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1.
J Mol Cell Cardiol ; 174: 38-46, 2023 01.
Artigo em Inglês | MEDLINE | ID: mdl-36372279

RESUMO

Cardiac fibrosis is regulated by the activation and phenotypic switching of quiescent cardiac fibroblasts to active myofibroblasts, which have extracellular matrix (ECM) remodeling and contractile functions which play a central role in cardiac remodeling in response to injury. Here, we show that expression and activity of the RNA binding protein HuR is increased in cardiac fibroblasts upon transformation to an active myofibroblast. Pharmacological inhibition of HuR significantly blunts the TGFß-dependent increase in ECM remodeling genes, total collagen secretion, in vitro scratch closure, and collagen gel contraction in isolated primary cardiac fibroblasts, suggesting a suppression of TGFß-induced myofibroblast activation upon HuR inhibition. We identified twenty-four mRNA transcripts that were enriched for HuR binding following TGFß treatment via photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation (PAR-CLIP). Eleven of these HuR-bound mRNAs also showed significant co-expression correlation with HuR, αSMA, and periostin in primary fibroblasts isolated from the ischemic-zone of infarcted mouse hearts. Of these, WNT1-inducible signaling pathway protein-1 (Wisp1; Ccn4), was the most significantly associated with HuR expression in fibroblasts. Accordingly, we found Wisp1 expression to be increased in cardiac fibroblasts isolated from the ischemic-zone of mouse hearts following ischemia/reperfusion, and confirmed Wisp1 expression to be HuR-dependent in isolated fibroblasts. Finally, addition of exogenous recombinant Wisp1 partially rescued myofibroblast-induced collagen gel contraction following HuR inhibition, demonstrating that HuR-dependent Wisp1 expression plays a functional role in HuR-dependent MF activity downstream of TGFß. In conclusion, HuR activity is necessary for the functional activation of primary cardiac fibroblasts in response to TGFß, in part through post-transcriptional regulation of Wisp1.


Assuntos
Proteínas de Sinalização Intercelular CCN , Proteína Semelhante a ELAV 1 , Miofibroblastos , Fator de Crescimento Transformador beta , Animais , Camundongos , Colágeno/metabolismo , Fibroblastos/metabolismo , Coração , Miofibroblastos/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Proteína Semelhante a ELAV 1/metabolismo , Proteínas de Sinalização Intercelular CCN/metabolismo
2.
Am J Physiol Heart Circ Physiol ; 321(1): H228-H241, 2021 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-34018851

RESUMO

Adipose tissue homeostasis plays a central role in cardiovascular physiology, and the presence of thermogenically active brown adipose tissue (BAT) has recently been associated with cardiometabolic health. We have previously shown that adipose tissue-specific deletion of HuR (Adipo-HuR-/-) reduces BAT-mediated adaptive thermogenesis, and the goal of this work was to identify the cardiovascular impacts of Adipo-HuR-/-. We found that Adipo-HuR-/- mice exhibit a hypercontractile phenotype that is accompanied by increased left ventricle wall thickness and hypertrophic gene expression. Furthermore, hearts from Adipo-HuR-/- mice display increased fibrosis via picrosirius red staining and periostin expression. To identify underlying mechanisms, we applied both RNA-seq and weighted gene coexpression network analysis (WGCNA) across both cardiac and adipose tissue to define HuR-dependent changes in gene expression as well as significant relationships between adipose tissue gene expression and cardiac fibrosis. RNA-seq results demonstrated a significant increase in proinflammatory gene expression in both cardiac and subcutaneous white adipose tissue (scWAT) from Adipo-HuR-/- mice that is accompanied by an increase in serum levels of both TNF-α and IL-6. In addition to inflammation-related genes, WGCNA identified a significant enrichment in extracellular vesicle-mediated transport and exosome-associated genes in scWAT, whose expression most significantly associated with the degree of cardiac fibrosis observed in Adipo-HuR-/- mice, implicating these processes as a likely adipose-to-cardiac paracrine mechanism. These results are significant in that they demonstrate the spontaneous onset of cardiovascular pathology in an adipose tissue-specific gene deletion model and contribute to our understanding of how disruptions in adipose tissue homeostasis may mediate cardiovascular disease.NEW & NOTEWORTHY The presence of functional brown adipose tissue in humans is known to be associated with cardiovascular health. Here, we show that adipocyte-specific deletion of the RNA binding protein HuR, which we have previously shown to reduce BAT-mediated thermogenesis, is sufficient to mediate a spontaneous development of cardiac hypertrophy and fibrosis. These results may have implications on the mechanisms by which BAT function and adipose tissue homeostasis directly mediate cardiovascular disease.


