Signaling Pathway In Cardiomyocytes Research Articles

1,25-Vitamin D3 protects against cooking oil fumes-derived PM2.5-induced cell damage through its anti-inflammatory effects in cardiomyocytes

The functional role of 1,25-vitamin D3 in cooking oil fumes (COFs)-derived PM2.5-induced cell damage is largely unexplored. The present study investigated the protective role of 1,25-vitamin D3 against cell injury by possible involvement of JAK/STAT and NF-κB signaling pathways in cardiomyocytes. Cell viability was measured using CCK-8 assay, and cell apoptosis was analyzed by flow cytometry, qRT-PCR and Western blot in cultured rat neonatal cardiomyocytes treated with 1,25-vitamin D3 and COFs-derived PM2.5. Expressions of JAK/STAT and NF-κB signaling pathway were measured by Western blot. The results suggested that treatment with COFs-derived PM2.5 significantly decreased cell viability and increased apoptosis and oxidative stress in cultured rat neonatal cardiomyocytes. 1,25-vitamin D3 pretreatment alleviated the cell injury by increasing cell viability and decreasing apoptosis in the cardiomyocytes. 1,25-vitamin D3 pretreatment also decreased the ROS level and inflammation in the cardiomyocytes. Furthermore, 1,25-vitamin D3 pretreatment alleviated COFs-derived PM2.5-evoked elevation of JAK/STAT and NF-κB signaling pathways. Our study showed that 1,25-vitamin D3 pretreatment protected cardiomyocytes from COFs-derived PM2.5-induced injury by decreasing ROS, apoptosis and inflammation level via activations of the JAK/STAT and NF-κB signaling pathways.

Irisin ameliorates angiotensin II-induced cardiomyocyte apoptosis through autophagy.

Cardiac hypertrophy is the main cause of heart failure and sudden death in patients. But the pathogenesis is unclear. Angiotensin II may contribute to cardiac hypertrophy in response to pressure overload. In angiotensin II-treated cardiomyocytes, there is a larger cross-sectional area, more apoptosis cells, and a reduction of irisin expression. An increase in P62, an autophagy flux index, as well as LC3II, were observed in cardiomyocytes after angiotensin II-induced injury. Surprisely, irisin supplementation increased LC3II expression and decreased P62 expression, consisted of results of RFP-GFP-LC3B adenovirus transfection, and reduced cardiomyocyte apoptosis, meanwhile, the protection of irisin was reversed by the autophagy inhibitor 3-methyladenine. In animal experiments, overexpression of irisin reduced cardiomyocyte apoptosis and alleviated myocardial hypertrophy caused by pressure overload. The above results indicate that irisin-induced protective autophagy and alleviated the apoptosis signaling pathway in cardiomyocytes, consequently reducing cardiomyocyte apoptosis after angiotensin II-induced injury. Hence, increasing irisin expression may be a new way to improve cardiac function and quality of life in patients with cardiac hypertrophy.

Disruption of Planar Cell Polarity Pathway Attributable to Valproic Acid-Induced Congenital Heart Disease through Hdac3 Participation in Mice.

