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Cardiomyopathies are complex heart muscle diseases that can be inherited or acquired. Dilated cardiomyopathy can result from mutations in LMNA, encoding the nuclear intermediate filament proteins lamin A/C. Some LMNA mutations lead to accumulation of the lamin A precursor, prelamin A, which is disease causing in a number of tissues, yet its impact upon the heart is unknown. Here, we discovered myocardial prelamin A accumulation occurred in a case of dilated cardiomyopathy, and we show that a potentially novel mouse model of cardiac-specific prelamin A accumulation exhibited a phenotype consistent with inflammatory cardiomyopathy, which we observed to be similar to HIV-associated cardiomyopathy, an acquired disease state. Numerous HIV protease therapies are known to inhibit ZMPSTE24, the enzyme responsible for prelamin A processing, and we confirmed that accumulation of prelamin A occurred in HIV+ patient cardiac biopsies. These findings (a) confirm a unifying pathological role for prelamin A common to genetic and acquired cardiomyopathies; (b) have implications for the management of HIV patients with cardiac disease, suggesting protease inhibitors should be replaced with alternative therapies (i.e., nonnucleoside reverse transcriptase inhibitors); and (c) suggest that targeting inflammation may be a useful treatment strategy for certain forms of inherited cardiomyopathy.

Peripartum cardiomyopathy (PPCM) and dilated cardiomyopathy (DCM) show similarities in clinical presentation. However, although DCM patients do not recover and slowly deteriorate further, PPCM patients show either a fast cardiac deterioration or complete recovery. The aim of this study was to assess if underlying cellular changes can explain the clinical similarities and differences in the two diseases. We, therefore, assessed sarcomeric protein expression, modification, titin isoform shift, and contractile behavior of cardiomyocytes in heart tissue of PPCM and DCM patients and compared these with nonfailing controls. Heart samples from ischemic heart disease (ISHD) patients served as heart failure control samples. Passive force was only increased in PPCM samples compared with controls, whereas PPCM, DCM, and ISHD samples all showed increased myofilament Ca 2+ sensitivity. Length-dependent activation was significantly impaired in PPCM compared with controls, no impairment was observed in ISHD samples, and DCM samples showed an intermediate response. Contractile impairments were caused by impaired protein kinase A (PKA)-mediated phosphorylation because exogenous PKA restored all parameters to control levels. Although DCM samples showed reexpression of EH-myomesin, an isoform usually only expressed in the heart before birth, PPCM and ISHD did not. The lack of EH-myomesin, combined with low PKA-mediated phosphorylation of myofilament proteins and increased compliant titin isoform, may explain the increase in passive force and blunted length-dependent activation of myofilaments in PPCM samples., (Copyright © 2017 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

MLP (muscle LIM protein)-deficient mice count among the first mouse models for dilated cardiomyopathy (DCM), yet the exact role of MLP in cardiac signalling processes is still enigmatic. Elevated PKCα signalling activity is known to be an important contributor to heart failure. Here we show that MLP directly inhibits the activity of PKCα. In end-stage DCM, PKCα is concentrated at the intercalated disc of cardiomyocytes, where it is sequestered by the adaptor protein CARP in a multiprotein complex together with PLCβ1. In mice deficient for both MLP and CARP the chronic PKCα signalling chain at the intercalated disc is broken and they remain healthy. Our results suggest that the main role of MLP in heart lies in the direct inhibition of PKCα and that chronic uninhibited PKCα activity at the intercalated disc in the absence of functional MLP leads to heart failure.

