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Endogenous Metabolites (endogenous + metabolite)
Selected AbstractsUse of the 1-mm micro-probe for metabolic analysis on small volume biological samplesJOURNAL OF CELLULAR AND MOLECULAR MEDICINE, Issue 8b 2009Natalie J. Serkova Abstract Endogenous metabolites are promising diagnostic end-points in cancer research. Clinical application of high-resolution NMR spectroscopy is often limited by extremely low volumes of human specimens. In the present study, the use of the Bruker 1-mm high-resolution TXI micro-probe was evaluated in the elucidation of metabolic profiles for three different clinical applications with limited sample sizes (body fluids, isolated cells and tissue biopsies). Sample preparation and 1H-NMR metabolite quantification protocols were optimized for following oncology-oriented applications: (i) to validate the absolute concentrations of citrate and spermine in human expressed prostatic specimens (EPS volumes 5 to 10 ,l: prostate cancer application); (ii) to establish the metabolic profile of isolated human lymphocytes (total cell count 4 = 106: chronic myelogenous leukaemia application); (iii) to assess the metabolic composition of human head-and-neck cancers from mouse xenografts (biopsy weights 20 to 70 mg: anti-cancer treatment application). In this study, the use of the Bruker 1-mm micro-probe provides a convenient way to measure and quantify endogenous metabolic profiles of samples with a very low volume/weight/cell count. [source] In vitro and in vivo antitumor effect of 2-methoxyestradiol on human melanomaINTERNATIONAL JOURNAL OF CANCER, Issue 5 2004Judit Dobos Abstract 2-methoxyestradiol (2ME2) is an endogenous metabolite of estradiol with estrogen-receptor-independent antitumor and antiangiogenic activity. We examined the effects of 2ME2 on the cellular proliferation of 8 human melanoma cell lines. We show that 2ME2 inhibited cell proliferation by inducing apoptosis and an arrest in the G2/M phase, and the mechanism of action involved microtubules, mitochondrial damage and caspase activation. In male SCID mice, 2ME2 was effective in reducing primary tumor weight and the number of liver metastases after intrasplenic injection of human melanoma cells. In the metastases, we found a significantly higher rate of apoptotic cells after 2ME2 treatment. These findings on the antitumor effect of 2ME2 in cell culture as well as in an animal model may have implications for designing alternative treatment options for patients with advanced malignant melanoma. © 2004 Wiley-Liss, Inc. [source] 2-methoxyestradiol induces apoptosis and cell cycle arrest in human chondrosarcoma cellsJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 8 2007Yi-Chin Fong Abstract 2-Methoxyestradiol (2ME) is an endogenous metabolite with estrogen receptor-independent anti-tumor activity. The current study seeks to determine the mechanism of anti-tumor activity of 2ME on human chondrosarcoma. 2ME caused a time- and dose-dependent cytotoxity in chondrosarcoma cells, while primary chondrocytes were minimally affected. Cells accumulated in G0/G1 phase in response to 2ME and DAPI stain indicated an induction of apoptosis. Bax, Cytochrome C, and Caspase-3 protein expression were increased, while p53 expression was decreased. A higher Bax/Bcl-2 ratio followed 2ME treatment. 2ME has a potentially promising role as a systemic therapy of chondrosarcoma when the mechanism of action is better delineated. Published by Wiley Periodicals, Inc. J Orthop Res 25:1106,1114, 2007 [source] Induction of endogenous pathways by antiepileptics and clinical implicationsFUNDAMENTAL & CLINICAL PHARMACOLOGY, Issue 5 2005M. Strolin Benedetti Abstract The aim of this study was to review modifications of the endogenous pathways (e.g. enzyme elevations, normal body constituent depletion or higher formation/excretion of endogenous metabolites) which could be ascribed to enzyme induction by antiepileptic drugs (AEDs). Information on older (e.g. phenobarbital, phenytoin and carbamazepine) and newer drugs (where information is available) is discussed together with clinical implications. The enzymes involved in the endogenous pathways and induced by the AEDs will not be limited to the hepatic microsomal enzymes; extrahepatic enzymes and/or enzymes present in other subcellular fractions will also be discussed, if pertinent. The induction of endogenous pathways by AEDs has been taken into account in the past, but much less emphasis has been given compared with the extensive literature on induction by AEDs of the metabolism of concomitantly administered drugs, either of the same or of different classes. Not all of the endogenous pathways examined and induced by AEDs appear to result in serious clinical