Cardiac remodeling and contractile dysfunction are leading causes in hypertrophy-associated heart failure (HF), increasing with a population's rising age. A hallmark of aged and diseased hearts is the accumulation of modified proteins caused by an impaired autophagy-lysosomal-pathway. Although, autophagy inducer rapamycin has been described to exert cardioprotective effects, it remains to be shown whether these effects can be attributed to improved cardiomyocyte autophagy and contractility. In vivo hypertrophy was induced by transverse aortic constriction (TAC), with mice receiving daily rapamycin injections beginning six weeks after surgery for four weeks. Echocardiographic analysis demonstrated TAC-induced HF and protein analyses showed abundance of modified proteins in TAC-hearts after 10 weeks, both reduced by rapamycin. In vitro, cardiomyocyte hypertrophy was mimicked by endothelin 1 (ET-1) and autophagy manipulated by silencing Atg5 in neonatal cardiomyocytes. ET-1 and siAtg5 decreased Atg5-Atg12 and LC3-II, increased natriuretic peptides, and decreased amplitude and early phase of contraction in cardiomyocytes, the latter two evaluated using ImageJ macro Myocyter recently developed by us. ET-1 further decreased cell contractility in control but not in siAtg5 cells. In conclusion, ET-1 decreased autophagy and cardiomyocyte contractility, in line with siAtg5-treated cells and the results of TAC-mice demonstrating a crucial role for autophagy in cardiomyocyte contractility and cardiac performance.

3',5'-Cyclic adenosine monophosphate (cAMP) is a ubiquitous second messenger which plays critical roles in cardiac function and disease. In adult mouse ventricular myocytes (AMVMs), several distinct functionally relevant microdomains with tightly compartmentalized cAMP signaling have been described. At least two types of microdomains reside in AMVM plasma membrane which are associated with caveolin-rich raft and non-raft sarcolemma, each with distinct cAMP dynamics and their differential regulation by receptors and cAMP degrading enzymes phosphodiesterases (PDEs). However, it is still unclear how cardiac disease such as hypertrophy leading to heart failure affects cAMP signals specifically in the non-raft membrane microdomains. To answer this question, we generated a novel transgenic mouse line expressing a highly sensitive Förster resonance energy transfer (FRET)-based biosensor E1-CAAX targeted to non-lipid raft membrane microdomains of AMVMs and subjected these mice to pressure overload induced cardiac hypertrophy. We could detect specific changes in PDE3-dependent compartmentation of β-adrenergic receptor induced cAMP in non-raft membrane microdomains which were clearly different from those occurring in caveolin-rich sarcolemma. This indicates differential regulation and distinct responses of these membrane microdomains to cardiac remodeling.

Injuries, high altitude, and endurance exercise lead to hypoxic conditions in skeletal muscle and sometimes to hypoxia-induced local tissue damage. Thus, regenerative myoblasts/satellite cells are exposed to different levels and durations of partial oxygen pressure depending on the spatial distance from the blood vessels. To date, it is unclear how hypoxia affects myoblasts proliferation, differentiation, and particularly fusion with normoxic myoblasts. To study this, we investigated how 21% and 2% oxygen affects C2C12 myoblast morphology, proliferation, and myogenic differentiation and evaluated the fusion of normoxic- or hypoxic-preconditioned C2C12 cells in 21% or 2% oxygen in vitro. Out data show that the long-term hypoxic culture condition does not affect the proliferation of C2C12 cells but leads to rounder cells and reduced myotube formation when compared with myoblasts exposed to normoxia. However, when normoxic- and hypoxic-preconditioned myoblasts were differentiated together, the resultant myotubes were significantly larger than the control myotubes. Whole transcriptome sequencing analysis revealed several novel candidate genes that are differentially regulated during the differentiation under normoxia and hypoxia in mixed culture conditions and may thus be involved in the increase in myotube size. Taken together, oxygen-dependent adaption and interaction of myoblasts may represent a novel approach for the development of innovative therapeutic targets.