Assuntos
Adipócitos/metabolismo , Cardiomegalia/genética , Proteína Semelhante a ELAV 1/genética , Miocárdio/metabolismo , Adipócitos/patologia , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Marrom/patologia , Tecido Adiposo Branco/metabolismo , Tecido Adiposo Branco/patologia , Animais , Cardiomegalia/metabolismo , Cardiomegalia/patologia , Proteína Semelhante a ELAV 1/metabolismo , Fibrose/genética , Fibrose/metabolismo , Fibrose/patologia , Camundongos , Camundongos Knockout , Miocárdio/patologia
3.
Clin Sci (Lond) ; 133(22): 2329-2344, 2019 11 29.
Artigo em Inglês | MEDLINE | ID: mdl-31777927

RESUMO

Adipose tissue is classically recognized as the primary site of lipid storage, but in recent years has garnered appreciation for its broad role as an endocrine organ comprising multiple cell types whose collective secretome, termed as adipokines, is highly interdependent on metabolic homeostasis and inflammatory state. Anatomical location (e.g. visceral, subcutaneous, epicardial etc) and cellular composition of adipose tissue (e.g. white, beige, and brown adipocytes, macrophages etc.) also plays a critical role in determining its response to metabolic state, the resulting secretome, and its potential impact on remote tissues. Compared with other tissues, the heart has an extremely high and constant demand for energy generation, of which most is derived from oxidation of fatty acids. Availability of this fatty acid fuel source is dependent on adipose tissue, but evidence is mounting that adipose tissue plays a much broader role in cardiovascular physiology. In this review, we discuss the impact of the brown, subcutaneous, and visceral white, perivascular (PVAT), and epicardial adipose tissue (EAT) secretome on the development and progression of cardiovascular disease (CVD), with a particular focus on cardiac hypertrophy and fibrosis.


Assuntos
Tecido Adiposo/fisiologia , Cardiomegalia/etiologia , Animais , Fibrose , Humanos
4.
Bioorg Med Chem Lett ; 28(3): 334-338, 2018 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-29290543

RESUMO

Prostaglandin E2 (PGE2) is a lipid mediator of inflammation and its inhibition has become a popular drug target due to its harmful physiological roles. Diarylheptanoids are one class of compounds that have shown successful inhibition of PGE2. This paper reports the synthesis and PGE2 inhibitory activity of a series of analogues of a naturally occurring diarylheptanoid. The most efficacious compounds were examined for dose-dependent PGE2 inhibition. Among several promising compounds, the lead candidate exhibited an IC50 value of 0.56 ng/µL or 1.7 µM with no detectable toxicity at the highest dose of 10 ng/µL.


Assuntos
Diarileptanoides/farmacologia , Dinoprostona/antagonistas & inibidores , Diarileptanoides/síntese química , Diarileptanoides/química , Relação Dose-Resposta a Droga , Humanos , Estrutura Molecular , Relação Estrutura-Atividade
5.
Biochim Biophys Acta ; 1849(6): 688-96, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25727182

RESUMO

Heat shock protein 70.3 (Hsp70.3) expression increases in response to cellular stress and plays a cytoprotective role. We have previously shown that Hsp70.3 expression is controlled through coordinated post-transcriptional regulation by miRNAs and alternative polyadenylation (APA), and APA-mediated shortening of the Hsp70.3 3'-UTR facilitates increased protein expression. A stress-induced increase in Hsp70.3 mRNA and protein expression is accompanied by alternative polyadenylation (APA)-mediated truncation of the 3'UTR of the Hsp70.3 mRNA transcript. However, the role that APA plays in stress-induced expression of Hsp70.3 remains unclear. Our results show that APA-mediated truncation of the Hsp70.3 3'UTR increases protein expression through enhanced polyribosome loading. Additionally, we demonstrate that the RNA binding protein HuR, which has been previously shown to play a role in mediating APA, is necessary for heat shock mediated increase in Hsp70.3 mRNA and protein. However, it is somewhat surprising to note that HuR does not play a role in APA of the Hsp70.3 mRNA, and these two regulatory events appear to be mutually exclusive regulators of Hsp70.3 expression. These results not only provide important insight to the regulation of stress response genes following heat shock, but also contribute an enhanced understanding of how alternative polyadenylation contributes to gene regulation.


