👤 Corrado Poggesi

Approximately 40% of hypertrophic cardiomyopathy (HCM) mutations are linked to the sarcomere protein cardiac myosin binding protein-C (cMyBP-C). These mutations are either classified as missense mutations or truncation mutations. One mutation whose nature has been inconsistently reported in the literature is the MYBPC3-c.772G > A mutation. Using patient-derived human induced pluripotent stem cells differentiated to cardiomyocytes (hiPSC-CMs), we have performed a mechanistic study of the structure-function relationship for this MYBPC3-c.772G > A mutation versus a mutation corrected, isogenic cell line. Our results confirm that this mutation leads to exon skipping and mRNA truncation that ultimately suggests ∼20% less cMyBP-C protein (i.e., haploinsufficiency). This, in turn, results in increased myosin recruitment and accelerated myofibril cycling kinetics. Our mechanistic studies suggest that faster ADP release from myosin is a primary cause of accelerated myofibril cross-bridge cycling due to this mutation. Additionally, the reduction in force generating heads expected from faster ADP release during isometric contractions is outweighed by a cMyBP-C phosphorylation mediated increase in myosin recruitment that leads to a net increase of myofibril force, primarily at submaximal calcium activations. These results match well with our previous report on contractile properties from myectomy samples of the patients from whom the hiPSC-CMs were generated, demonstrating that these cell lines are a good model to study this pathological mutation and extends our understanding of the mechanisms of altered contractile properties of this HCM MYBPC3-c.772G > A mutation. Show less

The 2 sarcomere genes most commonly associated with hypertrophic cardiomyopathy (HCM), MYBPC3 (myosin-binding protein C3) and MYH7 (β-myosin heavy chain), are indistinguishable at presentation, and genotype-phenotype correlations have been elusive. Based on molecular and pathophysiological differences, however, it is plausible to hypothesize a different behavior in myocardial performance, impacting lifetime changes in left ventricular (LV) function. We reviewed the initial and final echocardiograms of 402 consecutive HCM patients with pathogenic or likely pathogenic MYBPC3 (n=251) or MYH7 (n=151) mutations, followed over 9±8 years. At presentation, MYBPC3 patients were less frequently obstructive (15% versus 26%; MYBPC3-related HCM showed increased long-term prevalence of systolic dysfunction compared with MYH7, in spite of similar outcome. Such observations suggest different pathophysiology of clinical progression in the 2 subsets and may prove relevant for understanding of genotype-phenotype correlations in HCM. Show less

Genes associated with hypertrophic cardiomyopathy (HC) are not uniformly expressed in the atrial myocardium. Whether this may impact susceptibility to atrial fibrillation (AF) is unresolved. To analyze the prevalence and clinical correlates of AF in relation to genotype in a large HC cohort, prevalence and clinical profile of AF were assessed in 237 patients with HC, followed for 14 ± 10 years. Patients were divided into 3 genetic subgroups: (1) MYBPC3 (58%), (2) MYH7 (28%), and (3) "other genotypes" (14%; comprising TNNT2, TNNI3, TPM1, MYL2, complex genotypes, Z-line, and E-C coupling genes). Left atrial size was similar in the 3 subsets. AF occurred in 74 patients with HC (31%), with no difference among groups (31% in MYBPC3, 37% in MYH7 and 18% in other genotypes, p = 0.15), paroxysmal/persistent AF (12%, 18%, and 12%, respectively; p = 0.53), paroxysmal/persistent evolved to permanent (12%, 12%, and 3%, p = 0.36) or permanent AF (7%, 7%, and 3%, p = 0.82). Age at AF onset was younger in the group with other genotypes (37 ± 10 years) compared to the first 2 groups (53 ± 14 and 51 ± 17, respectively; p = 0.05) because of early onset associated with complex genotypes and a specific JPH2 mutation associated with abnormal intracellular calcium handling. At multivariate analysis, independent predictors of AF were atrial diameter (p ≤0.05) and age at diagnosis (p = 0.09), but not genetic subtype (p = 0.35). In conclusion, in patients with HC, genetic testing cannot be used in clinical decision making with regard to management strategies for AF. Genotype is not predictive of onset or severity of AF, which appears rather driven by hemodynamic determinants of atrial dilatation. Exceptions are represented by rare genes suggesting specific molecular pathways for AF in genetic cardiomyopathies. Show less

