RESUMO
The tumour suppressor p16/CDKN2A and the metabolic gene, methyl-thio-adenosine phosphorylase (MTAP), are frequently co-deleted in some of the most aggressive and currently untreatable cancers. Cells with MTAP deletion are vulnerable to inhibition of the metabolic enzyme, methionine-adenosyl transferase 2A (MAT2A), and the protein arginine methyl transferase (PRMT5). This synthetic lethality has paved the way for the rapid development of drugs targeting the MAT2A/PRMT5 axis. MAT2A and its liver- and pancreas-specific isoform, MAT1A, generate the universal methyl donor S-adenosylmethionine (SAM) from ATP and methionine. Given the pleiotropic role SAM plays in methylation of diverse substrates, characterising the extent of SAM depletion and downstream perturbations following MAT2A/MAT1A inhibition (MATi) is critical for safety assessment. We have assessed in vivo target engagement and the resultant systemic phenotype using multi-omic tools to characterise response to a MAT2A inhibitor (AZ'9567). We observed significant SAM depletion and extensive methionine accumulation in the plasma, liver, brain and heart of treated rats, providing the first assessment of both global SAM depletion and evidence of hepatic MAT1A target engagement. An integrative analysis of multi-omic data from liver tissue identified broad perturbations in pathways covering one-carbon metabolism, trans-sulfuration and lipid metabolism. We infer that these pathway-wide perturbations represent adaptive responses to SAM depletion and confer a risk of oxidative stress, hepatic steatosis and an associated disturbance in plasma and cellular lipid homeostasis. The alterations also explain the dramatic increase in plasma and tissue methionine, which could be used as a safety and PD biomarker going forward to the clinic.
Assuntos
Metionina Adenosiltransferase , S-Adenosilmetionina , Animais , Metionina Adenosiltransferase/genética , Metionina Adenosiltransferase/metabolismo , S-Adenosilmetionina/metabolismo , Masculino , Fígado/efeitos dos fármacos , Fígado/metabolismo , Ratos , Metionina/metabolismo , Ratos Sprague-Dawley , Purina-Núcleosídeo Fosforilase/metabolismo , Purina-Núcleosídeo Fosforilase/genética , Proteína-Arginina N-Metiltransferases/genética , Proteína-Arginina N-Metiltransferases/metabolismo , Proteína-Arginina N-Metiltransferases/antagonistas & inibidores , MultiômicaRESUMO
Untargeted metabolomics is an established approach in toxicology for characterising endogenous metabolic responses to xenobiotic exposure. Detecting the xenobiotic and its biotransformation products as part of the metabolomics analysis provides an opportunity to simultaneously gain deep insights into its fate and metabolism, and to associate the internal relative dose directly with endogenous metabolic responses. This integration of untargeted exposure and response measurements into a single assay has yet to be fully demonstrated. Here we assemble a workflow to discover and analyse pharmaceutical-related measurements from routine untargeted UHPLC-MS metabolomics datasets, derived from in vivo (rat plasma and cardiac tissue, and human plasma) and in vitro (human cardiomyocytes) studies that were principally designed to investigate endogenous metabolic responses to drug exposure. Our findings clearly demonstrate how untargeted metabolomics can discover extensive biotransformation maps, temporally-changing relative systemic exposure, and direct associations of endogenous biochemical responses to the internal dose.
Assuntos
Metabolômica , Xenobióticos , Humanos , Ratos , Animais , Metaboloma , BiotransformaçãoRESUMO
PURPOSE: We hypothesized that inhibition and trapping of PARP1 alone would be sufficient to achieve antitumor activity. In particular, we aimed to achieve selectivity over PARP2, which has been shown to play a role in the survival of hematopoietic/stem progenitor cells in animal models. We developed AZD5305 with the aim of achieving improved clinical efficacy and wider therapeutic window. This next-generation PARP inhibitor (PARPi) could provide a paradigm shift in clinical outcomes achieved by first-generation PARPi, particularly in combination. EXPERIMENTAL DESIGN: AZD5305 was tested in vitro for PARylation inhibition, PARP-DNA trapping, and antiproliferative abilities. In vivo efficacy was determined in mouse xenograft and PDX models. The potential for hematologic toxicity was evaluated in rat models, as monotherapy and combination. RESULTS: AZD5305 is a highly potent and selective inhibitor of PARP1 with 500-fold selectivity for PARP1 over PARP2. AZD5305 inhibits growth in cells with deficiencies in DNA repair, with minimal/no effects in other cells. Unlike first-generation PARPi, AZD5305 has minimal effects on hematologic parameters in a rat pre-clinical model at predicted clinically efficacious exposures. Animal models treated with AZD5305 at doses ≥0.1 mg/kg once daily achieved greater depth of tumor regression compared to olaparib 100 mg/kg once daily, and longer duration of response. CONCLUSIONS: AZD5305 potently and selectively inhibits PARP1 resulting in excellent antiproliferative activity and unprecedented selectivity for DNA repair deficient versus proficient cells. These data confirm the hypothesis that targeting only PARP1 can retain the therapeutic benefit of nonselective PARPi, while reducing potential for hematotoxicity. AZD5305 is currently in phase I trials (NCT04644068).
