Ger J M Stienen

VU University Medical Center, Amsterdamo, North Holland, Netherlands

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Publications (122)647.96 Total impact

  • G J M Stienen
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    ABSTRACT: Heart failure is a multi-factorial progressive disease in which eventually the contractile performance of the heart is insufficient to meet the demands of the body, even at rest. A distinction can be made on the basis of the cause of the disease in genetic and acquired heart failure and at the functional level between systolic and diastolic heart failure. Here the basic determinants of contractile function of myocardial cells will be reviewed and an attempt will be made to elucidate their role in the development of heart failure. The following topics are addressed: the tension generating capacity, passive tension, the rate of tension development, the rate of ATP utilisation, calcium sensitivity of tension development, phosphorylation of contractile proteins, length dependent activation and stretch activation. The reduction in contractile performance during systole can be attributed predominantly to a loss of cardiomyocytes (necrosis), myocyte disarray and a decrease in myofibrillar density all resulting in a reduction in the tension generating capacity and likely also to a mismatch between energy supply and demand of the myocardium. This leads to a decline in the ejection fraction of the heart. Diastolic dysfunction can be attributed to fibrosis and an increase in titin stiffness which result in an increase in stiffness of the ventricular wall and hampers the filling of the heart with blood during diastole. A large number of post translation modifications of regulatory sarcomeric proteins influence myocardial function by altering calcium sensitivity of tension development. It is still unclear whether in concert these influences are adaptive or maladaptive during the disease process.
    Journal of Muscle Research and Cell Motility 11/2014; · 1.36 Impact Factor
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    ABSTRACT: Cardiac troponin I (cTnI) is well known as a biomarker for the diagnosis of myocardial damage. However, because of its central role in the regulation of contraction and relaxation in heart muscle, cTnI may also be a potential target for the treatment of heart failure. Studies in rodent models of cardiac disease and human heart samples showed altered phosphorylation at various sites on cTnI (i.e. site-specific phosphorylation). This is caused by altered expression and/or activity of kinases and phosphatases during heart failure development. It is not known whether these (transient) alterations in cTnI phosphorylation are beneficial or detrimental. Knowledge of the effects of site-specific cTnI phosphorylation on cardiomyocyte contractility is therefore of utmost importance for the development of new therapeutic strategies in patients with heart failure. In this review we focus on the role of cTnI phosphorylation in the healthy heart upon activation of the beta-adrenergic receptor pathway (as occurs during increased stress and exercise) and as a modulator of the Frank-Starling mechanism. Moreover, we provide an overview of recent studies which aimed to reveal the functional consequences of changes in cTnI phosphorylation in cardiac disease.
    09/2014;
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    ABSTRACT: Several studies have indicated that diaphragm dysfunction develops in patients on mechanical ventilation (MV). Here, we tested the hypothesis that the contractility of sarcomeres - i.e. the smallest contractile unit in muscle - is affected in humans on MV. To this end, we compared diaphragm muscle fibers of 9 brain-dead organ donors (cases) who had been on MV for 26 ± 5 hours with diaphragm muscle fibers from 9 patients (controls) undergoing surgery for lung cancer who had been on MV less than 2 hours. In each diaphragm specimen we determined (1) muscle fiber cross-sectional area in cryosections by immunohistochemical methods, and (2) the contractile performance of permeabilized single muscle fibers by means of maximum specific force, kinetics of cross-bridge cycling by rate of tension redevelopment, myosin heavy chain content and concentration, and calcium sensitivity of force of slow-twitch and fast-twitch muscle fibers. In case subjects we noted no statistically significant decrease in outcomes compared to controls in slow-twitch or fast-twitch muscle fibers. These observations indicate that 26 hours of MV of humans is not invariably associated with changes in the contractile performance of sarcomeres in the diaphragm.
    American journal of physiology. Lung cellular and molecular physiology. 07/2014;
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    ABSTRACT: After lung transplantation, increased left ventricular (LV) filling can lead to LV failure, increasing the risk of post-operative complications and mortality. LV dysfunction in pulmonary arterial hypertension (PAH) is characterized by a reduced LV ejection fraction and impaired diastolic function.
