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1.
Molecules ; 26(16)2021 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-34443331

RESUMO

KD025, a ROCK2 isoform-specific inhibitor, has an anti-adipogenic activity which is not mediated by ROCK2 inhibition. To identify the target, we searched binding targets of KD025 by using the KINOMEscanTM screening platform, and we identified casein kinase 2 (CK2) as a novel target. KD025 showed comparable binding affinity to CK2α (Kd = 128 nM). By contrast, CK2 inhibitor CX-4945 and ROCK inhibitor fasudil did not show such cross-reactivity. In addition, KD025 effectively inhibited CK2 at a nanomolar concentration (IC50 = 50 nM). We examined if the inhibitory effect of KD025 on adipocyte differentiation is through the inhibition of CK2. Both CX-4945 and KD025 suppressed the generation of lipid droplets and the expression of proadipogenic genes Pparg and Cebpa in 3T3-L1 cells during adipocyte differentiation. Fasudil exerted no significant effect on the quantity of lipid droplets, but another ROCK inhibitor Y-27632 increased the expression of Pparg and Cebpa. Both CX-4945 and KD025 acted specifically in the middle stage (days 1-3) but were ineffective when treated at days 0-1 or the late stages, indicating that CX-4945 and KD025 may regulate the same target, CK2. The mRNA and protein levels of CK2α and CK2ß generally decreased in 3T3-L1 cells at day 2 but recovered thereafter. Other well-known CK2 inhibitors DMAT and quinalizarin inhibited effectively the differentiation of 3T3-L1 cells. Taken together, the results of this study confirmed that KD025 inhibits ROCK2 and CK2, and that the inhibitory effect on adipocyte differentiation is through the inhibition of CK2.


Assuntos
Adipócitos/citologia , Adipócitos/efeitos dos fármacos , Caseína Quinase II/antagonistas & inibidores , Diferenciação Celular/efeitos dos fármacos , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Quinases Associadas a rho/antagonistas & inibidores , Células 3T3-L1 , Animais , Regulação da Expressão Gênica/efeitos dos fármacos , Camundongos
2.
Molecules ; 26(15)2021 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-34361783

RESUMO

Amber-the fossilized resin of trees-is rich in terpenoids and rosin acids. The physiological effects, such as antipyretic, sedative, and anti-inflammatory, were used in traditional medicine. This study aims to clarify the physiological effects of amber extract on lipid metabolism in mouse 3T3-L1 cells. Mature adipocytes are used to evaluate the effect of amber extract on lipolysis by measuring the triglyceride content, glucose uptake, glycerol release, and lipolysis-related gene expression. Our results show that the amount of triacylglycerol, which is stored in lipid droplets in mature adipocytes, decreases following 96 h of treatment with different concentrations of amber extract. Amber extract treatment also decreases glucose uptake and increases the release of glycerol from the cells. Moreover, amber extract increases the expression of lipolysis-related genes encoding perilipin and hormone-sensitive lipase (HSL) and promotes the activity of HSL (by increasing HSL phosphorylation). Amber extract treatment also regulates the expression of other adipocytokines in mature adipocytes, such as adiponectin and leptin. Overall, our results indicate that amber extract increases the expression of lipolysis-related genes to induce lipolysis in 3T3-L1 cells, highlighting its potential for treating various obesity-related diseases.


Assuntos
Adipócitos/efeitos dos fármacos , Âmbar/farmacologia , Misturas Complexas/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Hipolipemiantes/farmacologia , Lipólise/efeitos dos fármacos , Células 3T3-L1 , Adipócitos/citologia , Adipócitos/metabolismo , Adiponectina/genética , Adiponectina/metabolismo , Âmbar/química , Animais , Diferenciação Celular , Misturas Complexas/química , Etanol/química , Glucose/metabolismo , Glicerol/metabolismo , Hipolipemiantes/química , Leptina/genética , Leptina/metabolismo , Gotículas Lipídicas/química , Gotículas Lipídicas/efeitos dos fármacos , Gotículas Lipídicas/metabolismo , Camundongos , Perilipina-1/genética , Perilipina-1/metabolismo , Fosforilação/efeitos dos fármacos , Esterol Esterase/genética , Esterol Esterase/metabolismo , Triglicerídeos/metabolismo
3.
Eur J Endocrinol ; 185(4): 539-552, 2021 Aug 27.
Artigo em Inglês | MEDLINE | ID: mdl-34342596

RESUMO

Objective: Sex steroid hormones like estrogens have a key role in the regulation of energy homeostasis and metabolism. In transwomen, gender-affirming hormone therapy like estradiol (in combination with antiandrogenic compounds) could affect metabolism as well. Given that the underlying pathophysiological mechanisms are not fully understood, this study assessed circulating estradiol-driven microRNAs (miRs) in transwomen and their regulation of genes involved in metabolism in mice. Methods: Following plasma miR-sequencing (seq) in a transwomen discovery (n = 20) and validation cohort (n = 30), we identified miR-224 and miR-452. Subsequent systemic silencing of these miRs in male C57Bl/6 J mice (n = 10) was followed by RNA-seq-based gene expression analysis of brown and white adipose tissue in conjunction with mechanistic studies in cultured adipocytes. Results: Estradiol in transwomen lowered plasma miR-224 and -452 carried in extracellular vesicles (EVs) while their systemic silencing in mice and cultured adipocytes increased lipogenesis (white adipose) but reduced glucose uptake and mitochondrial respiration (brown adipose). In white and brown adipose tissue, differentially expressed (miR target) genes are associated with lipogenesis (white adipose) and mitochondrial respiration and glucose uptake (brown adipose). Conclusion: This study identified an estradiol-drive post-transcriptional network that could potentially offer a mechanistic understanding of metabolism following gender-affirming estradiol therapy.


