Organization Of Cardiomyocytes Research Articles

A high-resolution cardiovascular magnetic resonance diffusion tensor map from ex-vivo C57BL/6 murine hearts.

BackgroundThe complex cardiac fiber structural organization and spatial arrangement of cardiomyocytes in laminar sheetlets contributes greatly to cardiac functional and contractile ejection patterns. This study presents the first comprehensive, ultra-high resolution, fully quantitative statistical tensor map of the fixed murine heart at isotropic resolution of 43 μm using diffusion tensor (DT) cardiovascular magnetic resonance (CMR).MethodsImaging was completed in approximately 12 hours using a six-directional encoding scheme, in five ex vivo healthy C57BL/6 mouse hearts. The tensor map constructed from this data provides an average description of the murine fiber architecture visualized with fiber tractography, and its population variability, using the latest advances in image tensor analysis and statistics.ResultsResults show that non-normalized cardiac tensor maps are associated with mean fractional anisotropy of 0.25 ± 0.07 and mean diffusivity of 8.9 ± 1.6 × 10−4 mm2/s. Moreover, average mid-ventricular helical angle distributions ranged between –41 ± 3° and +52 ± 5° and were highly correlated with transmural depth, in agreement with prior published results in humans and canines. Calculated variabilities of local myocyte orientations were 2.0° and 1.4°. Laminar sheet orientation variability was found to be less stable at 2.6°. Despite such variations, the murine heart seems to be highly structured, particularly when compared to canines and humans.ConclusionsThis tensor map has the potential to yield an accurate mean representation and identification of common or unique features of the cardiac myocyte architecture, to establish a baseline standard reference of DTI indices, and to improve detection of biomarkers, especially in pathological states or post-transgenetic modifications.

Extracellular matrix components of adipose derived stromal cells promote alignment, organization, and maturation of cardiomyocytes in vitro.

Adipose derived stromal cells (ADSC) are relevant therapeutic agents to treat myocardial infarction (MI) in clinical trials. Soluble factors secreted by ADSC, such as growth factors and cytokines, suppress inflammation and apoptosis while promoting angiogenesis and the proliferation of cardiomyocytes (CM). Moreover, ADSC synthesize extracellular matrix (ECM) components into the intercellular space which might contribute to their therapeutic capacity. Thus, ADSC might directly modulate the post-MI microenvironment through a combination of paracrine and juxtacrine signaling. In this study, the juxtacrine role of ADSC and ADSC-derived ECM on the organization and maturation of CM was investiagated. Human ADSC synthesized and deposited a heterogenous and complex mixture of ECM components such as collagen I, III, IV, fibronectin, elastin as well as the matricellular protein periostin. Cocultures of rodent CM with human ADSC or with human ADSC-derived ECM components enhanced the cardiomyocyte alignment, their intercellular connections and the maturation of their sarcomeres, while the proliferation rate of the CM was increased and their level of hypertrophy reduced. The use of human ADSC-derived ECM could serve both to augment in vitro tissue-engineered myocardial constructs and to improve myocardial remodeling after infarction.

Mechanosensitive kinases regulate stiffness-induced cardiomyocyte maturation.

Cells secrete and assemble extracellular matrix throughout development, giving rise to time-dependent, tissue-specific stiffness. Mimicking myocardial matrix stiffening, i.e. ~10-fold increase over 1 week, with a hydrogel system enhances myofibrillar organization of embryonic cardiomyocytes compared to static hydrogels, and thus we sought to identify specific mechanosensitive proteins involved. Expression and/or phosphorylation state of 309 unique protein kinases were examined in embryonic cardiomyocytes plated on either dynamically stiffening or static mature myocardial stiffness hydrogels. Gene ontology analysis of these kinases identified cardiogenic pathways that exhibited time-dependent up-regulation on dynamic versus static matrices, including PI3K/AKT and p38 MAPK, while GSK3β, a known antagonist of cardiomyocyte maturation, was down-regulated. Additionally, inhibiting GSK3β on static matrices improved spontaneous contraction and myofibril organization, while inhibiting agonist AKT on dynamic matrices reduced myofibril organization and spontaneous contraction, confirming its role in mechanically-driven maturation. Together, these data indicate that mechanically-driven maturation is at least partially achieved via active mechanosensing at focal adhesions, affecting expression and phosphorylation of a variety of protein kinases important to cardiomyogenesis.

