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1.
Life Sci ; 257: 118055, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32634429

RESUMO

AIMS: Human adipose derived mesenchymal stem cells (hAD-MSCs) as the most promising target for cell therapy and regenerative medicine, face senescence as a major drawback resulting in their limited proliferation and differentiation potentials. To evaluate the efficacy of miR-34a silencing as an anti-senescence strategy in hAD-MSCs, in this study common hallmarks of senescence were assessed after transient inhibition of miR-34a in hAD-MSCs. MATERIALS AND METHODS: The expression levels of miR-34a in hAD-MSCs at different passages were evaluated by real-time quantitative PCR. hAD-MSCs at passage 2 and passage 7 were transfected with miR-34a inhibitor. Doubling time assay, colony forming assay, and cell cycle analysis were performed to evaluate cell proliferation rate. The activity of senescence associated ß-galactosidase (SA-ß-gal) was assessed by histochemical staining. Moreover, the senescence associated molecular alterations including that of pro-senescence (P53, P21 and P16) and anti-senescence (SIRT1, HTERT and CD44) genes were examined by quantitative RT-PCR and western blot assays. To evaluate the differentiation potentials of MSCs, following adipogenic and osteogenic induction, the expression levels of lineage specific markers were analyzed by qPCR. KEY FINDINGS: Our results showed that inhibition of miR-34a enhances the proliferation, promotes the adipogenic and osteogenic differentiation potency, reduces the senescence associated-ß gal activity, and reverses the senescence associated molecular alterations in hAD-MSCs. SIGNIFICANCE: In this study, we showed that inhibition of miR-34a reduces the cellular senescence through the activation of SIRT1. Our findings support the silencing of miR-34a as an anti-senescence approach to improve the therapeutic potentials of hAD-MSCs.


Assuntos
Diferenciação Celular/fisiologia , Senescência Celular/fisiologia , Células-Tronco Mesenquimais/citologia , MicroRNAs/genética , Sirtuína 1/genética , Adipogenia/fisiologia , Tecido Adiposo/citologia , Inativação Gênica , Humanos , Receptores de Hialuronatos/genética , Osteogênese/fisiologia , Telomerase/genética
2.
Am J Physiol Endocrinol Metab ; 319(2): E363-E375, 2020 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-32603262

RESUMO

Bone morphogenetic protein (BMP) receptor signaling is critical for the regulation of the endocrine system and cardiovascular structure and function. The objective of this study was to investigate whether Bmp3b, a glycoprotein synthetized and secreted by adipose tissue, is necessary to regulate glucose and lipid metabolism, adipogenesis, and cardiovascular remodeling. Over the course of 4 mo, Bmp3b-knockout (Bmp3b-/-) mice gained more weight than wild-type (WT) mice. The plasma levels of cholesterol and triglycerides were higher in Bmp3b-/- mice than in WT mice. Bmp3b-/- mice developed insulin resistance and glucose intolerance. The basal heart rate was higher in Bmp3b-/- mice than in WT mice, and echocardiography revealed eccentric remodeling in Bmp3b-/- mice. The expression of adipogenesis-related genes in white adipose tissue was higher in Bmp3b-/- mice than in WT control mice. In vitro studies showed that Bmp3b modulates the activity of the C/ebpα promoter, an effect mediated by Smad2/3. The results of this study suggest that Bmp3b is necessary for the maintenance of homeostasis in terms of age-related weight gain, glucose metabolism, and left ventricular (LV) remodeling and function. Interventions that increase the level or function of BMP3b may decrease cardiovascular risk and pathological cardiac remodeling.


Assuntos
Adipogenia/fisiologia , Fator 10 de Diferenciação de Crescimento/deficiência , Fator 10 de Diferenciação de Crescimento/fisiologia , Síndrome Metabólica/etiologia , Adipócitos/patologia , Tecido Adiposo/patologia , Animais , Proteína Morfogenética Óssea 3/deficiência , Proteína Morfogenética Óssea 3/fisiologia , Dislipidemias/etiologia , Feminino , Intolerância à Glucose/etiologia , Cardiopatias/etiologia , Cardiopatias/fisiopatologia , Resistência à Insulina/fisiologia , Masculino , Síndrome Metabólica/fisiopatologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Obesidade/etiologia , Obesidade/patologia , Transdução de Sinais/fisiologia
3.
Mol Cell Biol ; 40(17)2020 08 14.
Artigo em Inglês | MEDLINE | ID: mdl-32601106

