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1.
JCI Insight ; 6(22)2021 Nov 22.
Artigo em Inglês | MEDLINE | ID: mdl-34806652

RESUMO

Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease with limited treatment options. Despite endothelial cells (ECs) comprising 30% of the lung cellular composition, the role of EC dysfunction in pulmonary fibrosis (PF) remains unclear. We hypothesize that sterol regulatory element-binding protein 2 (SREBP2) plays a critical role in the pathogenesis of PF via EC phenotypic modifications. Transcriptome data demonstrate that SREBP2 overexpression in ECs led to the induction of the TGF, Wnt, and cytoskeleton remodeling gene ontology pathways and the increased expression of mesenchymal genes, such as snail family transcriptional repressor 1 (snai1), α-smooth muscle actin, vimentin, and neural cadherin. Furthermore, SREBP2 directly bound to the promoter regions and transactivated these mesenchymal genes. This transcriptomic change was associated with an epigenetic and phenotypic switch in ECs, leading to increased proliferation, stress fiber formation, and ECM deposition. Mice with endothelial-specific transgenic overexpression of SREBP2 (EC-SREBP2[N]-Tg mice) that were administered bleomycin to induce PF demonstrated exacerbated vascular remodeling and increased mesenchymal transition in the lung. SREBP2 was also found to be markedly increased in lung specimens from patients with IPF. These results suggest that SREBP2, induced by lung injury, can exacerbate PF in rodent models and in human patients with IPF.

2.
Front Cell Dev Biol ; 9: 702974, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34595164

RESUMO

Endothelial cells (ECs) form the inner lining of blood vessels and are central to sensing chemical perturbations that can lead to oxidative stress. The degree of stress is correlated with divergent phenotypes such as quiescence, cell death, or senescence. Each possible cell fate is relevant for a different aspect of endothelial function, and hence, the regulation of cell fate decisions is critically important in maintaining vascular health. This study examined the oxidative stress response (OSR) in human ECs at the boundary of cell survival and death through longitudinal measurements, including cellular, gene expression, and perturbation measurements. 0.5 mM hydrogen peroxide (HP) produced significant oxidative stress, placed the cell at this junction, and provided a model to study the effectors of cell fate. The use of systematic perturbations and high-throughput measurements provide insights into multiple regimes of the stress response. Using a systems approach, we decipher molecular mechanisms across these regimes. Significantly, our study shows that heme oxygenase-1 (HMOX1) acts as a gatekeeper of cell fate decisions. Specifically, HP treatment of HMOX1 knockdown cells reversed the gene expression of about 51% of 2,892 differentially expressed genes when treated with HP alone, affecting a variety of cellular processes, including anti-oxidant response, inflammation, DNA injury and repair, cell cycle and growth, mitochondrial stress, metabolic stress, and autophagy. Further analysis revealed that these switched genes were highly enriched in three spatial locations viz., cell surface, mitochondria, and nucleus. In particular, it revealed the novel roles of HMOX1 on cell surface receptors EGFR and IGFR, mitochondrial ETCs (MTND3, MTATP6), and epigenetic regulation through chromatin modifiers (KDM6A, RBBP5, and PPM1D) and long non-coding RNA (lncRNAs) in orchestrating the cell fate at the boundary of cell survival and death. These novel aspects suggest that HMOX1 can influence transcriptional and epigenetic modulations to orchestrate OSR affecting cell fate decisions.

3.
Sci Transl Med ; 13(592)2021 05 05.
Artigo em Inglês | MEDLINE | ID: mdl-33952674

RESUMO

Pulmonary arterial hypertension (PAH) is a progressive disorder leading to occlusive vascular remodeling. Current PAH therapies improve quality of life but do not reverse structural abnormalities in the pulmonary vasculature. Here, we used high-throughput drug screening combined with in silico analyses of existing transcriptomic datasets to identify a promising lead compound to reverse PAH. Induced pluripotent stem cell-derived endothelial cells generated from six patients with PAH were exposed to 4500 compounds and assayed for improved cell survival after serum withdrawal using a chemiluminescent caspase assay. Subsequent validation of caspase activity and improved angiogenesis combined with data analyses using the Gene Expression Omnibus and Library of Integrated Network-Based Cellular Signatures databases revealed that the lead compound AG1296 was positively associated with an anti-PAH gene signature. AG1296 increased abundance of bone morphogenetic protein receptors, downstream signaling, and gene expression and suppressed PAH smooth muscle cell proliferation. AG1296 induced regression of PA neointimal lesions in lung organ culture and PA occlusive changes in the Sugen/hypoxia rat model and reduced right ventricular systolic pressure. Moreover, AG1296 improved vascular function and BMPR2 signaling and showed better correlation with the anti-PAH gene signature than other tyrosine kinase inhibitors. Specifically, AG1296 up-regulated small mothers against decapentaplegic (SMAD) 1/5 coactivators, cAMP response element-binding protein 3 (CREB3), and CREB5: CREB3 induced inhibitor of DNA binding 1 and downstream genes that improved vascular function. Thus, drug discovery for PAH can be accelerated by combining phenotypic screening with in silico analyses of publicly available datasets.


