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1.
Cell Rep ; 38(3): 110277, 2022 01 18.
Artigo em Inglês | MEDLINE | ID: mdl-35045290

RESUMO

Exosomes/small extracellular vesicles (sEVs) can serve as multifactorial mediators of cell-to-cell communication through their miRNA and protein cargo. Quantitative proteomic analysis of five cell lines representing metabolically important tissues reveals that each cell type has a unique sEV proteome. While classical sEV markers such as CD9/CD63/CD81 vary markedly in abundance, we identify six sEV markers (ENO1, GPI, HSPA5, YWHAB, CSF1R, and CNTN1) that are similarly abundant in sEVs of all cell types. In addition, each cell type has specific sEV markers. Using fat-specific Dicer-knockout mice with decreased white adipose tissue and increased brown adipose tissue, we show that these cell-type-specific markers can predict the changing origin of the serum sEVs. These results provide a valuable resource for understanding the sEV proteome of the cells and tissues important in metabolic homeostasis, identify unique sEV markers, and demonstrate how these markers can help in predicting the tissue of origin of serum sEVs.


Assuntos
Biomarcadores/sangue , Exossomos/metabolismo , Proteoma/metabolismo , Células 3T3 , Adiponectina/sangue , Tecido Adiposo/metabolismo , Animais , Camundongos
2.
J Biol Chem ; 294(3): 1059-1069, 2019 01 18.
Artigo em Inglês | MEDLINE | ID: mdl-30459233

RESUMO

FoxO proteins are major targets of insulin action, and FoxO1 mediates the effects of insulin on hepatic glucose metabolism. We reported previously that serpinB1 is a liver-secreted factor (hepatokine) that promotes adaptive ß-cell proliferation in response to insulin resistance in the liver-specific insulin receptor knockout (LIRKO) mouse. Here we report that FoxO1 plays a critical role in promoting serpinB1 expression in hepatic insulin resistance in a non-cell-autonomous manner. Mice lacking both the insulin receptor and FoxO1 (LIRFKO) exhibit reduced ß-cell mass compared with LIRKO mice because of attenuation of ß-cell proliferation. Although hepatic expression of serpinB1 mRNA and protein levels was increased in LIRKO mice, both the mRNA and protein levels returned to control levels in LIRFKO mice. Furthermore, liver-specific expression of constitutively active FoxO1 in transgenic mice induced an increase in hepatic serpinB1 mRNA and protein levels in refed mice. Conversely, serpinB1 mRNA and protein levels were reduced in mice lacking FoxO proteins in the liver. ChIP studies demonstrated that FoxO1 binds to three distinct sites located ∼9 kb upstream of the serpinb1 gene in primary mouse hepatocytes and that this binding is enhanced in hepatocytes from LIRKO mice. However, adenoviral expression of WT or constitutively active FoxO1 and insulin treatment are sufficient to regulate other FoxO1 target genes (IGFBP-1 and PEPCK) but not serpinB1 expression in mouse primary hepatocytes. These results indicate that liver FoxO1 promotes serpinB1 expression in hepatic insulin resistance and that non-cell-autonomous factors contribute to FoxO1-dependent effects on serpinB1 expression in the liver.


Assuntos
Proteína Forkhead Box O1/metabolismo , Regulação da Expressão Gênica , Hepatócitos/metabolismo , Fígado/metabolismo , Serpinas/biossíntese , Animais , Proteína Forkhead Box O1/genética , Hepatócitos/citologia , Proteína 1 de Ligação a Fator de Crescimento Semelhante à Insulina/genética , Proteína 1 de Ligação a Fator de Crescimento Semelhante à Insulina/metabolismo , Fígado/citologia , Masculino , Camundongos , Camundongos Transgênicos , Fosfoenolpiruvato Carboxiquinase (ATP)/genética , Fosfoenolpiruvato Carboxiquinase (ATP)/metabolismo , Serpinas/genética
3.
EMBO J ; 37(24)2018 12 14.
Artigo em Inglês | MEDLINE | ID: mdl-30446598

RESUMO

A finely tuned balance of self-renewal, differentiation, proliferation, and survival governs the pool size and regenerative capacity of blood-forming hematopoietic stem and progenitor cells (HSPCs). Here, we report that protein kinase C delta (PKCδ) is a critical regulator of adult HSPC number and function that couples the proliferative and metabolic activities of HSPCs. PKCδ-deficient mice showed a pronounced increase in HSPC numbers, increased competence in reconstituting lethally irradiated recipients, enhanced long-term competitive advantage in serial transplantation studies, and an augmented HSPC recovery during stress. PKCδ-deficient HSPCs also showed accelerated proliferation and reduced apoptosis, but did not exhaust in serial transplant assays or induce leukemia. Using inducible knockout and transplantation models, we further found that PKCδ acts in a hematopoietic cell-intrinsic manner to restrict HSPC number and bone marrow regenerative function. Mechanistically, PKCδ regulates HSPC energy metabolism and coordinately governs multiple regulators within signaling pathways implicated in HSPC homeostasis. Together, these data identify PKCδ as a critical regulator of HSPC signaling and metabolism that acts to limit HSPC expansion in response to physiological and regenerative demands.


