RESUMO
BACKGROUND: The transparent epidermis of Caenorhabditis elegans makes it an attractive model to study sperm motility and migration within an intact reproductive tract. C elegans synthesize specific F-series prostaglandins (PGFs) that are important for guiding sperm toward the spermatheca. These PGFs are synthesized from polyunsaturated fatty acid (PUFA) precursors, such as arachidonic acid (AA), via a novel pathway, independent of the classical cyclooxygenases (Cox) responsible for most PG synthesis. While the enzyme(s) responsible for PG synthesis has yet to be identified, the DAF-7 TGFß pathway has been implicated in modulating PG levels and sperm guidance. RESULTS: We find that the reduced PGF levels in daf-1 type I receptor mutants are responsible for the sperm guidance defect. The lower level of PGs in daf-1 mutants is due in part to the inaccessibility of AA. Finally, lipid analysis and assessment of sperm guidance in daf-1;daf-3 double mutants suggest DAF-3 suppresses PG production and sperm accumulation at the spermatheca. Our data suggest that DAF-3 functions in the nervous system, and possibly the germline, to affect sperm guidance. CONCLUSION: The C elegans TGFß pathway regulates many pathways to modulate PG metabolism and sperm guidance. These pathways likely function in the nervous system and possibly the germline.
Assuntos
Prostaglandinas/biossíntese , Motilidade dos Espermatozoides/genética , Fator de Crescimento Transformador beta/fisiologia , Animais , Animais Geneticamente Modificados , Caenorhabditis elegans , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Feminino , Masculino , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Motilidade dos Espermatozoides/efeitos dos fármacos , Espermatozoides/efeitos dos fármacos , Espermatozoides/metabolismo , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo , Fator de Crescimento Transformador beta/farmacologiaRESUMO
Prostaglandins are formed by enzymatic and non-enzymatic mechanisms. They have been detected in human ovarian follicular fluid (HFF), a medium rich in growth factors and nutrients important for oocyte growth and fertility. However, the comprehensive identification of HFF prostaglandins has not been addressed. Here we use hybrid triple quadrupole time-of-flight and triple quadrupole mass spectrometers to comprehensively analyze prostaglandins in HFF. We identified PGE1, PGE2, PGF2α, and other prostaglandins synthesized via prostaglandin-endoperoxide synthase (i.e. Cox) cascades. We also identified specific PGF2α isomers (F2-isoprostanes) and PGF3α analogs whose structures are inconsistent with Cox-dependent formation. A prospective cohort pilot study of infertility patient subtypes revealed two potential associations. F2-isoprostanes are decreased in the diminished ovarian reserve subtype and elevated PGF2α may be associated with decreased live birth. Other than PGF2α, only body mass index >25kg/m2 correlated with poor in vitro fertilization outcome. Our studies suggest that HFF contains prostaglandins formed from at least two mechanisms, which may correlate with distinct clinical parameters.
Assuntos
Líquido Folicular/metabolismo , Espectrometria de Massas , Prostaglandinas/metabolismo , Adulto , Feminino , Fertilidade , Líquido Folicular/fisiologia , Humanos , Nascido VivoRESUMO
The mechanisms that guide motile sperm through the female reproductive tract to oocytes are not well understood. We have shown that Caenorhabditis elegans oocytes synthesize sperm guiding F-series prostaglandins from polyunsaturated fatty acid (PUFA) precursors provided in yolk lipoprotein complexes. Here we use genetics and electrospray ionization tandem mass spectrometry to partially delineate F-series prostaglandin metabolism pathways. We show that omega-6 and omega-3 PUFAs, including arachidonic and eicosapentaenoic acids, are converted into more than 10 structurally related F-series prostaglandins, which function collectively and largely redundantly to promote sperm guidance. Disruption of omega-3 PUFA synthesis triggers compensatory up-regulation of prostaglandins derived from omega-6 PUFAs. C. elegans F-series prostaglandin synthesis involves biochemical mechanisms distinct from those in mammalian cyclooxygenase-dependent pathways, yet PGF(2α) stereoisomers are still synthesized. A comparison of F-series prostaglandins in C. elegans and mouse tissues reveals shared features. Finally, we show that a conserved cytochrome P450 enzyme, whose human homolog is implicated in Bietti's Crystalline Dystrophy, negatively regulates prostaglandin synthesis. These results support the model that multiple cyclooxygenase-independent prostaglandins function together to promote sperm motility important for fertilization. This cyclooxygenase-independent pathway for F-series synthesis may be conserved.