Light pollution worldwide promotes the progression of obesity, which is widely considered a consequence of circadian rhythm disruptions. However, the role of environmental light wavelength in mammalian obesity is not fully understood. Herein, mice fed a normal chow diet (NCD) or a high-fat diet (HFD) were exposed to daytime white (WL), blue (BL), green (GL), and red light (RL) for 8 weeks. Compared with WL and RL, BL significantly increased weight gain and white adipose tissue (WAT) weight, and it disrupted glucose homeostasis in mice fed with HFD but not NCD. The analysis of WAT found that BL significantly aggravated HFD-induced WAT hypertrophy, with a decrease in IL-10 and an increase in NLRP3, p-P65, p-IκB, TLR4, Cd36, Chrebp, Srebp-1c, Fasn, and Cpt1β relative to WL or RL. More interestingly, BL upregulated the expression of circadian clocks in the WAT, including Clock, Bmal1, Per1, Cry1, Cry2, Rorα, Rev-erbα, and Rev-erbβ compared with WL or RL. However, most of the changes had no statistical difference between BL and GL. Mechanistically, BL significantly increased plasma corticosterone (CORT) levels and glucocorticoid receptors in the WAT, which may account for the changes in circadian clocks. Further, in vitro study confirmed that CORT treatment did promote the expression of circadian clocks in 3T3-L1 cells, accompanied by an increase in Chrebp, Cd36, Hsp90, P23, NLRP3, and p-P65. Thus, daily BL, rather than RL exposure-induced CORT elevation, may drive changes in the WAT circadian clocks, ultimately exacerbating lipid dysmetabolism and adipocytic hypertrophy in the HFD-fed mice.

Cardiac hypertrophy is a common pathological condition and an independent risk factor that triggers cardiovascular morbidity. As an important epigenetic regulator, miRNA is widely involved in many biological processes. In this study, miRNAs expressed in rat hearts that underwent isoprenaline-induced cardiac hypertrophy were identified using high-throughput sequencing, and functional verification of typical miRNAs was performed using rat primary cardiomyocytes. A total of 623 miRNAs were identified, of which 33 were specifically expressed in cardiac hypertrophy rats. The enriched pathways of target genes of differentially expressed miRNAs included the FoxO signaling pathway, dopaminergic synapse, Wnt signaling pathway, MAPK (mitogen-activated protein kinase) signaling pathway, and Hippo signaling pathway. Subsequently, miR-144 was the most differentially expressed miRNA and was subsequently selected for in vitro validation. Inhibition of miR-144 expression in primary myocardial cells caused up-regulation of cardiac hypertrophy markers atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP). The dual luciferase reporter system showed that ANP may be a target gene of miR-144. Long non-coding RNA myocardial infarction associated transcript (LncMIAT) is closely related to heart disease, and here, we were the first to discover that LncMIAT may act as an miR-144 sponge in isoproterenol-induced cardiac hypertrophy. Taken together, these results enriched the understanding of miRNA in regulating cardiac hypertrophy and provided a reference for preventing and treating cardiac hypertrophy.

Pathological cardiac hypertrophy is one of the notable causes of heart failure. Circular RNAs (circRNAs) have been studied in association with cardiac hypertrophy; however, the mechanisms by which circRNAs regulate cardiac hypertrophy remain unclear. In this study, we identified a new circRNA, named circCacna1c, in cardiac hypertrophy. Adult male C57BL/6 mice and H9c2 cells were treated with isoprenaline hydrochloride (ISO) to establish a hypertrophy model. We found that circCacna1c was upregulated in ISO-induced hypertrophic heart tissue and H9c2 cells. Western blot and quantitative real-time polymerase chain reaction showed that silencing circCacna1c inhibited hypertrophic gene expression in ISO-induced H9c2 cells. Mechanistically, circCacna1c competitively bound to miR-29b-2-5p in a dual-luciferase reporter assay, which was downregulated in ISO-induced hypertrophic heart tissue and H9c2 cells. MiR-29b-2-5p inhibited the nuclear factor of activated T cells, cytoplasmic, calcineurin-dependent 1 (NFATc1) to control hypertrophic gene expression. After silencing circCacna1c, the expression of miR-29b-2-5p increased, which reduced hypertrophic gene expression by inhibiting NFATc1 expression. Together, these experiments indicate that circCacna1c promotes ISO-induced pathological hypertrophy through the miR-29b-2-5p/NFATc1 axis.

