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1.
J Am Heart Assoc ; 10(14): e019473, 2021 07 20.
Artigo em Inglês | MEDLINE | ID: mdl-34227403

RESUMO

Background It is known that dietary intake of polyunsaturated fatty acids may improve cardiac function. However, relatively high daily doses are required to achieve sufficient cardiac concentrations of beneficial omega-3 fatty acids. The liver X receptor (LXR) is a nuclear hormone receptor and a crucial regulator of lipid homeostasis in mammals. LXR activation has been shown to endogenously reprogram cellular lipid profiles toward increased polyunsaturated fatty acids levels. Here we studied whether LXR lipid reprogramming occurs in cardiac tissue and exerts cardioprotective actions. Methods and Results Male 129SV mice were treated with the LXR agonist AZ876 (20 µmol/kg per day) for 11 days. From day 6, the mice were injected with the nonselective ß-agonist isoproterenol for 4 consecutive days to induce diastolic dysfunction and subendocardial fibrosis while maintaining systolic function. Treatment with isoproterenol led to a marked impairment of global longitudinal strain and the E/e' ratio of transmitral flow to mitral annular velocity, which were both significantly improved by the LXR agonist. Histological examination showed a significant reduction in isoproterenol-induced subendocardial fibrosis by AZ876. Analysis of the cardiac lipid composition by liquid chromatography-high resolution mass spectrometry revealed a significant increase in cardiac polyunsaturated fatty acids levels and a significant reduction in saturated fatty acids by AZ876. Conclusions The present study provides evidence that the LXR agonist AZ876 prevents subendocardial damage, improves global longitudinal strain and E/e' in a mouse model of isoproterenol-induced cardiac damage, accompanied by an upregulation of cardiac polyunsaturated fatty acids levels. Cardiac LXR activation and beneficial endogenous cardiac lipid reprogramming may provide a new therapeutic strategy in cardiac disease with diastolic dysfunction.


Assuntos
Compostos de Anilina/farmacologia , Ácidos Graxos/metabolismo , Cardiopatias/prevenção & controle , Isoproterenol , Miocárdio/metabolismo , Tiazóis/farmacologia , Animais , Reprogramação Celular , Modelos Animais de Doenças , Fibrose , Cardiopatias/induzido quimicamente , Cardiopatias/patologia , Receptores X do Fígado/agonistas , Masculino , Camundongos , Camundongos da Linhagem 129 , Miocárdio/patologia
2.
Pflugers Arch ; 473(3): 533-546, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33580817

RESUMO

The cation channel transient receptor potential melastatin 4 (TRPM4) is a calcium-activated non-selective cation channel and acts in cardiomyocytes as a negative modulator of the L-type Ca2+ influx. Global deletion of TRPM4 in the mouse led to increased cardiac contractility under ß-adrenergic stimulation. Consequently, cardiomyocyte-specific inactivation of the TRPM4 function appears to be a promising strategy to improve cardiac contractility in heart failure patients. The aim of this study was to develop a gene therapy approach in mice that specifically silences the expression of TRPM4 in cardiomyocytes. First, short hairpin RNAmiR30 (shRNAmiR30) sequences against the TRPM4 mRNA were screened in vitro using lentiviral transduction for a stable expression of the shRNA cassettes. Western blot analysis identified three efficient shRNAmiR30 sequences out of six, which reduced the endogenous TRPM4 protein level by up to 90 ± 6%. Subsequently, the most efficient shRNAmiR30 sequences were delivered into cardiomyocytes of adult mice using adeno-associated virus serotype 9 (AAV9)-mediated gene transfer. Initially, the AAV9 vector particles were administered via the lateral tail vein, which resulted in a downregulation of TRPM4 by 46 ± 2%. Next, various optimization steps were carried out to improve knockdown efficiency in vivo. First, the design of the expression cassette was streamlined for integration in a self-complementary AAV vector backbone for a faster expression. Compared to the application via the lateral tail vein, intravenous application via the retro-orbital sinus has the advantage that the vector solution reaches the heart directly and in a high concentration, and eventually a TRPM4 knockdown efficiency of 90 ± 7% in the heart was accomplished by this approach. By optimization of the shRNAmiR30 constructs and expression cassette as well as the route of AAV9 vector application, a 90% reduction of TRPM4 expression was achieved in the adult mouse heart. In the future, AAV9-RNAi-mediated inactivation of TRPM4 could be a promising strategy to increase cardiac contractility in preclinical animal models of acute and chronic forms of cardiac contractile failure.


