Papers by Elisabeth Ehler
Cardiomyocytes are distinguished by a particularly regularly arranged cytoskeleton. Both the myof... more Cardiomyocytes are distinguished by a particularly regularly arranged cytoskeleton. Both the myofibrils, which perform the contractile work of the heart, and the intercalated discs, a special type of cell–cell contact that serves for mechanical and electrochemical connection between individual cardiomyocytes, are multiprotein complexes that must be assembled in a regular fashion during development to guarantee a fully functional heart. In heart disease such as hypertrophic cardiomyopathy, dilated cardiomyopathy or arrhythmogenic right ventricular cardiomyopathy, these structures can be compromised in their composition and thus function. The aim of this chapter is to discuss how cardiac cytoarchitecture is established during development and how it is altered in disease. Cardiomyocytes, which make up the contractile tissue of the heart, are characterised by a particularly regular cytoarchitecture that ensures that they deliver the maximum in contractile force from sliding actin and myosin molecules. They are arranged into a three-dimensional tissue in a way that guarantees a minimum of leakage of the signal that leads to a coordinated contraction throughout the heart chamber. This complex cytoarchitecture is mainly defined by the organisation of cytoskeletal proteins to multiprotein complexes such as, e.g., the myofibrils or the intercalated discs. This regular arrangement has fascinated researchers for decades and many studies were carried out over to years to investigate the assembly and maintenance of these structures either in cultured cardiomyocytes or in the animal in situ. M. Pluess • E. Ehler (*) Cardiovascular Division, BHF Centre of Research Excellence at King’s College London, London, UK Randall Division of Cell and Molecular Biophysics, New Hunt’s House, Guy’s Campus, Room 3.26A, London SE1 1UL, UK e-mail: [email protected] © Springer International Publishing Switzerland 2015 E. Ehler (ed.), Cardiac Cytoarchitecture, DOI 10.1007/978-3-319-15263-9_1 1 1.1 Cardiomyocytes During Development The heart is the first functional organ in developing mouse and chick embryos. It starts beating at embryonic day 9 in mouse and Hamburger–Hamilton stage 10 in chick and failure to function results in early embryonic lethality (Van Vliet et al. 2012). In the zebrafish embryo, the somites differentiate before the heart and also defects in heart function can be survived a bit longer due to the possibility of nutrient diffusion (Bakkers 2011). Remarkably this means that while the embryonic heart already has to carry out a certain amount of work, it is still being reshaped into a four-chambered organ in mammals and it also has to grow in size by a process called hyperplasia as the embryo grows (Rana et al. 2013). In contrast to other tissues, where new supply of cells comes from a less differentiated precursor population, all cardiomyocytes in the embryonic heart appear to possess functional myofibrils and can contract. Analysis for classical components of the myofibrils such as sarcomeric alpha-actinin, sarcomeric myosin heavy chain, titin or myomesin reveals no difference in sarcomeric protein expression throughout an embryonic ventricle (Ehler et al. 1999), suggesting that there is no numerous population of less differentiated cardiomyocyte precursor cells. However, it appears that the dynamic processes of contraction and cell division are not compatible with each other. Therefore, in order to allow for hyperplastic growth, the cardiomyocytes disassemble their myofibrils, undergo cell division and reassemble their myofibrils immediately after cytokinesis, while always remaining integrated in a tissue surrounded by beating neighbouring cardiomyocytes (Ahuja et al. 2004). Around postnatal day 4 in the mouse, cell division ceases and from then onwards the heart grows by increase of the size of the individual cardiomyocytes, a process called hypertrophy (Leu et al. 2001). Interestingly, this time of cessation of divisional activity of cardiomyocytes overlaps exactly with the time when successful repair of injury in the mouse heart cedes to happen (Porrello et al. 2011). If the tip of a mouse heart ventricle is cut off at postnatal day 0, the heart recovers fine and the missing tissue is replaced by neighbouring cardiomyocytes that are stimulated to undergo cell division following the injury. At postnatal day 7 this recovery is no longer possible and a similar scar is produced as in an injured adult heart, for example following myocardial infarction (Porrello et al. 2011). Ability to divide and to regenerate therefore seems to be tightly coupled and it is of huge interest to elucidate which signalling pathways need to be triggered to allow for regeneration in case of injury. At the same time, this arrest in cytokinetic activity in cardiomyocytes means that there are no “heart cancers”, at least not any caused by hyperplasia of adult ventricular cardiomyocytes. Why are embryonic cardiomyocytes able to undergo…
