Cyclosporin A Regulates Sodium-Calcium Exchanger (NCX1) Gene Expression in Vitro and Cardiac Hypertrophy in NCX1 Transgenic Mice

Pathological cardiac hypertrophy is considered a precursor to clinical heart failure. Understanding the transcriptional regulators that suppress the hypertrophic response may have profound implications for the treatment of heart disease. We report the generation of transgenic mice that overexpress the transcription factor CHF1/Hey2 in the myocardium. In response to the α-adrenergic agonist phenylephrine, they show marked attenuation in the hypertrophic response compared with wild-type controls, even though blood pressure is similar in both groups. Isolated myocytes from transgenic mice demonstrate a similar resistance to phenylephrine-induced hypertrophy in vitro, providing further evidence that the protective effect of CHF1/Hey2 is mediated at the myocyte level. Induction of the hypertrophy marker genes ANF, BNP, and β- MHC in the transgenic cells is concurrently suppressed in vivo and in vitro, demonstrating that the induction of hypertrophy-associated genes is repressed by CHF1/Hey2. Transfection of CHF1/Hey2 into neonatal cardiomyocytes suppresses activation of an ANF reporter plasmid by the transcription factor GATA4, which has previously been shown to activate a hypertrophic transcriptional program. Furthermore, CHF1/Hey2 binds GATA4 directly in coimmunoprecipitation assays and inhibits the binding of GATA4 to its recognition sequence within the ANF promoter. Our findings demonstrate that CHF1/Hey2 functions as an antihypertrophic gene, possibly through inhibition of a GATA4-dependent hypertrophic program.

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The Effect of a Novel Highly Selective Inhibitor of the Sodium/Calcium Exchanger (NCX) on Cardiac Arrhythmias in In Vitro and In Vivo Experiments

PLoS ONE ◽

10.1371/journal.pone.0166041 ◽

2016 ◽

Vol 11 (11) ◽

pp. e0166041 ◽

Cited By ~ 23

Author(s):

Zsófia Kohajda ◽

Nikolett Farkas-Morvay ◽

Norbert Jost ◽

Norbert Nagy ◽

Amir Geramipour ◽

...

Keyword(s):

Cardiac Arrhythmias ◽

Selective Inhibitor ◽

Sodium Calcium Exchanger ◽

In Vivo Experiments ◽

Sodium Calcium

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Stable Inhibition of Brain Synaptic Plasma Membrane Calcium ATPase in Rats Anesthetized with Halothane

Anesthesiology ◽

10.1097/00000542-199501000-00016 ◽

1995 ◽

Vol 82 (1) ◽

pp. 118-128 ◽

Cited By ~ 14

Author(s):

John J. Franks ◽

Jean-Louis Horn ◽

Piotr K. Janicki ◽

Gurkeerat Singh

Keyword(s):

Plasma Membrane ◽

Hydrolytic Activity ◽

Synaptic Plasma Membrane ◽

Sodium Calcium Exchanger ◽

Halothane Anesthesia ◽

Treatment Groups ◽

Sodium Calcium ◽

Recovery From Anesthesia ◽

Anesthetic Exposure

Background The authors recently showed that plasma membrane Ca(2+)-ATPase (PMCA) activity in cerebral synaptic plasma membrane (SPM) is diminished in a dose-related fashion during exposure in vitro to halothane, isoflurane, xenon, and nitrous oxide at clinically relevant partial pressures. They have now extended their work to in vivo studies, examining PMCA pumping in SPM obtained from control rats decapitated without anesthetic exposure, from rats decapitated during halothane anesthesia, and from rats decapitated after recovery from halothane anesthesia. Methods Three treatment groups were studied: 1) C, control rats that were decapitated without anesthetic exposure, 2) A, anesthetized rats exposed to 1 minimum effective dose (MED) for 20 min and then decapitated, and 3) R, rats exposed to 1 MED for 20 min and then decapitated after recovery from anesthesia, defined as beginning to groom. Plasma membrane Ca(2+)-ATPase pumping and Ca(2+)-dependent ATPase hydrolytic activity, as well as sodium-calcium exchanger activity and Na+-K+-ATPase hydrolytic activity, were assessed in cerebral SPM. In addition, halothane effect on smooth endoplasmic reticulum Ca(2+)-ATPase (SERCA) was examined. Results Plasma membrane Ca(2+)-ATPase transport of Ca2+ into SPM vesicles from anesthetized rats was reduced to 71% of control (P < 0.01) compared with 113% of control for the recovered group (NS). No depression by halothane of SERCA activity, sodium-calcium exchanger, or Na+-K+-ATPase activity was noted among the CAR treatment groups. Conclusions Plasma membrane Ca(2+)-ATPase is selectively and stably inhibited in cerebral SPM from rats killed while anesthetized with halothane, compared with rats killed without anesthesia or after recovery from anesthesia. The studies described in this report, in conjunction with previously reported inhibition of PMCA activity in vitro by a wide range of anesthetic agents, indicate a relationship between inhibition of PMCA and action of inhalational anesthetics.