Background:Valproic acid (VPA) exposure during pregnancy has been proven to contribute to congenital heart disease (CHD). Our previous findings implied that disruption of planar cell polarity (PCP) signaling pathway in cardiomyocytes might be a factor for the cardiac teratogenesis of VPA. In addition, the teratogenic ability of VPA is positively correlated to its histone deacetylase (HDAC) inhibition activity. This study aimed to investigate the effect of the VPA on cardiac morphogenesis, HDAC1/2/3, and PCP key genes (Vangl2/Scrib/Rac1), subsequently screening out the specific HDACs regulating PCP pathway.Methods:VPA was administered to pregnant C57BL mice at 700 mg/kg intraperitoneally on embryonic day 10.5. Dams were sacrificed on E15.5, and death/absorption rates of embryos were evaluated. Embryonic hearts were observed by hematoxylin-eosin staining to identify cardiac abnormalities. H9C2 cells (undifferentiated rat cardiomyoblasts) were transfected with Hdac1/2/3 specific small interfering RNA (siRNA). Based on the results of siRNA transfection, cells were transfected with Hdac3 expression plasmid and subsequently mock-treated or treated with 8.0 mmol/L VPA. Hdac1/2/3 as well as Vangl2/Scrib/Rac1 mRNA and protein levels were determined by real-time quantitative polymerase chain reaction and Western blotting, respectively. Total HDAC activity was detected by colorimetric assay.Results:VPA could induce CHD (P < 0.001) and inhibit mRNA or protein expression of Hdac1/2/3 as well as Vangl2/Scrib in fetal hearts, in association with total Hdac activity repression (all P < 0.05). In vitro, Hdac3 inhibition could significantly decrease Vangl2/Scrib expression (P < 0.01), while knockdown of Hdac1/2 had no influence (P > 0.05); VPA exposure dramatically decreased the expression of Vanlg2/Scrib together with Hdac activity (P < 0.01), while overexpression of Hdac3 could rescue the VPA-induced inhibition (P > 0.05).Conclusion:VPA could inhibit Hdac1/2/3, Vangl2/Scrib, or total Hdac activity both in vitro and in vivo and Hdac3 might participate in the process of VPA-induced cardiac developmental anomalies.

Relaxin protects cardiomyocytes against hypoxia-induced damage in in-vitro conditions: Involvement of Nrf2/HO-1 signaling pathway

Relaxin, a peptide hormone has emerged as a cardioprotective agent against the heart failure and has been found to protect cardiac muscle cells against hypoxia/reoxygenation injury under in vitro conditions. The present study was conducted to study its possible role in activating the Nrf2/HO-1 signaling pathway in cardiomyocytes, as a means to counter hypoxia associated oxidative damage and cell death. H9C2 cell line was induced with chemical hypoxia alone or together with relaxin. Hypoxia associated cellular damage and reactive oxygen species (ROS) production was accessed by Lactate dehydrogenase (LDH) release and DCFDA activity respectively. The anti-oxidative property of RLXH2 was measures by assessing the activities of different enzymes like Superoxide dismutase (SOD), Catalase (CAT) and Glutathione peroxidase (GSX). Expression levels of Nrf2 and HO-1 was studied by immunoblotting and quantitative real time PCR (qRT-PCR). Translocation of Nrf2 to nucleus was studied by immunoblotting. Our results found that hypoxia associated lactate dehydrogenase leakage and ROS production is countered by RLXH2 treatment. Similarly, RLXH2 was able to counter hypoxia induced oxidative damage as evident by increased activities of SOD, CAT and GSX. Furthermore, it was found that RLXH2 treatment induces translocation of Nrf2 from cytosol to nucleus and in turn enhances expression level of HO-1. Our results suggest that RLXH2 exerts cytoprotective action in cardiomyocytes against the hypoxia induced damage and activates Nrf2/HO-1 signaling pathway.

Blockade of RBP-J-Mediated Notch Signaling Pathway Exacerbates Cardiac Remodeling after Infarction by Increasing Apoptosis in Mice.

Background Ischemic heart disease (IHD) is the major cause of death in patients with cardiovascular disease. Cardiac remodeling is a common pathological change following myocardial infarction (MI), and cardiomyocyte apoptosis plays a key role in this change. Transcription factor recombination signal-binding protein-J (RBP-J)-mediated Notch signaling pathway has been implicated in several inherited cardiovascular diseases, including aortic valve diseases, but whether the RBP-J-mediated Notch signaling pathway plays a role in cardiomyocyte apoptosis after MI is unclear. Method We crossed RBP-Jfl/fl mice and Myh6-Cre/Esr1 transgenic mice to delete RBP-J in vivo and to partly inhibit the canonical Notch signaling pathway. MI was induced in mice by permanent ligation of the left anterior descending coronary artery followed by the knockout of RBP-J. Cardiac function and morphology were assessed by echocardiography and histological analysis 4 weeks after infarction. In addition, the expression and regulation of apoptosis-related molecules were examined by real time PCR and western blot. Results RBP-J knockout decreased the survival rate and deteriorated post-MI remodeling and function in mice, and this effect was associated with increased cardiomyocyte apoptosis. The potential mechanisms might be related to the downregulated expression of bcl-2, upregulated expression of bax, and cleaved-caspase 3 to exacerbate cardiomyocyte apoptosis. Conclusion These findings show that the RBP-J-mediated Notch signaling pathway in cardiomyocytes limits ventricular remodeling and improves cardiac function after MI. The RBP-J-mediated Notch signaling pathway has a protective role in cardiomyocyte apoptosis following cardiac injury.