Circulation Research, 2013
This article reviews the strengths and limitations of induced pluripotent stem cell-derived cardi... more This article reviews the strengths and limitations of induced pluripotent stem cell-derived cardiomyocytes (iPSC-CM) as models of cardiac arrhythmias. Specifically, the article attempts to answer the following questions: Which clinical arrhythmias can be modeled by iPSC-CM? How well can iPSC-CM model adult ventricular myocytes? What are the strengths and limitations of published iPSC-CM arrhythmia models? What new mechanistic insight has been gained? What is the evidence that would support using iPSC-CM to personalize antiarrhythmic drug therapy? The review also discusses the pros and cons of using the iPSC-CM technology for modeling specific genetic arrhythmia disorders, such as long QT syndrome, Brugada Syndrome, or Catecholaminergic Polymorphic Ventricular Tachycardia.
Heart, 2002
Objective: To investigate how permanent inhibition of guanylyl cyclase A receptor (GC-A) affects ... more Objective: To investigate how permanent inhibition of guanylyl cyclase A receptor (GC-A) affects cardiac function. Methods: Hearts of GC-A−/− and corresponding wild type mice (GC-A+/+) were characterised by histological, western blotting, and northern blotting analyses. Cardiac function was evaluated in isolated, working heart preparations. Results: At 4 months of age, GC-A−/− mice had global cardiac hypertrophy (about a 40% increase in cardiac weight) without interstitial fibrosis. Examination of heart function found a significant delay in the time of relaxation; all other parameters of cardiac contractility were similar to those in wild type mice. At 12 months, the hypertrophic changes were much more severe (about a 61% increase in cardiac weight), together with a shift in cardiac gene expression (enhanced concentrations of atrial natriuretic peptide (3.8-fold), B type natriuretic peptide (2-fold), β myosin heavy chain (1.6-fold) and α skeletal actin (1.7-fold) mRNA), increased expression of cytoskeletal tubulin and desmin (by 29.6% and 25.6%, respectively), and pronounced interstitial fibrosis. These changes were associated with significantly impaired cardiac contractility (+dP/dt decreased by about 10%) and relaxation (−dP/dt decreased by 21%), as well as depressed contractile responses to pressure load (all p < 0.05). Conclusions: Chronic hypertension in GC-A−/− mice is associated with progressive cardiac changes-namely, initially compensated cardiomyocyte hypertrophy, which is complicated by interstitial fibrosis and impaired cardiac contractility at later stages.
Journal of Muscle Research and Cell Motility, 1999
The development of myofibrils involves the formation of contractile filaments and their assembly ... more The development of myofibrils involves the formation of contractile filaments and their assembly into the strikingly regular structure of the sarcomere. We analysed this assembly process in cultured human skeletal muscle cells and in rat neonatal cardiomyocytes by immunofluorescence microscopy using antibodies directed against cytoskeletal and contractile proteins. In particular, the question in which temporal order the respective proteins are
The EMBO Journal, 2000
In polarized epithelial cells measles virus (MV) is predominantly released at the apical cell sur... more In polarized epithelial cells measles virus (MV) is predominantly released at the apical cell surface, irrespective of the sorting of its two envelope glycoproteins F and H. It has been reported previously that the viral matrix (M) protein modulates the fusogenic capacity of the viral envelope glycoproteins. Here, extant MV mutants and chimeras were used to determine the role of M protein in the transport of viral glycoproteins and release of progeny virions in polarized epithelial CaCo2 cells. In the absence of M, envelope glycoproteins are sorted to the basolateral surface, suggesting that they possess intrinsic basolateral sorting signals. However, interactions of M with the glycoprotein cytoplasmic tails allow M±glycoprotein co-segregation to the apical surface, suggesting a vectorial function of M to retarget the glycoproteins for apical virion release. Whereas this may allow virus airway shedding, the intrinsic sorting of the glycoproteins to the basolateral surface may account for systemic host infection by allowing ef®cient cell±cell fusion.