Assuntos
Micropartículas Derivadas de Células/genética , Estradiol/fisiologia , MicroRNAs/genética , Transexualidade , Adipócitos/efeitos dos fármacos , Adipócitos/fisiologia , Adipogenia/efeitos dos fármacos , Adipogenia/genética , Adulto , Animais , Micropartículas Derivadas de Células/efeitos dos fármacos , Micropartículas Derivadas de Células/metabolismo , Estudos de Coortes , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Metabolismo Energético/efeitos dos fármacos , Metabolismo Energético/genética , Estradiol/sangue , Estradiol/farmacologia , Vesículas Extracelulares/genética , Vesículas Extracelulares/metabolismo , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Homeostase/efeitos dos fármacos , Terapia de Reposição Hormonal , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/metabolismo , Pessoa de Meia-Idade , Interferência de RNA/efeitos dos fármacos , Pessoas Transgênero , Transexualidade/genética , Transexualidade/metabolismo , Adulto Jovem
4.
Nutrients ; 13(7)2021 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-34371822

RESUMO

Tetranectin (TN), a plasminogen-binding protein originally involved in fibrinolysis and bone formation, was later identified as a secreted adipokine from human and rat adipocytes and positively correlated with adipogenesis and lipid metabolism in adipocytes. To elucidate the nutritional regulation of adipogenic TN from diets containing different sources of fatty acids (saturated, n-6, n-3) in adipocytes, we cloned the coding region of porcine TN from a cDNA library and analyzed tissue expressions in weaned piglets fed with 2% soybean oil (SB, enriched in n-6 fatty acids), docosahexaenoic acid oil (DHA, an n-3 fatty acid) or beef tallow (BT, enriched in saturated and n-9 fatty acids) for 30 d. Compared with tissues in the BT- or SB-fed group, expression of TN was reduced in the adipose, liver and lung tissues from the DHA-fed group, accompanied with lowered plasma levels of triglycerides and cholesterols. This in vivo reduction was also confirmed in porcine primary differentiated adipocytes supplemented with DHA in vitro. Then, promoter analysis was performed. A 1956-bp putative porcine TN promoter was cloned and transcription binding sites for sterol regulatory-element binding protein (SREBP)-1c or forkhead box O proteins (FoxO) were predicted on the TN promoter. Mutating binding sites on porcine TN promoters showed that transcriptional suppression of TN by DHA on promoter activity was dependent on specific response elements for SREBP-1c or FoxO. The inhibited luciferase promoter activity by DHA on the TN promoter coincides with reduced gene expression of TN, SREBP-1c, and FoxO1 in human embryonic kidney HEK293T cells supplemented with DHA. To conclude, our current study demonstrated that the adipogenic TN was negatively regulated by nutritional modulation of DHA both in pigs in vivo and in humans/pigs in vitro. The transcriptional suppression by DHA on TN expression was partly through SREBP-1c or FoxO. Therefore, down-regulation of adipogenic tetranectin associated with fibrinolysis and adipogenesis may contribute to the beneficial effects of DHA on ameliorating obesity-induced metabolic syndromes such as atherosclerosis and adipose dysfunctions.


Assuntos
Adipogenia/efeitos dos fármacos , Ácidos Docosa-Hexaenoicos/farmacologia , Fatores de Transcrição Forkhead/metabolismo , Lectinas Tipo C/efeitos dos fármacos , Proteínas de Ligação a Elemento Regulador de Esterol/metabolismo , Adipócitos/efeitos dos fármacos , Animais , Fibrinólise/efeitos dos fármacos , Células HEK293 , Humanos , Fenômenos Fisiológicos da Nutrição/genética , Suínos
5.
Molecules ; 26(16)2021 Aug 19.
Artigo em Inglês | MEDLINE | ID: mdl-34443613

RESUMO

Adipogenesis is a complex process in which cell commitment and mitotic clonal expansion (MCE) are in-sequence crucial events leading to terminal adipocyte differentiation. The molecules able to block some key signals in this cascade can hamper adipogenesis becoming promising agents to counteract hyperplasia and hypertrophy of adipose tissue. Mono- and di-caffeoylquinic acid isomers are biologically active polyphenols, displaying in vitro and in vivo antioxidant, hepatoprotective, anti-diabetic and anti-obesity properties. Among these isomers, 3,5-dicaffeoylquinic acid (DCQA) has been reported to inhibit lipid accumulation in adipose cells more successfully than others. Thus, we investigated DCQA effects and molecular mechanisms on 3T3-L1 pre-adipocytes induced to differentiate with a hormonal cocktail (MDI). Oil Red O incorporation assessed that DCQA pre-treatment inhibited lipid accumulation in 3T3-L1 cells induced to differentiate for 10 days. At this time, an increased phosphorylation of both AMP-activated kinase and acetyl-CoA carboxylase, as well as a strong decrease in fatty acid synthase protein level, were registered by immunoblotting, thereby suggesting that DCQA treatment can reduce fatty acid anabolism in 3T3-L1 adipocytes. Furthermore, BrdU incorporation assay, performed 48 h after hormonal stimulation, revealed that DCQA treatment was also able to hinder the 3T3-L1 cell proliferation during the MCE, which is an essential step in the adipogenic process. Thus, we focused our attention on early signals triggered by the differentiation stimuli. In the first hours after hormonal cocktail administration, the activation of ERK1/2 and Akt kinases, or CREB and STAT3 transcription factors, was not affected by DCQA pre-treatment. Whereas 24 h after MDI induction, DCQA pre-treated cells showed increased level of the transcription factor Nrf2, that induced the expression of the antioxidant enzyme heme oxygenase 1 (HO-1). In control samples, the expression level of HO-1 was reduced 24 h after MDI induction in comparison with the higher amount of HO-1 protein found at 2 h. The HO-1 decrease was functional by allowing reactive oxygen species to boost and allowing cell proliferation induction at the beginning of MCE phase. Instead, in DCQA-treated cells the HO-1 expression was maintained at high levels for a further 24 h; in fact, its expression decreased only 48 h after MDI stimulation. The longer period in which HO-1 expression remained high led to a delay of the MCE phase, with a subsequent inhibition of both C/EBP-α expression and adipocyte terminal differentiation. In conclusion, DCQA counteracting an excessive adipose tissue expansion may become an attractive option in obesity treatment.