In cardiomyocytes columns of intermyofibrillar mitochondria run up to the intercalated disc (ID); half are collinear with those in the neighbouring cell, suggesting coordinated addition of sarcomeres and mitochondria both within and between cells during cardiomyocyte growth. Recent evidence for an association between sarcoplasmic reticulum (SR) and mitochondria indicates that the SR may be an intermediary in this coordinated behaviour. For this reason we have investigated the arrangement of SR and t tubules with respect to mitochondria and myofibrils, particularly at the ID. In the body of the cardiomyocyte the mitochondrial columns are frequently intersected by transverse tubules. In addition, we find that a majority of axial tubules are sandwiched between mitochondria and myofibril. No tubules are found at the ID. SR coats mitochondrial columns and fibrils throughout their length and reaches towards the peaks of the ID membrane where it attaches in the form of junctional (j)SR. These peripheral ID couplings are often situated between mitochondria and ID membrane, suggesting an SR connection between the two. In dilated cardiomyopathy (DCM) the mitochondria are somewhat disordered and clumped. In a mouse model for DCM, the muscle LIM protein KO, we find that there is a lack of mitochondria near the ID, suggesting the uncoupling of the myofibril/mitochondria organisation during growth. SR still coats the fibrils and reaches the ID folds in a jSR coupling. Unlike in control tissue, however, loops and long fingers of ID membrane penetrate into the proximal sarcomere suggesting a possible intermediary state in cardiomyocyte growth., Competing Interests: There are no sources of conflict.

Background: Studies of the functional consequences of DCM-causing mutations have been limited to a few cases where patients with known mutations had heart transplants. To increase the number of potential tissue samples for direct investigation we performed whole exon sequencing of explanted heart muscle samples from 30 patients that had a diagnosis of familial dilated cardiomyopathy and screened for potentially disease-causing mutations in 58 HCM or DCM-related genes., Results: We identified 5 potentially disease-causing OBSCN mutations in 4 samples; one sample had two OBSCN mutations and one mutation was judged to be not disease-related. Also identified were 6 truncating mutations in TTN, 3 mutations in MYH7, 2 in DSP and one each in TNNC1, TNNI3, MYOM1, VCL, GLA, PLB, TCAP, PKP2 and LAMA4. The mean level of obscurin mRNA was significantly greater and more variable in healthy donor samples than the DCM samples but did not correlate with OBSCN mutations. A single obscurin protein band was observed in human heart myofibrils with apparent mass 960 ± 60 kDa. The three samples with OBSCN mutations had significantly lower levels of obscurin immunoreactive material than DCM samples without OBSCN mutations (45±7, 48±3, and 72±6% of control level).Obscurin levels in DCM controls, donor heart and myectomy samples were the same., Conclusions: OBSCN mutations may result in the development of a DCM phenotype via haploinsufficiency. Mutations in the obscurin gene should be considered as a significant causal factor of DCM, alone or in concert with other mutations.

Myosin VI (MVI) is a unique unconventional myosin translocating, unlike other myosins, towards the minus end of actin filaments. It is involved in numerous cellular processes such as endocytosis, intracellular trafficking, cell migration, and transcription. In mammalian skeletal muscles it localizes mainly to sarcoplasmic reticulum and is also present within the muscle nuclei and at the neuromuscular junction (Karolczak et al. Histochem Cell Biol 2013; 23:219-228). We have also shown that in denervated rat hindlimb muscle the MVI expression level is significantly increased and its localization is changed, indicating an important role of MVI in striated muscle pathology. Here, we addressed this problem by examining the distribution and expression levels of myosin VI in biopsies of skeletal muscles from patients with different myopathies. We found that, particularly in myopathies associated with fiber atrophy, the amount of MVI was enhanced and its localization in affected fibers was changed. Also, since a mutation within the human MVI gene was shown to be associated with cardiomyopathy, we assessed MVI localization and expression level in cardiac muscle using wild type and MLP(-/-) mice, a dilated cardiomyopathy model. No significant difference in MVI expression level was observed for both types of animals. MVI was found at intercalated discs and also at the sarcoplasmic reticulum. In the knockout mice, it was also present in ring-like structures surrounding the nuclei. The data indicate that in striated muscle MVI could be engaged in sarcoplasmic reticulum maintenance and/or functioning, vesicular transport, signal transmission and possibly in gene transcription., (© 2014 Wiley Periodicals, Inc.)

