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1.
Nat Commun ; 11(1): 4779, 2020 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-32963246

RESUMO

Highly reproducible smoking-associated DNA methylation changes in whole blood have been reported by many Epigenome-Wide-Association Studies (EWAS). These epigenetic alterations could have important implications for understanding and predicting the risk of smoking-related diseases. To this end, it is important to establish if these DNA methylation changes happen in all blood cell subtypes or if they are cell-type specific. Here, we apply a cell-type deconvolution algorithm to identify cell-type specific DNA methylation signals in seven large EWAS. We find that most of the highly reproducible smoking-associated hypomethylation signatures are more prominent in the myeloid lineage. A meta-analysis further identifies a myeloid-specific smoking-associated hypermethylation signature enriched for DNase Hypersensitive Sites in acute myeloid leukemia. These results may guide the design of future smoking EWAS and have important implications for our understanding of how smoking affects immune-cell subtypes and how this may influence the risk of smoking related diseases.


Assuntos
Metilação de DNA/efeitos dos fármacos , Epigenoma , Fumar/efeitos adversos , Algoritmos , Grupo com Ancestrais do Continente Asiático , Sangue , Ilhas de CpG , Epigenômica/métodos , Grupos Étnicos , Feminino , Humanos , Linfócitos , Masculino , Pessoa de Meia-Idade , Modelos Estatísticos , Células Mieloides
2.
Ecotoxicol Environ Saf ; 203: 111031, 2020 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-32888610

RESUMO

Bone mineral density (BMD) changes were reported to be associated with excessive fluoride exposure and abnormal expression of RUNX2. However, whether the alteration of methylation status, a most commonly used marker for the alteration of gene expression in epidemiological investigation, of RUNX2 is associated with low-to-moderate fluoride exposure and BMD changes has not been reported. Our study aims to explore the role of RUNX2 promoter methylation in BMD changes induced by low-to-moderate fluoride exposure. A total of 1124 adults (413 men and 711 women) were recruited from Kaifeng City in 2017. We measured BMD using ultrasound bone densitometer. Concentrations of urinary fluoride (UF) were measured using ion-selective electrode, and the participants were grouped into control group (CG) and excessive fluoride group (EFG) according to the concentration of UF. We extracted DNA from fasting peripheral blood samples and then detected the promoter methylation levels of RUNX2 using quantitative methylation-specific PCR. Relationships between UF concentration, RUNX2 promoter methylation and BMD changes were analyzed using generalized linear model and logistic regression. Results showed in EFG (UF concentration > 1.6 mg/L), BMD was negatively correlated with UF concentration (ß: -0.14; 95%CI: -0.26, -0.01) and RUNX2 promoter methylation (ß: -0.13; 95%CI: -0.22, -0.03) in women. The methylation rate of RUNX2 promoter increased by 2.16% for each 1 mg/L increment in UF concentration of women in EFG (95%CI: 0.37, 3.96). No any significant associations between UF concentration, RUNX2 promoter methylation, and BMD were observed in the individuals in CG. Mediation analysis showed that RUNX2 promoter methylation mediated 18.2% (95% CI: 4.2%, 53.2%) of the association between UF concentration and BMD of women in EFG. In conclusion, excessive fluoride exposure (>1.6 mg/L) is associated with changes of BMD in women, and this association is mediated by RUNX2 promoter methylation.


Assuntos
Densidade Óssea/efeitos dos fármacos , Subunidade alfa 1 de Fator de Ligação ao Core/genética , Exposição Ambiental/análise , Fluoretos/toxicidade , Poluentes Químicos da Água/toxicidade , Absorciometria de Fóton , Adulto , Idoso , Densidade Óssea/genética , China , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Estudos Transversais , Metilação de DNA/efeitos dos fármacos , Feminino , Fluoretos/urina , Humanos , Masculino , Pessoa de Meia-Idade , Regiões Promotoras Genéticas , Inquéritos e Questionários , Poluentes Químicos da Água/urina
3.
Ecotoxicol Environ Saf ; 205: 111289, 2020 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-32949839

RESUMO

The Deepwater Horizon oil spill released 3.19 million barrels of crude oil into the Gulf of Mexico, making it the largest oil spill in U.S. history. Weathering and the application of dispersants can alter the solubility of compounds within crude oil, thus modifying the acute toxicity of the crude oil to aquatic life. The primary aim of our study was to determine the lasting impact of early-life stage sheepshead minnow (Cyprinodon variegatus variegatus) exposure to weathered, unweathered and dispersed crude oil on prey capture, male aggression, novel object interaction and global DNA methylation. Embryos were exposed from 1 to 10 dpf to water accommodations of crude oil and were raised to adulthood in artificial seawater. Our results suggest exposure to crude oil did not result in lasting impairment of complex behavioral responses of male sheepshead minnow. Exposure to dispersed weathered oil, however, decreased border dwelling in response to a novel object (i.e. decreased anxiety). Principal component analysis revealed that exposure to weathered oil had no overarching effect, but that unweathered crude oil increased variability in exploratory behaviors but decreased variability in anxiety-associated behaviors. Further work is needed to understand the effects of oil exposure on fish behavior and the potential ecological impact of subtle behavioral changes in fishes.