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Collagen promoter-GFP transgenic mice provide a model to study collagen gene expression during experimental intestinal inflammation and injury in vivo and in vitro

Gastroenterology ◽

10.1016/s0016-5085(08)80663-4 ◽

2001 ◽

Vol 120 (5) ◽

pp. A135

Author(s):

Christopher M. Dacosta ◽

James G. Simmons ◽

Charles Randall Fuller ◽

Kristen L. Williams ◽

Michael Breindl ◽

...

Keyword(s):

Gene Expression ◽

Transgenic Mice ◽

Intestinal Inflammation ◽

Collagen Gene ◽

Collagen Gene Expression ◽

Collagen Promoter

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Egr-1 negatively regulates expression of the sodium?calcium exchanger-1 in cardiomyocytes in vitro and in vivo

Cardiovascular Research ◽

10.1016/j.cardiores.2004.09.026 ◽

2005 ◽

Vol 65 (1) ◽

pp. 187-194 ◽

Cited By ~ 24

Author(s):

C WANG ◽

S DOSTANIC ◽

N SERVANT ◽

L CHALIFOUR

Keyword(s):

Sodium Calcium Exchanger ◽

Sodium Calcium

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TANK Promotes Pressure Overload Induced Cardiac Hypertrophy via Activating AKT Signaling Pathway

Frontiers in Cardiovascular Medicine ◽

10.3389/fcvm.2021.687540 ◽

2021 ◽

Vol 8 ◽

Author(s):

Yanan Pang ◽

Minglu Ma ◽

Dong Wang ◽

Xun Li ◽

Li Jiang

Keyword(s):

Transgenic Mice ◽

Cardiac Hypertrophy ◽

Knockout Mice ◽

Pressure Overload ◽

Akt Signaling ◽

Control Group ◽

Akt Inhibitor ◽

Aortic Banding ◽

Pathological Cardiac Hypertrophy

Background: TANK (TRAF family member associated NF-κB activator) acts as a member of scaffold proteins participated in the development of multiple diseases. However, its function in process of cardiac hypertrophy is still unknown.Methods and Results: In this study, we observed an increased expression of TANK in murine hypertrophic hearts after aortic banding, suggesting that TANK may be involved in the pathogenesis of cardiac hypertrophy. We generated cardiac-specific TANK knockout mice, and subsequently subjected to aortic banding for 4–8 weeks. TANK knockout mice showed attenuated cardiac hypertrophy and dysfunction compared to the control group. In contrast, cardiac-specific TANK transgenic mice showed opposite signs. Consistently, in vitro experiments revealed that TANK knockdown decreased the cell size and expression of hypertrophic markers. Mechanistically, AKT signaling was inhibited in TANK knockout mice, but activated in TANK transgenic mice after aortic banding. Blocking AKT signaling with a pharmacological AKT inhibitor alleviated the cardiac hypertrophy and dysfunction in TANK transgenic mice.Conclusions: Collectively, we identified TANK accelerates the progression of pathological cardiac hypertrophy and is a potential therapeutic target.

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Abstract 1118: Novel Lentiviral Vectors Bearing the Cardiac-Specific Promoter of the Sodium-Calcium Exchanger (NCX1) Report Cardiogenic Differentiation in Stem Cells

Circulation ◽

10.1161/circ.116.suppl_16.ii_224-d ◽

2007 ◽

Vol 116 (suppl_16) ◽

Author(s):

Andreas S Barth ◽

Eddy Kizana ◽

John Terrovitis ◽

Peihong Dong ◽

Rachel R Smith ◽

...

