Epicardium-derived Cells Research Articles

A Dynamic Notch Injury Response Activates Epicardium and Contributes to Fibrosis Repair

Transgenic Notch reporter mice express enhanced green fluorescent protein in cells with C-promoter binding factor-1 response element transcriptional activity (CBF1-RE(x)₄-EGFP), providing a unique and powerful tool for identifying and isolating "Notch-activated" progenitors. We asked whether, as in other tissues of this mouse, EGFP localized and functionally tagged adult cardiac tissue progenitors, and, if so, whether this cell-based signal could serve as a quantitative and qualitative biosensor of the injury repair response of the heart. In addition to scattered endothelial and interstitial cells, Notch-activated (EGFP(+)) cells unexpectedly richly populated the adult epicardium. We used fluorescence-activated cell sorting to isolate EGFP(+) cells and excluded hematopoietic (CD45(+)) and endothelial (CD31(+)) subsets. We analyzed EGFP(+)/CD45⁻/CD31⁻ cells, a small (<2%) but distinct subpopulation, by gene expression profiling and functional analyses. We called this mixed cell pool, which had dual multipotent stromal cell and epicardial lineage signatures, Notch-activated epicardial-derived cells (NECs). Myocardial infarction and thoracic aortic banding amplified the NEC pool, increasing fibroblast differentiation. Validating the functional vitality of clonal NEC lines, serum growth factors triggered epithelial-mesenchymal transition and the immobilized Notch ligand Delta-like 1-activated downstream target genes. Moreover, cardiomyocyte coculture and engraftment in NOD-SCID (nonobese diabetic-severe combined immunodeficiency) mouse myocardium increased cardiac gene expression in NECs. A dynamic Notch injury response activates adult epicardium, producing a multipotent cell population that contributes to fibrosis repair.

Desmosomes and Gap Junctions in Epicardium-Derived Cells. A Possible Role in Arrhythmogenic Right Ventricular Cardiomyopathy?

Background: Arrhythmogenic right ventricular cardiomyopathy (ARVC) is an inherited disease characterized by severe ventricular arrhythmias and progressive replacement of healthy myocardium with fibrous and adipose tissue. ARVC associates with mutations in genes coding for desmosomal proteins, most commonly plakophilin-2 (PKP2). It is unclear how disruption of desmosomes in cardiac myocytes leads to fibrosis and fat accumulation. We propose that cardiac-resident, desmosome-expressing, non-myocyte cells are a major source of fibroblasts and adipocytes in ARVC.

Epicardium-derived cells enhance proliferation, cellular maturation and alignment of cardiomyocytes

During heart development, cells from the proepicardial organ spread over the naked heart tube to form the epicardium. From here, epicardium-derived cells (EPDCs) migrate into the myocardium. EPDCs proved to be indispensable for the formation of the ventricular compact zone and myocardial maturation, by largely unknown mechanisms. In this study we investigated in vitro how EPDCs affect cardiomyocyte proliferation, cellular alignment and contraction, as well as the expression and cellular distribution of proteins involved in myocardial maturation. Embryonic quail EPDCs induced proliferation of neonatal mouse cardiomyocytes. This required cell–cell interactions, as proliferation was not observed in transwell cocultures. Western blot analysis showed elevated levels of electrical and mechanical junctions (connexin43, N-cadherin), sarcomeric proteins (Troponin-I, α-actinin), extracellular matrix (collagen I and periostin) in cocultures of EPDCs and cardiomyocytes. Immunohistochemistry indicated more membrane-bound expression of Cx43, N-cadherin, the mechanotransduction molecule focal adhesion kinase, and higher expression of the sarcoplasmic reticulum Ca 2+ ATPase (SERCA2a). Newly developed software for analysis of directionality in immunofluorescent stainings showed a quantitatively determined enhanced cellular alignment of cardiomyocytes. This was functionally related to increased contraction. The in vitro effects of EPDCs on cardiomyocytes were confirmed in three reciprocal in vivo models for EPDC-depletion (chicken and mice) in which downregulation of myocardial N-cadherin, Cx43, and FAK were observed. In conclusion, direct interaction of EPDCs with cardiomyocytes induced proliferation, correct mechanical and electrical coupling of cardiomyocytes, ECM-deposition and concurrent establishment of cellular array. These findings implicate that EPDCs are ideal candidates as adjuvant cells for cardiomyocyte integration during cardiac (stem) cell therapy.