Assuntos
Caenorhabditis elegans , Oócitos , Prostaglandinas F , Reprodução , Espermatozoides , Animais , Ácidos Araquidônicos/química , Ácidos Araquidônicos/metabolismo , Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/fisiologia , Sistema Enzimático do Citocromo P-450/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Ácidos Graxos Ômega-3/química , Ácidos Graxos Ômega-3/metabolismo , Ácidos Graxos Ômega-6/química , Ácidos Graxos Ômega-6/metabolismo , Feminino , Humanos , Masculino , Camundongos , Oócitos/metabolismo , Oócitos/fisiologia , Prostaglandinas F/biossíntese , Prostaglandinas F/química , Reprodução/genética , Reprodução/fisiologia , Motilidade dos Espermatozoides/genética , Espermatozoides/metabolismo , Espermatozoides/fisiologiaRESUMO
Mutations in VAPB/ALS8 are associated with amyotrophic lateral sclerosis (ALS) and spinal muscular atrophy (SMA), two motor neuron diseases that often include alterations in energy metabolism. We have shown that C. elegans and Drosophila neurons secrete a cleavage product of VAPB, the N-terminal major sperm protein domain (vMSP). Secreted vMSPs signal through Roundabout and Lar-like receptors expressed on striated muscle. The muscle signaling pathway localizes mitochondria to myofilaments, alters their fission/fusion balance, and promotes energy production. Here, we show that neuronal loss of the C. elegans VAPB homolog triggers metabolic alterations that appear to compensate for muscle mitochondrial dysfunction. When vMSP levels drop, cytoskeletal or mitochondrial abnormalities in muscle induce elevated DAF-16, the Forkhead Box O (FoxO) homolog, transcription factor activity. DAF-16 promotes muscle triacylglycerol accumulation, increases ATP levels in adults, and extends lifespan, despite reduced muscle mitochondria electron transport chain activity. Finally, Vapb knock-out mice exhibit abnormal muscular triacylglycerol levels and FoxO target gene transcriptional responses to fasting and refeeding. Our data indicate that impaired vMSP signaling to striated muscle alters FoxO activity, which affects energy metabolism. Abnormalities in energy metabolism of ALS patients may thus constitute a compensatory mechanism counterbalancing skeletal muscle mitochondrial dysfunction.
Assuntos
Esclerose Lateral Amiotrófica/metabolismo , Metabolismo Energético , Proteínas de Membrana/genética , Músculo Estriado/metabolismo , Atrofia Muscular Espinal/metabolismo , Esclerose Lateral Amiotrófica/genética , Esclerose Lateral Amiotrófica/patologia , Animais , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Drosophila/metabolismo , Fatores de Transcrição Forkhead , Humanos , Ligantes , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Knockout , Mitocôndrias/patologia , Neurônios Motores/metabolismo , Atrofia Muscular Espinal/genética , Atrofia Muscular Espinal/patologia , Transdução de Sinais , Fatores de Transcrição/metabolismo , Proteínas de Transporte VesicularRESUMO
This study was conducted to assess the value of a high resolution, high mass accuracy time-of-flight analyzer in combination with nanoliquid chromatography for the analysis of polyphenols and their metabolites. The goal was to create a method that utilizes small volumes of biological fluids and provides a significant improvement in sensitivity compared with existing methods. Accordingly, nanoLC-MS and nanoLC-pseudo-multiple reaction monitoring (MRM) methods were developed that had a lower limit of quantification of 0.5 nM for several polyphenols and were linear over 2-3 orders of magnitude (R(2)>0.999). Using urine samples, the ability to observe and quantify polyphenols in such a complex biological fluid depended on much narrower mass windows (0.050 amu or less) on a TOF analyzer than those used on a quadrupole analyzer (0.7 amu). Although a greater selectivity was possible with the low mass resolution of a triple quadrupole instrument using the MRM approach, for the daidzein metabolite O-DMA, a chromatographically resolvable second peak could only be substantially reduced by using a 0.01 amu mass window. The advantage of a TOF analyzer for product ion data is that the whole MSMS spectrum is collected at high mass accuracy and MRM experiments are conducted in silico after the analysis.