Assuntos
Adipócitos/citologia , Adipócitos/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Ácido Clorogênico/análogos & derivados , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Heme Oxigenase-1/metabolismo , Mitose/efeitos dos fármacos , Células 3T3-L1 , Adipócitos/metabolismo , Adipogenia/efeitos dos fármacos , Animais , Ácido Clorogênico/farmacologia , Camundongos
6.
Biomed Pharmacother ; 139: 111687, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-34243611

RESUMO

Obesity is one of the world's largest health problems, and 3-N-butylphthalide (NBP), a bioactive compound in celery, has been used in dieting and weight management programs. In this study, NBP prevented high-fat-diet-induced weight gain, reduced the food efficiency ratio, altered the blood biochemical profile, and reduced the obesity-related index. NBP reduced adiposity, white fat depots, liver weight, and hepatic steatosis in obese mice. NBP ameliorated the diabetic state by decreasing glucose levels and improving glucose and insulin tolerance. NBP increased uncoupling protein-1 expression in white adipose tissue and upregulated thermogenesis by enhancing mitochondrial respiration. NBP inhibited white adipocyte development by prohibiting lipid accumulation in human adipose-derived stem cells. NBP increased free fatty acid uptake and the oxygen consumption rate in beige adipocytes. Our results suggest that NBP could be used as functional natural supplement against obesity and its associated disorders.


Assuntos
Benzofuranos/farmacologia , Dieta Hiperlipídica/efeitos adversos , Metabolismo dos Lipídeos/efeitos dos fármacos , Lipídeos/fisiologia , Obesidade/metabolismo , Substâncias Protetoras/farmacologia , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Tecido Adiposo Branco/efeitos dos fármacos , Tecido Adiposo Branco/metabolismo , Adiposidade/efeitos dos fármacos , Animais , Glicemia/efeitos dos fármacos , Células Cultivadas , Fígado Gorduroso/metabolismo , Humanos , Fígado/efeitos dos fármacos , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , Termogênese/efeitos dos fármacos
7.
Aging (Albany NY) ; 13(13): 17489-17498, 2021 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-34232916

RESUMO

BACKGROUND AND PURPOSE: Obesity is becoming a major global health issue and is mainly induced by the accumulation of adipose tissues mediated by adipogenesis, which is reported to be regulated by peroxisome proliferator-activated receptor γ (PPARγ) and CCAAT enhancer-binding protein α (C/EBPα). Trichostatin A (TSA) is a novel histone deacetylase inhibitor (HDACI) that was recently reported to exert multiple pharmacological functions. The present study will investigate the inhibitory effect of TSA on adipogenesis, as well as the underlying mechanism. METHODS: The adipogenesis of 3T3-L1 cells was induced by stimulation with a differentiation cocktail (DMI) medium for 8 days. MTT assay was used to measure the cell viability and Oil Red O staining was used to evaluate the adipogenesis of 3T3-L1 cells. The total level of triglyceride and released glycerol were detected to evaluate the lipolysis during 3T3-L1 adipogenesis. The expression levels of Leptin, fatty acid-binding protein 4 (FABP4), and sterol regulatory element-binding protein (SREBP1C) were determined by qRT-PCR. qRT-PCR assay was utilized to detect the expression levels of PPARγ and C/EBPα in 3T3-L1 cells. A high-fat diet (HFD) was used to construct an obese mice model, followed by the treatment with TSA. HE staining was conducted to evaluate the pathological state of adipose tissues. Body weights and the weights of adipose tissues were recorded to evaluate the anti-obesity property of TSA. RESULTS: Firstly, the promoted lipid accumulation induced by DMI incubation was significantly reversed by the treatment with TSA in a dose-dependent manner. The elevated expression levels of Leptin, FABP4, SREBP1C, PPARγ, and C/EBPα induced by the stimulation with DMI incubation were dramatically inhibited by the introduction of TSA, accompanied by the upregulation of phosphorylated AMP-activated protein kinase (p-AMPK). Secondly, the inhibitory effect of TSA against the expression level of PPARγ and lipid accumulation was greatly abolished by an AMPK inhibitor. Lastly, the increased body weights and visceral adipocyte tissue weight, as well as the enlarged size of adipocytes induced by HFD were pronouncedly reversed by the administration of TSA. CONCLUSION: TSA inhibited adipogenesis in 3T3-L1 preadipocytes by activating the AMPK pathway.


