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1.
Nat Commun ; 10(1): 2901, 2019 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-31263101

RESUMO

Dysregulation of histone modifications promotes carcinogenesis by altering transcription. Breast cancers frequently overexpress the histone methyltransferase EZH2, the catalytic subunit of Polycomb Repressor Complex 2 (PRC2). However, the role of EZH2 in this setting is unclear due to the context-dependent functions of PRC2 and the heterogeneity of breast cancer. Moreover, the mechanisms underlying PRC2 overexpression in cancer are obscure. Here, using multiple models of breast cancer driven by the oncogene ErbB2, we show that the tyrosine kinase c-Src links energy sufficiency with PRC2 overexpression via control of mRNA translation. By stimulating mitochondrial ATP production, c-Src suppresses energy stress, permitting sustained activation of the mammalian/mechanistic target of rapamycin complex 1 (mTORC1), which increases the translation of mRNAs encoding the PRC2 subunits Ezh2 and Suz12. We show that Ezh2 overexpression and activity are pivotal in ErbB2-mediated mammary tumourigenesis. These results reveal the hitherto unknown c-Src/mTORC1/PRC2 axis, which is essential for ErbB2-driven carcinogenesis.


Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Epigênese Genética , Complexo Repressor Polycomb 2/genética , Receptor ErbB-2/metabolismo , Quinases da Família src/metabolismo , Trifosfato de Adenosina/metabolismo , Adulto , Animais , Neoplasias da Mama/patologia , Carcinogênese , Linhagem Celular Tumoral , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Feminino , Humanos , Glândulas Mamárias Humanas/metabolismo , Glândulas Mamárias Humanas/patologia , Alvo Mecanístico do Complexo 1 de Rapamicina/genética , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Camundongos , Camundongos Endogâmicos NOD , Camundongos Transgênicos , Pessoa de Meia-Idade , Mitocôndrias/genética , Mitocôndrias/metabolismo , Complexo Repressor Polycomb 2/metabolismo , Biossíntese de Proteínas , Receptor ErbB-2/genética , Quinases da Família src/genética
2.
J Agric Food Chem ; 67(16): 4493-4504, 2019 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-30938528

RESUMO

Expression of sodium-iodide symporter (NIS) is stimulated by sterol-regulatory-element-binding transcription factors (SREBFs) in mammary epithelial MCF-7 cells. Because conjugated linoleic acid (CLA) isomers have been shown to inhibit transcriptional activity of SREBFs in the mammary gland, the hypothesis was tested that CLA isomers inhibit NIS expression induced by all- trans retinoic acid (ATRA) in MCF-7 cells through inhibiting SREBF activity. c9t11-CLA and t10c12-CLA decreased ATRA-induced NIS-mRNA expression from 1.00 (ATRA alone) to 0.80 ± 0.12 (200 µM c9t11-CLA, P < 0.05) and 0.62 ± 0.10 (200 µM t10c12-CLA, P < 0.05), NIS-protein expression from 1.00 (ATRA alone) to 0.77 ± 0.08 (200 µM c9t11-CLA, P < 0.05) and 0.63 ± 0.05 (200 µM t10c12-CLA, P < 0.05), and NIS-promoter activity from 1.00 (ATRA alone) to 0.74 ± 0.13 (200 µM c9t11-CLA, P < 0.05) and 0.76 ± 0.13 (200 µM t10c12-CLA, P < 0.05); however, c9t11-CLA and t10c12-CLA increased the mRNA levels of SREBF isoforms and their target genes. In contrast, the mRNA expression of peroxisome-proliferator-activated receptor γ (PPARG) was strongly induced by ATRA alone but decreased by CLA isomers from 1.00 (ATRA alone) to 0.80 ± 0.06 (200 µM c9t11-CLA, P < 0.05) and 0.86 ± 0.06 (200 µM t10c12-CLA, P < 0.05). Overexpression of PPARγ in MCF-7 cells increased basal NIS-promoter activity, and treatment with the PPARγ ligand troglitazone stimulated ATRA-induced NIS-promoter activity. In conclusion, the results suggest that CLA isomers exert their effect on the expression of NIS by decreasing PPARG expression in MCF-7 cells.


Assuntos
Células Epiteliais/efeitos dos fármacos , Ácidos Linoleicos Conjugados/farmacologia , Glândulas Mamárias Humanas/metabolismo , Simportadores/genética , Tretinoína/farmacologia , Células Epiteliais/metabolismo , Expressão Gênica/efeitos dos fármacos , Humanos , Isomerismo , Células MCF-7 , Glândulas Mamárias Humanas/citologia , Glândulas Mamárias Humanas/efeitos dos fármacos , PPAR gama/genética , PPAR gama/metabolismo , Regiões Promotoras Genéticas/efeitos dos fármacos , Iodeto de Sódio/metabolismo , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismo , Simportadores/metabolismo
3.
Nat Commun ; 10(1): 1915, 2019 04 23.
Artigo em Inglês | MEDLINE | ID: mdl-31015424

RESUMO

Bromodomains (BRDs) are conserved protein interaction modules which recognize (read) acetyl-lysine modifications, however their role(s) in regulating cellular states and their potential as targets for the development of targeted treatment strategies is poorly understood. Here we present a set of 25 chemical probes, selective small molecule inhibitors, covering 29 human bromodomain targets. We comprehensively evaluate the selectivity of this probe-set using BROMOscan and demonstrate the utility of the set identifying roles of BRDs in cellular processes and potential translational applications. For instance, we discovered crosstalk between histone acetylation and the glycolytic pathway resulting in a vulnerability of breast cancer cell lines under conditions of glucose deprivation or GLUT1 inhibition to inhibition of BRPF2/3 BRDs. This chemical probe-set will serve as a resource for future applications in the discovery of new physiological roles of bromodomain proteins in normal and disease states, and as a toolset for bromodomain target validation.


