The angiotensin–calcineurin–NFAT pathway mediates stretch-induced up-regulation of matrix metalloproteinases-2/-9 in atrial myocytes
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During atrial fibrillation, arterial hypertension and systolic or diastolic heart failure, atrial myocytes are exposed to increased baseline stretch. Atrial stretch has been shown to induce cellular hypertrophy and extracellular matrix remodeling (ECM) via angiotensin-II dependent pathways and the matrix metalloproteinases system (MMPs). We hypothesized that atrial myocytes exposed to static stretch may increase their ECM remodeling activity via up-regulation of MMP-2/-9. We then tested the hypothesis that the membrane bound angiotensin-II type 1 (AT1) receptor and the intracellular calcineurin (Cn)-NFAT signaling pathway are potential mediators of stretch-induced MMP alterations, since Cn-NFAT is one important contributor to myocyte hypertrophy.
Methods and results
Neonatal rat atrial myocytes (NRAM) were cultured under conditions of static stretch by 21%. The differential effects of selective AT1 receptor blockade by losartan, Cn blockade by Cyclosporine-A (CsA) or NFAT inhibition by 11R-VIVIT (VIV), were analyzed. Stretch resulted in a significant up-regulation of active-MMP-2/-9 protein amount (active-MMP-2 ng/µg: control 8.95 ± 0.64 vs. stretch 13.11 ± 0.74 / active-MMP-9 ng/µg: control 1.45 ± 0.18 vs. stretch 1.94 ± 0.21, all n = 5) and enzyme activity (MMP-2 in %: control 1 ± 0.0 vs. stretch 1.87 ± 0.25, n = 7) associated with a significant increase of the membrane-type-1-MMP (MT1-MMP) protein expression (MT1-MMP in %: control 1 ± 0.0 vs. stretch 2.17 ± 0.21, n = 8). These observations were accompanied by an activation of the Cn-NFAT pathway (Cn-activity in nmol PO4 release/20 µg protein/30 min: control 0.37 ± 0.08 vs. stretch 0.65 ± 0.09, n = 3 / NFATc1-DNA binding activity in %: control 1 ± 0.0 vs. stretch 1.53 ± 0.17, n = 3). Losartan, CsA or VIV abolished stretch-induced alterations in MMP-2/-9 and MT1-MMP expression and enzyme activity by normalizing the Cn-activity and the DNA binding activity of NFATc1.
Our results present new insights in molecular mechanisms of ECM remodeling activity of atrial myocytes exposed to static stretch. The AT1-Cn-NFAT pathway is a potential mediator of MMP activation.
Keywordsmechanical stretch atrial fibrillation calcineurin matrix metalloproteinases
We thank the entire Institute of Laboratory Animal Science, University Hospital, RWTH Aachen in Germany for the helpful assistance in animal research. This work was supported in part by Deutsche Forschungsgemeinschaft (DFG) grant Lu 869/4-1 and IZKF Biomat. RWTH Aachen, Germany to A.L. and by the Network of Competence Atrial Fibrillation.
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