-
[show abstract]
[hide abstract]
ABSTRACT: Serum levels of cardiac troponins serve as biomarkers of myocardial injury. However, troponins are released into the serum only after damage to cardiac tissue has occurred. Here, we report development of a mouse model of doxorubicin (DOX)-induced chronic cardiotoxicity to aid in the identification of predictive biomarkers of early events of cardiac tissue injury. Male B6C3F(1) mice were administered intravenous DOX at 3mg/kg body weight, or an equivalent volume of saline, once a week for 4, 6, 8, 10, 12, and 14weeks, resulting in cumulative DOX doses of 12, 18, 24, 30, 36, and 42mg/kg, respectively. Mice were sacrificed a week following the last dose. A significant reduction in body weight gain was observed in mice following exposure to a weekly DOX dose for 1week and longer compared to saline-treated controls. DOX treatment also resulted in declines in red blood cell count, hemoglobin level, and hematocrit compared to saline-treated controls after the 2nd weekly dose until the 8th and 9th doses, followed by a modest recovery. All DOX-treated mice had significant elevations in cardiac troponin T concentrations in plasma compared to saline-treated controls, indicating cardiac tissue injury. Also, a dose-related increase in the severity of cardiac lesions was seen in mice exposed to 24mg/kg DOX and higher cumulative doses. Mice treated with cumulative DOX doses of 30mg/kg and higher showed a significant decline in heart rate, suggesting drug-induced cardiac dysfunction. Altogether, these findings demonstrate the development of DOX-induced chronic cardiotoxicity in B6C3F(1) mice.
Toxicology and Applied Pharmacology 11/2012; · 4.45 Impact Factor
-
Volodymyr Tryndyak,
Aline de Conti,
Tetyana Kobets,
Kristy Kutanzi,
Igor Koturbash,
Tao Han, James C Fuscoe,
John R Latendresse,
Stepan Melnyk,
Svitlana Shymonyak,
Leonard Collins,
Sharon A Ross,
Ivan Rusyn,
Frederick A Beland,
Igor P Pogribny
[show abstract]
[hide abstract]
ABSTRACT: Nonalcoholic fatty liver disease (NAFLD) is a major health problem and a leading cause of chronic liver disease in the United States and developed countries. In humans, genetic factors greatly influence individual susceptibility to NAFLD. The goals of this study were to compare the magnitude of interindividual differences in the severity of liver injury induced by methyl-donor deficiency among individual inbred strains of mice and to investigate the underlying mechanisms associated with the variability. Feeding mice a choline- and folate-deficient diet for 12 wk caused liver injury similar to NAFLD. The magnitude of liver injury varied among the strains, with the order of sensitivity being A/J ≈ C57BL/6J ≈ C3H/HeJ < 129S1/SvImJ ≈ CAST/EiJ < PWK/PhJ < WSB/EiJ. The interstrain variability in severity of NAFLD liver damage was associated with dysregulation of genes involved in lipid metabolism, primarily with a down-regulation of the peroxisome proliferator receptor α (PPARα)-regulated lipid catabolic pathway genes. Markers of oxidative stress and oxidative stress-induced DNA damage were also elevated in the livers but were not correlated with severity of liver damage. These findings suggest that the PPARα-regulated metabolism network is one of the key mechanisms determining interstrain susceptibility and severity of NAFLD in mice.- Tryndyak, V., de Conti, A., Kobets, T., Kutanzi, K., Koturbash, I., Han, T., Fuscoe, J. C., Latendresse, J. R., Melnyk, S., Shymonyak, S., Collins, L., Ross, S. A., Rusyn, I., Beland, F. A., Pogribny, I. P. Interstrain differences in the severity of liver injury induced by a choline- and folate-deficient diet in mice are associated with dysregulation of genes involved in lipid metabolism.