Transcription factor heterogeneity in proepicardial and epicardium-derived cell populations during heart development

Transcription factor heterogeneity in proepicardial and epicardium-derived cell populations during heart development

Cardiac malformations in Pdgfrα mutant embryos are associated with increased expression of WT1 and Nkx2.5 in the second heart field

Platelet-derived growth factor receptor alpha (Pdgfralpha) identifies cardiac progenitor cells in the posterior part of the second heart field. We aim to elucidate the role of Pdgfralpha in this region. Hearts of Pdgfralpha-deficient mouse embryos (E9.5-E14.5) showed cardiac malformations consisting of atrial and sinus venosus myocardium hypoplasia, including venous valves and sinoatrial node. In vivo staining for Nkx2.5 showed increased myocardial expression in Pdgfralpha mutants, confirmed by Western blot analysis. Due to hypoplasia of the primary atrial septum, mesenchymal cap, and dorsal mesenchymal protrusion, the atrioventricular septal complex failed to fuse. Impaired epicardial development and severe blebbing coincided with diminished migration of epicardium-derived cells and myocardial thinning, which could be linked to increased WT1 and altered alpha4-integrin expression. Our data provide novel insight for a possible role for Pdgfralpha in transduction pathways that lead to repression of Nkx2.5 and WT1 during development of posterior heart field-derived cardiac structures.

Transcription factor CHF1/Hey2 regulates coronary vascular maturation

The transcription factor CHF1/Hey2 has been implicated in a variety of cardiovascular developmental abnormalities including ventricular septal defect, deformed valves and cardiomyopathy. To date, its role in coronary vascular development remains unknown. We have found that KO mice developed coronary vascular abnormalities accompanied by a thin compact ventricular myocardium but grossly normal epicardial and subepicardial layers. The coronary vascular anomalies included dysmorphic large vessels and abnormal vascular structures at E15.5 and reduced recruitment of vascular smooth muscle cells into the coronary arteries at E18.5. In E18.5 KO hearts, the abnormal coronary veins demonstrated reduced expression of markers for vein identity. Whole-mount PECAM staining of the E18.5 KO hearts indicated that EphB4 negative vein networks were increased in the surface layers of the myocardium compared to those of the controls. CHF1/Hey2 was not expressed in the epicardium in vivo, and cultured epicardium-derived cells isolated from E12.5 wild-type mice showed no CHF1/Hey2 expression. KO mice with a myocardially expressed CHF1/Hey2 transgene partially rescued the vascular phenotypes. Quantitative RT-PCR analysis demonstrated that PDGF and Angiopoietin/Tie2 signaling pathways are altered in E12.5 KO hearts. Taken together, global CHF1/Hey2 deficiency caused impaired vascular formation, the reduced recruitment of vascular smooth muscle cells into coronary arteries and abnormally remodeled vein networks. These findings suggest that CHF1/Hey2 regulates the later steps of coronary vascular development in both a myocardial-dependent, non-cell autonomous fashion and likely a vascular cell-specific effect as well.

Epithelial–mesenchymal transition of epicardial mesothelium is a source of cardiac CD117-positive stem cells in adult human heart

Epithelial–mesenchymal transition is implicated in the remodelling of tissues during development and in the adult life. In the heart, it gives origin to progenitors of fibroblasts, coronary endothelium, smooth muscle cells, and cardiomyocytes. Moreover, epicardially-derived cells determine myocardial wall thickness and Purkinje fibre network. Recently, the presence of numerous cardiac stem cells in the subepicardium of the adult human heart has been described and the hypothesis that epicardially-derived cells can contribute to the population of cardiac stem cells in the adult heart has been advanced. In an effort to test this hypothesis and establish a possible link between epicardium, epicardially-derived cells and cardiac stem cells in the adult human heart we have examined epicardial mesothelial cells in the normal and pathological adult human heart with ischemic cardiomyopathy in vivo and we have induced and documented their epithelial–mesenchymal transition in vitro. Noticeably, epicardial cells were missing from the surface of pathological hearts and the cells with the expression of epithelial and mesenchymal markers populated thick subepicardial space. When the fragments of epicardium from the normal hearts were cultured on the specific substrate formed by extracellular matrix derived from cardiac fibroblasts, we obtained the outgrowth of the epithelial sheet with the mRNA and protein expression characteristic of epicardium. TGFβ induced cellular and molecular changes typical of epithelial–mesenchymal transition. Moreover, the epicardially-derived cells expressed CD117 antigen. Thus, this study provides evidence that cardiac stem cells can originate from epithelial–mesenchymal transition of the epicardial cells in the adult human heart.