    Journal of the American College of Cardiology 07/2014; 64(1):28-37. · 14.09 Impact Factor
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    ABSTRACT: The first mutation associated with hypertrophic cardiomyopathy (HCM) is the R403Q mutation in the gene encoding β-myosin heavy chain (β-MyHC). R403Q locates in the globular head of myosin (S1), responsible for interaction with actin, and thus motor function of myosin. Increased cross-bridge relaxation kinetics caused by the R403Q mutation might underlie increased energetic cost of tension generation; however, direct evidence is absent. Here we studied to what extent cross-bridge kinetics and energetics are related in single cardiac myofibrils and multicellular cardiac muscle strips of 3 HCM patients with the R403Q mutation and 9 sarcomere mutation-negative HCM patients (HCMsmn). Expression of R403Q was on average 41 ± 4 % of total MYH7 mRNA. Cross-bridge slow relaxation kinetics in single R403Q myofibrils was significantly higher (P < 0.0001) compared to HCMsmn myofibrils (0.47 ± 0.02 and 0.30 ± 0.02 s(-1), respectively). Moreover, compared to HCMsmn tension cost was significantly higher in the muscle strips of the three R403Q patients (2.93 ± 0.25 and 1.78 ± 0.10 μmol L s(-1) kN(-1) m(-2), respectively) which showed a positive linear correlation with relaxation kinetics in the corresponding myofibril preparations. This correlation suggests that faster cross-bridge relaxation kinetics results in an increase in energetic cost of tension generation in human HCM with the R403Q mutation compared to HCMsmn. Therefore, increased tension cost might contribute to HCM disease in patients carrying the R403Q mutation. This article is protected by copyright. All rights reserved.
    The Journal of Physiology 06/2014; · 4.38 Impact Factor
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    ABSTRACT: 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 energetic cost of contraction in human HCM disease development.
    Cardiovascular research. 05/2014;
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    American Journal of Respiratory and Critical Care Medicine 04/2014; 189(7):863-5. · 11.04 Impact Factor
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    ABSTRACT: Frank-Starling's law reflects the ability of the heart to adjust the force of its contraction to changes in ventricular filling, a property based on length-dependent myofilament activation (LDA). The threonine at amino acid 143 of cardiac troponin I (cTnI) is prerequisite for the length-dependent increase in Ca(2+)-sensitivity. Thr143 is a known target of protein kinase C (PKC) whose activity is increased in cardiac disease. Thr143 phosphorylation may modulate length-dependent myofilament activation in failing hearts. Therefore, we investigated if pseudo-phosphorylation at Thr143 modulates length-dependence of force using troponin exchange experiments in human cardiomyocytes. In addition, we studied effects of protein kinase A (PKA)-mediated cTnI phosphorylation at Ser23/24, which has been reported to modulate LDA. Isometric force was measured at various Ca(2+)-concentrations in membrane-permeabilized cardiomyocytes exchanged with recombinant wild-type (Wt) troponin or troponin mutated at the PKC site Thr143, or Ser23/24 into aspartic acid (D) or alanine (A) to mimic phosphorylation and dephosphorylation, respectively. In troponin-exchanged donor cardiomyocytes experiments were repeated after incubation with exogenous PKA. Pseudo-phosphorylation of Thr143 increased myofilament Ca(2+)-sensitivity compared to Wt without affecting LDA in failing and donor cardiomyocytes. Subsequent PKA treatment enhanced the length-dependent shift in Ca(2+)-sensitivity after Wt and 143D exchange. Exchange with Ser23/24 variants demonstrated that pseudo-phosphorylation of both Ser23 and Ser24 is needed to enhance the length-dependent increase in Ca(2+)-sensitivity. cTnI pseudo-phosphorylation did not alter length-dependent changes in maximal force. Thus phosphorylation at Thr143 enhances myofilament Ca(2+)-sensitivity without affecting LDA, while Ser23/24 bisphosphorylation is needed to enhance the length-dependent increase in myofilament Ca(2+)-sensitivity.
