Lex-SVM: exploring the potential of exon expression profiling for disease classification.

PubMed

Yuan, Xiongying; Zhao, Yi; Liu, Changning; Bu, Dongbo

2011-04-01

Exon expression profiling technologies, including exon arrays and RNA-Seq, measure the abundance of every exon in a gene. Compared with gene expression profiling technologies like 3' array, exon expression profiling technologies could detect alterations in both transcription and alternative splicing, therefore they are expected to be more sensitive in diagnosis. However, exon expression profiling also brings higher dimension, more redundancy, and significant correlation among features. Ignoring the correlation structure among exons of a gene, a popular classification method like L1-SVM selects exons individually from each gene and thus is vulnerable to noise. To overcome this limitation, we present in this paper a new variant of SVM named Lex-SVM to incorporate correlation structure among exons and known splicing patterns to promote classification performance. Specifically, we construct a new norm, ex-norm, including our prior knowledge on exon correlation structure to regularize the coefficients of a linear SVM. Lex-SVM can be solved efficiently using standard linear programming techniques. The advantage of Lex-SVM is that it can select features group-wisely, force features in a subgroup to take equal weihts and exclude the features that contradict the majority in the subgroup. Experimental results suggest that on exon expression profile, Lex-SVM is more accurate than existing methods. Lex-SVM also generates a more compact model and selects genes more consistently in cross-validation. Unlike L1-SVM selecting only one exon in a gene, Lex-SVM assigns equal weights to as many exons in a gene as possible, lending itself easier for further interpretation.

University of Texas Southwestern Medical Center (UTSW): Functional Signature Ontology Tool: Triplicate Measurements of Reporter Gene Expression in Response to Individual Genetic and Chemical Perturbations in HCT116 Cells | Office of Cancer Genomics

Cancer.gov

The goal of this project is to use an eight-gene expression profile to define functional signatures for small molecules and natural products with heretofore undefined mechanism of action. Two genes in the eight gene set are used as internal controls and do not vary across gene expression array data collected from the public domain. The remaining six genes are found to vary independently across a large collection of publically available gene expression array datasets.  Read the abstract

Differential gene expression in anterior pituitary glands from anestrous and cycling postpartum beef cows

USDA-ARS?s Scientific Manuscript database

Oligionucleotide microarrays (GeneChip Bovine Genome Arrays, Affymetrix Inc., Santa Clara, CA) were used to evaluate gene expression profiles in anterior pituitary glands collected from 4 anestrous and 4 cycling postpartum primiparous beef cows to provide insight into genes associated with transitio...

A functional gene array for detection of bacterial virulence elements

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jaing, C

2007-11-01

We report our development of the first of a series of microarrays designed to detect pathogens with known mechanisms of virulence and antibiotic resistance. By targeting virulence gene families as well as genes unique to specific biothreat agents, these arrays will provide important data about the pathogenic potential and drug resistance profiles of unknown organisms in environmental samples. To validate our approach, we developed a first generation array targeting genes from Escherichia coli strains K12 and CFT073, Enterococcus faecalis and Staphylococcus aureus. We determined optimal probe design parameters for microorganism detection and discrimination, measured the required target concentration, and assessedmore » tolerance for mismatches between probe and target sequences. Mismatch tolerance is a priority for this application, due to DNA sequence variability among members of gene families. Arrays were created using the NimbleGen Maskless Array Synthesizer at Lawrence Livermore National Laboratory. Purified genomic DNA from combinations of one or more of the four target organisms, pure cultures of four related organisms, and environmental aerosol samples with spiked-in genomic DNA were hybridized to the arrays. Based on the success of this prototype, we plan to design further arrays in this series, with the goal of detecting all known virulence and antibiotic resistance gene families in a greatly expanded set of organisms.« less

Transcript Profiling of Common Bean (Phaseolus vulgaris) Using the GeneChip Soybean Genome Array: Optimizing Analysis by Masking Biased Probes

USDA-ARS?s Scientific Manuscript database

Common bean (Phaseolus vulgaris) and soybean (Glycine max) both belong to the Phaseoleae tribe and share significant coding sequence homology. To evaluate the utility of the soybean GeneChip for transcript profiling of common bean, we hybridized cRNAs purified from nodule, leaf, and root of common b...

Novel molecular subtypes of serous and endometrioid ovarian cancer linked to clinical outcome.

PubMed

Tothill, Richard W; Tinker, Anna V; George, Joshy; Brown, Robert; Fox, Stephen B; Lade, Stephen; Johnson, Daryl S; Trivett, Melanie K; Etemadmoghadam, Dariush; Locandro, Bianca; Traficante, Nadia; Fereday, Sian; Hung, Jillian A; Chiew, Yoke-Eng; Haviv, Izhak; Gertig, Dorota; DeFazio, Anna; Bowtell, David D L

2008-08-15

The study aim to identify novel molecular subtypes of ovarian cancer by gene expression profiling with linkage to clinical and pathologic features. Microarray gene expression profiling was done on 285 serous and endometrioid tumors of the ovary, peritoneum, and fallopian tube. K-means clustering was applied to identify robust molecular subtypes. Statistical analysis identified differentially expressed genes, pathways, and gene ontologies. Laser capture microdissection, pathology review, and immunohistochemistry validated the array-based findings. Patient survival within k-means groups was evaluated using Cox proportional hazards models. Class prediction validated k-means groups in an independent dataset. A semisupervised survival analysis of the array data was used to compare against unsupervised clustering results. Optimal clustering of array data identified six molecular subtypes. Two subtypes represented predominantly serous low malignant potential and low-grade endometrioid subtypes, respectively. The remaining four subtypes represented higher grade and advanced stage cancers of serous and endometrioid morphology. A novel subtype of high-grade serous cancers reflected a mesenchymal cell type, characterized by overexpression of N-cadherin and P-cadherin and low expression of differentiation markers, including CA125 and MUC1. A poor prognosis subtype was defined by a reactive stroma gene expression signature, correlating with extensive desmoplasia in such samples. A similar poor prognosis signature could be found using a semisupervised analysis. Each subtype displayed distinct levels and patterns of immune cell infiltration. Class prediction identified similar subtypes in an independent ovarian dataset with similar prognostic trends. Gene expression profiling identified molecular subtypes of ovarian cancer of biological and clinical importance.

A systems biology approach using transcriptomic data reveals genes and pathways in porcine skeletal muscle affected by dietary lysine

USDA-ARS?s Scientific Manuscript database

Meeting the increasing market demands for pork products requires improvement of the feed efficiency of growing pigs. The use of Affymetrix Porcine Gene 1.0 ST array containing 19,211 genes in this study provides a comprehensive gene expression profile of skeletal muscle of finishing pigs in response...

An Array-Based Analysis of MicroRNA Expression Comparing Matched Frozen and Formalin-Fixed Paraffin-Embedded Human Tissue Samples

PubMed Central

Zhang, Xiao; Chen, Jiamin; Radcliffe, Tom; LeBrun, Dave P.; Tron, Victor A.; Feilotter, Harriet

2008-01-01

MicroRNAs (miRNAs) are small, noncoding RNAs that suppress gene expression at the posttranscriptional level via an antisense RNA-RNA interaction. miRNAs used for array-based profiling are generally purified from either snap-frozen or fresh samples. Because tissues found in most pathology departments are available only in formalin-fixed and paraffin-embedded (FFPE) states, we sought to evaluate miRNA derived from FFPE samples for microarray analysis. In this study, miRNAs extracted from matched snap-frozen and FFPE samples were profiled using the Agilent miRNA array platform (Agilent, Santa Clara, CA). Each miRNA sample was hybridized to arrays containing probes interrogating 470 human miRNAs. Seven cases were compared in either duplicate or triplicate. Intrachip and interchip analyses demonstrated that the processes of miRNA extraction, labeling, and hybridization from both frozen and FFPE samples are highly reproducible and add little variation to the results; technical replicates showed high correlations with one another (Kendall tau, 0.722 to 0.853; Spearman rank correlation coefficient, 0.891 to 0.954). Our results showed consistent high correlations between matched frozen and FFPE samples (Kendall tau, 0.669 to 0.815; Spearman rank correlation coefficient, 0.847 to 0.948), supporting the use of FFPE-derived miRNAs for array-based, gene expression profiling. PMID:18832457

Qualitative assessment of gene expression in affymetrix genechip arrays

NASA Astrophysics Data System (ADS)

Nagarajan, Radhakrishnan; Upreti, Meenakshi

2007-01-01

Affymetrix Genechip microarrays are used widely to determine the simultaneous expression of genes in a given biological paradigm. Probes on the Genechip array are atomic entities which by definition are randomly distributed across the array and in turn govern the gene expression. In the present study, we make several interesting observations. We show that there is considerable correlation between the probe intensities across the array which defy the independence assumption. While the mechanism behind such correlations is unclear, we show that scaling behavior and the profiles of perfect match (PM) as well as mismatch (MM) probes are similar and immune-to-background subtraction. We believe that the observed correlations are possibly an outcome of inherent non-stationarities or patchiness in the array devoid of biological significance. This is demonstrated by inspecting their scaling behavior and profiles of the PM and MM probe intensities obtained from publicly available Genechip arrays from three eukaryotic genomes, namely: Drosophila melanogaster (fruit fly), Homo sapiens (humans) and Mus musculus (house mouse) across distinct biological paradigms and across laboratories, with and without background subtraction. The fluctuation functions were estimated using detrended fluctuation analysis (DFA) with fourth-order polynomial detrending. The results presented in this study provide new insights into correlation signatures of PM and MM probe intensities and suggests the choice of DFA as a tool for qualitative assessment of Affymetrix Genechip microarrays prior to their analysis. A more detailed investigation is necessary in order to understand the source of these correlations.

Gene expression patterns in rainbow trout, Oncorhynchus mykiss, exposed to a suite of model toxicants

PubMed Central

Hook, Sharon E.; Skillman, Ann D.; Small, Jack A.; Schultz, Irvin R.

2008-01-01

The increased availability and use of DNA microarrays has allowed the characterization of gene expression patterns associated with exposure to different toxicants. An important question is whether toxicant induced changes in gene expression in fish are sufficiently diverse to allow for identification of specific modes of action and/or specific contaminants. In theory, each class of toxicant may generate a gene expression profile unique to its mode of toxic action. In this study, isogenic (cloned) rainbow trout Oncorhynchus mykiss were exposed to sublethal levels of a series of model toxicants with varying modes of action, including ethynylestradiol (xeno-estrogen), 2,2,4,4′-tetrabromodiphenyl ether (BDE-47, thyroid active), diquat (oxidant stressor), chromium VI, and benzo[a]pyrene (BaP) for a period of 1–3 weeks. An additional experiment measured trenbolone (anabolic steroid; model androgen) induced gene expression changes in sexually mature female trout. Following exposure, fish were euthanized, livers removed and RNA extracted. Fluorescently labeled cDNA were generated and hybridized against a commercially available Atlantic Salmon/Trout array (GRASP project, University of Victoria) spotted with 16,000 cDNA’s. The slides were scanned to measure abundance of a given transcript in each sample relative to controls. Data were analyzed via Genespring (Silicon Genetics) to identify a list of up- and downregulated genes, as well as to determine gene clustering patterns that can be used as “expression signatures”. The results indicate each toxicant exposure caused between 64 and 222 genes to be significantly altered in expression. Most genes exhibiting altered expression responded to only one of the toxicants and relatively few were co-expressed in multiple treatments. For example, BaP and Diquat, both of which exert toxicity via oxidative stress, upregulated 28 of the same genes, of over 100 genes altered by either treatment. Other genes associated with steroidogenesis, p450 and estrogen responsive genes appear to be useful for selectively identifying toxicant mode of action in fish, suggesting a link between gene expression profile and mode of toxicity. Our array results showed good agreement with quantitative real time polymerase chain reaction (qRT PCR), which demonstrates that the arrays are an accurate measure of gene expression. The specificity of the gene expression profile in response to a model toxicant, the link between genes with altered expression and mode of toxic action, and the consistency between array and qRT PCR results all suggest that cDNA microarrays have the potential to screen environmental contaminants for biomarkers and mode of toxic action. PMID:16488489

Gene expression patterns in rainbow trout, Oncorhynchus mykiss, exposed to a suite of model toxicants.