Actinidiachinensis planch polysaccharide protects against hypoxia‑induced apoptosis of cardiomyocytes invitro.

Cardiac hypertrophy is frequently accompanied by ischemic heart disease. Actinidia chinensis planch polysaccharide (ACP) is the main active compound from Actinidia chinensis planch. In the present study, a cardiac hypertrophy model was produced by treating cells with Angiotensin II (Ang II), which was used to investigate whether ACP protected against cardiac hypertrophy in vitro. It was demonstrated that ACP alleviated Ang II-induced cardiac hypertrophy. In addition, pretreatment with ACP prior to hypoxic culture reduced the disruption of the mitochondrial membrane potential as investigated by flow cytometry. Cell Counting kit-8 analysis demonstrated that ACP maintained the cell viability of cardiomyocytes. The flow cytometric analysis revealed that ACP inhibited hypoxia-induced apoptosis in cardiomyocytes treated with Ang II. Additionally, reverse transcription-polymerase chain reaction and western blotting assays demonstrated that ACP decreased the expression of apoptosis-associated genes including apoptosis-inducing factor mitochondria associated 1, the cysteinyl aspartate specific proteinases caspases-3/8/9, and cleaved caspases-3/8/9. The results of the present study also demonstrated that ACP inhibited the activation of the extracellular signal-regulated kinase 1/2 (ERK1/2) and phosphoinositide 3-kinase/protein kinase B (PI3K/AKT) signaling pathways. Furthermore, the specific activation of ERK1/2 and PI3K/AKT reversed the apoptotic-inhibitory effect of ACP. In conclusion, the protective effects of ACP against hypoxia-induced apoptosis may depend on depressing the ERK1/2 and PI3K/AKT signaling pathways in cardiomyocytes treated with Ang II.

Protective effect of miR378* on doxorubicin‐induced cardiomyocyte injury via calumenin

Doxorubicin (Dox) is a highly effective antitumor antibiotic, however myocardial toxicity severely limits its use clinically. The pathogenesis of doxorubicin‐induced cardiomyopathy is unclear. In Dox cardiomyopathy mice, there is a decline in cardiac function, a change in myocardial pathology and a reduction in miR378* expression. Expression changes in calumenin, an endoplasmic reticulum stress (ERS) chaperone protein and pathway factor, as well as apoptosis, were observed in cardiomyocytes after doxorubicin‐induced injury. However, miR378* increased calumenin expression, eased ERS, and reduced cardiomyocyte apoptosis, while, silencing miR378* reduced calumenin expression, aggravated ERS, and increased cardiomyocyte apoptosis. The above results indicate that miR378* alleviates ERS and inhibits the activation of the ERS‐mediated apoptosis signaling pathway in cardiomyocytes via regulating calumenin expression, thereby reducing cardiomyocyte apoptosis after doxorubicin‐induced injury. Increasing miR378* expression may be a new way to improve cardiac function and quality of life in patients with Dox cardiomyopathy.