Assuntos
Adipócitos/efeitos dos fármacos , Adipogenia/efeitos dos fármacos , Fármacos Antiobesidade/farmacologia , Inibidores de Histona Desacetilases/farmacologia , Ácidos Hidroxâmicos/farmacologia , Células 3T3-L1 , Tecido Adiposo/patologia , Animais , Peso Corporal/efeitos dos fármacos , Dieta Hiperlipídica , Proteínas de Ligação a Ácido Graxo/genética , Glicerol/metabolismo , Leptina/metabolismo , Lipólise/efeitos dos fármacos , Masculino , Camundongos , Camundongos Obesos , Obesidade/genética , Triglicerídeos/metabolismo
8.
Int J Mol Sci ; 22(13)2021 Jun 22.
Artigo em Inglês | MEDLINE | ID: mdl-34206506

RESUMO

Adipose-derived stem cells (ADSCs) came out from the regenerative medicine landscape for their ability to differentiate into several phenotypes, contributing to tissue regeneration both in vitro and in vivo. Dysregulation in stem cell recruitment and differentiation during adipogenesis is linked to a chronic low-grade inflammation and macrophage infiltration inside the adipose tissue, insulin resistance, cardiovascular disease and obesity. In the present paper we aimed to evaluate the role of metformin and vitamin D, alone or in combination, in modulating inflammation and autophagy in ADSCs during adipogenic commitment. ADSCs were cultured for 21 days in the presence of a specific adipogenic differentiation medium, together with metformin, or vitamin D, or both. We then analyzed the expression of FoxO1 and Heat Shock Proteins (HSP) and the secretion of proinflammatory cytokines IL-6 and TNF-α by ELISA. Autophagy was also assessed by specific Western blot analysis of ATG12, LC3B I, and LC3B II expression. Our results showed the ability of the conditioned media to modulate adipogenic differentiation, finely tuning the inflammatory response and autophagy. We observed a modulation in HSP mRNA levels, and a significant downregulation in cytokine secretion. Taken together, our findings suggest the possible application of these molecules in clinical practice to counteract uncontrolled lipogenesis and prevent obesity and obesity-related metabolic disorders.


Assuntos
Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Metformina/farmacologia , Vitamina D/farmacologia , Adipócitos/citologia , Diferenciação Celular/efeitos dos fármacos , Citocinas/metabolismo , Humanos , Mediadores da Inflamação/metabolismo , Células-Tronco Mesenquimais/citologia
9.
Int J Mol Sci ; 22(13)2021 Jun 23.
Artigo em Inglês | MEDLINE | ID: mdl-34201755

RESUMO

Diabetes mellitus is a debilitating disease, plaguing a significant number of people around the globe. Attempts to develop new drugs on well-defined atoxic metalloforms, which are capable of influencing fundamental cellular processes overcoming insulin resistance, has triggered an upsurge in molecular research linked to zinc metallodrugs. To that end, meticulous efforts were launched toward the design and synthesis of materials with insulin mimetic potential. Henceforth, trigonelline and N-(2-hydroxyethyl)-iminodiacetic acid (HEIDAH2) were selected as organic substrates seeking binding to zinc (Zn(II)), with new crystalline compounds characterized by elemental analysis, FT-IR, X-rays, thermogravimetry (TGA), luminescence, NMR, and ESI-MS spectrometry. Physicochemical characterization was followed by in vitro biochemical experiments, in which three out of the five zinc compounds emerged as atoxic, exhibiting bio-activity profiles reflecting enhanced adipogenic potential. Concurrently, well-defined qualitative-quantitative experiments provided links to genetic loci responsible for the observed effects, thereby unraveling their key involvement in signaling pathways in adipocyte tissue and insulin mimetic behavior. The collective results (a) signify the quintessential role of molecular studies in unearthing unknown facets of pathophysiological events in diabetes mellitus II, (b) reflect the close associations of properly configured molecular zincoforms to well-defined biological profiles, and (c) set the stage for further physicochemical-based development of efficient zinc antidiabetic metallodrugs.


Assuntos
Adipócitos/efeitos dos fármacos , Adipogenia , Insulina/farmacologia , Compostos Organometálicos/farmacocinética , Zinco/química , Células 3T3-L1 , Animais , Hipoglicemiantes/farmacologia , Camundongos , Transdução de Sinais
10.
FEBS Lett ; 595(16): 2085-2098, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-34197627

RESUMO

Brown and beige adipocytes dissipate energy by uncoupling protein 1 (UCP1)-dependent and UCP1-independent thermogenesis, which may be utilized to develop treatments against obesity. We have found that mRNA and protein expression of the alanine/serine/cysteine transporter-1 (ASC-1) was induced during adipocyte differentiation of human brown-prone deep neck and beige-competent subcutaneous neck progenitors, and SGBS preadipocytes. cAMP stimulation of differentiated adipocytes led to elevated uptake of serine, cysteine, and glycine, in parallel with increased oxygen consumption, augmented UCP1-dependent proton leak, increased creatine-driven substrate cycle-coupled respiration, and upregulation of thermogenesis marker genes and several respiratory complex subunits; these outcomes were impeded in the presence of the specific ASC-1 inhibitor, BMS-466442. Our data suggest that ASC-1-dependent consumption of serine, cysteine, and glycine is required for efficient thermogenic stimulation of human adipocytes.