The FASEB Journal 08/2012; · 5.71 Impact Factor
-
Tao Han,
Ching-Wei Chang,
Joshua C Kwekel,
Ying Chen,
Yun Ge,
Francisco Martinez-Murillo,
Donna Roscoe,
Zivana Težak,
Reena Philip,
Karen Bijwaard, James C Fuscoe
[show abstract]
[hide abstract]
ABSTRACT: Genotyping assays often require substantial amounts of DNA. To overcome the problem of limiting amounts of available DNA, Whole Genome Amplification (WGA) methods have been developed. The multiple displacement amplification (MDA) method using Φ29 polymerase has become the preferred choice due to its high processivity and low error rate. However, the uniformity and fidelity of the amplification process across the genome has not been extensively characterized.
To assess amplification uniformity, we used array-based comparative genomic hybridization (aCGH) to evaluate DNA copy number variations (CNVs) in DNAs amplified by two MDA kits: GenomiPhi and REPLI-g. The Agilent Human CGH array containing nearly one million probes was used in this study together with DNAs from a normal subject and 2 cystic fibrosis (CF) patients. Each DNA sample was amplified 4 independent times and compared to its native unamplified DNA. Komogorov distances and Phi correlations showed a high consistency within each sample group. Less than 2% of the probes showed more than 2-fold CNV introduced by the amplification process. The two amplification kits, REPLI-g and GenomiPhi, generate very similar amplified DNA samples despite the differences between the unamplified and amplified DNA samples. The results from aCGH analysis indicated that there were no obvious CNVs in the CFTR gene region due to WGA when compared to unamplified DNA. This was confirmed by quantitative real-time PCR copy number assays at 10 locations within the CFTR gene. DNA sequencing analysis of a 2-kb region within the CFTR gene showed no mutations introduced by WGA.
The relatively high uniformity and consistency of the WGA process, coupled with the low replication error rate, suggests that WGA DNA may be suitable for accurate genotyping. Regions of the genome that were consistently under-amplified were found to contain higher than average GC content. Because of the consistent differences between the WGA DNA and the native unamplified DNA, characterization of the genomic region of interest, as described here, will be necessary to ensure the reliability of genotyping results from WGA DNA.
BMC Genomics 06/2012; 13:217. · 4.07 Impact Factor
-
Robert N McBurney,
Wade M Hines,
Linda S VonTungeln,
Laura K Schnackenberg,
Richard D Beger,
Carrie L Moland,
Tao Han, James C Fuscoe,
Ching-Wei Chang,
James J Chen, [......],
Armin Graber,
Yu Guo,
Peter Juhasz,
Tricia Y Li,
Moira D Lynch,
Nicole M Morel,
Thomas N Plasterer,
Edward J Takach,
Chenhui Zeng,
Frederick A Beland
[show abstract]
[hide abstract]
ABSTRACT: The Liver Toxicity Biomarker Study is a systems toxicology approach to discover biomarkers that are indicative of a drug's potential to cause human idiosyncratic drug-induced liver injury. In phase I, the molecular effects in rat liver and blood plasma induced by tolcapone (a "toxic" drug) were compared with the molecular effects in the same tissues by dosing with entacapone (a "clean" drug, similar to tolcapone in chemical structure and primary pharmacological mechanism). Two durations of drug exposure, 3 and 28 days, were employed. Comprehensive molecular analysis of rat liver and plasma samples yielded marker analytes for various drug-vehicle or drug-drug comparisons. An important finding was that the marker analytes associated with tolcapone only partially overlapped with marker analytes associated with entacapone, despite the fact that both drugs have similar chemical structures and the same primary pharmacological mechanism of action. This result indicates that the molecular analyses employed in the study are detecting substantial "off-target" markers for the two drugs. An additional interesting finding was the modest overlap of the marker data sets for 3-day exposure and 28-day exposure, indicating that the molecular changes in liver and plasma caused by short- and long-term drug treatments do not share common characteristics.