Corin protects H2O2-induced apoptosis through PI3K/AKT and NF-κB pathway in cardiomyocytes

BackgroundThe functional role of corin in H2O2-induced apoptosis is largely unexplored. The present study investigated the protective role of corin against cell injury by possible involvement of PI3K/AKT and NF-kB signaling pathways in cardiomyocytes. MethodCardiomyocytes H9c2 and HL-1 cells were used in the study. Cell viability was measured using CCK-8 assay; cell apoptosis was analyzed by flow cytometry, TUNEL assay, and western blot; and cell migration was measured using wound healing assay. The fluorescent intensities of reactive oxygen species (ROS) were measured using a flow cytometer. Quantitative RT-PCR was used to measure the mRNA expression of corin. Western blot was used to measure the protein expression of corin, apoptosis-related proteins (Bax, cleaved-Caspase-3 and -9), and PI3K/AKT and NF-κB signaling pathway proteins. ResultsTreatment with H2O2 (150μM, 6h) significantly decreased cell viability and relative migration, increased apoptosis, and decreased the expression of corin in H9c2 and HL-1 cells. Overexpression of corin alleviated the H2O2-induced cell injury by increasing cell viability and migration and decreasing apoptosis in the cardiomyocytes. Overexpression of corin also decreased the ROS level in the cardiomyocytes likely through upregulating HIF-1α. These effects of corin on the cell injury might be mediated via the corin-induced activations of PI3K/AKT and NF-κB signaling pathways. ConclusionOverexpression of corin protected cardiomyocytes from H2O2-induced injury by decreasing apoptosis and ROS level via activations of the PI3K/AKT and NF-κB signaling pathways and upregulating HIF-1α.

Abstract P224: β-arrestin2 Stimulates SERCA2a SUMOylation and Activity in Cardiac Myocytes to Promote Contractility

Background: Heart failure (HF) is the most lethal disease worldwide and new treatments are needed. The β 1 -adrenergic receptor (AR) mediates the positive inotropy of catecholamines, partly via Sarco(Endo)plasmic Reticulum Ca 2+ -ATPase (SERCA)-2a activation. Agonist-activated β 1 ARs, however, are desensitized/downregulated in human HF due to the actions of the βarrestins (βarrestin1 and 2). βarrestins are GPCR adapter proteins and signal transducers. βarrestin1 is by far the predominant isoform in the heart and reduces contractility by desensitizing the β 1 AR, whereas βarrestin2 is expressed at negligible levels and is beneficial post-myocardial infarction (MI), as it combats inflammation and apoptosis. Herein, we sought to investigate whether cardiac βarrestin2 exerts any inotropic effects. Methods: We used βarrestin knockout (KO) mouse hearts and also performed intra-cardiac adenoviral gene transfer of βarrestin2 (Adβarrestin2) in post-MI mice in vivo. For mechanistic signaling studies we used the cardiomyocyte cell line H9c2. Results: SERCA2a SUMO (small ubiquitin-like modifier)ylation and activity and, consequently, cardiac contractility were increased in βarrestin1 KO`s vs. WT`s post-MI. The opposite was true for βarrestin2 KO`s post-MI. Additionally, βarrestin2, but not βarrestin1, was found to directly bind SERCA2a and induce its SUMOylation and activation in mouse hearts in vivo, as well as in cardiomyocytes in vitro acutely in response to β 1 AR stimulation. Interestingly, βarrestin2 did not affect the classic β 1 AR cAMP-dependent pro-contractile signaling pathway in cardiomyocytes, again contrary to βarrestin1. Importantly, and consistent with these findings, Adβarrestin2 gene transfer in post-MI mouse hearts in vivo resulted in enhanced cardiac function (post-MI ejection fraction of Adβarrestin2 vs. control (AdGFP) mice: 40.3 + 1.3% vs. 23.1 + 1.2%, respectively, p&lt;0.05, n=5). Conclusions: Cardiac β 1 AR-activated βarrestin2, but not βarrestin1, promotes SERCA2a SUMOylation and activity, increasing cardiac contractility. Given also its anti-inflammatory and anti-apoptotic effects post-MI, cardiac-specific βarrestin2 gene transfer may be a novel and safe inotropic therapy for both acute and chronic HF.

Midkine's Role in Cardiac Pathology.