Proceedings of the National Academy of Sciences, 1998
Protein phosphatase 2A (PP2A) is a multimeric enzyme, containing a catalytic subunit complexed wi... more Protein phosphatase 2A (PP2A) is a multimeric enzyme, containing a catalytic subunit complexed with two regulatory subunits. The catalytic subunit PP2A C is encoded by two distinct and unlinked genes, termed Cα and Cβ. The specific function of these two catalytic subunits is unknown. To address the possible redundancy between PP2A and related phosphatases as well as between Cα and Cβ, the Cα subunit gene was deleted by homologous recombination. Homozygous null mutant mice are embryonically lethal, demonstrating that the Cα subunit gene is an essential gene. As PP2A exerts a range of cellular functions including cell cycle regulation and cell fate determination, we were surprised to find that these embryos develop normally until postimplantation, around embryonic day 5.5/6.0. While no Cα protein is expressed, we find comparable expression levels of PP2A C at a time when the embryo is degenerating. Despite a 97% amino acid identity, Cβ cannot completely compensate for the absence of C...
Journal of Virology, 2002
Measles virus (MV) can infect the central nervous system and, in rare cases, causes subacute scle... more Measles virus (MV) can infect the central nervous system and, in rare cases, causes subacute sclerosing panencephalitis, characterized by a progressive degeneration of neurons. The route of MV transmission in neurons was investigated in cultured rat hippocampal slices by using MV expressing green fluorescent protein. MV infected hippocampal neurons and spread unidirectionally, in a retrograde manner, from CA1 to CA3 pyramidal cells and from there to the dentate gyrus. Spreading of infection depended on cell-to-cell contact and occurred without any detectable release of infectious particles. The role of the viral proteins in the retrograde MV transmission was determined by investigating their sorting in infected pyramidal cells. MV glycoproteins, the fusion protein (F) and hemagglutinin (H), the matrix protein (M), and the phosphoprotein (P), which is part of the viral ribonucleoprotein complex, were all sorted to the dendrites. While M, P, and H proteins remained more intracellular,...
Journal of Biological Chemistry, 2010
3 The abbreviations used are: RSK, 90-kDa ribosomal S6 kinase; ARVM, adult rat ventricular myocyt... more 3 The abbreviations used are: RSK, 90-kDa ribosomal S6 kinase; ARVM, adult rat ventricular myocytes; cMyBP-C, cardiac myosin-binding protein C; cTnI, cardiac troponin I; cTnI-Ala 2 , cTnI in which Ser 22 /Ser 23 are substituted by nonphosphorylatable Ala; CTK, C-terminal kinase; ET1, endothelin 1; ki, kinase-inactive; NHE, Na ϩ /H ϩ exchanger; NTK, N-terminal kinase; pCa,-log[Ca 2ϩ ]; pCa 50 , pCa at 50% maximum force.
FEBS Letters, 2000
NT2 cells are a transfectable human embryonal carcinoma cell line, that can be differentiated int... more NT2 cells are a transfectable human embryonal carcinoma cell line, that can be differentiated into postmitotic neuron-like cells (NT2N cells), and transplanted into rodent brains. Differentiation requires a 5-week-long treatment with retinoic acid prior to transplantation. Here, we show that this step can be omitted, and that undifferentiated NT2 cells migrate over long distances and differentiate into both neuron-and oligodendrocyte-like cell types upon grafting into brains of immunocompetent newborn mice. Grafted cells can be traced by fluorogold, with no evidence for tumor formation. Our approach provides an experimental model system which allows the immunohistological and biochemical study of neuronal and glial differentiation of human cells in vivo, and which may be suitable as an in vivo model for pharmacological studies.