Toxicologic Pathology 05/2012; 40(6):951-64. · 1.91 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Acrylamide (AA) is an industrial chemical that has been extensively investigated for central nervous system (CNS), reproductive, and genetic toxicity. However, AA effects on the liver, a major organ of drug metabolism, have not been adequately explored. In addition, the role of mitochondria in AA-mediated toxicity is still unclear. Changes in expression levels of genes associated with hepatic mitochondrial function of male transgenic Big Blue (BB) mice administered 500 mg/L AA or an equimolar concentration (600 mg/L) of its reactive metabolite glycidamide (GA) in drinking water for 3 and 4 wk, respectively, were examined. Transcriptional profiling of 542 mitochondria-related genes indicated a significant downregulation of genes associated with the 3-beta-hydroxysteroid dehydrogenase family in AA- and GA-treated mice, suggesting a possible role of both chemicals in altering hepatic steroid metabolism in BB mice. In addition, genes associated with lipid metabolism were altered by both treatments. Interestingly, only the parental compound (AA) significantly induced expression levels of genes associated with oxidative phosphorylation, in particular ATP synthase, which correlated with elevated ATP levels, indicating an increased energy demand in liver during AA exposure. Acrylamide-treated mice also showed significantly higher activity of glutathione S-transferase in association with decreased levels of reduced glutathione (GSH), which may imply an enhanced rate of conjugation of AA with GSH in liver. These results suggest different hepatic mechanisms of action of AA and GA and provide important insights into the involvement of mitochondria during their exposures.
Journal of Toxicology and Environmental Health Part A 03/2012; 75(6):324-39. · 1.83 Impact Factor
-
Yunfeng Tie,
Brooks McPhail,
Huixiao Hong,
Bruce A Pearce,
Laura K Schnackenberg,
Weigong Ge,
Dan A Buzatu,
Jon G Wilkes, James C Fuscoe,
Weida Tong,
Bruce A Fowler,
Richard D Beger,
Eugene Demchuk
[show abstract]
[hide abstract]
ABSTRACT: Polypharmacy increasingly has become a topic of public health concern, particularly as the U.S. population ages. Drug labels often contain insufficient information to enable the clinician to safely use multiple drugs. Because many of the drugs are bio-transformed by cytochrome P450 (CYP) enzymes, inhibition of CYP activity has long been associated with potentially adverse health effects. In an attempt to reduce the uncertainty pertaining to CYP-mediated drug-drug/chemical interactions, an interagency collaborative group developed a consensus approach to prioritizing information concerning CYP inhibition. The consensus involved computational molecular docking, spectral data-activity relationship (SDAR), and structure-activity relationship (SAR) models that addressed the clinical potency of CYP inhibition. The models were built upon chemicals that were categorized as either potent or weak inhibitors of the CYP3A4 isozyme. The categorization was carried out using information from clinical trials because currently available in vitro high-throughput screening data were not fully representative of the in vivo potency of inhibition. During categorization it was found that compounds, which break the Lipinski rule of five by molecular weight, were about twice more likely to be inhibitors of CYP3A4 compared to those, which obey the rule. Similarly, among inhibitors that break the rule, potent inhibitors were 2-3 times more frequent. The molecular docking classification relied on logistic regression, by which the docking scores from different docking algorithms, CYP3A4 three-dimensional structures, and binding sites on them were combined in a unified probabilistic model. The SDAR models employed a multiple linear regression approach applied to binned 1D ¹³C-NMR and 1D ¹⁵N-NMR spectral descriptors. Structure-based and physical-chemical descriptors were used as the basis for developing SAR models by the decision forest method. Thirty-three potent inhibitors and 88 weak inhibitors of CYP3A4 were used to train the models. Using these models, a synthetic majority rules consensus classifier was implemented, while the confidence of estimation was assigned following the percent agreement strategy. The classifier was applied to a testing set of 120 inhibitors not included in the development of the models. Five compounds of the test set, including known strong inhibitors dalfopristin and tioconazole, were classified as probable potent inhibitors of CYP3A4. Other known strong inhibitors, such as lopinavir, oltipraz, quercetin, raloxifene, and troglitazone, were among 18 compounds classified as plausible potent inhibitors of CYP3A4. The consensus estimation of inhibition potency is expected to aid in the nomination of pharmaceuticals, dietary supplements, environmental pollutants, and occupational and other chemicals for in-depth evaluation of the CYP3A4 inhibitory activity. It may serve also as an estimate of chemical interactions via CYP3A4 metabolic pharmacokinetic pathways occurring through polypharmacy and nutritional and environmental exposures to chemical mixtures.