Epicardium derived cells (EPDCs) in development, cardiac disease and repair of ischemia

The proepicardial-derived epicardium covers the myocardium and after a process of epithelial–mesenchymal transition (EMT) forms epicardium-derived cells (EPDCs). These cells migrate into the myocardium and show an essential role in the induction of the ventricular compact myocardium and the differentiation of the Purkinje fibres. EPDCs are furthermore the source of the interstitial fibroblast, the coronary smooth muscle cell and the adventitial fibroblast. The possible differentiation into cardiomyocytes, endothelial cells and the recently described telocyte and other cells in the cardiac stem cell niche needs further investigation. Surgically or genetically disturbed epicardial and EPDC differentiation leads to a spectrum of abnormalities varying from thin undifferentiated myocardium, which can be embryonic lethal, to a diminished coronary vascular bed with even absent main coronary arteries. The embryonic potential of EPDCs has been translated to both structural and functional congenital malformations and adult cardiac disease, like development of Ebstein’s malformation, arrhythmia and cardiomyopathies. Furthermore, the use of adult EPDCs as a stem cell source has been explored, showing in an animal model of myocardial ischemia the recapitulation of the embryonic program with improved function, angiogenesis and less adverse remodeling. Combining EPDCs and adult cardiomyocyte progenitor cells synergistically improved these results. The contribution of injected EPDCs was instructive rather than constructive. The finding of reactivation of the endogenous epicardium in ischemia with re-expression of developmental genes and renewed EMT marks the onset of a novel therapeutic focus.

Epicardial cell transformation

The embryonic epicardium, which invests all the heart surface by midgestation, derives from an extracardiac progenitor tissue called the proepicardium. The embryonic epicardium gives rise to a population of mesenchymal cells called the epicardially‐derived cells (EPDCs) that populate the space between the myocardium and the epicardium and later invade the ventricular myocardium. EPDCs contribute to the connective and vascular tissue of the heart. The transformation of epicardial cells into EPDCs is a critical step of the cardiac development, and it can be regarded as an epithelial‐mesenchymal transition (EMT), the same cellular process that accounts for the origin of the neural crest cells or the valvuloseptal tissue of the heart. We have just started to know how epicardial EMT is controlled at the molecular level, as well as the differences between the epicardial transformation and other embryonic EMTs. The Wilms' tumor suppressor gene Wt1 is probably a main element regulating epicardial EMT, since it is able to repress E‐cadherin and to activate Snail expression, regulating in this way the switch between epithelial and mesenchymal states in EPDCs. Wt1 also directly activates Raldh2 expression in the epicardium, thus controlling the retinoic acid synthesis in this tissue. We think that epicardial EMT should be regarded as a two‐step process, with a triggering signal in the proepicardium and the acquisition of an invasive behaviour when the primitive epicardium spreads over the heart surface. Later, a process of mesenchymal‐epithelial transformation in the most external epicardial cell layer gives rise to the definitive epicardial mesothelium.

Pulmonary atresia with intact ventricular septum: Second heart field derived myocardial and epicardial developmental clues

Pulmonary atresia with intact ventricular septum is a severe cardiac malformation that is ductus dependent and needs immediate postnatal care. The phenotype is characterized by three levels of pathology that are considered important for diagnosis and the choice of cardiac surgery. The pathology concerns an hypoplasia and hypertrophy of the right ventricle in which there is a variation in the involvement of the inlet, trabecular and outflow part of the right ventricle. There is also a variable degree of tricuspid valve pathology including hypoplasia and stenosis. The pulmonary valve is atretic which may have been preceded, in the fetal stage, by pulmonary stenosis. This latter phenomenon may be triggered by the severe coronary anomalies that are found in about 30% of the patients. This coronary anomaly is characterized by ventriculo-coronary arterial communications which is accompanied by sometimes severe main coronary artery pathology including interruptions of the main branches. In the latter situation the coronary myocardial perfusion becomes right ventricular dependent. Current cardiac developmental data show that the right ventricular myocardium is derived from the second heart field. Its anterior part contributes this myocardium to the outflow tract and is also employed by the neural crest cells to enter the heart. The posterior second heart field provides the epicardium derived cells (EPDC) that are important for myocardial wall compaction and formation of the coronary vasculature. We hypothesize that pulmonary atresia with intact ventricular septum without ventriculo-coronary arterial communications is primarily based on anterior SHF directed outflow tract septation anomalies, while pulmonary atresia with intact ventricular septum with ventriculo-coronary arterial communications has an additional major problem related to epicardium derived contribution. Developmentally they can therefore be considered as two different diseases that might need separate treatment protocols.