PubMed

Hook, Sharon E; Skillman, Ann D; Small, Jack A; Schultz, Irvin R

2006-05-25

The increased availability and use of DNA microarrays has allowed the characterization of gene expression patterns associated with exposure to different toxicants. An important question is whether toxicant induced changes in gene expression in fish are sufficiently diverse to allow for identification of specific modes of action and/or specific contaminants. In theory, each class of toxicant may generate a gene expression profile unique to its mode of toxic action. In this study, isogenic (cloned) rainbow trout Oncorhynchus mykiss were exposed to sublethal levels of a series of model toxicants with varying modes of action, including ethynylestradiol (xeno-estrogen), 2,2,4,4'-tetrabromodiphenyl ether (BDE-47, thyroid active), diquat (oxidant stressor), chromium VI, and benzo[a]pyrene (BaP) for a period of 1-3 weeks. An additional experiment measured trenbolone (anabolic steroid; model androgen) induced gene expression changes in sexually mature female trout. Following exposure, fish were euthanized, livers removed and RNA extracted. Fluorescently labeled cDNA were generated and hybridized against a commercially available Atlantic Salmon/Trout array (GRASP project, University of Victoria) spotted with 16,000 cDNA's. The slides were scanned to measure abundance of a given transcript in each sample relative to controls. Data were analyzed via Genespring (Silicon Genetics) to identify a list of up- and downregulated genes, as well as to determine gene clustering patterns that can be used as "expression signatures". The results indicate each toxicant exposure caused between 64 and 222 genes to be significantly altered in expression. Most genes exhibiting altered expression responded to only one of the toxicants and relatively few were co-expressed in multiple treatments. For example, BaP and Diquat, both of which exert toxicity via oxidative stress, upregulated 28 of the same genes, of over 100 genes altered by either treatment. Other genes associated with steroidogenesis, p450 and estrogen responsive genes appear to be useful for selectively identifying toxicant mode of action in fish, suggesting a link between gene expression profile and mode of toxicity. Our array results showed good agreement with quantitative real time polymerase chain reaction (qRT PCR), which demonstrates that the arrays are an accurate measure of gene expression. The specificity of the gene expression profile in response to a model toxicant, the link between genes with altered expression and mode of toxic action, and the consistency between array and qRT PCR results all suggest that cDNA microarrays have the potential to screen environmental contaminants for biomarkers and mode of toxic action.

Ebola virus infection induces irregular dendritic cell gene expression.

PubMed

Melanson, Vanessa R; Kalina, Warren V; Williams, Priscilla

2015-02-01

Filoviruses subvert the human immune system in part by infecting and replicating in dendritic cells (DCs). Using gene arrays, a phenotypic profile of filovirus infection in human monocyte-derived DCs was assessed. Monocytes from human donors were cultured in GM-CSF and IL-4 and were infected with Ebola virus Kikwit variant for up to 48 h. Extracted DC RNA was analyzed on SuperArray's Dendritic and Antigen Presenting Cell Oligo GEArray and compared to uninfected controls. Infected DCs exhibited increased expression of cytokine, chemokine, antiviral, and anti-apoptotic genes not seen in uninfected controls. Significant increases of intracellular antiviral and MHC I and II genes were also noted in EBOV-infected DCs. However, infected DCs failed to show any significant difference in co-stimulatory T-cell gene expression from uninfected DCs. Moreover, several chemokine genes were activated, but there was sparse expression of chemokine receptors that enabled activated DCs to home to lymph nodes. Overall, statistically significant expression of several intracellular antiviral genes was noted, which may limit viral load but fails to stop replication. EBOV gene expression profiling is of vital importance in understanding pathogenesis and devising novel therapeutic treatments such as small-molecule inhibitors.

Streptococcus pneumoniae Supragenome Hybridization Arrays for Profiling of Genetic Content and Gene Expression.

PubMed

Kadam, Anagha; Janto, Benjamin; Eutsey, Rory; Earl, Joshua P; Powell, Evan; Dahlgren, Margaret E; Hu, Fen Z; Ehrlich, Garth D; Hiller, N Luisa

2015-02-02

There is extensive genomic diversity among Streptococcus pneumoniae isolates. Approximately half of the comprehensive set of genes in the species (the supragenome or pangenome) is present in all the isolates (core set), and the remaining is unevenly distributed among strains (distributed set). The Streptococcus pneumoniae Supragenome Hybridization (SpSGH) array provides coverage for an extensive set of genes and polymorphisms encountered within this species, capturing this genomic diversity. Further, the capture is quantitative. In this manner, the SpSGH array allows for both genomic and transcriptomic analyses of diverse S. pneumoniae isolates on a single platform. In this unit, we present the SpSGH array, and describe in detail its design and implementation for both genomic and transcriptomic analyses. The methodology can be applied to construction and modification of SpSGH array platforms, as well to other bacterial species as long as multiple whole-genome sequences are available that collectively capture the vast majority of the species supragenome. Copyright © 2015 John Wiley & Sons, Inc.

Chemical-genetic profile analysis of five inhibitory compounds in yeast.

PubMed

Alamgir, Md; Erukova, Veronika; Jessulat, Matthew; Azizi, Ali; Golshani, Ashkan

2010-08-06

Chemical-genetic profiling of inhibitory compounds can lead to identification of their modes of action. These profiles can help elucidate the complex interactions between small bioactive compounds and the cell machinery, and explain putative gene function(s). Colony size reduction was used to investigate the chemical-genetic profile of cycloheximide, 3-amino-1,2,4-triazole, paromomycin, streptomycin and neomycin in the yeast Saccharomyces cerevisiae. These compounds target the process of protein biosynthesis. More than 70,000 strains were analyzed from the array of gene deletion mutant yeast strains. As expected, the overall profiles of the tested compounds were similar, with deletions for genes involved in protein biosynthesis being the major category followed by metabolism. This implies that novel genes involved in protein biosynthesis could be identified from these profiles. Further investigations were carried out to assess the activity of three profiled genes in the process of protein biosynthesis using relative fitness of double mutants and other genetic assays. Chemical-genetic profiles provide insight into the molecular mechanism(s) of the examined compounds by elucidating their potential primary and secondary cellular target sites. Our follow-up investigations into the activity of three profiled genes in the process of protein biosynthesis provided further evidence concerning the usefulness of chemical-genetic analyses for annotating gene functions. We termed these genes TAE2, TAE3 and TAE4 for translation associated elements 2-4.

A stochastic model for optimizing composite predictors based on gene expression profiles.

PubMed

Ramanathan, Murali

2003-07-01

This project was done to develop a mathematical model for optimizing composite predictors based on gene expression profiles from DNA arrays and proteomics. The problem was amenable to a formulation and solution analogous to the portfolio optimization problem in mathematical finance: it requires the optimization of a quadratic function subject to linear constraints. The performance of the approach was compared to that of neighborhood analysis using a data set containing cDNA array-derived gene expression profiles from 14 multiple sclerosis patients receiving intramuscular inteferon-beta1a. The Markowitz portfolio model predicts that the covariance between genes can be exploited to construct an efficient composite. The model predicts that a composite is not needed for maximizing the mean value of a treatment effect: only a single gene is needed, but the usefulness of the effect measure may be compromised by high variability. The model optimized the composite to yield the highest mean for a given level of variability or the least variability for a given mean level. The choices that meet this optimization criteria lie on a curve of composite mean vs. composite variability plot referred to as the "efficient frontier." When a composite is constructed using the model, it outperforms the composite constructed using the neighborhood analysis method. The Markowitz portfolio model may find potential applications in constructing composite biomarkers and in the pharmacogenomic modeling of treatment effects derived from gene expression endpoints.

Antibody profiling using a recombinant protein-based multiplex ELISA array accelerates recombinant vaccine development: Case study on red sea bream iridovirus as a reverse vaccinology model.

PubMed

Matsuyama, Tomomasa; Sano, Natsumi; Takano, Tomokazu; Sakai, Takamitsu; Yasuike, Motoshige; Fujiwara, Atushi; Kawato, Yasuhiko; Kurita, Jun; Yoshida, Kazunori; Shimada, Yukinori; Nakayasu, Chihaya

2018-05-03

Predicting antigens that would be protective is crucial for the development of recombinant vaccine using genome based vaccine development, also known as reverse vaccinology. High-throughput antigen screening is effective for identifying vaccine target genes, particularly for pathogens for which minimal antigenicity data exist. Using red sea bream iridovirus (RSIV) as a research model, we developed enzyme-linked immune sorbent assay (ELISA) based RSIV-derived 72 recombinant antigen array to profile antiviral antibody responses in convalescent Japanese amberjack (Seriola quinqueradiata). Two and three genes for which the products were unrecognized and recognized, respectively, by antibodies in convalescent serum were selected for recombinant vaccine preparation, and the protective effect was examined in infection tests using Japanese amberjack and greater amberjack (S. dumerili). No protection was provided by vaccines prepared from gene products unrecognized by convalescent serum antibodies. By contrast, two vaccines prepared from gene products recognized by serum antibodies induced protective immunity in both fish species. These results indicate that ELISA array screening is effective for identifying antigens that induce protective immune responses. As this method does not require culturing of pathogens, it is also suitable for identifying protective antigens to un-culturable etiologic agents. Copyright © 2018 Elsevier Ltd. All rights reserved.

Developmental transcriptional profiling reveals key insights into Triticeae reproductive development.

PubMed

Tran, Frances; Penniket, Carolyn; Patel, Rohan V; Provart, Nicholas J; Laroche, André; Rowland, Owen; Robert, Laurian S

2013-06-01

Despite their importance, there remains a paucity of large-scale gene expression-based studies of reproductive development in species belonging to the Triticeae. As a first step to address this deficiency, a gene expression atlas of triticale reproductive development was generated using the 55K Affymetrix GeneChip(®) wheat genome array. The global transcriptional profiles of the anther/pollen, ovary and stigma were analyzed at concurrent developmental stages, and co-expressed as well as preferentially expressed genes were identified. Data analysis revealed both novel and conserved regulatory factors underlying Triticeae floral development and function. This comprehensive resource rests upon detailed gene annotations, and the expression profiles are readily accessible via a web browser. © 2013 Her Majesty the Queen in Right of Canada as represented by the Minister of Agriculture and Agri-Food Canada.

Three gangliogliomas: results of GTG-banding, SKY, genome-wide high resolution SNP-array, gene expression and review of the literature.

PubMed

Xu, Li-Xin; Holland, Heidrun; Kirsten, Holger; Ahnert, Peter; Krupp, Wolfgang; Bauer, Manfred; Schober, Ralf; Mueller, Wolf; Fritzsch, Dominik; Meixensberger, Jürgen; Koschny, Ronald

2015-04-01

According to the World Health Organization gangliogliomas are classified as well-differentiated and slowly growing neuroepithelial tumors, composed of neoplastic mature ganglion and glial cells. It is the most frequent tumor entity observed in patients with long-term epilepsy. Comprehensive cytogenetic and molecular cytogenetic data including high-resolution genomic profiling (single nucleotide polymorphism (SNP)-array) of gangliogliomas are scarce but necessary for a better oncological understanding of this tumor entity. For a detailed characterization at the single cell and cell population levels, we analyzed genomic alterations of three gangliogliomas using trypsin-Giemsa banding (GTG-banding) and by spectral karyotyping (SKY) in combination with SNP-array and gene expression array experiments. By GTG and SKY, we could confirm frequently detected chromosomal aberrations (losses within chromosomes 10, 13 and 22; gains within chromosomes 5, 7, 8 and 12), and identify so far unknown genetic aberrations like the unbalanced non-reciprocal translocation t(1;18)(q21;q21). Interestingly, we report on the second so far detected ganglioglioma with ring chromosome 1. Analyses of SNP-array data from two of the tumors and respective germline DNA (peripheral blood) identified few small gains and losses and a number of copy-neutral regions with loss of heterozygosity (LOH) in germline and in tumor tissue. In comparison to germline DNA, tumor tissues did not show substantial regions with significant loss or gain or with newly developed LOH. Gene expression analyses of tumor-specific genes revealed similarities in the profile of the analyzed samples regarding different relevant pathways. Taken together, we describe overlapping but also distinct and novel genetic aberrations of three gangliogliomas. © 2014 Japanese Society of Neuropathology.

Altered retinal microRNA expression profiles in early diabetic retinopathy: an in silico analysis.