Assuntos
Antineoplásicos , Inibidores de Poli(ADP-Ribose) Polimerases , Humanos , Camundongos , Ratos , Animais , Linhagem Celular Tumoral , Ensaios Antitumorais Modelo de Xenoenxerto , Ftalazinas/farmacologia , Poli(ADP-Ribose) Polimerase-1 , Antineoplásicos/farmacologia , Reparo do DNARESUMO
The receptor tyrosine kinase, MERTK, plays an essential role in homeostasis of the retina via efferocytosis of shed outer nuclear segments of photoreceptors. The Royal College of Surgeons rat model of retinal degeneration has been linked to loss-of-function of MERTK, and together with the MERTK knock-out mouse, phenocopy retinitis pigmentosa in humans with MERTK mutations. Given recent efforts and interest in MERTK as a potential immuno-oncology target, development of a strategy to assess ocular safety at an early pre-clinical stage is critical. We have applied a state-of-the-art, multi-modal imaging platform to assess the in vivo effects of pharmacological inhibition of MERTK in mice. This involved the application of mass spectrometry imaging (MSI) to characterize the ocular spatial distribution of our highly selective MERTK inhibitor; AZ14145845, together with histopathology and transmission electron microscopy to characterize pathological and ultra-structural change in response to MERTK inhibition. In addition, we assessed the utility of a human retinal in vitro cell model to identify perturbation of phagocytosis post MERTK inhibition. We identified high localized total compound concentrations in the retinal pigment epithelium (RPE) and retinal lesions following 28 days of treatment with AZ14145845. These lesions were present in 4 of 8 treated animals, and were characterized by a thinning of the outer nuclear layer, loss of photoreceptors (PR) and accumulation of photoreceptor outer segments at the interface of the RPE and PRs. Furthermore, the lesions were very similar to that shown in the RCS rat and MERTK knock-out mouse, suggesting a MERTK-induced mechanism of PR cell death. This was further supported by the observation of reduced phagocytosis in the human retinal cell model following treatment with AZ14145845. Our study provides a viable, translational strategy to investigate the pre-clinical toxicity of MERTK inhibitors but is equally transferrable to novel chemotypes.
Assuntos
Fagocitose/efeitos dos fármacos , Células Fotorreceptoras de Vertebrados/efeitos dos fármacos , c-Mer Tirosina Quinase/antagonistas & inibidores , Animais , Linhagem Celular , Feminino , Humanos , Masculino , Espectrometria de Massas , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Imagem Multimodal , Células Fotorreceptoras de Vertebrados/patologia , Ratos , Ratos Long-Evans , Ratos Wistar , Degeneração Retiniana/induzido quimicamente , Epitélio Pigmentado da Retina/metabolismo , Distribuição Tecidual , c-Mer Tirosina Quinase/genéticaRESUMO
OBJECTIVE: This study validated the newly adapted electronic SNAPPS (eSNAPPS) against the original paper SNAPPS. Subsequently, the study estimated the prevalence of PFP in running participants and spectators attending three mass-participant running events in the United Kingdom by using the eSNAPPS tool. DESIGN: This study had two parts. Firstly, a validation of the original paper version of the SNAPPS tool. Secondly, if validation was achieved, eSNAPPS was used in a prevalence study. PARTICIPANTS: A convenience sample of running participants and spectators aged 18-40 years attending the mass participation running events. MAIN OUTCOME MEASURE: The 12-month prevalence of PFP. RESULTS: eSNAPPS was valid in identifying those with PFP (ICC 0.99 for Overall agreement, p < 0.0001). In the prevalence study, a total of 1080 running participants and spectators completed the eSNAPPS. The overall prevalence of PFP was 17.4% (95%CI: 15.2%, 19.8%); 20.5% of males (16.5, 24.9) and 15.7% of females (13.1, 18.7) had PFP. Prevalence was 17.4% (15.2, 19.8) in spectators and 16.7% in running participants (14.5, 19.0). CONCLUSION: The overall PFP prevalence in this study was slightly smaller than those previously reported in the literature. Findings also show that there were similar prevalence estimates in spectators and running participants.