Molecules 01/2012; 17(3):3407-60. · 2.39 Impact Factor
-
Brooks McPhail,
Yunfeng Tie,
Huixiao Hong,
Bruce A Pearce,
Laura K Schnackenberg,
Weigong Ge,
Luis G Valerio, James C Fuscoe,
Weida Tong,
Dan A Buzatu,
Jon G Wilkes,
Bruce A Fowler,
Eugene Demchuk,
Richard D Beger
[show abstract]
[hide abstract]
ABSTRACT: An interagency collaboration was established to model chemical interactions that may cause adverse health effects when an exposure to a mixture of chemicals occurs. Many of these chemicals--drugs, pesticides, and environmental pollutants--interact at the level of metabolic biotransformations mediated by cytochrome P450 (CYP) enzymes. In the present work, spectral data-activity relationship (SDAR) and structure-activity relationship (SAR) approaches were used to develop machine-learning classifiers of inhibitors and non-inhibitors of the CYP3A4 and CYP2D6 isozymes. The models were built upon 602 reference pharmaceutical compounds whose interactions have been deduced from clinical data, and 100 additional chemicals that were used to evaluate model performance in an external validation (EV) test. SDAR is an innovative modeling approach that relies on discriminant analysis applied to binned nuclear magnetic resonance (NMR) spectral descriptors. In the present work, both 1D ¹³C and 1D ¹⁵N-NMR spectra were used together in a novel implementation of the SDAR technique. It was found that increasing the binning size of 1D ¹³C-NMR and ¹⁵N-NMR spectra caused an increase in the tenfold cross-validation (CV) performance in terms of both the rate of correct classification and sensitivity. The results of SDAR modeling were verified using SAR. For SAR modeling, a decision forest approach involving from 6 to 17 Mold2 descriptors in a tree was used. Average rates of correct classification of SDAR and SAR models in a hundred CV tests were 60% and 61% for CYP3A4, and 62% and 70% for CYP2D6, respectively. The rates of correct classification of SDAR and SAR models in the EV test were 73% and 86% for CYP3A4, and 76% and 90% for CYP2D6, respectively. Thus, both SDAR and SAR methods demonstrated a comparable performance in modeling a large set of structurally diverse data. Based on unique NMR structural descriptors, the new SDAR modeling method complements the existing SAR techniques, providing an independent estimator that can increase confidence in a structure-activity assessment. When modeling was applied to hazardous environmental chemicals, it was found that up to 20% of them may be substrates and up to 10% of them may be inhibitors of the CYP3A4 and CYP2D6 isoforms. The developed models provide a rare opportunity for the environmental health branch of the public health service to extrapolate to hazardous chemicals directly from human clinical data. Therefore, the pharmacological and environmental health branches are both expected to benefit from these reported models.
Molecules 01/2012; 17(3):3383-406. · 2.39 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The effects of 12-week exposure to zidovudine (AZT) at 400, 500, and 600 mg/kg/d were examined on expression of 542 mitochondria-related genes and mitochondrial DNA (mtDNA) copy number in the liver of male and female B6C3F(1) mice to understand mitochondrial role in sex-related differences in development of lactic acidosis. Plasma lactate levels and hematologic parameters were also examined. Results indicated increased red blood cell (RBC) count in vehicle-treated controls, whereas a dose-related decline in the RBC count was noted in AZT-treated mice compared to the basal levels before treatments began. These decreases were associated with significant dose-related increases in mean corpuscular volume and mean corpuscular hemoglobin levels. This effect was greater in AZT-treated females compared to males. In both sexes, 12-week AZT or vehicle exposure significantly reduced plasma lactate levels compared to the basal levels. Results also showed modest, but significant, changes in the expression of genes associated with apoptosis and lipid metabolism at 600 mg/kg/d AZT. Neither drug nor sex influenced hepatic mtDNA copy number. Altogether, 12-week AZT exposure as high as 600 mg/kg/d did not impair hepatic mitochondria or induce lactic acidosis in B6C3F(1) mice. However, AZT-mediated hematologic toxicity appeared to be greater in females compared to males.
