Modulator Of Cardiac Hypertrophy Research Articles

Expression and localization of intermediate filament synemin isoforms in human pluripotent stem cells and their cardiac derivates

Synemin is an intermediate filament family involved in the cytoarchitecture of skeletal, cardiac and smooth muscle cells. In mammals, the SYNM gene encodes 3 isoforms: H (α, 170 kDa), M (β, 140 kDa) and L (Low, 41 kDa) through alternative splicing. In mouse, H and M isoform expression begins in the embryo while L isoform is only found in adult. Mouse embryonic pluripotent stem cells express M isoform as well as Oct-3/4 and SSEA-1. Two missense mutations have been recorded in patients with dilated cardiomyopathy and, α and β synemin levels increased in failing human hearts. Little is known about synemin expression in human induced pluripotent stem cells (hiPSC) and their cardiac derivatives. The aims are: (1) To characterize expression of synemin isoforms in hiPSC differentiation towards the cardiac lineage; (2) To analyze the impact of synemin in a human in vitro model of cardiac hypertrophy using hiPSC-derived cardiomyocytes (hiPSC-CMs). Expression of synemin isoforms was investigated in 2 lines of healthy donor-derived hiPSC and hiPSC-CMs by RT-qPCR, Western blot and immunofluorescent analysis. Synemin and α-actinin immunostaining was carried out on hiPSC-CMs treated 7 days with 200 nM AngII or 100 nM Iso following by an estimate of cell size and contraction measurements (ImageJ). Two different lines of healthy donor-derived hiPSC express the 3 isoforms of synemin at mRNA and protein levels. Synemin protein is detected in the cytoplasm and potentially in the nucleus of hiPSC expressing Nanog and Oct3/4. In hiPSC-CMs at 30 days of differentiation, expression of α, β and L isoforms increased with a fold change of 35, 80 and 114 respectively compared with hiPSC. Immunostaining reveals a cytoplasmic localization of synemin with striated pattern. An increase of synemin protein expression and nuclear localization was shown in hiPSC-CMs treated by hypertrophic agents. We showed for the first time that hiPSC and hiPSC-CMs express all known synemin isoforms. Moreover, subcellular localization of this protein change under hypertrophic conditions. Based on the literature, these data could suggest a potential role of synemin in cell motility, proliferation and also in the modulation of cardiac hypertrophy. To better understand these functions we have generated two synemin KO hiPSC lines using Dual-Guide CRISPR-Cas9 system of which characterization is ongoing.

Role of Known Transient Receptor Potential Vanilloid Channels in Modulating Cardiac Mechanobiology.

The transient receptor potential (TRP) channels have been described in almost every mammalian cell type. Several members of the Vanilloid (TRPV) subtype have been found to play important roles in modulating cardiac structure and function through Ca2+ handling in response to systemic and local mechanobiological cues. In this review, we will consider the most studied TRPV channels in the cardiovascular field; transient receptor potential vanilloid 1 as a modulator of cardiac hypertrophy; transient receptor potential vanilloid 2 as a structural and functional protein; transient receptor potential vanilloid 3 in the development of hypertrophy and myocardial fibrosis; and transient receptor potential vanilloid 4 in its roles modulating the fibrotic and functional responses of the heart to pressure overload. Lastly, we will also review the potential overlapping roles of these channels with other TRP proteins as well as the advances in translational and clinical arenas associated with TRPV channels.

Abstract 205: Mlip is a New Integrator of Physiological Stress Required For Proper Cardiac Adaptation