Disease mechanisms regarding hypertrophic cardiomyopathy (HCM) are largely unknown and disease onset varies. Sarcomere mutations might induce energy depletion for which until now there is no direct evidence at sarcomere level in human HCM. This study investigated if mutations in genes encoding myosin-binding protein C (MYBPC3) and myosin heavy chain (MYH7) underlie changes in the energetic cost of contraction in the development of human HCM disease. Energetic cost of contraction was studied in vitro by measurements of force development and ATPase activity in cardiac muscle strips from 26 manifest HCM patients (11 MYBPC3mut, 9 MYH7mut, and 6 sarcomere mutation-negative, HCMsmn). In addition, in vivo, the ratio between external work (EW) and myocardial oxygen consumption (MVO2) to obtain myocardial external efficiency (MEE) was determined in 28 pre-hypertrophic mutation carriers (14 MYBPC3mut and 14 MYH7mut) and 14 healthy controls using [(11)C]-acetate positron emission tomography and cardiovascular magnetic resonance imaging. Tension cost (TC), i.e. ATPase activity during force development, was higher in MYBPC3mut and MYH7mut compared with HCMsmn at saturating [Ca(2+)]. TC was also significantly higher in MYH7mut at submaximal, more physiological [Ca(2+)]. EW was significantly lower in both mutation carrier groups, while MVO2 did not differ. MEE was significantly lower in both mutation carrier groups compared with controls, showing the lowest efficiency in MYH7 mutation carriers. We provide direct evidence that sarcomere mutations perturb the energetic cost of cardiac contraction. Gene-specific severity of cardiac abnormalities may underlie differences in disease onset and suggests that early initiation of metabolic treatment may be beneficial, in particular, in MYH7 mutation carriers. Show less

Familial hypertrophic cardiomyopathy (HCM), frequently caused by sarcomeric gene mutations, is characterized by cellular dysfunction and asymmetric left-ventricular (LV) hypertrophy. We studied whether cellular dysfunction is due to an intrinsic sarcomere defect or cardiomyocyte remodelling. Cardiac samples from 43 sarcomere mutation-positive patients (HCMmut: mutations in thick (MYBPC3, MYH7) and thin (TPM1, TNNI3, TNNT2) myofilament genes) were compared with 14 sarcomere mutation-negative patients (HCMsmn), eight patients with secondary LV hypertrophy due to aortic stenosis (LVHao) and 13 donors. Force measurements in single membrane-permeabilized cardiomyocytes revealed significantly lower maximal force generating capacity (Fmax) in HCMmut (21 ± 1 kN/m²) and HCMsmn (26 ± 3 kN/m²) compared with donor (36 ± 2 kN/m²). Cardiomyocyte remodelling was more severe in HCMmut compared with HCMsmn based on significantly lower myofibril density (49 ± 2 vs. 63 ± 5%) and significantly higher cardiomyocyte area (915 ± 15 vs. 612 ± 11 μm²). Low Fmax in MYBPC3mut, TNNI3mut, HCMsmn, and LVHao was normalized to donor values after correction for myofibril density. However, Fmax was significantly lower in MYH7mut, TPM1mut, and TNNT2mut even after correction for myofibril density. In accordance, measurements in single myofibrils showed very low Fmax in MYH7mut, TPM1mut, and TNNT2mut compared with donor (respectively, 73 ± 3, 70 ± 7, 83 ± 6, and 113 ± 5 kN/m²). In addition, force was lower in MYH7mut cardiomyocytes compared with MYBPC3mut, HCMsmn, and donor at submaximal [Ca²⁺]. Low cardiomyocyte Fmax in HCM patients is largely explained by hypertrophy and reduced myofibril density. MYH7 mutations reduce force generating capacity of sarcomeres at maximal and submaximal [Ca²⁺]. These hypocontractile sarcomeres may represent the primary abnormality in patients with MYH7 mutations. Show less

High-myofilament Ca(2+) sensitivity has been proposed as a trigger of disease pathogenesis in familial hypertrophic cardiomyopathy (HCM) on the basis of in vitro and transgenic mice studies. However, myofilament Ca(2+) sensitivity depends on protein phosphorylation and muscle length, and at present, data in humans are scarce. To investigate whether high myofilament Ca(2+) sensitivity and perturbed length-dependent activation are characteristics for human HCM with mutations in thick and thin filament proteins. Cardiac samples from patients with HCM harboring mutations in genes encoding thick (MYH7, MYBPC3) and thin (TNNT2, TNNI3, TPM1) filament proteins were compared with sarcomere mutation-negative HCM and nonfailing donors. Cardiomyocyte force measurements showed higher myofilament Ca(2+) sensitivity in all HCM samples and low phosphorylation of protein kinase A (PKA) targets compared with donors. After exogenous PKA treatment, myofilament Ca(2+) sensitivity was similar (MYBPC3mut, TPM1mut, sarcomere mutation-negative HCM), higher (MYH7mut, TNNT2mut), or even significantly lower (TNNI3mut) compared with donors. Length-dependent activation was significantly smaller in all HCM than in donor samples. PKA treatment increased phosphorylation of PKA-targets in HCM myocardium and normalized length-dependent activation to donor values in sarcomere mutation-negative HCM and HCM with truncating MYBPC3 mutations but not in HCM with missense mutations. Replacement of mutant by wild-type troponin in TNNT2mut and TNNI3mut corrected length-dependent activation to donor values. High-myofilament Ca(2+) sensitivity is a common characteristic of human HCM and partly reflects hypophosphorylation of PKA targets compared with donors. Length-dependent sarcomere activation is perturbed by missense mutations, possibly via posttranslational modifications other than PKA hypophosphorylation or altered protein-protein interactions, and represents a common pathomechanism in HCM. Show less