RESUMO

Transcription factors C/EBPß and C/EBPδ are induced within hours after initiation of adipogenesis in culture. They directly promote the expression of master adipogenic transcription factors peroxisome proliferator-activated receptor γ (PPARγ) and C/EBPα and are required for adipogenesis in vivo However, the mechanism that controls the induction of C/EBPß and C/EBPδ remains elusive. We previously showed that histone methyltransferases MLL3/MLL4 and associated PTIP are required for the induction of PPARγ and C/EBPα during adipogenesis. Here, we show MLL3/MLL4/PTIP-associated protein PAGR1 (also known as PA1) cooperates with phosphorylated CREB and ligand-activated glucocorticoid receptor to directly control the induction of C/EBPß and C/EBPδ in the early phase of adipogenesis. Deletion of Pagr1 in white and brown preadipocytes prevents the induction of C/EBPß and C/EBPδ and leads to severe defects in adipogenesis. Adipogenesis defects in PAGR1-deficient cells can be rescued by the ectopic expression of C/EBPß or PPARγ. Finally, the deletion of Pagr1 in Myf5+ precursor cells impairs brown adipose tissue and muscle development. Thus, by controlling the induction of C/EBPß and C/EBPδ, PAGR1 plays a critical role in adipogenesis.


Assuntos
Adipogenia/fisiologia , Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Proteína delta de Ligação ao Facilitador CCAAT/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ligação a DNA/metabolismo , Histona-Lisina N-Metiltransferase/metabolismo , Células 3T3-L1 , Adipócitos/metabolismo , Tecido Adiposo Marrom/metabolismo , Animais , Diferenciação Celular/fisiologia , Histona Metiltransferases/metabolismo , Camundongos , Camundongos Knockout , PPAR gama/metabolismo , Ligação Proteica
4.
Nat Commun ; 11(1): 2303, 2020 05 08.
Artigo em Inglês | MEDLINE | ID: mdl-32385276

RESUMO

White adipose tissue (WAT) expansion in obesity occurs through enlargement of preexisting adipocytes (hypertrophy) and through formation of new adipocytes (adipogenesis). Adipogenesis results in WAT hyperplasia, smaller adipocytes and a metabolically more favourable form of obesity. How obesogenic WAT hyperplasia is induced remains, however, poorly understood. Here, we show that the mechanosensitive cationic channel Piezo1 mediates diet-induced adipogenesis. Mice lacking Piezo1 in mature adipocytes demonstrated defective differentiation of preadipocyte into mature adipocytes when fed a high fat diet (HFD) resulting in larger adipocytes, increased WAT inflammation and reduced insulin sensitivity. Opening of Piezo1 in mature adipocytes causes the release of the adipogenic fibroblast growth factor 1 (FGF1), which induces adipocyte precursor differentiation through activation of the FGF-receptor-1. These data identify a central feed-back mechanism by which mature adipocytes control adipogenesis during the development of obesity and suggest Piezo1-mediated adipocyte mechano-signalling as a mechanism to modulate obesity and its metabolic consequences.


Assuntos
Adipócitos/metabolismo , Fator 1 de Crescimento de Fibroblastos/metabolismo , Canais Iônicos/metabolismo , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/metabolismo , Adipogenia/fisiologia , Tecido Adiposo Branco/metabolismo , Animais , Calorimetria , Células Cultivadas , Feminino , Fator 1 de Crescimento de Fibroblastos/genética , Citometria de Fluxo , Humanos , Imuno-Histoquímica , Marcação In Situ das Extremidades Cortadas , Insulina/sangue , Interleucina-6/sangue , Canais Iônicos/genética , Masculino , Camundongos , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Transdução de Sinais/genética , Transdução de Sinais/fisiologia
5.
Nat Commun ; 11(1): 2306, 2020 05 08.
Artigo em Inglês | MEDLINE | ID: mdl-32385399

RESUMO

During ß-adrenergic stimulation of brown adipose tissue (BAT), p38 phosphorylates the activating transcription factor 2 (ATF2) which then translocates to the nucleus to activate the expression of Ucp1 and Pgc-1α. The mechanisms underlying ATF2 target activation are unknown. Here we demonstrate that p62 (Sqstm1) binds to ATF2 to orchestrate activation of the Ucp1 enhancer and Pgc-1α promoter. P62Δ69-251 mice show reduced expression of Ucp1 and Pgc-1α with impaired ATF2 genomic binding. Modulation of Ucp1 and Pgc-1α expression through p62 regulation of ATF2 signaling is demonstrated in vitro and in vivo in p62Δ69-251 mice, global p62-/- and Ucp1-Cre p62flx/flx mice. BAT dysfunction resulting from p62 deficiency is manifest after birth and obesity subsequently develops despite normal food intake, intestinal nutrient absorption and locomotor activity. In summary, our data identify p62 as a master regulator of BAT function in that it controls the Ucp1 pathway through regulation of ATF2 genomic binding.