Rac1 signaling is critical to cardiomyocyte polarity and embryonic heart development.

BackgroundDefects in cardiac septation are the most common form of congenital heart disease, but the mechanisms underlying these defects are still poorly understood. The small GTPase Rac1 is implicated in planar cell polarity of epithelial cells in Drosophila; however, its role in mammalian cardiomyocyte polarity is not clear. We tested the hypothesis that Rac1 signaling in the second heart field regulates cardiomyocyte polarity, chamber septation, and right ventricle development during embryonic heart development.Methods and ResultsMice with second heart field–specific deficiency of Rac1 (Rac1SHF) exhibited ventricular and atrial septal defects, a thinner right ventricle myocardium, and a bifid cardiac apex. Fate‐mapping analysis showed that second heart field contribution to the interventricular septum and right ventricle was deficient in Rac1SHF hearts. Notably, cardiomyocytes had a spherical shape with disrupted F‐actin filaments in Rac1SHF compared with elongated and well‐aligned cardiomyocytes in littermate controls. Expression of Scrib, a core protein in planar cell polarity, was lost in Rac1SHF hearts with decreased expression of WAVE and Arp2/3, leading to decreased migratory ability. In addition, Rac1‐deficient neonatal cardiomyocytes displayed defects in cell projections, lamellipodia formation, and cell elongation. Furthermore, apoptosis was increased and the expression of Gata4, Tbx5, Nkx2.5, and Hand2 transcription factors was decreased in the Rac1SHF right ventricle myocardium.ConclusionsDeficiency of Rac1 in the second heart field impairs elongation and cytoskeleton organization of cardiomyocytes and results in congenital septal defects, thin right ventricle myocardium, and a bifid cardiac apex. Our study suggests that Rac1 signaling is critical to cardiomyocyte polarity and embryonic heart development.

Scrib:Rac1 interactions are required for the morphogenesis of the ventricular myocardium.

AimsThe organization and maturation of ventricular cardiomyocytes from the embryonic to the adult form is crucial for normal cardiac function. We have shown that a polarity protein, Scrib, may be involved in regulating the early stages of this process. Our goal was to establish whether Scrib plays a cell autonomous role in the ventricular myocardium, and whether this involves well-known polarity pathways.Methods and resultsDeletion of Scrib in cardiac precursors utilizing Scribflox mice together with the Nkx2.5-Cre driver resulted in disruption of the cytoarchitecture of the forming trabeculae and ventricular septal defects. Although the majority of mice lacking Scrib in the myocardium survived to adulthood, they developed marked cardiac fibrosis. Scrib did not physically interact with the planar cell polarity (PCP) protein, Vangl2, in early cardiomyocytes as it does in other tissues, suggesting that the anomalies did not result from disruption of PCP signalling. However, Scrib interacted with Rac1 physically in embryonic cardiomyocytes and genetically to result in ventricular abnormalities, suggesting that this interaction is crucial for the development of the early myocardium.ConclusionsThe Scrib–Rac1 interaction plays a crucial role in the organization of developing cardiomyocytes and formation of the ventricular myocardium. Thus, we have identified a novel signalling pathway in the early, functioning, heart muscle. These data also show that the foetus can recover from relatively severe abnormalities in prenatal ventricular development, although cardiac fibrosis can be a long-term consequence.