Assuntos
Comportamento Animal/efeitos dos fármacos , Peixes Listrados/fisiologia , Larva/efeitos dos fármacos , Poluição por Petróleo/efeitos adversos , Petróleo/toxicidade , Poluentes Químicos da Água/toxicidade , Animais , Metilação de DNA/efeitos dos fármacos , Ecologia , Golfo do México , Peixes Listrados/genética , Larva/genética , Larva/fisiologia , Masculino , Água do Mar/química , Tempo (Meteorologia)
4.
Ann Hematol ; 99(10): 2405-2416, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32813071

RESUMO

Predictive factors of response to hypomethylating agents (HMA) in elderly acute myeloid leukemia (AML) patients remain unclear in the real-life setting and no direct comparison between azacitidine (AZA) and decitabine (DEC) has been carried out. We retrospectively evaluated 110 AML patients treated with HMA (78 AZA, 32 DEC) as first-line therapy outside of clinical trials. Median age was 75 years (range 58-87). The median overall survival (OS) of the entire cohort was 8.0 months (95% CI 6.1-10), without significant differences among the subgroups: AZA 8.8 months vs DEC 6.3 months (p = 0.291). HMA treatment yielded an overall response rate (ORR) of 40% (AZA 37% vs DEC 47%, p = 0.237). A stable disease (SD) after 4 HMA cycles was not associated with a worse survival outcome compared with an early optimal response. Factors independently associated with a better OS were transfusion independence during treatment (p = 0.049), achievement of an optimal response to treatment (p < 0.001), and a baseline hemoglobin level ≥ 9.25 (p = 0.018). A bone marrow (BM) blast count ≥ 30% (p < 0.001) and a therapy-related AML (p = 0.008) remain poor survival predictors. Of the available biologic features, an adverse risk category according to the ELN classification was significantly associated with a shorter survival over the intermediate risk category (p = 0.034). Disease progression remains the primary cause of death. Infectious complications were more severe (p = 0.036) and occurred earlier (p = 0.006) in the DEC group compared with that of the AZA group. In conclusion, clinical prognostic factors associated to response and survival have been identified without significant associations concerning overall outcomes between the two HMAs.


Assuntos
Antimetabólitos Antineoplásicos/uso terapêutico , Azacitidina/uso terapêutico , Metilação de DNA/efeitos dos fármacos , DNA de Neoplasias/efeitos dos fármacos , Decitabina/uso terapêutico , Leucemia Mieloide Aguda/tratamento farmacológico , Idoso , Idoso de 80 Anos ou mais , Medula Óssea/patologia , Causas de Morte , Contagem de Células , DNA de Neoplasias/química , Progressão da Doença , Intervalo Livre de Doença , Feminino , Hemoglobinas/análise , Humanos , Infecções/etiologia , Infecções/mortalidade , Estimativa de Kaplan-Meier , Leucemia Mieloide Aguda/mortalidade , Masculino , Pessoa de Meia-Idade , Células-Tronco Neoplásicas , Modelos de Riscos Proporcionais , Estudos Retrospectivos , Resultado do Tratamento
5.
Aquat Toxicol ; 226: 105580, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32712368

RESUMO

Bisphenol A (BPA), a well-known estrogenic endocrine disruptor, is ubiquitously present in the environment, possessing the potential to interfere with the reproductive endocrine system in male mammals. However, there are limited studies on the reproductive toxicity in male aquatic animals associated with epigenetic modifications. In order to evaluate the potential effects of BPA on reproduction and better understand the underlying mechanism, adult male rare minnow (Gobiocypris rarus) were exposed to 15 µg L-1 BPA over a period of 63 d. Results showed that BPA induced congestion of blood vessels and infiltration of inflammatory cells after 21 d exposure, and decreased sperm fertilization after 63 d exposure. The genome DNA methylation levels were significantly increased throughout the treatment, and a strong positive stain were found in the spermatocyte, spermatid and sperm. The H3K4me3 level in all types of germ cell were increased by 21 d exposure while decreased following 63 d exposure. The positive stain of H3K9me3 was decreased in sperms while increased in spermatids by 21 d exposure. In addition, the H3K9me3 level was significantly increased after 63 d exposure, and a strong positive stain were found in spermatocytes, spermatids, and sperms. Our result also revealed that the transcripts of DNA methyltransferase genes (dnmt1 and dnmt3-8) and histone methyltransferase genes (mll2-5, setdb1-2 and ezh2) were also markedly changed under BPA exposure for 21-63 d. These findings indicated that BPA had toxicity in male reproductive, and DNA/histone methylation might play a vital role in the regulation of BPA-triggered the decreased of sperm quality.


