Aims Expression of cardiac ankyrin repeat protein (CARP) is augmented in heart failure due to dilated or ischaemic cardiomyopathy. It is unclear whether CARP is upregulated in heart failure due to arrhythmogenic right ventricular cardiomyopathy (ARVC). In the present study, we investigated the expression pattern of CARP and the correlation between CARP and the well‐known heart failure marker pro‐atrial natriuretic peptide (proANP) in ARVC failing hearts. Methods and results Gene microarray analysis demonstrated increased CARP expression in ARVC failing hearts compared with non‐failing control hearts, which was further validated by real‐time RT–PCR, western blot, and ELISA at the mRNA and protein levels. Fractionation experiments revealed that the upregulation of CARP expression is restricted to the nuclei of residual cardiac cells in ARVC failing hearts. Regression analysis showed a positive correlation between CARP and proANP in ARVC failing hearts. Conclusion Augmented CARP expression may be a common molecular event in failing hearts regardless of cardiomyopathic aetiology. The upregulation of nuclear CARP expression and positive correlation between cardiac CARP and proANP suggests that CARP may be used as a genetic marker existing in the nuclei in contrast to proANP existing in the cytosol of cardiac cells in heart failure patients.

As proteins are the ultimate biological determinants of phenotype of disease, we screened altered proteins associated with heart failure due to arrhythmogenic right ventricular cardiomyopathy (ARVC) to identify biomarkers potential for rapid diagnosis of heart failure. By 2-dimensional gel electrophoresis and mass spectrometry, we identified five commonly altered proteins with more than 1.5 fold changes in eight ARVC failing hearts using eight non-failing hearts as reference. Noticeably, one of the altered proteins, heat shock protein 70 (HSP70), was increased by 1.64 fold in ARVC failing hearts compared with non-failing hearts. The increase of cardiac HSP70 was further validated by Western blot, immunochemistry, and enzyme-linked immunosorbent assay (ELISA) in failing hearts due to not only ARVC, but also dilated (DCM, n = 18) and ischemic cardiomyopathy (ICM, n = 8). Serum HSP70 was also observed to be significantly increased in heart failure patients derived from the three forms of cardiomyopathies. In addition, we observed hypoxia/serum depletion stimulation induced significantly elevation of intracellular and extracellular HSP70 in cultured neonatal rat cardiomyocytes. For the first time to our knowledge, we revealed and clearly demonstrated significant up-regulation of cardiac and serum HSP70 in ARVC heart failure patients. Our results indicate that elevated HSP70 is the common feature of heart failure due to ARVC, DCM, and ICM, which suggests that HSP70 may be used as a biomarker for the presence of heart failure due to cardiomyopathies of different etiologies and may hold diagnostic/prognostic potential in clinical practice.

Apolipoprotein D (Apo D) is reported to be in close association with developing and mature blood vessels, and involved in enhanced smooth muscle cell migration after injury. This study was designed to clarify the expression pattern of Apo D and the possibility of Apo D as a new marker in human end-stage heart failure. Individual RNA samples obtained from independent left ventricular tissue of six heart failure patients derived from cardiomyopathies of different aetiologies during cardiac transplantation and six non-failing control subjects were hybridized to the gene microarray containing, in total, 35 000 well-characterized Homo sapiens genes. Apo D was one of the highly expressed genes (3.3-fold upregulated) detected by microarray, which was further confirmed by quantitative real-time reverse transcriptase polymerase chain reaction (RT-PCR) (5.88-fold upregulated) in failing hearts compared with non-failing hearts. Both Western blotting and immunohistochemistry analyses also demonstrated the higher levels of Apo D protein in failing hearts. Importantly, we observed elevated levels of plasma Apo D in heart failure patients compared with non-failing control subjects. We demonstrated, for the first time to our knowledge, that Apo D was highly expressed in the mRNA and protein levels in human failing hearts compared with non-failing hearts. Furthermore, our finding of elevated plasma Apo D levels in patients with heart failure provides clues that Apo D may act not only as a cardiac molecular marker but also as a circulating biomarker in patients with heart failure.