Assuntos
Fator 2 Ativador da Transcrição/metabolismo , Proteína Sequestossoma-1/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Adipogenia/fisiologia , Tecido Adiposo Marrom/diagnóstico por imagem , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Branco/diagnóstico por imagem , Tecido Adiposo Branco/metabolismo , Animais , Núcleo Celular/metabolismo , Imagem por Ressonância Magnética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Obesidade/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Tomografia Computadorizada com Tomografia por Emissão de Pósitrons , Ligação Proteica , Proteína Sequestossoma-1/genética , Proteína Desacopladora 1/metabolismo
6.
Metabolism ; 108: 154250, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32335074

RESUMO

BACKGROUND: Chronic steroid treatment causes an increase in visceral adiposity and osteoporosis. It is believed that steroids may alter a balance between differentiation of mesenchymal stem cells (MSCs) into either adipocytes or osteoblasts; however, the detailed molecular mechanisms are unclear. We previously identified Dexras1 as a critical factor that potentiates adipogenesis in response to glucocorticoids. Thus, in this study, we investigated the role of Dexras1 in maintaining the balance between chronic steroid treatment-associated adipogenesis and osteoporosis. MATERIAL AND METHODS: We treated wild type (WT) and Dexras1 knockout (KO) mice with dexamethasone for five weeks followed by 60% HFD for additional two weeks with dexamethasone. The changes of glucocorticoid-induced body weight gain and osteoporosis were analyzed. Bone marrow derived stromal cells (BMSCs) and mouse embryonic fibroblasts (MEFs) extracted from WT and Dexras1 KO mice, as well as MC3T3-E1 pre-osteoblasts and osteoclasts differentiated from RAW264.7 were analyzed to further define the role of Dexras1 in osteoblasts and osteoclasts. RESULTS: Dual-energy X-ray absorptiometry and micro-computed tomography analyses in murine femurs revealed that Dexras1 deficiency was associated with increased osteogenesis, concurrent with reduced adipogenesis. Furthermore, Dexras1 deficiency promoted osteogenesis of BMSCs and MEFs in vitro, suggesting that Dexras1 deficiency prevents steroid-induced osteoporosis. We also observed that Dexras1 downregulated SMAD signaling pathways, which reduced the osteogenic differentiation capacity of pre-osteoblast MC3T3-E1 cells into mature osteoblasts. CONCLUSION: We propose that Dexras1 is critical for maintaining the equilibrium between adipogenesis and osteogenesis upon steroid treatment.


Assuntos
Adipogenia/fisiologia , Osteogênese/fisiologia , Proteínas ras/metabolismo , Células 3T3 , Adipócitos/metabolismo , Animais , Diferenciação Celular/fisiologia , Linhagem Celular , Fêmur/metabolismo , Glucocorticoides/metabolismo , Masculino , Células-Tronco Mesenquimais/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Osteoblastos/metabolismo , Osteoclastos/metabolismo , Osteoporose/metabolismo , Células RAW 264.7 , Transdução de Sinais/fisiologia
7.
Anim Genet ; 51(3): 351-357, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32253788

RESUMO

Intramuscular fat (IMF) content is a critical factor affecting meat flavor, juiciness, tenderness, and color. Therefore, the improvement of IMF content is one of the hotspots of animal science research. Fat deposition is the result of a combination of increased number of fat cells and cellular hypertrophy. In addition, transcription factors can influence the number of adipocytes and regulate lipid metabolism. The progress of the transcription factors regulating adipocyte differentiation in beef cattle, including IMF cell sources, and promoting or inhibiting adipogenic differentiation of transcription factors is reviewed in this paper.


Assuntos
Adipócitos/metabolismo , Adipogenia/fisiologia , Bovinos/fisiologia , Músculo Esquelético/metabolismo , Fatores de Transcrição/genética , Animais , Bovinos/genética , Diferenciação Celular , Fatores de Transcrição/metabolismo
8.
Metabolism ; 105: 154189, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32105664

RESUMO

BACKGROUND: Sprouty (SPRY) proteins play critical roles in controlling cell proliferation, differentiation, and survival by inhibiting receptor tyrosine kinase (RTK)-mediated extracellular signal-regulated kinase (ERK) signaling. Recent studies have demonstrated that SPRY4 negatively regulates angiogenesis and tumor growth. However, whether SPRY4 regulates osteogenic and/or adipogenic differentiation of mesenchymal stem cells remains to be explored. RESULTS: In this study, we investigated the expression pattern of Spry4 and found that its expression was regulated during the differentiation of mouse marrow stromal progenitor cells and increased in the metaphysis of ovariectomized mice. In vitro loss-of-function and gain-of-function studies demonstrated that SPRY4 inhibited osteogenic differentiation and stimulated adipogenic differentiation of progenitor cells. In vivo experiments showed that silencing of Spry4 in the marrow of C57BL/6 mice blocked fat accumulation and promoted osteoblast differentiation in ovariectomized mice. Mechanistic investigations revealed the inhibitory effect of SPRY4 on canonical wingless-type MMTV integration site (Wnt) signaling and ERK pathway. ERK1/2 was shown to interact with low-density lipoprotein receptor-related protein 6 (LRP6) and activate the canonical Wnt signaling pathway. Inactivation of Wnt signaling attenuated the inhibition of adipogenic differentiation and stimulation of osteogenic differentiation by Spry4 small interfering RNA (siRNA). Finally, promoter study revealed that ß-catenin transcriptionally inhibited the expression of Spry4. CONCLUSIONS: Our study for the first time suggests that a novel SPRY4-ERK1/2-Wnt/ß-catenin regulatory loop exists in marrow stromal progenitor cells and plays a key role in cell fate determination. It also highlights the potential of SPRY4 as a novel therapeutic target for the treatment of metabolic bone disorders such as osteoporosis.