Assuntos
Compostos Benzidrílicos/toxicidade , Cyprinidae/metabolismo , Metilação de DNA/efeitos dos fármacos , Disruptores Endócrinos/toxicidade , Histonas/metabolismo , Fenóis/toxicidade , Espermatozoides/efeitos dos fármacos , Poluentes Químicos da Água/toxicidade , Animais , DNA/metabolismo , Feminino , Humanos , Masculino , Análise do Sêmen , Espermatozoides/metabolismo , Testículo/efeitos dos fármacos
6.
Artigo em Inglês | MEDLINE | ID: mdl-32645824

RESUMO

Environmental lead (Pb) exposure is closely associated with pathogenesis of a range of neurological disorders, including Alzheimer's disease (AD), Parkinson's disease (PD), amyotrophic lateral sclerosis (ALS), attention deficit/hyperactivity disorder (ADHD), etc. Epigenetic machinery modulates neural development and activities, while faulty epigenetic regulation contributes to the diverse forms of CNS (central nervous system) abnormalities and diseases. As a potent epigenetic modifier, lead is thought to cause neurological disorders through modulating epigenetic mechanisms. Specifically, increasing evidence linked aberrant DNA methylations, histone modifications as well as ncRNAs (non-coding RNAs) with AD cases, among which circRNA (circular RNA) stands out as a new and promising field for association studies. In 23-year-old primates with developmental lead treatment, Zawia group discovered a variety of epigenetic changes relating to AD pathogenesis. This is a direct evidence implicating epigenetic basis in lead-induced AD animals with an entire lifespan. Additionally, some epigenetic molecules associated with AD etiology were also known to respond to chronic lead exposure in comparable disease models, indicating potentially interlaced mechanisms with respect to the studied neurotoxic and pathological events. Of note, epigenetic molecules acted via globally or selectively influencing the expression of disease-related genes. Compared to AD, the association of lead exposure with other neurological disorders were primarily supported by epidemiological survey, with fewer reports connecting epigenetic regulators with lead-induced pathogenesis. Some pharmaceuticals, such as HDAC (histone deacetylase) inhibitors and DNA methylation inhibitors, were developed to deal with CNS disease by targeting epigenetic components. Still, understandings are insufficient regarding the cause-consequence relations of epigenetic factors and neurological illness. Therefore, clear evidence should be provided in future investigations to address detailed roles of novel epigenetic factors in lead-induced neurological disorders, and efforts of developing specific epigenetic therapeutics should be appraised.


Assuntos
Doença de Alzheimer , DNA/genética , Chumbo/efeitos adversos , Doenças do Sistema Nervoso/induzido quimicamente , Animais , Metilação de DNA/efeitos dos fármacos , Epigênese Genética , Inibidores de Histona Desacetilases , Doenças do Sistema Nervoso/genética , Doenças Neurodegenerativas/genética
7.
PLoS One ; 15(7): e0235481, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32614880

RESUMO

In general, beef cattle long-distance transportation from cow-calf operations to feedlots or from feedlots to abattoirs is a common situation in the beef industry. The aim of this study was to determine the effect of rumen-protected methionine (RPM) supplementation on a proposed gene network for muscle fatigue, creatine synthesis (CKM), and reactive oxygen species (ROS) metabolism after a transportation simulation in a test track. Angus × Simmental heifers (n = 18) were stratified by body weight (408 ± 64 kg; BW) and randomly assigned to dietary treatments: 1) control diet (CTRL) or 2) control diet + 8 gr/hd/day of top-dressed rumen-protected methionine (RPM). After an adaptation period to Calan gates, animals received the mentioned dietary treatment consisting of Bermuda hay ad libitum and a soy hulls and corn gluten feed based supplement. After 45 days of supplementation, animals were loaded onto a trailer and transported for 22 hours (long-term transportation). Longissimus muscle biopsies, BW and blood samples were obtained on day 0 (Baseline), 43 (Pre-transport; PRET), and 46 (Post-transport; POST). Heifers' average daily gain did not differ between baseline and PRET. Control heifer's shrink was 10% of BW while RPM heifers shrink was 8%. Serum cortisol decreased, and glucose and creatine kinase levels increased after transportation, but no differences were observed between treatments. Messenger RNA was extracted from skeletal muscle tissue and gene expression analysis was performed by RT-qPCR. Results showed that AHCY and DNMT3A (DNA methylation), SSPN (Sarcoglycan complex), and SOD2 (Oxidative Stress-ROS) were upregulated in CTRL between baseline and PRET and, decreased between pre and POST while they remained constant for RPM. Furthermore, CKM was not affected by treatments. In conclusion, RPM supplementation may affect ROS production and enhance DNA hypermethylation, after a long-term transportation.