Aging and diseases are generally a result of the inability of tissues to properly adapt to stress. The heart is particularly vulnerable to disequilibrium in homeostasis as its regenerative capacities are limited. Many molecular players have been identified as cardiac gate-keepers and integrators of stress, all essential to preserve cardiac function. However, the molecular basis of the relationship between aging and the pathogenesis of cardiac dysfunction remains poorly understood. Muscle enriched A-type lamin interacting protein (MLIP) is a unique mammalian protein of unknown function that was recently identified through its interaction with A-type lamins in the heart and as a modulator of cardiac hypertrophy in vitro. Here we report that young Mlip deficient mice develop dilated cardiomyopathy, manifested by an increase in heart mass with reduced cardiac function. Through global gene expression profiling of the Mlip deficient hearts, we identified a deregulation in mTOR signalling, a key stress sensing canonical pathway. Analysis of mTOR regulatory proteins (AMPK and AKT) revealed hyperactivation of the mTOR pathway and a deregulated integration of these two stress/nutrient sensors. These data support the notion that Mlip deficient hearts have impaired cardiac adaptation due to deregulated mTOR activity resulting in maladaptive remodeling, and the development of dilated cardiomyopathy. These results are further supported by a genetic association between Mlip and early response to pro-hypertrophic stimulus. Collectively, these results demonstrate that Mlip is required for normal integration of physiological stress (postnatal cardiac growth, isoproterenol-induced hypertrophy) through the regulation of the AMPK/Akt/mTOR pathway to maintain cardiac homeostasis in the adult heart.

Quantitative Phosphoproteomic Study of Pressure-Overloaded Mouse Heart Reveals Dynamin-Related Protein 1 as a Modulator of Cardiac Hypertrophy

Pressure-overload stress to the heart causes pathological cardiac hypertrophy, which increases the risk of cardiac morbidity and mortality. However, the detailed signaling pathways induced by pressure overload remain unclear. Here we used phosphoproteomics to delineate signaling pathways in the myocardium responding to acute pressure overload and chronic hypertrophy in mice. Myocardial samples at 4 time points (10, 30, 60 min and 2 weeks) after transverse aortic banding (TAB) in mice underwent quantitative phosphoproteomics assay. Temporal phosphoproteomics profiles showed 360 phosphorylation sites with significant regulation after TAB. Multiple mechanical stress sensors were activated after acute pressure overload. Gene ontology analysis revealed differential phosphorylation between hearts with acute pressure overload and chronic hypertrophy. Most interestingly, analysis of the cardiac hypertrophy pathway revealed phosphorylation of the mitochondrial fission protein dynamin-related protein 1 (DRP1) by prohypertrophic kinases. Phosphorylation of DRP1 S622 was confirmed in TAB-treated mouse hearts and phenylephrine (PE)-treated rat neonatal cardiomyocytes. TAB-treated mouse hearts showed phosphorylation-mediated mitochondrial translocation of DRP1. Inhibition of DRP1 with the small-molecule inhibitor mdivi-1 reduced the TAB-induced hypertrophic responses. Mdivi-1 also prevented PE-induced hypertrophic growth and oxygen consumption in rat neonatal cardiomyocytes. We reveal the signaling responses of the heart to pressure stress in vivo and in vitro. DRP1 may be important in the development of cardiac hypertrophy.

Role of Renin Angiotensin System inhibitors on Toll‐like receptors expression in cardiac hypertrophy induced by renal ischemia/reperfusion

ObjectiveThe heart is a target organ of many regulatory systems that modulates heart trophism, such as cardiac renin‐angiotensin system (RAS). Literature data confirm the direct relationship between inflammatory responses, Toll‐Like receptors (TLR) and many cardiovascular diseases. This work aims to investigate the role of RAS inhibitors on TLR expression in a cardiac hypertrophy model induced by renal ischemia/reperfusion.MethodsC57bl/6 young, male mice (n=72) were treated with Losartan (Los ‐ 10 mg/kg/day) or Enalapril (Ena ‐ 40 mg/kg/day) for 5 days before or after surgery, and kept under observation for 12, 15 or 20 days to investigate the role of TLR in cardiac hypertrophy, evaluated by morphologic parameters and gene expression.ResultsOur model was able to induce cardiac hypertrophy in 12d mice that was prevented and reversed with Los and Ena administration based on ANF/α‐actin gene expression. TLR gene expression was evaluated in animal treated with Los and Ena, that were able to reduce TLR mRNA levels in 15d and 20d mice, respectively.ConclusionTaken together our data suggest an important role for local RAS as a modulator of cardiac hypertrophy through regulation of TLR expression. Financial Support: FAPESP and UFABC.