Assuntos
Apoptose , Medula Óssea/enzimologia , Proliferação de Células , Células-Tronco Hematopoéticas/enzimologia , Proteína Quinase C-delta/metabolismo , Transdução de Sinais , Animais , Células-Tronco Hematopoéticas/citologia , Camundongos , Camundongos Knockout , Proteína Quinase C-delta/genética
5.
Nature ; 542(7642): 450-455, 2017 02 23.
Artigo em Inglês | MEDLINE | ID: mdl-28199304

RESUMO

Adipose tissue is a major site of energy storage and has a role in the regulation of metabolism through the release of adipokines. Here we show that mice with an adipose-tissue-specific knockout of the microRNA (miRNA)-processing enzyme Dicer (ADicerKO), as well as humans with lipodystrophy, exhibit a substantial decrease in levels of circulating exosomal miRNAs. Transplantation of both white and brown adipose tissue-brown especially-into ADicerKO mice restores the level of numerous circulating miRNAs that are associated with an improvement in glucose tolerance and a reduction in hepatic Fgf21 mRNA and circulating FGF21. This gene regulation can be mimicked by the administration of normal, but not ADicerKO, serum exosomes. Expression of a human-specific miRNA in the brown adipose tissue of one mouse in vivo can also regulate its 3' UTR reporter in the liver of another mouse through serum exosomal transfer. Thus, adipose tissue constitutes an important source of circulating exosomal miRNAs, which can regulate gene expression in distant tissues and thereby serve as a previously undescribed form of adipokine.


Assuntos
Tecido Adiposo/metabolismo , Regulação da Expressão Gênica , MicroRNAs/sangue , MicroRNAs/metabolismo , Comunicação Parácrina , Regiões 3' não Traduzidas/genética , Adipocinas/metabolismo , Tecido Adiposo/transplante , Tecido Adiposo Marrom/citologia , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Marrom/transplante , Tecido Adiposo Branco/metabolismo , Tecido Adiposo Branco/transplante , Animais , Exossomos/genética , Fatores de Crescimento de Fibroblastos/sangue , Fatores de Crescimento de Fibroblastos/genética , Genes Reporter/genética , Teste de Tolerância a Glucose , Fígado/metabolismo , Masculino , Camundongos , MicroRNAs/genética , Modelos Biológicos , Especificidade de Órgãos/genética , RNA Mensageiro/genética , Ribonuclease III/deficiência , Ribonuclease III/genética , Transcrição Gênica
6.
Aging (Albany NY) ; 8(6): 1201-22, 2016 06.
Artigo em Inglês | MEDLINE | ID: mdl-27241713

RESUMO

Aging increases the risk of type 2 diabetes, and this can be prevented by dietary restriction (DR). We have previously shown that DR inhibits the downregulation of miRNAs and their processing enzymes - mainly Dicer - that occurs with aging in mouse white adipose tissue (WAT). Here we used fat-specific Dicer knockout mice (AdicerKO) to understand the contributions of adipose tissue Dicer to the metabolic effects of aging and DR. Metabolomic data uncovered a clear distinction between the serum metabolite profiles of Lox control and AdicerKO mice, with a notable elevation of branched-chain amino acids (BCAA) in AdicerKO. These profiles were associated with reduced oxidative metabolism and increased lactate in WAT of AdicerKO mice and were accompanied by structural and functional changes in mitochondria, particularly under DR. AdicerKO mice displayed increased mTORC1 activation in WAT and skeletal muscle, where Dicer expression is not affected. This was accompanied by accelerated age-associated insulin resistance and premature mortality. Moreover, DR-induced insulin sensitivity was abrogated in AdicerKO mice. This was reverted by rapamycin injection, demonstrating that insulin resistance in AdicerKO mice is caused by mTORC1 hyperactivation. Our study evidences a DR-modulated role for WAT Dicer in controlling metabolism and insulin resistance.


Assuntos
Tecido Adiposo Branco/metabolismo , Envelhecimento/metabolismo , RNA Helicases DEAD-box/metabolismo , Metabolismo Energético/fisiologia , Resistência à Insulina/fisiologia , Longevidade/genética , Ribonuclease III/metabolismo , Tecido Adiposo Branco/efeitos dos fármacos , Envelhecimento/genética , Animais , RNA Helicases DEAD-box/genética , Metabolismo Energético/efeitos dos fármacos , Longevidade/efeitos dos fármacos , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Metabolômica , Camundongos , Camundongos Knockout , Mitocôndrias/metabolismo , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/metabolismo , Ribonuclease III/genética , Sirolimo/farmacologia
7.
J Clin Invest ; 126(3): 837-53, 2016 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-26808499