Assuntos
Suplementos Nutricionais , Metionina/farmacologia , Fadiga Muscular/efeitos dos fármacos , Nutrigenômica/métodos , Transportes/métodos , Animais , Bovinos , Creatina/metabolismo , Metilação de DNA/efeitos dos fármacos , Feminino , Espécies Reativas de Oxigênio/metabolismo , Estresse Fisiológico
8.
Mol Immunol ; 125: 172-177, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32707536

RESUMO

Recent studies suggest that microbiome derived 3(3,4-dihydroxy-phenyl) propionic acid (DHCA) attenuates IL-6 cytokine production through downregulation of the epigenetic modifier DNA Methyltransferase 1 (DNMT1) expression and inhibition of DNA methylation at the 5'-C-phosphate-G-3' (CpG)-rich IL6 sequence introns 1 and 3 in a mouse model of depression. In this study, we extended the investigation of DHCA epigenetic mechanisms in IL-6 expression in human peripheral blood mononuclear cells (PBMC). Using Lucia Luciferase reporter gene system we identified CpG-rich sequences in which of methylation is influenced by DHCA similar to what observed in response to treatment with the DNA methylation inhibitor 5-aza-2'-deoxycytidine. Correlation study showed that DNA methylation at select CpG motifs in the IL-6 promoter correlates with IL-6 gene expression. Our study suggests that DHCA is effective in reducing IL-6 expression in human PBMCs, in part, by regulation of methylation in the IL-6 promoter region.


Assuntos
Regulação da Expressão Gênica/efeitos dos fármacos , Monócitos/efeitos dos fármacos , Propionatos/farmacologia , Células Cultivadas , Metilação de DNA/efeitos dos fármacos , Metilação de DNA/genética , Regulação para Baixo , Regulação da Expressão Gênica/genética , Humanos , Interleucina-6/biossíntese , Microbiota , Neuroimunomodulação/efeitos dos fármacos , Neuroimunomodulação/genética
9.
PLoS One ; 15(7): e0235556, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32614916

RESUMO

To gain a better insight into the selenium nanoparticle (nSe) benefits/toxicity, this experiment was carried out to address the behavior of bitter melon seedlings to nSe (0, 1, 4, 10, 30, and 50 mgL-1) or bulk form (selenate). Low doses of nSe increased biomass accumulation, while concentrations of 10 mgL-1 and above were associated with stem bending, impaired root meristem, and severe toxicity. Responses to nSe were distinct from that of bulk in that the nano-type exhibited a higher efficiency to stimulate growth and organogenesis than the bulk. The bulk form displayed higher phytotoxicity than the nano-type counterpart. According to the MSAP-based analysis, nSe mediated substantial variation in DNA cytosine methylation, reflecting the epigenetic modification. By increasing the concentration of nSe, the expression of the WRKY1 transcription factor linearly up-regulated (mean = 7.9-fold). Transcriptions of phenylalanine ammonia-lyase (PAL) and 4-Coumarate: CoA-ligase (4CL) genes were also induced. The nSe treatments at low concentrations enhanced the activity of leaf nitrate reductase (mean = 52%) in contrast with the treatment at toxic concentrations. The toxic concentration of nSe increased leaf proline concentration by 80%. The nSe supplement also stimulated the activities of peroxidase (mean = 35%) and catalase (mean = 10%) enzymes. The nSe-treated seedlings exhibited higher PAL activity (mean = 39%) and soluble phenols (mean = 50%). The nSe toxicity was associated with a disrupted differentiation of xylem conducting tissue. The callus formation and performance of the explants originated from the nSe-treated seedlings had a different trend than that of the control. This experiment provides new insights into the nSe-associated advantage/ cytotoxicity and further highlights the necessity of designing convincing studies to introduce novel methods for plant cell/tissue cultures and agriculture.


Assuntos
Metilação de DNA/efeitos dos fármacos , Epigênese Genética/efeitos dos fármacos , Momordica charantia/metabolismo , Nanopartículas/toxicidade , Selênio/química , Citosina/metabolismo , Momordica charantia/efeitos dos fármacos , Momordica charantia/crescimento & desenvolvimento , Nanopartículas/química , Nitrato Redutase/genética , Nitrato Redutase/metabolismo , Fenóis/metabolismo , Fenilalanina Amônia-Liase/genética , Fenilalanina Amônia-Liase/metabolismo , Folhas de Planta/enzimologia , Folhas de Planta/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Raízes de Plantas/anatomia & histologia , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/metabolismo , Prolina/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Regulação para Cima/efeitos dos fármacos
10.
PLoS One ; 15(6): e0234641, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32574164

RESUMO

Chondrocytes, comparable to many cells from the connective tissue, dedifferentiate and end up in a similar fibroblastoid cell type, marked by the loss of the specific expression pattern. Here, chondrocytes isolated from osteoarthritic (OA) patients were investigated. The OA chondrocytes used in this work were not affected by the loss of specific gene expression upon cell culture. The mRNA levels of known cartilage markers, such as SOX5, SOX6, SOX9, aggrecan and proteoglycan 4, remained unchanged. Since chondrocytes from OA and healthy tissue show different DNA methylation patterns, the underlying mechanisms of cartilage marker maintenance were investigated with a focus on the epigenetic modification by DNA methylation. The treatment of dedifferentiated chondrocytes with the DNA methyltransferase inhibitor 5-aza-2´-deoxycytidine (5-aza-dC) displayed no considerable impact on the maintenance of marker gene expression observed in the dedifferentiated state, while the chondrogenic differentiation capacity was compromised. On the other hand, the pre-cultivation with 5-aza-dC improved the osteogenesis and adipogenesis of OA chondrocytes. Contradictory to these effects, the DNA methylation levels were not reduced after treatment for four weeks with 1 µM 5-aza-dC. In conclusion, 5-aza-dC affects the differentiation capacity of OA chondrocytes, while the global DNA methylation level remains stable. Furthermore, dedifferentiated chondrocytes isolated from late-stage OA patients represent a reliable cell source for in vitro studies and disease models without the need for additional alterations.


