Changes in Cx43 and NaV1.5 Expression Precede the Occurrence of Substantial Fibrosis in Calcineurin-Induced Murine Cardiac Hypertrophy

Abstract

In mice, the calcium-dependent phosphatase calcineurin A (CnA) induces a transcriptional pathway leading to pathological cardiac hypertrophy. Interestingly, induction of CnA has been frequently noticed in human hypertrophic and failing hearts. Independently, the arrhythmia vulnerability of such hearts has been regularly associated with remodeling of parameters determining electrical conduction (expression level of connexin43 (Cx43) and NaV1.5, connective tissue architecture), for which the precise molecular basis and sequence of events is still unknown. Recently, we observed reduced Cx43 and NaV1.5 expression in 4-week old mouse hearts, overexpressing a constitutively active form of CnA (MHC-CnA model), but the order of events is still unknown. Therefore, three key parameters of conduction (Cx43, NaV1.5 and connective tissue expression) were characterized in MHC-CnA ventricles versus wild-type (WT) during postnatal development on a weekly basis. At postnatal week 1, CnA overexpression induced cardiac hypertrophy in MHC-CnA. Moreover, protein and RNA levels of both Cx43 and NaV1.5 were reduced by at least 50% as compared to WT. Cx43 immunoreactive signal was reduced at week 2 in MHC-CnA. At postnatal week 3, Cx43 was less phosphorylated and RNA level of Cx43 normalized to WT values, although the protein level was still reduced. Additionally, MHC-CnA hearts displayed substantial fibrosis relative to WT, which was accompanied by increased RNA levels for genes previously associated with fibrosis such as Col1a1, Col1a2, Col3a1, Tgfb1, Ctgf, Timp1 and microRNA miR-21. In MHC-CnA, reduction in Cx43 and NaV1.5 expression thus coincided with overexpression of CnA and hypertrophy development and preceded significant presence of fibrosis. At postnatal week 4 the alterations in conductional parameters observed in the MHC-CnA model lead to abnormal conduction and arrhythmias, similar to those observed in cardiac remodeling in heart failure patients. The MHC-CnA model, therefore, provides for a unique model to resolve the molecular origin of conductional remodeling in detail.

CTGF knockout does not affect cardiac hypertrophy and fibrosis formation upon chronic pressure overload

“…Moreover, we observed an increased expression of CTGF in control hearts after 16 weeks of pressure overload stimulation. This finding mirrors data from other studies that identified high CTGF expression levels under different cardiovascular pathological conditions as seen in patients and in experimental animal models, including applied pressure overload [8][9][10][11][12][13][14]. These studies, including our control hearts, show that CTGF is upregulated in the fibrotic heart suggesting the connotation of CTGF as a pro-fibrotic factor.…”

“…Increased levels of CTGF have been found in patients with heart failure, ischemia and coronary artery disease [8][9][10]. Similar elevations were observed in injury-induced animal models of cardiac hypertrophy and fibrosis [10][11][12][13][14]. The implication of CTGF in fibrosis development/maintenance, but also the close relationship with the pro-fibrotic cytokine transforming growth factor beta-1 (TGF-β1), is well documented [6,15].…”

“…RT-qPCR on ventricular tissue was performed using TaqMan Gene Expression Assays (Applied Biosystems by Life Technologies Corp., Carlsbad, CA, USA) as described earlier [14]. Relative mRNA expression levels were determined for CTGF, CCN3, CCN5, brain natriuretic peptide (BNP), Collagen 1a1, Collagen 1a2, Collagen 3a1, TGF-β1, plasminogen activator inhibitor-1 (PAI-1), heat shock protein 47 (Hsp47), Cx43 and sodium channel Na V 1.5 (all from Applied Biosystems by Life Technologies Corp.).…”

The Complex Network of the Brugada Syndrome

“…Nava described the complete syndrome in 1987/1998 (1)(2)(3) and by a simple analysis of the ECG demonstrated that the ECG pattern was due to a right ventricular conduction disturbance linked to a structural abnormality of the right ventricle. These evidencebased theories received a lot of heavy criticism (and other unfair definitions and boycott), because experts have insisted the syndrome was idiopathic, linked to ion channel abnormalities in a totally normal heart.…”

“…)-linked transcription factors. Indeed, work from our group and colleagues has associated common variation at the SCN5A-SCN10A locus with the risk for the Brugada syndrome compared with healthy controls(2,3). This locus is associated with a TBX3/5 binding site thought to regulate SCN5A transcription.It is therefore appealing to investigate how TBX3/5 may influence Connexin-43 expression and in turn influence the phenotype of the Brugada syndrome.Dr.…”