Assuntos
Adipogenia/genética , Adipogenia/fisiologia , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Sistema de Sinalização das MAP Quinases/genética , Sistema de Sinalização das MAP Quinases/fisiologia , Células-Tronco Mesenquimais/fisiologia , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/fisiologia , Osteogênese/genética , Osteogênese/fisiologia , Via de Sinalização Wnt/genética , Via de Sinalização Wnt/fisiologia , beta Catenina/genética , beta Catenina/fisiologia , Animais , Medula Óssea/metabolismo , Feminino , Proteína-6 Relacionada a Receptor de Lipoproteína de Baixa Densidade/genética , Proteína-6 Relacionada a Receptor de Lipoproteína de Baixa Densidade/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Ovariectomia , RNA Interferente Pequeno/farmacologia
9.
Obesity (Silver Spring) ; 28(2): 293-302, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31970913

RESUMO

OBJECTIVE: The objectives of this study were to assess the role of mitochondrial pyruvate carriers (MPCs) in adipocyte development in vitro and determine whether MPCs are regulated in vivo by high-fat feeding in male and female C57BL/6J mice. METHODS: This study utilized small interfering RNA-mediated knockdown to assess the requirement of MPC1 for adipogenesis in the 3T3-L1 model system. Treatment with UK-5099, a potent pharmacological MPC inhibitor, was also used to assess the loss of MPC activity. Western blot analysis was performed on adipose tissue samples from mice on a low-fat diet or a high-fat diet (HFD) for 12 weeks. RESULTS: The loss of MPC expression via small interfering RNA-mediated knockdown or pharmacological inhibition did not affect adipogenesis of 3T3-L1 cells. In vivo studies indicated that expression of MPCs was significantly decreased in brown adipose tissue of male mice, but not female, on an HFD. CONCLUSIONS: Although MPCs are essential for pyruvate transport, MPCs are not required for adipogenesis in vitro, suggesting that other substrates can be used for energy production when the MPC complex is not functional. Also, a significant decrease in MPC1 and 2 expression occurred in brown fat, but not white fat, of male mice fed an HFD.


Assuntos
Adipogenia/fisiologia , Tecido Adiposo Marrom/metabolismo , Dieta Hiperlipídica/métodos , Transportadores de Ácidos Monocarboxílicos/metabolismo , Obesidade/fisiopatologia , Animais , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL
10.
FASEB J ; 34(2): 2792-2811, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31912559

RESUMO

While adipose tissue is required to maintain glucose metabolism, excessive calorie intake induces obesity via mechanisms including accelerated proliferation and differentiation of preadipocytes, leading to insulin resistance. Here, we investigated the role of myoferlin (MYOF), a ferlin family protein, in regulating glucose metabolism by mainly focusing on its unknown role in adipose tissue. Whereas young MYOF knockout (KO) mice on a normal diet showed aggravated glucose tolerance and insulin sensitivity, those on a high-fat diet (HFD) showed preserved glucose tolerance with an attenuated gain of body weight, reduced visceral fat deposits, and less severe fatty liver. The Adipose MYOF expression was reduced by aging but was restored by an HFD along with the retained expression of NFAT transcription factors. Loss-of-function of MYOF in preadipocytes suppressed proliferation and differentiation into mature adipocytes along with the decreased expression of genes involved in adipogenesis. The MYOF expression in preadipocytes was reduced with differentiation. Attenuated obesity in MYOF KO mice on an HFD was also accompanied with increased oxygen consumption by an unidentified mechanism and with reduced adipose inflammation due to less inflammatory macrophages. These insights suggest that the multifunctional roles of MYOF involve the regulation of preadipocyte function and affect glucose metabolism bidirectionally depending on consumed calories.