RESUMO

Abnormal fibroblast function underlies poor wound healing in patients with diabetes; however, the mechanisms that impair wound healing are poorly defined. Here, we evaluated fibroblasts from individuals who had type 1 diabetes (T1D) for 50 years or more (Medalists, n = 26) and from age-matched controls (n = 7). Compared with those from controls, Medalist fibroblasts demonstrated a reduced migration response to insulin, lower VEGF expression, and less phosphorylated AKT (p-AKT), but not p-ERK, activation. Medalist fibroblasts were also functionally less effective at wound closure in nude mice. Activation of the δ isoform of protein kinase C (PKCδ) was increased in postmortem fibroblasts from Medalists, fibroblasts from living T1D subjects, biopsies of active wounds of living T1D subjects, and granulation tissues from mice with streptozotocin-induced diabetes. Diabetes-induced PKCD mRNA expression was related to a 2-fold increase in the mRNA half-life. Pharmacologic inhibition and siRNA-mediated knockdown of PKCδ or expression of a dominant-negative isoform restored insulin signaling of p-AKT and VEGF expression in vitro and improved wound healing in vivo. Additionally, increasing PKCδ expression in control fibroblasts produced the same abnormalities as those seen in Medalist fibroblasts. Our results indicate that persistent PKCδ elevation in fibroblasts from diabetic patients inhibits insulin signaling and function to impair wound healing and suggest PKCδ inhibition as a potential therapy to improve wound healing in diabetic patients.


Assuntos
Diabetes Mellitus Tipo 1/enzimologia , Pé Diabético/enzimologia , Fibroblastos/fisiologia , Proteína Quinase C-delta/fisiologia , Idoso , Idoso de 80 Anos ou mais , Animais , Hipóxia Celular , Movimento Celular , Proliferação de Células , Células Cultivadas , Diabetes Mellitus Tipo 1/complicações , Diabetes Mellitus Tipo 1/patologia , Pé Diabético/patologia , Feminino , Técnicas de Silenciamento de Genes , Meia-Vida , Humanos , Insulina/fisiologia , Masculino , Camundongos Nus , Pessoa de Meia-Idade , Proteína Quinase C-delta/antagonistas & inibidores , Inibidores de Proteínas Quinases/farmacologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Cicatrização
8.
J Clin Endocrinol Metab ; 101(3): 1225-34, 2016 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26756119

RESUMO

CONTEXT: HIV patients are at an increased risk for cardiometabolic disease secondary to depot-specific alterations in adipose function, but mechanisms remain poorly understood. OBJECTIVE: The endoribonuclease Dicer has been linked to the modulation of brown and white adipocyte differentiation. We previously demonstrated that Dicer knockout mice undergo transformation of brown adipose tissue to white adipose tissue and develop a lipodystrophic phenotype. We hypothesized reduced Dicer and brown adipose tissue gene expression from nonlipomatous sc fat among HIV patients with a lipodystrophic phenotype. DESIGN: Eighteen HIV (nine with and without lipodystrophic changes in fat distribution, characterized by excess dorsocervical adipose tissue [DCAT]) and nine non-HIV subjects underwent punch biopsy of abdominal sc fat to determine expression of Dicer and other adipose-related genes. RESULTS: HIV subjects with long-duration antiretroviral use demonstrated excess DCAT vs non-HIV subjects (9.8 ± 1.0 vs 6.6 ± 0.8 cm(2), P = .02) with similar body mass index. Dicer expression was decreased in abdominal sc fat of HIV vs non-HIV (4.88 [1.91, 11.93] vs 17.69 [10.72, 47.91], P = .01), as were PPARα, ZIC1, PRDM16, DIO2, and HSP60 (all P ≤ .03). Moreover, the expression of Dicer (2.49 [0.02, 4.88] vs 11.20 [4.83, 21.45], P = .006), brown fat (PPARα [P = .002], ZIC1 [P = .004], LHX8 [P = .03], PRDM16 [P = .0008], PAT2 [P = .008], P2RX5 [P = .02]), beige fat (TMEM26 [P = .004], CD137 [P = .008]), and other genes (DIO2 [P = .002], leptin [P = .003], HSP60 [P = .0004]) was further decreased in abdominal sc fat comparing HIV subjects with vs without excess DCAT. Down-regulation of Dicer in the abdominal sc fat correlated with the down-regulation of all brown and beige fat genes (all P ≤ .01). CONCLUSION: Our results demonstrate dysfunctional sc adipose tissue marked by reduced Dicer in relationship to the down-regulation of brown and beige fat-related genes in lipodystrophic HIV patients and may provide a novel mechanism for metabolic dysregulation. A strategy to increase browning of white adipose tissue may improve cardiometabolic health in HIV.