Assuntos
Inquéritos Epidemiológicos/métodos , Internet , Síndrome da Dor Patelofemoral/epidemiologia , Adolescente , Adulto , Fenômenos Biomecânicos , Feminino , Humanos , Masculino , Síndrome da Dor Patelofemoral/diagnóstico , Síndrome da Dor Patelofemoral/etiologia , Prevalência , Corrida/lesões , Reino Unido/epidemiologia , Adulto JovemRESUMO
Imaging mass cytometry (IMC) offers the opportunity to image metal- and heavy halogen-containing xenobiotics in a highly multiplexed experiment with other immunochemistry-based reagents to distinguish uptake into different tissue structures or cell types. However, in practice, many xenobiotics are not amenable to this analysis, as any compound which is not bound to the tissue matrix will delocalize during aqueous sample-processing steps required for IMC analysis. Here, we present a strategy to perform IMC experiments on a water-soluble polysarcosine-modified dendrimer drug-delivery system (S-Dends). This strategy involves two consecutive imaging acquisitions on the same tissue section using the same instrumental platform, an initial laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MSI) experiment followed by tissue staining and a standard IMC experiment. We demonstrated that settings can be found for the initial ablation step that leave sufficient residual tissue for subsequent antibody staining and visualization. This workflow results in lateral resolution for the S-Dends of 2 µm followed by imaging of metal-tagged antibodies at 1 µm.
Assuntos
Citometria por Imagem , Água , Sistemas de Liberação de Medicamentos , Espectrometria de Massas , Coloração e RotulagemRESUMO
INTRODUCTION: Accurate assessment of muscle insulin sensitivity requires measurement of insulin concentration in interstitial fluid (ISF), but has proved difficult. We aimed to optimise measurement of ISF insulin concentrations in rat muscles in vivo using microdialysis. METHODS: Factorial experimental design experiments were performed in vitro to determine optimal conditions for insulin recovery with microdialysis probes. These conditions were tested in vivo, adjusted appropriately and used in lean and obese Zucker rats to compare ISF insulin concentrations basally and during hyperinsulinaemic-euglycaemic (HE) clamp. RESULTS: Optimal conditions in vivo were: a 100kDa microdialysis probe inserted in muscle, perfused with 1% BSA, 1.5mM glucose in 0.9% sodium chloride at 1µl/min. Samples were collected into siliconised glass microvials. As a reference for insulin, we established a protocol of inulin infusion, beginning at -80min and reaching equilibrium within 60min. HE clamp, beginning at 0min, increased ISF insulin concentration from 122±56 basally to 429±180pmol/l (P<0.05) in lean rats and from 643±165 to 1087±243pmol/l (P=0.07) in obese rats; ISF insulin concentrations were significantly higher throughout in obese rats. The difference between ISF and plasma insulin concentration (ISF:plasma ratio) was substantially higher in obese rats, but fell to similar values in obese and lean rats during HE clamp. DISCUSSION: Optimising insulin recovery with microdialysis allowed accurate measurement of basal ISF insulin in muscle of lean and obese Zucker rats and indicates insulin transport across capillaries is impaired in obese rats, basally and during hyperinsulinaemia.
Assuntos
Líquido Extracelular/química , Insulina/análise , Microdiálise/métodos , Músculo Esquelético/química , Animais , Capilares/metabolismo , Técnica Clamp de Glucose , Hiperinsulinismo/sangue , Hiperinsulinismo/metabolismo , Insulina/sangue , Resistência à Insulina , Inulina/farmacologia , Masculino , Ratos , Ratos Wistar , Ratos Zucker , Reprodutibilidade dos TestesRESUMO
Both fresh and processed foods make up vital parts of the food supply. Processed food contributes to both food security (ensuring that sufficient food is available) and nutrition security (ensuring that food quality meets human nutrient needs). This ASN scientific statement focuses on one aspect of processed foods: their nutritional impacts. Specifically, this scientific statement 1) provides an introduction to how processed foods contribute to the health of populations, 2) analyzes the contribution of processed foods to "nutrients to encourage" and "constituents to limit" in the American diet as recommended by the Dietary Guidelines for Americans, 3) identifies the responsibilities of various stakeholders in improving the American diet, and 4) reviews emerging technologies and the research needed for a better understanding of the role of processed foods in a healthy diet. Analyses of the NHANES 2003-2008 show that processed foods provide both nutrients to encourage and constituents to limit as specified in the 2010 Dietary Guidelines for Americans. Of the nutrients to encourage, processed foods contributed 55% of dietary fiber, 48% of calcium, 43% of potassium, 34% of vitamin D, 64% of iron, 65% of folate, and 46% of vitamin B-12. Of the constituents to limit, processed foods contributed 57% of energy, 52% of saturated fat, 75% of added sugars, and 57% of sodium. Diets are more likely to meet food guidance recommendations if nutrient-dense foods, either processed or not, are selected. Nutrition and food science professionals, the food industry, and other stakeholders can help to improve the diets of Americans by providing a nutritious food supply that is safe, enjoyable, affordable, and sustainable by communicating effectively and accurately with each other and by working together to improve the overall knowledge of consumers.