Reply to Kass and Takimoto: Cardiac hypertrophy without cGKI and PDE5 in cardiac myocytes

Concerning the letter by Kass and Takimoto (1) in PNAS regarding our recent publication (2), we would like to make the following comments. First, we agree that our results showing a “lack of importance” for cGMP-dependent protein kinase I (cGKI) as a modulator of cardiac hypertrophy only refers to the “basal” condition as we clearly stated in the manuscript (1, 2). We purposely did not use guanylyl cyclase stimulators or phosphodiesterase (PDE) inhibitors in these studies. However, we do note that, particularly under the stress conditions of isoproterenol infusion or aortic constriction, atrial natriuretic peptide is substantially increased. This provides substantial drive on cGMP synthesis. It is possible that cGKI needs to be induced by very high cGMP to have protective effects, although induction is not a requirement for the importance of this kinase in other tissues. The point here is that ablation of cGKI from cardiomyocytes and most other tissues (except vasculature) does not potentiate the ability of transverse aortic constriction (TAC) or isoproterenol (ISO) to cause hypertrophy. To us, this is an observation that needs to be reconciled with current dogma. The degrees of hypertrophy obtained in our studies were equal to or even higher than those obtained in many other studies.

Calcineurin and cardiac function: is more or less better for the heart?

calcineurin (cn) is a Ca2+-calmodulin-activated serine-threonine phosphatase that is ubiquitously expressed and plays an important role in transducing Ca2+-dependent signals. Cn is a heterodimer composed of calmodulin binding catalytic subunit A (CnA) and a Ca2+ binding regulatory subunit B (CnB) ([

RALT Specifically Halts Maladaptive Cardiac Hypertrophy

The adult heart enlarges and increases in weight in response to ischemic stress and hypertension, a process termed cardiac hypertrophy. However, the heart also increases in size after regular physiological exercise. Cardiac hypertrophy is recognized as an important process in the development of heart failure. Initial results pointed to a clear signaling pattern of physiological hypertrophy after training versus pathological hypertrophy after damage. Although the signaling mechanisms involved have been studied for more than a decade, there is no clear distinction at the molecular level of physiological and pathological hypertrophy.1 Rather, the balance between signaling pathways preventing deterioration and pathways activated in association with preserving left ventricular function is regarded as critical. Accordingly, the terms adaptive and maladaptive were introduced to reflect these processes, respectively.2 A potential treatment avenue could be to selectively identify pathways involved in maladaptive hypertrophy that could be blocked. Alternatively, enhancing signaling pathways specifically involved in adaptive hypertrophy might allow us to conserve left ventricular function. Cai et al have identified a new target in cardiac hypertrophy: the receptor-associated late transducer (RALT).3 RALT appears to be a specific antagonist of maladaptive hypertrophy (Figure). Mechanistically, RALT functions as a negative regulator of epidermal growth factor receptor-mediated signaling not only in the heart but in other organs as well. The investigators used angiotensin II and isoproterenol as stimuli for stress-induced, maladaptive left ventricular remodeling in transgenic mice overexpressing RALT under the control of the α-myosin heavy …

N‐terminal pro‐B‐type natriuretic peptide in patients with growth hormone disturbances

Acromegaly is associated with hypertrophic cardiomyopathy, hypertension and subsequent congestive heart failure. Impairment of cardiac function has also been associated with growth hormone deficiency (GHD). B-type natriuretic peptides (BNPs) have emerged as strong diagnostic and prognostic risk markers. They are cardioprotective hormones that compensate heart disease by promoting natriuresis and modulation of cardiac hypertrophy in response to volume expansion and ventricular wall stretch. To investigate N-terminal pro-BNP (NT-proBNP) in patients with GH disturbances before and after treatment, and in healthy gender- and age-matched controls. Ten acromegalic patients (age 48 +/- 12 years) and 10 patients with GHD (age 41 +/- 14 years) were studied. None had symptoms or signs of cardiovascular disease except for hypertension. Serum NT-proBNP was measured before and 3, 6, 12 and 24 months after treatment. Baseline NT-proBNP was lower in acromegalic patients [median (interquartile range) 24.3 (17.8-33.0) pg/ml] than in 20 healthy matched controls [57.9 (35.4-92.2) pg/ml; P < 0.001]. NT-proBNP increased during treatment (P = 0.002), concomitant with a decrease in IGF-I (P < 0.001). After 3 months of treatment NT-proBNP peaked, with a fourfold increase to 96.4 (57.8-113.2) pg/ml. NT-proBNP did not differ in patients with GHD compared with controls (P = 0.19), and did not change during treatment (P = 0.39). Untreated patients with acromegaly had low NT-proBNP levels that increased fourfold after treatment. This is in contrast to other conditions with cardiac hypertrophy. Further studies are required to determine the significance of this novel finding in relation to acromegalic cardiac disease.