Assuntos
Condrócitos/patologia , Decitabina/farmacologia , Osteoartrite/patologia , Adipogenia/efeitos dos fármacos , Adipogenia/genética , Biomarcadores/metabolismo , Cartilagem Articular/efeitos dos fármacos , Cartilagem Articular/metabolismo , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Forma Celular/efeitos dos fármacos , Forma Celular/genética , Células Cultivadas , Condrócitos/efeitos dos fármacos , Condrócitos/metabolismo , Colágeno Tipo II/metabolismo , DNA (Citosina-5-)-Metiltransferases/metabolismo , Metilação de DNA/efeitos dos fármacos , Metilação de DNA/genética , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Osteoartrite/genética , Osteogênese/efeitos dos fármacos , Osteogênese/genética
11.
Ecotoxicol Environ Saf ; 201: 110802, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32531573

RESUMO

Extended exposure to inorganic arsenic through contaminated drinking water has been linked with increased incidence of diabetes mellitus. The most common exposure occurs through the consumption of contaminated drinking water mainly through geogenic sources of inorganic arsenic. Epigenetic modifications are important mechanisms through which environmental pollutants could exert their toxic effects. Bisulfite sequencing polymerase chain reaction method followed by Sanger sequencing was performed for DNA methylation analysis. Our results showed that sodium arsenite treatment significantly reduced insulin secretion in pancreatic islets. It was revealed that the methylation of glucose transporter 2 (Glut2) gene was changed at two cytosine-phosphate-guanine (CpG) sites (-1743, -1734) in the promoter region of the sodium arsenite-treated group comparing to the control. No changes were observed in the methylation status of peroxisome proliferator-activated receptor-gamma (PPARγ), pancreatic and duodenal homeobox 1 (Pdx1) and insulin 2 (Ins2) CpG sites in the targeted regions. Measuring the gene expression level showed increase in Glut2 expression, while the expression of insulin (INS) and Pdx1 were significantly affected by sodium arsenite treatment. This study revealed that exposure to sodium arsenite changed the DNA methylation pattern of Glut2, a key transporter of glucose entry into the pancreatic beta cells (ß-cells). Our data suggested possible epigenetic-mediated toxicity mechanism for arsenite-induced ß-cells dysfunction. Further studies are needed to dissect the precise epigenetic modulatory activity of sodium arsenite that affect the biogenesis of insulin.


Assuntos
Arsenitos/toxicidade , Metilação de DNA/efeitos dos fármacos , Transportador de Glucose Tipo 2/genética , Insulina/metabolismo , Ilhotas Pancreáticas/efeitos dos fármacos , Compostos de Sódio/toxicidade , Poluentes Químicos da Água/toxicidade , Animais , Epigênese Genética/efeitos dos fármacos , Proteínas de Homeodomínio/genética , Técnicas In Vitro , Insulina/genética , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/metabolismo , Ilhotas Pancreáticas/metabolismo , Masculino , Regiões Promotoras Genéticas , Ratos , Ratos Wistar , Transativadores/genética
12.
PLoS One ; 15(6): e0233784, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32492024

RESUMO

Recent advances in somatic cell nuclear transfer (SCNT) in canines facilitate the production of canine transgenic models. Owing to the importance of stable and strong promoter activity in transgenic animals, we tested human elongation factor 1α (hEF1α) and cytomegalovirus (CMV) promoter sequences in SCNT transgenic dogs. After transfection, transgenic donor fibroblasts with the hEF1α-enhanced green fluorescence protein (EGFP) transgene were successfully isolated using fluorescence-activated cell sorting (FACS). We obtained four puppies, after SCNT, and identified three puppies as being transgenic using PCR analysis. Unexpectedly, EGFP regulated by hEF1α promoter was not observed at the organismal and cellular levels in these transgenic dogs. EGFP expression was rescued by the inhibition of DNA methyltransferases, implying that the hEF1α promoter is silenced by DNA methylation. Next, donor cells with CMV-EGFP transgene were successfully established and SCNT was performed. Three puppies of six born puppies were confirmed to be transgenic. Unlike hEF1α-regulated EGFP, CMV-regulated EGFP was strongly detectable at both the organismal and cellular levels in all transgenic dogs, even after 19 months. In conclusion, our study suggests that the CMV promoter is more suitable, than the hEF1α promoter, for stable transgene expression in SCNT-derived transgenic canine model.