Assuntos
Antineoplásicos/farmacologia , Células Epiteliais/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/farmacologia , Acetilação , Sequência de Aminoácidos , Antineoplásicos/química , Linhagem Celular Tumoral , Epigênese Genética , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Glucose/deficiência , Transportador de Glucose Tipo 1/antagonistas & inibidores , Transportador de Glucose Tipo 1/genética , Transportador de Glucose Tipo 1/metabolismo , Glicólise/efeitos dos fármacos , Glicólise/genética , Ensaios de Triagem em Larga Escala , Histonas/genética , Histonas/metabolismo , Humanos , Glândulas Mamárias Humanas/metabolismo , Glândulas Mamárias Humanas/patologia , Proteínas Nucleares/antagonistas & inibidores , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Transdução de Sinais , Bibliotecas de Moléculas Pequenas/química , Relação Estrutura-Atividade
4.
Int J Mol Sci ; 20(5)2019 Mar 12.
Artigo em Inglês | MEDLINE | ID: mdl-30871110

RESUMO

We first demonstrated that long-term increased polyamine (spermine, spermidine, putrescine) intake elevated blood spermine levels in mice and humans, and lifelong consumption of polyamine-rich chow inhibited aging-associated increase in aberrant DNA methylation, inhibited aging-associated pathological changes, and extend lifespan of mouse. Because gene methylation status is closely associated with aging-associated conditions and polyamine metabolism is closely associated with regulation of gene methylation, we investigated the effects of extracellular spermine supplementation on substrate concentrations and enzyme activities involved in gene methylation. Jurkat cells and human mammary epithelial cells were cultured with spermine and/or D,L-alpha-difluoromethylornithine (DFMO), an inhibitor of ornithine decarboxylase. Spermine supplementation inhibited enzymatic activities of adenosylmethionine decarboxylase in both cells. The ratio of decarboxylated S-adenosylmethionine to S-adenosyl-L-methionine increased by DFMO and decreased by spermine. In Jurkat cells cultured with DFMO, the protein levels of DNA methyltransferases (DNMTs) 1, 3A and 3B were not changed, however the activity of the three enzymes markedly decreased. The protein levels of these enzymes were not changed by addition of spermine, DNMT 3A and especially 3B were activated. We show that changes in polyamine metabolism dramatically affect substrate concentrations and activities of enzymes involved in gene methylation.


Assuntos
DNA (Citosina-5-)-Metiltransferases/metabolismo , Espermina/metabolismo , Adenosilmetionina Descarboxilase/metabolismo , Linhagem Celular Tumoral , Células Cultivadas , Metilação de DNA/fisiologia , Metilases de Modificação do DNA/metabolismo , Eflornitina/metabolismo , Células Epiteliais/metabolismo , Humanos , Células Jurkat , Glândulas Mamárias Humanas/metabolismo , Ornitina Descarboxilase/metabolismo , Poliaminas/metabolismo , Putrescina/metabolismo , S-Adenosilmetionina/análogos & derivados , S-Adenosilmetionina/metabolismo , Espermidina/metabolismo
5.
J Dairy Sci ; 102(4): 3071-3081, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30712927

RESUMO

Maternal milk is the primary source of nutrition for suckling mammals, and its yield and composition are important determinants of survival during the early neonatal period. The objective of this study was to examine whether parenteral administration of l-Arg to twin-bearing ewes, during mid to late pregnancy, influenced prepartum maternal mammary gland development and subsequent lactation performance in the early postpartum period (14 d). At 80 d of pregnancy, multiparous Romney ewes were housed indoors in group pens, split into 2 cohorts, and fed a lucerne-based pellet diet, formulated to meet 100% of National Research Council-recommended requirements for twin-bearing pregnant ewes, once a day. Cohort 1 was administered l-Arg (72.7 mg/kg of live weight via i.v, 3 times a day) from d 100 of pregnancy until d 140. At d 140, ewes were euthanized and maternal mammary tissues were collected for analysis of the biochemical indices total DNA, RNA, protein, protein synthetic efficiency (protein:RNA), cell size (protein:DNA), transcriptional efficiency (RNA:DNA), and the abundance of mammalian target of rapamycin (mTOR) and mTORSer2448 protein. Cohort 2 was administered an identical l-Arg regimen as cohort 1, but from d 100 until parturition. Milk was collected over a 14-d period (d 1, 4, 7, 10, and 14) to assess milk yield and composition. In cohort 1, total mammary DNA (cell number) tended to be higher in l-Arg ewes, with no change in total mammary RNA or protein content, biochemical indices of protein synthetic efficiency, cell size or transcriptional efficiency, or mTOR protein abundance or phosphorylation. In cohort 2, milk composition analysis from l-Arg ewes showed lower (d 7-14) milk somatic cell counts, greater crude protein percentage from d 7 to 10 but lower at d 14, and altered absolute concentrations of some free AA (d 7 and 14) compared with controls. We propose that parenteral administration of l-Arg during late pregnancy is associated with increased mammary gland cellular content and decreased somatic cell counts during early lactation.