P778Physiological and pharmacological characterization of human engineered heart tissue

Objective: Human induced pluripotent stem cell (iPS cell)-derived cardiomyocytes represent a valuable tool in cardiovascular research of tremendous potential. However, these cells are still immature and characterized by poor sarcomeric organization and cellular orientation in 2D cell culture. Therefore, the measurement of contractile force, the most important and best understood function of cardiomyocytes in vivo, is not established for these cells. This study describes the generation of three-dimensional, strip-format, force generating engineered heart tissues (EHTs) from human iPS cell-derived cardiomyocytes and presents a characterization of physiological and pharmacological parameters based on force development. Methods and results: Cardiomyocyte differentiation of human induced pluripotent cells was achieved by a growth factor-based three stage protocol. Strip-format EHTs were generated from dissociated cardiomyocytes in fibrin matrix between flexible silicone posts. Within two weeks after casting coherently beating human EHTs were formed and EHTs displayed a regular beating pattern for several weeks. Histological analysis revealed a high degree of sarcomeric organization and alignment of cardiomyocytes in EHTs. Functional analysis was performed by measuring force response to calcium concentration, pre-load, pacing frequency, beta-adrenergic and muscarinic agonists, modulators of sodium, calcium and potassium channels and revealed concentration-dependent effects. Comparison with native human heart tissue suggests an overall high level of similarity and minor differences. Conclusions: This study demonstrates feasibility to characterize human iPS cell-derived cardiomyocytes in EHTs by measuring contractile force. The analysis suggests high levels of similarity between EHTs and native human heart tissue. Human EHTs are a promising platform for automated toxicology screens in future drug development and for in vitro experiments on human cardiomyocytes in general.

Kidney Protection against Ischemia/Reperfusion Injury by Myofibrillogenesis Regulator-1

Background/Aims: Ischemia/reperfusion (I/R) injury is characterized by cytoskeletal reorganization and loss of polarity in proximal tubule epithelial cells. Previously, we showed that myofibrillogenesis regulator (MR)-1 promoted actin organization in cardiomyocytes. MR-1 is also expressed in the kidney. Methods: In this study, we investigated MR-1 expression in acute renal failure induced by I/R in Sprague-Dawley rats. We determined the MR-1 expression and the ratio of fibrous actin (F-actin) to globular actin (G-actin). HK-2 cells were treated with or without hypoxia/reoxygenation (H/R), and MR-1 levels were increased by adenoviral overexpression or silenced by RNA interference. Results: I/R and H/R resulted in cellular injury and decreases of MR-1, the F-/G-actin ratio, and myosin light chain (MLC)-2. MR-1 overexpression attenuated H/R-induced cell injury and loss of surface membrane polarity of actin. MR-1 overexpression also increased the expression and phosphorylation of MLC-2 and MLC kinase, which were decreased in MR-1-silenced and H/R-treated cells. Conclusion: Together, these data show that MR-1 promoted actin polarity on the membrane surface and protected HK-2 cells from H/R injury. The mechanism might involve the rapid organization of F-actin through the upregulation and phosphorylation of MLC-2.

Unchanged mitochondrial organization and compartmentation of high-energy phosphates in creatine-deficient GAMT−/−mouse hearts

Disruption of the creatine kinase (CK) system in hearts of CK-deficient mice leads to changes in the ultrastructure and regulation of mitochondrial respiration. We expected to see similar changes in creatine-deficient mice, which lack the enzyme guanidinoacetate methyltransferase (GAMT) to produce creatine. The aim of this study was to characterize the changes in cardiomyocyte mitochondrial organization, regulation of respiration, and intracellular compartmentation associated with GAMT deficiency. Three-dimensional mitochondrial organization was assessed by confocal microscopy. On populations of permeabilized cardiomyocytes, we recorded ADP and ATP kinetics of respiration, competition between mitochondria and pyruvate kinase for ADP produced by ATPases, ADP kinetics of endogenous pyruvate kinase, and ATP kinetics of ATPases. These data were analyzed by mathematical models to estimate intracellular compartmentation. Quantitative analysis of morphological and kinetic data as well as derived model fits showed no difference between GAMT-deficient and wild-type mice. We conclude that inactivation of the CK system by GAMT deficiency does not alter mitochondrial organization and intracellular compartmentation in relaxed cardiomyocytes. Thus, our results suggest that the healthy heart is able to preserve cardiac function at a basal level in the absence of CK-facilitated energy transfer without compromising intracellular organization and the regulation of mitochondrial energy homeostasis. This raises questions on the importance of the CK system as a spatial energy buffer in unstressed cardiomyocytes.