Keyword(s):

Stem Cells ◽

Troponin I ◽

Lentiviral Vectors ◽

Neonatal Rat ◽

Luciferase Reporter ◽

Sodium Calcium Exchanger ◽

Α Subunit ◽

Sodium Calcium ◽

Cardiogenic Differentiation

Background: Cardiosphere-derived resident cardiac stem cells (CDCs) are readily isolated from adult hearts and confer functional benefit in animal models of heart failure. To study cardiogenic differentiation in CDCs, we developed a method to genetically label and selectively enrich for cells that have differentiated down the cardiomyocyte lineage. Methods and Results: Lentiviral vectors achieved significantly higher transduction efficiency (>90%) and longer-term transgene expression in human CDCs than any of the nine adeno-associated viral serotypes tested (AAV 1– 6, 2.5, 8, 9). To define the best reporter of cardiogenic differentiation, five cardiac-specific promoters (sodium-calcium exchanger [NCX1], L-type calcium channel α-subunit, α-myosin heavy chain, troponin I and myosin ventricular light chain) were subcloned upstream of GFP in lentiviral vectors. Cardiac specificity was assessed by transducing neonatal rat ventricular myocytes (NRVMs), HEK293, cardiac fibroblasts, skeletal myoblasts, endothelial and vascular smooth muscle cells with each vector and then measuring GFP fluorescence by flow cytometry. The cardiac NCX1-promoter conveyed the highest degree of cardiac specificity, with expression limited to NRVMs. At a multiplicity of infection of 35, NCX1-GFP vectors did not affect cell death or apoptosis of CDCs. Expression persisted for up to 6 months in vitro . NCX1-GFP positive CDCs subpopulations, were FACS-sorted and expanded in vitro , demonstrating enhanced expression of a variety of cardiac markers by real-time PCR. The utility of lentiviral vectors bearing the cardiac NCX1-promoter was further exemplified by treating CDCs with cardiogenic sulfonylhydrazones (SHZ). SHZ potently activated a NCX1-firefly luciferase reporter (211 ± 21% greater than control cells) and upregulated cardiac transcripts (including BNP, troponin I, α-tropomyosin, GATA-4). Conclusion: Lentiviral vectors carrying the NCX1 promoter represent a useful tool for genetically marking stem cells and their progeny that have differentiated down the cardiomyocyte lineage. The ability to selectively enrich for cardiomyocytes and their precursors has potential relevance for the development of cell-based therapies.

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Abstract P141: Cardiac TRH Partly Mediates Angiotensin II-induced Fibrotic and Hypertrophy Effects in "in vivo" and "in vitro" Models

Hypertension ◽

10.1161/hyp.66.suppl_1.p141 ◽

2015 ◽

Vol 66 (suppl_1) ◽

Author(s):

Silvia I García ◽

Ludmila S Peres Diaz ◽

Maia Aisicovich ◽

Mariano L Schuman ◽

María S Landa

Keyword(s):