Correlation between left ventricular hypertrophy and GAA trinucleotide repeat length in Friedreich's ataxia.

Friedreich's ataxia (FA), the most common inherited ataxia, is associated frequently with cardiac hypertrophy, and death is often cardiac related. Recently, the disease has been associated with a mutation that consists of an unstable expansion of GAA repeats in the first intron of the gene encoding frataxin on chromosome 9. We studied 44 consecutive patients with FA, determined the size of GAA expansions in the frataxin gene, and examined the relation between the genotype and cardiac phenotype assessed by M-mode and two-dimensional echocardiography. All the patients were homozygous for the mutation. The size of the GAA expansion on the smaller allele varied from 270 to 1200. We found a correlation between the size of GAA expansion and the left ventricular wall thickness (r = .51, P < .001) and the left ventricular mass index (r = .45, P = .002). Left ventricular hypertrophy was observed in 81% of patients with a number of GAA repeats above the median value of 770 compared with only 14% in the other group (P = .002). These data demonstrate that in FA, the severity of left ventricular hypertrophy is related to the number of GAA repeats. These results suggest that abnormalities of the gene encoding frataxin, a protein of unknown function highly expressed in the normal heart, may play an important role in the modulation of cardiac hypertrophy.

Modulation of cardiac hypertrophy by estrogens.

Gender-specific differences in heart disease have long been known but it has only been since the advent of molecular biology that it has become possible to investigate the molecular mechanisms. Most biochemical work in the last 50 years has focused on the characterization of the steroid hormones involved in gender specificity. More recently, the cloning of the steroid receptors and characterization of the signaling pathways through these proteins has given new insights into the mechanisms underlying the mode of action of steroid hormones. It has also become clear that the steroid receptors can be classified into families (receptors for thyroid hormone, glucocorticoids, estrogens, androgens, retinoic acid, and so called orphan receptors of mostly unknown function). The structures of these receptors show very close resemblance and all are DNA-binding proteins acting as transcription factors. Some (if not all) act as repressors of transcription of some genes in the native state and are converted to activators (or perhaps repressors of other genes) upon binding of the cognate hormone. Naturally, classical target tissues for estrogens and androgens have been studied first and only in very recent years has it been recognized that estrogens and androgens act on a much wider spectrum of tissues. In the cardiovascular field, the beneficial effect of estrogen replacement therapy in postmenopausal women which reduces the incidence of cardiovascular disease by some 40% and the lower incidence of cardiovascular disease in premenopausal women have mostly been explained by the beneficial action of estrogens on the lipid profile (increase in HDL and decrease in LDL cholesterol). Recently, functional estrogen receptors have also been shown in vascular smooth muscle cells and in the endothelium. Our own group has characterized the presence of estrogen receptors in the myocardium and in cardiac fibroblasts. We have also shown that these receptors are transcriptionally active because they are able to drive a minigene composed of a triple estrogen responsive DNA regulatory element (promoter) coupled to the firefly luciferase gene which serves as a reporter by way of its ability to drive a light-emitting reaction. We are in the process of characterizing the target genes for estrogen in the myocardium. A specific series of immediate-early genes is induced by estradiol (the major premenopausal estrogen) and we have also characterized a number of tissue-specific genes whose expression is driven by estrogens in the myocardium. The ultimate goal of these investigations is to explore the use of estrogens in the treatment of cardiac hypertrophy (and failure) by way of their properties to counteract (at least some of) the pathological switches in gene expression in these disease entities.