Assuntos
Hipertensão Pulmonar , Células-Tronco Pluripotentes Induzidas , Hipertensão Arterial Pulmonar , Animais , Proliferação de Células , Simulação por Computador , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Células Endoteliais , Humanos , Hipertensão Pulmonar/tratamento farmacológico , Artéria Pulmonar , Qualidade de Vida , Ratos , Tirfostinas
4.
Cell Stem Cell ; 28(1): 96-111.e7, 2021 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-33142114

RESUMO

The embryonic endocardium is essential for early heart development as it functions to induce trabecular myocardium, the first heart tissue to form, and is the source of the cells that make up the valves and a portion of the coronary vasculature. With this potential, human endocardial cells could provide unique therapeutic opportunities that include engineering biological valves and cell-based therapy strategies to replace coronary vasculature in damaged hearts. To access human endocardial cells, we generated a human pluripotent stem cell (hPSC)-derived endothelial population that displays many characteristics of endocardium, including expression of the cohort of genes that identifies this lineage in vivo, the capacity to induce a trabecular fate in immature cardiomyocytes in vitro, and the ability to undergo an endothelial-to-mesenchymal transition. Analyses of the signaling pathways required for development of the hPSC-derived endocardial cells identified a novel role for BMP10 in the specification of this lineage from cardiovascular mesoderm.


Assuntos
Endocárdio , Células-Tronco Pluripotentes , Proteínas Morfogenéticas Ósseas , Diferenciação Celular , Humanos , Miocárdio , Transdução de Sinais
6.
Cell Stem Cell ; 27(4): 574-589.e8, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-32810435

RESUMO

Hypoplastic left heart syndrome (HLHS) is a complex congenital heart disease characterized by abnormalities in the left ventricle, associated valves, and ascending aorta. Studies have shown intrinsic myocardial defects but do not sufficiently explain developmental defects in the endocardial-derived cardiac valve, septum, and vasculature. Here, we identify a developmentally impaired endocardial population in HLHS through single-cell RNA profiling of hiPSC-derived endocardium and human fetal heart tissue with an underdeveloped left ventricle. Intrinsic endocardial defects contribute to abnormal endothelial-to-mesenchymal transition, NOTCH signaling, and extracellular matrix organization, key factors in valve formation. Endocardial abnormalities cause reduced cardiomyocyte proliferation and maturation by disrupting fibronectin-integrin signaling, consistent with recently described de novo HLHS mutations associated with abnormal endocardial gene and fibronectin regulation. Together, these results reveal a critical role for endocardium in HLHS etiology and provide a rationale for considering endocardial function in regenerative strategies.


Assuntos
Síndrome do Coração Esquerdo Hipoplásico , Células-Tronco Pluripotentes Induzidas , Endocárdio , Humanos , Miocárdio , Transdução de Sinais
7.
Circulation ; 142(4): 365-379, 2020 07 28.
Artigo em Inglês | MEDLINE | ID: mdl-32393053

RESUMO

BACKGROUND: Metabolic disorders such as obesity and diabetes mellitus can cause dysfunction of endothelial cells (ECs) and vascular rarefaction in adipose tissues. However, the modulatory role of ECs in adipose tissue function is not fully understood. Other than vascular endothelial growth factor-vascular endothelial growth factor receptor-mediated angiogenic signaling, little is known about the EC-derived signals in adipose tissue regulation. We previously identified Argonaute 1 (AGO1; a key component of microRNA-induced silencing complex) as a crucial regulator in hypoxia-induced angiogenesis. In this study, we intend to determine the AGO1-mediated EC transcriptome, the functional importance of AGO1-regulated endothelial function in vivo, and the relevance to adipose tissue function and obesity. METHODS: We generated and subjected mice with EC-AGO1 deletion (EC-AGO1-knockout [KO]) and their wild-type littermates to a fast food-mimicking, high-fat high-sucrose diet and profiled the metabolic phenotypes. We used crosslinking immunoprecipitation- and RNA-sequencing to identify the AGO1-mediated mechanisms underlying the observed metabolic phenotype of EC-AGO1-KO. We further leveraged cell cultures and mouse models to validate the functional importance of the identified molecular pathway, for which the translational relevance was explored using human endothelium isolated from healthy donors and donors with obesity/type 2 diabetes mellitus. RESULTS: We identified an antiobesity phenotype of EC-AGO1-KO, evident by lower body weight and body fat, improved insulin sensitivity, and enhanced energy expenditure. At the organ level, we observed the most significant phenotype in the subcutaneous and brown adipose tissues of KO mice, with greater vascularity and enhanced browning and thermogenesis. Mechanistically, EC-AGO1 suppression results in inhibition of thrombospondin-1 (THBS1/TSP1), an antiangiogenic and proinflammatory cytokine that promotes insulin resistance. In EC-AGO1-KO mice, overexpression of TSP1 substantially attenuated the beneficial phenotype. In human endothelium isolated from donors with obesity or type 2 diabetes mellitus, AGO1 and THBS1 are expressed at higher levels than the healthy controls, supporting a pathological role of this pathway. CONCLUSIONS: Our study suggests a novel mechanism by which ECs, through the AGO1-TSP1 pathway, control vascularization and function of adipose tissues, insulin sensitivity, and whole-body metabolic state.