Assuntos
Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Branco/metabolismo , RNA Helicases DEAD-box/genética , Infecções por HIV/complicações , Síndrome de Lipodistrofia Associada ao HIV/genética , Ribonuclease III/genética , Gordura Subcutânea/metabolismo , Adipogenia/genética , Tecido Adiposo Marrom/patologia , Tecido Adiposo Branco/patologia , Dorso , Distribuição da Gordura Corporal , Estudos de Casos e Controles , RNA Helicases DEAD-box/metabolismo , Metabolismo Energético/genética , Expressão Gênica , Infecções por HIV/genética , Infecções por HIV/patologia , HIV-1 , Síndrome de Lipodistrofia Associada ao HIV/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Pescoço , Ribonuclease III/metabolismo , Gordura Subcutânea/patologia
9.
Sci Transl Med ; 6(247): 247ra103, 2014 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-25080478

RESUMO

White, beige, and brown adipocytes are developmentally and functionally distinct but often occur mixed together within individual depots. To target white, beige, and brown adipocytes for diagnostic or therapeutic purposes, a better understanding of the cell surface properties of these cell types is essential. Using a combination of in silico, in vitro, and in vivo methods, we have identified three new cell surface markers of adipose cell types. The amino acid transporter ASC-1 is a white adipocyte-specific cell surface protein, with little or no expression in brown adipocytes, whereas the amino acid transporter PAT2 and the purinergic receptor P2RX5 are cell surface markers expressed in classical brown and beige adipocytes in mice. These markers also selectively mark brown/beige and white adipocytes in human tissue. Thus, ASC-1, PAT2, and P2RX5 are membrane surface proteins that may serve as tools to identify and target white and brown/beige adipocytes for therapeutic purposes.


Assuntos
Adipócitos Marrons/metabolismo , Adipócitos Brancos/metabolismo , Sistema y+ de Transporte de Aminoácidos/metabolismo , Sistemas de Transporte de Aminoácidos Neutros/metabolismo , Membrana Celular/metabolismo , Receptores Purinérgicos P2X5/metabolismo , Simportadores/metabolismo , Adipócitos Marrons/efeitos dos fármacos , Adipócitos Brancos/efeitos dos fármacos , Agonistas de Receptores Adrenérgicos beta 3/farmacologia , Sistema y+ de Transporte de Aminoácidos/genética , Sistemas de Transporte de Aminoácidos Neutros/genética , Animais , Biomarcadores/metabolismo , Membrana Celular/efeitos dos fármacos , Temperatura Baixa , Biologia Computacional , Modelos Animais de Doenças , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Humanos , Masculino , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Obesidade/genética , Obesidade/metabolismo , Receptores Purinérgicos P2X5/genética , Simportadores/genética , Fatores de Tempo
10.
J Clin Invest ; 124(8): 3339-51, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24983316

RESUMO

miRNAs are important regulators of biological processes in many tissues, including the differentiation and function of brown and white adipocytes. The endoribonuclease dicer is a major component of the miRNA-processing pathway, and in adipose tissue, levels of dicer have been shown to decrease with age, increase with caloric restriction, and influence stress resistance. Here, we demonstrated that mice with a fat-specific KO of dicer develop a form of lipodystrophy that is characterized by loss of intra-abdominal and subcutaneous white fat, severe insulin resistance, and enlargement and "whitening" of interscapular brown fat. Additionally, KO of dicer in cultured brown preadipocytes promoted a white adipocyte-like phenotype and reduced expression of several miRNAs. Brown preadipocyte whitening was partially reversed by expression of miR-365, a miRNA known to promote brown fat differentiation; however, introduction of other miRNAs, including miR-346 and miR-362, also contributed to reversal of the loss of the dicer phenotype. Interestingly, fat samples from patients with HIV-related lipodystrophy exhibited a substantial downregulation of dicer mRNA expression. Together, these findings indicate the importance of miRNA processing in white and brown adipose tissue determination and provide a potential link between this process and HIV-related lipodystrophy.


Assuntos
Adipócitos Marrons/metabolismo , Adipócitos Brancos/citologia , Lipodistrofia/genética , Lipodistrofia/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Adipócitos Marrons/citologia , Animais , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Estudos de Coortes , RNA Helicases DEAD-box/deficiência , RNA Helicases DEAD-box/genética , RNA Helicases DEAD-box/metabolismo , Modelos Animais de Doenças , Regulação para Baixo , Metabolismo Energético , Feminino , Síndrome de Lipodistrofia Associada ao HIV/genética , Síndrome de Lipodistrofia Associada ao HIV/metabolismo , Síndrome de Lipodistrofia Associada ao HIV/patologia , Humanos , Resistência à Insulina , Lipodistrofia/patologia , Masculino , Camundongos , Camundongos Knockout , Processamento Pós-Transcricional do RNA , Ribonuclease III/deficiência , Ribonuclease III/genética , Ribonuclease III/metabolismo
11.
Cell Metab ; 17(5): 644-656, 2013 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-23583168

RESUMO

Fat distribution is closely linked to metabolic disease risk. Distribution varies with sex, genetic background, disease state, certain drugs and hormones, development, and aging. Preadipocyte replication and differentiation, developmental gene expression, susceptibility to apoptosis and cellular senescence, vascularity, inflammatory cell infiltration, and adipokine secretion vary among depots, as do fatty-acid handling and mechanisms of enlargement with positive-energy and loss with negative-energy balance. How interdepot differences in these molecular, cellular, and pathophysiological properties are related is incompletely understood. Whether fat redistribution causes metabolic disease or whether it is a marker of underlying processes that are primarily responsible is an open question.