Assuntos
Cromatografia Líquida/métodos , Espectrometria de Massas/métodos , Nanotecnologia/métodos , Polifenóis/análise , Polifenóis/metabolismo , Animais , Limite de Detecção , Camundongos , Polifenóis/sangue , Polifenóis/urinaRESUMO
The extracellular cellular matrix (ECM) maintains tissue structure and regulates signaling functions by continuous degradation and remodeling. Inflammation or other disease conditions activate proteases including matrix metalloproteinases (MMPs) that degrade ECM proteins and in particular generate fragments of collagen and elastin, some of which are biologically active ECM peptides or matrikines. Stepwise degradation of collagen by MMP 8, 9 and prolyl endopeptidase release the matrikine proline-glycine-proline (PGP) and its product acetyl-PGP (AcPGP). These peptides are considered as potential biomarkers and therapeutic targets for many disease conditions such as chronic lung disease, heart disease, and cancer. However, there is no published, validated method for the measurement of PGP and AcPGP in plasma and therefore, we developed a sensitive, selective and reliable, isotope dilution LC-multiple reaction monitoring MS method for their determination in human plasma. The chromatographic separation of PGP and AcPGP was achieved in 3 min using Jupiter column with a gradient consisting of acidified acetonitrile and water at a flow rate of 0.5 ml/min. The limit of detection (LOD) for PGP and AcPGP was 0.01 ng/ml and the limit of quantification (LOQ) was 0.05 ng/ml and 0.1 ng/ml, respectively. Precision and accuracy values for all analytes were within 20 % except for the lowest QC of 0.01 ng/ml. The mean extraction recoveries of these analytes were > 90 % using a Phenomenex Phree cartridge and the matrix effect was < 15 % for all the QCs for PGP and AcPGP except the lowest QC. The stability of PGP and AcPGP was > 90 % in several tested conditions including autosampler use, storage at -80 °C, and after 6 times freeze-thaw cycles. Using this method, we successfully extracted and determined PGP levels in human plasma from healthy and COPD subjects. Therefore, this method is suitable for quantification of these peptides in the clinical setting.
Assuntos
Glicina , Prolina , Humanos , Cromatografia Líquida , Espectrometria de Massas em Tandem , Peptídeos , ColágenoRESUMO
Innexins are the subunits of invertebrate gap junctions. Here we show that the innexin INX-14 promotes sperm guidance to the fertilization site in the Caenorhabditis elegans hermaphrodite reproductive tract. inx-14 loss causes cell nonautonomous defects in sperm migration velocity and directional velocity. Results from genetic and immunocytochemical analyses provide strong evidence that INX-14 acts in transcriptionally active oocyte precursors in the distal gonad, not in transcriptionally inactive oocytes that synthesize prostaglandin sperm-attracting cues. Somatic gonadal sheath cell interaction is necessary for INX-14 function, likely via INX-8 and INX-9 expressed in sheath cells. However, electron microscopy has not identified gap junctions in oocyte precursors, suggesting that INX-14 acts in a channel-independent manner or INX-14 channels are difficult to document. INX-14 promotes prostaglandin signaling to sperm at a step after F-series prostaglandin synthesis in oocytes. Taken together, our results support the model that INX-14 functions in a somatic gonad/germ cell signaling mechanism essential for sperm function. We propose that this mechanism regulates the transcription of a factor(s) that modulates prostaglandin metabolism, transport, or activity in the reproductive tract.