Assuntos
Tecido Adiposo Marrom/metabolismo , Proteínas Argonauta/metabolismo , Suscetibilidade a Doenças , Endotélio/metabolismo , Fatores de Iniciação em Eucariotos/metabolismo , Doenças Metabólicas/etiologia , Doenças Metabólicas/metabolismo , Adulto , Animais , Proteínas Argonauta/genética , Dieta Hiperlipídica , Modelos Animais de Doenças , Metabolismo Energético , Fatores de Iniciação em Eucariotos/genética , Feminino , Perfilação da Expressão Gênica , Marcação de Genes , Loci Gênicos , Humanos , Resistência à Insulina , Masculino , Doenças Metabólicas/diagnóstico , Camundongos , Camundongos Knockout , Pessoa de Meia-Idade , Modelos Biológicos , Obesidade , Fenótipo
8.
J Vis Exp ; (158)2020 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-32364545

RESUMO

It has been shown that endocardial endothelial cells (EECs) and coronary endothelial cells (CECs) differ in origin, development, markers, and functions. Consequently, these two cell populations play unique roles in cardiac diseases. Current studies involving isolated endothelial cells investigate cell populations consisting of both EECs and CECs. This protocol outlines a method to independently isolate these two cell populations for cell-specific characterization. Following the collection of the left and right ventricular free wall, endothelial cells from the outer surface and inner surface are separately liberated using a digestion buffer solution. The sequential digestion of the outer surface and the inner endocardial layer retained separation of the two endothelial cell populations. The separate isolation of EECs and CECs is further verified through the identification of markers specific to each population. Based on previously published single cell RNA profiling in the mouse heart, the Npr3, Hapln1, and Cdh11 gene expression is unique to EECs; while Fabp4, Mgll, and Cd36 gene expression is unique to CECs. qPCR data revealed enriched expression of these characteristic markers in their respective samples, indicating successful EEC and CEC isolation, as well as maintenance of cell phenotype, enabling further cell-specific functional analysis.


Assuntos
Vasos Coronários/citologia , Endocárdio/citologia , Endotélio Vascular/citologia , Ventrículos do Coração/citologia , Coração/fisiologia , Animais , Biomarcadores/metabolismo , Células Cultivadas , Vasos Coronários/metabolismo , Endocárdio/metabolismo , Endotélio Vascular/metabolismo , Perfilação da Expressão Gênica , Ventrículos do Coração/metabolismo , Ratos , Ratos Sprague-Dawley
9.
EMBO Mol Med ; 12(5): e11303, 2020 05 08.
Artigo em Inglês | MEDLINE | ID: mdl-32324970

RESUMO

Endothelial dysfunction is critically involved in the pathogenesis of pulmonary arterial hypertension (PAH) and that exogenously administered microRNA may be of therapeutic benefit. Lower levels of miR-483 were found in serum from patients with idiopathic pulmonary arterial hypertension (IPAH), particularly those with more severe disease. RNA-seq and bioinformatics analyses showed that miR-483 targets several PAH-related genes, including transforming growth factor-ß (TGF-ß), TGF-ß receptor 2 (TGFBR2), ß-catenin, connective tissue growth factor (CTGF), interleukin-1ß (IL-1ß), and endothelin-1 (ET-1). Overexpression of miR-483 in ECs inhibited inflammatory and fibrogenic responses, revealed by the decreased expression of TGF-ß, TGFBR2, ß-catenin, CTGF, IL-1ß, and ET-1. In contrast, inhibition of miR-483 increased these genes in ECs. Rats with EC-specific miR-483 overexpression exhibited ameliorated pulmonary hypertension (PH) and reduced right ventricular hypertrophy on challenge with monocrotaline (MCT) or Sugen + hypoxia. A reversal effect was observed in rats that received MCT with inhaled lentivirus overexpressing miR-483. These results indicate that PAH is associated with a reduced level of miR-483 and that miR-483 might reduce experimental PH by inhibition of multiple adverse responses.