Assuntos
Dieta/efeitos adversos , Fast Foods/análise , Manipulação de Alimentos , Alimentos em Conserva/análise , Dieta/economia , Fast Foods/economia , Fast Foods/normas , Abastecimento de Alimentos/economia , Abastecimento de Alimentos/normas , Alimentos em Conserva/economia , Alimentos em Conserva/normas , Indústria de Processamento de Alimentos , Guias como Assunto , Promoção da Saúde , Humanos , Política Nutricional , Ciências da Nutrição , Valor Nutritivo , Cooperação do Paciente , Papel Profissional , Sociedades Científicas , Estados Unidos , Recursos HumanosRESUMO
Previous evidence suggests soy genistein may be protective against prostate cancer, but whether this protection involves an estrogen receptor (ER)-dependent mechanism is unknown. To test the hypothesis that phytoestrogens may act through ERα or ERß to play a protective role against prostate cancer, we bred transgenic mice lacking functional ERα or ERß with transgenic adenocarcinoma of mouse prostate (TRAMP) mice. Dietary genistein reduced the incidence of cancer in ER wild-type (WT)/transgenic adenocarcinoma of mouse prostate mice but not in ERα knockout (KO) or ERßKO mice. Cancer incidence was 70% in ERWT mice fed the control diet compared with 47% in ERWT mice fed low-dose genistein (300 mg/kg) and 32% on the high-dose genistein (750 mg/kg). Surprisingly, genistein only affected the well differentiated carcinoma (WDC) incidence but had no effect on poorly differentiated carcinoma (PDC). No dietary effects have been observed in either of the ERKO animals. We observed a very strong genotypic influence on PDC incidence, a protective effect in ERαKO (only 5% developed PDC), compared with 19% in the ERWT, and an increase in the incidence of PDC in ERßKO mice to 41%. Interestingly, immunohistochemical analysis showed ERα expression changing from nonnuclear in WDC to nuclear in PDC, with little change in ERß location or expression. In conclusion, genistein is able to inhibit WDC in the presence of both ERs, but the effect of estrogen signaling on PDC is dominant over any dietary treatment, suggesting that improved differential targeting of ERα vs. ERß would result in prevention of advanced prostate cancer.
Assuntos
Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/metabolismo , Neoplasias da Próstata/metabolismo , Animais , Modelos Animais de Doenças , Receptor alfa de Estrogênio/genética , Receptor beta de Estrogênio/genética , Genisteína/uso terapêutico , Imuno-Histoquímica , Masculino , Camundongos , Camundongos Knockout , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genéticaRESUMO
Colon cancer is the third most commonly diagnosed type of cancer in the United States. Lifestyle and dietary patterns influence colon cancer risk both positively and negatively. Among the dietary factors, several plant-derived compounds have been found to afford colon cancer protection. These compounds potentially influence all aspects of colonic cellular regulation and develop complex interrelationships with the colonic microbiome. Increasing understanding of the role of microorganisms in determining the colonic environment has led to awareness of this important interrelationship among dietary factors and the microbial population. Plant-derived polyphenols are active mediators of cellular events, target key carcinogenic pathways, and modulate colonic microbial populations. In turn, the colonic microorganisms metabolize dietary compounds and mediate cellular events. In addition, the role of estrogen receptors in colon cancer and the importance of dietary components that mediate estrogen receptor-ß are increasingly being discovered. Hence, dietary bioactive compounds and the intestinal microbiota create a complex milieu that directly affects the carcinogenic events of the colon. These relationships must be carefully characterized in future research to provide dietary recommendations that will reduce colon cancer risk.
Assuntos
Neoplasias do Colo/microbiologia , Neoplasias do Colo/prevenção & controle , Suplementos Nutricionais/análise , Microbiota , Compostos Fitoquímicos/administração & dosagem , Animais , Neoplasias do Colo/epidemiologia , Humanos , Microbiota/efeitos dos fármacos , Fatores de RiscoRESUMO
The incidence of inflammatory bowel diseases has increased during recent decades. Within the colon, the families of mucins (MUC) and trefoil factors (TFF) facilitate mucosal protection. Probiotic administration influences the intestinal MUC layer. Additionally, food components may affect gut microflora or have direct effects on the MUC barrier. Our objective was to determine whether diet and/or Lactobacillus rhamnosus GG (LGG) would mediate dextran sodium sulfate (DSS)-induced colitis by altering expression of the MUC and TFF genes. C57BL/6 mice were fed diets containing 20% (wt:wt) casein, soy, or whey proteins with or without LGG for 12 d. Seven days after starting LGG diets, the mice were given 2% DSS in drinking water for 4 d. Two additional casein groups with or without LGG were given tap water, for a total of 8 groups. One day after the DSS treatment, the mice were killed and the colon and cecum tissues and cecum contents were collected and analyzed by qRT-PCR. Whey protein significantly increased cecal LGG content compared with the other diets. In the casein diet groups, MUC1 and TFF-3 expression in colon was significantly induced by DSS independent of LGG. Compared with other DSS-treated groups, soy protein decreased MUC-1 and TFF-3 in the colon. Similarly, soy protein decreased the impact of DSS on inflammatory scores, TNFα gene expression, and colon shortening. There was no overall effect of LGG on these measurements. In conclusion, soy protein suppressed the DSS-induced inflammatory stimulation of MUC, TFF, and TNFα gene expression independently of LGG.