The M-band is the prominent cytoskeletal structure that cross-links the myosin and titin filaments in the middle of the sarcomere. To investigate M-band alterations in heart disease, we analyzed the expression of its main components, proteins of the myomesin family, in mouse and human cardiomyopathy. Cardiac function was assessed by echocardiography and compared to the expression pattern of myomesins evaluated with RT-PCR, Western blot, and immunofluorescent analysis. Disease progression in transgenic mouse models for dilated cardiomyopathy (DCM) was accompanied by specific M-band alterations. The dominant splice isoform in the embryonic heart, EH-myomesin, was strongly up-regulated in the failing heart and correlated with a decrease in cardiac function (R = -0.86). In addition, we have analyzed the expressions of myomesins in human myocardial biopsies (N = 40) obtained from DCM patients, DCM patients supported by a left ventricular assist device (LVAD), hypertrophic cardiomyopathy (HCM) patients and controls. Quantitative RT-PCR revealed that the EH-myomesin isoform was up-regulated 41-fold (P < 0.001) in the DCM patients compared to control patients. In DCM hearts supported by a LVAD and HCM hearts, the EH-myomesin expression was comparable to controls. Immunofluorescent analyses indicate that EH-myomesin was enhanced in a cell-specific manner, leading to a higher heterogeneity of the myocytes' cytoskeleton through the myocardial wall. We suggest that the up-regulation of EH-myomesin denotes an adaptive remodeling of the sarcomere cytoskeleton in the dilated heart and might serve as a marker for DCM in mouse and human myocardium.

Beta-catenin is a component of the intercalated disc in cardiomyocytes, but can also be involved in signalling and activation of gene transcription. We wanted to determine how long-term changes in beta-catenin expression levels would affect mature cardiomyocytes. Conditional transgenic mice that either lacked beta-catenin or that expressed a non-degradable form of beta-catenin in the adult ventricle were created. While mice lacking beta-catenin in the ventricle do not have an overt phenotype, mice expressing a non-degradable form develop dilated cardiomyopathy and do not survive beyond 5 months. A detailed analysis could reveal that this phenotype is correlated with a distinct localisation of beta-catenin in adult cardiomyocytes, which cannot be detected in the nucleus, no matter how much protein is present. Our report is the first study that addresses long-term effects of either the absence of beta-catenin or its stabilisation on ventricular cardiomyocytes and it suggests that beta-catenin's role in the nucleus may be of little significance in the healthy adult heart.

The contractile tissue of the heart is composed of individual cells, making specific cell-cell contacts necessary to ensure mechanical and electrochemical coupling during beating. These contact sites, termed the intercalated discs, have gained increased attention recently due to their potential involvement in cardiac disease. This article discusses how the intercalated discs are assembled during heart development and how they are affected in cardiomyopathy, with particular emphasis on dilated cardiomyopathy. A model is proposed to relate the alterations that are seen at a molecular level with changes in function observed in that kind of cardiac disease.

The unique cytoarchitecture of cardiomyocytes arises by complex interactions of different filamentous structures of the cytoskeleton. Intermediate filaments of the non-sarcomeric cytoskeleton are not essential for development but important for maintenance of myofibrils. Myofibrils consist of contractile proteins involved in force generation and the muscle cytoskeleton framework. The latter is essential for proper assembly and maintenance as well as for interaction with other cardiomyocytes or the extracellular matrix, thus being involved in force transmission. The information for sarcomere assembly is encoded in the proteins and some domains essential for faithful incorporation have been identified by epitope tagging experiments. Many KO mutations result in embryonic lethal phenotypes and new techniques e.g. using cardiomyocytes derived from ES cell-lines will have to be developed that allow to study such mutations in cardiomyocytes rather than whole organisms. Alterations in the expression levels of several proteins of the muscle cytoskeleton or impairment of their function by point mutations can result in increased mechanical stress in the cardiomyocytes which finally leads to cellular responses such as the development of dilated cardiomyopathy (DCM). MLP (muscle-LIM-protein) deficient mice develop DCM and changes in the mechanical coupling of cardiomyocytes result in alterations at the intercalated disks and enhanced accumulation of adherens junction proteins. Therefore, controlled interactions between proteins of the muscle cytoskeleton and contractile proteins are essential to ensure proper cardiac function and a more detailed insight in these processes might provide new tools to improve the contractile efficiency of the cardiomyocytes and thus working output in cardiomyopathies.