Assuntos
Hipertensão Pulmonar , MicroRNAs , Animais , Modelos Animais de Doenças , Humanos , Hipertensão Pulmonar/genética , Hipóxia , MicroRNAs/genética , Monocrotalina , Ratos
10.
Nat Commun ; 9(1): 292, 2018 01 18.
Artigo em Inglês | MEDLINE | ID: mdl-29348663

RESUMO

The optimal expression of endothelial nitric oxide synthase (eNOS), the hallmark of endothelial homeostasis, is vital to vascular function. Dynamically regulated by various stimuli, eNOS expression is modulated at transcriptional, post-transcriptional, and post-translational levels. However, epigenetic modulations of eNOS, particularly through long non-coding RNAs (lncRNAs) and chromatin remodeling, remain to be explored. Here we identify an enhancer-associated lncRNA that enhances eNOS expression (LEENE). Combining RNA-sequencing and chromatin conformation capture methods, we demonstrate that LEENE is co-regulated with eNOS and that its enhancer resides in proximity to eNOS promoter in endothelial cells (ECs). Gain- and Loss-of-function of LEENE differentially regulate eNOS expression and EC function. Mechanistically, LEENE facilitates the recruitment of RNA Pol II to the eNOS promoter to enhance eNOS nascent RNA transcription. Our findings unravel a new layer in eNOS regulation and provide novel insights into cardiovascular regulation involving endothelial function.


Assuntos
Células Endoteliais/metabolismo , Elementos Facilitadores Genéticos/genética , Regulação Enzimológica da Expressão Gênica , Óxido Nítrico Sintase Tipo III/genética , RNA Longo não Codificante/genética , Animais , Células Cultivadas , Perfilação da Expressão Gênica , Humanos , Masculino , Camundongos Endogâmicos C57BL , Óxido Nítrico Sintase Tipo III/metabolismo , Regiões Promotoras Genéticas/genética , RNA Polimerase II/metabolismo , Transcrição Genética
11.
J Am Soc Nephrol ; 28(11): 3251-3261, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28696247

RESUMO

CKD is an independent risk factor for cardiovascular disease (CVD). The accumulation of uremic toxins in CKD induces oxidative stress and endothelial dysfunction. MicroRNA-92a (miR-92a) is induced by oxidative stress in endothelial cells (ECs) and involved in angiogenesis and atherosclerosis. We investigated a role for oxidative stress-responsive miR-92a in CKD. Our study of patients at three clinical sites showed increased serum miR-92a level with decreased kidney function. In cultured ECs, human CKD serum or uremic toxins (such as indoxyl sulfate), compared with non-CKD serum, induced the levels of miR-92a and suppressed the expression of miR-92a targets, including key endothelial-protective molecules. The antioxidant N-acetylcysteine inhibited these vasculopathic properties. In rats, adenine-induced CKD associated with increased levels of miR-92a in aortas, serum, and CD144+ endothelial microparticles. Furthermore, CD144+ microparticles from human uremic serum contained more miR-92a than those from control serum. Additional analysis showed a positive correlation between serum levels of miR-92a and indoxyl sulfate in a cohort of patients with ESRD undergoing hemodialysis. Collectively, our findings suggest that the uremic toxins accumulated in CKD can upregulate miR-92a in ECs, which impairs EC function and predisposes patients to CVD.


Assuntos
Células Endoteliais/fisiologia , MicroRNAs/fisiologia , Insuficiência Renal Crônica/fisiopatologia , Animais , Feminino , Humanos , Masculino , MicroRNAs/sangue , Pessoa de Meia-Idade , Ratos , Ratos Wistar , Insuficiência Renal Crônica/sangue
12.
Sci Rep ; 6: 38579, 2016 12 06.
Artigo em Inglês | MEDLINE | ID: mdl-27922125

RESUMO

Estrogen, via estrogen receptor alpha (ERα), exerts several beneficial effects on metabolism and energy homeostasis by controlling size, enzymatic activity and hormonal content of adipose tissue. The actions of estrogen on sympathetic ganglia, which are key players in the browning process, are less well known. In the present study we show that ERß influences browning of subcutaneous adipose tissue (SAT) via its actions both on sympathetic ganglia and on the SAT itself. A 3-day-treatment with a selective ERß agonist, LY3201, induced browning of SAT in 1-year-old obese WT and ERα-/- female mice. Browning was associated with increased expression of ERß in the nuclei of neurons in the sympathetic ganglia, increase in tyrosine hydroxylase in both nerve terminals in the SAT and sympathetic ganglia neurons and an increase of ß3-adrenoceptor in the SAT. LY3201 had no effect on browning in young female or male mice. In the case of young females browning was already maximal while in males there was very little expression of ERß in the SAT and very little expression of the ß3-adrenoceptor. The increase in both sympathetic tone and responsiveness of adipocytes to catecholamines reveals a novel role for ERß in controlling browning of adipose tissue.