AIDS research and treatment 01/2012; 2012:317695.
-
[show abstract]
[hide abstract]
ABSTRACT: MicroRNAs (MiRNAs) are small noncoding RNAs that function as regulators of gene expression to control cell growth and differentiation. In this study, we analyzed miRNA and mRNA expression in the livers of rats treated with a carcinogenic dose of comfrey (Symphytum officinale) for 12 weeks. Groups of six rats were fed a normal diet or a diet containing 8% comfrey root. The animals were sacrificed 1 day after the last treatment and the livers were isolated for miRNA expression analysis using LC Sciences miRNA microarrays and for mRNA expression analysis using Affymetrix rat genome microarrays. MiRNA expression levels were significantly changed by comfrey treatment. The treated samples were separated clearly from the control samples in both principal component analysis (PCA) and hierarchical clustering analysis (HCA). Quantitative measurements of seven miRNAs using TaqMan real-time PCR were consistent with the microarray results in terms of fold-change and the direction of the change in expression. Forty-five miRNAs (P < 0.01) and 1,921 mRNAs (q = 0) were significantly changed by comfrey treatment. Using a target prediction algorithm, 434 differentially expressed genes (DEGs) were predicted to be targeted by the differentially expressed miRNAs (DEMs). The DEM-targeted DEGs were more likely to be involved in carcinogenesis than the DEGs that were not targeted by the DEMs. The nontargeted DEGs were enriched in noncancer-related biological processes. Our data suggest that comfrey may exert its carcinogenic effects by disturbing miRNA expression resulting in altered mRNA levels of the DEM-targeted genes that are functionally associated with carcinogenesis.
Environmental and Molecular Mutagenesis 03/2011; 52(6):469-78. · 3.71 Impact Factor
-
Wen Zou,
Sufian F Al-Khaldi,
William S Branham,
Tao Han, James C Fuscoe,
Jing Han,
Steven L Foley,
Joshua Xu,
Hong Fang,
Carl E Cerniglia,
Rajesh Nayak
[show abstract]
[hide abstract]
ABSTRACT: Rapid, accurate and inexpensive analysis of the disease-causing potential of foodborne pathogens is an important consideration in food safety and biodefense, particularly in developing countries. The objective of this study is to demonstrate the use of a robust and inexpensive microarray platform to assay the virulence gene profiles in Salmonella from food and/or the food animal environment, and then use ArrayTrack™ for data analysis.
The spotted array consisted of 69 selected Salmonella-specific virulence gene probes (65bp each). These probes were printed on poly-L-lysine-coated slides. Genomic DNA was digested with Sau3AI, labeled with Cy3 dye, hybridized to the gene probes, and the images were captured and analyzed by GenePix 4000B and ArrayTrack™, a free software developed by Food and Drug Administration (FDA) researchers.
Nearly 58% of the virulence-associated genes tested were present in all Salmonella strains tested. In general, genes belonging to inv, pip, prg, sic, sip, spa or ttr families were detected in more than 90% of the isolates, while the iacP, avrA, invH, rhuM, sirA, sopB, sopE or sugR genes were detected in 40 to 80% of the isolates. The gene variability was independent of the Salmonella serotype.
This hybridization array presents an accurate and cost-effective method for evaluating the disease-causing potential of Salmonella in outbreak investigations by targeting a selective set of Salmonella-associated virulence genes.