PubMed

Xiong, Fen; Du, Xinhua; Hu, Jianyan; Li, Tingting; Du, Shanshan; Wu, Qiang

2014-07-01

MicroRNAs (miRNAs) - as negative regulators of target genes - are associated with various human diseases, but their precise role(s) in diabetic retinopathy (DR) remains to be elucidated. The aim of this study was to elucidate the involvement of miRNAs in early DR using in silico analysis to explore their gene expression patterns. We used the streptozotocin (STZ)-induced diabetic rat to investigate the roles of miRNAs in early DR. Retinal miRNA expression profiles from diabetic versus healthy control rats were examined by miRNA array analysis. Based on several bioinformatic systems, specifically, gene ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, we identified signatures of the potential pathological processes, gene functions, and signaling pathways that are influenced by dysregulated miRNAs. We used quantitative real-time polymerase chain reaction (qRT-PCR) to validate six (i.e. those with significant changes in expression levels) of the 17 miRNAs that were detected in the miRNA array. We also describe the significant role of the miRNA-gene network, which is based on the interactions between miRNAs and target genes. GO analysis of the 17 miRNAs detected in the miRNA array analysis revealed the most prevalent miRNAs to be those related to biological processes, olfactory bulb development and axonogenesis. These miRNAs also exert significant influence on additional pathways, including the mitogen-activated protein and calcium signaling pathways. Six of the seventeen miRNAs were chosen for qRT-PCR validation. With the exception of a slight difference in miRNA-350, our results are in close agreement with the differential expressions detected by array analysis. This study, which describes miRNA expression during the early developmental phases of DR, revealed extensive miRNA interactions. Based on both their target genes and signaling pathways, we suggest that miRNAs perform critical regulatory functions during the early stages of DR evolution.

High resolution array CGH and gene expression profiling of alveolar soft part sarcoma

PubMed Central

Selvarajah, Shamini; Pyne, Saumyadipta; Chen, Eleanor; Sompallae, Ramakrishna; Ligon, Azra H.; Nielsen, Gunnlaugur P.; Dranoff, Glenn; Stack, Edward; Loda, Massimo; Flavin, Richard

2014-01-01

Purpose Alveolar soft part sarcoma (ASPS) is a soft tissue sarcoma with poor prognosis, and little molecular evidence for its origin, initiation and progression. The aim of this study was to elucidate candidate molecular pathways involved in tumor pathogenesis. Experimental Design We employed high-throughput array comparative genomic hybridization and cDNA-Mediated Annealing, Selection, Ligation, and Extension Assay to profile the genomic and expression signatures of primary and metastatic ASPS from 17 tumors derived from 11 patients. We used an integrative bioinformatics approach to elucidate the molecular pathways associated with ASPS progression. Fluorescence in situ hybridization was performed to validate the presence of the t(X;17)(p11.2;q25) ASPL-TFE3 fusion and hence confirm the aCGH observations. Results FISH analysis identified the ASPL-TFE3 fusion in all cases. ArrayCGH revealed a higher number of numerical aberrations in metastatic tumors relative to primaries, but failed to identify consistent alterations in either group. Gene expression analysis highlighted 1,063 genes which were differentially expressed between the two groups. Gene set enrichment analysis identified 16 enriched gene sets (p < 0.1) associated with differentially expressed genes. Notable among these were several stem cell gene expression signatures and pathways related to differentiation. In particular, the paired box transcription factor PAX6 was up-regulated in the primary tumors, along with several genes whose mouse orthologs have previously been implicated in Pax6-DNA binding during neural stem cell differentiation. Conclusion In addition to suggesting a tentative neural line of differentiation for ASPS, these results implicate transcriptional deregulation from fusion genes in the pathogenesis of ASPS. PMID:24493828

Gene expression profiles in chondrosarcoma cells subjected to cyclic stretching and hydrostatic pressure. A cDNA array study.

PubMed

Karjalainen, Hannu M; Sironen, Reijo K; Elo, Mika A; Kaarniranta, Kai; Takigawa, Masaharu; Helminen, Heikki J; Lammi, Mikko J

2003-01-01

Mechanical forces have a profound effect on cartilage tissue and chondrocyte metabolism. Strenuous loading inhibits the cellular metabolism, while optimal level of loading at correct frequency raises an anabolic response in chondrocytes. In this study, we used Atlas Human Cancer cDNA array to investigate mRNA expression profiles in human chondrosarcoma cells stretched 8% for 6 hours at a frequency of 0.5 Hz. In addition, cultures were exposed to continuous and cyclic (0.5 Hz) 5 MPa hydrostatic pressure. Cyclic stretch had a more profound effect on the gene expression profiles than 5 MPa hydrostatic pressure. Several genes involved with the regulation of cell cycle were increased in stretched cells, as well as mRNAs for PDGF-B, glucose-1-phosphate uridylyltransferase, Tiam1, cdc37 homolog, Gem, integrin alpha6, and matrix metalloproteinase-3. Among down-regulated genes were plakoglobin, TGF-alpha, retinoic acid receptor-alpha and Wnt8b. A smaller number of changes was detected after pressure treatments. Plakoglobin was increased under cyclic and continuous 5 MPa hydrostatic pressure, while mitogen-activated protein kinase-9, proliferating cell nuclear antigen, Rad6, CD9 antigen, integrins alphaE and beta8, and vimentin were decreased. Cyclic and continuous pressurization induces a number of specific changes. In conclusion, a different set of genes were affected by three different types of mechanical stimuli applied on chondrosarcoma cells.

Novel mouse model recapitulates genome and transcriptome alterations in human colorectal carcinomas.

PubMed

McNeil, Nicole E; Padilla-Nash, Hesed M; Buishand, Floryne O; Hue, Yue; Ried, Thomas

2017-03-01

Human colorectal carcinomas are defined by a nonrandom distribution of genomic imbalances that are characteristic for this disease. Often, these imbalances affect entire chromosomes. Understanding the role of these aneuploidies for carcinogenesis is of utmost importance. Currently, established transgenic mice do not recapitulate the pathognonomic genome aberration profile of human colorectal carcinomas. We have developed a novel model based on the spontaneous transformation of murine colon epithelial cells. During this process, cells progress through stages of pre-immortalization, immortalization and, finally, transformation, and result in tumors when injected into immunocompromised mice. We analyzed our model for genome and transcriptome alterations using ArrayCGH, spectral karyotyping (SKY), and array based gene expression profiling. ArrayCGH revealed a recurrent pattern of genomic imbalances. These results were confirmed by SKY. Comparing these imbalances with orthologous maps of human chromosomes revealed a remarkable overlap. We observed focal deletions of the tumor suppressor genes Trp53 and Cdkn2a/p16. High-level focal genomic amplification included the locus harboring the oncogene Mdm2, which was confirmed by FISH in the form of double minute chromosomes. Array-based global gene expression revealed distinct differences between the sequential steps of spontaneous transformation. Gene expression changes showed significant similarities with human colorectal carcinomas. Pathways most prominently affected included genes involved in chromosomal instability and in epithelial to mesenchymal transition. Our novel mouse model therefore recapitulates the most prominent genome and transcriptome alterations in human colorectal cancer, and might serve as a valuable tool for understanding the dynamic process of tumorigenesis, and for preclinical drug testing. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Using the methylome to identify aggressive Barrett’s esophagus — EDRN Public Portal

Cancer.gov

OVERALL STRATEGY: Our strategy will consist of using HumanMethylation450 arrays to identify methylation profiles and/or candidate methylated genes that distinguish BE from BE+LGD, BE+HGD and EAC (Aim 1). We will then assess whether these genes are predictive markers for aggressive BE (Aim 2)

Integrative Genomics Reveals Mechanisms of Copy Number Alterations Responsible for Transcriptional Deregulation in Colorectal Cancer

PubMed Central

Camps, Jordi; Nguyen, Quang Tri; Padilla-Nash, Hesed M.; Knutsen, Turid; McNeil, Nicole E.; Wangsa, Danny; Hummon, Amanda B.; Grade, Marian; Ried, Thomas; Difilippantonio, Michael J.

2016-01-01

To evaluate the mechanisms and consequences of chromosomal aberrations in colorectal cancer (CRC), we used a combination of spectral karyotyping, array comparative genomic hybridization (aCGH), and array-based global gene expression profiling on 31 primary carcinomas and 15 established cell lines. Importantly, aCGH showed that the genomic profiles of primary tumors are recapitulated in the cell lines. We revealed a preponderance of chromosome breakpoints at sites of copy number variants (CNVs) in the CRC cell lines, a novel mechanism of DNA breakage in cancer. The integration of gene expression and aCGH led to the identification of 157 genes localized within high-level copy number changes whose transcriptional deregulation was significantly affected across all of the samples, thereby suggesting that these genes play a functional role in CRC. Genomic amplification at 8q24 was the most recurrent event and led to the overexpression of MYC and FAM84B. Copy number dependent gene expression resulted in deregulation of known cancer genes such as APC, FGFR2, and ERBB2. The identification of only 36 genes whose localization near a breakpoint could account for their observed deregulated expression demonstrates that the major mechanism for transcriptional deregulation in CRC is genomic copy number changes resulting from chromosomal aberrations. PMID:19691111

Chemical-genetic profile analysis in yeast suggests that a previously uncharacterized open reading frame, YBR261C, affects protein synthesis

PubMed Central

Alamgir, Md; Eroukova, Veronika; Jessulat, Matthew; Xu, Jianhua; Golshani, Ashkan

2008-01-01

Background Functional genomics has received considerable attention in the post-genomic era, as it aims to identify function(s) for different genes. One way to study gene function is to investigate the alterations in the responses of deletion mutants to different stimuli. Here we investigate the genetic profile of yeast non-essential gene deletion array (yGDA, ~4700 strains) for increased sensitivity to paromomycin, which targets the process of protein synthesis. Results As expected, our analysis indicated that the majority of deletion strains (134) with increased sensitivity to paromomycin, are involved in protein biosynthesis. The remaining strains can be divided into smaller functional categories: metabolism (45), cellular component biogenesis and organization (28), DNA maintenance (21), transport (20), others (38) and unknown (39). These may represent minor cellular target sites (side-effects) for paromomycin. They may also represent novel links to protein synthesis. One of these strains carries a deletion for a previously uncharacterized ORF, YBR261C, that we term TAE1 for Translation Associated Element 1. Our focused follow-up experiments indicated that deletion of TAE1 alters the ribosomal profile of the mutant cells. Also, gene deletion strain for TAE1 has defects in both translation efficiency and fidelity. Miniaturized synthetic genetic array analysis further indicates that TAE1 genetically interacts with 16 ribosomal protein genes. Phenotypic suppression analysis using TAE1 overexpression also links TAE1 to protein synthesis. Conclusion We show that a previously uncharacterized ORF, YBR261C, affects the process of protein synthesis and reaffirm that large-scale genetic profile analysis can be a useful tool to study novel gene function(s). PMID:19055778

Chemical-genetic profile analysis in yeast suggests that a previously uncharacterized open reading frame, YBR261C, affects protein synthesis.

PubMed

Alamgir, Md; Eroukova, Veronika; Jessulat, Matthew; Xu, Jianhua; Golshani, Ashkan

2008-12-03

Functional genomics has received considerable attention in the post-genomic era, as it aims to identify function(s) for different genes. One way to study gene function is to investigate the alterations in the responses of deletion mutants to different stimuli. Here we investigate the genetic profile of yeast non-essential gene deletion array (yGDA, approximately 4700 strains) for increased sensitivity to paromomycin, which targets the process of protein synthesis. As expected, our analysis indicated that the majority of deletion strains (134) with increased sensitivity to paromomycin, are involved in protein biosynthesis. The remaining strains can be divided into smaller functional categories: metabolism (45), cellular component biogenesis and organization (28), DNA maintenance (21), transport (20), others (38) and unknown (39). These may represent minor cellular target sites (side-effects) for paromomycin. They may also represent novel links to protein synthesis. One of these strains carries a deletion for a previously uncharacterized ORF, YBR261C, that we term TAE1 for Translation Associated Element 1. Our focused follow-up experiments indicated that deletion of TAE1 alters the ribosomal profile of the mutant cells. Also, gene deletion strain for TAE1 has defects in both translation efficiency and fidelity. Miniaturized synthetic genetic array analysis further indicates that TAE1 genetically interacts with 16 ribosomal protein genes. Phenotypic suppression analysis using TAE1 overexpression also links TAE1 to protein synthesis. We show that a previously uncharacterized ORF, YBR261C, affects the process of protein synthesis and reaffirm that large-scale genetic profile analysis can be a useful tool to study novel gene function(s).

Using mummichog (Fundulus heteroclitus) arrays to monitor the effectiveness of remediation at a superfund site in Charleston, South Carolina, U.S.A.

