Molecular mechanisms of inflammation and calcification in aortic valve stenosis (2012)

<p>Aortic valve stenosis is a slowly progressive disorder with a spectrum of disease ranging from aortic sclerosis to severe destroyed valvular architecture leading to critical outflow obstruction. The diseased valve is characterized by inflammation, as an initiating event, pathological remodeling of extracellular matrix and pronounced calcification, which all eventually cause restricted leaflet mobility. Compelling evidence obtained from both experimental animal models and human studies provided detailed histopathological picture of disease development. This implies endothelial cell-, macrophage activation and inflammation-dependent calcification paradigm, in which phenotypic transdifferentiation of valvular myofibroblasts towards osteogenic phenotype takes place to further enhance the structural and compositional changes of the valve leaflets. The prognosis of symptomatic patients with severe stenosis is poor without surgical valve replacement. To date there is no medical treatment for this condition other than surgical valve replacement.</p><p>Based on inflammation associated calcification, observed in early stages, within this thesis the molecular mechanisms of calcified aortic valve stenosis with focus on inflammation and subsequent calcification were characterized. The aim of the project was to determine the role of proinflammatory signaling through the leukotriene pathway in aortic stenosis. Surgically explanted human aortic valves were subjected for macroscopic dissection followed by Taqman qPCR in order to correlate the gene expression pattern with the echocardiographic parameters quantifying the stenosis severity. This study established a macroscopic dissection technique as a model for in vivo disease development, where different parts of stenotic aortic valve represent the entire disease spectrum from early signs to advanced stages. The inflammatory environment within the affected aortic valve stimulates the 5-lipoxygenase pathway leading to production of potent inflammatory mediators, leukotrienes. Several components of the 5-lipoxygenas pathway were correlated with the stenosis severity. As inflamed valvular tissue demonstrates signs of micro- and macrocalcification, the next step was to determine the spatio-temporal distribution of genes with osteoinductive and osteoresorptive pontential. These findings along with the plasma measures of proteins taking part in bone turnover point to a correlation between their tissue-, systemic levels and the stenosis severity. Furthermore, the intracellular effects of LTC4 in vitro cell culture of valvular insterstitial cells and smooth muscle cells from human coronary artery were characterized. The results of these experiments demonstrated cascade of events leading to activation of cell death pathways, activation of the nuclear enzyme PARP-1, upregulation of CysLT1 receptor expression in response to proinflammatory stimuli coupled to nuclear calcium signaling. Moreover, in a separate experiment a conceptual model of phenotypic plasticity of the valvular insterstitial cells due to epigenetic alteration was provided, which might be connected to stenosis progression. The amount of methylated DNA within the 5-lipoxygenase promoter region was significantly lower in the calcified part compared with non-calcified, which was confirmed by accompanied increase in expression of 5-lipoxygenase leading to enhanced inflammatory activity in the calcified region of the valve.</p><p>In conclusion, the findings of this thesis integrate observations of molecular research using quantitative gene expression data with clinical variables in terms of echocardiographic parameters. The results point to leukotrienes as potential mediators of inflammation in aortic stenosis. Furthermore, the bone turnover both at tissue and systemic levels is associated with stenosis severity. In addition, the thesis also provides mechanistic insight into the direct role of leukotrienes in the pathophysiological process of aortic stenosis. Finally, translational implication of our data suggests possibility for pharmacological intervention using leukotriene receptor antagonists with a potential to retard the hemodynamic progression.</p><h3>List of scientific papers</h3><p>I. Edit Nagy, Daniel Andersson, Kenneth Caidahl, Maria J. Eriksson, Per Eriksson, Anders Franco-Cereceda, Göran K Hansson, and Magnus Bäck Upregulation of the 5-lipoxygenase pathway in human aortic valves correlates with severity of stenosis and leads to leukotriene-induced effects on valvular myofibroblasts. Circulation. 2011;123:1316-25. <br><a href="https://doi.org/10.1161/CIRCULATIONAHA.110.966846">https://doi.org/10.1161/CIRCULATIONAHA.110.966846</a><br><br> </p><p>II. Edit Nagy, Kenneth Caidahl, Anders Franco-Cereceda and Magnus Bäck. Increased transcript level of poly(ADP-ribose) polymerase (PARP-1) in human tricuspid compared with bicuspid aortic valves correlates with the stenosis severity. Biochem Biophys Res Commun. 2012;420:671-5. <br><a href="https://doi.org/10.1016/j.bbrc.2012.03.064">https://doi.org/10.1016/j.bbrc.2012.03.064</a><br><br> </p><p>III. Alison Eaton, Edit Nagy, Jérémy Fauconnier, Mathilde Pacault, Magnus Bäck. Cysteinyl-leukotriene signaling through perinuclear CysLT1 receptors on vascular smooth muscle cells transduces nuclear calcium signaling and alterations of gene expression. J Mol Med. 2012; Apr 20. [Epub ahead of print]. <br><a href="https://doi.org/10.1007/s00109-012-0904-1">https://doi.org/10.1007/s00109-012-0904-1</a><br><br> </p><p>IV. Edit Nagy, Magnus Bäck. Epigenetic regulation of 5-lipoxygenase in the phenotypic plasticity of valvular interstitial cells associated with aortic valve stenosis. FEBS Letters. 2012; 586,1325-1329. <br><a href="https://doi.org/10.1016/j.febslet.2012.03.039">https://doi.org/10.1016/j.febslet.2012.03.039</a><br><br> </p><p>V. Edit Nagy, Per Eriksson, Mohamed Yousri, Kenneth Caidahl, Erik Ingelsson, Göran K. Hansson, Anders Franco-Cereceda, Magnus Bäck. The role of valvular osteoclasts in calcification and aortic valve stenosis severity. [Manuscript]</p>