    AJP Heart and Circulatory Physiology 02/2014; · 4.01 Impact Factor
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    ABSTRACT: Protein kinase C (PKC)-mediated phosphorylation of troponin I (cTnI) at Ser42/44 is increased in heart failure. While studies in rodents demonstrated that PKC-mediated Ser42/44 phosphorylation decreases maximal force and ATPase activity, PKC incubation of human cardiomyocytes did not affect maximal force. We investigated whether Ser42/44 pseudo-phosphorylation affects force development and ATPase activity using troponin exchange in human myocardium. Additionally, we studied if pseudo-phosphorylated Ser42/44 modulates length-dependent activation of force, which is regulated by protein kinase A (PKA)-mediated cTnI-Ser23/24 phosphorylation. Isometric force was measured in membrane-permeabilized cardiomyocytes exchanged with human recombinant wild-type troponin or troponin mutated at Ser42/44 or Ser23/24 into aspartic acid (D) or alanine (A) to mimic phosphorylation and dephosphorylation, respectively. In troponin-exchanged donor cardiomyocytes experiments were repeated after PKA incubation. ATPase activity was measured in troponin-exchanged cardiac muscle strips. Compared to wild-type, 42D/44D decreased Ca2+-sensitivity without affecting maximal force in failing and donor cardiomyocytes. In donor myocardium, 42D/44D did not affect maximal ATPase activity or tension cost. Interestingly, 42D/44D blunted the length-dependent increase in Ca2+-sensitivity induced upon PKA-mediated phosphorylation. Since the drop in Ca2+-sensitivity at physiological Ca2+-concentrations is relatively large phosphorylation of Ser42/44 may result in a decrease of force and associated ATP utilization in the human heart.
    Archives of Biochemistry and Biophysics 01/2014; · 3.37 Impact Factor
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    ABSTRACT: Right ventricular (RV) diastolic function is impaired in patients with pulmonary arterial hypertension (PAH). Our previous study showed that elevated cardiomyocyte stiffness and myofilament Ca(2+) sensitivity underlie diastolic dysfunction in PAH. This study investigates protein modifications contributing to cellular diastolic dysfunction in PAH.
    Journal of the American Heart Association. 01/2014; 3(3).
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    ABSTRACT: Mutations in the MYBPC3 gene, encoding cardiac myosin binding protein C (cMyBP-C) are frequent causes of hypertrophic cardiomyopathy (HCM). Previously, we have presented evidence for reduced cMyBP-C expression (haploinsufficiency), in patients with a truncation mutation in MYBPC3. In mice, lacking cMyBP-C cross-bridge kinetics was accelerated. In this study, we investigated whether cross-bridge kinetics was altered in myectomy samples from HCM patients harboring heterozygous MYBPC3 mutations (MYBPC3mut). Isometric force and the rate of force redevelopment (k tr) at different activating Ca(2+) concentrations were measured in mechanically isolated Triton-permeabilized cardiomyocytes from MYBPC3mut (n = 18) and donor (n = 7) tissue. Furthermore, the stretch activation response of cardiomyocytes was measured in tissue from eight MYBPC3mut patients and five donors to assess the rate of initial force relaxation (k 1) and the rate and magnitude of the transient increase in force (k 2 and P 3, respectively) after a rapid stretch. Maximal force development of the cardiomyocytes was reduced in MYBPC3mut (24.5 ± 2.3 kN/m(2)) compared to donor (34.9 ± 1.6 kN/m(2)). The rates of force redevelopment in MYBPC3mut and donor over a range of Ca(2+) concentrations were similar (k tr at maximal activation: 0.63 ± 0.03 and 0.75 ± 0.09 s(-1), respectively). Moreover, the stretch activation parameters did not differ significantly between MYBPC3mut and donor (k 1: 8.5±0.5 and 8.8 ± 0.4 s(-1); k 2: 0.77 ± 0.06 and 0.74 ± 0.09 s(-1); P 3: 0.08 ± 0.01 and 0.09 ± 0.01, respectively). Incubation with protein kinase A accelerated k 1 in MYBPC3mut and donor to a similar extent. Our experiments indicate that, at the cMyBP-C expression levels in this patient group (63 ± 6 % relative to donors), cross-bridge kinetics are preserved and that the depressed maximal force development is not explained by perturbation of cross-bridge kinetics.