Assuntos
Arginina/metabolismo , Leite/metabolismo , Ovinos/metabolismo , Fenômenos Fisiológicos da Nutrição Animal , Animais , Contagem de Células , Estudos de Coortes , Suplementos Nutricionais/análise , Feminino , Humanos , Lactação , Glândulas Mamárias Humanas/metabolismo , Leite/química , Gravidez , Fenômenos Fisiológicos da Nutrição Pré-Natal , Ovinos/genética , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , Gêmeos
6.
Microb Pathog ; 128: 268-275, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30630066

RESUMO

The goal of current investigations was to reveal the molecular mechanism triggered through feeding a diet with high-concentrate to dairy cows for subacute ruminal acidosis (SARA) induction and to examine the oxidative stress parameters in their mammary epithelial tissue. In an eighteen-weeks feeding trial, 12 Holstein Friesian cows with a standard weight of 455 ±â€¯28 kg were evenly divided into two groups and given either a low-concentrate (LC, forage to concentrate ratio = 6:4) or a high-concentrate (HC, forage to concentrate ratio = 4:6) diet. A remarkable reduction in ruminal pH also increased ruminal lipopolysaccharide (LPS) concentration that was observed in the high-concentrate group of cows at 4 h post-feeding in the morning. Moreover, reduced milk yield was observed in the HC group. The relative mRNA abundance of glutathione peroxidase (GPX) 1 and 3 and superoxide dismutase (SOD) 1 and 2 were down-regulated in high-concentrate fed animals than in the LC, while mRNA was expressed with no change in the of SOD3 among groups. In addition, genes responsible for oxidative stress e.g., ERK, JNK, and p38 were also showed dramatically high mRNA intensity in HC group. The protein concentration of ERK, pERK, pJNK, with pp38, were up-regulated significantly as JNK & p38 showed no big difference. While Nrf2 and pNrf2 were down-regulated considerably in HC group. The total antioxidant capacity (T-AOC) was significantly decreased but of Malondialdehyde (MDA) concentration was raised in HC group than in LC. We thus proposed that higher levels of endogenous LPS may affect the Mitogen-activated protein kinases (MAPK) and nuclear factor erythroid 2-related factor 2 (Nrf2)-dependent antioxidant response.


Assuntos
Ração Animal , Dieta/veterinária , Lipopolissacarídeos/farmacologia , Glândulas Mamárias Humanas/efeitos dos fármacos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Acidose/veterinária , Animais , Antioxidantes/metabolismo , Bovinos , Feminino , Expressão Gênica/efeitos dos fármacos , Glutationa Peroxidase/metabolismo , Humanos , Concentração de Íons de Hidrogênio , Inflamação , Lipopolissacarídeos/sangue , Sistema de Sinalização das MAP Quinases , Glândulas Mamárias Humanas/metabolismo , Leite/metabolismo , RNA Mensageiro/metabolismo , Transdução de Sinais/genética , Superóxido Dismutase/metabolismo , Superóxido Dismutase-1/metabolismo
7.
Neoplasia ; 21(1): 132-145, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30550871

RESUMO

The organization of the extracellular matrix has a profound impact on cancer development and progression. The matrix becomes aligned throughout tumor progression, providing "highways" for tumor cell invasion. Aligned matrix is associated with breast density and is a negative prognostic factor in several cancers; however, the underlying mechanisms regulating this reorganization remain poorly understood. Deletion of the tumor suppressor Pten in the stroma was previously shown to promote extracellular matrix expansion and tumor progression. However, it was unknown if PTEN also regulated matrix organization. To address this question, a murine model with fibroblast-specific Pten deletion was used to examine how PTEN regulates matrix remodeling. Using second harmonic generation microscopy, Pten deletion was found to promote collagen alignment parallel to the mammary duct in the normal gland and further remodeling perpendicular to the tumor edge in tumor-bearing mice. Increased alignment was observed with Pten deletion in vitro using fibroblast-derived matrices. PTEN loss was associated with fibroblast activation and increased cellular contractility, as determined by traction force microscopy. Inhibition of contractility abrogated the increased matrix alignment observed with PTEN loss. Murine mammary adenocarcinoma cells cultured on aligned matrices derived from Pten-/- fibroblasts migrated faster than on matrices from wild-type fibroblasts. Combined, these data demonstrate that PTEN loss in fibroblasts promotes extracellular matrix deposition and alignment independently from cancer cell presence, and this reorganization regulates cancer cell behavior. Importantly, stromal PTEN negatively correlated with collagen alignment and high mammographic density in human breast tissue, suggesting parallel function for PTEN in patients.


Assuntos
Matriz Extracelular/metabolismo , Glândulas Mamárias Animais/metabolismo , PTEN Fosfo-Hidrolase/metabolismo , Células Estromais/metabolismo , Animais , Densidade da Mama , Linhagem Celular Tumoral , Movimento Celular , Colágeno/metabolismo , Feminino , Fibroblastos/metabolismo , Técnicas de Inativação de Genes , Humanos , Glândulas Mamárias Animais/patologia , Glândulas Mamárias Humanas/metabolismo , Glândulas Mamárias Humanas/patologia , Camundongos , Camundongos Transgênicos , PTEN Fosfo-Hidrolase/genética
8.
Anim Sci J ; 90(2): 214-221, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30556368

RESUMO

The aim of this study was to determine the effects of isonitrogenous and isocaloric diets containing different qualities of forages and concentrate content on milk fat composition and genes that encode mammary lipogenic enzymes in dairy cows. A total of 20 Holstein cows were assigned to 1 of 2 treatment diets composed of either mixed forages (MF, starch : 21.50%) or corn stover forage (CS, starch : 25.39%). Mammary tissue biopsies were performed to analyze the mRNA expression of lipogenic enzymes. Dry matter intake, body weight, milk protein, and lactose were not affected by treatments. The milk yield, fat content and saturated fatty acid (SFA) and short- and medium-chain fatty acid (SMFA) contents in milk were lower in the CS diet than in the MF diet, but the unsaturated FA and long-chain FA contents were higher. Genes involved in de novo FA synthesis, FA uptake and transport, and Δ9-desaturation were lower in the CS treatment than in the MF treatment. No effects on the nuclear transcription factors were observed between the two treatments. The data indicated that corn stover diet reduced the milk yield, fat content, SMFA, and SFA contents in milk, as well as the gene expression of mammary lipogenic enzymes in dairy cows.