Assuntos
Tecido Adiposo Marrom/metabolismo , Receptor beta de Estrogênio/agonistas , Obesidade/metabolismo , Gordura Subcutânea Abdominal/metabolismo , Tecido Adiposo Marrom/efeitos dos fármacos , Tecido Adiposo Branco/metabolismo , Fatores Etários , Animais , Benzopiranos/farmacologia , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/genética , Receptor beta de Estrogênio/metabolismo , Feminino , Expressão Gênica , Masculino , Camundongos , Camundongos Knockout , Camundongos Obesos , Modelos Biológicos , Obesidade/genética , Fatores Sexuais , Gordura Subcutânea Abdominal/efeitos dos fármacos , Sistema Nervoso Simpático/efeitos dos fármacos
13.
Proc Natl Acad Sci U S A ; 113(42): 11883-11888, 2016 10 18.
Artigo em Inglês | MEDLINE | ID: mdl-27688768

RESUMO

The aryl hydrocarbon receptor (AhR) is now recognized as an important physiological regulator in the immune and reproductive systems, and in the development of the liver and vascular system. AhR regulates cell cycle, cell proliferation, and differentiation through interacting with other signaling pathways, like estrogen receptor α (ERα), androgen receptor (AR), and Notch signaling. In the present study, we investigated Notch and estrogen signaling in AhR-/- mice. We found low fertility with degenerative changes in the testes, germ cell apoptosis, and a reduced number of early spermatids. There was no change in aromatase, AR, ERα, or ERß expression in the testis and no detectable change in serum estrogen levels. However, expression of Notch receptors (Notch1 and Notch3) and their target Hairy and Enhancer of Split homolog 1 (HES1) was reduced. In addition, the testosterone level was slightly reduced in the serum. In the mammary fat pad, AhR appeared to regulate estrogen signaling because, in AhR-/- males, there was significant growth of the mammary ducts with high expression of ERα in the ductal epithelium. The enhanced mammary ductal growth appears to be related to overexpression of ERα accompanied by a high proliferation index, whereas the reduced fertility appears to be related defects in Notch signaling that leads to reduced expression of HES1 and, consequently, early maturation of spermatocytes and a depletion of primary spermatids. Previous reports have indicated that AhR pathway is associated with infertility in men. Our results provide a mechanistic explanation for this defect.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/deficiência , Receptor alfa de Estrogênio/metabolismo , Receptores de Hidrocarboneto Arílico/deficiência , Receptores Notch/metabolismo , Transdução de Sinais , Animais , Apoptose/genética , Aromatase/metabolismo , Biomarcadores , Proliferação de Células , Receptor alfa de Estrogênio/genética , Receptor beta de Estrogênio/genética , Receptor beta de Estrogênio/metabolismo , Feminino , Fertilidade/genética , Deleção de Genes , Expressão Gênica , Células Germinativas/metabolismo , Imuno-Histoquímica , Masculino , Glândulas Mamárias Animais/crescimento & desenvolvimento , Glândulas Mamárias Animais/metabolismo , Camundongos , Camundongos Knockout , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo , Receptores de Progesterona/genética , Receptores de Progesterona/metabolismo , Espermatócitos/metabolismo , Testículo/metabolismo
14.
J Mol Cell Biol ; 8(6): 518-529, 2016 12.
Artigo em Inglês | MEDLINE | ID: mdl-27436752