Cardiomyocyte architectural plasticity in fetal, neonatal, and adult pig hearts delineated with diffusion tensor MRI

Cardiomyocyte organization is a critical determinant of coordinated cardiac contractile function. Because of the acute opening of the pulmonary circulation, the relative workload of the left ventricle (LV) and right ventricle (RV) changes substantially immediately after birth. We hypothesized that three-dimensional cardiomyocyte architecture might be required to adapt rapidly to accommodate programmed perinatal changes of cardiac function. Isolated fixed hearts from pig fetuses or pigs at midgestation, preborn, postnatal day 1 (P1), postnatal day 5, postnatal day 14 (P14), and adulthood (n = 5 for each group) were acquired for diffusion-weighted magnetic resonance imaging. Cardiomyocyte architecture was visualized by three-dimensional fiber tracking and was quantitatively evaluated by the measured helix angle (α(h)). Upon the completion of MRI, hearts were sectioned and stained with hematoxylin/eosin (H&E) to evaluate cardiomyocyte alignment, with picrosirius red to evaluate collagen content, and with anti-Ki67 to evaluate postnatal cell proliferation. The helical architecture of cardiomyocyte was observed as early as the midgestational period. Postnatal changes of cardiomyocyte architecture were observed from P1 to P14, which primary occurred in the septum and RV free wall (RVFW). In the septum, the volume ratio of LV- vs. RV-associated cardiomyocytes rapidly changed from RV-LV balanced pattern at birth to LV dominant pattern by P14. In the RVFW, subendocardial α(h) decreased by ~30° from P1 to P14. These findings indicate that the helical architecture of cardiomyocyte is developed as early as the midgestation period. Substantial and rapid adaptive changes in cardiac microarchitecture suggested considerable developmental plasticity of cardiomyocyte form and function in the postnatal period in response to altered cardiac mechanical function.

Engineering of oriented myocardium on three-dimensional micropatterned collagen-chitosan hydrogel.

Surface topography and electrical field stimulation are important guidance cues that aid the organization and contractility of cardiomyocytes in vivo. We report here on the use of these biomimetic cues in vitro to engineer an implantable contractile cardiac tissue. Photocrosslinkable collagen-chitosan hydrogels with microgrooves of 10 µm, 20 µm and 100 µm in width were fabricated using polydimethylsiloxane (PDMS) molds. The hydrogels were seeded with cardiomyocytes, placed into a bioreactor array with the microgrooves aligned with the electrical field lines, and stimulated with biphasic square pulses at 1 Hz and 2.5 V/cm. At Day 6, cardiomyocytes were aligned in the direction of the microgrooves. When cultivated without electrical stimulation, the excitation threshold of engineered cardiac tissues using micropatterned hydrogels was significantly lower than using smooth hydrogels, thus showing the importance of cell alignment to cardiac function. The success rate of achieving beating constructs was higher with the application of electrical stimulation. In addition, formation of dense contractile cardiac organoids was observed in groups with both biomimetic cues. The cultivation of cardiomyocytes on hydrogels with 10 µm grooves yielded 100% beating tissues with or without electrical stimulation, thus suggesting a smaller groove width is necessary for cells to communicate and form proper gap junctions. However, electrical field stimulation further increased cell density and enhanced tissue morphology which may be essential for the integration of the tissue construct to the native heart tissue upon implantation. The biodegradability of the hydrogel substrate allows for the rapid translation of the engineered, oriented cardiac tissue to clinical applications.