Assuntos
Clonagem de Organismos/veterinária , Citomegalovirus/genética , Técnicas de Transferência Nuclear/veterinária , Fator 1 de Elongação de Peptídeos/genética , Regiões Promotoras Genéticas/genética , Ativação Transcricional/genética , Animais , Animais Geneticamente Modificados , Azacitidina/farmacologia , Células Cultivadas , DNA (Citosina-5-)-Metiltransferases/antagonistas & inibidores , Metilação de DNA/efeitos dos fármacos , Cães , Transferência Embrionária/veterinária , Feminino , Fibroblastos/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas de Fluorescência Verde/genética , Humanos , Gravidez , Transfecção , Transgenes
13.
Prostate ; 80(12): 977-985, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32542727

RESUMO

BACKGROUND: Recently, resveratrol (Res) has been suggested to suppress the migration and invasion of prostate cancer (PCa). In the present study, we aimed to investigate the effects of Res on genomic DNA methylation, as well as the migration and invasion of PCa cells. METHODS: The suppression by Res of the growth of PCa cells was verified through a cytotoxicity assay. In addition, the effects of Res on 5-methylcytosine (5mC), 5-hydroxymethylcytosine (5hmC), and ten-eleven translocation 1 (TET1) levels were assessed, and the cell migration and invasion were also determined. The expressions of TET1, tissue inhibitor of metalloproteinases (TIMP) 2, TIMP3, MMP2, and MMP9 were detected through Western blot analysis. Afterward, TET1 was silenced using lentiviral short hairpin RNA to examine the effect of TET1 on the Res-triggered inhibition of migration and invasion of PCa cells. RESULTS: Our results showed that Res upregulated the 5hmC and TET1 levels and downregulated the 5mC level. Moreover, Res also inhibited the migration and invasion of PCa cells, promoted the demethylation of TIMP2 and TIMP3 to upregulate their expressions, and suppressed the expressions of MMP2 and MMP9. The silencing of TET1 in the presence of Res showed that Res could exert its effect through TET1. CONCLUSIONS: Our findings indicated that Res inhibited the migration and invasion of PCa cells via the TET1/TIMP2/TIMP3 pathway, which might potentially serve as a target for the treatment of PCa.


Assuntos
Oxigenases de Função Mista/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Resveratrol/farmacologia , Inibidor Tecidual de Metaloproteinase-2/metabolismo , Inibidor Tecidual de Metaloproteinase-3/metabolismo , 5-Metilcitosina/análogos & derivados , 5-Metilcitosina/metabolismo , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Metilação de DNA/efeitos dos fármacos , Células HEK293 , Humanos , Masculino , Oxigenases de Função Mista/biossíntese , Oxigenases de Função Mista/genética , Invasividade Neoplásica , Células PC-3 , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Proteínas Proto-Oncogênicas/biossíntese , Proteínas Proto-Oncogênicas/genética , Resveratrol/farmacocinética , Inibidor Tecidual de Metaloproteinase-2/biossíntese , Inibidor Tecidual de Metaloproteinase-2/genética , Inibidor Tecidual de Metaloproteinase-3/biossíntese , Inibidor Tecidual de Metaloproteinase-3/genética , Regulação para Cima
14.
Toxicology ; 441: 152507, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32512035

RESUMO

Vorinostat was approved as the first histone deacetylase inhibitor for the management of cutaneous T cell lymphoma. However, it's in vivo genetic and epigenetic effects on non-cancerous cells remain poorly understood. As genetic and epigenetic changes play a critical role in the pathogenesis of carcinogenesis, we investigated whether vorinostat induces genetic and epigenetic alterations in mouse bone marrow cells. Bone marrow cells were isolated 24 h following the last oral administration of vorinostat at the doses of 25, 50, or 100 mg/kg/day for five days (approximately equal to the recommended human doses). The cells were then used to assess clastogenicity and aneugenicity by the micronucleus test complemented by fluorescence in situ hybridization assay; DNA strand breaks, oxidative DNA strand breaks, and DNA methylation by the modified comet assay; apoptosis by annexin V/PI staining analysis and the occurrence of the hypodiploid DNA content; and DNA damage/repair gene expression by polymerase chain reaction (PCR) Array. The expression of the mRNA transcripts were also confirmed by real-time PCR and western blot analysis. Vorinostat caused structural chromosomal damage, numerical chromosomal abnormalities, DNA strand breaks, oxidative DNA strand breaks, DNA hypomethylation, and programed cell death in a dose-dependent manner. Furthermore, the expression of numerous genes implicated in DNA damage/repair were altered after vorinostat treatment. Accordingly, the genetic/epigenetic mechanism(s) of action of vorinostat may play a role in its carcinogenicity and support the continued study and development of new compounds with lower toxicity.


Assuntos
Antineoplásicos/toxicidade , Células da Medula Óssea/efeitos dos fármacos , Vorinostat/toxicidade , Animais , Antineoplásicos/administração & dosagem , Apoptose/efeitos dos fármacos , Aberrações Cromossômicas/induzido quimicamente , Ensaio Cometa , Metilação de DNA/efeitos dos fármacos , Reparo do DNA/efeitos dos fármacos , Relação Dose-Resposta a Droga , Regulação para Baixo , Masculino , Camundongos , Estresse Oxidativo/efeitos dos fármacos , Vorinostat/administração & dosagem
15.
Nat Commun ; 11(1): 2316, 2020 05 08.
Artigo em Inglês | MEDLINE | ID: mdl-32385268