    Pflügers Archiv - European Journal of Physiology 11/2013; · 4.87 Impact Factor
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    ABSTRACT: Hypertrophic cardiomyopathy (HCM) is an autosomal dominant heart disease mostly due to mutations in genes encoding sarcomeric proteins. HCM is characterised by asymmetric hypertrophy of the left ventricle (LV) in the absence of another cardiac or systemic disease. At present it lacks specific treatment to prevent or reverse cardiac dysfunction and hypertrophy in mutation carriers and HCM patients. Previous studies have indicated that sarcomere mutations increase energetic costs of cardiac contraction and cause myocardial dysfunction and hypertrophy. By using a translational approach, we aim to determine to what extent disturbances of myocardial energy metabolism underlie disease progression in HCM. Hypertrophic obstructive cardiomyopathy (HOCM) patients and aortic valve stenosis (AVS) patients will undergo a positron emission tomography (PET) with acetate and cardiovascular magnetic resonance imaging (CMR) with tissue tagging before and 4 months after myectomy surgery or aortic valve replacement + septal biopsy. Myectomy tissue or septal biopsy will be used to determine efficiency of sarcomere contraction in-vitro, and results will be compared with in-vivo cardiac performance. Healthy subjects and non-hypertrophic HCM mutation carriers will serve as a control group. Our study will reveal whether perturbations in cardiac energetics deteriorate during disease progression in HCM and whether these changes are attributed to cardiac remodelling or the presence of a sarcomere mutation per se. In-vitro studies in hypertrophied cardiac muscle from HOCM and AVS patients will establish whether sarcomere mutations increase ATP consumption of sarcomeres in human myocardium. Our follow-up imaging study in HOCM and AVS patients will reveal whether impaired cardiac energetics are restored by cardiac surgery.
    Netherlands heart journal: monthly journal of the Netherlands Society of Cardiology and the Netherlands Heart Foundation 10/2013; · 1.41 Impact Factor
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    ABSTRACT: The role of right ventricular (RV) diastolic stiffness in pulmonary arterial hypertension (PAH) is not well-established. Therefore, we investigated the presence and possible underlying mechanisms of RV diastolic stiffness in PAH-patients. Single-beat RV pressure-volume analyses were performed in 21 PAH-patients and 7 controls to study RV diastolic stiffness. Data presented as mean±SEM. RV diastolic stiffness (β) was significantly increased in PAH-patients (PAH: 0.050±0.005 vs. control: 0.029±0.003; p<0.05) and closely associated to disease severity. Subsequently, we searched for possible underlying mechanisms, using RV tissue of PAH-patients undergoing heart-lung transplantation and non-failing donors. Histological analyses revealed increased cardiomyocyte cross-sectional areas (PAH: 453±31 vs. control: 218±21 μm(2); p<0.001), indicating RV hypertrophy. In addition, the amount of RV fibrosis was enhanced in PAH tissue (PAH: 9.6±0.7 vs. control: 7.2±0.6%; p<0.01). To investigate the contribution of stiffening of the sarcomere (the contractile apparatus of RV cardiomyocytes) to RV diastolic stiffness, we isolated and membrane-permeabilized single RV cardiomyocytes. Passive tension at different sarcomere lengths was significantly higher in PAH compared to controls (+200%; pinteraction<0.001), indicating stiffening of RV sarcomeres. An important regulator of sarcomeric stiffening is the sarcomeric protein titin. Therefore, we investigated titin isoform composition and phosphorylation. No alterations were observed in titin isoform composition (N2BA/N2B ratio PAH: 0.78±0.07 vs. control 0.91±0.08), but titin phosphorylation in RV-tissue of PAH-patients was significantly reduced (PAH: 0.16±0.01 vs. control 0.20±0.01 a.u.;p<0.05). RV diastolic stiffness is significantly increased in PAH-patients, with important contributions from increased collagen and intrinsic stiffening of the RV cardiomyocyte sarcomeres.