Application of Regularized Richardson-Lucy Algorithm for Deconvolution of Confocal Microscopy Images

While confocal microscopes have considerably smaller contribution of out-of-focus light than widefield microscopes, the confocal images can still be enhanced by deconvolution if the optical and data acquisition effects are accounted for. Several deconvolution algorithms have been proposed for 3D microscopy. In this work we analyze the Richardson-Lucy iterative algorithm that is derived for Poisson noise and combined with total variation (TV) regularization. The influence of TV regularization on deconvolution process is determined by one parameter. However, the choice of regularization parameters is often unknown while it has considerable effect on the result of deconvolution process. The aims of this work were: to find good estimates of regularization parameter from the input; to develop an open source software package that would allow testing different deconvolution algorithms and that would be easy to use in practice. For that, we derived a formula to estimate this regularization parameter automatically from the images as the algorithm progresses. To assess the effectiveness of this algorithm, synthetic images were composed on the basis of confocal images of rat cardiomyocytes. From the analysis of deconvolved results, we have determined under which conditions our estimation of TV regularization parameter gives good results. The estimated TV regularization parameter can be monitored during deconvolution process and used as a stopping criterion. As a result, we propose a practical method to deconvolve confocal microscope images that uses estimated regularization parameter depending on the input image. We applied the deconvolution algorithm to study mitochondrial organization in rat cardiomyocytes. An open source software for deconvolving 3D images is available in http://sysbio.ioc.ee/software/.

3D Confocal Microscope Image Enhancement by Richardson-Lucy Deconvolution Algorithm with Total Variation Regularization: Parameters Estimation

Deconvolution is an efficient tool for enhancing both fluorescence and confocal microscopy images. Although in confocal microscopy the point spread function is rather small and images are much sharper compared to fluorescence microscopy, deconvolution can considerably improve image contrast and reduce noise.Several deconvolution methods have been proposed for 3D microscopy. In this work, we used Richardson-Lucy (RL) iterative algorithm assuming Poisson noise. In the presence of noise, the RL algorithm does not always give a optimal solution. To reduce the effects of noise, the RL algorithm is combined with total variation regularization. As it has been shown before, total variation (TV) with a carefully chosen regularization parameter reduces intensity oscillations in homogeneous areas and helps enhance contrast on edges.The aim of this work was to find good estimates to deconvolution algorithm parameters from the input to obtain optimal results. The test images generated from experimental data by smoothing out the noise but keeping typical structures. The analysis showed that optimal algorithm parameter Lambda is strongly correlating with the noise level. On the other hand finding the optimal parameter value is a very tedious process and so we derived formula estimating the value from the input image. The estimated Lambda turned out to give a more robust stopping criterion then the popular criterion using the threshold level for the relative change between two successive iteration steps.We applied the deconvolution algorithm to study mitochondrial organization in rat cardiomyocytes. An open source software for deconvolving 3D images is available in http://sysbio.ioc.ee/software/.

Mammalian Formin Fhod3 Regulates Actin Assembly and Sarcomere Organization in Striated Muscles

Actin filament assembly in nonmuscle cells is regulated by the actin polymerization machinery, including the Arp2/3 complex and formins. However, little is known about the regulation of actin assembly in muscle cells, where straight actin filaments are organized into the contractile unit sarcomere. Here, we show that Fhod3, a myocardial formin that localizes to thin actin filaments in a striated pattern, regulates sarcomere organization in cardiomyocytes. RNA interference-mediated depletion of Fhod3 results in a marked reduction in filamentous actin and disruption of the sarcomeric structure. These defects are rescued by expression of wild-type Fhod3 but not by that of mutant proteins carrying amino acid substitution for conserved residues for actin assembly. These findings suggest that actin dynamics regulated by Fhod3 are critical for sarcomere organization in striated muscle cells.