Assuntos
Colite Ulcerativa/dietoterapia , Colite Ulcerativa/genética , Lacticaseibacillus rhamnosus , Mucina-1/genética , Mucinas/genética , Probióticos/administração & dosagem , Proteínas de Soja/administração & dosagem , Fator de Necrose Tumoral alfa/genética , Animais , Sequência de Bases , Caseínas/administração & dosagem , Ceco/metabolismo , Ceco/microbiologia , Colite Ulcerativa/induzido quimicamente , Colite Ulcerativa/terapia , Colo/metabolismo , Colo/microbiologia , Sulfato de Dextrana/toxicidade , Modelos Animais de Doenças , Feminino , Expressão Gênica , Camundongos , Camundongos Endogâmicos C57BL , Proteínas do Leite/administração & dosagem , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fator Trefoil-3 , Proteínas do Soro do LeiteRESUMO
Many botanical compounds have been proposed to prevent cancer. We investigated the cancer treatment and prevention abilities of apigenin, baicalein, curcumin, epigallocatechin 3-gallate (EGCG), genistein, quercetin, and resveratrol both in vivo in transgenic adenocarcinoma of the mouse prostate (TRAMP) mice as well as in vitro in prostate cancer cell lines. In our experiments, these seven compounds act similarly to the Hedgehog antagonist cyclopamine, a teratogenic plant alkaloid, which had been previously shown to "cure" prostate cancer in a mouse xenograft model. With IC(50) values ranging from <1 to 25 mumol/L, these compounds can inhibit Gli1 mRNA concentration by up to 95% and downregulate Gli reporter activity by 80%. We show that four compounds, genistein, curcumin, EGCG, and resveratrol, inhibit Hedgehog signaling as monitored by real-time reverse transcription-PCR analysis of Gli1 mRNA concentration or by Gli reporter activity. Three compounds, apigenin, baicalein, and quercetin, decreased Gli1 mRNA concentration but not Gli reporter activity. Our results show that these compounds are also able to reduce or delay prostate cancer in vivo in TRAMP mice. All seven compounds, when fed in combination as pure compounds or as crude plant extracts, inhibit well-differentiated carcinoma of the prostate by 58% and 81%, respectively. In vitro, we show that all seven compounds also inhibit growth in human and mouse prostate cancer cell lines. Mechanistically, we propose the Hedgehog signaling pathway to be a direct or indirect target of these compounds. These botanicals at pharmacologic concentrations are potentially safer and less expensive alternatives to cyclopamine and its pharmaceutical analogues for cancer therapy.
Assuntos
Proteínas Hedgehog/metabolismo , Neoplasias da Próstata/metabolismo , Transdução de Sinais , Animais , Linhagem Celular Tumoral , Humanos , Concentração Inibidora 50 , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fitoterapia/métodos , RNA Mensageiro/metabolismo , Membro 25 de Receptores de Fatores de Necrose Tumoral/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
High-moisture extrusion of soy protein isolate generates a highly palatable meat substitute. No systematic evaluation of the nutritional quality of soy processed in this manner has been performed. This study compared the growth rate of male and female mice fed diets containing soy protein isolate without extrusion or with high-moisture extrusion. Other measures of overall growth and animal health were examined. Minor differences in the parameters were observed. Overall, the extruded soy protein was equally nutritious as the unextruded soy protein for the animals. Hence, high-moisture extrusion may be considered a useful method to generate high-quality protein foods. A longer term feeding trial may be recommended to further define the nutritional adequacy of this protein.
