of microRNAs in subcutaneous adipose tissue of critical limb ischemia patients and discusses the potential use of microRNAs as biomarker to predict the risk of amputation in these patients. In conclusion, this thesis provides novel insights in the role of 14q32 microRNAs in processes of vascular remodelling. Experimental studies have identified 14q32 microRNAs as potential therapeutic targets for treatment and prevention of atherosclerosis, restenosis and peripheral arterial disease. Show less