Cell tracking using iron oxide fails to distinguish dead from living transplanted cells in the infarcted heart

Recently, debate has arisen about the usefulness of cell tracking using iron oxide-labeled cells. Two important issues in determining the usefulness of cell tracking with MRI are generally overlooked; first, the effect of graft rejection in immunocompetent models, and second, the necessity for careful histological confirmation of the fate of the labeled cells in the presence of iron oxide. Therefore, both iron oxide-labeled living as well as dead epicardium-derived cells (EPDCs) were investigated in ischemic myocardium of immunodeficient non-obese diabetic (NOD)/acid: non-obese diabetic severe combined immunodeficient (NOD/scid) mice with 9.4T MRI until 6 weeks after surgery, at which time immunohistochemical analysis was performed. In both groups, voids on MRI scans were observed that did not change in number, size, or localization over time. Based on MRI, no distinction could be made between living and dead injected cells. Prussian blue staining confirmed that the hypointense spots on MRI corresponded to iron-loaded cells. However, in the dead-EPDC recipients, all iron-positive cells appeared to be macrophages, while the living-EPDC recipients also contained engrafted iron-loaded EPDCs. Iron labeling is inadequate for determining the fate of transplanted cells in the immunodeficient host, since dead cells produce an MRI signal indistinguishable from incorporated living cells.

Localization and origin of cardiac CD117-positive cells: identification of a population of epicardially-derived cells in adult human heart.

During heart morphogenesis, epicardial cells undergo epithelial-mesenchymal transition giving origin to a population of epicardially derived cells that play a crucial role in the development of most cardiac cell lineages. Considering the hypothesis that epithelial-mesenchymal transition of epicardial mesothelium can generate cardiac primitive cells in the adult heart, we have examined in vivo and in vitro the epicardium and subepicardium of normal human adult hearts and of pathological hearts from patients with chronic ischemic heart failure for the presence of CD117-positive cells with epithelial and mesenchymal markers expression. The number of CD117-positive cells increased significantly in the subepicardium of pathological hearts and sloped down towards myocardium, remaining still elevated with respect to normal hearts. While cells with typical epithelial proteins expression formed an intact layer on the surface of the normal hearts, CD117-positive cells were localized mainly in the subepicardium and expressed mesenchymal markers in the pathological hearts. Epithelial-mesenchymal transition, induced in vitro by several growth factors known to accumulate in the ischemic myocardium, gave origin to epicardially-derived cells with CD117 expression. These data support the hypothesis of epicardial origin of cardiac primitive cells in the adult human heart.

Genetic fate mapping demonstrates contribution of epicardium-derived cells to the annulus fibrosis of the mammalian heart

The annulus fibrosis electrically insulates the atria and ventricles, allowing the timed sequential beating of these structures that is necessary for efficient heart function. Abnormal development of the annulus fibrosis leads to persistence of accessory electrical pathways from atria to ventricles, providing the anatomical substrate for re-entrant cardiac arrhythmias such as Wolff-Parkinson-White syndrome. To better understand the development of the annulus fibrosis and the etiology of these cardiac arrhythmias, we used Cre-LoxP technology to assess the contribution of epicardium derived cells (EPDCs) to the annulus fibrosis. We found that EPDCs migrated into the region of the forming annulus fibrosis, marked by the protein periostin. These EPDCs also stained positive for procollagen I, suggesting that the EPDCs themselves synthesize proteins of the annulus fibrosis. To further test the hypothesis that EPDCs contribute to cells that synthesize the annulus fibrosis, we purified genetically marked EPDCs from the atrioventricular region and measured gene expression by quantitative PCR. These EPDCs were highly enriched for mRNAs encoding periostin, procollagen I, fibronectin I, vimentin, discoidin domain receptor 2, and tenascin C, markers of fibroblasts and components of the annulus fibrosis. In addition, these EPDCs were highly enriched for Snail, Smad1, Slug, and Twist1, markers for epithelial-to-mesenchymal transition (EMT), and a metalloprotease, Mmp2, that contributes to cellular migration. Our work provides for the first time definitive evidence that epicardium contributes to formation of the mammalian annulus fibrosis through EMT. Abnormalities of this differentiation process may underlie development of some forms of re-entrant atrioventricular tachycardia.