Developing Hearts Need Their SPEG

Myocardial formation is a particularly bad time to be missing building blocks and tools essential for myofibrillogenesis. Prenatal development and early postnatal development are characterized by intense cardiomyocyte formation and contractile apparatus assembly critical for normal growth and maturation. In this rapidly evolving milieu of cardiac formation, disturbances of cardiomyocyte organization or contractile performance usually have consequences leading to cardiomyopathic damage or lethality.1,2 Sensitivity of the developing heart to perturbation makes it a fertile playground for genetic engineering to demonstrate phenotypic effects resulting from assembling myocardium with missing or malfunctioning molecular pieces. Deliberate DNA recombination mischief with a knock-in mouse described by Liu et al3 now finds another way to jumble the puzzle by disruption of the striated preferentially expressed gene (SPEG). Loss of SPEG results in a postnatal dilated cardiomyopathic phenotype, combined with neonatal lethality, that indicates that appropriate myocardial development requires SPEG participation. These findings raise questions regarding what role SPEG fills in the construction of healthy myocardium and whether alterations in SPEG are linked to cardiomyopathic remodeling in the mature heart. Article p 261 SPEG is a member of a family of myosin light-chain kinases (MLCKs) that are required for cytoskeletal remodeling in myocytes. The MLCK domains of family members trace their evolutionary origins from mammals all the way back to Unc-89, an obscurin-MLCK found in Drosophila and Caenorhabditis elegans .4 Certain MLCK members are implicated in the regulation of the actin-based cytoskeleton and sarcomeric reorganization via phosphorylation of downstream targets such as myosin light-chain 2V. …

Non–Cell-Autonomous Roles for the Planar Cell Polarity Gene Vangl2 in Development of the Coronary Circulation

Establishment of cellular polarity is essential for the development of many tissues. In this study, we describe defects in the formation of the coronary vasculature in the loop-tail (Lp) mutant in which the planar cell polarity (PCP) gene, Vangl2, is disrupted. Although Vangl2 is expressed exclusively in the myocardial cells of the developing heart, the coronary vessels do not develop an intact smooth muscle layer, and there are enlarged, ectopic vessels on the surface of the heart. Reduced fibronectin deposition in the subepicardial space is associated with limited migration of epicardially derived cells (EPDCs) into the ventricular myocardium and likely contributes to these defects. Analysis of cardiomyocytes shows that the actin cytoskeleton is disrupted and the cytoarchitecture of the ventricular myocardium is abnormal in Lp/Lp hearts. Moreover, activation of RhoA/Rho kinase signaling is disrupted in these cells. Conditional inhibition of myocardial Rho kinase activity disrupts the organization of the cardiomyocytes and formation of the coronary vessels to produce the same spectrum of defects as seen in Lp. These data suggest that Vangl2 and Rho kinase act cell autonomously in the myocardium to regulate the organization of cardiomyocytes but also have non-cell-autonomous effects on the formation of the coronary vasculature.

Disruption of Planar Cell Polarity Signaling Results in Congenital Heart Defects and Cardiomyopathy Attributable to Early Cardiomyocyte Disorganization

The Drosophila scribble gene regulates apical-basal polarity and is implicated in control of cellular architecture and cell growth control. Mutations in mammalian Scrib (circletail; Crc mutant) also result in abnormalities suggestive of roles in planar cell polarity regulation. We show that Crc mutants develop heart malformations and cardiomyopathy attributable to abnormalities in cardiomyocyte organization within the early heart tube. N-Cadherin is lost from the cardiomyocyte cell membrane and cell-cell adhesion is disrupted. This results in abnormalities in heart looping and formation of both the trabeculae and compact myocardium, which ultimately results in cardiac misalignment defects and ventricular noncompaction. Thus, these late abnormalities arise from defects occurring at the earliest stages of heart development. Mislocalization of Vangl2 in Crc/Crc cardiomyocytes suggests Scrib is acting in the planar cell polarity pathway in this tissue. Moreover, double heterozygosity for mutations in both Scrib and Vangl2 can cause cardiac defects similar to those found in homozygous mutants for each gene but without other major defects. We propose that heterozygosity for mutations in different genes in the planar cell polarity pathway may be an important mechanism for congenital heart defects and cardiomyopathy in humans.