Midkine (MDK) is a heparin-binding growth factor that is normally expressed in mid-gestational development mediating mesenchymal and epithelial interactions. As organisms age, expression of MDK diminishes; however, in adults, MDK expression is associated with acute and chronic pathologic conditions such as myocardial infarction and heart failure (HF). The role of MDK is not clear in cardiovascular disease and currently there is no consensus if it plays a beneficial or detrimental role in HF. The lack of clarity in the literature is exacerbated by differing roles that circulating and myocardial MDK play in signaling pathways in cardiomyocytes (some of which have yet to be elucidated). Of particular interest, serum MDK is elevated in adults with chronic heart failure and higher circulating MDK is associated with worse cardiac function. In addition, pediatric HF patients have higher levels of myocardial MDK. This review focuses on what is known about the effect of exogenous versus myocardial MDK in various cardiac disease models in an effort to better clarify the role of midkine in HF.

\u03b21-adrenergic receptor O-glycosylation regulates N-terminal cleavage and signaling responses in cardiomyocytes

β1-adrenergic receptors (β1ARs) mediate catecholamine actions in cardiomyocytes by coupling to both Gs/cAMP-dependent and Gs-independent/growth-regulatory pathways. Structural studies of the β1AR define ligand-binding sites in the transmembrane helices and effector docking sites at the intracellular surface of the β1AR, but the extracellular N-terminus, which is a target for post-translational modifications, typically is ignored. This study identifies β1AR N-terminal O-glycosylation at Ser37/Ser41 as a mechanism that prevents β1AR N-terminal cleavage. We used an adenoviral overexpression strategy to show that both full-length/glycosylated β1ARs and N-terminally truncated glycosylation-defective β1ARs couple to cAMP and ERK-MAPK signaling pathways in cardiomyocytes. However, a glycosylation defect that results in N-terminal truncation stabilizes β1ARs in a conformation that is biased toward the cAMP pathway. The identification of O-glycosylation and N-terminal cleavage as novel structural determinants of β1AR responsiveness in cardiomyocytes could be exploited for therapeutic advantage.

Time course of changes in oxidative stress and stress-induced proteins in cardiomyocytes exposed to doxorubicin and prevention by vitamin C.

We previously reported that Vitamin C (Vit C) protects against doxorubicin (Dox)-induced cardiotoxicity by reducing oxidative stress, p38 mitogen-activated kinase (MAPK) and p53 activation and rescuing cell death in isolated adult cardiomyocytes. The pattern of activation and the role of oxidative stress as well as down-stream mechanisms for such protection remain elusive. Therefore the present study aims to analyze time-dependant generation of reactive oxygen species (ROS) and the activation of stress induced signalling pathways in cardiomyocytes treated with Dox and Vit C. The data provides further understanding of heart pathophysiology in response to Dox at the cellular level, and may help to optimize the timing of various therapeutic approaches. Cardiomyocytes isolated from adult Sprague-Dawley rats were exposed to Dox (10 μM), Vit C (25 μM), and Dox + Vit C for different time intervals up to 24 h. p38-JNK (SB203580) and p53 (pifithrin-α) inhibitors were used to determine the role of each respective signalling protein. Dox administration to cardiomyocytes increased the levels of ROS in a time-dependent manner that followed the activation of stress-induced proteins p53, p38 and JNK MAPKs, culminating in an increase in autophagy and apoptosis markers. Dox-induced increase in ROS was alleviated by Vit C adjuvant treatment at all time-points and this was also correlated with blunting of the activation of the studied signaling pathways leading to the prevention of apoptosis and preservation of cell viability. Protective effect of Vit C against the activation of stress induced proteins, autophagy and apoptosis was mainly attributed to its antioxidant properties even though blockage of p38, JNK and p53 by pharmacological inhibitors also suppressed Dox-induced apoptosis. ROS is defined as a key inducer of cardiomyocyte damage under Dox exposure; Vit C could effectively counteract all Dox-induced changes in cardiomyocytes and may potentially be used as an antioxidant adjuvant therapy to protect against Dox-induced cardiomyopathy.

Potential phytoestrogen alternatives exert cardio-protective mechanisms via estrogen receptors.