Gene Expression ◽

Body Weight ◽

Cardiac Hypertrophy ◽

Structural Changes ◽

In Vitro Models ◽

Heart Weight ◽

Saline Control ◽

Control Group

Cardiac TRH (cTRH) is overexpressed in the hypertrophied ventricle (LV) of the SHR. Additionally in vivo siRNA-TRH treatment induced downregulation of LV-TRH preventing cardiac hypertrophy and fibrosis demonstrating that TRH is involved in hypertrophic and fibrotic processes. Moreover, in a normal heart, the increase of LV TRH expression alone could induce structural changes where fibrosis and hypertrophy could be involved, independently of any other system alterations. Is well-known the cardiac hypertrophy/ fibrotic effects induced by AII, raising the question of whether specific LV cTRH inhibition might attenuates AII induced cardiac hypertrophy and fibrosis in mice. We challenged C57 mice with AII (osmotic pumps,14 days; 2 mg/kg) to induce cardiac hypertrophy vs saline. Groups were divided and , simultaneously to pump surgery, injected intracardiac with siRNA-TRH and siRNA-Con as its control. Body weight, water consume and SABP were measured daily. As expected, AII significantly increased SABP (p<0.05) in both groups treated , although cardiac hypertrophy (heart weight/body weight) was only evident in the group with the cardiac TRH system undamaged, suggesting that the cardiac TRH system function as a necessary mediator of the AII-induced hypertrophic effect. As hypothesized, we found an AII-induced increase of TRH (p<0.05) gene expression (real-t PCR) confirmed by immunofluorescence that was not observed in the group AII+siRNA-TRH demonstrating the specific siRNA treatment efficiency. Furthermore, AII significantly increase (p<0.05) BNP (hypertrophic marker), III collagen and TGFB (fibrosis markers) expressions only in the group with AII with the cardiac TRH system intact. On the contrary, the group with AII and the cTRH system inhibited, shows genes expressions similar to the saline control group. We confirmed these results by immunofluorescence. Similar fibrotic results were observed with NIH3T3 cell culture where we demonstrated that AII induced TRH gene expression (p<0.05) and its inhibition impedes AII-induced increase of TGFB and III/I collagens expressions telling us about the role of the cTRH in the AII fibrosis effects. Our results point out that the cardiac TRH is involved in the AII-induced hypertrophic and fibrotic effects.

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Abstract 1828: Dyxin/Lmcd1 Mediates Cardiac Hypertrophy Both In Vitro And In Vivo

Circulation ◽

10.1161/circ.118.suppl_18.s_393-a ◽

2008 ◽

Vol 118 (suppl_18) ◽

Author(s):

Derk Frank ◽

Robert Frauen ◽

Christiane Hanselmann ◽

Christian Kuhn ◽

Rainer Will ◽

...

Keyword(s):

Transgenic Mice ◽

Cardiac Hypertrophy ◽

Neonatal Rat ◽

Heart Weight ◽

Cardiomyocyte Hypertrophy ◽

The Novel ◽

Rat Cardiomyocytes ◽

A Genome

In order to identify new molecular mediators of cardiomyocyte hypertrophy, we performed a genome wide mRNA microarray screen of biomechanically stretched neonatal rat cardiomyocytes (NRCM). We found the novel sarcomeric LIM protein Dyxin/Lmcd1 being significantly upregulated (5.6x, p<0.001). Moreover, Dyxin was also significantly induced in several mouse models of myocardial hypertrophy including aortic banding, calcineurin overexpression and angiotensin stimulation, suggesting a potential role as a mediator of cardiac hypertrophy. To further test this hypothesis, we adenovirally overexpressed Dyxin in NRCM which potently induced cellular hypertrophy (150%, p<0.001) and the hypertrophic gene program (ANF, BNP). Consistent with an induction of calcineurin signalling, the calcineurin-responsive gene Rcan1– 4 (MCIP1.4) was found significantly upregulated (3.2x, p<0.001). Conversely, knockdown of Dyxin (−75% on protein level) via miRNA completely blunted the hypertrophic response to hypertrophic stimuli, including stretch and PE (both p<0.001). Furthermore, PE-mediated activation of calcineurin signaling (Upregulation of Rcan1– 4 by 7.3x, p<0.001) was completely blocked by knockdown of Dyxin. To confirm these results in vivo, we next generated transgenic mice with cardiac-restricted overexpression of Dyxin using the α -MHC promoter. Despite normal cardiac function as assessed by echocardiography, adult transgenic mice displayed significant cardiac hypertrophy in morphometrical analyses (3.9 vs. 3.5 mg/g LV/heart weight, n=8–11, p<0.05). This finding was supplemented by a robust induction of the hypertrophic gene program including ANF (3.7-fold, n=6, p=0.01) and α -skeletal actin (2.8-fold, n=6, p<0.05). Likewise, Rcan1– 4 was found upregulated (+112%, n=5, p<0.05), Taken together, we show that the novel sarcomeric z-disc protein Dyxin/Lmcd1 is significantly upregulated in several models of cardiac hypertrophy and potently induces cardiomyocyte hypertrophy both in vitro and in vivo. Mechanistically, Lmcd1/Dyxin appears to signal through the calcineurin pathway.

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