Assuntos
Ração Animal , Bovinos/metabolismo , Bovinos/fisiologia , Dieta/veterinária , Ácidos Graxos/metabolismo , Lactação , Lipogênese/genética , Glândulas Mamárias Humanas/enzimologia , Glândulas Mamárias Humanas/metabolismo , Leite/metabolismo , Acetil-CoA Carboxilase/genética , Acetil-CoA Carboxilase/metabolismo , Ração Animal/análise , Animais , Coenzima A Ligases/genética , Coenzima A Ligases/metabolismo , Proteína 3 Ligante de Ácido Graxo/genética , Proteína 3 Ligante de Ácido Graxo/metabolismo , Ácidos Graxos/biossíntese , Feminino , Expressão Gênica , Humanos , Lipase Lipoproteica/genética , Lipase Lipoproteica/metabolismo , Proteínas do Leite/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Amido , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismo , Zea mays
9.
Oncogene ; 38(14): 2437-2450, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30531835

RESUMO

Tumor progression is regulated by a complex interplay between neoplastic cells and the tumor microenvironment. Tumor-associated macrophages have been shown to promote breast cancer progression in advanced disease and more recently, in early stage cancers. However, little is known about the macrophage-derived factors that promote tumor progression in early stage lesions. Using a p53-null model of early stage mammary tumor progression, we found that Gas6 is highly expressed in pre-invasive lesions associated with increased infiltrating macrophages, as compared with those with few recruited macrophages. We show that F4/80+CD11b+ macrophages produce Gas6 in premalignant lesions in vivo, and that macrophage-derived Gas6 induces a tumor-like phenotype ex vivo. Using a 3-D co-culture system, we show that macrophage-derived Gas6 activates its receptor Axl and downstream survival signals including Akt and STAT3, which was accompanied by altered E-cadherin expression to induce a malignant morphology. In vivo studies demonstrated that deletion of stromal Gas6 delays early stage progression and decreases tumor formation, while tumor growth in established tumors remains unaffected. These studies suggest that macrophage-derived Gas6 is a critical regulator of the transition from premalignant to invasive cancer, and may lead to the development of unique biomarkers of neoplastic progression for patients with early stage breast cancer, including ductal carcinoma in situ.


Assuntos
Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Glândulas Mamárias Humanas/metabolismo , Glândulas Mamárias Humanas/patologia , Animais , Biomarcadores Tumorais/metabolismo , Caderinas/metabolismo , Carcinoma Intraductal não Infiltrante/metabolismo , Carcinoma Intraductal não Infiltrante/patologia , Proliferação de Células/fisiologia , Progressão da Doença , Feminino , Humanos , Macrófagos/metabolismo , Macrófagos/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/fisiologia , Microambiente Tumoral/fisiologia
10.
Clin Sci (Lond) ; 132(24): 2583-2598, 2018 12 21.
Artigo em Inglês | MEDLINE | ID: mdl-30545896

RESUMO

Estrogens generated within endocrine organs and the reproductive system act as ligands for at least three types of estrogen receptors. Estrogen receptors α (ERα) and ß (ERß) belong to the so-called classical family of estrogen receptors, whereas the G protein-coupled receptor GPR30, also known as GPER-1, has been described as a novel estrogen receptor sited in the cell membrane of target cells. Furthermore, these receptors are under stimulation of a family of exogenous estrogens, known as phytoestrogens, which are a diverse group of non-steroidal plant compounds derived from plant food consumed by humans and animals. Because phytoestrogens are omnipresent in our daily diet, they are becoming increasingly important in both human health and disease. Recent evidence indicates that in addition to classical estrogen receptors, phytoestrogens also activate GPER-1 a relevant observation since GPER-1 is involved in several physiopathological disorders and especially in estrogen-dependent diseases such as breast cancer.The first estrogen receptors discovered were the classical ERα and ERß, but from an evolutionary point of view G protein-coupled receptors trace their origins in history to over a billion years ago suggesting that estrogen receptors like GPER-1 may have been the targets of choice for ancient phytoestrogens and/or estrogens.This review provides a comprehensive and systematic literature search on phytoestrogens and its relationship with classical estrogen receptors and GPER-1 including its role in breast cancer, an issue still under discussion.


Assuntos
Anticarcinógenos/administração & dosagem , Neoplasias da Mama/metabolismo , Antagonistas de Estrogênios/administração & dosagem , Glândulas Mamárias Humanas/efeitos dos fármacos , Fitoestrógenos/administração & dosagem , Receptores Acoplados a Proteínas-G/agonistas , Animais , Anticarcinógenos/efeitos adversos , Neoplasias da Mama/induzido quimicamente , Neoplasias da Mama/epidemiologia , Neoplasias da Mama/prevenção & controle , Transformação Celular Neoplásica/induzido quimicamente , Transformação Celular Neoplásica/metabolismo , Exposição Dietética/efeitos adversos , Antagonistas de Estrogênios/efeitos adversos , Feminino , Humanos , Glândulas Mamárias Humanas/metabolismo , Glândulas Mamárias Humanas/patologia , Fitoestrógenos/efeitos adversos , Fatores de Proteção , Receptores Estrogênicos/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo , Medição de Risco , Fatores de Risco , Transdução de Sinais/efeitos dos fármacos
11.
BMC Cancer ; 18(1): 1264, 2018 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-30563501