The 17 beta-estradiol (E2) is a sex hormone that is most abundant and most active estrogen in premenopausal women. The importance of E2 in providing cardioprotection and reducing the occurrence of heart disease in women of reproductive age has been well recognized. There are three subtype of estrogen receptors (ERs), including ERα, ERβ and GPR30 have been identified and accumulating evidence reveal their roles on E2-mediated genomic and nongenomic pathway in cardiomyocytes against various cardiac insults. In this review, we focus on the estrogen and ERs mediated signaling pathways in cardiomyocytes that determines cardio-protection against various stresses and further discuss the clinical implication of ERs and phytoestrogens. Further we provide some insights on phytoeostrogens which may play as alternatives in estrogen replacement therapies.

Abstract 12941: MURC/Cavin-4 Deficiency Reduces Infarct Size and Preserves Cardiac Function in a Mouse Model of Ischemia-Reperfusion Injury

Introduction: Cardiac ischemia-reperfusion (I/R) injury is considered clinically important issue in cardioprotective strategy for ischemic heart disease. Muscle-restricted coiled-coil protein (MURC)/Cavin-4 is a member of the cavin family, which is a component of caveola. MURC/Cavin-4 is involved in myofibrillar organization by regulating Rho/ROCK signaling pathway and cardiac hypertrophy through ERK1/2 activation. MURC/Cavin-4 mutations are observed in patients with dilated cardiomyopathy. However, the role of MURC/Cavin-4 in cardiac I/R injury remains unknown. Methods and Results: Cardiac I/R was performed on MURC/Cavin-4-/- mice and WT mice. Mice were subjected to 1 hour ischemia by ligating left anterior descending artery, followed by 24 hours reperfusion. MURC/Cavin-4 mRNA expression was continuously increased after reperfusion in WT mouse hearts. Compared with WT mice, MURC/Cavin-4-/- mice exhibited significantly smaller infarct size (IS), which was assessed as proportion of the area at risk (AAR) (IS/AAR[%], MURC/Cavin-4-/- vs. WT: 52.7 ± 3.1 vs. 69.8 ± 12.3, p &lt; 0.05). Echocardiography showed that MURC/Cavin-4 deficiency significantly preserved cardiac function (LVEF[%], MURC/Cavin-4-/- vs. WT: 54.2 ± 5.6 vs. 36.2 ± 2.0, p &lt; 0.01; FS[%], MURC/Cavin-4-/- vs. WT: 27.0 ± 3.7 vs. 16.9 ± 1.1, p &lt; 0.01, respectively). Hydrogen peroxide (H 2 O 2 ), the most abundant form of reactive oxygen species (ROS) produced during I/R, causes inflammation, apoptosis and subsequent tissue damages. In neonatal rat cardiomyocytes, MURC/Cavin-4 siRNA reduced cell death induced by H 2 O 2 compared with control siRNA, as determined by TUNEL staining (TUNEL positive cells[%],MURC/Cavin-4 siRNA vs. control siRNA: 45.1 ± 7.2 vs. 65.7 ± 12.1, p &lt; 0.05). Western blot analysis showed that p38 MAPK activation by H 2 O 2 was inhibited in cardiomyocytes transfected with MURC/Cavin-4 siRNA compared with control siRNA. Akt/PKB signaling was not influenced by MURC/Cavin-4 siRNA transfection. Conclusions: Our results indicate that MURC/Cavin-4 plays a crucial role in the development of cardiac I/R injury through ROS-induced apoptosis signaling pathway in cardiomyocytes. MURC/Cavin-4 might be a novel therapeutic target for cardiac I/R injury.

Abstract 299: Natural Product Derivative Bio Promotes Recovery After Myocardial Infarction via Unique Modulation of the Cardiac Microenvironment