Cardiomyopathy has become one of the leading causes of death in patients with Duchenne muscular dystrophy (DMD). We recently reported that the inhibition of the interaction between the receptor activator of nuclear factor κB ligand (RANKL) and receptor activator of nuclear factor κB (RANK) significantly improves muscle and bone functions in dystrophin-deficient mdx mice. RANKL and RANK are also expressed in cardiac muscle. Here, we investigate whether anti-RANKL treatment prevents cardiac hypertrophy and dysfunction in dystrophic mdx mice. Anti-RANKL treatment significantly reduced LV hypertrophy and heart mass, and maintained cardiac function in mdx mice. Anti-RANKL treatment also inhibited NFκB and PI3K, two mediators implicated in cardiac hypertrophy. Furthermore, anti-RANKL treatment increased SERCA activity and the expression of RyR, FKBP12, and SERCA2a, leading possibly to an improved Ca2+ homeostasis in dystrophic hearts. Interestingly, preliminary post hoc analyses suggest that denosumab, a human anti-RANKL, reduced left ventricular hypertrophy in two patients with DMD. Taken together, our results indicate that anti-RANKL treatment prevents the worsening of cardiac hypertrophy in mdx mice and could potentially maintain cardiac function in teenage or adult patients with DMD.

Heart failure is a leading cause of death that develops subsequent to deleterious hypertrophic cardiac remodelling. MAPK pathways play a key role in coordinating the induction of gene expression during hypertrophy. Induction of the immediate early gene (IEG) response including activator protein 1 (AP-1) complex factors is a necessary and early event in this process. How MAPK and IEG expression are coupled during cardiac hypertrophy is not resolved. Here, in vitro, in rodent models and in human samples, we demonstrate that MAPK-stimulated IEG induction depends on the mitogen and stress-activated protein kinase (MSK) and its phosphorylation of histone H3 at serine 28 (pH3S28). pH3S28 in IEG promoters in turn recruits Brg1, a BAF60 ATP-dependent chromatin remodelling complex component, initiating gene expression. Without MSK activity and IEG induction, the hypertrophic response is suppressed. These studies provide new mechanistic insights into the role of MAPK pathways in signalling to the epigenome and regulation of gene expression during cardiac hypertrophy.

Disturbances in cardiac lipid metabolism are associated with the development of cardiac hypertrophy and heart failure. Spontaneously hypertensive rats (SHRs), a genetic model of primary hypertension and pathological left ventricular (LV) hypertrophy, have high levels of diacylglycerols in cardiomyocytes early in development. However, the exact effect of lipids and pathways that are involved in their metabolism on the development of cardiac dysfunction in SHRs is unknown. Therefore, we used SHRs and Wistar Kyoto (WKY) rats at 6 and 18 weeks of age to analyze the impact of perturbations of processes that are involved in lipid synthesis and degradation in the development of LV hypertrophy in SHRs with age. Triglyceride levels were higher, whereas free fatty acid (FA) content was lower in the LV in SHRs compared with WKY rats. The expression of de novo FA synthesis proteins was lower in cardiomyocytes in SHRs compared with corresponding WKY controls. The higher expression of genes that are involved in TG synthesis in 6-week-old SHRs may explain the higher TG content in these rats. Adenosine monophosphate-activated protein kinase phosphorylation and peroxisome proliferator-activated receptor α protein content were lower in cardiomyocytes in 18-week-old SHRs, suggesting a lower rate of β-oxidation. The decreased protein content of α/β-hydrolase domain-containing 5, adipose triglyceride lipase (ATGL) activator, and increased content of G0/G1 switch protein 2, ATGL inhibitor, indicating a lower rate of lipolysis in the heart in SHRs. In conclusion, the present study showed that the development of LV hypertrophy and myocardial dysfunction in SHRs is associated with triglyceride accumulation, attributable to a lower rate of lipolysis and β-oxidation in cardiomyocytes.