Assuntos
Adipócitos/metabolismo , Adipogenia/fisiologia , Adiposidade/fisiologia , Glucose/metabolismo , Proteínas de Membrana/metabolismo , Proteínas Musculares/metabolismo , Tecido Adiposo Marrom/metabolismo , Animais , Diferenciação Celular , Inflamação/metabolismo , Resistência à Insulina/fisiologia , Macrófagos/metabolismo , Masculino , Camundongos Endogâmicos C57BL
11.
FASEB J ; 34(1): 474-493, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31914704

RESUMO

The RhoA/ROCK-mediated actin cytoskeleton dynamics have been implicated in adipogenesis. The two ROCK isoforms, ROCK1 and ROCK2, are highly homologous. The contribution of ROCK2 to adipogenesis in vivo has not been elucidated. The present study aimed at the in vivo and in vitro roles of ROCK2 in the regulation of adipogenesis and the development of obesity. We performed molecular, histological, and metabolic analyses in ROCK2+/- and ROCK2+/KD mouse models, the latter harboring an allele with a kinase-dead (KD) mutation. Both ROCK2+/- and ROCK2+/KD mouse models showed a lean body mass phenotype during aging, associated with increased amounts of beige cells in subcutaneous white adipose tissue (sWAT) and increased thermogenic gene expression in all fat depots. ROCK2+/- mice on a high-fat diet showed increased energy expenditure accompanying by reduced obesity, and improved insulin sensitivity. In vitro differentiated ROCK2+/- stromal-vascular (SV) cells revealed increased beige adipogenesis associated with increased thermogenic gene expressions. Treatment with a selective ROCK2 inhibitor, KD025, to inhibit ROCK2 activity in differentiated SV cells reproduced the pro-beige phenotype of ROCK2+/- SV cells. In conclusion, ROCK2 activity-mediated actin cytoskeleton dynamics contribute to the inhibition of beige adipogenesis in WAT, and also promotes age-related and diet-induced fat mass gain and insulin resistance.


Assuntos
Adipogenia/fisiologia , Tecido Adiposo Marrom/fisiologia , Tecido Adiposo Branco/fisiologia , Resistência à Insulina , Obesidade/fisiopatologia , Termogênese/fisiologia , Quinases Associadas a rho/fisiologia , Animais , Diferenciação Celular , Dieta Hiperlipídica/efeitos adversos , Metabolismo Energético , Camundongos , Camundongos Knockout , Obesidade/etiologia , Transdução de Sinais
12.
Nat Commun ; 11(1): 213, 2020 01 10.
Artigo em Inglês | MEDLINE | ID: mdl-31924774

RESUMO

Human antigen R (HuR) is an essential regulator of RNA metabolism, but its function in metabolism remains unclear. This study identifies HuR as a major repressor during adipogenesis. Knockdown and overexpression of HuR in primary adipocyte culture enhances and inhibits adipogenesis in vitro, respectively. Fat-specific knockout of HuR significantly enhances adipogenic gene program in adipose tissues, accompanied by a systemic glucose intolerance and insulin resistance. HuR knockout also results in depot-specific phenotypes: it can repress myogenesis program in brown fat, enhance inflammation program in epidydimal white fat and induce browning program in inguinal white fat. Mechanistically, HuR may inhibit adipogenesis by recognizing and modulating the stability of hundreds of adipocyte transcripts including Insig1, a negative regulator during adipogenesis. Taken together, our work establishes HuR as an important posttranscriptional regulator of adipogenesis and provides insights into how RNA processing contributes to adipocyte development.


Assuntos
Adipogenia/genética , Adipogenia/fisiologia , Proteína Semelhante a ELAV 1/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Tecido Adiposo/patologia , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Branco/metabolismo , Animais , Proteína Semelhante a ELAV 1/genética , Regulação da Expressão Gênica , Técnicas de Inativação de Genes , Intolerância à Glucose/metabolismo , Humanos , Inflamação , Resistência à Insulina , Peptídeos e Proteínas de Sinalização Intracelular , Masculino , Proteínas de Membrana , Camundongos Endogâmicos C57BL , Camundongos Knockout
13.
Gene ; 733: 144265, 2020 Apr 05.
Artigo em Inglês | MEDLINE | ID: mdl-31805318

RESUMO

To explore its roles in adipogenesis, the levels of genomic 5mC methylation were examined across the adipocyte differentiation of 3 T3-L1 cells. This led to the identification of an up-regulating 5mC profile during the process. To further explore the regulation, gene expression assay was performed with a set of 5mC metabolic enzymes. Among them, TET2 was found to be the most regulated 5mC demethylase, in addition to a well-investigated 5mC methylase DNMT1. In the process, the expression of Tet2 increased for over 16-fold, suggesting its implications in the differentiation. Therefore, loss-of-function and gain-of-function assays were performed with Tet2. It was found that in relative to the differentiation of wild-type cells, knockdown of Tet2 expression led to greatly enhanced differentiation process, while over-expression of the gene resulted in repressed differentiation. Pathway study found that during the differentiation, TET2 demethylates Adrb3 promoter to up-regulate its expression. This led to enhanced lipolysis and decreased lipid production. To the upstream pathway, vitamin C treatment was found to enhance the activity of TETs, decrease 5mC levels and repress lipid production. Taken together, TET2 was characterized as an anti-adipogenic demethylase in adipocyte differentiation of 3 T3-L1 cells.