Assuntos
Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Adrenérgicos/farmacologia , Sistema y+ de Transporte de Aminoácidos/metabolismo , Aminoácidos/metabolismo , Termogênese , Transporte Biológico/efeitos dos fármacos , Humanos , Termogênese/efeitos dos fármacos
11.
Molecules ; 26(14)2021 Jul 12.
Artigo em Inglês | MEDLINE | ID: mdl-34299508

RESUMO

Although the hypoglycemic potential of brewer's yeast extract has been reported, there is limited information pertaining to the hypoglycemic ingredients of Saccharomyces pastorianus extract and their mechanisms of action available. This study aimed to investigate the in vivo and in vitro hypoglycemic effect of S. pastorianus extract and to elucidate its molecular mechanisms. S. pastorianus extract was mainly composed of proteins followed by carbohydrates. In diabetic rats, oral administration of S. pastorianus extract significantly reduced the levels of plasma glucose and enhanced the activity of hepatic glucose-6-phosphatase dehydrogenase. Treatment with S. pastorianus extract increased the localization of type 4 glucose transporter (GLUT4), PTP, and insulin receptor at 3T3-L1 cell membranes and raised the levels of P38 MAPK, PI3K, and AKT in the cytosol. In agreement with these results, pretreatment of 3T3-L1 cells with inhibitors of PTP, PI3K, Akt/PKB, and p38 MAPK inhibited glucose uptake induced by application of S. pastorianus extract. Most importantly, a 54 kDa protein with hypoglycemic activity was identified and suggested as the major ingredient contributing to the hypoglycemic activity of S. pastorianus extract. In summary, these results clearly confirm the hypoglycemic activity of S. pastorianus extract and provide critical insights into the underlying molecular mechanisms.


Assuntos
Hipoglicemiantes/farmacologia , Saccharomyces/metabolismo , Células 3T3-L1 , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Animais , Transporte Biológico/efeitos dos fármacos , Metabolismo dos Carboidratos/efeitos dos fármacos , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/metabolismo , Glucose/metabolismo , Transportador de Glucose Tipo 4/metabolismo , Insulina/metabolismo , Resistência à Insulina/fisiologia , Masculino , Camundongos , Proteínas de Transporte de Monossacarídeos/metabolismo , Ratos , Ratos Sprague-Dawley , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
12.
Molecules ; 26(14)2021 Jul 18.
Artigo em Inglês | MEDLINE | ID: mdl-34299620

RESUMO

Type 2 diabetes mellitus (T2DM) is linked to insulin resistance and a loss of insulin sensitivity, leading to millions of deaths worldwide each year. T2DM is caused by reduced uptake of glucose facilitated by glucose transporter 4 (GLUT4) in muscle and adipose tissue due to decreased intracellular translocation of GLUT4-containing vesicles to the plasma membrane. To treat T2DM, novel medications are required. Through a fluorescence microscopy-based high-content screen, we tested more than 600 plant extracts for their potential to induce GLUT4 translocation in the absence of insulin. The primary screen in CHO-K1 cells resulted in 30 positive hits, which were further investigated in HeLa and 3T3-L1 cells. In addition, full plasma membrane insertion was examined by immunostaining of the first extracellular loop of GLUT4. The application of appropriate inhibitors identified PI3 kinase as the most important signal transduction target relevant for GLUT4 translocation. Finally, from the most effective hits in vitro, four extracts effectively reduced blood glucose levels in chicken embryos (in ovo), indicating their applicability as antidiabetic pharmaceuticals or nutraceuticals.


Assuntos
Glicemia/efeitos dos fármacos , Glucose/metabolismo , Hipoglicemiantes/farmacologia , Insulina/farmacologia , Extratos Vegetais/farmacologia , Células 3T3-L1 , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Animais , Células CHO , Linhagem Celular , Linhagem Celular Tumoral , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Cricetulus , Diabetes Mellitus Tipo 2 , Transportador de Glucose Tipo 4/metabolismo , Células HeLa , Humanos , Resistência à Insulina/fisiologia , Camundongos , Transporte Proteico/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos
13.
Int J Mol Sci ; 22(11)2021 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-34198827

RESUMO

The objective of this study was to investigate molecular mechanisms underlying the ability of carnosic acid to attenuate an early increase in reactive oxygen species (ROS) levels during MDI-induced adipocyte differentiation. The levels of superoxide anion and ROS were determined using dihydroethidium (DHE) and 2'-7'-dichlorofluorescin diacetate (DCFH-DA), respectively. Both superoxide anion and ROS levels peaked on the second day of differentiation. They were suppressed by carnosic acid. Carnosic acid attenuates the translation of NADPH (nicotinamide adenine dinucleotide phosphate) oxidase 4 (Nox4), p47phox, and p22phox, and the phosphorylation of nuclear factor-kappa B (NF-κB) and NF-κB inhibitor (IkBa). The translocation of NF-κB into the nucleus was also decreased by carnosic acid. In addition, carnosic acid increased the translation of heme oxygenase-1 (HO-1), γ-glutamylcysteine synthetase (γ-GCSc), and glutathione S-transferase (GST) and both the translation and nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2). Taken together, these results indicate that carnosic acid could down-regulate ROS level in an early stage of MPI-induced adipocyte differentiation by attenuating ROS generation through suppression of NF-κB-mediated translation of Nox4 enzyme and increasing ROS neutralization through induction of Nrf2-mediated translation of phase II antioxidant enzymes such as HO-1, γ-GCS, and GST, leading to its anti-adipogenetic effect.


Assuntos
ATPases Associadas a Diversas Atividades Celulares/genética , Abietanos/farmacologia , DNA Helicases/genética , Heme Oxigenase-1/genética , Proteínas de Membrana/genética , NADPH Oxidase 4/genética , Inibidor de NF-kappaB alfa/genética , Células 3T3-L1 , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Animais , Antioxidantes/farmacologia , Diferenciação Celular/efeitos dos fármacos , Grupo dos Citocromos b/genética , Etídio/análogos & derivados , Etídio/farmacologia , Fluoresceínas/farmacologia , Glutationa Transferase/genética , Camundongos , NADPH Oxidases/genética , Biossíntese de Proteínas/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo
14.
Molecules ; 26(13)2021 Jun 23.
Artigo em Inglês | MEDLINE | ID: mdl-34201708