PubMed

Roling, Jonathan A; Bain, Lisa J; Gardea-Torresdey, Jorge; Key, Peter B; Baldwin, William S

2007-06-01

We previously developed a cDNA array for mummichogs (Fundulus heteroclitus), an estuarine minnow, that is targeted for identifying differentially expressed genes from exposure to polycyclic aromatic hydrocarbons and several metals, including chromium. A chromium-contaminated Superfund site at Shipyard Creek in Charleston, South Carolina, USA, is undergoing remediation, providing us a unique opportunity to study the utility of arrays for monitoring the effectiveness of site remediation. Mummichogs were captured in Shipyard Creek in Charleston prior to remediation (2000) and after remediation began (2003 and 2005). Simultaneously, mummichogs were collected from a reference site at the Winyah Bay National Estuarine Research Reserve (NERR) in Georgetown, South Carolina, USA. The hepatic gene expression pattern of fish captured at Shipyard Creek in 2000 showed wide differences from the fish captured at NERR in 2000. Interestingly, as remediation progressed the gene expression pattern of mummichogs captured at Shipyard Creek became increasingly similar to those captured at NERR. The arrays acted as multidimensional biomarkers as the number of differentially expressed genes dropped from 22 in 2000 to four in 2003, and the magnitude of differential expression dropped from 3.2-fold in 2000 to no gene demonstrating a difference over 1.5-fold in 2003. Furthermore, the arrays indicated changes in the bioavailability of chromium caused by hydraulic dredging in the summer of 2005. This research is, to our knowledge, the first report using arrays as biomarkers for a weight-of-evidence hazard assessment and demonstrates that arrays can be used as multidimensional biomarkers to monitor site mitigation because the gene expression profile is associated with chromium bioavailability and body burden.

Informatic selection of a neural crest-melanocyte cDNA set for microarray analysis

PubMed Central

Loftus, S. K.; Chen, Y.; Gooden, G.; Ryan, J. F.; Birznieks, G.; Hilliard, M.; Baxevanis, A. D.; Bittner, M.; Meltzer, P.; Trent, J.; Pavan, W.

1999-01-01

With cDNA microarrays, it is now possible to compare the expression of many genes simultaneously. To maximize the likelihood of finding genes whose expression is altered under the experimental conditions, it would be advantageous to be able to select clones for tissue-appropriate cDNA sets. We have taken advantage of the extensive sequence information in the dbEST expressed sequence tag (EST) database to identify a neural crest-derived melanocyte cDNA set for microarray analysis. Analysis of characterized genes with dbEST identified one library that contained ESTs representing 21 neural crest-expressed genes (library 198). The distribution of the ESTs corresponding to these genes was biased toward being derived from library 198. This is in contrast to the EST distribution profile for a set of control genes, characterized to be more ubiquitously expressed in multiple tissues (P < 1 × 10−9). From library 198, a subset of 852 clustered ESTs were selected that have a library distribution profile similar to that of the 21 neural crest-expressed genes. Microarray analysis demonstrated the majority of the neural crest-selected 852 ESTs (Mel1 array) were differentially expressed in melanoma cell lines compared with a non-neural crest kidney epithelial cell line (P < 1 × 10−8). This was not observed with an array of 1,238 ESTs that was selected without library origin bias (P = 0.204). This study presents an approach for selecting tissue-appropriate cDNAs that can be used to examine the expression profiles of developmental processes and diseases. PMID:10430933

DNA Microarray Profiling Highlights Nrf2-Mediated Chemoprevention Targeted by Wasabi-Derived Isothiocyanates in HepG2 Cells.

PubMed

Trio, Phoebe Zapanta; Kawahara, Atsuyoshi; Tanigawa, Shunsuke; Sakao, Kozue; Hou, De-Xing

2017-01-01

6-MSITC and 6-MTITC are sulforaphane (SFN) analogs found in Japanese Wasabi. As we reported previously, Wasabi isothiocyanates (ITCs) are activators of Nrf2-antioxidant response element pathway, and also inhibitors of pro-inflammatory cyclooxygenase-2. This study is the first to assess the global changes in transcript levels by Wasabi ITCs, comparing with SFN, in HepG2 cells. We performed comparative gene expression profiling by treating HepG2 cells with ITCs, followed by DNA microarray analyses using HG-U133 plus 2.0 oligonucleotide array. Partial array data on selected gene products were confirmed by RT-PCR and Western blotting. Ingenuity Pathway Analysis (IPA) was used to identify functional subsets of genes and biologically significant network pathways. 6-MTITC showed the highest number of differentially altered (≥2 folds) gene expression, of which 114 genes were upregulated and 75 were downregulated. IPA revealed that Nrf2-mediated pathway, together with glutamate metabolism, is the common significantly modulated pathway across treatments. Interestingly, 6-MSITC exhibited the most potent effect toward Nrf2-mediated pathway. Our data suggest that 6-MSITC could exert chemopreventive role against cancer through its underlying antioxidant activity via the activation of Nrf2-mediated subsequent induction of cytoprotective genes.

Microarray expression profiling identifies genes with altered expression in HDL-deficient mice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Callow, Matthew J.; Dudoit, Sandrine; Gong, Elaine L.

2000-05-05

Based on the assumption that severe alterations in the expression of genes known to be involved in HDL metabolism may affect the expression of other genes we screened an array of over 5000 mouse expressed sequence tags (ESTs) for altered gene expression in the livers of two lines of mice with dramatic decreases in HDL plasma concentrations. Labeled cDNA from livers of apolipoprotein AI (apo AI) knockout mice, Scavenger Receptor BI (SR-BI) transgenic mice and control mice were co-hybridized to microarrays. Two-sample t-statistics were used to identify genes with altered expression levels in the knockout or transgenic mice compared withmore » the control mice. In the SR-BI group we found 9 array elements representing at least 5 genes to be significantly altered on the basis of an adjusted p value of less than 0.05. In the apo AI knockout group 8 array elements representing 4 genes were altered compared with the control group (p < 0.05). Several of the genes identified in the SR-BI transgenic suggest altered sterol metabolism and oxidative processes. These studies illustrate the use of multiple-testing methods for the identification of genes with altered expression in replicated microarray experiments of apo AI knockout and SR-BI transgenic mice.« less

Expression Profiling Smackdown: Human Transcriptome Array HTA 2.0 vs. RNA-Seq

PubMed Central

Palermo, Meghann; Driscoll, Heather; Tighe, Scott; Dragon, Julie; Bond, Jeff; Shukla, Arti; Vangala, Mahesh; Vincent, James; Hunter, Tim

2014-01-01

The advent of both microarray and massively parallel sequencing have revolutionized high-throughput analysis of the human transcriptome. Due to limitations in microarray technology, detecting and quantifying coding transcript isoforms, in addition to non-coding transcripts, has been challenging. As a result, RNA-Seq has been the preferred method for characterizing the full human transcriptome, until now. A new high-resolution array from Affymetrix, GeneChip Human Transcriptome Array 2.0 (HTA 2.0), has been designed to interrogate all transcript isoforms in the human transcriptome with >6 million probes targeting coding transcripts, exon-exon splice junctions, and non-coding transcripts. Here we compare expression results from GeneChip HTA 2.0 and RNA-Seq data using identical RNA extractions from three samples each of healthy human mesothelial cells in culture, LP9-C1, and healthy mesothelial cells treated with asbestos, LP9-A1. For GeneChip HTA 2.0 sample preparation, we chose to compare two target preparation methods, NuGEN Ovation Pico WTA V2 with the Encore Biotin Module versus Affymetrix's GeneChip WT PLUS with the WT Terminal Labeling Kit, on identical RNA extractions from both untreated and treated samples. These same RNA extractions were used for the RNA-Seq library preparation. All analyses were performed in Partek Genomics Suite 6.6. Expression profiles for control and asbestos-treated mesothelial cells prepared with NuGEN versus Affymetrix target preparation methods (GeneChip HTA 2.0) are compared to each other as well as to RNA-Seq results.

Magnesium supplementation, metabolic and inflammatory markers, and global genomic and proteomic profiling: a randomized, double-blind, controlled, crossover trial in overweight individuals.

PubMed

Chacko, Sara A; Sul, James; Song, Yiqing; Li, Xinmin; LeBlanc, James; You, Yuko; Butch, Anthony; Liu, Simin

2011-02-01

Dietary magnesium intake has been favorably associated with reduced risk of metabolic outcomes in observational studies; however, few randomized trials have introduced a systems-biology approach to explore molecular mechanisms of pleiotropic metabolic actions of magnesium supplementation. We examined the effects of oral magnesium supplementation on metabolic biomarkers and global genomic and proteomic profiling in overweight individuals. We undertook this randomized, crossover, pilot trial in 14 healthy, overweight volunteers [body mass index (in kg/m(2)) ≥25] who were randomly assigned to receive magnesium citrate (500 mg elemental Mg/d) or a placebo for 4 wk with a 1-mo washout period. Fasting blood and urine specimens were collected according to standardized protocols. Biochemical assays were conducted on blood specimens. RNA was extracted and subsequently hybridized with the Human Gene ST 1.0 array (Affymetrix, Santa Clara, CA). Urine proteomic profiling was analyzed with the CM10 ProteinChip array (Bio-Rad Laboratories, Hercules, CA). We observed that magnesium treatment significantly decreased fasting C-peptide concentrations (change: -0.4 ng/mL after magnesium treatment compared with +0.05 ng/mL after placebo treatment; P = 0.004) and appeared to decrease fasting insulin concentrations (change: -2.2 μU/mL after magnesium treatment compared with 0.0 μU/mL after placebo treatment; P = 0.25). No consistent patterns were observed across inflammatory biomarkers. Gene expression profiling revealed up-regulation of 24 genes and down-regulation of 36 genes including genes related to metabolic and inflammatory pathways such as C1q and tumor necrosis factor-related protein 9 (C1QTNF9) and pro-platelet basic protein (PPBP). Urine proteomic profiling showed significant differences in the expression amounts of several peptides and proteins after treatment. Magnesium supplementation for 4 wk in overweight individuals led to distinct changes in gene expression and proteomic profiling consistent with favorable effects on several metabolic pathways. This trial was registered at clinicaltrials.gov as NCT00737815.

Expression profiling analysis: Uncoupling protein 2 deficiency improves hepatic glucose, lipid profiles and insulin sensitivity in high-fat diet-fed mice by modulating expression of genes in peroxisome proliferator-activated receptor signaling pathway.

PubMed

Zhou, Mei-Cen; Yu, Ping; Sun, Qi; Li, Yu-Xiu

2016-03-01

Uncoupling protein 2 (UCP2), which was an important mitochondrial inner membrane protein associated with glucose and lipid metabolism, widely expresses in all kinds of tissues including hepatocytes. The present study aimed to explore the impact of UCP2 deficiency on glucose and lipid metabolism, insulin sensitivity and its effect on the liver-associated signaling pathway by expression profiling analysis. Four-week-old male UCP2-/- mice and UCP2+/+ mice were randomly assigned to four groups: UCP2-/- on a high-fat diet, UCP2-/- on a normal chow diet, UCP2+/+ on a high-fat diet and UCP2+/+ on a normal chow diet. The differentially expressed genes in the four groups on the 16th week were identified by Affymetrix gene array. The results of intraperitoneal glucose tolerance test and insulin tolerance showed that blood glucose and β-cell function were improved in the UCP2-/- group on high-fat diet. Enhanced insulin sensitivity was observed in the UCP2-/- group. The differentially expressed genes were mapped to 23 pathways (P < 0.05). We concentrated on the 'peroxisome proliferator-activated receptor (PPAR) signaling pathway' (P = 3.19 × 10(-11)), because it is closely associated with the regulation of glucose and lipid profiles. In the PPAR signaling pathway, seven genes (PPARγ, Dbi, Acsl3, Lpl, Me1, Scd1, Fads2) in the UCP2-/- mice were significantly upregulated. The present study used gene arrays to show that activity of the PPAR signaling pathway involved in the improvement of glucose and lipid metabolism in the liver of UCP2-deficient mice on a long-term high-fat diet. The upregulation of genes in the PPAR signaling pathway could explain our finding that UCP2 deficiency ameliorated insulin sensitivity. The manipulation of UCP2 protein expression could represent a new strategy for the prevention and treatment of diabetes.

Gene expression profiling and pathway analysis in MCF-7 and MDA-MB-231 human breast cancer cell lines treated with dioscin

USDA-ARS?s Scientific Manuscript database

The long-term goal of our study is to understand the genetic and epigenetic mechanisms of breast cancer metastasis in human and to discover new possible genetic markers for use in clinical practice. We have used microarray technology (Human OneArray microarray, phylanxbiotech.com) to compare gene ex...