A New Direction for Cardiac Regeneration Therapy

Background— Adult human epicardium-derived cells (EPDCs), transplanted into the infarcted heart, are known to improve cardiac function, mainly through paracrine protection of the surrounding tissue. We hypothesized that this effect might be further improved if these supportive EPDCs were combined with cells that could possibly supply the ischemic heart with new cardiomyocytes. Therefore, we transplanted EPDCs together with cardiomyocyte progenitor cells that can generate mature cardiomyocytes in vitro. Methods and Results— EPDCs and cardiomyocyte progenitor cells were isolated from human adult atrial appendages, expanded in culture, and transplanted separately or together into the infarcted mouse myocardium (total cell number, 4�10 5 ). Cardiac function was determined 6 weeks later (9.4T MRI). Coculturing increased proliferation rate and production of several growth factors, indicating a mutual effect. Cotransplantation resulted in further improvement of cardiac function compared with single cell-type recipients ( P &lt;0.05), which themselves demonstrated better function than vehicle-injected controls ( P &lt;0.05). However, in contrast to our hypothesis, no graft-derived cardiomyocytes were observed within the 6-week survival, supporting that not only EPDCs but also cardiomyocyte progenitor cells acted in a paracrine manner. Because injected cell number and degree of engraftment were similar between groups, the additional functional improvement in the cotransplantation group cannot be explained by an increased amount of secreted factors but rather by an altered type of secretion. Conclusion— EPDCs and cardiomyocyte progenitor cells synergistically improve cardiac function after myocardial infarction, probably instigated by complementary paracrine actions. Our results demonstrate for the first time that synergistically acting cells hold great promise for future clinical regeneration therapy.

Epicardium-Derived Cells as Progenitors of Cardiac Fibroblasts: A Possible Role in Arrhythmogenic Right Ventricular Cardiomyopathy?

Epicardium-Derived Cells as Progenitors of Cardiac Fibroblasts: A Possible Role in Arrhythmogenic Right Ventricular Cardiomyopathy?

The Many Faces of Hypertrophic Cardiomyopathy: From Developmental Biology to Clinical Practice

Hypertrophic cardiomyopathy (HCM) is the most common genetic heart disease, characterized by complex pathophysiology, heterogeneous morphology, and variable clinical manifestations over time. Besides cardiac hypertrophy, the HCM phenotype is characterized by a host of manifestations, including mitral valve and subvalvar abnormalities, subaortic and mid-ventricular left ventricular (LV) obstruction, microvascular dysfunction, myocardial fibrosis, disarray, atrial remodeling, myocardial bridging of epicardial coronary arteries, LV apical aneurysms, and autonomic nervous system abnormalities. Such heterogeneous phenotype still lacks a comprehensive explanation, which cannot be accounted solely by genetic heterogeneity, despite the large number of genes and mutations involved. It is likely that pre-natal and acquired features deriving from the primary genetic defect interact with the environment to produce the final result evident in each patient. Based on novel insights provided by cardiac developmental biology, a common lineage ancestry of several HCM manifestations might be traced back to the pluripotent epicardium-derived cells, which early during heart development differentiate into interstitial fibroblasts, coronary smooth muscle cells, and atrio-ventricular endocardial cushions as mesenchymal cells. To date, the different faces of HCM have not been sufficiently liked or explained. We here attempt to address these issues by describing the various components of the disease, their origin, interaction, and clinical significance.