The Journal of Infection in Developing Countries 01/2011; 5(2):94-105. · 1.19 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Drug-induced liver injury (DILI) is the primary adverse event that results in the withdrawal of drugs from the market and a frequent reason for the failure of drug candidates in the pre-clinical or clinical phases of drug development. This paper presents an approach for identifying potential liver toxicity genomic biomarkers from a liver toxicity biomarker study involving the paired compounds entacapone ("non-liver toxic drug") and tolcapone ("hepatotoxic drug"). Molecular analysis of the rat liver and plasma samples, combined with statistical analysis, revealed many similarities and differences between the in vivo biochemical effects of the two drugs. Six hundred and ninety-five genes and 61 pathways were selected based on the classification scheme. Of the 61 pathways, 5 were specific to treatment with tolcapone. Two of the 12 animals in the tolcapone group were found to have high ALT, AST, or TBIL levels. The gene Vars2 (valyl-tRNA synthetase 2) was identified in both animals and the pathway to which it belongs, the aminoacyl-tRNA biosynthesis pathway, was one of the three most significant tolcapone-specific pathways identified.
International Journal of Molecular Sciences 01/2011; 12(7):4609-24. · 2.60 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: A methyl-deficient diet induces liver injury similar to human nonalcoholic steatohepatitis, one of the main risk factors for the development of hepatocellular carcinoma. Previous studies have demonstrated that this diet perturbs DNA methylation by causing a profound loss of global cytosine methylation, predominantly at heavily methylated repetitive sequences. However, whether methyl deficiency affects the methylation status of gene promoters has not been explored.
Mouse gene expression and CpG island microarrays were used to characterize the gene expression and CpG island methylation profiles in the livers of C57BL/6J mice fed a methyl-deficient diet. We detected 164 genes that were differentially expressed and exhibited an inverse relationship between the gene expression and the extent of CpG island methylation. Furthermore, these genes were associated with altered lipid and glucose metabolism, DNA damage and repair, apoptosis, the development of fibrosis, and liver tissue remodeling. Although there were both increased and decreased levels of CpG island methylation, the number of hypomethylated genes was substantially greater than the number of hypermethylated genes.
The results this study demonstrate that pairing methylation profiles with gene expression profiles is a powerful approach to identify dysregulated high-priority fundamental pathophysiological pathways associated with disease development.
Molecular Nutrition & Food Research 10/2010; 55(3):411-8. · 4.30 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Comfrey has been consumed by humans as a vegetable and a tea and used as an herbal medicine for more than 2000 years. Comfrey, however, produces hepatotoxicity in livestock and humans and carcinogenicity in experimental animals. Comfrey contains as many as 14 pyrrolizidine alkaloids (PA), including 7-acetylintermedine, 7-acetyllycopsamine, echimidine, intermedine, lasiocarpine, lycopsamine, myoscorpine, symlandine, symphytine, and symviridine. The mechanisms underlying comfrey-induced genotoxicity and carcinogenicity are still not fully understood. The available evidence suggests that the active metabolites of PA in comfrey interact with DNA in liver endothelial cells and hepatocytes, resulting in DNA damage, mutation induction, and cancer development. Genotoxicities attributed to comfrey and riddelliine (a representative genotoxic PA and a proven rodent mutagen and carcinogen) are discussed in this review. Both of these compounds induced similar profiles of 6,7-dihydro-7-hydroxy-1-hydroxymethyl-5H-pyrrolizine (DHP)-derived DNA adducts and similar mutation spectra. Further, the two agents share common mechanisms of drug metabolism and carcinogenesis. Overall, comfrey is mutagenic in liver, and PA contained in comfrey appear to be responsible for comfrey-induced toxicity and tumor induction.