Assuntos
Proteínas ELAV/genética , Proteínas de Choque Térmico HSP70/biossíntese , Poliadenilação/genética , Biossíntese de Proteínas , Regiões 3' não Traduzidas , Animais , Proteínas ELAV/metabolismo , Regulação da Expressão Gênica , Proteínas de Choque Térmico HSP70/genética , Resposta ao Choque Térmico/genética , Camundongos , MicroRNAs/genética , Processamento de Proteína Pós-Traducional/genética , RNA Mensageiro/biossíntese , RNA Mensageiro/genética
6.
Am J Physiol Gastrointest Liver Physiol ; 311(3): G423-30, 2016 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-27390324

RESUMO

Divalent metal-ion transporter-1 (DMT1), the principal mechanism by which nonheme iron is taken up at the intestinal brush border, is energized by the H(+)-electrochemical potential gradient. The provenance of the H(+) gradient in vivo is unknown, so we have explored a role for brush-border Na(+)/H(+) exchanger (NHE) isoforms by examining iron homeostasis and intestinal iron handling in mice lacking NHE2 or NHE3. We observed modestly depleted liver iron stores in NHE2-null (NHE2(-/-)) mice stressed on a low-iron diet but no change in hematological or blood iron variables or the expression of genes associated with iron metabolism compared with wild-type mice. Ablation of NHE3 strongly depleted liver iron stores, regardless of diet. We observed decreases in blood iron variables but no overt anemia in NHE3-null (NHE3(-/-)) mice on a low-iron diet. Intestinal expression of DMT1, the apical surface ferrireductase cytochrome b reductase-1, and the basolateral iron exporter ferroportin was upregulated in NHE3(-/-) mice, and expression of liver Hamp1 (hepcidin) was suppressed compared with wild-type mice. Absorption of (59)Fe from an oral dose was substantially impaired in NHE3(-/-) compared with wild-type mice. Our data point to an important role for NHE3 in generating the H(+) gradient that drives DMT1-mediated iron uptake at the intestinal brush border.


Assuntos
Regulação da Expressão Gênica/fisiologia , Ferro/metabolismo , Microvilosidades/fisiologia , Trocadores de Sódio-Hidrogênio/metabolismo , Animais , Transporte Biológico , Humanos , Mucosa Intestinal/metabolismo , Camundongos , Camundongos Knockout , Oócitos , Sódio/metabolismo , Trocador 3 de Sódio-Hidrogênio , Trocadores de Sódio-Hidrogênio/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Desequilíbrio Hidroeletrolítico , Xenopus
7.
Am J Physiol Gastrointest Liver Physiol ; 309(8): G635-47, 2015 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-26294671

RESUMO

Divalent metal-ion transporter-1 (DMT1) is a widely expressed iron-preferring membrane-transport protein that serves a critical role in erythroid iron utilization. We have investigated its role in intestinal metal absorption by studying a mouse model lacking intestinal DMT1 (i.e., DMT1(int/int)). DMT1(int/int) mice exhibited a profound hypochromic-microcytic anemia, splenomegaly, and cardiomegaly. That the anemia was due to iron deficiency was demonstrated by the following observations in DMT1(int/int) mice: 1) blood iron and tissue nonheme-iron stores were depleted; 2) mRNA expression of liver hepcidin (Hamp1) was depressed; and 3) intraperitoneal iron injection corrected the anemia, and reversed the changes in blood iron, nonheme-iron stores, and hepcidin expression levels. We observed decreased total iron content in multiple tissues from DMT1(int/int) mice compared with DMT1(+/+) mice but no meaningful change in copper, manganese, or zinc. DMT1(int/int) mice absorbed (64)Cu and (54)Mn from an intragastric dose to the same extent as did DMT1(+/+) mice but the absorption of (59)Fe was virtually abolished in DMT1(int/int) mice. This study reveals a critical function for DMT1 in intestinal nonheme-iron absorption for normal growth and development. Further, this work demonstrates that intestinal DMT1 is not required for the intestinal transport of copper, manganese, or zinc.