Assuntos
Técnicas de Transferência de Genes , Miócitos Cardíacos/metabolismo , Canais de Cátion TRPM , Animais , Dependovirus , Vetores Genéticos , Masculino , Camundongos , Interferência de RNA , RNA Interferente Pequeno , Transdução Genética/métodos
3.
Elife ; 72018 08 23.
Artigo em Inglês | MEDLINE | ID: mdl-30136928

RESUMO

Amyotrophic lateral sclerosis (ALS) is characterized by the progressive degeneration of motoneurons in the primary motor cortex (pMO) and in spinal cord. However, the pathogenic process involves multiple subnetworks in the brain and functional MRI studies demonstrate an increase in functional connectivity in areas connected to pMO despite the ongoing neurodegeneration. The extent and the structural basis of the motor subnetwork remodeling in experimentally tractable models remain unclear. We have developed a new retrograde AAV9 to quantitatively map the projections to pMO in the SOD1(G93A) ALS mouse model. We show an increase in the number of neurons projecting from somatosensory cortex to pMO at presymptomatic stages, followed by an increase in projections from thalamus, auditory cortex and contralateral MO (inputs from 20 other structures remains unchanged) as disease advances. The stage- and structure-dependent remodeling of projection to pMO in ALS may provide insights into the hyperconnectivity observed in ALS patients.


Assuntos
Esclerose Lateral Amiotrófica/fisiopatologia , Dependovirus/metabolismo , Córtex Motor/fisiopatologia , Esclerose Lateral Amiotrófica/patologia , Animais , Espinhas Dendríticas/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Feminino , Humanos , Masculino , Camundongos , Córtex Motor/patologia , Proteínas Mutantes/metabolismo , Rede Nervosa/patologia , Rede Nervosa/fisiopatologia , Dobramento de Proteína , Células Piramidais/metabolismo , Células Piramidais/patologia , Superóxido Dismutase/metabolismo , Tálamo/patologia , Tálamo/fisiopatologia
4.
Circ Res ; 114(10): 1623-39, 2014 May 09.
Artigo em Inglês | MEDLINE | ID: mdl-24812351

RESUMO

Genetically encoded Ca(2+) indicators constitute a powerful set of tools to investigate functional aspects of Ca(2+) signaling in isolated cardiomyocytes, cardiac tissue, and whole hearts. Here, we provide an overview of the concepts, experiences, state of the art, and ongoing developments in the use of genetically encoded Ca(2+) indicators for cardiac cells and heart tissue. This review is supplemented with in vivo viral gene transfer experiments and comparisons of available genetically encoded Ca(2+) indicators with each other and with the small molecule dye Fura-2. In the context of cardiac myocytes, we provide guidelines for selecting a genetically encoded Ca(2+) indicator. For future developments, we discuss improvements of a broad range of properties, including photophysical properties such as spectral spread and biocompatibility, as well as cellular and in vivo applications.


Assuntos
Sinalização do Cálcio/genética , Corantes Fluorescentes , Miócitos Cardíacos/química , Miócitos Cardíacos/fisiologia , Transgenes , Animais , Diagnóstico por Imagem/métodos , Técnicas de Transferência de Genes , Humanos , Miócitos Cardíacos/metabolismo
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