Journal of Toxicology and Environmental Health Part B 10/2010; 13(7-8):509-26. · 4.72 Impact Factor
-
Leming Shi,
Gregory Campbell,
Wendell D Jones,
Fabien Campagne,
Zhining Wen,
Stephen J Walker,
Zhenqiang Su,
Tzu-Ming Chu,
Federico M Goodsaid,
Lajos Pusztai, [......],
Lun Yang,
Xiao Zeng,
Jialu Zhang,
Li Zhang,
Min Zhang,
Chen Zhao,
Raj K Puri,
Uwe Scherf,
Weida Tong,
Russell D Wolfinger
[show abstract]
[hide abstract]
ABSTRACT: Gene expression data from microarrays are being applied to predict preclinical and clinical endpoints, but the reliability of these predictions has not been established. In the MAQC-II project, 36 independent teams analyzed six microarray data sets to generate predictive models for classifying a sample with respect to one of 13 endpoints indicative of lung or liver toxicity in rodents, or of breast cancer, multiple myeloma or neuroblastoma in humans. In total, >30,000 models were built using many combinations of analytical methods. The teams generated predictive models without knowing the biological meaning of some of the endpoints and, to mimic clinical reality, tested the models on data that had not been used for training. We found that model performance depended largely on the endpoint and team proficiency and that different approaches generated models of similar performance. The conclusions and recommendations from MAQC-II should be useful for regulatory agencies, study committees and independent investigators that evaluate methods for global gene expression analysis.
Nature Biotechnology 08/2010; 28(8):827-38. · 29.50 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The field of toxicogenomics, which is becoming an important sub-discipline of toxicology, resulted from the natural convergence of the field of conventional toxicological research and the emergent field of functional genomics. One technology that has played a significant role in the field of toxicogenomics (in addition to many others) is the gene expression microarray. In this chapter, the authors provide an example of the application of gene expression microarrays to the field of toxicogenomics by detailing the strategy that was used for obtaining, analyzing, and interpreting gene expression data generated from RNA isolated from the liver of toxicant-exposed rats.
Methods in molecular biology (Clifton, N.J.) 01/2010; 597:227-41.
-
[show abstract]
[hide abstract]
ABSTRACT: Age- and sex-related susceptibility to adverse drug reactions and disease is a key concern in understanding drug safety and disease progression. We hypothesize that the underlying suite of hepatic genes expressed at various life cycle stages will impact susceptibility to adverse drug reactions. Understanding the basal liver gene expression patterns is a necessary first step in addressing this hypothesis and will inform our assessments of adverse drug reactions as the liver plays a central role in drug metabolism and biotransformation. Untreated male and female F344 rats were sacrificed at 2, 5, 6, 8, 15, 21, 52, 78, and 104 weeks of age. Liver tissues were collected for histology and gene expression analysis. Whole-genome rat microarrays were used to query global expression profiles.
An initial list of differentially expressed genes was selected using criteria based upon p-value (p < 0.05) and fold-change (+/- 1.5). Three dimensional principal component analyses revealed differences between males and females beginning at 2 weeks with more divergent profiles beginning at 5 weeks. The greatest sex-differences were observed between 8 and 52 weeks before converging again at 104 weeks. K-means clustering identified groups of genes that displayed age-related patterns of expression. Various adult aging-related clusters represented gene pathways related to xenobiotic metabolism, DNA damage repair, and oxidative stress.
These results suggest an underlying role for genes in specific clusters in potentiating age- and sex-related differences in susceptibility to adverse health effects. Furthermore, such a comprehensive picture of life cycle changes in gene expression deepens our understanding and informs the utility of liver gene expression biomarkers.
BMC Genomics 01/2010; 11:675. · 4.07 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The rapid advances in proteomic analyses coupled with the completion of multiple genomes have led to an increased demand for determining protein functions. The first step is classification or prediction into families. A method was developed for the prediction of protein family based only on protein sequence using support vector machine (SVM) models. In these models, the amino acids were classified into three categories (apolar, polar, and charged). Consecutive fragments ranging from one to five were annotated by amino acid type to define the protein features of each protein. SVM models were constructed based on the protein features of a training set of proteins and then examined with an independent set of proteins. The approach was tested for 20 protein families from the iProClass database of Protein Information Resources (PIR). For two-class SVM models, an average prediction accuracy of 0.9985 was achieved, while for multi-class SVM models an accuracy of 0.9941 was achieved. This study demonstrates that SVM based methods can accurately recognize and predict the protein family to which a sequence belongs based solely on its primary amino acid sequence.