Rationale: Myocardial infarction (MI) induces cardiac remodeling, which is regulated by the cardiac microenvironment and results in scarring and loss of cardiac function. Targeting the microenvironment represents a novel therapeutic approach. Objective: To investigate the therapeutic effect of a natural compound derivative, BIO [(2’Z,3’E)-6-Bromoindirubin-3′-oxime], on cardiac microenvironment cells and remodeling post-MI. Methods and Results: Using a series of co-culture studies, BIO was shown to induce proliferation in cardiomyocytes and, conversely, inhibit proliferation in cardiac fibroblasts. In addition, BIO produced anti-fibrotic effects in the fibroblasts, such as reduced motility and expression changes in fibrosis-related factors. In macrophages, BIO inhibited the expression of pro-inflammatory inducible nitric oxide synthase. Interestingly, BIO modulated molecular crosstalk between cardiac fibroblasts and differentiating macrophages to increase expression of anti-inflammatory M2 macrophage markers. In the optically transparent zebrafish-based model of heart failure, BIO induced cardiomyocyte proliferation to recover survival rate. BIO is a known glycogen synthase kinase-3β inhibitor, but these effects could not be recapitulated using the classical inhibitor, lithium chloride; indicating novel, potential therapeutic effects of BIO on remodeling. We characterized these novel effects of BIO as differential modulation of p27 expression and potent induction of interleukin-10 in microenvironment cells. In rat MI model, BIO reduced fibrosis and improved cardiac performance. Histological analysis revealed a greater presence of anti-inflammatory M2 macrophages in the infarction zone. BIO treatment also reduced serum levels of pro-fibrotic interleukin-6. Conclusions: BIO differentially activates signaling pathways in cardiomyocytes, cardiac fibroblasts and cardiac macrophages to reduce fibrosis, increase cardiomyocyte proliferation and promote recovery post-MI.

Disrupted desmosomes drive fibrosis

![Figure][1] Compared with control cells (left), fibronectin is up-regulated in cardiomyocytes lacking PKP2 (right). [Dubash et al.][2] describe how the loss of desmosomal proteins activates a profibrotic signaling pathway in cardiomyocytes. Plakophilin-2 (PKP2) is a key structural

DNA damage-inducible transcript 4 (DDIT4) mediates methamphetamine-induced autophagy and apoptosis through mTOR signaling pathway in cardiomyocytes

Methamphetamine (METH) is an amphetamine-like psychostimulant that is commonly abused. Previous studies have shown that METH can induce damages to the nervous system and recent studies suggest that METH can also cause adverse and potentially lethal effects on the cardiovascular system. Recently, we demonstrated that DNA damage-inducible transcript 4 (DDIT4) regulates METH-induced neurotoxicity. However, the role of DDIT4 in METH-induced cardiotoxicity remains unknown. We hypothesized that DDIT4 may mediate METH-induced autophagy and apoptosis in cardiomyocytes. To test the hypothesis, we examined DDIT4 protein expression in cardiomyocytes and in heart tissues of rats exposed to METH with Western blotting. We also determined the effects on METH-induced autophagy and apoptosis after silencing DDIT4 expression with synthetic siRNA with or without pretreatment of a mTOR inhibitor rapamycin in cardiomyocytes using Western blot analysis, fluorescence microscopy and TUNEL staining. Our results showed that METH exposure increased DDIT4 expression and decreased phosphorylation of mTOR that was accompanied with increased autophagy and apoptosis both in vitro and in vivo. These effects were normalized after silencing DDIT4. On the other hand, rapamycin promoted METH-induced autophagy and apoptosis in DDIT4 knockdown cardiomyocytes. These results suggest that DDIT4 mediates METH-induced autophagy and apoptosis through mTOR signaling pathway in cardiomyocytes.

C1q/TNF-related protein-1 functions to protect against acute ischemic injury in the heart.