Assuntos
Gorduras/metabolismo , Doenças Metabólicas/metabolismo , Tecido Adiposo/metabolismo , Animais , Metabolismo Energético , Humanos
12.
Cell Metab ; 16(3): 336-47, 2012 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-22958919

RESUMO

Excess adipose tissue is associated with metabolic disease and reduced life span, whereas caloric restriction decreases these risks. Here we show that as mice age, there is downregulation of Dicer and miRNA processing in adipose tissue resulting in decreases of multiple miRNAs. A similar decline of Dicer with age is observed in C. elegans. This is prevented in both species by caloric restriction. Decreased Dicer expression also occurs in preadipocytes from elderly humans and can be produced in cells by exposure to oxidative stress or UV radiation. Knockdown of Dicer in cells results in premature senescence, and fat-specific Dicer knockout renders mice hypersensitive to oxidative stress. Finally, Dicer loss-of-function mutations in worms reduce life span and stress tolerance, while intestinal overexpression of Dicer confers stress resistance. Thus, regulation of miRNA processing in adipose-related tissues plays an important role in longevity and the ability of an organism to respond to environmental stress and age-related disease.


Assuntos
Tecido Adiposo/metabolismo , Envelhecimento/metabolismo , Regulação Enzimológica da Expressão Gênica/fisiologia , Longevidade/fisiologia , MicroRNAs/metabolismo , Estresse Oxidativo/genética , Ribonuclease III/metabolismo , Animais , Evolução Biológica , Caenorhabditis elegans , Técnicas de Cultura de Células , Primers do DNA/genética , Humanos , Longevidade/genética , Camundongos , Camundongos Knockout , Ribonuclease III/genética
13.
Endocrinology ; 152(12): 4571-80, 2011 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-22009727

RESUMO

Substance P (SP), encoded by the tachykinin 1 (Tac1) gene, is the most potent tachykinin ligand for the high-affinity neurokinin-1 receptor (NK-1R). We previously reported that NK-1R-deficient mice show less weight gain and reduced circulating levels of leptin and insulin in response to a high-fat diet (HFD) and demonstrated the presence of functional NK-1R in isolated human preadipocytes. Here we assessed the effects of SP on weight gain in response to HFD and determined glucose metabolism in Tac1-deficient (Tac1(-/-)) mice. The effect of SP on the expression of molecules that may predispose to reduced glucose uptake was also determined in isolated human mesenteric, omental, and sc preadipocytes. We show that although weight accumulation in response to HFD was similar between Tac1(-/-) mice and wild-type littermates, Tac1(-/-) mice demonstrated lower glucose and leptin and increased adiponectin blood levels and showed improved responses to insulin challenge after HFD. SP stimulated phosphorylation of c-Jun N-terminal kinase, protein kinase C, mammalian target of rapamycin, and inhibitory serine insulin receptor substrate-1 phosphorylation in human preadipocytes in vitro. Preincubation of human mesenteric preadipocytes with the protein kinase C pseudosubstrate inhibitor reduced insulin receptor substrate 1 phosphorylation in response to SP. Lastly, SP also induced insulin receptor substrate-1 phosphorylation in mature human sc adipocytes. Our results demonstrate an important role for SP in adipose tissue responses and obesity-associated pathologies. These novel SP effects on molecules that enhance insulin resistance at the adipocyte level may reflect an important role for this peptide in the pathophysiology of type 2 diabetes.


Assuntos
Glucose/metabolismo , Insulina/metabolismo , Receptores da Neurocinina-1/fisiologia , Transdução de Sinais , Substância P/fisiologia , Adipócitos/metabolismo , Animais , Gorduras na Dieta/administração & dosagem , Humanos , Resistência à Insulina , Camundongos , Camundongos Knockout , Taquicininas/deficiência , Taquicininas/fisiologia , Aumento de Peso
14.
Mol Endocrinol ; 25(5): 799-809, 2011 May.
Artigo em Inglês | MEDLINE | ID: mdl-21436255