RESUMO

Among the four prostaglandin E2 receptors, EP3 receptor is the one most abundantly expressed in white adipose tissue (WAT). The mouse EP3 gene gives rise to three isoforms, namely EP3α, EP3ß, and EP3γ, which differ only at their C-terminal tails. To date, functions of EP3 receptor and its isoforms in WAT remain incompletely characterized. In this study, we found that the expression of all EP3 isoforms were downregulated in WAT of both db/db and high-fat diet-induced obese mice. Genetic ablation of three EP3 receptor isoforms (EP3-/- mice) or EP3α and EP3γ isoforms with EP3ß intact (EP3ß mice) led to an obese phenotype with increased food intake, decreased motor activity, reduced insulin sensitivity, and elevated serum triglycerides. Since the differentiation of preadipocytes and mouse embryonic fibroblasts to adipocytes was markedly facilitated by either pharmacological blockade or genetic deletion/inhibition of EP3 receptor via the cAMP/PKA/PPARγ pathway, increased adipogenesis may contribute to obesity in EP3-/- and EP3ß mice. Moreover, both EP3-/- and EP3ß mice had increased lipolysis in WAT mainly due to the activated cAMP/PKA/hormone-sensitive lipase pathway. Taken together, our findings suggest that EP3 receptor and its α and γ isoforms are involved in both adipogenesis and lipolysis and influence food intake, serum lipid levels, and insulin sensitivity.


Assuntos
Adipogenia , Tecido Adiposo Branco/metabolismo , Lipólise , Receptores de Prostaglandina E Subtipo EP3/metabolismo , Adipócitos/metabolismo , Adipócitos/patologia , Animais , Diferenciação Celular , Deleção de Genes , Inflamação/metabolismo , Inflamação/patologia , Resistência à Insulina , Lipoproteínas VLDL/metabolismo , Camundongos , Camundongos Obesos , Obesidade/metabolismo , Obesidade/patologia , Fenótipo , Isoformas de Proteínas/metabolismo , Ratos Sprague-Dawley , Transdução de Sinais , Triglicerídeos/metabolismo
15.
Adipocyte ; 5(2): 238-42, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27386163

RESUMO

The recent discovery of browning of white adipose tissue (WAT) has raised great research interest because of its significant potential in counteracting obesity and type II diabetes. However, the mechanisms underlying browning are still poorly understood. Liver X receptors (LXRs) are one class of nuclear receptors, which play a vital role in regulating cholesterol, triglyceride and glucose metabolism. Following our previous finding that LXRs serve as repressors of UCP1 in classic brown adipose tissue in female mice, we found that LXRs, especially LXRß, also repress the browning process of subcutaneous adipose tissue (SAT) in male rodents fed a normal diet. Depletion of LXRs activated thyrotropin releasing hormone positive neurons in the paraventricular area of the hypothalamus, and thus stimulated secretion of thyroid-stimulating hormone from the pituitary. Consequently production of thyroid hormones in the thyroid gland and circulating thyroid hormone level were increased. Moreover, the activity of thyroid signaling in SAT was markedly increased. One unexpected finding of our study is that LXRs are indispensable in the thyroid hormone negative feedback loop at the level of the hypothalamus. LXRs maintain expression of thyroid receptors in the brain and when they are inactivated there is no negative feedback of thyroid hormone in the hypothalamus. Together, our findings have uncovered the basis of increased energy expenditure in male LXR knock-out mice and provided support for targeting LXRs in treatment of obesity.

16.
Proc Natl Acad Sci U S A ; 113(27): 7614-9, 2016 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-27335465

RESUMO

The etiology of peripheral squamous cell lung cancer (PSCCa) remains unknown. Here, we show that this condition spontaneously develops in mice in which the genes for two oxysterol receptors, Liver X Receptor (LXR) α (Nr1h3) and ß (Nr1h2), are inactivated. By 1 y of age, most of these mice have to be euthanized because of severe dyspnea. Starting at 3 mo, the lungs of LXRα,ß(Dko) mice, but not of LXRα or LXRß single knockout mice, progressively accumulate foam cells, so that by 1 y, the lungs are covered by a "golden coat." There is infiltration of inflammatory cells and progressive accumulation of lipid in the alveolar wall, type 2 pneumocytes, and macrophages. By 14 mo, there are three histological lesions: one resembling adenomatous hyperplasia, one squamous metaplasia, and one squamous cell carcinoma characterized by expression of transformation-related protein (p63), sex determining region Y-box 2 (Sox2), cytokeratin 14 (CK14), and cytokeratin 13 (CK13) and absence of thyroid transcription factor 1 (TTF1), and prosurfactant protein C (pro-SPC). RNA sequencing analysis at 12 mo confirmed a massive increase in markers of M1 macrophages and lymphocytes. The data suggest a previously unidentified etiology of PSCCa: cholesterol dysregulation and M1 macrophage-predominant lung inflammation combined with damage to, and aberrant repair of, lung tissue, particularly the peripheral parenchyma. The results raise the possibility that components of the LXR signaling may be useful targets in the treatment of PSCCa.