consequences (e.g. induction of hepatic ALP, increased excretion of d -glucaric acid or of 6, -hydroxycortisol). In some cases, induction of some pathways (e.g. increase of high-density lipoprotein cholesterol or of conjugated bilirubin) might even be a beneficial side-effect, however enzyme induction is considered rather a detrimental aspect for an AED, as induction is generally a broad and a non-specific phenomenon. The new AEDs have generally less induction potential than the older agents. Yet some (felbamate, topiramate, oxcarbazepine and lamotrigine) have the potential for inducing enzymes, whereas others (levetiracetam, gabapentin and vigabatrin) appear to be completely devoid of enzyme inducing characteristics, at least as far as the enzymes investigated are concerned. [source] Multidrug resistance,associated proteins are crucial for the viability of activated rat hepatic stellate cells,,HEPATOLOGY, Issue 2 2008Rebekka A. Hannivoort Hepatic stellate cells (HSCs) survive and proliferate in the chronically injured liver. ATP-binding cassette (ABC) transporters play a crucial role in cell viability by transporting toxic metabolites or xenobiotics out of the cell. ABC transporter expression in HSCs and its relevance to cell viability and/or activation have not been reported so far. The aim of this study was to investigate the expression, regulation, and function of multidrug resistance,associated protein (Mrp)-type and multidrug resistance protein (Mdr),type ABC transporters in activated rat HSCs. Rat HSCs were exposed to cytokines or oxidative stress. ABC transporter expression was determined by quantitative polymerase chain reaction and immunohistochemistry. HSCs were exposed to the Mdr inhibitors verapamil and PSC-833 and the Mrp inhibitor MK571. Mdr and Mrp transporter function was evaluated with flow cytometry. Apoptosis was determined by activated caspase-3 and acridine orange staining, and necrosis was determined by Sytox green nuclear staining. An in vivo model of carbon tetrachloride (CCl4),induced liver fibrosis was used. With respect to hepatocytes, activated HSCs expressed high levels of Mrp1 and comparable levels of Mrp3, Mrp4, Mdr1a, and Mdr1b but not the hepatocyte-specific transporters bile salt export pump, Mrp2, and Mrp6. Mrp1 protein staining correlated with desmin staining in livers from CCl4 -treated rats. Mrp1 expression increased upon activation of HSCs. Cytokines induced Mdr1b expression only. Oxidative stress was not a major regulator of Mdr and Mrp transporter expression. Activated HSCs became necrotic when exposed to the Mrp inhibitors. Conclusion: Activated HSCs contain relatively high levels of Mrp1. Mrp-type transporters are required for the viability of activated HSCs. Mrp-dependent export of endogenous metabolites is important for the survival of activated HSCs in chronic liver diseases. (HEPATOLOGY 2008.) [source] Metabolomics in the assessment of chemical-induced reproductive and developmental outcomes using non-invasive biological fluids: application to the study of butylbenzyl phthalateJOURNAL OF APPLIED TOXICOLOGY, Issue 8 2009Susan Sumner Abstract This study was conducted to evaluate the use of metabolomics for improving our ability to draw correlations between early life exposures and reproductive and/or developmental outcomes. Pregnant CD rats were exposed by gavage daily during gestation to vehicle or to butylbenzyl phthalate (BBP) in vehicle at a level known to induce effects in the offspring and at a level previously not shown to induce effects. Urine was collected for 24,h (on dry ice using all glass metabolism chambers) from dams on gestational day 18 (during exposure) and on post natal day (pnd) 21, and from pnd 25 pups. Traditional phenotypic anchors were measured in pups (between pnd 0 and pnd 26). Metabolomics of urine collected from dams exposed to vehicle or BBP exhibited different patterns for endogenous metabolites. Even three weeks after gestational exposure, metabolic profiles of endogenous compounds in urine could differentiate dams that received the vehicle, low dose or high dose of BBP. Metabolic profiles could differentiate male from female pups, pups born to dams receiving the vehicle, low or high BBP dose, and pups with observable adverse reproductive effects from pups with no observed effects. Metabolites significant to the separation of dose groups and their relationship with effects measured in the study were mapped to biochemical pathways for determining mechanistic relevance. The application of metabolomics to understanding the mechanistic link between low levels of environmental exposure and disease/dysfunction holds huge promise, because this technology is ideal for the analysis of biological fluids in human populations. Copyright © 2009 John Wiley & Sons, Ltd. [source] Development and validation of a gas chromatography/mass spectrometry method for the metabolic profiling of human colon tissueRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 4 2009Mainak Mal In this study, a gas chromatography/mass spectrometry (GC/MS) method was developed and validated for the metabolic profiling of human colon tissue. Each colon tissue sample (20,mg) was ultra-sonicated with 1,mL of a mixture of chloroform/methanol/water in the ratio of 20:50:20 (v/v/v), followed by centrifugation, collection of supernatant, drying, removal of moisture using anhydrous toluene and finally derivatization using N -methyl- N -trifluoroacetamide (MSTFA) with 1% trimethylchlorosilane (TMCS). A volume of 1,µL of the derivatized mixture was injected into the GC/MS system. A total of 53 endogenous metabolites were separated and identified in the GC/MS chromatogram, all of which were selected to evaluate the sample stability and precision of the method. Of the identified endogenous metabolites 19 belonging to diverse chemical classes and covering a wide range of the GC retention times (Rt) were selected to investigate the quantitative linearity of the method. The developed GC/MS method demonstrated good reproducibility with intra- and inter-day precision within relative standard deviation (RSD) of ±15%. The metabolic profiles of the intact tissue were determined to be stable (100,±,15%) for up to 90 days at ,80°C. Satisfactory results were also obtained in the case of other stability-indicating studies such as freeze/thaw cycle stability, bench-top stability and autosampler stability. The developed method showed a good linear response for each of the 19 analytes tested (r2,>,0.99). Our GC/MS metabolic profiling method was successfully applied to discriminate biopsied colorectal cancer (CRC) tissue from their matched normal tissue obtained from six CRC patients using orthogonal partial least-squares discriminant analysis [two latent variables, R2Y,=,0.977 and Q2 (cumulative),=,0.877]. Copyright © 2009 John Wiley & Sons, Ltd. [source] Liquid chromatography/mass spectrometry for metabonomics investigation of the biochemical effects induced by aristolochic acid in rats: the use of information-dependent acquisition for biomarker identificationRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 6 2008Wan Chan The toxic effects of oral administrations of nephrotoxic and carcinogenic aristolochic acid (AA) to male Sprague-Dawley rats were investigated by using high-performance liquid chromatography coupled with a quadrupole time-of-flight mass spectrometer. Analysis of the urine and plasma samples revealed distinct changes in the biochemical patterns in the AA-dosed rats. After peak finding and alignment, principal component analysis (PCA) and partial least-squares discriminant analysis (PLS-DA) were used for multivariate data analysis. Potential biomarkers were studied by high-resolution mass spectrometry (MS) and tandem mass spectrometry (MS/MS) analyses. The MS/MS spectra of all endogenous metabolites satisfying the pre-defined criteria were acquired in a single information-dependent acquisition (IDA) analysis, demonstrating that IDA was an efficient approach for structural elucidation in metabonomic studies. Citric acid and a glucuronide-containing metabolite were observed as potential biomarkers in rat urine. A significant increase in plasma creatinine concentration was also observed in the AA-dosed rats, which indicated that AA induced an adverse effect on the renal clearance function. Copyright © 2008 John Wiley & Sons, Ltd. [source] UPLC/MSE; a new approach for generating molecular fragment information for biomarker structure elucidationRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 13 2006Robert S. Plumb A new approach to obtain fragmentation information in liquid chromatography/mass spectrometry (LC/MS) studies of small molecules in complex mixtures is presented using simultaneous acquisition of exact mass at high and low collision energy, MSE. LC/MS-TOF and LC/MS/MS-TOF are powerful tools for the analysis of complex mixtures, especially those for biological fluids allowing the elucidation of elemental composition and fragmentation information. In this example the composition of rat urine was studied using this new approach, allowing the structures of several endogenous components to be confirmed in one analytical run by the simultaneous acquisition of exact mass precursor and fragment ion data. The spectral data obtained using this new approach are comparable to those obtained by conventional LC/MS/MS as exemplified by the identification of endogenous metabolites present in rat urine. Copyright © 2006 John Wiley & Sons, Ltd. [source] | |||||||||||||