European Journal of Cell Biology, 2010
Skeletal muscle differentiation is a complex process: It is characterised by changes in gene expr... more Skeletal muscle differentiation is a complex process: It is characterised by changes in gene expression and protein composition. Simultaneously, a dramatic remodelling of the cytoskeleton and associated cell-matrix contacts, the costameres, occurs. The expression and localisation of the protein ponsin at cell-matrix contacts marks the establishment of costameres. In this report we show that skeletal muscle cells are characterised by a novel ponsin isoform, which contains a large insertion in its carboxy-terminus. This skeletal muscle-specific module binds the adapter proteins Nck1 and Nck2, and increased co-localisation of ponsin with Nck2 is observed at remodelling cell-matrix contacts of differentiating skeletal muscle cells. Since this ponsin insertion can be phosphorylated, it may adjust the interaction affinity with Nck adapter proteins. The novel ponsin isoform and its interaction with Nck1/2 provide exciting insight into the convergence of signalling pathways at the costameres, and its crucial role for skeletal muscle differentiation and re-generation.
Endocrinology, 2001
The intestinal peptides, guanylin and uroguanylin, may have an important role in the endocrine co... more The intestinal peptides, guanylin and uroguanylin, may have an important role in the endocrine control of renal function. Both peptides and their receptor, guanylyl cyclase C (GC-C), are also expressed within the kidney, suggesting that they may act locally in an autocrine/paracrine fashion. However, their physiological regulation within the kidney has not been studied. To begin to address this issue, we evaluated the distribution of uroguanylin and guanylin messenger RNA (mRNA) in the mouse nephron and the regulation of renal expression by changes in dietary salt/water intake. Expression was determined in 1) wild-type mice, 2) two strains of receptor-guanylyl cyclase-deficient mice (ANP-receptor-deficient, GC-AϪ/Ϫ, and GC-C-deficient mice); and 3) cultured renal epithelial (M-1) cells, by RT-PCR, Northern blotting and immunocytochemistry. Renal uroguanylin messenger RNA expression was higher than
Histochemistry and Cell Biology, 2012
Myosin VI (MVI) is a unique unconventional motor moving backwards on actin filaments. In non-musc... more Myosin VI (MVI) is a unique unconventional motor moving backwards on actin filaments. In non-muscle cells, it is involved in cell migration, endocytosis and intracellular trafficking, actin cytoskeleton dynamics, and possibly in gene transcription. An important role for MVI in striated muscle functioning was suggested in a report showing that a point mutation (H236R) within the MVI gene was associated with cardiomyopathy (Mohiddin et al., J Med Genet 41:309-314, 2004). Here, we have addressed MVI function in striated muscle by examining its expression and distribution in rat hindlimb skeletal muscle. We found that MVI was present predominantly at the muscle fiber periphery, and it was also localized within muscle nuclei. Analysis of both the hindlimb and cardiac muscle longitudinal sections revealed *3 lm striation pattern, corresponding to the sarcoplasmic reticulum. Moreover, MVI was detected in the sarcoplasmic reticulum fractions isolated from skeletal and cardiac muscle. The protein also localized to the postsynaptic region of the neuromuscular junction. In denervated muscle, the defined MVI distribution pattern was abolished and accompanied by significant increase in its amount in the muscle fibers. In addition, we have identified several novel potential MVI-binding partners, which seem to aid our observations that in striated muscle MVI could be involved in postsynaptic trafficking as well as in maintenance of and/or transport within the sarcoplasmic reticulum and non-sarcomeric cytoskeleton.