During osteoarthritis (OA), hypertrophy-like chondrocytes contribute to the disease process. TGF-β's signaling pathways can contribute to a hypertrophy(-like) phenotype in chondrocytes, especially at high doses of TGF-β. In this study, we examine which transcription factors (TFs) are activated and involved in TGF-β-dependent induction of a hypertrophy-like phenotype in human OA chondrocytes. We found that TGF-β, at levels found in synovial fluid in OA patients, induces hypertrophic differentiation, as characterized by increased expression of RUNX2, COL10A1, COL1A1, VEGFA and IHH. Using luciferase-based TF activity assays, we observed that the expression of these hypertrophy genes positively correlated to SMAD3:4, STAT3 and AP1 activity. Blocking these TFs using specific inhibitors for ALK-5-induced SMAD signaling (5 µM SB-505124), JAK-STAT signaling (1 µM Tofacitinib) and JNK signaling (10 µM SP-600125) led to the striking observation that only SB-505124 repressed the expression of hypertrophy factors in TGF-β-stimulated chondrocytes. Therefore, we conclude that ALK5 kinase activity is essential for TGF-β-induced expression of crucial hypertrophy factors in chondrocytes.

Cost- and time-intensive porcine translational disease models offer great opportunities to test drugs and therapies for pathological cardiac hypertrophy and can be supported by porcine cell culture models that provide further insights into basic disease mechanisms. Cardiac progenitor cells (CPCs) residing in the adult heart have been shown to differentiate in vitro into cardiomyocytes and could contribute to cardiac regeneration. Therefore, it is important to evaluate their changes on the cellular level caused by disease. We successfully isolated Isl1+Sca1+cKit+ porcine CPCs (pCPCs) from pig hearts and stimulated them with endothelin-1 (ET-1) and angiotensin II (Ang II) in vitro. We also performed a cardiac reprogramming transfection and tested the same conditions. Our results show that undifferentiated Isl1+Sca1+cKit+ pCPCs were significantly upregulated in GATA4, MEF2c, and miR-29a gene expressions and in BNP and MCP-1 protein expressions with Ang II stimulation, but they showed no significant changes in miR-29a and MCP-1 when stimulated with ET-1. Differentiated Isl1+Sca1+cKit+ pCPCs exhibited significantly higher levels of MEF2c, GATA4, miR-29a, and miR-21 as well as Cx43 and BNP with Ang II stimulation. pMx-MGT-transfected Isl1+Sca1+cKit+ pCPCs showed significant elevations in MEF2c, GATA4, and BNP expressions when stimulated with ET-1. Our model demonstrates that in vitro stimulation leads to successful Isl1+Sca1+cKit+ pCPC hypertrophy with upregulation of cardiac remodeling associated genes and profibrotic miRNAs and offers great possibilities for further investigations of disease mechanisms and treatment.

Dysbindin, a schizophrenia susceptibility marker and an essential constituent of BLOC-1 (biogenesis of lysosome-related organelles complex-1), has recently been associated with cardiomyocyte hypertrophy through the activation of Myozap-RhoA-mediated SRF signaling. We employed sandy mice (Dtnbp1_KO), which completely lack Dysbindin protein because of a spontaneous deletion of introns 5-7 of the Dtnbp1 gene, for pathophysiological characterization of the heart. Unlike in vitro, the loss-of-function of Dysbindin did not attenuate cardiac hypertrophy, either in response to transverse aortic constriction stress or upon phenylephrine treatment. Interestingly, however, the levels of hypertrophy-inducing interaction partner Myozap as well as the BLOC-1 partners of Dysbindin like Muted and Pallidin were dramatically reduced in Dtnbp1_KO mouse hearts. Taken together, our data suggest that Dysbindin's role in cardiomyocyte hypertrophy is redundant in vivo, yet essential to maintain the stability of its direct interaction partners like Myozap, Pallidin and Muted.