RESUMO

BACKGROUND: Obesity is associated with oxidative stress, a major factor in carcinogenesis, and with high leptin concentration. The aim of this study was to determine the effects of leptin on the antioxidant response in three human mammary epithelial cells each presenting a different neoplastic status: healthy human mammary epithelial cells (HMEC), oestrogen-receptor positive MCF-7 cells and triple-negative MDA-MB-231 cells. METHODS: This in vitro kinetic study characterized the cell antioxidant response after 1, 6 and 24 h in the presence of leptin (10 or 100 ng/ml).The antioxidant response was defined in terms of cell glutathione content, gene expression and catalytic activity of antioxidant enzymes (i.e. glutathione peroxidase 1 (Gpx1), glutathione reductase (GR), glutathione S transferase (GST), heme-oxygenase 1 (HO-1) and cyclooxygenase-2 (COX-2)). Oxidative stress occurrence was assessed by lipid hydro peroxide (HPLIP) and isoprostane concentrations in culture media at 24 h. RESULTS: At both concentrations used, leptin induced ROS production in all cell models, contributing to various antioxidant responses linked to neoplastic cell status. HMEC developed a highly inducible antioxidant response based on antioxidant enzyme activation and an increase in cell GSH content at 10 ng/ml of leptin. However, at 100 ng/ml of leptin, activation of antioxidant response was lower. Conversely, in tumour cells, MCF-7 and MDA-MB-231, leptin did not induce an efficient antioxidant response, at either concentration, resulting in an increase of lipid peroxidation products. CONCLUSIONS: Leptin can modulate the oxidative status of mammary epithelial cells differently according to their neoplastic state. These novel results shed light on oxidative status changes in mammary cells in the presence of leptin.


Assuntos
Antioxidantes/administração & dosagem , Leptina/administração & dosagem , Glândulas Mamárias Humanas/metabolismo , Obesidade/metabolismo , Antioxidantes/metabolismo , Carcinogênese/efeitos dos fármacos , Ciclo-Oxigenase 2/metabolismo , Feminino , Glutationa Peroxidase/metabolismo , Glutationa Redutase/metabolismo , Glutationa Transferase/metabolismo , Heme Oxigenase-1/metabolismo , Humanos , Leptina/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Células MCF-7 , Glândulas Mamárias Humanas/efeitos dos fármacos , Obesidade/tratamento farmacológico , Obesidade/enzimologia , Obesidade/patologia , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Superóxido Dismutase/metabolismo
12.
BMC Cancer ; 18(1): 1273, 2018 Dec 19.
Artigo em Inglês | MEDLINE | ID: mdl-30567518

RESUMO

BACKGROUND: Breast cancer is the most common malignancy in women affecting one out of eight females throughout their lives. Autotaxin (ATX) is upregulated in breast cancer which results in increased lysophosphatidic acid (LPA) formation within the tumor. This study's aim was to identify the role of different mammary cell populations within the ATX-LPA axis. METHODS: Epithelial-cell-adhesion-molecule-positive (EpCAM) and -negative cells from breast tumors, adipose-derived stem cells (ADSCs) of tumor-adjacent and tumor-distant mammary fat were isolated and compared to healthy ADSCs, mammary epithelial cells (HMECs), and mesenchymal cells (MES) of healthy mammary tissue (n = 4 each) and further to well-established breast (cancer) cell lines. RESULTS: mRNA expression analyses revealed that ADSCs and MES largely expressed LPA receptor 1 (LPAR1) while epithelial cells mainly expressed LPAR6. LPA 18:1 activated all the cell populations and cell lines by rise in cytosolic free calcium concentrations. MES and ADSCs expressed ATX whereas epithelial cells did not. ADSCs revealed the highest expression in ATX with a significant decline after adipogenic differentiation in healthy ADSCs, whereas ATX expression increased in ADSCs from tumor patients. Breast (cancer) cell lines did not express ATX. Transmigration of MES was stimulated by LPA whereas an inhibitory effect was observed in epithelial cells with no differences between tumors and healthy cells. Triple-negative breast cancer (TNBC) cell lines were also stimulated and the transmigration partly inhibited using the LPA receptor antagonist Ki16425. CONCLUSIONS: We here show that each mammary cell population plays a different role in the ATX-LPA axis with ADSCs and adipocytes being the main source of ATX in tumor patients in our experimental setting. Inhibitors of this axis may therefore present a valuable target for pharmacological therapies.


Assuntos
Lisofosfolipídeos/genética , Diester Fosfórico Hidrolases/genética , Receptores de Ácidos Lisofosfatídicos/genética , Neoplasias de Mama Triplo Negativas/genética , Adipócitos/metabolismo , Adipócitos/patologia , Diferenciação Celular/genética , Linhagem Celular Tumoral , Linhagem da Célula/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Glândulas Mamárias Humanas/metabolismo , Células-Tronco Mesenquimais/metabolismo , RNA Mensageiro/genética , Neoplasias de Mama Triplo Negativas/metabolismo , Neoplasias de Mama Triplo Negativas/patologia
13.
BMC Bioinformatics ; 19(1): 540, 2018 Dec 22.
Artigo em Inglês | MEDLINE | ID: mdl-30577750

RESUMO

BACKGROUND: DNA methylation of CpG dinucleotides is an essential epigenetic modification that plays a key role in transcription. Widely used DNA enrichment-based methods offer high coverage for measuring methylated CpG dinucleotides, with the lowest cost per CpG covered genome-wide. However, these methods measure the DNA enrichment of methyl-CpG binding, and thus do not provide information on absolute methylation levels. Further, the enrichment is influenced by various confounding factors in addition to methylation status, for example, CpG density. Computational models that can accurately derive absolute methylation levels from DNA enrichment data are needed. RESULTS: We developed "MeDEStrand," a method that uses a sigmoid function to estimate and correct the CpG bias from enrichment results to infer absolute DNA methylation levels. Unlike previous methods, which estimate CpG bias based on reads mapped at the same genomic loci, MeDEStrand processes the reads for the positive and negative DNA strands separately. We compared the performance of MeDEStrand to that of three other state-of-the-art methods "MEDIPS," "BayMeth," and "QSEA" on four independent datasets generated using immortalized cell lines (GM12878 and K562) and human primary cells (foreskin fibroblasts and mammary epithelial cells). Based on the comparison of the inferred absolute methylation levels from MeDIP-seq data and the corresponding reduced-representation bisulfite sequencing data from each method, MeDEStrand showed the best performance at high resolution of 25, 50, and 100 base pairs. CONCLUSIONS: The MeDEStrand tool can be used to infer whole-genome absolute DNA methylation levels at the same cost of enrichment-based methods with adequate accuracy and resolution. R package MeDEStrand and its tutorial is freely available for download at https://github.com/jxu1234/MeDEStrand.git .