    Circulation 09/2013; · 15.20 Impact Factor
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    ABSTRACT: Nebulin-a giant sarcomeric protein-plays a pivotal role in skeletal muscle contractility by specifying thin filament length and function. Although mutations in the gene encoding nebulin (NEB) are a frequent cause of nemaline myopathy, the most common non-dystrophic congenital myopathy, the mechanisms by which mutations in NEB cause muscle weakness remain largely unknown. To better understand these mechanisms, we have generated a mouse model in which Neb exon 55 is deleted (Neb(ΔExon55)) to replicate a founder mutation seen frequently in patients with nemaline myopathy with Ashkenazi Jewish heritage. Neb(ΔExon55) mice are born close to Mendelian ratios, but show growth retardation after birth. Electron microscopy studies show nemaline bodies-a hallmark feature of nemaline myopathy-in muscle fibres from Neb(ΔExon55) mice. Western blotting studies with nebulin-specific antibodies reveal reduced nebulin levels in muscle from Neb(ΔExon55) mice, and immunofluorescence confocal microscopy studies with tropomodulin antibodies and phalloidin reveal that thin filament length is significantly reduced. In line with reduced thin filament length, the maximal force generating capacity of permeabilized muscle fibres and single myofibrils is reduced in Neb(ΔExon55) mice with a more pronounced reduction at longer sarcomere lengths. Finally, in Neb(ΔExon55) mice the regulation of contraction is impaired, as evidenced by marked changes in crossbridge cycling kinetics and by a reduction of the calcium sensitivity of force generation. A novel drug that facilitates calcium binding to the thin filament significantly augmented the calcium sensitivity of submaximal force to levels that exceed those observed in untreated control muscle. In conclusion, we have characterized the first nebulin-based nemaline myopathy model, which recapitulates important features of the phenotype observed in patients harbouring this particular mutation, and which has severe muscle weakness caused by thin filament dysfunction.
    Brain 05/2013; · 10.23 Impact Factor
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    ABSTRACT: AIMS: 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.Methods and ResultsCardiac 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), 8 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&emsp14;kN/m(2)) and HCMsmn (26±3&emsp14;kN/m(2)) compared to donor (36±2&emsp14;kN/m(2)). Cardiomyocyte remodelling was more severe in HCMmut compared to HCMsmn based on significantly lower myofibrillar density (49±2 versus 63±5%) and significantly higher cardiomyocyte area (915±15 versus 612±11&emsp14;µm(2)). Low Fmax in MYBPC3mut, TNNI3mut, HCMsmn and LVHao was normalized to donor values after correction for myofibrillar density. However, Fmax was significantly lower in MYH7mut, TPM1mut and TNNT2mut even after correction for myofibrillar density. In accordance, measurements in single myofibrils showed very low Fmax in MYH7mut, TPM1mut and TNNT2mut compared to donor (respectively, 73±3, 70±7, 83±6 and 113±5&emsp14;kN/m(2)). In addition, force was lower in MYH7mut cardiomyocytes compared to MYBPC3mut, HCMsmn and donor at submaximal [Ca(2+)]. CONCLUSIONS: Low cardiomyocyte Fmax in HCM patients is largely explained by hypertrophy and reduced myofibrillar density. MYH7 mutations reduce force generating capacity of sarcomeres at maximal and submaximal [Ca(2+)]. These hypocontractile sarcomeres may represent the primary abnormality in patients with MYH7 mutations.
    Cardiovascular Research 05/2013; · 5.81 Impact Factor
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    ABSTRACT: BACKGROUND: Nemaline myopathy-the most common non-dystrophic congenital myopathy-is caused by mutations in thin filament genes, of which the nebulin gene is the most frequently affected one. The nebulin gene codes for the giant sarcomeric protein nebulin, which plays a crucial role in skeletal muscle contractile performance. Muscle weakness is a hallmark feature of nemaline myopathy patients with nebulin mutations, and is caused by changes in contractile protein function, including a lower calcium-sensitivity of force generation. To date no therapy exists to treat muscle weakness in nemaline myopathy. Here, we studied the ability of the novel fast skeletal muscle troponin activator, CK-2066260, to augment force generation at submaximal calcium levels in muscle cells from nemaline myopathy patients with nebulin mutations. METHODS: Contractile protein function was determined in permeabilised muscle cells isolated from frozen patient biopsies. The effect of 5 μM CK-2066260 on force production was assessed. RESULTS: Nebulin protein concentrations were severely reduced in muscle cells from these patients compared to controls, while myofibrillar ultrastructure was largely preserved. Both maximal active tension and the calcium-sensitivity of force generation were lower in patients compared to controls. Importantly, CK-2066260 greatly increased the calcium-sensitivity of force generation-without affecting the cooperativity of activation-in patients to levels that exceed those observed in untreated control muscle. CONCLUSIONS: Fast skeletal troponin activation is a therapeutic mechanism to augment contractile protein function in nemaline myopathy patients with nebulin mutations and with other neuromuscular diseases.
