Clin Res Cardiol (2021) DOI DOI https://doi.org/10.1007/s00392-021-01843-w

Human hypertrophic cardiomyopathy is associated with sarcomeric protein hypophosphorylation and oxidation.

R. Hassoun1, H. Budde1, P. Steinwascher1, M. Herwig2, M. Tangos1, D. Kolijn2, Á. Kovács2, A. Mügge1, K. Jaquet1, N. Hamdani2, for the study group: AG13

1Molecular and Experimental Cardiology, Ruhr University Bochum, St-Joseph Hospital, Bochum; 2Molecular and Experimental Cardiology, Physiology, Ruhr University Bochum, St-Joseph Hospital, Bochum;

Introduction Familial Hypertrophic Cardiomyopathy (FHC), an inherited disorder of the myocardium with a prevalence of 1:500 is associated with pronounced hypertrophy of the left ventricle and diastolic dysfunction. The main target genes encode myosin heavy chain and myosin binding protein C and among others TNNI3 is also affected. It encodes the inhibitory subunit (cTnI) of cardiac troponin which regulates muscle contraction. One cTnI- variant associated with FHC is p.cTnIR145G. Methods and results At first we studied the effect of β-adrenergic stimulation using isoproterenol (ISO) and β2-adrenoceptor agonist ICI 118,551 (ICI) or β1-adrenoceptor agonist CGP-20712A (CGP) on the global phosphorylation of TnI and also on sites specific phosphorylation of TnI at position serine 23/24 in adult rat cardiomyocytes using adenovirus-driven expression of human cTnI-wild type (WT) and cTnI-R145G. Our data showed that ISO, ISO / ICI and ISO/CGP treatment increases TnI phosphorylation at serine 23/24 in WT expressing cells and p.cTnI-R145G though to a different extent. ISO/CGP treatment reduces significantly the phosphorylation at S23/24 in p.cTnI-R145G compared to WT. Global TnI phosphorylation using Pro Qdiamond was unchanged between the two groups, however again upon ISO/CGP treatment a significantly lower raise in phosphorylation of  both WT and p.cTnI-R145G is observed as with ISO or ISO/ICI. The results indicate that p.cTnI-R145G expression influences the phosphorylation of cTnI in response to β-adrenergic stimulation dependent on the receptor subtype. In addition, we investigated the interplay TnI PKA phosphorylation and oxidative stress and thereby the cardiomyocyte function. TnI phosphorylation at serine 23/24 is increased upon treatment with PKA using either isolated recombinant protein TnI or human hypertrophic cardiomyopathy (HCM) tissues. This phosphorylation was then reduced after treatment with oxidized glutathione enzyme in both recombinant protein TnI and in human HCM tissues suggesting reduced affinity of PKA to serine 23/24 in the presence of TnI oxidation. In human single skinned cardiomyocyte from HCM patients, PKA reduced Ca2+ sensitivity of force production, which was high compared to non-failing human cardiomyocyte. A further significant increase was observed upon subsequent treatment of the cardiomyocytes with oxidized glutathione enzyme, which could be explained by reduced TnI phosphorylation and increased TnI oxidation. Human HCM tissues showed high level of oxidative stress parameters and lower level of reduced glutathione. Conclusion Oxidative stress-induced posttranslational modifications phosphorylation or oxidation of TnI modulated contractile function and thereby contribute to deterioration of cardiac function in HCM.

https://dgk.org/kongress_programme/jt2021/aP848.html