Global Identification and Characterization of Transcriptionally Active Regions in the Rice Genome

PubMed Central

Stolc, Viktor; Deng, Wei; He, Hang; Korbel, Jan; Chen, Xuewei; Tongprasit, Waraporn; Ronald, Pamela; Chen, Runsheng; Gerstein, Mark; Wang Deng, Xing

2007-01-01

Genome tiling microarray studies have consistently documented rich transcriptional activity beyond the annotated genes. However, systematic characterization and transcriptional profiling of the putative novel transcripts on the genome scale are still lacking. We report here the identification of 25,352 and 27,744 transcriptionally active regions (TARs) not encoded by annotated exons in the rice (Oryza. sativa) subspecies japonica and indica, respectively. The non-exonic TARs account for approximately two thirds of the total TARs detected by tiling arrays and represent transcripts likely conserved between japonica and indica. Transcription of 21,018 (83%) japonica non-exonic TARs was verified through expression profiling in 10 tissue types using a re-array in which annotated genes and TARs were each represented by five independent probes. Subsequent analyses indicate that about 80% of the japonica TARs that were not assigned to annotated exons can be assigned to various putatively functional or structural elements of the rice genome, including splice variants, uncharacterized portions of incompletely annotated genes, antisense transcripts, duplicated gene fragments, and potential non-coding RNAs. These results provide a systematic characterization of non-exonic transcripts in rice and thus expand the current view of the complexity and dynamics of the rice transcriptome. PMID:17372628

BeadArray Expression Analysis Using Bioconductor

PubMed Central

Ritchie, Matthew E.; Dunning, Mark J.; Smith, Mike L.; Shi, Wei; Lynch, Andy G.

2011-01-01

Illumina whole-genome expression BeadArrays are a popular choice in gene profiling studies. Aside from the vendor-provided software tools for analyzing BeadArray expression data (GenomeStudio/BeadStudio), there exists a comprehensive set of open-source analysis tools in the Bioconductor project, many of which have been tailored to exploit the unique properties of this platform. In this article, we explore a number of these software packages and demonstrate how to perform a complete analysis of BeadArray data in various formats. The key steps of importing data, performing quality assessments, preprocessing, and annotation in the common setting of assessing differential expression in designed experiments will be covered. PMID:22144879

Tissue Gene Expression Analysis Using Arrayed Normalized cDNA Libraries

PubMed Central

Eickhoff, Holger; Schuchhardt, Johannes; Ivanov, Igor; Meier-Ewert, Sebastian; O'Brien, John; Malik, Arif; Tandon, Neeraj; Wolski, Eryk-Witold; Rohlfs, Elke; Nyarsik, Lajos; Reinhardt, Richard; Nietfeld, Wilfried; Lehrach, Hans

2000-01-01

We have used oligonucleotide-fingerprinting data on 60,000 cDNA clones from two different mouse embryonic stages to establish a normalized cDNA clone set. The normalized set of 5,376 clones represents different clusters and therefore, in almost all cases, different genes. The inserts of the cDNA clones were amplified by PCR and spotted on glass slides. The resulting arrays were hybridized with mRNA probes prepared from six different adult mouse tissues. Expression profiles were analyzed by hierarchical clustering techniques. We have chosen radioactive detection because it combines robustness with sensitivity and allows the comparison of multiple normalized experiments. Sensitive detection combined with highly effective clustering algorithms allowed the identification of tissue-specific expression profiles and the detection of genes specifically expressed in the tissues investigated. The obtained results are publicly available (http://www.rzpd.de) and can be used by other researchers as a digital expression reference. [The sequence data described in this paper have been submitted to the EMBL data library under accession nos. AL360374–AL36537.] PMID:10958641

TRANSCRIPTIONAL PROFILING USING THE FLOWTHROUGH GENOSENSOR FINAL CRADA REPORT C/ORNL97-00472

DOE Office of Scientific and Technical Information (OSTI.GOV)

Doktycz, M. J.; Yang, H.

1999-06-01

A Cooperative Research and Development Agreement (CRADA) between Lockheed Martin Energy Research Corporation (Contractor) and Gene Logic, Inc., (Participant) was carried out to evaluate the technical feasibility study of the application of the flowthrough genosensor for gene expression (transcriptional) profiling, over the current industry practice of using flat surface hybridization arrays to monitor the relative abundance of individual mRNA species in a cell. Various parameters, including substrate preparation, flow rates, hybridization conditions and sample concentrations, were evaluated on the flowthrough genosensor. The superiority of the flowthrough genosensor, in terms of hybridization rate and sensitivity were established.

HuMiChip: Development of a Functional Gene Array for the Study of Human Microbiomes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tu, Q.; Deng, Ye; Lin, Lu

Microbiomes play very important roles in terms of nutrition, health and disease by interacting with their hosts. Based on sequence data currently available in public domains, we have developed a functional gene array to monitor both organismal and functional gene profiles of normal microbiota in human and mouse hosts, and such an array is called human and mouse microbiota array, HMM-Chip. First, seed sequences were identified from KEGG databases, and used to construct a seed database (seedDB) containing 136 gene families in 19 metabolic pathways closely related to human and mouse microbiomes. Second, a mother database (motherDB) was constructed withmore » 81 genomes of bacterial strains with 54 from gut and 27 from oral environments, and 16 metagenomes, and used for selection of genes and probe design. Gene prediction was performed by Glimmer3 for bacterial genomes, and by the Metagene program for metagenomes. In total, 228,240 and 801,599 genes were identified for bacterial genomes and metagenomes, respectively. Then the motherDB was searched against the seedDB using the HMMer program, and gene sequences in the motherDB that were highly homologous with seed sequences in the seedDB were used for probe design by the CommOligo software. Different degrees of specific probes, including gene-specific, inclusive and exclusive group-specific probes were selected. All candidate probes were checked against the motherDB and NCBI databases for specificity. Finally, 7,763 probes covering 91.2percent (12,601 out of 13,814) HMMer confirmed sequences from 75 bacterial genomes and 16 metagenomes were selected. This developed HMM-Chip is able to detect the diversity and abundance of functional genes, the gene expression of microbial communities, and potentially, the interactions of microorganisms and their hosts.« less

Cell-specific prediction and application of drug-induced gene expression profiles.

PubMed

Hodos, Rachel; Zhang, Ping; Lee, Hao-Chih; Duan, Qiaonan; Wang, Zichen; Clark, Neil R; Ma'ayan, Avi; Wang, Fei; Kidd, Brian; Hu, Jianying; Sontag, David; Dudley, Joel

2018-01-01

Gene expression profiling of in vitro drug perturbations is useful for many biomedical discovery applications including drug repurposing and elucidation of drug mechanisms. However, limited data availability across cell types has hindered our capacity to leverage or explore the cell-specificity of these perturbations. While recent efforts have generated a large number of drug perturbation profiles across a variety of human cell types, many gaps remain in this combinatorial drug-cell space. Hence, we asked whether it is possible to fill these gaps by predicting cell-specific drug perturbation profiles using available expression data from related conditions--i.e. from other drugs and cell types. We developed a computational framework that first arranges existing profiles into a three-dimensional array (or tensor) indexed by drugs, genes, and cell types, and then uses either local (nearest-neighbors) or global (tensor completion) information to predict unmeasured profiles. We evaluate prediction accuracy using a variety of metrics, and find that the two methods have complementary performance, each superior in different regions in the drug-cell space. Predictions achieve correlations of 0.68 with true values, and maintain accurate differentially expressed genes (AUC 0.81). Finally, we demonstrate that the predicted profiles add value for making downstream associations with drug targets and therapeutic classes.

Cell-specific prediction and application of drug-induced gene expression profiles

PubMed Central

Hodos, Rachel; Zhang, Ping; Lee, Hao-Chih; Duan, Qiaonan; Wang, Zichen; Clark, Neil R.; Ma'ayan, Avi; Wang, Fei; Kidd, Brian; Hu, Jianying; Sontag, David

2017-01-01

Gene expression profiling of in vitro drug perturbations is useful for many biomedical discovery applications including drug repurposing and elucidation of drug mechanisms. However, limited data availability across cell types has hindered our capacity to leverage or explore the cell-specificity of these perturbations. While recent efforts have generated a large number of drug perturbation profiles across a variety of human cell types, many gaps remain in this combinatorial drug-cell space. Hence, we asked whether it is possible to fill these gaps by predicting cell-specific drug perturbation profiles using available expression data from related conditions--i.e. from other drugs and cell types. We developed a computational framework that first arranges existing profiles into a three-dimensional array (or tensor) indexed by drugs, genes, and cell types, and then uses either local (nearest-neighbors) or global (tensor completion) information to predict unmeasured profiles. We evaluate prediction accuracy using a variety of metrics, and find that the two methods have complementary performance, each superior in different regions in the drug-cell space. Predictions achieve correlations of 0.68 with true values, and maintain accurate differentially expressed genes (AUC 0.81). Finally, we demonstrate that the predicted profiles add value for making downstream associations with drug targets and therapeutic classes. PMID:29218867

The von Hippel-Lindau (VHL) tumor-suppressor gene is down-regulated by selenium deficiency in Caco-2 cells and rat colon mucosa.

PubMed

Uthus, Eric; Begaye, Adrienne; Ross, Sharon; Zeng, Huawei

2011-08-01

To test the hypothesis that selenium affects DNA methylation and hence gene regulation, we employed a methylation array (Panomics) in the human colonic epithelial Caco-2 cell model. The array profiles DNA methylation from promoter regions of 82 human genes. After conditioning cells to repeatedly reduced concentrations of fetal bovine serum, a serum-free culture was established. Se-methylselenocysteine (SeMSC) was added at 0 (deficient Se) or 250 (control Se) nM to cells maintained in DMEM. After 7 days, cells were collected and stored at -80 °C until analysis; experiments were replicated three times. Glutathione peroxidase activity was significantly decreased in cells grown in low SeMSC. Cells grown in 250 nM SeMSC had maximal GPx activity. Genomic DNA from cells grown in the low-SeMSC media and media containing 250 nM SeMSC was incubated with methylation-binding protein followed by isolation of methylated DNA. The methylated DNA was labeled with biotin and hybridized to the methylation array. Thus, genes with promoter methylation will produce a higher chemiluminescence signal than those genes with no promoter methylation. Of the genes profiled, the von Hippel-Lindau (VHL) gene was most different as indicated by quantification following chemiluminescence detection demonstrating that the promoter region of VHL was hypermethylated in cells from the low-SeMSC media. To determine whether promoter methylation affected transcription, we isolated RNA from replicate samples and performed real-time RT PCR. VHL (mRNA) was down-regulated (fold change significantly <1) in cells grown in low SeMSC compared to cells grown in 250 nM SeMSC (control; fold change = 1). We also show that (mRNA) Vhl expression is significantly reduced in mucosa from rats fed a diet deficient in Se. Our results suggest that low Se status affects DNA promoter region methylation and that this can result in down-regulation of the tumor suppressor gene VHL.

Global gene expression profiles of Phytophthora ramorum strain pr102 in response to plant host and tissue differentiation

Treesearch

Caroline M. Press; Niklaus J. Grunwald

2008-01-01

The release of the draft genome sequence of P. ramorum strain Pr102, enabled the construction of an oligonucleotide microarray of the entire genome of Pr102. The array contains 344,680 features (oligos) that represent the transcriptome of Pr102. P. ramorum RNA was extracted from mycelium and sporangia and used to compare gene...

TIPMaP: a web server to establish transcript isoform profiles from reliable microarray probes.

PubMed

Chitturi, Neelima; Balagannavar, Govindkumar; Chandrashekar, Darshan S; Abinaya, Sadashivam; Srini, Vasan S; Acharya, Kshitish K

2013-12-27

Standard 3' Affymetrix gene expression arrays have contributed a significantly higher volume of existing gene expression data than other microarray platforms. These arrays were designed to identify differentially expressed genes, but not their alternatively spliced transcript forms. No resource can currently identify expression pattern of specific mRNA forms using these microarray data, even though it is possible to do this. We report a web server for expression profiling of alternatively spliced transcripts using microarray data sets from 31 standard 3' Affymetrix arrays for human, mouse and rat species. The tool has been experimentally validated for mRNAs transcribed or not-detected in a human disease condition (non-obstructive azoospermia, a male infertility condition). About 4000 gene expression datasets were downloaded from a public repository. 'Good probes' with complete coverage and identity to latest reference transcript sequences were first identified. Using them, 'Transcript specific probe-clusters' were derived for each platform and used to identify expression status of possible transcripts. The web server can lead the user to datasets corresponding to specific tissues, conditions via identifiers of the microarray studies or hybridizations, keywords, official gene symbols or reference transcript identifiers. It can identify, in the tissues and conditions of interest, about 40% of known transcripts as 'transcribed', 'not-detected' or 'differentially regulated'. Corresponding additional information for probes, genes, transcripts and proteins can be viewed too. We identified the expression of transcripts in a specific clinical condition and validated a few of these transcripts by experiments (using reverse transcription followed by polymerase chain reaction). The experimental observations indicated higher agreements with the web server results, than contradictions. The tool is accessible at http://resource.ibab.ac.in/TIPMaP. The newly developed online tool forms a reliable means for identification of alternatively spliced transcript-isoforms that may be differentially expressed in various tissues, cell types or physiological conditions. Thus, by making better use of existing data, TIPMaP avoids the dependence on precious tissue-samples, in experiments with a goal to establish expression profiles of alternative splice forms--at least in some cases.