    Journal of Medical Genetics 04/2013; · 5.70 Impact Factor
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    ABSTRACT: Rationale: High myofilament Ca(2+)-sensitivity has been proposed as trigger of disease pathogenesis in familial hypertrophic cardiomyopathy (HCM) based on in vitro and transgenic mice studies. However, myofilament Ca(2+)-sensitivity depends on protein phosphorylation and muscle length and at present data in human are scarce. Objective:To investigate if high myofilament Ca(2+)-sensitivity and perturbed length-dependent activation is characteristic for human HCM with mutations in thick and thin filament proteins. Methods and Results:: Cardiac samples from HCM patients harboring mutations in genes encoding thick (MYH7, MYBPC3) and thin (TNNT2, TNNI3, TPM1) filament proteins were compared with sarcomere mutation-negative HCM (HCMsmn) and non-failing donors. Cardiomyocyte force measurements showed higher myofilament Ca(2+)-sensitivity in all HCM samples and low phosphorylation of protein kinase A (PKA)-targets compared to donors. After exogenous PKA treatment, myofilamentCa(2+)-sensitivity was either similar (MYBPC3mut, TPM1mut, HCMsmn), higher (MYH7mut, TNNT2mut) or even significantly lower (TNNI3mut) compared to 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 HCMsmn and HCM with truncating MYBPC3 mutations, but not in HCM with missense mutations. Replacement of mutant by wild-type troponin in TNNT2mu and TNNI3mut corrected length-dependent activation to donor values. Conclusions: High myofilament Ca(2+)-sensitivity is a common characteristic of human HCM and partly reflects hypophosphorylation of PKA-targets compared to donors. Length-dependent sarcomere activation is perturbed by missense mutations, possibly via post-translational modifications other than PKA-hypophosphorylation or altered protein-protein interactions, and represents a common pathomechanism in HCM.
    Circulation Research 03/2013; · 11.86 Impact Factor
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    ABSTRACT: Nemaline myopathy is the most common congenital skeletal muscle disease, and mutations in the nebulin gene account for 50% of all cases. Recent studies suggest that the disease severity might be related to the nebulin expression levels. Considering that mutations in the nebulin gene are typically recessive, one would expect that a single functional nebulin allele would maintain nebulin protein expression which would result in preserved skeletal muscle function. We investigated skeletal muscle function of heterozygous nebulin knock-out (i.e., nebulin +/À) mice using a multidisciplinary approach including protein and gene expression analysis and combined in vivo and in vitro force measurements. Skeletal muscle anatomy and energy metabolism were studied strictly non-invasively using magnetic resonance imaging and 31P-magnetic resonance spectroscopy. Maximal force production was reduced by around 16% in isolated muscle of nebulin +/À mice while in vivo force generating capacity was preserved. Muscle weakness was associated with a shift toward a slower proteomic phenotype, but was not related to nebulin protein deficiency or to an impaired energy metabolism. Further studies would be warranted in order to determine the mechanisms leading to a mild skeletal muscle phenotype resulting from the expression of a single nebulin allele.