Basic Research in Cardiology, 2021
Titin truncating variants are a well-established cause of cardiomyopathy; however, the role of ti... more Titin truncating variants are a well-established cause of cardiomyopathy; however, the role of titin missense variants is less well understood. Here we describe the generation of a mouse model to investigate the underlying disease mechanism of a previously reported titin A178D missense variant identified in a family with non-compaction and dilated cardiomyopathy. Heterozygous and homozygous mice carrying the titin A178D missense variant were characterised in vivo by echocardiography. Heterozygous mice had no detectable phenotype at any time point investigated (up to 1 year). By contrast, homozygous mice developed dilated cardiomyopathy from 3 months. Chronic adrenergic stimulation aggravated the phenotype. Targeted transcript profiling revealed induction of the foetal gene programme and hypertrophic signalling pathways in homozygous mice, and these were confirmed at the protein level. Unsupervised proteomics identified downregulation of telethonin and four-and-a-half LIM domain 2, a...
Biophysical Reviews, 2021
Biophysical reviews, 2018
It has been known for several decades that mutations in genes that encode for proteins involved i... more It has been known for several decades that mutations in genes that encode for proteins involved in the control of actomyosin interactions such as the troponin complex, tropomyosin and MYBP-C and thus regulate contraction can lead to hereditary hypertrophic cardiomyopathy. In recent years, it has become apparent that actin-binding proteins not directly involved in the regulation of contraction also can exhibit changed expression levels, show altered subcellular localisation or bear mutations that might lead to hereditary cardiomyopathies. The aim of this review is to look beyond the troponin/tropomyosin mechanism and to give an overview of the different types of actin-associated proteins and their potential roles in cardiomyocytes. It will then discuss recent findings relevant to their involvement in heart disease.
Cardiovascular research, Jan 10, 2018
The pathology of heart failure is characterized by poorly contracting and dilated ventricles. At ... more The pathology of heart failure is characterized by poorly contracting and dilated ventricles. At the cellular level, this is associated with lengthening of individual cardiomyocytes and loss of sarcomeres. While it is known that the transcription factor myocyte enhancer factor -2 (MEF2) is involved in this cardiomyocyte remodeling, the underlying mechanism remains to be elucidated. Here, we aim to mechanistically link MEF2 target genes with loss of sarcomeres during cardiomyocyte remodeling. Neonatal rat cardiomyocytes overexpressing MEF2 elongated and lost their sarcomeric structure. We identified myotonic dystrophy protein kinase (DMPK) as direct MEF2 target gene involved in this process. Adenoviral overexpression of DMPK E, the isoform upregulated in heart failure, resulted in severe loss of sarcomeres in vitro and transgenic mice overexpressing DMPK E displayed disruption of sarcomere structure and cardiomyopathy in vivo. Moreover, we found a decreased expression of sarcomeric g...
Developmental cell, Feb 5, 2018
Mechanical properties are cues for many biological processes in health or disease. In the heart, ... more Mechanical properties are cues for many biological processes in health or disease. In the heart, changes to the extracellular matrix composition and cross-linking result in stiffening of the cellular microenvironment during development. Moreover, myocardial infarction and cardiomyopathies lead to fibrosis and a stiffer environment, affecting cardiomyocyte behavior. Here, we identify that single cardiomyocyte adhesions sense simultaneous (fast oscillating) cardiac and (slow) non-muscle myosin contractions. Together, these lead to oscillating tension on the mechanosensitive adaptor protein talin on substrates with a stiffness of healthy adult heart tissue, compared with no tension on embryonic heart stiffness and continuous stretching on fibrotic stiffness. Moreover, we show that activation of PKC leads to the induction of cardiomyocyte hypertrophy in a stiffness-dependent way, through activation of non-muscle myosin. Finally, PKC and non-muscle myosin are upregulated at the costamere...
The American Journal of Pathology, 2017
If citing, it is advised that you check and use the publisher's definitive version for pagination... more If citing, it is advised that you check and use the publisher's definitive version for pagination, volume/issue, and date of publication details. And where the final published version is provided on the Research Portal, if citing you are again advised to check the publisher's website for any subsequent corrections.
Journal of muscle research and cell motility, Jun 21, 2016
In cardiomyocytes columns of intermyofibrillar mitochondria run up to the intercalated disc (ID);... more 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. Th...
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Papers by Elisabeth Ehler