Novel link between angiotensin II and Wnt signalling in myocardial fibrogenesis in inflammatory cardiomyopathy

Abstract

Rationale: Identification of genetic or epigenetic factors regulating the pathophysiology of inflammatory dilated cardiomyopathy (iDCM) might result in novel treatment strategies against inflammatory heart diseases. Angiotensin (Ang) II contributes to fibrogenesis through the production of pro-fibrotic factors and has been implicated in the pathogenesis of diverse cardiovascular diseases. Wnt signalling controls heart development, but on the other hand - plays a pivotal role in adult cardiac remodelling, mainly in the fibrosis formation. However, the link between Ang II and Wnt pathways and their particular roles in the progression of the iDCM remain unknown yet. Methods and results: Experimental autoimmune myocarditis (EAM), mimicking human iDCM, was induced in BALB/c wild-type (wt) and Agtr1-/- mice using alpha-myosin heavy chain peptide/complete Freund's adjuvant immunization. Healthy heart fibroblasts (HF) or heart-infiltrating inflammatory CD133+ progenitors (representing main source of cardiac myofibroblasts) were isolated at the peak of EAM, expanded and treated with Transforming Growth Factor-beta (TGF-β), Wnt proteins or their inhibitors. We demonstrated for the first time that Wnt signalling is activated in wt myocardium (within infiltrating inflammatory cells) in the acute phase and persist in TGF-β-mediated fibrotic phase of the EAM (mainly within myofibroblasts). Wt HF or heart-infiltrating inflammatory CD133+ progenitors upon stimulation with TGF-β differentiated into pathogenic myofibroblasts. This process is regulated by Wnt signalling. Blocking of Wnt activity with natural or chemical inhibitors abrogated myofibroblast differentiation of both, wt HF or inflammatory CD133+ progenitors in vitro, and in vivo- that prevented myocardial fibrosis formation. Moreover, we showed that Agtr1-/- mice develop similar extents of myocarditis, but with significantly reduced post-inflammatory fibrosis compared to wt animals. Specific chimera mice revealed that bone marrow cell compartment is indeed responsible for the fibrosis formation. Importantly, Wnt signalling was not activated in Agtr1-/- heart following the EAM induction. Furthermore, heart-infiltrating inflammatory CD133+ progenitors derived from Agtr1-/- inflamed heart showed significantly reduced TGF-β-mediated myofibroblast differentiation and lack of Wnt signalling activation. Conclusion: Our results clearly define Ang II and Wnt signalling as main mediators of myocardial fibrogenesis in the EAM model. We assume that specific targeting of Ang II/Wnt signalling components might be suitable for therapeutic interventions.