RESUMO

Vitamin A metabolite retinoic acid (RA) regulates life-sustaining differentiation processes and metabolic homeostasis. The aldehyde dehydrogenase-1 (Aldh1) family of enzymes (Aldh1a1, a2, and a3) catalyzes RA production from retinaldehyde and thereby controls concentrations of this transcriptionally active metabolite. The hierarchy of Aldh1 functions in adipose tissue has not been elucidated. We hypothesized that Aldh1 enzymes produce endogenous RA and regulate adipogenesis and fat formation in a fat depot-specific manner. We demonstrate that adipogenesis in vitro is accompanied by RA production generated primarily by Aldh1a1. In Aldh1a1-deficient adipocytes, adipogenesis is impaired compared with wild-type adipocytes due to markedly reduced expression of PPARγ regulated through zinc-finger protein 423 (ZFP423)-dependent mechanisms. These effects were recovered to some extent either by RA stimulation or overexpression of any of the Aldh1 enzymes in Aldh1a1(-/-) cells arguing that Aldh1a1 plays a dominant role in autocrine RA production. In vivo studies in C57/BL6 and Aldh1a1(-/-) mice on a regular diet revealed that multiple Aldh1 enzymes regulate differences in the formation of sc and visceral fat. In Aldh1a1(-/-) mice, visceral fat essentially lacked all Aldh1 expression. This loss of RA-producing enzymes was accompanied by 70% decreased expression of ZFP423, PPARγ, and Fabp4 in visceral fat of Aldh1a1(-/-) vs. wild-type mice and by the predominant loss of visceral fat. Subcutaneous fat of Aldh1a1(-/-) mice expressed Aldh1a3 for RA production that was sufficient to maintain expression of ZFP423 and PPARγ and sc fat mass. Our data suggest a paradigm for regulation of fat depots through the concerted action of Aldh1 enzymes that establish RA-dependent tandem regulation of transcription factors ZFP423 and PPARγ in a depot-specific manner.


Assuntos
Adipogenia , Isoenzimas/metabolismo , Retinal Desidrogenase/metabolismo , Células 3T3-L1 , Adipócitos/citologia , Adipócitos/enzimologia , Adulto , Família Aldeído Desidrogenase 1 , Animais , Distribuição da Gordura Corporal , Fator de Ligação a CCAAT/metabolismo , Proteínas de Ligação ao Cálcio , Diferenciação Celular , Citocinas/metabolismo , Proteínas de Ligação a DNA/metabolismo , Feminino , Genes Reporter , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Gordura Intra-Abdominal/metabolismo , Luciferases/biossíntese , Camundongos , Camundongos Endogâmicos C57BL , Nicotinamida Fosforribosiltransferase/metabolismo , PPAR gama/genética , PPAR gama/metabolismo , Elementos de Resposta , Gordura Subcutânea/metabolismo , Fatores de Transcrição/metabolismo , Transcrição Gênica , Tretinoína/metabolismo
15.
Gerontology ; 57(1): 66-75, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-20110661

RESUMO

Fat mass and fat tissue distribution change dramatically throughout life. In old age, fat becomes dysfunctional and is redistributed from subcutaneous to intra-abdominal visceral depots as well as other ectopic sites, including bone marrow, muscle and the liver. These changes are associated with increased risk of metabolic syndrome. Fat tissue is a nutrient storage, endocrine and immune organ that undergoes renewal throughout the lifespan. Preadipocytes, which account for 15-50% of cells in fat tissue, give rise to new fat cells. With aging, declines in preadipocyte proliferation and differentiation likely contribute to increased systemic exposure to lipotoxic free fatty acids. Age-related fat tissue inflammation is related to changes that occur in preadipocytes and macrophages in a fat depot-dependent manner. Fat tissue inflammation frequently leads to further reduction in adipogenesis with aging, more lipotoxicity and activation of cellular stress pathways that, in turn, exacerbate inflammatory responses of preadipocytes and immune cells, establishing self-perpetuating cycles that lead to systemic dysfunction. In this review, we will consider how inherent, age-related, depot-dependent alterations in preadipocyte function contribute to age-related fat tissue redistribution and metabolic dysfunction.


Assuntos
Adipócitos/citologia , Células-Tronco Adultas/citologia , Envelhecimento/patologia , Adipogenia/fisiologia , Tecido Adiposo/fisiologia , Adulto , Idoso , Envelhecimento/fisiologia , Distribuição da Gordura Corporal , Feminino , Humanos , Mediadores da Inflamação/fisiologia , Metabolismo dos Lipídeos , Masculino , Pessoa de Meia-Idade , Modelos Biológicos
16.
Obesity (Silver Spring) ; 18(10): 1875-80, 2010 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-20300084

RESUMO

To elucidate cellular mechanisms of sex-related differences in fat distribution, we determined body fat distribution (dual-energy X-ray absorptiometry and single-slice abdominal computed tomography (CT)), adipocyte size, adipocyte number, and proportion of early-differentiated adipocytes (aP2(+)CD68(-)) in the stromovascular fraction (SVF) in the upper and lower body of normal-weight healthy men (n = 12) and premenopausal women (n = 20) (age: 18-49 years, BMI: 18-26 kg/m(2)). Women had more subcutaneous and less visceral fat than men. The proportion of early differentiated adipocytes in the subcutaneous adipose tissue SVF of women was greater than in men (P = 0.01), especially in the femoral depot, although in vitro adipogenesis, as assessed by peroxisome proliferator activated receptor-γ (PPARγ) expression, was not increased in femoral preadipocytes cultured from women compared with men. In women, differentiation of femoral preadipocytes was less than that of abdominal subcutaneous preadipocytes (P = 0.04), and femoral subcutaneous preadipocytes tended to be more resistant to tumor necrosis factor-α (TNFα)-induced apoptosis (P = 0.06). Thus, turnover and utilization of the preadipocyte pool may be reduced in lower vs. the upper-body fat in women. Collectively, these data indicate that the microenvironment, rather than differences in inherent properties of preadipocytes between genders, may explain the gynoid obesity phenotype and higher percent body fat in women compared to men.