Assuntos
Proteínas de Caenorhabditis elegans/fisiologia , Caenorhabditis elegans/metabolismo , Junções Comunicantes/metabolismo , Espermatozoides/citologia , Animais , Caenorhabditis elegans/citologia , Cromatografia Líquida , Ácidos Graxos/administração & dosagem , Feminino , Masculino , Microinjeções , Prostaglandinas/metabolismo , Interferência de RNA , Transdução de Sinais , Espectrometria de Massas por Ionização por Electrospray , Espectrometria de Massas em Tandem , Fatores de Transcrição/metabolismoRESUMO
In this study, a comparative, untargeted metabolomics approach was applied to compare urinary metabolite profiles of rats fed irradiated and non-irradiated diets. γ-Irradiated and non-irradiated NIH 7001 diet was given orally to animals beginning 5 days after exposure to the carcinogen N-methyl-N-nitrosourea and continued for 120 days. There was a 36% reduction in mammary tumor incidence in rats consuming the γ-irradiated diet, compared to rats receiving the non-irradiated form of the same diet. Urine samples from rats fed with γ-irradiated and non-irradiated diets were analyzed using nanoLC-MS/MS on a Q-TOF mass spectrometer, collecting positive and negative ion data. Data processing involved feature detection and alignment with MS-DIAL, normalization, mean-centering and Pareto scaling, and univariate and multivariate statistical analysis using MetaboAnalyst, and pathway analysis with Mummichog. Unsupervised Principal Component Analysis and supervised Partial Least Squares-Discriminant Analysis of both negative and positive ions revealed separation of the two groups. The top 25 metabolites from variable importance in projection scores >1 showed their contributions in discriminating urines the γ-irradiated diet fed group from non-irradiated control diet group. Consumption of the γ-irradiated diet led to alteration of several gut microbial metabolites such as phenylacetylglycine, indoxyl sulfate, kynurenic acid, hippurate and betaine in the urine. This study provides insights into metabolic changes in rat urine in response to a γ-irradiated diet which may be associated with mammary cancer prevention.
RESUMO
Dietary isoflavones, capable of influencing reproductive parameters in domestic cats (Felis catus), have been detected in commercial diets fed to captive cheetahs (Acinonyx jubatus). However, the absorptive and metabolic capacity of cheetahs towards isoflavones has not yet been studied. Experiments were designed to describe the plasma concentration-time curve, metabolite profile, and urinary and fecal excretion of genistein and daidzein in cheetahs following consumption of isoflavones. Four adult cheetahs were administered a single oral bolus of genistein and daidzein, and five juvenile cheetahs consuming a milk replacer formula found to contain isoflavones were also included. Urine was collected from all animals, and blood and feces were also collected from adult cheetahs following isoflavone exposure. Samples were analyzed for isoflavone metabolite concentration by liquid chromatography-electrospray ionization-multiple reaction ion monitoring mass spectrometry and high-performance liquid chromatography. Sulfate conjugates were the primary metabolites detected of both genistein and daidzein (60-80% of total isoflavones present) in the plasma and urine of cheetahs. A smaller proportion of daidzein was detected as conjugates in the urine of juvenile cheetahs, compared to adult cheetahs. Other metabolites included unconjugated genistein and daidzein, O-desmethylangolensin, and dihydrodaidzein, but not equol. Only 33% of the ingested genistein dose, and 9% of daidzein, was detected in plasma from adult cheetahs. However, of the ingested dose, 67% of genistein and 45% of daidzein were detected in the feces of adults. This study revealed that cheetahs appear efficient in their conjugation of absorbed dietary isoflavones and only a small fraction of ingested dose is absorbed. However, the capacity of the cheetah to conjugate genistein and daidzein with sulfate moieties appears lower than reported in the domestic cat. This may confer greater opportunity for biologic activity of isoflavones in the cheetah than would be predicted from findings in the domestic cat. However, further investigation is required.
Assuntos
Acinonyx , Ração Animal/análise , Dieta/veterinária , Isoflavonas/metabolismo , Isoflavonas/farmacocinética , Absorção , Envelhecimento , Fenômenos Fisiológicos da Nutrição Animal , Animais , Fezes/química , Isoflavonas/química , Isoflavonas/urina , Masculino , Estrutura MolecularRESUMO
Prostaglandins comprise a family of lipid signaling molecules derived from polyunsaturated fatty acids and are involved in a wide array of biological processes, including fertilization. Prostaglandin-endoperoxide synthase (a.k.a. cyclooxygenase or Cox) initiates prostaglandin synthesis from 20-carbon polyunsaturated fatty acids, such as arachidonic acid. Oocytes of Caenorhabditis elegans (C. elegans) have been shown to secrete sperm-guidance cues prostaglandins, independent of Cox enzymes. Both prostaglandin synthesis and signal transduction in C. elegans are environmentally modulated pathways that regulate sperm guidance to the fertilization site. Environmental factors such as food triggers insulin and TGF-ß secretion and their levels regulate tissue-specific prostaglandin synthesis in C. elegans. This novel PG pathway is abundant in mouse and human ovarian follicular fluid, where their functions, mechanism of synthesis and pathways remain to be established. Given the importance of prostaglandins in reproductive processes, a better understanding of how diets and other environmental factors influence their synthesis and function may lead to new strategies towards improving fertility in mammals.