Assuntos
Metilação de DNA , DNA/análise , Epigênese Genética , Fibroblastos/metabolismo , Genoma Humano , Glândulas Mamárias Humanas/metabolismo , Análise de Sequência de DNA/métodos , Células Cultivadas , Ilhas de CpG , DNA/metabolismo , Fibroblastos/citologia , Genômica/métodos , Humanos , Glândulas Mamárias Humanas/citologia
14.
Yakugaku Zasshi ; 138(6): 829-836, 2018.
Artigo em Japonês | MEDLINE | ID: mdl-29863055

RESUMO

 Intrinsic serotonin (5-hydroxytryptamine; 5-HT) synthesized within the mammary epithelium has an important physiological role in milk volume homeostasis in many species including mice, cows, and humans. During lactation, mammary epithelial cells activate 5-HT synthesis by tryptophan hydroxylase 1 (TPH1). TPH1 catalyzes the rate-limiting step in 5-HT biosynthesis within mammary glands. 5-HT synthesized in mammary glands is released into both the apical (milk) and basolateral spaces by a vesicular monoamine transporter. 5-HT released into milk is incorporated by the apical membrane-expressed serotonin reuptake transporter and degraded by the monoamine oxidase A enzyme. Suckling maintains 5-HT at low levels in milk. When the mammary gland becomes filled with milk, 5-HT provides a negative feedback signal that suppresses further milk synthesis in the mammary epithelium. Our research, using human mammary epithelial MCF-12A cells, shows that the expression of ß-casein, a differentiation marker, is suppressed via 5-HT-mediated inhibition of signal transducer and activator of transcription 5. Additionally, our results show that reduced ß-casein expression in MCF-12A cells is associated with 5-HT7 receptor expression. Furthermore, we show that 5-HT7 receptor-mediated suppression of ß-casein expression is involved in the activation of protein kinase A and protein-tyrosine phosphatase 1B. Thus, this mechanism might be associated with the feedback signals by 5-HT within the mammary epithelium. Hence, further research that builds on our findings should include the elucidation of the physiological roles of 5-HT present in milk synthesized by mammary epithelial cells in vivo and its effects on nursing infants.


Assuntos
Lactação/efeitos dos fármacos , Serotonina/fisiologia , Animais , Caseínas/metabolismo , Feminino , Homeostase , Humanos , Glândulas Mamárias Animais/metabolismo , Glândulas Mamárias Humanas/metabolismo , Monoaminoxidase/fisiologia , Receptores de Serotonina/fisiologia , Serotonina/biossíntese , Triptofano Hidroxilase/fisiologia
15.
Biochim Biophys Acta Mol Cell Biol Lipids ; 1863(9): 1057-1067, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-29906613

RESUMO

Fatty acid-binding proteins (FABPs) are involved in binding and storing hydrophobic ligands such as long-chain fatty acids, as well as transporting them to the appropriate compartments in the cell. Epidermal fatty acid-binding protein (FABP5) is an intracellular lipid-binding protein that is abundantly expressed in adipocytes and macrophages. Previous studies have revealed that the FABP5 expression level is closely related to malignancy in various types of cancer. However, its precise functions in the metabolisms of cancer cells remain unclear. Here, we revealed that FABP5 knockdown significantly induced downregulation of the genes expression, such as hormone-sensitive lipase (HSL), monoacylglycerol lipase (MAGL), elongation of long-chain fatty acid member 6 (Elovl6), and acyl-CoA synthetase long-chain family member 1 (ACSL1), which are involved in altered lipid metabolism, lipolysis, and de novo FA synthesis in highly aggressive prostate and breast cancer cells. Moreover, we demonstrated that FABP5 induced inflammation and cytokine production through the nuclear factor-kappa B signaling pathway activated by reactive oxygen species and protein kinase C in PC-3 and MDA-MB-231 cells. Thus, FABP5 might regulate lipid quality and/or quantity to promote aggressiveness such as cell growth, invasiveness, survival, and inflammation in prostate and breast cancer cells. In the present study, we have revealed for the first time that high expression of FABP5 plays a critical role in alterations of lipid metabolism, leading to cancer development and metastasis in highly aggressive prostate and breast cancer cells.


Assuntos
Proteínas de Ligação a Ácido Graxo/genética , Ácidos Graxos/biossíntese , Regulação Neoplásica da Expressão Gênica , Glândulas Mamárias Humanas/metabolismo , NF-kappa B/genética , Próstata/metabolismo , Acetiltransferases/genética , Acetiltransferases/metabolismo , Linhagem Celular Tumoral , Coenzima A Ligases/genética , Coenzima A Ligases/metabolismo , Citocinas/genética , Citocinas/metabolismo , Proteínas de Ligação a Ácido Graxo/antagonistas & inibidores , Proteínas de Ligação a Ácido Graxo/metabolismo , Feminino , Humanos , Gotículas Lipídicas/química , Gotículas Lipídicas/metabolismo , Lipólise , Masculino , Glândulas Mamárias Humanas/patologia , Monoacilglicerol Lipases/genética , Monoacilglicerol Lipases/metabolismo , NF-kappa B/metabolismo , Próstata/patologia , Proteína Quinase C/genética , Proteína Quinase C/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Esterol Esterase/genética , Esterol Esterase/metabolismo
16.
Int J Mol Sci ; 19(6)2018 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-29875329

