Angiotensin II receptor 1 controls profibrotic Wnt/β-catenin signalling in experimental autoimmune myocarditis

Aims Angiotensin (Ang) II signalling has been suggested to promote cardiac fibrosis in inflammatory heart diseases; how- ever, the underlying mechanisms remain obscure. Using Agtr1a-/- mice with genetic deletion of angiotensin receptor type 1 (ATR1) and the experimental autoimmune myocarditis (EAM) model, we aimed to elucidate the role of Ang II-ATR1 pathway in development of heart-specific autoimmunity and post-inflammatory fibrosis. .................................................................................................................................................................................................... Methods EAM was induced in wild-type (WT) and Agtr1a-/- mice by subcutaneous injections with alpha myosin heavy chain and results peptide emulsified in complete Freund’s adjuvant. Agtr1a-/- mice developed myocarditis to a similar extent as WT controls at day 21 but showed reduced fibrosis and better systolic function at day 40. Crisscross bone marrow chi- maera experiments proved that ATR1 signalling in the bone marrow compartment was critical for cardiac fibrosis. Heart infiltrating, bone-marrow-derived cells produced Ang II, but lack of ATR1 in these cells reduced transforming growth factor beta (TGF-b)-mediated fibrotic responses. At the molecular level, Agtr1a-/- heart-inflammatory cells showed impaired TGF-b-mediated phosphorylation of Smad2 and TAK1. In WT cells, TGF-b induced formation of RhoA-GTP and RhoA-A-kinase anchoring protein-Lbc (AKAP-Lbc) complex. In Agtr1a-/- cells, stabilization of RhoA- GTP and interaction of RhoA with AKAP-Lbc were largely impaired. Furthermore, in contrast to WT cells, Agtr1a-/- cells stimulated with TGF-b failed to activate canonical Wnt pathway indicated by suppressed activity of glycogen synthase kinase-3 (GSK-3)b and nuclear b-catenin translocation and showed reduced expression of Wnts. In line with these in vitro findings, b-catenin was detected in inflammatory regions of hearts of WT, but not Agtr1a-/- mice and expression of canonical Wnt1 and Wnt10b were lower in Agtr1a-/- hearts. .................................................................................................................................................................................................... Conclusion Ang II-ATR1 signalling is critical for development of post-inflammatory fibrotic remodelling and dilated cardiomyop- athy. Our data underpin the importance of Ang II-ATR1 in effective TGF-b downstream signalling response includ- ing activation of profibrotic Wnt/b-catenin pathway.