Diastolic dysfunction is general to all idiopathic dilated (IDCM) and hypertrophic cardiomyopathy (HCM) patients. Relaxation deficits may result from increased actin-myosin formation during diastole due to altered tropomyosin position, which blocks myosin binding to actin in the absence of Ca2+. We investigated whether ADP-stimulated force development (without Ca2+) can be used to reveal changes in actin-myosin blockade in human cardiomyopathy cardiomyocytes. Cardiac samples from HCM patients, harboring thick-filament (MYH7mutv MYBPC3mut) and thin-filament (TNNT2mutv TNNI3mut) mutations, and IDCM were compared with sarcomere mutation-negative HCM (HCMsmn) and nonfailing donors. Myofilament ADP sensitivity was higher in IDCM and HCM compared with donors, whereas it was lower for MYBPC3. Increased ADP sensitivity in IDCM, HCMsmn, and MYH7mut was caused by low phosphorylation of myofilament proteins, as it was normalized to donors by protein kinase A (PKA) treatment. Troponin exchange experiments in a TNNT2mut sample corrected the abnormal actin-myosin blockade. In MYBPC3trunc samples, ADP sensitivity highly correlated with cardiac myosin-binding protein-C (cMyBP-C) protein level. Incubation of cardiomyocytes with cMyBP-C antibody against the actin-binding N-terminal region reduced ADP sensitivity, indicative of cMyBP-C's role in actin-myosin regulation. In the presence of Ca2+, ADP increased myofilament force development and sarcomere stiffness. Enhanced sarcomere stiffness in sarcomere mutation-positive HCM samples was irrespective of the phosphorylation background. In conclusion, ADP-stimulated contraction can be used as a tool to study how protein phosphorylation and mutant proteins alter accessibility of myosin binding on actin. In the presence of Ca2+, pathologic [ADP] and low PKA-phosphorylation, high actin-myosin formation could contribute to the impaired myocardial relaxation observed in cardiomyopathies.

ADP, Cardiomyopathy, Hypertrophy, Idiopathic dilated, Three-state model
dx.doi.org/10.1073/pnas.1513843112, hdl.handle.net/1765/81538
Proceedings of the National Academy of Sciences of the United States of America
Department of Cardiology

Sequeira, V, Najafi, A, Wijnker, P.J.M, Dos Remedios, C.G, Michels, M, Kuster, D.W.D, & van der Velden, J. (2015). ADP-stimulated contraction: A predictor of thin-filament activation in cardiac disease. Proceedings of the National Academy of Sciences of the United States of America, 112(50), E7003–E7012. doi:10.1073/pnas.1513843112