RESUMO

Since seminal descriptions of signal transducer and activator of transcription 3 (STAT3) as a signal transducer and transcriptional regulator, which is most usually activated by phosphorylation of a specific tyrosine residue, a staggering wealth of research has delineated the key role of this transcription factor as a mediator of mammary gland postlactational regression (involution), and paradoxically, a pro-survival factor in breast cancer and some breast cancer cell lines. STAT3 is a critical regulator of lysosomal-mediated programmed cell death (LM-PCD) during mammary gland involution, where uptake of milk fat globules, and consequent high levels of free fatty acids, cause permeabilisation of lysosomal vesicle membranes, in turn leading to cathepsin protease leakage and cell death. A recent proteomic screen of STAT3-induced changes in lysosomal membrane protein components has highlighted wide-ranging effects of STAT3, which may coordinate LM-PCD via the stimulation of endocytosis, intracellular trafficking, and lysosome biogenesis. In parallel, STAT3 regulates the acute phase response during the first phase of involution, and it contributes to shaping the pro-tumourigenic 'wound healing' signature of the gland during the second phase of this process. STAT3 activation during involution is important across species, although some differences exist in the progression of involution in dairy cows. In breast cancer, a number of upstream regulators can lead to STAT3 activation and the effects of phosphorylation of STAT3 are equally wide-ranging. Recent studies have implicated microRNAs in some regulatory pathways. In this review, we will examine the multifaceted role of STAT3 in mammary gland involution and tumourigenesis, incorporating a review of these fundamental processes in tandem with a discussion of recent developments in this field.


Assuntos
Neoplasias da Mama/etiologia , Neoplasias da Mama/metabolismo , Transformação Celular Neoplásica/metabolismo , Glândulas Mamárias Humanas/metabolismo , Fator de Transcrição STAT3/metabolismo , Animais , Biomarcadores , Neoplasias da Mama/patologia , Transformação Celular Neoplásica/genética , Microambiente Celular , Feminino , Regulação da Expressão Gênica , Humanos , Glândulas Mamárias Humanas/patologia , Fator de Transcrição STAT3/antagonistas & inibidores , Fator de Transcrição STAT3/genética , Transdução de Sinais
17.
PLoS One ; 13(5): e0196854, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29718989

RESUMO

The bioactive lipid sphingosine-1-phosphate (S1P) is a main regulator of cell survival, proliferation, motility, and platelet aggregation, and it is essential for angiogenesis and lymphocyte trafficking. In that S1P acts as a second messenger intra- and extracellularly, it might promote cancer progression. The main cause is found in the high S1P concentration in the blood, which encourage cancer cells to migrate through the endothelial barrier into the blood vessels. The irreversible degradation of S1P is solely caused by the sphingosine-1-phosphate lyase (SGPL1). SGPL1 overexpression reduces cancer cell migration and therefore silences the endogenous S1P siren, which promotes cancer cell attraction-the main reason for metastasis. Since our previous metabolomics studies revealed an increased SGPL1 activity in association with successful breast cancer cell treatment in vitro, we further investigated expression and localization of SGPL1. Expression analyses confirmed a very low SGPL1 expression in all breast cancer samples, regardless of their subtype. Additionally, we were able to prove a novel SGPL expression in the cytoplasm membrane of non-tumorigenic breast cells by fusing three independent methods. The general SGPL1 downregulation and the loss of the plasma membrane expression resulted in S1P dependent stimulation of migration in the breast cancer cell lines MCF-7 and BT-20. Not only S1P stimulated migration could be repressed by overexpressing the natural SGPL1 variant not but also more general migratory activity was significantly reduced. Here, for the first time, we report on the SGPL1 plasma membrane location in human, non-malignant breast epithelial cell lines silencing the extracellular S1P siren in vitro, and thereby regulating pivotal cellular functions. Loss of this plasma membrane distribution as well as low SGPL1 expression levels could be a potential prognostic marker and a viable target for therapy. Therefore, the precise role of SGPL1 for cancer treatment should be evaluated.


Assuntos
Aldeído Liases/fisiologia , Membrana Celular/metabolismo , Lisofosfolipídeos/metabolismo , Glândulas Mamárias Humanas/metabolismo , Esfingosina/análogos & derivados , Aldeído Liases/metabolismo , Linhagem Celular Tumoral , Células Epiteliais/metabolismo , Regulação da Expressão Gênica , Humanos , Lisofosfolipídeos/fisiologia , Células MCF-7 , Metástase Neoplásica , Esfingosina/metabolismo , Esfingosina/fisiologia
18.
BMC Vet Res ; 14(1): 168, 2018 May 23.
Artigo em Inglês | MEDLINE | ID: mdl-29792195