RESUMO
The transforming growth factor beta superfamily encompasses a large family of ligands that are well conserved across many organisms. They are regulators of a number of physiological and pathological processes. The model nematode Caenorhabditis elegans has been instrumental in identifying key components of the transforming growth factor beta (TGFß) pathway. In C. elegans, the TGFß homolog DAF-7 signals through the DAF-1 Type I and DAF-4 Type II receptors to phosphorylate downstream R-SMADs DAF-8 and DAF-14. These R-SMADs translocate into the nucleus to inhibit Co-SMAD DAF-3. Many of the roles of the canonical DAF-7 pathway, involving both DAF-1 and DAF-3, have been identified using targeted genetic studies. Few have assessed the global transcriptomic changes in response to these genes, especially in adult animals. In this study, we performed RNA sequencing on wild type, daf-1, and daf-1; daf-3 adult hermaphrodites. To assess the overall trends of the data, we identified differentially expressed genes (DEGs) and performed gene ontology analysis to identify the types of downstream genes that are differentially expressed. Hierarchical clustering showed that the daf-1; daf-3 double mutants are transcriptionally more similar to wild type than daf-1 mutants. Analysis of the DEGs showed a disproportionally high number of genes whose expression is increased in daf-1 mutants, suggesting that DAF-1 acts as a general repressor of gene expression in wild type animals. Gene ontology analysis of the DEGs produced many significantly enriched terms, including Molting Cycle, Response to Topologically Incorrect Protein, and Response to Biotic Stimulus. Understanding the direct and indirect targets of the DAF-7 TGFß pathway through this RNA-seq dataset can provide insight into novel roles of the multifunctional signaling pathway, as well as identify novel genes that may participate in previously reported functions of TGFß signaling.
Assuntos
Proteínas de Caenorhabditis elegans/genética , Receptores de Superfície Celular/genética , Proteínas Smad/genética , Transcriptoma , Fator de Crescimento Transformador beta/genética , Animais , Caenorhabditis elegans , Proteínas de Caenorhabditis elegans/metabolismo , Mutação , Receptores de Superfície Celular/metabolismo , Transdução de Sinais , Proteínas Smad/metabolismoRESUMO
We previously discovered that Caenorhabditis elegans synthesizes Cox-independent F-series prostaglandins (PGs). To delineate the Cox-independent prostaglandin pathways and evaluate their role in sperm motility in C. elegans, we developed a novel biochemical method for the rapid production of F-series PGs using arachidonic acid as the substrate and worm lysate as source of enzyme(s). Among the four F2-series PGs produced in the reaction, three of them were identified as 8-isoPGF2α, 5iPF2 VI, and PGF2α based on their retention times and MS/MS spectral comparison with standards using LC-MS/MS. PG production was not markedly affected by specific antioxidants, or Cox, Lox, and Cyp inhibitors, suggesting that these PGs are formed through a novel, biologically regulated mechanism in C. elegans. This study also assessed the ability of 8-isoPGF2α, 5iPF2 VI, PGF2α, and a mixture containing these PGs in a 0.5/0.08/1 ratio that reflects their synthetic composition to modulate sperm motility in fat-2 mutants. PGF2α and the PG mixture at 25 µM concentration significantly stimulated sperm velocity by 28% and 38%, whereas 8-isoPGF2α and 5iPF2 VI reduced the velocity by 21% and 30%, respectively, compared to vehicle control. These results indicate that the sperm motility effects of PGs are structure- and composition-dependent in C. elegans.