Epirubicin (EPI) is one of the most widely used anthracycline chemotherapy drugs, yet its cardiotoxicity severely limits its clinical application. Altered intracellular Ca2+ homeostasis has been shown to contribute to EPI-induced cell death and hypertrophy in the heart. While store-operated Ca2+ entry (SOCE) has recently been linked with cardiac hypertrophy and heart failure, its role in EPI-induced cardiotoxicity remains unknown. Using a publicly available RNA-seq dataset of human iPSC-derived cardiomyocytes, gene analysis showed that cells treated with 2 µM EPI for 48 h had significantly reduced expression of SOCE machinery genes, e.g., Orai1, Orai3, TRPC3, TRPC4, Stim1, and Stim2. Using HL-1, a cardiomyocyte cell line derived from adult mouse atria, and Fura-2, a ratiometric Ca2+ fluorescent dye, this study confirmed that SOCE was indeed significantly reduced in HL-1 cells treated with EPI for 6 h or longer. However, HL-1 cells presented increased SOCE as well as increased reactive oxygen species (ROS) production at 30 min after EPI treatment. EPI-induced apoptosis was evidenced by disruption of F-actin and increased cleavage of caspase-3 protein. The HL-1 cells that survived to 24 h after EPI treatment demonstrated enlarged cell sizes, up-regulated expression of brain natriuretic peptide (a hypertrophy marker), and increased NFAT4 nuclear translocation. Treatment by BTP2, a known SOCE blocker, decreased the initial EPI-enhanced SOCE, rescued HL-1 cells from EPI-induced apoptosis, and reduced NFAT4 nuclear translocation and hypertrophy. This study suggests that EPI may affect SOCE in two phases: the initial enhancement phase and the following cell compensatory reduction phase. Administration of a SOCE blocker at the initial enhancement phase may protect cardiomyocytes from EPI-induced toxicity and hypertrophy.

Intracellular Ca2+ overload secondary to chronic hemodynamic stimuli promotes the recruitment of Ca2+-dependent signaling implicated in cardiomyocyte hypertrophy. The present study tested the hypothesis that sympathetic-mediated hypertrophy of neonatal rat ventricular cardiomyocytes (NRVMs) translated to an increase in calcium influx secondary to the upregulation of CaV1.2 channel subunits. Confocal imaging of norepinephrine (NE)-treated NRVMs revealed a hypertrophic response compared to untreated NRVMs. L-type CaV1.2 peak current density was increased 4-fold following a 24-h stimulation with NE. NE-treated NRVMs exhibited a significant upregulation of CaVα2δ1 and CaVβ3 protein levels without significant changes of CaVα1C and CaVβ2 protein levels. Pre-treatment with the β1-blocker metoprolol failed to inhibit hypertrophy or CaVβ3 upregulation whereas CaVα2δ1 protein levels were significantly reduced. NE promoted the phosphorylation of ERK 1/2, and the response was attenuated by the β1-blocker. U0126 pre-treatment suppressed NE-induced ERK1/2 phosphorylation but failed to attenuate hypertrophy. U0126 inhibition of ERK1/2 phosphorylation prevented NE-mediated upregulation of CaVα2δ1, whereas CaVβ3 protein levels remained elevated. Thus, β1-adrenergic receptor-mediated recruitment of the ERK1/2 plays a seminal role in the upregulation of CaVα2δ1 in NRVMs independent of the concomitant hypertrophic response. However, the upregulation of CaVβ3 protein levels may be directly dependent on the hypertrophic response of NRVMs.

The cardiac expression of the mitochondrial uncoupling protein (UCP)-2 is increased in patients with heart failure. However, the underlying causes as well as the possible consequences of these alterations during the transition from hypertrophy to heart failure are still unclear. To investigate the role of UCP-2 mechanistically, expression of UCP-2 was silenced by small interfering RNA in adult rat ventricular cardiomyocytes. We demonstrate that a downregulation of UCP-2 by siRNA in cardiomyocytes preserves contractile function in the presence of angiotensin II. Furthermore, silencing of UCP-2 was associated with an upregulation of glucose transporter type (Glut)-4, increased glucose uptake, and reduced intracellular lactate levels, indicating improvement of the oxidative glucose metabolism. To study this adaptation in vivo, spontaneously hypertensive rats served as a model for cardiac hypertrophy due to pressure overload. During compensatory hypertrophy, we found low UCP-2 levels with an upregulation of Glut-4, while the decompensatory state with impaired function was associated with an increase of UCP-2 and reduced Glut-4 expression. By blocking the aldosterone receptor with spironolactone, both cardiac function as well as UCP-2 and Glut-4 expression levels of the compensated phase could be preserved. Furthermore, we were able to confirm this by left ventricular (LV) biopsies of patients with end-stage heart failure. The results of this study show that UCP-2 seems to impact the cardiac glucose metabolism during the transition from hypertrophy to failure by affecting glucose uptake through Glut-4. We suggest that the failing heart could benefit from low UCP-2 levels by improving the efficiency of glucose oxidation. For this reason, UCP-2 inhibition might be a promising therapeutic strategy to prevent the development of heart failure.