Assuntos
Metilação de DNA , Proteínas de Ligação a DNA/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Células 3T3-L1 , 5-Metilcitosina/metabolismo , Adipócitos/metabolismo , Adipogenia/genética , Adipogenia/fisiologia , Animais , Diferenciação Celular/genética , Proteínas de Ligação a DNA/genética , Epigênese Genética , Expressão Gênica/genética , Lipólise/genética , Camundongos , Regiões Promotoras Genéticas , Proteínas Proto-Oncogênicas/genética , Receptores Adrenérgicos beta 3/genética , Receptores Adrenérgicos beta 3/metabolismo , Fatores de Transcrição/genética
14.
Animal ; 14(2): 312-321, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31581971

RESUMO

In the livestock industry, subcutaneous and visceral fat pads are considered as wastes, while intramuscular fat or marbling fat is essential for improving flavor and palatability of meat. Thus, strategies for optimizing fat deposition are needed. Intramuscular adipocytes provide sites for lipid deposition and marbling formation. In the present article, we addressed the origin and markers of intramuscular adipocyte progenitors - fibro-adipogenic progenitors (FAPs), as well as the latest progresses in mechanisms regulating the proliferation and differentiation of intramuscular FAPs. Finally, by targeting intramuscular FAPs, possible nutritional manipulations to improve marbling fat deposition are discussed. Despite recent progresses, the properties and regulation of intramuscular FAPs in livestock remain poorly understood and deserve further investigation.


Assuntos
Adipogenia/fisiologia , Tecido Adiposo/metabolismo , Gado/fisiologia , Adipócitos/metabolismo , Animais , Diferenciação Celular , Proliferação de Células , Carne , Músculo Esquelético/fisiologia , Nutrientes , Células-Tronco/fisiologia
15.
J Cell Physiol ; 235(2): 1601-1614, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31304602

RESUMO

Obesity-associated conditions represent major global health and financial burdens and understanding processes regulating adipogenesis could lead to novel intervention strategies. This study shows that adhesion G-protein coupled receptor 56 (GPR56) gene transcripts are reduced in abdominal visceral white adipose tissue derived from obese Zucker rats versus lean controls. Immunostaining in 3T3-L1 preadipocytes reveals both mitotic cell restricted surface and low level general expression patterns of Gpr56. Gpr56 transcripts are differentially expressed in 3T3-L1 cells during adipogenesis. Transient knockdown (KD) of Gpr56 in 3T3-L1 cells dramatically inhibits differentiation through reducing the accumulation of both neutral cellular lipids (56%) and production of established adipogenesis Pparγ 2 (60-80%), C/ebpα (40-78%) mediator, and Ap2 (56-80%) marker proteins. Furthermore, genome editing of Gpr56 in 3T3-L1 cells created CW2.2.4 and RM4.2.5.5 clones (Gpr56 -/- cells) with compound heterozygous deletion frameshift mutations which abolish adipogenesis. Genome edited cells have sustained levels of the adipogenesis inhibitor ß-catenin, reduced proliferation, reduced adhesion, altered profiles, and or abundance of extracellular matrix component gene transcripts for fibronectin, types I, III, and IV collagens and loss of actin stress fibers. ß-catenin KD alone is insufficient to restore adipogenesis in Gpr56 -/- cells. Together these data show that Gpr56 is required for adipogenesis in 3T3-L1 cells. This report is the first demonstration that Gpr56 participates in regulation of the adipogenesis developmental program. Modulation of the levels of this protein and/or its biological activity may represent a novel target for development of therapeutic agents for the treatment of obesity.


Assuntos
Adipócitos/metabolismo , Adipogenia/fisiologia , Receptores Acoplados a Proteínas-G/metabolismo , Células 3T3-L1 , Animais , Técnicas de Silenciamento de Genes , Masculino , Camundongos , Obesidade/metabolismo , Ratos , Ratos Zucker
16.
J Cell Physiol ; 235(2): 1821-1837, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31432516