RESUMO

Our early-life environment has a profound influence on developing organs that impacts metabolic function and determines disease susceptibility across the life-course. Using a rat model for exposure to an endocrine disrupting chemical (EDC), we show that early-life chemical exposure causes metabolic dysfunction in adulthood and reprograms histone marks in the developing liver to accelerate acquisition of an adult epigenomic signature. This epigenomic reprogramming persists long after the initial exposure, but many reprogrammed genes remain transcriptionally silent with their impact on metabolism not revealed until a later life exposure to a Western-style diet. Diet-dependent metabolic disruption was largely driven by reprogramming of the Early Growth Response 1 (EGR1) transcriptome and production of metabolites in pathways linked to cholesterol, lipid and one-carbon metabolism. These findings demonstrate the importance of epigenome:environment interactions, which early in life accelerate epigenomic aging, and later in adulthood unlock metabolically restricted epigenetic reprogramming to drive metabolic dysfunction.


Assuntos
Epigenoma/genética , Animais , Metilação de DNA/efeitos dos fármacos , Metilação de DNA/genética , Proteína 1 de Resposta de Crescimento Precoce/genética , Disruptores Endócrinos/toxicidade , Epigênese Genética/efeitos dos fármacos , Epigênese Genética/genética , Epigenômica/métodos , Feminino , Interação Gene-Ambiente , Estudo de Associação Genômica Ampla , Masculino , Ratos
16.
PLoS Genet ; 16(5): e1008823, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32453789

RESUMO

The development of type 2 diabetes mellitus (T2DM) depends on interactions between genetic and environmental factors, and a better understanding of gene-diet interactions in T2DM will be useful for disease prediction and prevention. Ascorbic acid has been proposed to reduce the risk of T2DM. However, the links between ascorbic acid and metabolic consequences are not fully understood. Here, we report that glucose transporter 10 (GLUT10) maintains intracellular levels of ascorbic acid to promote adipogenesis, white adipose tissue (WAT) development and protect mice from high-fat diet (HFD)-induced metabolic dysregulation. We found genetic polymorphisms in SLC2A10 locus are suggestively associated with a T2DM intermediate phenotype in non-diabetic Han Taiwanese. Additionally, mice carrying an orthologous human Glut10G128E variant (Glut10G128E mice) with compromised GLUT10 function have reduced adipogenesis, reduced WAT development and increased susceptibility to HFD-induced metabolic dysregulation. We further demonstrate that GLUT10 is highly expressed in preadipocytes, where it regulates intracellular ascorbic acid levels and adipogenesis. In this context, GLUT10 increases ascorbic acid-dependent DNA demethylation and the expression of key adipogenic genes, Cebpa and Pparg. Together, our data show GLUT10 regulates adipogenesis via ascorbic acid-dependent DNA demethylation to benefit proper WAT development and protect mice against HFD-induced metabolic dysregulation. Our findings suggest that SLC2A10 may be an important HFD-associated susceptibility locus for T2DM.


Assuntos
Tecido Adiposo Branco/metabolismo , Ácido Ascórbico/metabolismo , Metilação de DNA , Diabetes Mellitus Tipo 2/genética , Dieta Hiperlipídica/efeitos adversos , Proteínas Facilitadoras de Transporte de Glucose/genética , Células 3T3-L1 , Adipogenia , Adulto , Idoso , Animais , Proteínas Estimuladoras de Ligação a CCAAT/genética , Metilação de DNA/efeitos dos fármacos , Diabetes Mellitus Tipo 2/metabolismo , Modelos Animais de Doenças , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas Facilitadoras de Transporte de Glucose/metabolismo , Hemoglobina A Glicada/metabolismo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Mutação , PPAR gama/genética
18.
Aquat Toxicol ; 224: 105479, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32417751

RESUMO

Lead (Pb) is a toxic environmental pollutant that is frequently present in effluents from urban, mining, and industrial sources. The combinatorial effects of heavy metal exposure and temperature in aquatic organisms have received considerable attention as heat stress occurs simultaneously in conjunction with several contaminants in a natural environment. In this study, we examined the potential effects of Pb exposure in conditions of thermal stress (34 °C) in zebrafish (Danio rerio) embryos. Thermal stress at 34 °C induced a dramatic decrease in the survival rate, although exposure to Pb at 26 °C decreased the survival rate of the embryos. Malformations, such as the curved body shape, were increased in response to exposure to a combination of Pb and heat stress. The combination of Pb and heat stress also caused a decrease in the heart rate. Moreover, Pb and high-temperature exposure induced the upregulation of SOD, CAT, TNF-α, IL-1ß, p53, and BAX transcripts, and downregulation of Dnmt1 and Dnmt3b transcripts. Thermal stress enhanced transcriptional responses of eight indicator genes following Pb toxicity. The induction of cell death in response to combined exposures was also confirmed in the body of zebrafish by fluorescence intensity image analysis. These data indicated that thermal stress enhanced the poisonous effects of Pb exposure on antioxidant defense, inflammation, and apoptotic mechanisms. Transcriptional inhibition of DNA methylation-related genes might serve as a crucial factor contributing to the possibility of epigenetic adaptation by altering combined stress. We suggest that a careful evaluation of the potential effects of climate change (especially temperature) should be considered when investigating the toxic levels of metal pollution, such as Pb, in an aquatic environment.