New insights into the molecular basis of desmoplakinand desmin-related cardiomyopathies

Desmosomes are intercellular adhesive complexes that anchor the intermediate filament cytoskeleton to the cell membrane in epithelia and cardiac muscle cells. The desmosomal component desmoplakin plays a key role in tethering various intermediate filament networks through its C-terminal plakin repeat domain. To gain better insight into the cytoskeletal organization of cardiomyocytes, we investigated the association of desmoplakin with desmin by cell transfection, yeast two-hybrid, and/or in vitro binding assays. The results indicate that the association of desmoplakin with desmin depends on sequences within the linker region and C-terminal extremity of desmoplakin, where the B and C subdomains contribute to efficient binding; a potentially phosphorylatable serine residue in the C-terminal extremity of desmoplakin affects its association with desmin; the interaction of desmoplakin with non-filamentous desmin requires sequences contained within the desmin C-terminal rod portion and tail domain in yeast, whereas in in vitro binding studies the desmin tail is dispensable for association; and mutations in either the C-terminus of desmoplakin or the desmin tail linked to inherited cardiomyopathy seem to impair desmoplakindesmin interaction. These studies increase our understanding of desmoplakin-intermediate filament interactions, which are important for maintenance of cytoarchitecture in cardiomyocytes, and give new insights into the molecular basis of desmoplakin- and desmin-related human diseases.

Mechanics of the Normal Heart

Even though studies on isolated papillary muscles and cardiomyocytes can be applied to the mechanics of a beating heart, it is not always easy for physicians to relate these findings to clinical medicine. Thus, it is important to extend the studies to intact heart either in simulations or in animal models and even better to validate the results with human subjects. Advances in engineering and computer technology have allowed us to bridge the gap between physiology and mechanics. Cardiomyocyte stress/strain relates to muscle energy expenditure, which dictates oxygen and substrate utilization. Appreciation of this sequential relationship by clinicians will facilitate the logical development and assessment of therapies. Theory of finite element analysis (FEA) can predict cardiac mechanics under normal and pathologic conditions. Imaging studies provide an avenue to relate these predictions indirectly to experimental studies. In this fashion, we can understand the mechanical basis for the micro- and macroanatomical twisting motion of the beating heart. The purposes of this manuscript are: (1) to examine the terms that are traditionally used to describe mechanical stresses and strain within the ventricle, (2) to explore the three-dimensional organization of cardiomyocytes that influences global ventricular function, (3) to apply mechanical measures to both single cardiomyofibrils and the intact ventricle (4) to evaluate mathematical and computer models used to characterize cardiac mechanics, and (5) to outline the clinical methods available to measure ventricular function and relate findings from FEA to pathologic conditions.

Trypanosoma cruzi disrupts myofibrillar organization and intracellular calcium levels in mouse neonatal cardiomyocytes

Immunofluorescence studies of normal and Trypanosoma cruzi-infected primary cultures of heart muscle cells were performed to gather information about the arrangement of myofibrillar components during the intracellular life cycle of this parasite. By using a panel of monoclonal antibodies against various myofibrillar proteins, a progressive disruption and loss of contractile proteins (such myosin and actin) of the host cell was detected during infection. The host cell formed a loose network of myofibrillar proteins around the parasites. Breakdown of the myofibrils occurred in regions where the parasites were present, and heavily infected cells showed myofibrillar proteins at their periphery. In parallel, we investigated the effect of T. cruzi infection on intracellular calcium levels by using a Ca2+ fluorescent indicator (confocal microscopy). Infected cardiomyocytes displayed a marked impairment in contractility, and calcium influxes became irregular and less intense when compared with those of non-infected cells. Our results demonstrate that T. cruzi infection dramatically affects calcium fluxes and causes myofibrillar breakdown disturbing cardiomyocyte contractility.

ENDOTHELIAL-CARDIOMYOCYTE INTERACTIONS IN CARDIAC DEVELOPMENT AND REPAIR

Communication between endothelial cells and cardiomyocytes regulates not only early cardiac development but also adult cardiomyocyte function, including the contractile state. In the normal mammalian myocardium, each cardiomyocyte is surrounded by an intricate network of capillaries and is next to endothelial cells. Cardiomyocytes depend on endothelial cells not only for oxygenated blood supply but also for local protective signals that promote cardiomyocyte organization and survival. While endothelial cells direct cardiomyocytes, cardiomyocytes reciprocally secrete factors that impact endothelial cell function. Understanding how endothelial cells communicate with cardiomyocytes will be critical for cardiac regeneration, in which the ultimate goal is not simply to improve systolic function transiently but to establish new myocardium that is both structurally and functionally normal in the long term.