Assuntos
Proteínas de Transporte de Cátions/metabolismo , Cobre/metabolismo , Absorção Intestinal/fisiologia , Ferro/metabolismo , Manganês/metabolismo , Anemia Hipocrômica/genética , Anemia Hipocrômica/patologia , Animais , Proteínas de Transporte de Cátions/genética , Transportador de Cobre 1 , Regulação da Expressão Gênica/fisiologia , Homeostase/fisiologia , Camundongos , Camundongos Knockout , Zinco/metabolismo
8.
bioRxiv ; 2023 Jan 18.
Artigo em Inglês | MEDLINE | ID: mdl-36711986

RESUMO

Myocardial ischemia/reperfusion (I/R) injury and the resulting cardiac remodeling is a common cause of heart failure. The RNA binding protein Human Antigen R (HuR) has been previously shown to reduce cardiac remodeling following both I/R and cardiac pressure overload, but the full extent of the HuR-dependent mechanisms within cells of the myocardium have yet to be elucidated. In this study, we applied a novel small molecule inhibitor of HuR to define the functional role of HuR in the acute response to I/R injury and gain a better understanding of the HuR-dependent mechanisms during post-ischemic myocardial remodeling. Our results show an early (two hours post-I/R) increase in HuR activity that is necessary for early inflammatory gene expression by cardiomyocytes in response to I/R. Surprisingly, despite the reductions in early inflammatory gene expression at two hours post-I/R, HuR inhibition has no effect on initial infarct size at 24-hours post-I/R. However, in agreement with previously published work, we do see a reduction in pathological remodeling and preserved cardiac function at two weeks post-I/R upon HuR inhibition. RNA-sequencing analysis of neonatal rat ventricular myocytes (NRVMs) at two hours post-LPS treatment to model damage associated molecular pattern (DAMP)-mediated activation of toll like receptors (TLRs) demonstrates a broad HuR-dependent regulation of pro-inflammatory chemokine and cytokine gene expression in cardiomyocytes. We show that conditioned media from NRVMs pre-treated with HuR inhibitor loses the ability to induce inflammatory gene expression in bone marrow derived macrophages (BMDMs) compared to NRVMs treated with LPS alone. Functionally, HuR inhibition in NRVMs also reduces their ability to induce endocrine migration of peripheral blood monocytes in vitro and reduces post-ischemic macrophage infiltration to the heart in vivo. In summary, these results suggest a HuR-dependent expression of pro-inflammatory gene expression by cardiomyocytes that leads to subsequent monocyte recruitment and macrophage activation in the post-ischemic myocardium.

9.
Adipocyte ; 9(1): 335-345, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-32713230

RESUMO

The goal of this study was to define the functional role of adipocyte-specific expression of the RNA binding protein Human antigen R (HuR). Mice with an adipocyte-specific deletion of HuR (Adipo-HuR-/- ) were generated by crossing HuR floxed (HuRfl/fl ) mice with mice expressing adiponectin-driven cre-recombinase (Adipoq-cre). Our results show that Adipo-HuR-/- mice display a lean phenotype compared to wild-type littermate controls. HuR deletion results in a diet-independent reduction in percent body fat composition along with an increase in energy expenditure. Functionally, Adipo-HuR-/- mice show a significant impairment in acute adaptive thermogenesis (six hours at 4°C), but uncoupling protein 1 (UCP1) protein expression in brown adipose tissue (BAT) is unchanged compared to control. Pharmacological inhibition of HuR also results in a marked decline in core body temperature following acute cold challenge independent of UCP1 protein expression. Among the 588 HuR-dependent genes in BAT identified by RNA-seq analysis, gene ontology analysis shows a significant enrichment in mediators of calcium transport and signalling, almost all of which are decreased in Adipo-HuR-/- mice compared to control. In conclusion, adipocyte expression of HuR plays a central role in metabolic homoeostasis and mediates UCP1-independent thermogenesis in BAT, potentially through post-transcriptional control of intracellular calcium transport.Abbreviations: Adipo-HuR-/-: Adipocyte-specific HuR deletion mice; BAT: Brown adipose tissue; HuR: Human antigen R; UCP1: Uncoupling protein 1.