Assuntos
Caenorhabditis elegans/metabolismo , Prostaglandina-Endoperóxido Sintases/metabolismo , Prostaglandinas/metabolismo , Motilidade dos Espermatozoides , Animais , Caenorhabditis elegans/enzimologia , Cromatografia Líquida , Masculino , Espectrometria de Massas em TandemRESUMO
The chemopreventive efficacy of cranberry juice concentrate in an experimental model of urinary bladder cancer was evaluated using female Fischer-344 rats. The animals received N-butyl-N-(4-hydroxybutyl)-nitrosamine (OH-BBN) for a period of eight weeks. Cranberry juice concentrate was administered at doses of 1.0 or 0.5 ml/rat/day beginning one week after the final OH-BBN treatment and continuing until the end of the study. The urinary bladders of all the rats were weighed and examined grossly for lesions, and all masses were submitted for pathological evaluation. A dose-dependent preventive effect of cranberry treatment was observed, with a reduced number of urinary bladder cancers (38%) in the 1.0 ml/rat/day group versus the control group. The cranberry extract neither affected body weight gain nor caused other signs of toxicity. For the metabolic studies, serum and urine were collected at 4 and 12 h after the administration of the cranberry juice concentrate and were analyzed by LC-MS/MS. Quercetin and its methylated derivative were detected in the urine samples. However, no quercetin was detected in the serum samples, indicating its poor bioavailability. These data suggest that components of cranberries may be effective in preventing urinary bladder carcinogenesis.
Assuntos
Fitoterapia , Extratos Vegetais/uso terapêutico , Neoplasias da Bexiga Urinária/tratamento farmacológico , Vaccinium macrocarpon/química , Animais , Butilidroxibutilnitrosamina , Cromatografia Líquida , Feminino , Ratos , Ratos Endogâmicos F344 , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Neoplasias da Bexiga Urinária/induzido quimicamente , Neoplasias da Bexiga Urinária/patologiaRESUMO
For the first time, a rapid, sensitive and simple liquid chromatography/tandem mass spectrometry (LC-MS/MS) method using an atmospheric pressure chemical ionization (APCI) source for the quantification of PD168393 in rat serum was developed and validated. Serum samples were pretreated with methanol for protein precipitation. The chromatographic separation was performed on a Jupiter-C5 column (250 mm x 2.0 mm i.d.) pre-equilibrated with 0.1% formic acid. The tandem mass spectrometer was tuned in the multiple reaction monitoring mode to monitor the m/z transitions 369/313 for PD168393 and m/z 343/308 for the internal standard triazolam, using positive ion mode. The MS/MS response was linear over the concentration range from 2 ng/mL to 5000 ng/mL, with a lower limit of quantification (LLQ) of 2 ng/mL. At the lowest quality control (4 ng/mL), the intra- and inter-day precisions (CV%) for PD168393 were less than 10% and the accuracies were between 92% and 111%. The validated method can be used in most or all stages of the screening and optimizing process for future method validation of pharmacokinetic studies.
Assuntos
Cromatografia Líquida/métodos , Quinazolinas/sangue , Espectrometria de Massas em Tandem/métodos , Animais , Quinazolinas/química , Ratos , Padrões de Referência , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
BACKGROUND: The onset of menopause marks a pivotal time in which the incidence of hypertension and of cardiovascular disease (CVD) begins to increase dramatically in women. Before menopause, the incidences of these diseases are significantly lower in women than in age-matched men. After menopause, the rates of these diseases in women eventually approximate those in men. The loss of endogenous estrogen at menopause has been traditionally believed to be the primary factor involved in these changes. OBJECTIVE: This review summarizes recent findings regarding the effectiveness of botanicals in the treatment of some menopausal symptoms and other symptoms of aging (eg, rise in arterial pressure, cognitive decline, insulin resistance, and hyperlipidemia). METHODS: Articles were selected for inclusion in this review based on the significance of the research and contribution to the current understanding of how each botanical elicits cardioprotective effects. To this end, PubMed and MEDLINE databases were searched, using terms that included the name of the specific botanical along with the relevant aspects of its action(s), such as blood pressure, glycemic control, and lipids. Most of the articles used were published within the past 5 years, although some older articles that were seminal in advancing the current understanding of botanicals were also included. RESULTS: Soy has been found to lower plasma lipid concentrations and arterial pressure in postmenopausal women and age-matched men, and to have protective effects in heart disease and atherosclerosis of the carotid and coronary circulation. Soy was also found to lower fasting insulin concentrations and glycosylated hemoglobin concentrations. Grape seed extract, another frequently used botanical, contains polyphenols that have been found to reduce arterial pressure and salt-sensitive hypertension in estrogendepleted animal models. CONCLUSION: Several botanical compounds have been found to have beneficial effects in the treatment of the symptoms of menopause and other symptoms of aging, including CVD, cognitive decline, and metabolic diseases.