RESUMO

BACKGROUND: Mounting evidences observed that subacute ruminal acidosis (SARA) induced by high concentration (HC) diet increases the translocation of histamine from digestive tract into circulation causing a diverse of diseases in dairy cows. However, it is largely unknown how it does affect the function of mammary gland and milk quality. Hence, this study aims to observe the effects of histamine derived from the digestive tract on the inflammatory response and casein synthesis in the mammary glands during SARA. Twelve cows fitted rumen fistula were randomly divided into either control group administrated low concentration (LC) diet (60% forage, n = 6) or treatment group administrated HC diet (40% forage, n = 6) for 18 weeks. RESULTS: Our data showed that HC diet resulted in significant declines in rumen pH value, milk yield and milk quality, as well as longer duration of averaged pH value below 5.6 per day (more than 180 min) compared to LC diet, these findings confirmed SARA occurence. Our study also observed that SARA increased the content of histamine in rumen fluid, plasma, liver and mammary gland, and enhanced the mRNA expression of histamine specific receptor in the mammary gland. Additionally, we found that the mRNA expression of inflammatory response genes in mammary glands was increased, which was consistent with the protein expression results, showing that the protein kinase C(PKC) / nuclear factor kappa B (NF-κB) or protein kinase A (PKA) / NF-κB signalling pathways of the inflammatory response were activated. The mRNA expression of mTOR, P70S6K and αS1 in mammary glands were significantly decreased with the protein expression of mTOR, P70S6K and αS1-casein, and the phosphorylation levels of the mTOR and P70S6K proteins were also decreased. CONCLUSIONS: Our study showed that the milk protein of lactating cows is depressed after long-term feeding of HC at the individual level, which was paralleled at the gene and protein levels. The inflammatory response in mammary gland caused by histamine derived from the digestive tract is related to the decline of casein synthesis. Our findings point to a new link between the inflammatory response and casein synthesis, but the understanding of the molecular mechanisms involved in this process will require further research.


Assuntos
Acidose/veterinária , Caseínas/metabolismo , Doenças dos Bovinos/metabolismo , Histamina/metabolismo , Glândulas Mamárias Humanas/metabolismo , Gastropatias/veterinária , Acidose/metabolismo , Animais , Western Blotting/veterinária , Bovinos , Dieta/veterinária , Ensaio de Imunoadsorção Enzimática/veterinária , Feminino , Humanos , Lactação/metabolismo , Leite/química , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Rúmen/metabolismo , Gastropatias/metabolismo
19.
Proc Natl Acad Sci U S A ; 115(19): E4426-E4432, 2018 05 08.
Artigo em Inglês | MEDLINE | ID: mdl-29686092

RESUMO

Because of the ubiquitous adaptability of our material culture, some human populations have occupied extreme environments that intensified selection on existing genomic variation. By 32,000 years ago, people were living in Arctic Beringia, and during the Last Glacial Maximum (LGM; 28,000-18,000 y ago), they likely persisted in the Beringian refugium. Such high latitudes provide only very low levels of UV radiation, and can thereby lead to dangerously low levels of biosynthesized vitamin D. The physiological effects of vitamin D deficiency range from reduced dietary absorption of calcium to a compromised immune system and modified adipose tissue function. The ectodysplasin A receptor (EDAR) gene has a range of pleiotropic effects, including sweat gland density, incisor shoveling, and mammary gland ductal branching. The frequency of the human-specific EDAR V370A allele appears to be uniquely elevated in North and East Asian and New World populations due to a bout of positive selection likely to have occurred circa 20,000 y ago. The dental pleiotropic effects of this allele suggest an even higher occurrence among indigenous people in the Western Hemisphere before European colonization. We hypothesize that selection on EDAR V370A occurred in the Beringian refugium because it increases mammary ductal branching, and thereby may amplify the transfer of critical nutrients in vitamin D-deficient conditions to infants via mothers' milk. This hypothesized selective context for EDAR V370A was likely intertwined with selection on the fatty acid desaturase (FADS) gene cluster because it is known to modulate lipid profiles transmitted to milk from a vitamin D-rich diet high in omega-3 fatty acids.


Assuntos
Clima Frio , Receptor Edar , Ácidos Graxos/metabolismo , Troca Materno-Fetal/fisiologia , Leite Humano/metabolismo , Seleção Genética/fisiologia , Vitamina D/metabolismo , Alelos , Receptor Edar/genética , Receptor Edar/metabolismo , Feminino , Humanos , Masculino , Glândulas Mamárias Humanas/anatomia & histologia , Glândulas Mamárias Humanas/metabolismo , Gravidez
20.
Exp Anim ; 67(3): 361-371, 2018 Jul 30.
Artigo em Inglês | MEDLINE | ID: mdl-29526866

RESUMO

The aim of this study is to investigate the changes with age on morphology and sex hormone receptor expression in the mammary glands of male Sprague-Dawley rats, focusing on male-specific cells, "oxyphilic cells", observed after sexual maturity. The mammary glands of male rats at 14, 21, 35, 50, 75 and 100 days old were examined by gross observation, microscopic observation using whole mount specimens, histological and immunohistochemical sections. Grossly, mammary glands showed brown color at 50-100 days old. In whole mount specimens, terminal end buds (TEBs) were observed at 14-50 days old and the number of TEBs was highest at 35 days old. Histologically, the male mammary glands contained small epithelial cells with scanty cytoplasm at 14-35 days old while ductal and lobular epithelial cells were changed into oxyphilic cells with abundant cytoplasm at 50-100 days old. Immunohistochemicaly, androgen receptor (AR), estrogen receptor (ER) and progesterone receptor (PgR) expressions were found in both mammary glands found at a young age and oxyphilic cells. In oxyphilic cells, AR expression was dominant compared to ER and PgR expressions and increased with age. From these results, the development at 50-100 days old might be strongly related to AR. Ultrastructural observation of oxyphilic cells confirmed a number of lipid droplets, deformed and/or enlarged mitochondria, lysosomes and peroxisomes in their cytoplasm.


Assuntos
Glândulas Mamárias Humanas/crescimento & desenvolvimento , Glândulas Mamárias Humanas/metabolismo , Células Oxífilas/metabolismo , Receptores Androgênicos/metabolismo , Receptores Estrogênicos/metabolismo , Receptores de Progesterona/metabolismo , Envelhecimento/metabolismo , Animais , Humanos , Imuno-Histoquímica , Gotículas Lipídicas/metabolismo , Lisossomos/patologia , Masculino , Glândulas Mamárias Humanas/citologia , Mitocôndrias/patologia , Células Oxífilas/ultraestrutura , Peroxissomos/patologia , Ratos Sprague-Dawley , Maturidade Sexual
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