Assuntos
Adipócitos , Adipogenia , Distribuição da Gordura Corporal , Gordura Intra-Abdominal , Obesidade , Caracteres Sexuais , Gordura Subcutânea , Adipócitos/citologia , Adipócitos/metabolismo , Adiposidade , Adolescente , Adulto , Apoptose , Feminino , Fêmur , Humanos , Gordura Intra-Abdominal/citologia , Gordura Intra-Abdominal/metabolismo , Masculino , Pessoa de Meia-Idade , Obesidade/metabolismo , Obesidade/patologia , PPAR gama/metabolismo , Valores de Referência , Gordura Subcutânea/citologia , Gordura Subcutânea/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Adulto Jovem
17.
J Gerontol A Biol Sci Med Sci ; 65(3): 242-51, 2010 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-20106964

RESUMO

Fat distribution changes with aging. Inherent changes in fat cell progenitors may contribute because fat cells turn over throughout life. To define mechanisms, gene expression was profiled in preadipocytes cultured from epididymal and perirenal depots of young and old rats. 8.4% of probe sets differed significantly between depots, particularly developmental genes. Only 0.02% differed with aging, despite using less stringent criteria than for comparing depots. Twenty-five genes selected based on fold change with aging were analyzed in preadipocytes from additional young, middle-aged, and old animals by polymerase chain reaction. Thirteen changed significantly with aging, 13 among depots, and 9 with both. Genes involved in inflammation, stress, and differentiation changed with aging, as occurs in fat tissue. Age-related changes were greater in perirenal than epididymal preadipocytes, consistent with larger declines in replication and adipogenesis in perirenal preadipocytes. Thus, age-related changes in preadipocyte gene expression differ among depots, potentially contributing to fat redistribution and dysfunction.


Assuntos
Adipócitos/metabolismo , Envelhecimento/genética , Proteínas de Transporte/genética , Regulação da Expressão Gênica no Desenvolvimento , Lectinas Tipo C/genética , Proteínas de Membrana/genética , RNA/genética , Adipócitos/citologia , Envelhecimento/metabolismo , Animais , Western Blotting , Distribuição da Gordura Corporal , Proteínas de Transporte/biossíntese , Células Cultivadas , Lectinas Tipo C/biossíntese , Masculino , Proteínas de Membrana/biossíntese , Proteínas dos Microtúbulos , Reação em Cadeia da Polimerase , Prognóstico , Ratos
18.
Am J Physiol Gastrointest Liver Physiol ; 296(5): G1012-9, 2009 May.
Artigo em Inglês | MEDLINE | ID: mdl-19282377

RESUMO

White adipose tissue is intimately involved in the regulation of immunity and inflammation. We reported that human mesenteric preadipocytes express the substance P (SP)-mediated neurokinin-1 receptor (NK-1R), which signals proinflammatory responses. Here we tested the hypothesis that SP promotes proliferation and survival of human mesenteric preadipocytes and investigated responsible mechanism(s). Preadipocytes were isolated from mesenteric fat biopsies during gastric bypass surgery. Proliferative and antiapoptotic responses were delineated in 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS), bromodeoxyuridine (BrdU), caspase-3, and TUNEL assays, as well as Western immunoanalysis. SP (10(-7) M) increased MTS and proliferation (BrdU) and time dependently (15-30 min) induced Akt, EGF receptor, IGF receptor, integrin alphaVbeta3, phosphatidylinositol 3-kinase, and PKC-theta phosphorylation. Furthermore, pharmacological antagonism of Akt and PKC-theta activation significantly attenuated SP-induced preadipocyte proliferation. Exposure of preadipocytes to the proapoptotic Fas ligand (FasL, 100 microM) resulted in nuclear DNA fragmentation (TUNEL assay), as well as increased cleaved poly (ADP-ribose) polymerase, cleaved caspase-7, and caspase-3 expression. Cotreatment with SP almost completely abolished these responses in a NK-1R-dependent fashion. SP (10(-7) M) also time dependently stimulated expression 4E binding protein 1 and phosphorylation of p70 S6 kinase, which increased protein translation efficiency. SP increases preadipocyte viability, reduces apoptosis, and stimulates proliferation, possibly via cell cycle upregulation and increased protein translation efficiency. SP-induced proliferative and antiapoptotic pathways in fat depots may contribute to development of the creeping fat and inflammation characteristic of Crohn's disease.