Assuntos
Metabolismo dos Lipídeos , Receptores X do Fígado/fisiologia , Neoplasias Pulmonares/etiologia , Pulmão/metabolismo , Neoplasias de Células Escamosas/etiologia , Células Epiteliais Alveolares/metabolismo , Animais , Fibroblastos/metabolismo , Homeostase , Pulmão/patologia , Macrófagos/metabolismo , Camundongos Endogâmicos C57BL , Pneumonia/etiologia , Análise de Sequência de RNA
17.
Proc Natl Acad Sci U S A ; 112(45): 14006-11, 2015 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-26504234

RESUMO

The recent discovery of browning of white adipose tissue (WAT) has raised great research interest because of its significant potential in counteracting obesity and type 2 diabetes. Browning is the result of the induction in WAT of a newly discovered type of adipocyte, the beige cell. When mice are exposed to cold or several kinds of hormones or treatments with chemicals, specific depots of WAT undergo a browning process, characterized by highly activated mitochondria and increased heat production and energy expenditure. However, the mechanisms underlying browning are still poorly understood. Liver X receptors (LXRs) are one class of nuclear receptors, which play a vital role in regulating cholesterol, triglyceride, and glucose metabolism. Following our previous finding that LXRs serve as repressors of uncoupling protein-1 (UCP1) in classic brown adipose tissue in female mice, we found that LXRs, especially LXRß, also repress the browning process of subcutaneous adipose tissue (SAT) in male rodents fed a normal diet. Depletion of LXRs activated thyroid-stimulating hormone (TSH)-releasing hormone (TRH)-positive neurons in the paraventricular nucleus area of the hypothalamus and thus stimulated secretion of TSH from the pituitary. Consequently, production of thyroid hormones in the thyroid gland and circulating thyroid hormone level were increased. Moreover, the activity of thyroid signaling in SAT was markedly increased. Together, our findings have uncovered the basis of increased energy expenditure in male LXR knockout mice and provided support for targeting LXRs in treatment of obesity.


Assuntos
Tecido Adiposo Branco/metabolismo , Encéfalo/metabolismo , Metabolismo Energético/fisiologia , Mitocôndrias/metabolismo , Receptores Nucleares Órfãos/metabolismo , Hormônios Tireóideos/metabolismo , Análise de Variância , Animais , Composição Corporal/fisiologia , Ensaio de Imunoadsorção Enzimática , Perfilação da Expressão Gênica , Imuno-Histoquímica , Receptores X do Fígado , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores Nucleares Órfãos/genética , Hormônio Liberador de Tireotropina/metabolismo
18.
Proc Natl Acad Sci U S A ; 112(27): 8397-402, 2015 Jul 07.
Artigo em Inglês | MEDLINE | ID: mdl-26100911

RESUMO

The antidiuretic hormone arginine vasopressin is a systemic effector in urinary concentration. However, increasing evidence suggests that other locally produced factors may also play an important role in the regulation of water reabsorption in renal collecting ducts. Recently, prostaglandin E2 (PGE2) receptor EP4 has emerged as a potential therapeutic target for the treatment of nephrogenic diabetes insipidus, but the underlying mechanism is unknown. To evaluate the role of EP4 in regulating water homeostasis, mice with renal tubule-specific knockout of EP4 (Ksp-EP4(-/-)) and collecting duct-specific knockout of EP4 (AQP2-EP4(-/-)) were generated using the Cre-loxP recombination system. Urine concentrating defect was observed in both Ksp-EP4(-/-) and AQP2-EP4(-/-) mice. Decreased aquaporin 2 (AQP2) abundance and apical membrane targeting in renal collecting ducts were evident in Ksp-EP4(-/-) mice. In vitro studies demonstrated that AQP2 mRNA and protein levels were significantly up-regulated in mouse primary inner medullary collecting duct (IMCD) cells after pharmacological activation or adenovirus-mediated overexpression of EP4 in a cAMP/cAMP-response element binding protein-dependent manner. In addition, EP4 activation or overexpression also increased AQP2 membrane accumulation in a mouse IMCD cell line (IMCD3) stably transfected with the AQP2 gene, mainly through the cAMP/protein kinase A and extracellular signal-regulated kinase pathways. In summary, the EP4 receptor in renal collecting ducts plays an important role in regulating urinary concentration under physiological conditions. The ability of EP4 to promote AQP2 membrane targeting and increase AQP2 abundance makes it a potential therapeutic target for the treatment of clinical disorders including acquired and congenital diabetes insipidus.