Look Who’s Talking

See related article, pages 431–441 Given the increasing prevalence of patients with congestive heart failure, there is a strong interest in identifying and isolating cardiac progenitor cells capable of differentiating into cardiomyocytes that could be used to repair a damaged, failing heart. Recent work in the zebrafish model system has suggested that the epicardium, the nonmyocyte epithelial layer of cells covering the heart, may play a critical role in the regeneration of injured myocardium.1 The epicardium has a distinct embryological origin from both the myocardium and endocardium. It arises from a mass of mesothelial cells termed the proepicardium, located on the wall of the embryonic pericardial cavity just dorsal and caudal to the developing heart.2 During early embryonic development, cells of the proepicardium migrate onto the heart at the atrioventricular junction and then spread over the surface of the heart to form a primitive epicardium. Some cells of the epicardium then undergo a mesenchymal transformation and invade into the subepicardial space. There these epicardial-derived cells further differentiate into cells that form the coronary vasculature as well as cardiac fibroblasts. Through this multistep differentiation pathway, the proepicardium serves as an important source for the noncardiomyocyte cellular component of the heart. Over the last few years, the pathways that regulate the development and differentiation of the proepicardium have begun to be revealed. First, the formation of the proepicardium …

Human epicardium: ultrastructural ancestry of mesothelium and mesenchymal cells

The human sub-epicardial area contains an unexplored cellular population under a layer of mesothelial cells. Transmission electron microscopy revealed the existence of interstitial Cajal-like cells (ICLCs), isolated smooth muscle cells (iSMC) and mesenchymal cells besides other well-known cells. The presence of iSMC in the sub-epicardial space is quite unique and could explain why epicardial-derived cells isolated from human epicardium generate smooth muscle cells in culture. Mesenchymal cells, guided by ICLCs, were found migrating from sub-epicardial area in the mesothelial layer. These findings suggest that epithelial-mesenchymal transition is not a common process involved in cardiac regeneration in vivo.

Landmarks and Lineages in the Developing Heart

See related article, pages 1267–1274 One of the overarching concepts underpinning mammalian embryonic development is that of “regulation.” This implies that the lineage potential of a particular cell is broader than its actual fate. Although the influential experiments in this area performed by Hans Driesch in the late 1800s concerned the fate of individual embryonic blastomeres, the concept of regulative development has pervaded all aspects of embryology, overlapping with the concept of regenerative fields. In the context of the heart, a good example is that of frog embryo cells fated to form the dorsal mesocardium and dorsal pericardial (splanchnic) mesoderm but which can “regulate” and form myocardial tissue if the normal heart is injured or extirpated.1 As we delve into the molecular mechanisms of developmental processes, we understand that limitations to cell fate are often set, particularly in the embryo, by geographical or environmental parameters, such as the source and strength of a secreted inductive signal, and these may change with time. Anatomic patterns or landmarks provide vital clues to how we should think about development and indeed disease. For example, segmentation of the embryonic paraxial mesoderm into repetitive units called somites, which bear one of the main progenitor populations for the musculoskeletal system, has been widely studied and it is known that somite segmentation is controlled by a network of synchronized molecular oscillators coupled by the Delta-Notch intercellular signaling system.2 The heart has also long been considered to have a segmental prepattern based, in part, on the series of swellings and constrictions that become evident during early heart tube formation and early fate-mapping experiments suggesting that these were the precursor structures of the chambers and their flanking regions, including the atrioventricular canal (AVC). This notion has been perpetuated as fact in embryology and medical textbooks with …

Developmental origins of hypertrophic cardiomyopathy phenotypes: a unifying hypothesis

The majority of genetic mutations associated with hypertrophic cardiomyopathy (HCM) occur in genes encoding sarcomeric proteins, which are expressed only in cardiomyocytes. However, some manifestations of the HCM phenotype, such as myocardial disarray, interstitial fibrosis, mitral valve abnormalities, and microvascular remodeling, indicate the involvement of other cell lineages. The link between sarcomeric gene defects and these 'extended' HCM phenotypes remains elusive. Based on novel insights provided by cardiac developmental biology, we propose that a common lineage ancestry of the diverse HCM phenotypes not involving the cardiomyocyte can be traced to the pluripotent epicardium-derived cells (EPDCs). During cardiac colonization, EPDCs differentiate into interstitial fibroblasts, coronary smooth-muscle cells, and atrioventricular endocardial cushions as mesenchymal cells. We propose that the cross-talk between healthy EPDCs and abnormally contracting cardiomyocytes might account for the diverse manifestations of HCM, by a putative mechanism of mechanotransduction leading to abnormal gene expression and differentiation.