Assuntos
Tecido Adiposo/metabolismo , Proteína Semelhante a ELAV 1/genética , Metabolismo Energético/genética , Regulação da Expressão Gênica , Termogênese/genética , Proteína Desacopladora 1/genética , Animais , Biomarcadores , Deleção de Genes , Masculino , Camundongos , Camundongos Knockout
10.
JCI Insight ; 4(4)2019 02 21.
Artigo em Inglês | MEDLINE | ID: mdl-30668549

RESUMO

RNA binding proteins represent an emerging class of proteins with a role in cardiac dysfunction. We show that activation of the RNA binding protein human antigen R (HuR) is increased in the failing human heart. To determine the functional role of HuR in pathological cardiac hypertrophy, we created an inducible cardiomyocyte-specific HuR-deletion mouse and showed that HuR deletion reduces left ventricular hypertrophy, dilation, and fibrosis while preserving cardiac function in a transverse aortic constriction (TAC) model of pressure overload-induced hypertrophy. Assessment of HuR-dependent changes in global gene expression suggests that the mechanistic basis for this protection occurs through a reduction in fibrotic signaling, specifically through a reduction in TGF-ß (Tgfb) expression. Finally, pharmacological inhibition of HuR at a clinically relevant time point following the initial development of pathological hypertrophy after TAC also yielded a significant reduction in pathological progression, as marked by a reduction in hypertrophy, dilation, and fibrosis and preserved function. In summary, this study demonstrates a functional role for HuR in the progression of pressure overload-induced cardiac hypertrophy and establishes HuR inhibition as a viable therapeutic approach for pathological cardiac hypertrophy and heart failure.


Assuntos
Proteína Semelhante a ELAV 1/metabolismo , Insuficiência Cardíaca/patologia , Hipertrofia Ventricular Esquerda/tratamento farmacológico , Miocárdio/patologia , Animais , Cardiotônicos/farmacologia , Cardiotônicos/uso terapêutico , Modelos Animais de Doenças , Proteína Semelhante a ELAV 1/antagonistas & inibidores , Proteína Semelhante a ELAV 1/genética , Fibrose , Insuficiência Cardíaca/tratamento farmacológico , Ventrículos do Coração/citologia , Ventrículos do Coração/efeitos dos fármacos , Ventrículos do Coração/patologia , Humanos , Hipertrofia Ventricular Esquerda/genética , Hipertrofia Ventricular Esquerda/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Miocárdio/citologia , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/patologia , RNA-Seq , Remodelação Ventricular/efeitos dos fármacos
12.
Cell Signal ; 28(11): 1735-41, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27521603

RESUMO

The RNA binding protein Human antigen R (HuR) interacts with specific AU-rich domains in target mRNAs and is highly expressed in many cell types, including cardiomyocytes. However, the role of HuR in cardiac physiology is largely unknown. Our results show that HuR undergoes cytoplasmic translocation, indicative of its activation, in hypertrophic cardiac myocytes. Specifically, HuR cytoplasmic translocation is significantly increased in NRVMs (neonatal rat ventricular myocytes) following treatment with phenylephrine or angiotensin II, agonists of two independent Gαq-coupled GPCRs known to induce hypertrophy. This Gq-mediated HuR activation is dependent on p38 MAP kinase, but not canonical Gq-PKC signaling. Furthermore, we show that HuR activation is necessary for Gq-mediated hypertrophic growth of NRVMs as siRNA-mediated knockdown of HuR inhibits hypertrophy as measured by cell size and expression of ANF (atrial natriuretic factor). Additionally, HuR overexpression is sufficient to induce hypertrophic cell growth. To decipher the downstream mechanisms by which HuR translocation promotes cardiomyocyte hypertrophy, we assessed the role of HuR in the transcriptional activity of NFAT (nuclear factor of activated T cells), the activation of which is a hallmark of cardiac hypertrophy. Using an NFAT-luciferase reporter assay, we show an acute inhibition of NFAT transcriptional activity following pharmacological inhibition of HuR. In conclusion, our results identify HuR as a novel mediator of cardiac hypertrophy downstream of the Gq-p38 MAPK pathway, and suggest modulation of NFAT activity as a potential mechanism.


Assuntos
Cardiomegalia/enzimologia , Cardiomegalia/patologia , Miócitos Cardíacos/enzimologia , Miócitos Cardíacos/patologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Animais Recém-Nascidos , Fator Natriurético Atrial/metabolismo , Cardiomegalia/genética , Núcleo Celular/metabolismo , Proliferação de Células , Proteína Semelhante a ELAV 1/antagonistas & inibidores , Proteína Semelhante a ELAV 1/metabolismo , Técnicas de Silenciamento de Genes , Fatores de Transcrição NFATC/metabolismo , Ratos Sprague-Dawley , Ativação Transcricional/genética
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