Assuntos
Sistema Cardiovascular/efeitos dos fármacos , Suplementos Nutricionais , Isoflavonas/farmacologia , Fitoterapia , Extratos Vegetais/farmacologia , Animais , Pressão Sanguínea/efeitos dos fármacos , Cognição/efeitos dos fármacos , Feminino , Flavonoides/farmacologia , Genisteína/farmacologia , Extrato de Sementes de Uva , Humanos , Masculino , Fenóis/farmacologia , Fitoestrógenos/farmacologia , Extratos Vegetais/uso terapêutico , Polifenóis , Proantocianidinas/farmacologia , Proantocianidinas/uso terapêutico , Pueraria , Proteínas de Soja/farmacologia , Proteínas de Soja/uso terapêutico , Vasoconstrição/efeitos dos fármacosRESUMO
Asthma is a chronic inflammatory disease process involving the conductive airways of the human lung. The dysregulated inflammatory response in this disease process may involve multiple cell-cell interactions mediated by signaling molecules, including lipid mediators. Extracellular vesicles (EVs) are lipid membrane particles that are now recognized as critical mediators of cell-cell communication. Here, we compared the lipid composition and presence of specific lipid mediators in airway EVs purified from the bronchoalveolar lavage (BAL) fluid of healthy controls and asthmatic subjects with and without second-hand smoke (SHS) exposure. Airway exosome concentrations were increased in asthmatics, and correlated with blood eosinophilia and serum IgE levels. Frequencies of HLA-DR+ and CD54+ exosomes were also significantly higher in asthmatics. Lipidomics analysis revealed that phosphatidylglycerol, ceramide-phosphates, and ceramides were significantly reduced in exosomes from asthmatics compared to the non-exposed control groups. Sphingomyelin 34:1 was more abundant in exosomes of SHS-exposed asthmatics compared to healthy controls. Our results suggest that chronic airway inflammation may be driven by alterations in the composition of lipid mediators within airway EVs of human subjects with asthma.
Assuntos
Asma/patologia , Vesículas Extracelulares/metabolismo , Adulto , Idoso , Líquido da Lavagem Broncoalveolar/química , Estudos de Casos e Controles , Ceramidas/metabolismo , Análise Discriminante , Regulação para Baixo , Exossomos/metabolismo , Feminino , Antígenos HLA-DR/metabolismo , Humanos , Imunoglobulina E/sangue , Molécula 1 de Adesão Intercelular/metabolismo , Masculino , Pessoa de Meia-Idade , Fosfatidilgliceróis/metabolismo , Esfingomielinas/metabolismo , Poluição por Fumaça de TabacoRESUMO
The tumor suppressor protein Merlin is proteasomally degraded in breast cancer. We undertook an untargeted metabolomics approach to discern the global metabolomics profile impacted by Merlin in breast cancer cells. We discerned specific changes in glutathione metabolites that uncovered novel facets of Merlin in impacting the cancer cell metabolome. Concordantly, Merlin loss increased oxidative stress causing aberrant activation of Hedgehog signaling. Abrogation of GLI-mediated transcription activity compromised the aggressive phenotype of Merlin-deficient cells indicating a clear dependence of cells on Hedgehog signaling. In breast tumor tissues, GLI1 expression enhanced tissue identification and discriminatory power of Merlin, cumulatively presenting a powerful substantiation of the relationship between these two proteins. We have uncovered, for the first time, details of the tumor cell metabolomic portrait modulated by Merlin, leading to activation of Hedgehog signaling. Importantly, inhibition of Hedgehog signaling offers an avenue to target the vulnerability of tumor cells with loss of Merlin.
