The Role of β-adrenergic Signalling and Natriuretic Peptide Receptors in Cardiac Myofibroblast Proliferation and Differentiation

Abstract

Uncontrolled proliferation of cardiac myofibroblasts (CMFs) leads to increased extracellular matrix (ECM) remodelling, resulting in cardiac fibrosis. Cardiac fibrosis occurs in many forms of heart disease and increases the risk of progression to heart failure. Treatment with atrial natriuretic peptide (ANP) was shown to decrease CMF proliferation. Chronic treatment of adult mice with a synthetic catecholamine, isoproterenol (ISO), induced cyclin dependent kinase 1 (CDK1) expression in non-cardiomyocytes. The aims of this study were to develop and characterize an in vitro cell culture system for adult mouse CMFs, and determine the effects of ISO treatment on the status of CDK1 expression, and proliferation and differentiation of CMFs. We also investigated the effect of genetic ablation of the atrial natriuretic peptide (ANP) receptor 1 (Npr1) on CMF proliferation and their sensitization to further treatment with catecholamines. Results showed that CDK1 expression, DNA synthesis and proliferation increase in adult mouse CMFs in response to ISO treatment. Analysis of cAMP levels in CMFs treated with ISO in the presence or absence of β adrenergic receptor (βAR) subtype specific blockers indicated that the β2AR response may be predominant in these cells. Consistent with this, the β2AR specific blocker ICI-118,551 and the non-selective βAR blocker carvedilol effectively blocked ISO induced CMF proliferation. ISO is also able to modulate pro-fibrotic gene expression program in CMFs by significantly increasing the transcription of genes such as collagen type 3 α1, connective tissue growth factor, α-smooth muscle actin and plasminogen activator inhibitor. Furthermore, pharmacological inhibition of CDK1 using RO-3306 effectively decreased CMF proliferation. In contrast, genetic ablation of both copies of Npr1 significantly increased CMF proliferation compared to wild type CMFs. Unexpectedly, CMFs lacking either one or both copies of Npr1 receptor revealed blunted mitogenic responses to ISO treatment compared to wild type cells. Collectively, these results suggest that CDK1 levels may be increased in response to the over-activated adrenergic signalling seen in heart failure. As such, it may present a possible drug target for inhibiting the expression of pro-fibrotic genes and CF proliferation. In addition, β2AR specific blockers, ANP and other synthetic Npr1 ligands may be effective in amelioration of excessive cardiac fibrosis commonly seen with many cardiovascular diseases.