Assuntos
Apoptose/efeitos dos fármacos , Metilação de DNA/efeitos dos fármacos , Embrião não Mamífero/efeitos dos fármacos , Resposta ao Choque Térmico/efeitos dos fármacos , Chumbo/toxicidade , Estresse Oxidativo/efeitos dos fármacos , Poluentes Químicos da Água/toxicidade , Peixe-Zebra , Animais , Antioxidantes/metabolismo , Apoptose/genética , Embrião não Mamífero/imunologia , Embrião não Mamífero/patologia , Resposta ao Choque Térmico/genética , Resposta ao Choque Térmico/imunologia , Temperatura Alta , Inflamação , Regulação para Cima , Proteínas de Peixe-Zebra/genética
19.
Proc Natl Acad Sci U S A ; 117(23): 13033-13043, 2020 06 09.
Artigo em Inglês | MEDLINE | ID: mdl-32461362

RESUMO

Trichomonas vaginalis is a common sexually transmitted parasite that colonizes the human urogenital tract causing infections that range from asymptomatic to highly inflammatory. Recent works have highlighted the importance of histone modifications in the regulation of transcription and parasite pathogenesis. However, the nature of DNA methylation in the parasite remains unexplored. Using a combination of immunological techniques and ultrahigh-performance liquid chromatography (UHPLC), we analyzed the abundance of DNA methylation in strains with differential pathogenicity demonstrating that N6-methyladenine (6mA), and not 5-methylcytosine (5mC), is the main DNA methylation mark in T. vaginalis Genome-wide distribution of 6mA reveals that this mark is enriched at intergenic regions, with a preference for certain superfamilies of DNA transposable elements. We show that 6mA in T. vaginalis is associated with silencing when present on genes. Interestingly, bioinformatics analysis revealed the presence of transcriptionally active or repressive intervals flanked by 6mA-enriched regions, and results from chromatin conformation capture (3C) experiments suggest these 6mA flanked regions are in close spatial proximity. These associations were disrupted when parasites were treated with the demethylation activator ascorbic acid. This finding revealed a role for 6mA in modulating three-dimensional (3D) chromatin structure and gene expression in this divergent member of the Excavata.


Assuntos
Adenina/metabolismo , Cromatina/química , Metilação de DNA/genética , Trichomonas vaginalis/genética , Ácido Ascórbico/farmacologia , Técnicas de Cultura de Células , Cromatina/genética , Cromatina/metabolismo , Biologia Computacional , Metilação de DNA/efeitos dos fármacos , Elementos de DNA Transponíveis/genética , Regulação da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Conformação Molecular , Análise de Sequência de DNA
20.
Am J Physiol Cell Physiol ; 319(2): C268-C276, 2020 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-32459505

RESUMO

DNA methylation, a critical epigenetic mechanism, plays an important role in governing gene expressions during biological processes such as aging, which is well known to be accelerated in hyperglycemia (diabetes). In the present study, we investigated the effects of glucose on whole genome DNA methylation in small [human retinal microvascular endothelial cells (HRECs)] and large [human umbilical vein endothelial cells (HUVECs)] vessel endothelial cell (EC) lines exposed to basal or high glucose-containing media for variable lengths of time. Using the Infinium EPIC array, we obtained 773,133 CpG sites (probes) for analysis. Unsupervised clustering of the top 5% probes identified four distinct clusters within EC groups, with significant methylation differences attributed to EC types and the duration of cell culture rather than glucose stimuli alone. When comparing the ECs incubated for 2 days versus 7 days, hierarchical clustering analyses [methylation change >10% and false discovery rate (FDR) <0.05] identified 17,354 and 128 differentially methylated CpGs for HUVECs and HRECs, respectively. Predominant DNA hypermethylation was associated with the length of culture and was enriched for gene enhancer elements and regions surrounding CpG shores and shelves. We identified 88 differentially methylated regions (DMRs) for HUVECs and 8 DMRs for HRECs (all FDR <0.05). Pathway enrichment analyses of DMRs highlighted involvement of regulators of embryonic development (i.e., HOX genes) and cellular differentiation [transforming growth factor-ß (TGF-ß) family members]. Collectively, our findings suggest that DNA methylation is a complex process that involves tightly coordinated, cell-specific mechanisms. Such changes in methylation overlap genes critical for cellular differentiation and embryonic development.


Assuntos
Envelhecimento/genética , Ilhas de CpG/genética , Metilação de DNA/genética , Células Endoteliais/metabolismo , Envelhecimento/patologia , Ilhas de CpG/efeitos dos fármacos , DNA/genética , Metilação de DNA/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Epigênese Genética , Regulação da Expressão Gênica/efeitos dos fármacos , Genes Homeobox/genética , Genoma Humano/genética , Glucose/farmacologia , Células Endoteliais da Veia Umbilical Humana , Humanos , Regiões Promotoras Genéticas/efeitos dos fármacos
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