RESUMO

Caffeine is a plant alkaloid present in food and beverages consumed worldwide. It has high lipid solubility with recognized actions in the central nervous system and in peripheral tissues, notably the adipose depots. However, the literature is scant regarding caffeine's influence on adipocyte functions other than lipolysis, such as glucose incorporation into lipids (lipogenesis) and amine oxidation. The objective of this study was to explore the direct effects of caffeine and of isobutylmethylxanthine (IBMX) on these adipocyte functions. Glucose transport into fat cells freshly isolated from mice, rats, or humans was monitored by determining [3H]-2-deoxyglucose (2-DG) uptake, while the incorporation of radiolabeled glucose into cell lipids was used as an index of lipogenic activity. Oxidation of benzylamine by primary amine oxidase (PrAO) was inhibited by increasing doses of caffeine in human adipose tissue preparations with an inhibition constant (Ki) in the millimolar range. Caffeine inhibited basal and insulin-stimulated glucose transport as well as lipogenesis in rodent adipose cells. The antilipogenic action of caffeine was also observed in adipocytes from mice genetically invalidated for PrAO activity, indicating that PrAO activity was not required for lipogenesis inhibition. These caffeine inhibitory properties were extended to human adipocytes: relative to basal 2-DG uptake, set at 1.0 ± 0.2 for 6 individuals, 0.1 mM caffeine tended to reduce uptake to 0.83 ± 0.08. Insulin increased uptake by 3.86 ± 1.11 fold when tested alone at 100 nM, and by 3.21 ± 0.80 when combined with caffeine. Our results reinforce the recommendation of caffeine's potential in the treatment or prevention of obesity complications.


Assuntos
Adipócitos/efeitos dos fármacos , Aminas Biogênicas/metabolismo , Cafeína/farmacologia , Glucose/metabolismo , Lipogênese/efeitos dos fármacos , Monoaminoxidase/metabolismo , Tecido Adiposo/efeitos dos fármacos , Tecido Adiposo/metabolismo , Animais , Benzilaminas/metabolismo , Transporte Biológico/efeitos dos fármacos , Desoxiglucose/metabolismo , Humanos , Insulina/metabolismo , Lipólise/efeitos dos fármacos , Camundongos , Ratos , Xantinas/farmacologia
15.
FASEB J ; 35(7): e21728, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34110658

RESUMO

Proliferation and differentiation of preadipocytes, and other cell types, is accompanied by an increase in glucose uptake. Previous work showed that a pulse of high glucose was required during the first 3 days of differentiation in vitro, but was not required after that. The specific glucose metabolism pathways required for adipocyte differentiation are unknown. Herein, we used 3T3-L1 adipocytes as a model system to study glucose metabolism and expansion of the adipocyte metabolome during the first 3 days of differentiation. Our primary outcome measures were GLUT4 and adiponectin, key proteins associated with healthy adipocytes. Using complete media with 0 or 5 mM glucose, we distinguished between developmental features that were dependent on the differentiation cocktail of dexamethasone, insulin, and isobutylmethylxanthine alone or the cocktail plus glucose. Cocktail alone was sufficient to activate the capacity for 2-deoxglucose uptake and glycolysis, but was unable to support the expression of GLUT4 and adiponectin in mature adipocytes. In contrast, 5 mM glucose in the media promoted a transient increase in glucose uptake and glycolysis as well as a significant expansion of the adipocyte metabolome and proteome. Using genetic and pharmacologic approaches, we found that the positive effects of 5 mM glucose on adipocyte differentiation were specifically due to increased expression of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 (PFKFB3), a key regulator of glycolysis and the ancillary glucose metabolic pathways. Our data reveal a critical role for PFKFB3 activity in regulating the cellular metabolic remodeling required for adipocyte differentiation and maturation.


Assuntos
Adipócitos/metabolismo , Glucose/metabolismo , Fosfofrutoquinase-2/metabolismo , Células 3T3-L1 , Adipócitos/efeitos dos fármacos , Adiponectina/metabolismo , Animais , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/fisiologia , Linhagem Celular , Dexametasona/farmacologia , Transportador de Glucose Tipo 4/metabolismo , Glicólise/efeitos dos fármacos , Glicólise/fisiologia , Insulina/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Xantinas/farmacologia
16.
Int J Mol Sci ; 22(10)2021 May 19.
Artigo em Inglês | MEDLINE | ID: mdl-34069744

RESUMO

Bisphenol A (BPA) is an endocrine-disrupting chemical used in the production of plastics, and is linked to developmental, reproductive, and metabolic disorders including obesity. Manufacturers have begun using 'BPA-free' alternatives instead of BPA in many consumer products. However, these alternatives have had much less testing and oversight, yet they are already being mass-produced and used across industries from plastics to food-contact coatings. Here, we used human female adipose-derived stem cells (hASCs), a type of adult mesenchymal stem cell, to compare the effects of BPA and BPA alternatives on adipogenesis or fat cell development in vitro. We focused on two commonly used BPA replacements, bisphenol AF (BPAF) and tetramethyl bisphenol F (TMBPF; monomer of the new valPure V70 food-contact coating). Human ASCs were differentiated into adipocytes using chemically defined media in the presence of control differentiation media with and without 17ß-estradiol (E2; 10 µM), or with increasing doses of BPA (0, 0.1 and 1 µM), BPAF (0, 0.1, 1 and 10 nM), or TMBPF (0, 0.01 and 0.1 µM). After differentiation, the cells were stained and imaged to visualize and quantify the accumulation of lipid vacuoles and number of developing fat cells. Treated cells were also examined for cell viability and apoptosis (programmed cell death) using the respective cellular assays. Similar to E2, BPA at 0.1 µM and BPAF at 0.1 nM, significantly increased adipogenesis and lipid production by 20% compared to control differentiated cells (based on total lipid vacuole number to cell number ratios), whereas higher levels of BPA and BPAF significantly decreased adipogenesis (p < 0.005). All tested doses of TMBPF significantly reduced adipogenesis and lipid production by 30-40%, likely at least partially through toxic effects on stem cells, as viable cell numbers decreased and apoptosis levels increased throughout differentiation. These findings indicate that low, environmentally-relevant doses of BPA, BPAF, and TMBPF have significant effects on fat cell development and lipid accumulation, with TMBPF having non-estrogenic, anti-adipogenic effects. These and other recent results may provide a potential cellular mechanism between exposure to bisphenols and human obesity, and underscore the likely impact of these chemicals on fat development in vivo.


