Project description:When applied in toxicological studies, the recently developed gene expression profiling techniques using microarrays, which brought forth the new field of toxicogenomics, facilitate the interpretation of a toxic compound?s mechanism of action. In this study, we investigated whether genotoxic carcinogens at doses known to induce liver tumors in the 2-year rat bioassay deregulate a common set of genes in a short-term in vivo study and, if so, whether these deregulated genes represent defined biological pathways. Rats were dosed with the four genotoxic hepatocarcinogens Dimethylnitrosamine [CAS:62-75-9;CHEBI:21778] (4 mg/kg/day), 2-nitrofluorene [CAS:607-57-8;CHEBI:1224] (44 mg/kg/day), Aflatoxin B1 [CAS:1162-65-8;CHEBI:2504] (0.24 mg/kg/day), and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone [CAS:64091-91-4;CHEBI:32692] (NNK, 20 mg/kg/day). After treatment for up to 14 days, the expression profiles of the livers were analyzed on Affymetrix RG_U34A microarrays. Among the significantly upregulated genes were a set of target genes of the tumor suppressor protein p53, indicating a DNA damage response. Such a response was expected and, therefore, confirmed the validity of our approach. In addition, the gene expression changes suggest a specific detoxification response, the activation of proliferative and survival signaling pathways, and some cell structural changes. These responses were strong throughout the 14 day time course for 2-nitrofluorene and aflatoxin B1; in the case of dimethylnitrosamine and NNK, the effects were weakly detectable at day 1 and then increased with time. For dimethylnitrosamine and aflatoxin B1, which caused observable inflammation in vivo, we found a corresponding upregulation of inflammatory genes at the same time points. Thus, by the toxicogenomic analysis of short-term in vivo studies, we identified genes and pathways commonly deregulated by genotoxic carcinogens, which may be indicative for the early events in tumorigenesis and, thus, predictive of later tumor development.
Project description:we assessed characteristic molecular and proteomic signatures in rat liver treated with drugs (pyrazinamide, ranitidine, enalapril, carbamazepine, and chlorpromazine) that are known to cause DILI in humans. In the present study, we assessed the characteristic gene expression signature for DILI in a rat model. Rats were administered representative drugs that are already known to induce DILI in humans and transcriptomic changes in rat liver were analyzed. The representative drugs, which induce three types (hepatocellular, mixed, and cholestatic) of DILI, that were used in this study were pyrazinamide (PZA, 150~1500 mg/kg), ranitidine (RAN, 209.5~2095 mg/kg), enalapril (ENA, 148.65~1486.5 mg/kg), carbamazepine (CBZ, 97.85~978.5 mg/kg), and chlorpromazine (CPZ, 7.1~71 mg/kg).
Project description:Androgens are required for prostate development, growth and physiology, by activating the androgen receptor (AR) upon activation by testosterone and dihydrotestosterone (DHT), the AR undergoes conformational changes, dimerizes and translocates to the cell nucleus regulation important genes releted to cell survival. Understanding the mechanisms of androgen regulation in the prostate gland is important, because the prostate is affected by several different diseases, in particular prostate cancer (PCa). Several ways exist to treat prostate cancer and promote epithelial cell death. Treatments involving androgen manipulation include surgical castration (bilateral orchiectomy), antiandrogens (usually AR antagonists), or substances that inhibit androgen synthesis (5 alpha-reductase inhibitors, gonadotrophin-releasing hormone blockers). 17 beta-estradiol exerts anti-androgen effects by blocking the hypothalamic production of gonadotropin-releasing hormone and thereby inhibiting the production of testosterone by the testes , but also acts locally via interactions with either of the estrogen receptors found in the gland. It is known that the kinetics of apoptosis are different in the rat ventral prostate (VP) of castrated rats (Cas group) and in rats subjected to 17 beta-estradiol high dose (group E2) or their combination (group Cas+E2), with an evident additive effect in the latter situation (Garcia-Florez et al, 2005). The microarray approach was done to figure out what genes are expressed and how the cells of ventral prostate gland responses when the androgen is not available comparing three diferent androgen deprivation methods (sirurgical castration, high dose of 17-beta estradiol and both treatment combined). Forty-eight 21-day-old male Wistar rats were obtained from the Multidisciplinary Center for Biological Research (CEMIB), University of Campinas. The animals were kept under normal light conditions (12-h light:dark cycle) and received filtered tap water and Purina rodent chow ad libitum. On the 90th day after birth, the rats were divided in four groups (n=3) and assigned to different treatment groups. To cause androgen deprivation, we utilized three different procedures with different effects on epithelial cell apoptosis. Animals in the first group were castrated (Cas) by orchiectomy via scrotal incision under ketamine (150 mg/Kg body weight) and xylazin (10 mg/kg body weight) anesthesia. Animals in the second group received a 25 mg/Kg body weight dose of 17β-estradiol diluted in corn oil (E2 group). The third group received a combination of both treatments (Cas+E2 group) (combined orchiectomy and 17β-estradiol). In the control group (Ct; normal androgen and estrogen), the animals received only the vehicle. Three days after the treatments, the rats were killed by anesthetic overdose, and the ventral prostate was dissected out for the microarray and immunohistochemistry analyses.