Assuntos
Proteínas Alimentares , Valor Nutritivo , Proteínas de Soja/isolamento & purificação , Animais , Desenvolvimento Ósseo/efeitos dos fármacos , Dieta Vegetariana , Proteínas Alimentares/administração & dosagem , Feminino , Manipulação de Alimentos/métodos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Tamanho do Órgão/efeitos dos fármacos , Caracteres Sexuais , Proteínas de Soja/administração & dosagem , Água/análise , Aumento de Peso/efeitos dos fármacosRESUMO
We determined if soy isoflavones have dose-related estrogenic and methylation effects. Thirty-four healthy premenopausal women were randomized to 40 mg or 140 mg isoflavones daily through one menstrual cycle. Breast specific and systemic estrogenic effects were assessed measuring the estrogenic marker complement (C)3 and changes in cytology, whereas methylation assessment of 5 cancer related genes (p16, RASSF1A, RARbeta2, ER, and CCND2) was performed on intraductal specimens. Serum genistein significantly increased after consuming both isoflavone doses. Cytology did not significantly change at either isoflavone dose. Serum C3 levels posttreatment were inversely related to change in serum genistein (r =-0.76, P = 0.0045) in women consuming low but not high dose isoflavones. The RAR beta 2 hypermethylation increase posttreatment correlated with the posttreatment genistein level considering the entire group (r = 0.67, P = 0.0017) and those receiving high-dose isoflavones (r = 0.68, P = 0.021). At the low but not the high isoflavone dose, CCND2 hypermethylation increase correlated with posttreatment genistein levels (r = 0.79, P = 0.011). In summary, the inverse correlation between C3 and genistein suggests an antiestrogenic effect. Isoflavones induced dose-specific changes in RARbeta2 and CCND2 gene methylation, which correlated with genistein levels. This work provides novel insights into estrogenic and methylation effects of dietary isoflavones.
Assuntos
Mama/química , Metilação de DNA/genética , Antagonistas de Estrogênios/administração & dosagem , Glycine max/química , Isoflavonas/administração & dosagem , Pré-Menopausa , Adulto , Líquidos Corporais/química , Mama/efeitos dos fármacos , Neoplasias da Mama/genética , Complemento C3/análise , Ciclina D2 , Ciclinas/genética , Metilação de DNA/efeitos dos fármacos , Método Duplo-Cego , Feminino , Genisteína/sangue , Humanos , Isoflavonas/sangue , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Regiões Promotoras Genéticas/efeitos dos fármacos , Estudos Prospectivos , Receptores do Ácido Retinoico/genéticaRESUMO
OBJECTIVES: Marginal intake of dietary zinc can be associated with increased risk of cardiovascular diseases. In the current study we hypothesized that vascular dysfunction and associated inflammatory events are activated during a zinc deficient state. DESIGN: We tested this hypothesis using both vascular endothelial cells and mice lacking the functional LDL-receptor gene. RESULTS: Zinc deficiency increased oxidative stress and NF-kappaB DNA binding activity, and induced COX-2 and E-selectin gene expression, as well as monocyte adhesion in cultured endothelial cells. The NF-kappaB inhibitor CAPE significantly reduced the zinc deficiency-induced COX-2 expression, suggesting regulation through NF-kappaB signaling. PPAR can inhibit NF-kappaB signaling, and our previous data have shown that PPAR transactivation activity requires adequate zinc. Zinc deficiency down-regulated PPARalpha expression in cultured endothelial cells. Furthermore, the PPARgamma agonist rosiglitazone was unable to inhibit the adhesion of monocytes to endothelial cells during zinc deficiency, an event which could be reversed by zinc supplementation. Our in vivo data support the importance of PPAR dysregulation during zinc deficiency. For example, rosiglitazone induced inflammatory genes (e.g., MCP-1) only during zinc deficiency, and adequate zinc was required for rosiglitazone to down-regulate pro-inflammatory markers such as iNOS. In addition, rosiglitazone increased IkappaBalpha protein expression only in zinc adequate mice. Finally, plasma data from LDL-R-deficient mice suggest an overall pro-inflammatory environment during zinc deficiency and support the concept that zinc is required for proper anti-inflammatory or protective functions of PPAR. CONCLUSIONS: These studies suggest that zinc nutrition can markedly modulate mechanisms of the pathology of inflammatory diseases such as atherosclerosis.