Assuntos
Adipócitos/metabolismo , Adipogenia/efeitos dos fármacos , Células-Tronco Mesenquimais/efeitos dos fármacos , Adipócitos/efeitos dos fármacos , Adipogenia/fisiologia , Tecido Adiposo/efeitos dos fármacos , Células-Tronco Adultas/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Compostos Benzidrílicos/efeitos adversos , Diferenciação Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Disruptores Endócrinos/toxicidade , Estradiol/farmacologia , Feminino , Humanos , Obesidade/metabolismo , Fenóis/efeitos adversos , Células-Tronco/efeitos dos fármacos
17.
Life Sci ; 282: 119668, 2021 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-34087283

RESUMO

AIMS: Berteroin (5-methylthiopentyl isothiocyanate) is a naturally occurring sulforaphane analog containing a non-oxidized sulfur atom in cruciferous vegetables. The objectives of the present study were to determine the effects of berteroin on lipid metabolism in hepatocytes and adipocytes and to elucidate the mechanisms involved. MAIN METHODS: The effect of berteroin on lipid metabolism were evaluated in liver X receptor α agonist-stimulated HepG2 cells and adipocyte differentiation-induced 3T3-L1 cells using MTT assay, western blot, real time polymerase chain reaction, oil red O staining, and triglyceride assay. KEY FINDINGS: T0901317 treatment increased the expression of sterol regulatory element binding protein (SREBP)-1c, a major transcription factor that mediates lipogenesis, and berteroin pretreatment significantly inhibited the expressions of T0901317-induced SREBP-1c and lipogenic genes. Especially, berteroin had a greater inhibitory effect on T0901317-induced SREBP-1c activation than sulforaphane, AICAR, or metformin. Berteroin also markedly suppressed lipid droplet formations and triglyceride accumulations caused by both T0901317 stimulation in HepG2 hepatocytes and differentiation induction in 3T3-L1 preadipocytes. However, berteroin significantly increased the expression of mitochondrial fatty acid oxidation-related genes (carnitine palmitoyltransferase 1 (CPT-1) and peroxisome proliferator-activated receptor gamma coactivator-1α) and the phosphorylation of adenosine monophosphate-activated protein kinase (AMPK) in HepG2 cells. Interestingly, effects of berteroin on the expressions of SREBP-1c protein and CPT-1 mRNA were remarkably prevented by compound C (an AMPK inhibitor). SIGNIFICANCE: Our results suggest berteroin-inhibited hepatic lipid accumulation and adipocyte differentiation might be mediated by AMPK activation and that berteroin might be useful for the prevention, amelioration, and treatment of metabolic diseases, including hepatic steatosis.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Adipogenia/efeitos dos fármacos , Isotiocianatos/farmacologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Fígado/efeitos dos fármacos , Células 3T3-L1 , Adipócitos/citologia , Adipócitos/efeitos dos fármacos , Adipócitos/metabolismo , Animais , Células Hep G2 , Humanos , Fígado/metabolismo , Camundongos
18.
Int J Mol Sci ; 22(11)2021 May 24.
Artigo em Inglês | MEDLINE | ID: mdl-34074061

RESUMO

BACKGROUND: Erythritol, a sugar alcohol, is widely used as a substitute for sugar in diets for patients with diabetes or obesity. METHODS: In this study, we aimed to investigate the effects of erythritol on metabolic disorders induced by a high-fat diet in C57BL/6J mice, while focusing on changes in innate immunity. RESULTS: Mice that were fed a high-fat diet and administered water containing 5% erythritol (Ery group) had markedly lower body weight, improved glucose tolerance, and markedly higher energy expenditure than the control mice (Ctrl group) (n = 6). Furthermore, compared with the Ctrl group, the Ery group had lesser fat deposition in the liver, smaller adipocytes, and significantly better inflammatory findings in the small intestine. The concentrations of short-chain fatty acids (SCFAs), such as acetic acid, propanoic acid, and butanoic acid, in the serum, feces, and white adipose tissue of the Ery group were markedly higher than those in the Ctrl group. In flow cytometry experiments, group 3 innate lymphoid cell (ILC3) counts in the lamina propria of the small intestine and ILC2 counts in the white adipose tissue of the Ery group were markedly higher than those in the Ctrl group. Quantitative real-time reverse transcription polymerase chain reaction analyses showed that the Il-22 expression in the small intestine of the Ery group was markedly higher than that in the Ctrl group. CONCLUSIONS: Erythritol markedly decreased metabolic disorders such as diet-induced obesity, glucose intolerance, dyslipidemia, and fat accumulation in the mouse liver by increasing SCFAs and modulating innate immunity.