Obesity is associated with an increased risk of cardiovascular disease. C1q/TNF-related protein (CTRP)-1 is a poorly characterized adipokine that is up-regulated in association with ischemic heart disease. We investigated the role of CTRP1 in myocardial ischemia injury. CTRP1-knockout mice showed increased myocardial infarct size, cardiomyocyte apoptosis, and proinflammatory gene expression after I/R compared with wild-type (WT) mice. In contrast, systemic delivery of CTRP1 attenuated myocardial damage after I/R in WT mice. Treatment of cardiomyocytes with CTRP1 led to reduction of hypoxia-reoxygenation-induced apoptosis and lipopolysaccharide-stimulated expression of proinflammatory cytokines, which was reversed by inhibition of sphingosine-1-phosphate (S1P) signaling. Treatment of cardiomyocytes with CTRP1 also resulted in the increased production of cAMP, which was blocked by suppression of S1P signaling. The antiapoptotic and anti-inflammatory actions of CTRP1 were cancelled by inhibition of adenylyl cyclase or knockdown of adiponectin receptor 1. Furthermore, blockade of S1P signaling reversed CTRP1-mediated inhibition of myocardial infarct size, apoptosis, and inflammation after I/R in vivo. These data indicate that CTRP1 protects against myocardial ischemic injury by reducing apoptosis and inflammatory response through activation of the S1P/cAMP signaling pathways in cardiomyocytes, suggesting that CTRP1 plays a crucial role in the pathogenesis of ischemic heart disease.

A novel miR-371a-5p-mediated pathway, leading to BAG3 upregulation in cardiomyocytes in response to epinephrine, is lost in Takotsubo cardiomyopathy.

Molecular mechanisms protecting cardiomyocytes from stress-induced death, including tension stress, are essential for cardiac physiology and defects in these protective mechanisms can result in pathological alterations. Bcl2-associated athanogene 3 (BAG3) is expressed in cardiomyocytes and is a component of the chaperone-assisted autophagy pathway, essential for homeostasis of mechanically altered cells. BAG3 ablation in mice results in a lethal cardiomyopathy soon after birth and mutations of this gene have been associated with different cardiomyopathies including stress-induced Takotsubo cardiomyopathy (TTC). The pathogenic mechanism leading to TTC has not been defined, but it has been suggested that the heart can be damaged by excessive epinephrine (epi) spillover in the absence of a protective mechanism. The aim of this study was to provide more evidence for a role of BAG3 in the pathogenesis of TTC. Therefore, we sequenced BAG3 gene in 70 TTC patients and in 81 healthy donors with the absence of evaluable cardiovascular disease. Mutations and polymorphisms detected in the BAG3 gene included a frequent nucleotide change g2252c in the BAG3 3′-untranslated region (3′-UTR) of Takotsubo patients (P<0.05), resulting in loss of binding of microRNA-371a-5p (miR-371a-5p) as evidenced by dual-luciferase reporter assays and argonaute RNA-induced silencing complex catalytic component 2/pull-down assays. Moreover, we describe a novel signaling pathway in cardiomyocytes that leads to BAG3 upregulation on exposure to epi through an ERK-dependent upregulation of miR-371a-5p. In conclusion, the presence of a g2252c polymorphism in the BAG3 3′-UTR determines loss of miR-371a-5p binding and results in an altered response to epi, potentially representing a new molecular mechanism that contributes to TTC pathogenesis.

Nucleostemin exerts anti-apoptotic function via p53 signaling pathway in cardiomyocytes.

Cardiomyocytes apoptosis following reperfusion injury causes irreversible damage to cardiac function. Understanding the mechanisms underlying cardiomyocytes death under these conditions can be helpful to identify strategies to abrogate such detrimental effects. Stem cell-specific proteins and regulatory pathways become important in understanding reparative processes in the myocardium. One such regulatory protein named nucleostemin (NS) has vital roles in cardiac ischemia. Although the relationship between NS and cell apoptosis has been studied, it is unknown how NS is controlled and how it participates in cardiomyocytes apoptosis induced by ischemia reperfusion (I/R). In the present study, we aimed to investigate the direct role of NS in myocardial I/R. In vivo, NS was highly expressed in cardiac tissues after I/R. Double immunofluorescent staining showed that NS located in the nucleolar of cardiomyocytes and correlated with cardiomyocytes apoptosis. Furthermore, in vitro primary rat cardiomyocytes increased NS expression induced by hypoxia-reoxygenation (H/R) treatment, in line with results in vivo. Suppression of NS expression by siNS promoted the expression of terminal deoxynucleotide transferase-mediated dUTP nick end labeling (TUNEL)-positive cells p53 and cleaved caspase-3, which demonstrates I/R may require increased expression of NS to suppress p53 activation and maintain cardiomyocytes survival.