Assuntos
Células Endoteliais/metabolismo , Regulação da Expressão Gênica , Inflamação/metabolismo , NF-kappa B/metabolismo , Receptores Ativados por Proliferador de Peroxissomo/metabolismo , Zinco/deficiência , Animais , Aterosclerose/etiologia , Ácidos Cafeicos/farmacologia , Adesão Celular/efeitos dos fármacos , Células Cultivadas , Ciclo-Oxigenase 2/metabolismo , Modelos Animais de Doenças , Células Endoteliais/efeitos dos fármacos , Masculino , Camundongos , Monócitos/efeitos dos fármacos , Monócitos/fisiologia , NF-kappa B/antagonistas & inibidores , NF-kappa B/genética , Estresse Oxidativo , Receptores Ativados por Proliferador de Peroxissomo/agonistas , Receptores Ativados por Proliferador de Peroxissomo/genética , Álcool Feniletílico/análogos & derivados , Receptores de LDL/deficiência , Rosiglitazona , Tiazolidinedionas/farmacologia , Zinco/fisiologiaRESUMO
There have been many trials describing the effects of polyunsaturated fatty acids (PUFA) on fecundity, neonatal development, and maternal behavior in humans, but few controlled studies in rodents. We examined the effects of a maternal diet high in omega 3 (N-3) or omega 6 (N-6) PUFA on NIH Swiss mice. Female mice were ad libitum fed one of three complete and balanced diets (N-3, enriched in menhaden oil; N-6, enriched in corn oil; C, control diet, Purina 5015) from age 4 wk until the end of the study. Mice were bred at approximately 19 wk and 27 wk of age, providing a total of 838 pups from 129 litters in two experiments. After weaning their pups from parity 1, behavior of dams was assessed on elevated-plus and open-field mazes. Although the fraction of male pups from the N-3 and C groups was not different from 0.5, dams on the N-6 diet birthed more daughters than sons (213 vs. 133; P < 0.001). Although maternal stress has been reported to favor birth of daughters, the behavior of N-6 dams was not different from controls. By contrast, the N-3 dams displayed greater anxiety, spending less time in the open arms and more time in the closed arms of the elevated maze and traveling less distance and exhibiting less exploratory behavior in the open field (P < 0.05). N-3 dams tended to produce smaller litters than C dams, and N-3-suckled pups gained less weight (P < 0.05). In conclusion, the N-3 diet had negative effects on murine fecundity and maternal behavior, whereas the N-6 diet favored birth of daughters.
Assuntos
Dieta , Ácidos Graxos Ômega-3/administração & dosagem , Ácidos Graxos Ômega-6/administração & dosagem , Comportamento Materno/efeitos dos fármacos , Razão de Masculinidade , Animais , Animais Recém-Nascidos , Comportamento Animal/efeitos dos fármacos , Peso ao Nascer/efeitos dos fármacos , Ácidos Graxos Insaturados/administração & dosagem , Feminino , Masculino , Camundongos , Camundongos Endogâmicos , GravidezRESUMO
The HER2 proto-oncogene, a member of the epidermal growth factor receptor family, is overexpressed in 20-30% of breast cancers. Genistein, the main soy isoflavone, interacts with estrogen receptors (ER) and it is also a potent tyrosine kinase inhibitor. Previously, our laboratory found that genistein delayed mammary tumor onset in transgenic mice that overexpress HER2 gene. Our goal was to define the mechanism through which genistein affects mammary tumorigenesis in HER2 overexpressing mice. We hypothesized that genistein inhibits HER2 activation and expression through ER-dependent and ER-independent mechanisms. Genistein inhibited total HER2 protein expression and tyrosine phosphorylation in BT-474, an ERalpha (-) and ERbeta (+) human breast cancer cell line, however, E2 had no effect. Taken together, these data suggest that genistein has an ER-independent inhibitory effect, presumably, through tyrosine kinase inhibition activity. Genistein at 1.0 microM mimicked E2 and down-regulated HER2 protein phosphorylation when BT-474 was co-transfected with ERalpha, but not ERbeta. Although E2 and overexpression of HER2 can promote mammary tumorigenesis, an inverse relationship between ER expression and HER2 overexpression has been found in human breast cancer. We cloned a 500-bp promoter region upstream of the HER2 transcription initiation site. Co-transfection with ERalpha, but not with ERbeta, down-regulated HER2 promoter reporter in BT-474. At concentrations > or =1 microM, genistein inhibited HER2 promoter reporter in the absence of ERalpha. In conclusion, genistein at > or =1 microM inhibited HER2 protein expression, phosphorylation, and promoter activity through an ER-independent mechanism. In the presence of ERalpha, genistein mimicked E2 and inhibited HER2 protein phosphorylation. These data support genistein's chemo-prevention and potential chemo-therapeutic roles in breast cancer.
Assuntos
Anticarcinógenos/farmacologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Genes erbB-2/efeitos dos fármacos , Genisteína/farmacologia , Receptor ErbB-2/metabolismo , Receptores de Estrogênio/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Estradiol/farmacologia , Feminino , Humanos , Fosforilação/efeitos dos fármacos , Regiões Promotoras Genéticas/efeitos dos fármacos , Proteínas Tirosina Quinases/antagonistas & inibidores , Proto-Oncogene Mas , Receptor ErbB-2/genéticaRESUMO
Zinc is a structural and functional component of PPAR and zinc deficiency may be associated with an increased risk for cardiovascular diseases. We tested the hypothesis that zinc deficiency compromises lipid metabolism in rosiglitazone (RSG)-treated mice lacking the LDL-receptor (LDL-R) gene. LDL-R-deficient mice were maintained for 3 wk on low-fat (7 g/100 g) diets that were either zinc deficient or zinc adequate. Subsequently, diets were adjusted to a high-fat (HF) (15 g/100 g) regimen for 1 wk to produce a biological environment of mild oxidative and inflammatory stress. One-half of the mice within each zinc group was gavaged daily with the PPARgamma agonist RSG starting 2 d prior to the HF feeding. Selected lipid parameters were studied. Zinc deficiency increased plasma total cholesterol, which was also elevated by RSG. Zinc deficiency also caused an increased lipoprotein-cholesterol distribution toward the non-HDL fraction (VLDL, intermediate density lipoprotein, LDL). Plasma total fatty acids tended to increase during zinc deficiency and RSG treatment resulted in similar changes in the fatty acid profile in zinc-deficient mice. Fatty acid translocase (FAT/CD36) expression in abdominal aorta was upregulated by RSG only in zinc-deficient mice. In contrast, RSG treatment markedly increased lipoprotein lipase (LPL) expression only in zinc-adequate mice. In vitro studies confirmed that adequate zinc is required for RSG-induced PPARgamma activity to transactivate target genes. These data suggest that in this atherogenic mouse model treated with RSG, lipid metabolism can be compromised during zinc deficiency and that adequate dietary zinc may be considered during therapy with the antidiabetic medicine RSG.