Transcriptional profiling of root-knot nematode induced feeding sites in cowpea (Vigna unguiculata L. Walp.) using a soybean genome array.

PubMed

Das, Sayan; Ehlers, Jeffrey D; Close, Timothy J; Roberts, Philip A

2010-08-19

The locus Rk confers resistance against several species of root-knot nematodes (Meloidogyne spp., RKN) in cowpea (Vigna unguiculata). Based on histological and reactive oxygen species (ROS) profiles, Rk confers a delayed but strong resistance mechanism without a hypersensitive reaction-mediated cell death process, which allows nematode development but blocks reproduction. Responses to M. incognita infection in roots of resistant genotype CB46 and a susceptible near-isogenic line (null-Rk) were investigated using a soybean Affymetrix GeneChip expression array at 3 and 9 days post-inoculation (dpi). At 9 dpi 552 genes were differentially expressed in incompatible interactions (infected resistant tissue compared with non-infected resistant tissue) and 1,060 genes were differentially expressed in compatible interactions (infected susceptible tissue compared with non-infected susceptible tissue). At 3 dpi the differentially expressed genes were 746 for the incompatible and 623 for the compatible interactions. When expression between infected resistant and susceptible genotypes was compared, 638 and 197 genes were differentially expressed at 9 and 3 dpi, respectively. In comparing the differentially expressed genes in response to nematode infection, a greater number and proportion of genes were down-regulated in the resistant than in the susceptible genotype, whereas more genes were up-regulated in the susceptible than in the resistant genotype. Gene ontology based functional categorization revealed that the typical defense response was partially suppressed in resistant roots, even at 9 dpi, allowing nematode juvenile development. Differences in ROS concentrations, induction of toxins and other defense related genes seem to play a role in this unique resistance mechanism.

BEAT: Bioinformatics Exon Array Tool to store, analyze and visualize Affymetrix GeneChip Human Exon Array data from disease experiments

PubMed Central

2012-01-01

Background It is known from recent studies that more than 90% of human multi-exon genes are subject to Alternative Splicing (AS), a key molecular mechanism in which multiple transcripts may be generated from a single gene. It is widely recognized that a breakdown in AS mechanisms plays an important role in cellular differentiation and pathologies. Polymerase Chain Reactions, microarrays and sequencing technologies have been applied to the study of transcript diversity arising from alternative expression. Last generation Affymetrix GeneChip Human Exon 1.0 ST Arrays offer a more detailed view of the gene expression profile providing information on the AS patterns. The exon array technology, with more than five million data points, can detect approximately one million exons, and it allows performing analyses at both gene and exon level. In this paper we describe BEAT, an integrated user-friendly bioinformatics framework to store, analyze and visualize exon arrays datasets. It combines a data warehouse approach with some rigorous statistical methods for assessing the AS of genes involved in diseases. Meta statistics are proposed as a novel approach to explore the analysis results. BEAT is available at http://beat.ba.itb.cnr.it. Results BEAT is a web tool which allows uploading and analyzing exon array datasets using standard statistical methods and an easy-to-use graphical web front-end. BEAT has been tested on a dataset with 173 samples and tuned using new datasets of exon array experiments from 28 colorectal cancer and 26 renal cell cancer samples produced at the Medical Genetics Unit of IRCCS Casa Sollievo della Sofferenza. To highlight all possible AS events, alternative names, accession Ids, Gene Ontology terms and biochemical pathways annotations are integrated with exon and gene level expression plots. The user can customize the results choosing custom thresholds for the statistical parameters and exploiting the available clinical data of the samples for a multivariate AS analysis. Conclusions Despite exon array chips being widely used for transcriptomics studies, there is a lack of analysis tools offering advanced statistical features and requiring no programming knowledge. BEAT provides a user-friendly platform for a comprehensive study of AS events in human diseases, displaying the analysis results with easily interpretable and interactive tables and graphics. PMID:22536968

Molecular Targeted Therapies of Childhood Choroid Plexus Carcinoma

DTIC Science & Technology

2013-10-01

Microarray intensities were analyzed in PGS, using the benign human choroid plexus papilloma (CPP) samples as an expression baseline reference. This...additional human and mouse CPC genomic profiles (timeframe: months 1-5). The goal of these studies is to expand our number of genomic profiles (DNA and...mRNA arrays) of both human and mouse CPCs to provide a comprehensive dataset with which to identify key candidate oncogenes, tumor suppressor genes

Mitogen- and Stress-Activated Protein Kinase 1 Regulates Status Epilepticus-Evoked Cell Death in the Hippocampus

PubMed Central

Choi, Yun-Sik; Horning, Paul; Aten, Sydney; Karelina, Kate; Alzate-Correa, Diego; Arthur, J. Simon C.; Hoyt, Kari R.; Obrietan, Karl

2017-01-01

Mitogen-activated protein kinase (MAPK) signaling has been implicated in a wide range of neuronal processes, including development, plasticity, and viability. One of the principal downstream targets of both the extracellular signal-regulated kinase/MAPK pathway and the p38 MAPK pathway is Mitogen- and Stress-activated protein Kinase 1 (MSK1). Here, we sought to understand the role that MSK1 plays in neuroprotection against excitotoxic stimulation in the hippocampus. To this end, we utilized immunohistochemical labeling, a MSK1 null mouse line, cell viability assays, and array-based profiling approaches. Initially, we show that MSK1 is broadly expressed within the major neuronal cell layers of the hippocampus and that status epilepticus drives acute induction of MSK1 activation. In response to the status epilepticus paradigm, MSK1 KO mice exhibited a striking increase in vulnerability to pilocarpine-evoked cell death within the CA1 and CA3 cell layers. Further, cultured MSK1 null neurons exhibited a heighted level of N-methyl-D-aspartate-evoked excitotoxicity relative to wild-type neurons, as assessed using the lactate dehydrogenase assay. Given these findings, we examined the hippocampal transcriptional profile of MSK1 null mice. Affymetrix array profiling revealed that MSK1 deletion led to the significant (>1.25-fold) downregulation of 130 genes and an upregulation of 145 genes. Notably, functional analysis indicated that a subset of these genes contribute to neuroprotective signaling networks. Together, these data provide important new insights into the mechanism by which the MAPK/MSK1 signaling cassette confers neuroprotection against excitotoxic insults. Approaches designed to upregulate or mimic the functional effects of MSK1 may prove beneficial against an array of degenerative processes resulting from excitotoxic insults. PMID:28870089

Fidelity and enhanced sensitivity of differential transcription profiles following linear amplification of nanogram amounts of endothelial mRNA

NASA Technical Reports Server (NTRS)

Polacek, Denise C.; Passerini, Anthony G.; Shi, Congzhu; Francesco, Nadeene M.; Manduchi, Elisabetta; Grant, Gregory R.; Powell, Steven; Bischof, Helen; Winkler, Hans; Stoeckert, Christian J Jr;

Temporal Changes in Gene Expression in Rainbow Trout Exposed to Ethynyl Estradiol*

PubMed Central

Skillman, Ann D.; Small, Jack A.; Schultz, Irvin R.

2007-01-01

We examined changes in the genomic response during continuous exposure to the xenoestrogen ethynylestradiol. Isogenic rainbow trout Oncorhynchus mykiss were exposed to nominal concentrations of 100 ng/L ethynyl estradiol (EE2) for a period of three weeks. At fixed time points within the exposure, fish were euthanized, livers harvested and RNA extracted. Fluorescently labeled cDNA were generated and hybridized against a commercially available Salmonid array (GRASP project, University of Victoria, Canada) spotted with 16,000 cDNAs. The slides were scanned to measure abundance of a given transcript in each sample relative to controls. Data were analyzed via Genespring (Silicon Genetics) to identify a list of up and down regulated genes, and to determine gene clustering patterns that can be used as “expression signatures”. Gene ontology was determined using the annotation available from the GRASP website. Our analysis indicates each exposure time period generated specific gene expression profiles. Changes in gene expression were best understood by grouping genes by their gene expression profiles rather than examining fold change at a particular time point. Many of the genes commonly used as biomarkers of exposure to xenoestrogens were not induced initially and did not have gene expression profiles typical of the majority of genes with altered expression. PMID:17215170

Temporal changes in gene expression in rainbow trout exposed to ethynyl estradiol.

PubMed

Hook, Sharon E; Skillman, Ann D; Small, Jack A; Schultz, Irvin R

2007-02-01

We examined changes in the genomic response during continuous exposure to the xenoestrogen ethynyl estradiol. Isogenic rainbow trout Oncorhynchus mykiss were exposed to nominal concentrations of 100 ng/L ethynyl estradiol (EE2) for a period of 3 weeks. At fixed time points within the exposure, fish were euthanized, livers harvested and RNA extracted. Fluorescently labeled cDNA were generated and hybridized against a commercially available Salmonid array (GRASP project, University of Victoria, Canada) spotted with 16,000 cDNAs. The slides were scanned to measure abundance of a given transcript in each sample relative to controls. Data were analyzed via Genespring (Silicon Genetics) to identify a list of up and down regulated genes, and to determine gene clustering patterns that can be used as "expression signatures". Gene ontology was determined using the annotation available from the GRASP website. Our analysis indicates each exposure time period generated specific gene expression profiles. Changes in gene expression were best understood by grouping genes by their gene expression profiles rather than examining fold change at a particular time point. Many of the genes commonly used as biomarkers of exposure to xenoestrogens were not induced initially and did not have gene expression profiles typical of the majority of genes with altered expression.

Alteration of gene expression profiling including GPR174 and GNG2 is associated with vasovagal syncope.

PubMed

Huang, Yu-Juan; Zhou, Zai-wei; Xu, Miao; Ma, Qing-wen; Yan, Jing-bin; Wang, Jian-yi; Zhang, Quo-qin; Huang, Min; Bao, Liming

2015-03-01

Vasovagal syncope (VVS) causes accidental harm for susceptible patients. However, pathophysiology of this disorder remains largely unknown. In an effort to understanding of molecular mechanism for VVS, genome-wide gene expression profiling analyses were performed on VVS patients at syncope state. A total of 66 Type 1 VVS child patients and the same number healthy controls were enrolled in this study. Peripheral blood RNAs were isolated from all subjects, of which 10 RNA samples were randomly selected from each groups for gene expression profile analysis using Gene ST 1.0 arrays (Affymetrix). The results revealed that 103 genes were differently expressed between the patients and controls. Significantly, two G-proteins related genes, GPR174 and GNG2 that have not been related to VVS were among the differently expressed genes. The microarray results were confirmed by qRT-PCR in all the tested individuals. Ingenuity pathway analysis and gene ontology annotation study showed that the differently expressed genes are associated with stress response and apoptosis, suggesting that the alteration of some gene expression including G-proteins related genes is associated with VVS. This study provides new insight into the molecular mechanism of VVS and would be helpful to further identify new molecular biomarkers for the disease.

MOLECULAR CHARACTERIZATION OF ENDOCRINE DISRUPTION IN FISH USING CDNA ARRAYS.

EPA Science Inventory

We are developing cDNA macroarrays to measure the induction of gene expression in sheepshead minnows and largemouth bass exposed to anthropogenic chemicals that can mimic the action of endogenous hormones. For sheepshead minnows exposed in aqua, we observed similar genetic profil...