RESUMO

Extracellular matrix (ECM) has a marked influence on adipose tissue development. Adipose tissue formation is initiated with proliferation of preadipocytes and migration before undergoing further differentiation into mature adipocytes. Previous studies showed that collagen I (col I) provides a good substratum for 3T3-L1 preadipocytes to grow and migrate. However, it remains unclear whether and how col I regulates adipogenic differentiation of preadipocytes. This study reports that lipid accumulation, representing in vitro adipogenesis of the 3T3-L1 preadipocytes or the mouse primary adipocyte precursor cells derived from subcutaneous adipose tissue in the inguinal region is inhibited by the culture on col I, owing to downregulation of adipogenic factors. Previous study shows that col I enhances 3T3-L1 cell migration via stimulating the nuclear translocation of yes-associated protein (YAP). In this study, we report that downregulation of YAP is associated with in vitro adipogenesis of preadipocytes as well as with in vivo adipose tissue of high-fat diet fed mice. Increased expression of YAP in the cells cultured on col I-coated dishes is correlated with repression of adipogenic differentiation processes. The inactivation of YAP using YAP inhibitor, verteporfin, or YAP small-interfering RNA enhanced adipogenic differentiation and reversed the inhibitory effect of col I. Activation of YAP either by the transfection of YAP plasmid or the silence of large tumor suppressor 1 (LATS1), an inhibitory kinase of YAP, inhibited adipogenic differentiation. The results indicate that col I inhibits adipogenic differentiation via YAP activation in vitro.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adipócitos/metabolismo , Adipogenia/fisiologia , Proteínas de Ciclo Celular/metabolismo , Diferenciação Celular/fisiologia , Colágeno Tipo I/metabolismo , Células 3T3-L1 , Animais , Dieta Hiperlipídica , Camundongos , Células-Tronco/metabolismo
17.
J Cell Physiol ; 235(2): 891-899, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31240708

RESUMO

Adipose tissue is a complex and heterogenic tissue exhibiting high variability and appears to have multiple functions, especially in metabolic regulation. Change in carbohydrate source is reported to have a profound effect in the regular functioning of adipocytes. Here, we analyzed the role of two monosaccharides namely, glucose (5.2 and 25 nM), galactose (25 mM), and two disaccharides namely, lactose and sucrose (both at 25 mM) in the adipocyte differentiation process and its utilization by adipocytes as an energy source. The change in cell morphology, adipocyte-specific gene expression, and protein levels were analyzed at three different time points: 2, 6, and 48 hr. Oil Red O staining at Day 8 of differentiation showed that no other carbohydrates were able to increase lipid content as better as 25 mM glucose. Gene expression pattern was altered by the change in glucose concentration and sucrose was able to mimic the effect of glucose even though, the lipid synthesis was solely promoted by high glucose levels. Galactose and lactose did not show any effect in promoting adipocyte differentiation. The expression of PPAR γ was high in the presence of sucrose and galactose, possibly of adipogenic cocktail in enhancing the expression rather than the effect of carbohydrate. Acarbose, a potent glucosidase inhibitor was able to inhibit the lipid content in adipocytes grown with sucrose as a carbohydrate source and shows the possibility of its direct utilization. Lactate production by cells upon differentiation also proved the possible uptake of glucose after sucrose cleavage.


Assuntos
Adipócitos/metabolismo , Adipogenia/fisiologia , Galactose/metabolismo , Glucose/metabolismo , Lactose/metabolismo , Sacarose/metabolismo , Células 3T3-L1 , Acarbose/farmacologia , Tecido Adiposo/citologia , Tecido Adiposo/metabolismo , Animais , Linhagem Celular , Meios de Cultura/farmacologia , Expressão Gênica/efeitos dos fármacos , Glucosidases/antagonistas & inibidores , Camundongos , Obesidade/patologia
18.
Plast Reconstr Surg ; 145(1): 116-126, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31881612

RESUMO

BACKGROUND: Radiofibrosis of breast tissue compromises breast reconstruction by interfering with tissue viability and healing. Autologous fat transfer may reduce radiotherapy-related tissue injury, but graft survival is compromised by the fibrotic microenvironment. Elevated expression of receptor for hyaluronan-mediated motility (RHAMM; also known as hyaluronan-mediated motility receptor, or HMMR) in wounds decreases adipogenesis and increases fibrosis. The authors therefore developed RHAMM peptide mimetics to block RHAMM profibrotic signaling following radiation. They propose that this blocking peptide will decrease radiofibrosis and establish a microenvironment favoring adipose-derived stem cell survival using a rat mammary fat pad model. METHODS: Rat mammary fat pads underwent a one-time radiation dose of 26 Gy. Irradiated (n = 10) and nonirradiated (n = 10) fat pads received a single intramammary injection of a sham injection or peptide NPI-110. Skin changes were examined clinically. Mammary fat pad tissue was processed for fibrotic and adipogenic markers using quantitative polymerase chain reaction and immunohistochemical analysis. RESULTS: Clinical assessments and molecular analysis confirmed radiation-induced acute skin changes and radiation-induced fibrosis in rat mammary fat pads. Peptide treatment reduced fibrosis, as detected by polarized microscopy of picrosirius red staining, increased collagen ratio of 3:1, reduced expression of collagen-1 crosslinking enzymes lysyl-oxidase, transglutaminase 2, and transforming growth factor ß1 protein, and increased adiponectin, an antifibrotic adipokine. RHAMM was expressed in stromal cell subsets and was downregulated by the RHAMM peptide mimetic. CONCLUSION: Results from this study predict that blocking RHAMM function in stromal cell subsets can provide a postradiotherapy microenvironment more suitable for fat grafting and breast reconstruction.