Project description:In this study we aimed to identify the molecular pathways modified by the false positive genotoxins Quercetin, 8-Hydroxyquinoline and 17beta-Estradiol that may explain their in vitro genotoxic and their in vivo non-genotoxic effects, by combining in vitro transcriptomics with phenotypic data. The effects of the false positive genotoxins were compared to the effects of the true genotoxins Benzo[a]pyrene and Aflatoxin B1 and the non-genotoxins 2,3,7,8-Tetrachlorodibenzodioxin, Cyclosporin A and Ampicillin C. <br><br>A custom CDF for use with the processed data file is available on the FTP site for this experiment.
Project description:Purpose : Identification of novel microRNA biomarkers in urine and plasma from rats with kidney or liver damage micoRNA-SEQ was used to analyze changes in miRNA profiles of tissue, plasma and urine samples of rats treated with either a nephrotoxicant (cisplatin) or one of two hepatotoxicants (Acetaminophen [APAP] or Carbon Tetrachloride [CCL4]).
Project description:Phenotypic plasticity, the ability of one genotype to express different phenotypes in response to changing environmental conditions, is one of the most common phenomena characterising the living world and is not only relevant for the ecology but also for the evolution of species. Daphnia, the waterflea, is a textbook example for predator induced phenotypic plastic defences including changes in life-history, behaviour and morphology. However, the analysis of molecular mechanisms underlying these inducible defences is still in its early stages.<br><br>We exposed Daphnia magna to chemical cues of the predator Triops cancriformis to identify key processes underlying plastic defensive trait formation. D. magna is known to develop an array of morphological changes in the presence of T. cancriformis including changes of carapace morphology and cuticle hardening. To get a more comprehensive idea of this phenomenon, we studied four different genotypes originating from habitats with different predation history, reaching from predator-free to temporary habitats containing T. cancriformis.<br><br>We analysed the morphologies as well as proteomes of predator-exposed and control animals. Three genotypes showed morphological changes when the predator was present. Using a high-throughput proteomics approach, we found 294 proteins which were significantly altered in their abundance after predator exposure in a general or genotype dependant manner. Proteins connected to genotype dependant responses were related to the cuticle, protein synthesis and calcium binding whereas the yolk protein vitellogenin increased in abundance in all genotypes, indicating their involvement in a more general response. Furthermore, genotype dependant responses at the proteome level correlated well with local adaptation to Triops predation.<br><br>Altogether, our study provides new insights concerning genotype dependant and general molecular processes involved in predator-induced phenotypic plasticity in D. magna.
Project description:An insulating myelin sheath ensures saltatory conduction of mechanosensory A afferents. Myelin damage results in the electrical instability of A fibers and the ability to generate pain in response to light touch/pressure (mechanical allodynia). We have hypothesized and then established that the release of T cell epitopes of myelin basic protein (MBP) enables nociceptive circuitry in myelinated fibers. Thus, mass spectrometry analysis of the rat sciatic nerve proteome followed by bioinformatics examination of the datasets revealed a loss of MBP and activation of T-helper cell signaling in the nerves undergoing chronic constriction injury (CCI). Matrix metalloproteinase-9 (MMP-9) proteolysis resulted in the MBP digest peptides, including the MBP84-104 and MBP68-86 regions, which exhibit prominent immunogenic epitopes. Myelin-forming Schwann cells and paranodal areas accumulated MHCII, MMP-9 and the degraded MBP at the sciatic nerve injury site. Administration of the immunodominant MBP84-104 and MBP68-86 peptides but not of the control peptides in a naïve rat sciatic nerve produced robust mechanical allodynia. Allodynia was accompanied by the T cell infiltration and an increase in MHCII, IL-17A and TNF- levels at the nerve injection site and the segmental ganglia. The pro-nociceptive activity of the synthetic MBP84-104 diminished in athymic nude rats lacking T cells. SB-3CT, an antagonist of MMP-9, inhibited mechanical allodynia, neuroinflammation and spinal sensitization after CCI. Collectively, our novel data implicate, for the first time, MMP-mediated cleavage of MBP and the resulting MBP digest fragments as a major cause of neuropathic pain. Gene extression profiling of total RNAs extracted from rat sciatic nerves, dorsal root ganglion and spinal cords after MBP84-104 peptide injection
Project description:This study looks at the effect of glycomacropeptide (GMP) treatment on rats with trinitrobenzene sulfuric acid (TNBS) induced colitis.<br><br> Female Wistar rats (175-225 g) were randomly assigned to three different groups. A control group C (n=3) that did not receive the TNBS challenge but 0.25 ml of phosphate buffered saline (PBS) intrarectally, and two groups (group T (n=3) and group GMP (n=5) that received the TNBS challenge as described below. Group GMP received 500 mg kg-1 day-1 of GMP in 1% methylcellulose p.o, starting two days before the TNBS challenge. Treatment was administered to rats for five days after the TNBS challenge using an oesophageal catheter. The T group received the vehicle. Colitis was induced by the method of Morris et al. with minor modifications (13). Briefly, animals were fasted overnight and anaesthetized with halothane. Under these conditions, animals were given 10 mg of TNBS dissolved in 0.25 ml of 50% ethanol (v v-1) by means of a Teflon cannula inserted 8 cm through the anus. Animals were kept in a head-down position for an additional 30 s and returned to their cage. Distal colon samples were obtained. For TNBS treated animals samples were obtained avoiding necrotic tissue.
Project description:Identification of novel pathophysiological mechanisms of carotid artery disease at systemic level by studying the gene expression profile of peripheral blood of affected patients with respect to control subjects.