Assuntos
Aquaporina 2/genética , Capacidade de Concentração Renal/genética , Túbulos Renais Coletores/metabolismo , Receptores de Prostaglandina E Subtipo EP4/genética , Animais , Aquaporina 2/metabolismo , Western Blotting , Linhagem Celular , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Células Cultivadas , AMP Cíclico/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Dinoprostona/análogos & derivados , Dinoprostona/biossíntese , Dinoprostona/farmacologia , Túbulos Renais Coletores/citologia , Sistema de Sinalização das MAP Quinases , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Microscopia de Fluorescência , Cultura Primária de Células , Pirrolidinonas/farmacologia , Interferência de RNA , Receptores de Prostaglandina E Subtipo EP4/agonistas , Receptores de Prostaglandina E Subtipo EP4/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Água/metabolismo
19.
Hepatology ; 59(5): 1779-90, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24806753

RESUMO

UNLABELLED: FAM3A belongs to a novel cytokine-like gene family, and its physiological role remains largely unknown. In our study, we found a marked reduction of FAM3A expression in the livers of db/db and high-fat diet (HFD)-induced diabetic mice. Hepatic overexpression of FAM3A markedly attenuated hyperglycemia, insulin resistance, and fatty liver with increased Akt (pAkt) signaling and repressed gluconeogenesis and lipogenesis in the livers of those mice. In contrast, small interfering RNA (siRNA)-mediated knockdown of hepatic FAM3A resulted in hyperglycemia with reduced pAkt levels and increased gluconeogenesis and lipogenesis in the livers of C57BL/6 mice. In vitro study revealed that FAM3A was mainly localized in the mitochondria, where it increases adenosine triphosphate (ATP) production and secretion in cultured hepatocytes. FAM3A activated Akt through the p110α catalytic subunit of PI3K in an insulin-independent manner. Blockade of P2 ATP receptors or downstream phospholipase C (PLC) and IP3R and removal of medium calcium all significantly reduced FAM3A-induced increase in cytosolic free Ca(2+) levels and attenuated FAM3A-mediated PI3K/Akt activation. Moreover, FAM3A-induced Akt activation was completely abolished by the inhibition of calmodulin (CaM). CONCLUSION: FAM3A plays crucial roles in the regulation of glucose and lipid metabolism in the liver, where it activates the PI3K-Akt signaling pathway by way of a Ca(2+) /CaM-dependent mechanism. Up-regulating hepatic FAM3A expression may represent an attractive means for the treatment of insulin resistance, type 2 diabetes, and nonalcoholic fatty liver disease (NAFLD).


Assuntos
Classe Ia de Fosfatidilinositol 3-Quinase/fisiologia , Citocinas/fisiologia , Gluconeogênese , Lipogênese , Fígado/metabolismo , Proteínas Proto-Oncogênicas c-akt/fisiologia , Transdução de Sinais/fisiologia , Trifosfato de Adenosina/metabolismo , Animais , Diabetes Mellitus Tipo 2/complicações , Dieta Hiperlipídica , Fígado Gorduroso/etiologia , Células Hep G2 , Humanos , Hiperglicemia/etiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Receptores Purinérgicos P2/fisiologia
20.
Proc Natl Acad Sci U S A ; 111(6): 2277-82, 2014 Feb 11.
Artigo em Inglês | MEDLINE | ID: mdl-24464484

RESUMO

The farnesoid X receptor (FXR) is a ligand-activated transcription factor belonging to the nuclear receptor superfamily. FXR is mainly expressed in liver and small intestine, where it plays an important role in bile acid, lipid, and glucose metabolism. The kidney also has a high FXR expression level, with its physiological function unknown. Here we demonstrate that FXR is ubiquitously distributed in renal tubules. FXR agonist treatment significantly lowered urine volume and increased urine osmolality, whereas FXR knockout mice exhibited an impaired urine concentrating ability, which led to a polyuria phenotype. We further found that treatment of C57BL/6 mice with chenodeoxycholic acid, an FXR endogenous ligand, significantly up-regulated renal aquaporin 2 (AQP2) expression, whereas FXR gene deficiency markedly reduced AQP2 expression levels in the kidney. In vitro studies showed that the AQP2 gene promoter contained a putative FXR response element site, which can be bound and activated by FXR, resulting in a significant increase of AQP2 transcription in cultured primary inner medullary collecting duct cells. In conclusion, the present study demonstrates that FXR plays a critical role in the regulation of urine volume, and its activation increases urinary concentrating capacity mainly via up-regulating its target gene AQP2 expression in the collecting ducts.


Assuntos
Capacidade de Concentração Renal/genética , Receptores Citoplasmáticos e Nucleares/genética , Animais , Aquaporina 2/genética , Aquaporina 2/metabolismo , Sequência de Bases , Primers do DNA , Rim/metabolismo , Masculino , Camundongos , Camundongos Knockout , Reação em Cadeia da Polimerase , Receptores Citoplasmáticos e Nucleares/metabolismo
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