High-resolution single-nucleotide polymorphism array-profiling in myeloproliferative neoplasms identifies novel genomic aberrations

PubMed Central

Stegelmann, Frank; Bullinger, Lars; Griesshammer, Martin; Holzmann, Karlheinz; Habdank, Marianne; Kuhn, Susanne; Maile, Carmen; Schauer, Stefanie; Döhner, Hartmut; Döhner, Konstanze

2010-01-01

Single-nucleotide polymorphism arrays allow for genome-wide profiling of copy-number alterations and copy-neutral runs of homozygosity at high resolution. To identify novel genetic lesions in myeloproliferative neoplasms, a large series of 151 clinically well characterized patients was analyzed in our study. Copy-number alterations were rare in essential thrombocythemia and polycythemia vera. In contrast, approximately one third of myelofibrosis patients exhibited small genomic losses (less than 5 Mb). In 2 secondary myelofibrosis cases the tumor suppressor gene NF1 in 17q11.2 was affected. Sequencing analyses revealed a mutation in the remaining NF1 allele of one patient. In terms of copy-neutral aberrations, no chromosomes other than 9p were recurrently affected. In conclusion, novel genomic aberrations were identified in our study, in particular in patients with myelofibrosis. Further analyses on single-gene level are necessary to uncover the mechanisms that are involved in the pathogenesis of myeloproliferative neoplasms. PMID:20015882

Profiling Pre-MicroRNA and Mature MicroRNA Expressions Using a Single Microarray and Avoiding Separate Sample Preparation

PubMed Central

Gan, Lin; Denecke, Bernd

2013-01-01

Mature microRNA is a crucial component in the gene expression regulation network. At the same time, microRNA gene expression and procession is regulated in a precise and collaborated way. Pre-microRNAs mediate products during the microRNA transcription process, they can provide hints of microRNA gene expression regulation or can serve as alternative biomarkers. To date, little effort has been devoted to pre-microRNA expression profiling. In this study, three human and three mouse microRNA profile data sets, based on the Affymetrix miRNA 2.0 array, have been re-analyzed for both mature and pre-microRNA signals as a primary test of parallel mature/pre-microRNA expression profiling on a single platform. The results not only demonstrated a glimpse of pre-microRNA expression in human and mouse, but also the relationship of microRNA expressions between pre- and mature forms. The study also showed a possible application of currently available microRNA microarrays in profiling pre-microRNA expression in a time and cost effective manner. PMID:27605179

Stress amplifies sex differences in primate prefrontal profiles of gene expression.

PubMed

Lee, Alex G; Hagenauer, Megan; Absher, Devin; Morrison, Kathleen E; Bale, Tracy L; Myers, Richard M; Watson, Stanley J; Akil, Huda; Schatzberg, Alan F; Lyons, David M

2017-11-02

Stress is a recognized risk factor for mood and anxiety disorders that occur more often in women than men. Prefrontal brain regions mediate stress coping, cognitive control, and emotion. Here, we investigate sex differences and stress effects on prefrontal cortical profiles of gene expression in squirrel monkey adults. Dorsolateral, ventrolateral, and ventromedial prefrontal cortical regions from 18 females and 12 males were collected after stress or no-stress treatment conditions. Gene expression profiles were acquired using HumanHT-12v4.0 Expression BeadChip arrays adapted for squirrel monkeys. Extensive variation between prefrontal cortical regions was discerned in the expression of numerous autosomal and sex chromosome genes. Robust sex differences were also identified across prefrontal cortical regions in the expression of mostly autosomal genes. Genes with increased expression in females compared to males were overrepresented in mitogen-activated protein kinase and neurotrophin signaling pathways. Many fewer genes with increased expression in males compared to females were discerned, and no molecular pathways were identified. Effect sizes for sex differences were greater in stress compared to no-stress conditions for ventromedial and ventrolateral prefrontal cortical regions but not dorsolateral prefrontal cortex. Stress amplifies sex differences in gene expression profiles for prefrontal cortical regions involved in stress coping and emotion regulation. Results suggest molecular targets for new treatments of stress disorders in human mental health.

Identification of specific gene expression profiles in fibroblasts derived from middle ear cholesteatoma.

PubMed

Yoshikawa, Mamoru; Kojima, Hiromi; Wada, Kota; Tsukidate, Toshiharu; Okada, Naoko; Saito, Hirohisa; Moriyama, Hiroshi

2006-07-01

To investigate the role of fibroblasts in the pathogenesis of cholesteatoma. Tissue specimens were obtained from our patients. Middle ear cholesteatoma-derived fibroblasts (MECFs) and postauricular skin-derived fibroblasts (SFs) as controls were then cultured for a few weeks. These fibroblasts were stimulated with interleukin (IL) 1alpha and/or IL-1beta before gene expression assays. We used the human genome U133A probe array (GeneChip) and real-time polymerase chain reaction to examine and compare the gene expression profiles of the MECFs and SFs. Six patients who had undergone tympanoplasty. The IL-1alpha-regulated genes were classified into 4 distinct clusters on the basis of profiles differentially regulated by SF and MECF using a hierarchical clustering analysis. The messenger RNA expressions of LARC (liver and activation-regulated chemokine), GMCSF (granulocyte-macrophage colony-stimulating factor), epiregulin, ICAM1 (intercellular adhesion molecule 1), and TGFA (transforming growth factor alpha) were more strongly up-regulated by IL-1alpha and/or IL-1beta in MECF than in SF, suggesting that these fibroblasts derived from different tissues retained their typical gene expression profiles. Fibroblasts may play a role in hyperkeratosis of middle ear cholesteatoma by releasing molecules involved in inflammation and epidermal growth. These fibroblasts may retain tissue-specific characteristics presumably controlled by epigenetic mechanisms.

Comparative prion disease gene expression profiling using the prion disease mimetic, cuprizone

PubMed Central

Moody, Laura R; Herbst, Allen J; Yoo, Han Sang; Vanderloo, Joshua P

2009-01-01

Identification of genes expressed in response to prion infection may elucidate biomarkers for disease, identify factors involved in agent replication, mechanisms of neuropathology and therapeutic targets. Although several groups have sought to identify gene expression changes specific to prion disease, expression profiles rife with cell population changes have consistently been identified. Cuprizone, a neurotoxicant, qualitatively mimics the cell population changes observed in prion disease, resulting in both spongiform change and astrocytosis. The use of cuprizone-treated animals as an experimental control during comparative expression profiling allows for the identification of transcripts whose expression increases during prion disease and remains unchanged during cuprizone-triggered neuropathology. In this study, expression profiles from the brains of mice preclinically and clinically infected with Rocky Mountain Laboratory (RML) mouse-adapted scrapie agent and age-matched controls were profiled using Affymetrix gene arrays. In total, 164 genes were differentially regulated during prion infection. Eighty-three of these transcripts have been previously undescribed as differentially regulated during prion disease. A 0.4% cuprizone diet was utilized as a control for comparative expression profiling. Cuprizone treatment induced spongiosis and astrocyte proliferation as indicated by glial fibrillary acidic protein (Gfap) transcriptional activation and immunohistochemistry. Gene expression profiles from brain tissue obtained from cuprizone-treated mice identified 307 differentially regulated transcript changes. After comparative analysis, 17 transcripts unaffected by cuprizone treatment but increasing in expression from preclinical to clinical prion infection were identified. Here we describe the novel use of the prion disease mimetic, cuprizone, to control for cell population changes in the brain during prion infection. PMID:19535908

Analysis of gene expression profiles in tympanic membrane following perforation using PCR Array in rats--preliminary investigation.

PubMed

Hassmann-Poznańska, Elżbieta; Taranta, Andrzej; Bialuk, Izabela; Poznańska, Maria; Zajączkiewicz, Hanna; Winnicka, Maria Małgorzata

2013-10-01

The goal of this work was to identify genes, known to be involved in the skin wound healing, that express differentially in the healthy and injured tympanic membrane (TM), and designate the molecules potentially beneficial for treatment of TM perforation. The molecular mechanisms controlling the course of TM regeneration are far from being elucidated. Twenty rats had their tympanic membranes perforated, while four served as a control. Animals were sacrificed on either days 1, 2, 3, 5 and 10 post injury, and TMs were immediately dissected and frozen in liquid nitrogen. Total TM RNA was isolated and reversely transcribed. qPCR was performed using Rat Wound Healing RT(2) Profiler PCR Array (QIAGEN) containing primers for 84 genes. Statistically significant changes in the expression of 42 genes were found in various stages of TM healing. The increased expression of genes taking part in the inflammatory reaction (interleukin 6, granulocyte and macrophage chemotactic proteins) was observed from day 2. The expression of several genes of extracellular matrix components and their remodeling enzymes was also changed. Among growth factor genes: Vegfa, Igf1 and Hbegf showed increased expression at the beginning of the healing process, while Hgf expression was highest on day 3. Several changes in the expression of genes involved in remodeling of extracellular matrix point to important role of connective tissue in TM healing. The molecules accelerating this process, like HbEGF and HGF, seem to be good candidates for further evaluation of their possible use in clinical treatment. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Molecular Profiles for Lung Cancer Pathogenesis and Detection in US Veterans

DTIC Science & Technology

2011-10-01

expression data was analyzed using the BRB-ArrayTools v .4.1.0 developed by Dr. Richard Simon and the BRB-ArrayTools Development Team and then normalized...14. Spira A, Beane J, Shah V , et al. Effects of cigarette smoke on the human airway epithelial cell transcriptome. Proc Natl Acad Sci U S A 2004;101...10143-10148 15. Spira A, Beane JE, Shah V , et al. Airway epithelial gene expression in the diagnostic evaluation of smokers with suspect lung cancer

Molecular Profiles for Lung Cancer Pathogenesis and Detection in US Veterans

DTIC Science & Technology

2011-10-01

using the BRB-ArrayTools v .4.1.0 developed by Dr. Richard Simon and the BRB-ArrayTools Development Team and then normalized and log-transformed by...C. V Figure 23. Expression of genes located at chromosome 3q26-3q29. RPKM values were obtained for...airway epithelium of cigarette smokers. Am J Respir Cell Mol Biol 2003;29:331-343 14. Spira A, Beane J, Shah V , et al. Effects of cigarette smoke on

Molecular Profiles for Lung Cancer Pathogenesis and Detection in US Veterans

DTIC Science & Technology

2011-10-01

data was analyzed using the BRB-ArrayTools v .4.1.0 developed by Dr. Richard Simon and the BRB-ArrayTools Development Team and then normalized and log...Spira A, Beane J, Shah V , et al. Effects of cigarette smoke on the human airway epithelial cell transcriptome. Proc Natl Acad Sci U S A 2004;101:10143...10148 15. Spira A, Beane JE, Shah V , et al. Airway epithelial gene expression in the diagnostic evaluation of smokers with suspect lung cancer. Nat

Customizing microarrays for neuroscience drug discovery.

PubMed

Girgenti, Matthew J; Newton, Samuel S

2007-08-01

Microarray-based gene profiling has become the centerpiece of gene expression studies in the biological sciences. The ability to now interrogate the entire genome using a single chip demonstrates the progress in technology and instrumentation that has been made over the last two decades. Although this unbiased approach provides researchers with an immense quantity of data, obtaining meaningful insight is not possible without intensive data analysis and processing. Custom developed arrays have emerged as a viable and attractive alternative that can take advantage of this robust technology and tailor it to suit the needs and requirements of individual investigations. The ability to simplify data analysis, reduce noise and carefully optimize experimental conditions makes it a suitable tool that can be effectively utilized in neuroscience drug discovery efforts. Furthermore, incorporating recent advancements in fine focusing gene profiling to include specific cellular phenotypes can help resolve the complex cellular heterogeneity of the brain. This review surveys the use of microarray technology in neuroscience paying special attention to customized arrays and their potential in drug discovery. Novel applications of microarrays and ancillary techniques, such as laser microdissection, FAC sorting and RNA amplification, have also been discussed. The notion that a hypothesis-driven approach can be integrated into drug development programs is highlighted.