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    ABSTRACT: Nemaline myopathy is the most common congenital skeletal muscle disease, and mutations in the nebulin gene account for 50% of all cases. Recent studies suggest that the disease severity might be related to the nebulin expression levels. Considering that mutations in the nebulin gene are typically recessive, one would expect that a single functional nebulin allele would maintain nebulin protein expression which would result in preserved skeletal muscle function. We investigated skeletal muscle function of heterozygous nebulin knock-out (i.e., nebulin(+/-)) mice using a multidisciplinary approach including protein and gene expression analysis and combined in vivo and in vitro force measurements. Skeletal muscle anatomy and energy metabolism were studied strictly non-invasively using magnetic resonance imaging and 31P-magnetic resonance spectroscopy. Maximal force production was reduced by around 16% in isolated muscle of nebulin(+/-) mice while in vivo force generating capacity was preserved. Muscle weakness was associated with a shift toward a slower proteomic phenotype, but was not related to nebulin protein deficiency or to an impaired energy metabolism. Further studies would be warranted in order to determine the mechanisms leading to a mild skeletal muscle phenotype resulting from the expression of a single nebulin allele.
    Neuromuscular Disorders 01/2013; · 3.46 Impact Factor
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    ABSTRACT: OBJECTIVE: To investigate whether sarcomeric dysfunction contributes to muscle weakness in facioscapulohumeral muscular dystrophy (FSHD). METHODS: Sarcomeric function was evaluated by contractile studies on demembranated single muscle fibers obtained from quadriceps muscle biopsies of 4 patients with FSHD and 4 healthy controls. The sarcomere length dependency of force was determined together with measurements of thin filament length using immunofluorescence confocal scanning laser microscopy. X-ray diffraction techniques were used to study myofilament lattice spacing. RESULTS: FSHD muscle fibers produced only 70% of active force compared to healthy controls, a reduction which was exclusive to type II muscle fibers. Changes in force were not due to changes in thin filament length or sarcomere length. Passive force was increased 5- to 12-fold in both fiber types, with increased calcium sensitivity of force generation and decreased myofilament lattice spacing, indicating compensation by the sarcomeric protein titin. CONCLUSIONS: We have demonstrated a reduction in sarcomeric force in type II FSHD muscle fibers, and suggest compensatory mechanisms through titin stiffening. Based on these findings, we propose that sarcomeric dysfunction plays a critical role in the development of muscle weakness in FSHD.
    Neurology 01/2013; · 8.30 Impact Factor

Publication Stats

2k Citations
647.96 Total Impact Points

Institutions

  • 2003–2014
    • VU University Medical Center
      • • Institute for Cardiovascular Research (ICaR-VU)
      • • Department of Physiology
      • • Department of Anesthesiology
      Amsterdamo, North Holland, Netherlands
  • 2002–2014
    • VU University Amsterdam
      • • Department of Physics and Astronomy
      • • Institute for Cardiovascular Research VU
      • • Laboratory for Physiology
      Amsterdamo, North Holland, Netherlands
  • 2013
    • Hannover Medical School
      Hanover, Lower Saxony, Germany
  • 2010–2013
    • University of Padova
      • Department of Biomedical Sciences - DSB
      Padua, Veneto, Italy
  • 2009
    • The University of Arizona
      • Department of Molecular and Cellular Biology
      Tucson, AZ, United States
  • 2008
    • Istituto di Ricovero e Cura a Carattere Scientifico San Raffaele Pisana
      Roma, Latium, Italy
  • 2000–2008
    • University of Debrecen
      • • Department of Clinical Pharmacology
      • • Institute of Cardiology
      • • Department of Physiology
      Debrecen, Hajdu-Bihar, Hungary
  • 2007
    • Erasmus Universiteit Rotterdam
      • Department of Cardiology
      Rotterdam, South Holland, Netherlands
    • University of Illinois at Chicago
      • Department of Physiology and Biophysics (Chicago)
      Chicago, IL, United States
  • 1990–2007
    • University of Amsterdam
      • • Department of Plant Physiology
      • • Department of Cardiology
      • • Laboratory for Physiology
      Amsterdamo, North Holland, Netherlands
  • 2006
    • University of Florence
      • Dipartimento di Medicina Sperimentale e Clinica
      Florence, Tuscany, Italy
  • 2004
    • University of Leuven
      • Division of Experimental Cardiology
      Louvain, Flanders, Belgium