Journal of computational biology: a journal of computational molecular cell biology 12/2009; 16(12):1671-88. · 1.69 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Genome-wide association studies (GWAS) use dense maps of single nucleotide polymorphisms (SNPs) that cover the entire human genome to search for genetic markers with different allele frequencies between cases and controls. Given the complexity of GWAS, it is not surprising that only a small portion of associated SNPs in the initial GWAS results were successfully replicated in the same populations. Each of the steps in a GWAS has the potential to generate spurious associations. In addition, there are batch effects in the genotyping experiments and in genotype calling that can cause both Type I and Type II errors. Decreasing or eliminating the various sources of spurious associations and batch effects is vital for reliably translating GWAS findings to clinical practice and personalized medicine. Here we review and discuss the variety of sources of spurious associations and batch effects in GWAS and provide possible solutions to the problems.
11/2009: pages 191 - 201; , ISBN: 9780470685983
-
[show abstract]
[hide abstract]
ABSTRACT: Usnic acid is a lichen metabolite used as a weight-loss dietary supplement due to its uncoupling action on mitochondria. However, its use has been associated with severe liver disorders in some individuals. Animal studies conducted thus far evaluated the effects of usnic acid on mitochondria primarily by measuring the rate of oxygen consumption and/or ATP generation. To obtain further insight into usnic acid-mediated effects on mitochondria, we examined the expression levels of 542 genes associated with mitochondrial structure and functions in liver of B6C3F(1) female mice using a mitochondria-specific microarray. Beginning at 8 weeks of age, mice received usnic acid at 0, 60, 180, and 600 ppm in ground, irradiated 5LG6 diet for 14 days. Microarray analysis showed a significant effect of usnic acid on the expression of several genes only at the highest dose of 600 ppm. A prominent finding of the study was a significant induction of genes associated with complexes I through IV of the electron transport chain. Moreover, several genes involved in fatty acid oxidation, the Krebs cycle, apoptosis, and membrane transporters were over-expressed. Usnic acid is a lipophilic weak acid that can diffuse through mitochondrial membranes and cause a proton leak (uncoupling). The up-regulation of complexes I-IV may be a compensatory mechanism to maintain the proton gradient across the mitochondrial inner membrane. In addition, induction of fatty acid oxidation and the Krebs cycle may be an adaptive response to uncoupling of mitochondria.
Mitochondrion 03/2009; 9(2):149-58. · 3.62 Impact Factor
-
Robert N McBurney,
Wade M Hines,
Linda S Von Tungeln,
Laura K Schnackenberg,
Richard D Beger,
Carrie L Moland,
Tao Han, James C Fuscoe,
Ching-Wei Chang,
James J Chen, [......],
Armin Graber,
Yu Guo,
Peter J Juhasz,
Tricin Y Li,
Moira D Lynch,
Nicole M Morel,
Thomas N Plasterer,
Edward J Takach,
Chenhui Zeng,
Frederick A Beland
[show abstract]
[hide abstract]
ABSTRACT: Drug-induced liver injury (DILI) is the primary adverse event that results in withdrawal of drugs from the market and a frequent reason for the failure of drug candidates in development. The Liver Toxicity Biomarker Study (LTBS) is an innovative approach to investigate DILI because it compares molecular events produced in vivo by compound pairs that (a) are similar in structure and mechanism of action, (b) are associated with few or no signs of liver toxicity in preclinical studies, and (c) show marked differences in hepatotoxic potential. The LTBS is a collaborative preclinical research effort in molecular systems toxicology between the National Center for Toxicological Research and BG Medicine, Inc., and is supported by seven pharmaceutical companies and three technology providers. In phase I of the LTBS, entacapone and tolcapone were studied in rats to provide results and information that will form the foundation for the design and implementation of phase II. Molecular analysis of the rat liver and plasma samples combined with statistical analyses of the resulting datasets yielded marker analytes, illustrating the value of the broad-spectrum, molecular systems analysis approach to studying pharmacological or toxicological effects.
Toxicologic Pathology 02/2009; 37(1):52-64. · 1.91 Impact Factor