The C1q/tumor necrosis factor-alpha-related protein 9 (CTRP9) has been reported to exert cardioprotective effects, but its role in the right ventricle (RV) remains unclear. To investigate the role of CTRP9 in RV hypertrophy and failure, we performed pulmonary artery banding in weanling rats to induce compensatory RV hypertrophy seven weeks after surgery and RV failure 22 weeks after surgery. CTRP9 expression, signal transduction and mechanisms involved in protective CTRP9 effects were analyzed in rat and human RV tissue and cardiac cells. We demonstrate that CTRP9 was induced during compensatory RV hypertrophy but almost lost at the stage of RV failure. RV but not left ventricular (LV) cardiomyocytes or RV endothelial cells demonstrated increased intracellular reactive oxygen species (ROS) and apoptosis activation at this stage. Exogenous CTRP9 induced AMP-activated protein kinase (AMPK)-dependent transcriptional activation of the anti-oxidant thioredoxin-1 (Trx1) and superoxide dismutase-2 (SOD2) and reduced phenylephrine-induced ROS. Combined knockdown of adiponectin receptor-1 (AdipoR1) and AdipoR2 or knockdown of calreticulin attenuated CTRP9-mediated anti-oxidant effects. Immunoprecipitation showed an interaction of AdipoR1 with AdipoR2 and the co-receptor T-cadherin, but no direct interaction with calreticulin. Thus, CTRP9 mediates cardioprotective effects through inhibition of ROS production induced by pro-hypertrophic agents via AMPK-mediated activation of anti-oxidant enzymes.

Pathological cardiac remodeling correlates with chronic neurohumoral stimulation and abnormal Ca2+ signaling in cardiomyocytes. Store-operated calcium entry (SOCE) has been described in adult and neonatal murine cardiomyocytes, and Orai1 proteins act as crucial ion-conducting constituents of this calcium entry pathway that can be engaged not only by passive Ca2+ store depletion but also by neurohumoral stimuli such as angiotensin-II. In this study, we, therefore, analyzed the consequences of Orai1 deletion for cardiomyocyte hypertrophy in neonatal and adult cardiomyocytes as well as for other features of pathological cardiac remodeling including cardiac contractile function in vivo. Cellular hypertrophy induced by angiotensin-II in embryonic cardiomyocytes from Orai1-deficient mice was blunted in comparison to cells from litter-matched control mice. Due to lethality of mice with ubiquitous Orai1 deficiency and to selectively analyze the role of Orai1 in adult cardiomyocytes, we generated a cardiomyocyte-specific and temporally inducible Orai1 knockout mouse line (Orai1CM-KO). Analysis of cardiac contractility by pressure-volume loops under basal conditions and of cardiac histology did not reveal differences between Orai1CM-KO mice and controls. Moreover, deletion of Orai1 in cardiomyocytes in adult mice did not protect them from angiotensin-II-induced cardiac remodeling, but cardiomyocyte cross-sectional area and cardiac fibrosis were enhanced. These alterations in the absence of Orai1 go along with blunted angiotensin-II-induced upregulation of the expression of Myoz2 and a lack of rise in angiotensin-II-induced STIM1 and Orai3 expression. In contrast to embryonic cardiomyocytes, where Orai1 contributes to the development of cellular hypertrophy, the results obtained from deletion of Orai1 in the adult myocardium reveal a protective function of Orai1 against the development of angiotensin-II-induced cardiac remodeling, possibly involving signaling via Orai3/STIM1-calcineurin-NFAT related pathways.