Assuntos
Lipídeos/fisiologia , Receptores de LDL/deficiência , Tiazolidinedionas/farmacologia , Zinco/deficiência , Zinco/farmacologia , Animais , Colesterol/sangue , Ingestão de Energia , Ácidos Graxos não Esterificados/sangue , Lipoproteínas/sangue , Fígado/efeitos dos fármacos , Fígado/metabolismo , Masculino , Camundongos , Receptores de LDL/metabolismo , RosiglitazonaRESUMO
BACKGROUND: Prostate cancer is an important public health problem. It is an excellent candidate disease for chemoprevention because prostate cancer is typically slow growing and is usually diagnosed in elderly males. Pygeum africanum (Prunus africana or Rosaceae) is an African prune (plum) tree found in tropical Africa. An extract from the bark of Pygeum africanum has been used in Europe as a prevention and treatment of prostate disorders including benign prostatic hypertrophy (BPH). More recently in the USA, the phytotherapeutic preparations of Pygeum africanum and Saw palmetto have been marketed for prostate health including prostate cancer prevention and treatment. METHODS: The anti-cancer potential of Pygeum africanum has been tested both in vitro (PC-3 and LNCaP cells) and in vivo (TRAMP mouse model). RESULTS: In tissue culture, ethanolic extracts (30%) of Pygeum africanum inhibited the growth of PC-3 and LNCaP cells; induced apoptosis and altered cell kinetics; down regulated ERalpha and PKC-alpha protein, and demonstrated good binding ability to both mouse uterine estrogen receptors and LNCaP human androgen receptors. TRAMP mice fed Pygeum africanum showed a significant reduction (P = 0.034) in prostate cancer incidence (35%) compared to casein fed mice (62.5%). CONCLUSION: Pygeum africanum, which is widely used in Europe and USA for treatment of BPH, has a significant role in regulation of prostate cancer both in vitro and in vivo and therefore may be a useful supplement for people at high risk for developing prostate cancer.
Assuntos
Antineoplásicos/uso terapêutico , Carcinoma/tratamento farmacológico , Fitosteróis/uso terapêutico , Fitoterapia , Neoplasias da Próstata/tratamento farmacológico , Prunus africana , Animais , Linhagem Celular Tumoral , Avaliação Pré-Clínica de Medicamentos , Humanos , Masculino , Camundongos , Extratos Vegetais/uso terapêuticoRESUMO
Low zinc concentration can be associated with an increased risk of cardiovascular diseases. In the current study, we hypothesize that zinc deficiency can increase and zinc supplementation can decrease proatherosclerotic events in LDL receptor knock-out (LDL-R-/-) mice fed a moderate-fat diet. Mice were fed either a zinc-deficient (0 micromol Zn/g), a control (0.45 micromol Zn/g), or a zinc-supplemented (1.529 micromol Zn/g) diet for 4 wk. Mice fed the zinc-deficient diet had significantly increased concentrations of cholesterol and triacylglycerides in the VLDL and HDL fractions. Zinc supplementation decreased these lipid variables compared with control mice. We detected significantly higher concentrations of glutathione reductase mRNA in the thoracic aortae of zinc-deficient mice. Furthermore, inflammatory markers, such as nuclear factor-kappaB and vascular cell adhesion molecule-1, were significantly increased in zinc-deficient mice compared with mice of the control or supplemented groups. In addition, zinc deficiency significantly reduced the DNA binding activity of peroxisome proliferator activate receptors (PPARs) in liver extracts. Interestingly, mRNA expression levels of PPARgamma were significantly increased in thoracic aortae of zinc-deficient mice, indicating an adaptation process to decreased PPAR signaling. These data provide in vivo evidence of zinc deficiency inducing proinflammatory events in an atherogenic mouse model. These data also suggest that adequate zinc may be a critical component in protective PPAR signaling during atherosclerosis.