Assuntos
Adaptação Fisiológica , Neurofibromina 2/genética , Proteínas Hedgehog/genética , Proteínas Hedgehog/metabolismo , Humanos , Células MCF-7 , Metaboloma , Neurofibromina 2/metabolismo , Estresse Oxidativo , Transdução de Sinais , Proteína GLI1 em Dedos de Zinco/genética , Proteína GLI1 em Dedos de Zinco/metabolismoRESUMO
Specific and quantitative analyses of the bioactive components and their metabolites in body fluids are essential to assess the interaction between groups of compounds in dietary supplements and preparations of psychoactives. Reverse-phase LC separations combined with tandem mass spectrometry provide the necessary specificity and sensitivity. In this paper, applications of these methods are described for the analysis of isoflavones, salvinorin A, synephrine isomers and their metabolites in serum, urine and aqueous humor.
Assuntos
Suplementos Nutricionais , Flavonoides/química , Flavonoides/metabolismo , Fenóis/química , Fenóis/metabolismo , Animais , Cromatografia Líquida , Diterpenos/análise , Diterpenos/metabolismo , Diterpenos/farmacocinética , Diterpenos Clerodânicos , Cães , Flavonoides/farmacocinética , Espectrometria de Massas , Fenóis/farmacocinética , Polifenóis , Psicotrópicos/química , Psicotrópicos/metabolismo , Espectrometria de Massas por Ionização por Electrospray , Sinefrina/análise , Sinefrina/metabolismo , Sinefrina/farmacocinéticaRESUMO
In the present study, anti-proliferative activities of cranberry derived flavonoids and some of their in vivo metabolites were evaluated using a panel of human bladder tumor cell lines (RT4, SCABER, and SW-780) and non-tumorigenic immortalized human uroepithelial cells (SV-HUC). Among the compounds tested, quercetin 3-O-glucoside, isorhamnetin (3'-O-methylquercetin), myricetin and quercetin showed strong concentration-dependent cell growth inhibitory activities in bladder cancer cells with IC50 values in a range of 8-92 µM. Furthermore, isorhamnetin and myricetin had very low inhibitory activity against SV-HUC even at very high concentrations (>200 µM) compared to bladder cancer cells, indicating that their cytotoxicity is selective for cancer cells. To determine whether the differential cell growth inhibitory effects of isomeric flavonoids quercetin 3-O-glucoside (active) and hyperoside (quercetin 3-O-galactoside) (inactive) are related to their metabolism by the cancer cells, SW-780 cells were incubated with these compounds and their metabolism was examined by LC-MS/MS. Compared to quercetin 3-O-glucoside, hyperoside undergoes relatively less metabolic biotransformation (methylation, glucuronidation and quinone formation). These data suggest that isorhamnetin and quercetin 3-O-glucoside may be the active forms of quercetin in prevention of bladder cancer in vivo and emphasize the importance of metabolism for the prevention of bladder cancer by diets rich in cranberries.
Assuntos
Flavonoides/farmacologia , Inibidores do Crescimento/farmacologia , Extratos Vegetais/farmacologia , Neoplasias da Bexiga Urinária/fisiopatologia , Vaccinium macrocarpon/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Flavonoides/química , Flavonoides/metabolismo , Frutas/química , Frutas/metabolismo , Inibidores do Crescimento/química , Inibidores do Crescimento/metabolismo , Humanos , Estrutura Molecular , Extratos Vegetais/química , Extratos Vegetais/metabolismo , Espectrometria de Massas em Tandem , Neoplasias da Bexiga Urinária/tratamento farmacológico , Neoplasias da Bexiga Urinária/metabolismo , Vaccinium macrocarpon/metabolismoRESUMO
Metabolomics, a systems biology discipline representing analysis of known and unknown pathways of metabolism, has grown tremendously over the past 20 years. Because of its comprehensive nature, metabolomics requires careful consideration of the question(s) being asked, the scale needed to answer the question(s), collection and storage of the sample specimens, methods for extraction of the metabolites from biological matrices, the analytical method(s) to be employed and the quality control of the analyses, how collected data are correlated, the statistical methods to determine metabolites undergoing significant change, putative identification of metabolites and the use of stable isotopes to aid in verifying metabolite identity and establishing pathway connections and fluxes. This second part of a comprehensive description of the methods of metabolomics focuses on data analysis, emerging methods in metabolomics and the future of this discipline. Copyright © 2016 John Wiley & Sons, Ltd.