During the development of hypertrophic cardiomyopathy, the heart returns to fetal energy metabolism where cells utilize more glucose instead of fatty acids as a source of energy. Metabolism of glucose can increase synthesis of the extracellular glycosaminoglycan hyaluronan, which has been shown to be involved in the development of cardiac hypertrophy and fibrosis. The aim of this study was to investigate hyaluronan metabolism in cardiac tissue from patients with hypertrophic cardiomyopathy in relation to cardiac growth. NMR and qRT-PCR analysis of human cardiac tissue from hypertrophic cardiomyopathy patients and healthy control hearts showed dysregulated glucose and hyaluronan metabolism in the patients. Gas phase electrophoresis revealed a higher amount of low molecular mass hyaluronan and larger cardiomyocytes in cardiac tissue from patients with hypertrophic cardiomyopathy. Histochemistry showed high concentrations of hyaluronan around individual cardiomyocytes in hearts from hypertrophic cardiomyopathy patients. Experimentally, we could also observe accumulation of low molecular mass hyaluronan in cardiac hypertrophy in a rat model. In conclusion, the development of hypertrophic cardiomyopathy with increased glucose metabolism affected both hyaluronan molecular mass and amount. The process of regulating cardiomyocyte size seems to involve fragmentation of hyaluronan.

While high levels of saturated fatty acids are associated with impairment of cardiovascular functions, n-3 polyunsaturated fatty acids (PUFAs) have been shown to exert protective effects. However the molecular mechanisms underlying this evidence are not completely understood. In the present study we have used rat H9c2 ventricular cardiomyoblasts as a cellular model of lipotoxicity to highlight the effects of palmitate, a saturated fatty acid, on genetic and epigenetic modulation of fatty acid metabolism and fate, and the ability of PUFAs, eicosapentaenoic acid, and docosahexaenoic acid, to contrast the actions that may contribute to cardiac dysfunction and remodeling. Treatment with a high dose of palmitate provoked mitochondrial depolarization, apoptosis, and hypertrophy of cardiomyoblasts. Palmitate also enhanced the mRNA levels of sterol regulatory element-binding proteins (SREBPs), a family of master transcription factors for lipogenesis, and it favored the expression of genes encoding key enzymes that metabolically activate palmitate and commit it to biosynthetic pathways. Moreover, miR-33a, a highly conserved microRNA embedded in an intronic sequence of the SREBP2 gene, was co-expressed with the SREBP2 messenger, while its target carnitine palmitoyltransferase-1b was down-regulated. Manipulation of the levels of miR-33a and SREBPs allowed us to understand their involvement in cell death and hypertrophy. The simultaneous addition of PUFAs prevented the effects of palmitate and protected H9c2 cells. These results may have implications for the control of cardiac metabolism and dysfunction, particularly in relation to dietary habits and the quality of fatty acid intake.

Reduced levels of the sensory nerve neuropeptide substance P (SP) have been reported in the diabetic rat heart, the consequence being a loss of cardioprotection in response to ischemic post-conditioning. We considered whether this loss of SP also predisposes the heart to non-ischemic diabetic cardiomyopathy in the form of fibrosis and hypertrophy. We report that diabetic Leprdb/db mice have reduced serum SP and that administration of exogenous replacement SP ameliorated cardiac fibrosis. Cardiac hypertrophy did not occur in Leprdb/db mice. Cardiac fibroblasts exposed to high glucose converted to a myofibroblast phenotype and produced excess extracellular matrix proteins; this was prevented by the presence of SP in the culture media. Cardiac fibroblasts exposed to high glucose produced increased amounts of the receptor for advanced glycation end products, reactive oxygen species and inflammatory cytokines, all of which were prevented by SP. Cultured macrophages assumed an M1 pro-inflammatory phenotype in response to high glucose as indicated by increased TNF-α, CCL2, and IL-6. SP promoted a shift to the reparative M2 macrophage phenotype characterized by arginase-1 and IL-10. Leprdb/db mice showed increased left ventricular M1 phenotype macrophages and an increase in the M1/M2 ratio. Replacement SP in Leprdb/db mice restored a favorable M1 to M2 balance. Together these findings indicate that a loss of SP predisposes the diabetic heart to developing fibrosis. The anti-fibrotic actions of replacement SP involve direct effects on cardiac fibroblasts and macrophages to oppose adverse phenotype changes. This study identifies the potential of replacement SP to treat diabetic cardiomyopathy.