Assuntos
Adipócitos/metabolismo , Apoptose , Proliferação de Células , Gordura Intra-Abdominal/metabolismo , Transdução de Sinais , Substância P/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adipócitos/efeitos dos fármacos , Adipócitos/enzimologia , Adipócitos/patologia , Apoptose/efeitos dos fármacos , Caspase 3/metabolismo , Caspase 7/metabolismo , Ciclo Celular , Proteínas de Ciclo Celular , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular , Células Cultivadas , Receptores ErbB/metabolismo , Proteína Ligante Fas/metabolismo , Humanos , Integrina alfaVbeta3/metabolismo , Gordura Intra-Abdominal/efeitos dos fármacos , Gordura Intra-Abdominal/enzimologia , Gordura Intra-Abdominal/patologia , Isoenzimas/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Inibidores de Fosfoinositídeo-3 Quinase , Fosfoproteínas/metabolismo , Fosforilação , Poli(ADP-Ribose) Polimerases/metabolismo , Proteína Quinase C/metabolismo , Proteína Quinase C-theta , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptor IGF Tipo 1/metabolismo , Receptores da Neurocinina-1/metabolismo , Proteínas Quinases S6 Ribossômicas 70-kDa/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fatores de Tempo , Fator de Necrose Tumoral alfa/metabolismo
19.
Am J Physiol Endocrinol Metab ; 293(6): E1810-9, 2007 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-17911345

RESUMO

Fat depot sizes peak in middle age but decrease by advanced old age. This phenomenon is associated with ectopic fat deposition, decreased adipocyte size, impaired differentiation of preadipocytes into fat cells, decreased adipogenic transcription factor expression, and increased fat tissue inflammatory cytokine generation. To define the mechanisms contributing to impaired adipogenesis with aging, we examined the release of TNFalpha, which inhibits adipogenesis, and the expression of CCAAT/enhancer-binding protein (C/EBP) homologous protein (CHOP), which blocks activity of adipogenic C/EBP family members, in preadipocytes cultured from young, middle-aged, and old rats. Medium conditioned by fat tissue, as well as preadipocytes, from old rats impeded lipid accumulation by preadipocytes from young animals. More TNFalpha was released by preadipocytes from old than young rats. Differences in TNFalpha-converting enzyme, TNFalpha degradation, or the presence of macrophages in cultures were not responsible. TNFalpha induced rat preadipocyte CHOP expression. CHOP was higher in undifferentiated preadipocytes from old than younger animals. Overexpression of CHOP in young rat preadipocytes inhibited lipid accumulation. TNFalpha short interference RNA reduced CHOP and partially restored lipid accumulation in old rat preadipocytes. CHOP normally increases during late differentiation, potentially modulating the process. This late increase in CHOP was not affected substantially by aging: CHOP was similar in differentiating preadipocytes and fat tissue from old and young animals. Hypoglycemia, which normally causes an adaptive increase in CHOP, was less effective in inducing CHOP in preadipocytes from old than younger animals. Thus increased TNFalpha release by undifferentiated preadipocytes with elevated basal CHOP contributes to impaired adipogenesis with aging.


Assuntos
Adipócitos/metabolismo , Adipogenia/efeitos dos fármacos , Envelhecimento/fisiologia , Fator de Transcrição CHOP/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Proteínas ADAM/metabolismo , Proteína ADAM17 , Adipócitos/citologia , Adipócitos/efeitos dos fármacos , Tecido Adiposo/citologia , Tecido Adiposo/metabolismo , Animais , Western Blotting , Células Cultivadas , Técnicas de Cocultura , Meios de Cultivo Condicionados/farmacologia , Epididimo/citologia , Rim/citologia , Masculino , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/genética , Ratos , Ratos Endogâmicos BN , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fator de Transcrição CHOP/genética , Transfecção , Fator de Necrose Tumoral alfa/genética
20.
Am J Physiol Endocrinol Metab ; 292(1): E298-307, 2007 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-16985259

RESUMO

Anatomically separate fat depots differ in size, function, and contribution to pathological states, such as the metabolic syndrome. We isolated preadipocytes from different human fat depots to determine whether the basis for this variation is partly attributable to differences in inherent properties of fat cell progenitors. We found that genome-wide expression profiles of primary preadipocytes cultured in parallel from abdominal subcutaneous, mesenteric, and omental fat depots were distinct. Interestingly, visceral fat was not homogeneous. Preadipocytes from one of the two main visceral depots, mesenteric fat, had an expression profile closer to that of subcutaneous than omental preadipocytes, the other main visceral depot. Expression of genes that regulate early development, including homeotic genes, differed extensively among undifferentiated preadipocytes isolated from different fat depots. These profiles were confirmed by real-time PCR analysis of preadipocytes from additional lean and obese male and female subjects. We made preadipocyte strains from single abdominal subcutaneous and omental preadipocytes by expressing telomerase. Depot-specific developmental gene expression profiles persisted for 40 population doublings in these strains. Thus, human fat cell progenitors from different regions are effectively distinct, consistent with different fat depots being separate mini-organs.


Assuntos
Tecido Adiposo/metabolismo , Perfilação da Expressão Gênica/métodos , Genes Controladores do Desenvolvimento , Células-Tronco/metabolismo , Adulto , Linhagem Celular Transformada , Análise por Conglomerados , Feminino , Humanos , Gordura Intra-Abdominal/metabolismo , Masculino , Análise em Microsséries , Especificidade de Órgãos , Gordura Subcutânea/metabolismo , Telomerase/genética
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