Assuntos
Dieta Hiperlipídica/efeitos adversos , Eritritol/farmacologia , Intolerância à Glucose/dietoterapia , Imunidade Inata/efeitos dos fármacos , Intestino Delgado/efeitos dos fármacos , Obesidade/tratamento farmacológico , Adipócitos/citologia , Adipócitos/efeitos dos fármacos , Tecido Adiposo/metabolismo , Tecido Adiposo Branco/metabolismo , Animais , Glicemia/efeitos dos fármacos , Glicemia/metabolismo , Metabolismo Energético/efeitos dos fármacos , Eritritol/administração & dosagem , Ácidos Graxos Voláteis/sangue , Ácidos Graxos Voláteis/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Intolerância à Glucose/metabolismo , Imunidade Inata/genética , Inflamação/dietoterapia , Inflamação/genética , Inflamação/metabolismo , Interleucinas/genética , Interleucinas/metabolismo , Intestino Delgado/imunologia , Intestino Delgado/metabolismo , Fígado/efeitos dos fármacos , Fígado/enzimologia , Fígado/metabolismo , Fígado/patologia , Linfócitos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Membrana Mucosa/efeitos dos fármacos , Membrana Mucosa/metabolismo , Obesidade/genética , Obesidade/metabolismo
19.
Int J Mol Sci ; 22(9)2021 May 05.
Artigo em Inglês | MEDLINE | ID: mdl-34063048

RESUMO

Pazopanib is a multikinase inhibitor with anti-tumor activity. As of now, the anti-obesity effect and mode of action of pazopanib are unknown. In this study, we investigated the effects of pazopanib on lipid accumulation, lipolysis, and expression of inflammatory cyclooxygenase (COX)-2 in differentiating and differentiated 3T3-L1 cells, a murine preadipocyte. Of note, pazopanib at 10 µM markedly decreased lipid accumulation and triglyceride (TG) content during 3T3-L1 preadipocyte differentiation with no cytotoxicity. Furthermore, pazopanib inhibited not only expression of CCAAT/enhancer-binding protein-α (C/EBP-α), peroxisome proliferator-activated receptor-γ (PPAR-γ), and perilipin A but also phosphorylation of signal transducer and activator of transcription (STAT)-3 during 3T3-L1 preadipocyte differentiation. In addition, pazopanib treatment increased phosphorylation of cAMP-activated protein kinase (AMPK) and its downstream effector ACC during 3T3-L1 preadipocyte differentiation. However, in differentiated 3T3-L1 adipocytes, pazopanib treatment did not stimulate glycerol release and hormone-sensitive lipase (HSL) phosphorylation, hallmarks of lipolysis. Moreover, pazopanib could inhibit tumor necrosis factor (TNF)-α-induced expression of COX-2 in both 3T3-L1 preadipocytes and differentiated cells. In summary, this is the first report that pazopanib has strong anti-adipogenic and anti-inflammatory effects in 3T3-L1 cells, which are mediated through regulation of the expression and phosphorylation of C/EBP-α, PPAR-γ, STAT-3, ACC, perilipin A, AMPK, and COX-2.


Assuntos
Adipócitos/citologia , Adipócitos/metabolismo , Diferenciação Celular/efeitos dos fármacos , Ciclo-Oxigenase 2/metabolismo , Indazóis/farmacologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Pirimidinas/farmacologia , Sulfonamidas/farmacologia , Células 3T3-L1 , Adenilato Quinase/metabolismo , Adipócitos/efeitos dos fármacos , Adipogenia/efeitos dos fármacos , Animais , Proteína alfa Estimuladora de Ligação a CCAAT/metabolismo , Morte Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Ácido Graxo Sintases/metabolismo , Glicerol/metabolismo , Leptina/metabolismo , Camundongos , PPAR gama/metabolismo , Perilipina-1/metabolismo , Fosforilação/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Resistina/metabolismo , Fator de Transcrição STAT3/metabolismo , Esterol Esterase/metabolismo , Triglicerídeos/metabolismo , Fator de Necrose Tumoral alfa/farmacologia
20.
Int J Mol Sci ; 22(11)2021 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-34072724

RESUMO

At the onset of lactation, dairy cows suffer from insulin resistance, insulin deficiency or both, similar to human diabetes, resulting in lipolysis, ketosis and fatty liver. This work explored the combined effects of different levels of magnesium (0.1, 0.3, 1 and 3 mM) and insulin (25, 250 and 25,000 pM) on metabolic pathways and the expression of magnesium-responsive genes in a bovine adipocyte model. Magnesium starvation (0.1 mM) and low insulin (25 pM) independently decreased or tended to decrease the accumulation of non-polar lipids and uptake of the glucose analog 6-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-6-deoxyglucose (6-NBDG). Activity of glycerol 3-phosphate dehydrogenase (GPDH) was highest at 25 pM insulin and 3 mM magnesium. Expression of SLC41A1 and SLC41A3 was reduced at 0.1 mM magnesium either across insulin concentrations (SLC41A1) or at 250 pM insulin (SLC41A3). MAGT1 expression was reduced at 3 mM magnesium. NIPA1 expression was reduced at 3 mM and 0.1 mM magnesium at 25 and 250 pM insulin, respectively. Expression of SLC41A2, CNNM2, TRPM6 and TRPM7 was not affected. We conclude that magnesium promotes lipogenesis in adipocytes and inversely regulates the transcription of genes that increase vs. decrease cytosolic magnesium concentration. The induction of GAPDH activity by surplus magnesium at low insulin concentration can counteract excessive lipomobilization.


Assuntos
Adipócitos/metabolismo , Metabolismo Energético , Regulação da Expressão Gênica , Homeostase , Insulina/metabolismo , Magnésio/metabolismo , Adipócitos/efeitos dos fármacos , Animais , Bovinos , Células Cultivadas , Metabolismo Energético/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Glucose/metabolismo , Insulina/farmacologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Magnésio/farmacologia , Álcool Oxidorredutases Dependentes de NAD(+) e NADP(+)/metabolismo
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