Assuntos
Tecido Adiposo/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Fibrose/metabolismo , Receptores de Hialuronatos/metabolismo , Lesões Experimentais por Radiação/metabolismo , Adipogenia/efeitos dos fármacos , Adipogenia/fisiologia , Animais , Biomarcadores/metabolismo , Modelos Animais de Doenças , Fibrose/tratamento farmacológico , Peptídeos/farmacologia
19.
Genes (Basel) ; 11(1)2019 12 23.
Artigo em Inglês | MEDLINE | ID: mdl-31878002

RESUMO

Vulpinic acid, a naturally occurring methyl ester of pulvinic acid, has been reported to exert anti-fungal, anti-cancer, and anti-oxidative effects. However, its metabolic action has not been implicated yet. Here, we show that vulpinic acid derived from a mushroom, Pulveroboletus ravenelii controls the cell fate of mesenchymal stem cells and preadipocytes by inducing the acetylation of histone H3 and α-tubulin, respectively. The treatment of 10T1/2 mesenchymal stem cells with vulpinic acid increased the expression of Wnt6, Wnt10a, and Wnt10b, which led to osteogenesis inhibiting the adipogenic lineage commitment, through the upregulation of H3 acetylation. By contrast, treatment with vulpinic acid promoted the terminal differentiation of 3T3-L1 preadipocytes into mature adipocytes. In this process, the increase in acetylated tubulin was accompanied, while acetylated H3 was not altered. As excessive generation of adipocytes occurs, the accumulation of lipid drops was not concentrated, but dispersed into a number of adipocytes. Consistently, the expressions of lipolytic genes were upregulated and inflammatory factors were downregulated in adipocytes exposed to vulpinic acid during adipogenesis. These findings reveal the multiple actions of vulpinic acid in two stages of differentiation, promoting the osteogenesis of mesenchymal stem cells and decreasing hypertrophic adipocytes, which can provide experimental evidence for the novel metabolic advantages of vulpinic acid.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Furanos/farmacologia , Fenilacetatos/farmacologia , Células-Tronco/efeitos dos fármacos , Células 3T3-L1 , Adipócitos/metabolismo , Adipogenia/fisiologia , Animais , Furanos/metabolismo , Lipólise/fisiologia , Células-Tronco Mesenquimais , Camundongos , Osteogênese/fisiologia , Fenilacetatos/metabolismo , Transdução de Sinais , Células-Tronco/metabolismo
20.
Nat Cell Biol ; 21(12): 1490-1503, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31768046

RESUMO

Tendon injuries cause prolonged disability and never recover completely. Current mechanistic understanding of tendon regeneration is limited. Here, we use single-cell transcriptomics to identify a tubulin polymerization-promoting protein family member 3-expressing (Tppp3+) cell population as potential tendon stem cells. Through inducible lineage tracing, we demonstrate that these cells can generate new tenocytes and self-renew upon injury. A fraction of Tppp3+ cells expresses platelet-derived growth factor receptor alpha (Pdfgra). Ectopic platelet-derived growth factor-AA (PDGF-AA) protein induces new tenocyte production while inactivation of Pdgfra in Tppp3+ cells blocks tendon regeneration. These results support Tppp3+Pdgfra+ cells as tendon stem cells. Unexpectedly, Tppp3-Pdgfra+ fibro-adipogenic progenitors coexist in the tendon stem cell niche and give rise to fibrotic cells, revealing a clandestine origin of fibrotic scars in healing tendons. Our results explain why fibrosis occurs in injured tendons and present clinical challenges to enhance tendon regeneration without a concurrent increase in fibrosis by PDGF application.


Assuntos
Moléculas de Adesão Celular/metabolismo , Fibrose/metabolismo , Fator de Crescimento Derivado de Plaquetas/metabolismo , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Regeneração/fisiologia , Células-Tronco/metabolismo , Tendões/metabolismo , Adipogenia/fisiologia , Animais , Diferenciação Celular/fisiologia , Proliferação de Células/fisiologia , Células Cultivadas , Modelos Animais de Doenças , Fibrose/fisiopatologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Transdução de Sinais/fisiologia , Células-Tronco/fisiologia , Traumatismos dos Tendões/metabolismo , Traumatismos dos Tendões/fisiopatologia , Tendões/fisiopatologia , Tenócitos/metabolismo , Tenócitos/fisiologia
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