Gene expression profiles in peripheral blood mononuclear cells of Chinese nickel refinery workers with high exposures to nickel and control subjects

PubMed Central

Arita, Adriana; Muñoz, Alexandra; Chervona, Yana; Niu, Jingping; Qu, Qingshan; Zhao, Najuan; Ruan, Ye; Kiok, Kathrin; Kluz, Thomas; Sun, Hong; Clancy, Hailey A.; Shamy, Magdy; Costa, Max

2012-01-01

Background Occupational exposure to nickel (Ni) is associated with an increased risk of lung and nasal cancers. Ni compounds exhibit weak mutagenic activity, alter the cell’s epigenetic homeostasis, and activate signaling pathways. However, changes in gene expression associated with Ni exposure have only been investigated in vitro. This study was conducted in a Chinese population to determine whether occupational exposure to Ni was associated with differential gene expression profiles in the peripheral blood mononuclear cells (PBMCs) of Ni-refinery workers when compared to referents. Methods Eight Ni-refinery workers and ten referents were selected. PBMC RNA was extracted and gene expression profiling was performed using Affymetrix exon arrays. Differentially expressed genes between both groups were identified in a global analysis. Results There were a total of 2756 differentially expressed genes (DEG) in the Ni-refinery workers relative to the control subjects (FDR adjusted p<0.05) with 770 up-regulated genes and 1986 down-regulated genes. DNA repair and epigenetic genes were significantly overrepresented (p< 0.0002) among the DEG. Of 31 DNA repair genes, 29 were repressed in the high exposure group and two were overexpressed. Of the 16 epigenetic genes 12 were repressed in the high exposure group and 4 were overexpressed. Conclusions The results of this study indicate that occupational exposure to Ni is associated with alterations in gene expression profiles in PBMCs of subjects. Impact Gene expression may be useful in identifying patterns of deregulation that precede clinical identification of Ni-induced cancers. PMID:23195993

Comparison of the Predictive Accuracy of DNA Array-Based Multigene Classifiers across cDNA Arrays and Affymetrix GeneChips

PubMed Central

Stec, James; Wang, Jing; Coombes, Kevin; Ayers, Mark; Hoersch, Sebastian; Gold, David L.; Ross, Jeffrey S; Hess, Kenneth R.; Tirrell, Stephen; Linette, Gerald; Hortobagyi, Gabriel N.; Symmans, W. Fraser; Pusztai, Lajos

2005-01-01

We examined how well differentially expressed genes and multigene outcome classifiers retain their class-discriminating values when tested on data generated by different transcriptional profiling platforms. RNA from 33 stage I-III breast cancers was hybridized to both Affymetrix GeneChip and Millennium Pharmaceuticals cDNA arrays. Only 30% of all corresponding gene expression measurements on the two platforms had Pearson correlation coefficient r ≥ 0.7 when UniGene was used to match probes. There was substantial variation in correlation between different Affymetrix probe sets matched to the same cDNA probe. When cDNA and Affymetrix probes were matched by basic local alignment tool (BLAST) sequence identity, the correlation increased substantially. We identified 182 genes in the Affymetrix and 45 in the cDNA data (including 17 common genes) that accurately separated 91% of cases in supervised hierarchical clustering in each data set. Cross-platform testing of these informative genes resulted in lower clustering accuracy of 45 and 79%, respectively. Several sets of accurate five-gene classifiers were developed on each platform using linear discriminant analysis. The best 100 classifiers showed average misclassification error rate of 2% on the original data that rose to 19.5% when tested on data from the other platform. Random five-gene classifiers showed misclassification error rate of 33%. We conclude that multigene predictors optimized for one platform lose accuracy when applied to data from another platform due to missing genes and sequence differences in probes that result in differing measurements for the same gene. PMID:16049308

Comparison of global brain gene expression profiles between inbred long-sleep and inbred short-sleep mice by high-density gene array hybridization.

PubMed

Xu, Y; Ehringer, M; Yang, F; Sikela, J M

2001-06-01

Inbred long-sleep (ILS) and short-sleep (ISS) mice show significant central nervous system-mediated differences in sleep time for sedative dose of ethanol and are frequently used as a rodent model for ethanol sensitivity. In this study, we have used complementary DNA (cDNA) array hybridization methodology to identify genes that are differentially expressed between the brains of ILS and ISS mice. To carry out this analysis, we used both the gene discovery array (GDA) and the Mouse GEM 1 Microarray. GDA consists of 18,378 nonredundant mouse cDNA clones on a single nylon filter. Complex probes were prepared from total brain mRNA of ILS or ISS mice by using reverse transcription and 33P labeling. The labeled probes were hybridized in parallel to the gene array filters. Data from GDA experiments were analyzed with SQL-Plus and Oracle 8. The GEM microarray includes 8,730 sequence-verified clones on a glass chip. Two fluorescently labeled probes were used to hybridize a microarray simultaneously. Data from GEM experiments were analyzed by using the GEMTools software package (Incyte). Differentially expressed genes identified from each method were confirmed by relative quantitative reverse transcription-polymerase chain reaction (RT-PCR). A total of 41 genes or expressed sequence tags (ESTs) display significant expression level differences between brains of ILS and ISS mice after GDA, GEM1 hybridization, and quantitative RT-PCR confirmation. Among them, 18 clones were expressed higher in ILS mice, and 23 clones were expressed higher in ISS mice. The individual gene or EST's function and mapping information have been analyzed. This study identified 41 genes that are differentially expressed between brains of ILS and ISS mice. Some of them may have biological relevance in mediation of phenotypic variation between ILS and ISS mice for ethanol sensitivity. This study also demonstrates that parallel gene expression comparison with high-density cDNA arrays is a rapid and efficient way to discover potential genes and pathways involved in alcoholism and alcohol-related physiologic processes.

Changes in global gene expression profiles induced by HPV 16 E6 oncoprotein variants in cervical carcinoma C33-A cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zacapala-Gómez, Ana Elvira, E-mail: zak_ana@yahoo.com.mx; Del Moral-Hernández, Oscar, E-mail: odelmoralh@gmail.com; Villegas-Sepúlveda, Nicolás, E-mail: nvillega@cinvestav.mx

We analyzed the effects of the expression of HPV 16 E6 oncoprotein variants (AA-a, AA-c, E-A176/G350, E-C188/G350, E-G350), and the E-Prototype in global gene expression profiles in an in vitro model. E6 gene was cloned into an expression vector fused to GFP and was transfected in C33-A cells. Affymetrix GeneChip Human Transcriptome Array 2.0 platform was used to analyze the expression of over 245,000 coding transcripts. We found that HPV16 E6 variants altered the expression of 387 different genes in comparison with E-Prototype. The altered genes are involved in cellular processes related to the development of cervical carcinoma, such asmore » adhesion, angiogenesis, apoptosis, differentiation, cell cycle, proliferation, transcription and protein translation. Our results show that polymorphic changes in HPV16 E6 natural variants are sufficient to alter the overall gene expression profile in C33-A cells, explaining in part the observed differences in oncogenic potential of HPV16 variants. - Highlights: • Amino acid changes in HPV16 E6 variants modulate the transciption of specific genes. • This is the first comparison of global gene expression profile of HPV 16 E6 variants. • Each HPV 16 E6 variant appears to have its own molecular signature.« less

FULL-GENOME ANALYSIS OF ALTERNATIVE SPLICING IN MOUSE LIVER AFTER HEPATOTOXICANT EXPOSURE

EPA Science Inventory

Alternative splicing plays a role in determining gene function and protein diversity. We have employed whole genome exon profiling using Affymetrix Mouse Exon 1.0 ST arrays to understand the significance of alternative splicing on a genome-wide scale in response to multiple toxic...

Marek's Disease Virus-Induced Immunosuppression: Array Analysis of Chicken Immune Response Gene Expression Profiling

USDA-ARS?s Scientific Manuscript database

Marek’s disease (MD) is a lymphoproliferative disease of chickens induced by a highly cell-associated oncogenic alpha-herpesvirus, Marek’s disease virus (MDV). MDV replicates in chicken lymphocytes and establishes a latency infection within CD4+ T cells. Host-virus interaction, immune responses to...

COMPARISON OF PM-INDUCED GENE EXPRESSION PROFILES BETWEEN BRONCHIAL EPITHELIAL CELLS AND NASAL EPITHELIAL CELLS IN HUMAN

EPA Science Inventory

Epidemiologic studies have linked exposures to particulate matter (PM) and increased pulmonary mortality and morbidity. Bronchial epithelial cells (BEC) are the primary target of PM. PM exposure induces a wide array of biological responses in BEC. Primary human BEC, however, need...

Gene expression profile of mouse prostate tumors reveals dysregulations in major biological processes and identifies potential murine targets for preclinical development of human prostate cancer therapy.

PubMed

Haram, Kerstyn M; Peltier, Heidi J; Lu, Bin; Bhasin, Manoj; Otu, Hasan H; Choy, Bob; Regan, Meredith; Libermann, Towia A; Latham, Gary J; Sanda, Martin G; Arredouani, Mohamed S

2008-10-01

Translation of preclinical studies into effective human cancer therapy is hampered by the lack of defined molecular expression patterns in mouse models that correspond to the human counterpart. We sought to generate an open source TRAMP mouse microarray dataset and to use this array to identify differentially expressed genes from human prostate cancer (PCa) that have concordant expression in TRAMP tumors, and thereby represent lead targets for preclinical therapy development. We performed microarrays on total RNA extracted and amplified from eight TRAMP tumors and nine normal prostates. A subset of differentially expressed genes was validated by QRT-PCR. Differentially expressed TRAMP genes were analyzed for concordant expression in publicly available human prostate array datasets and a subset of resulting genes was analyzed by QRT-PCR. Cross-referencing differentially expressed TRAMP genes to public human prostate array datasets revealed 66 genes with concordant expression in mouse and human PCa; 56 between metastases and normal and 10 between primary tumor and normal tissues. Of these 10 genes, two, Sox4 and Tubb2a, were validated by QRT-PCR. Our analysis also revealed various dysregulations in major biologic pathways in the TRAMP prostates. We report a TRAMP microarray dataset of which a gene subset was validated by QRT-PCR with expression patterns consistent with previous gene-specific TRAMP studies. Concordance analysis between TRAMP and human PCa associated genes supports the utility of the model and suggests several novel molecular targets for preclinical therapy.

Gene expression profile differences in left and right liver lobes from mid-gestation fetal baboons: a cautionary tale

PubMed Central

Cox, Laura A; Schlabritz-Loutsevitch, Natalia; Hubbard, Gene B; Nijland, Mark J; McDonald, Thomas J; Nathanielsz, Peter W

2006-01-01

Interpretation of gene array data presents many potential pitfalls in adult tissues. Gene array techniques applied to fetal tissues present additional confounding pitfalls. The left lobe of the fetal liver is supplied with blood containing more oxygen than the right lobe. Since synthetic activity and cell function are oxygen dependent, we hypothesized major differences in mRNA expression between the fetal right and left liver lobes. Our aim was to demonstrate the need to evaluate RNA samples from both lobes. We performed whole genome expression profiling on left and right liver lobe RNA from six 90-day gestation baboon fetuses (term 180 days). Comparing right with left, we found 875 differentially expressed genes – 312 genes were up-regulated and 563 down-regulated. Pathways for damaged DNA binding, endonuclease activity, interleukin binding and receptor activity were up-regulated in right lobe; ontological pathways related to cell signalling, cell organization, cell biogenesis, development, intracellular transport, phospholipid metabolism, protein biosynthesis, protein localization, protein metabolism, translational regulation and vesicle mediated transport were down-regulated in right lobe. Molecular pathway analysis showed down-regulation of pathways related to heat shock protein binding, ion channel and transporter activities, oxygen binding and transporter activities, translation initiation and translation regulator activities. Genes involved in amino acid biosynthesis, lipid biosynthesis and oxygen transport were also differentially expressed. This is the first demonstration of RNA differences between the two lobes of the fetal liver. The data support the argument that a complete interpretation of gene expression in the developing liver requires data from both lobes. PMID:16484296

Profiling of the yak skeletal muscle tissue gene expression and comparison with the domestic cattle by genome array.

PubMed

Wang, H B; Zan, L S; Zhang, Y Y

2014-01-01

Of all the mammals of the world, the yak lives at the highest altitude area of more than 3000 m. Comparison between yak and cattle of the low-altitude areas will be informative in studying animal adaptation to higher altitudes. To investigate the molecular mechanism involved in meat quality differences between the two Chinese special varieties Qinghai yak and Qinchuan cattle, 12 chemical-physical characteristics of the longissimus dorsi muscle related to meat quality were compared at the age of 36 months, and the gene expression profiles were constructed by utilizing the bovine genome array. Significant analysis of microarrays was used to identify the differentially expressed genes. Gene ontology and pathway analysis were performed by a free Web-based Molecular Annotation System 2.0. The results reveal ~11 000 probes representing about 10 000 genes that were detected in both the Qinghai yak and Qinchuan cattle. A total of 1922 genes were shown to be differentially expressed, 633 probes were upregulated and 1259 probes were downregulated in the muscle tissue of Qinghai yak that were mainly involved in ubiquitin-mediated proteolysis, muscle growth regulation, glucose metabolism, immune response and so on. Quantitative real-time PCR (qRT-PCR) was performed to validate some differentially expressed genes identified by microarray. Further analysis implied that animals living at a high altitude may supply energy by more active protein catabolism and glycolysis compared with those living in the plain areas. Our results establish the groundwork for further studies on yaks' meat quality and will be beneficial in improving the yaks' breeding by molecular biotechnology.

Genomic profiling of CHEK2*1100delC-mutated breast carcinomas.

PubMed

Massink, Maarten P G; Kooi, Irsan E; Martens, John W M; Waisfisz, Quinten; Meijers-Heijboer, Hanne

2015-11-09

CHEK2*1100delC is a moderate-risk breast cancer susceptibility allele with a high prevalence in the Netherlands. We performed copy number and gene expression profiling to investigate whether CHEK2*1100delC breast cancers harbor characteristic genomic aberrations, as seen for BRCA1 mutated breast cancers. We performed high-resolution SNP array and gene expression profiling of 120 familial breast carcinomas selected from a larger cohort of 155 familial breast tumors, including BRCA1, BRCA2, and CHEK2 mutant tumors. Gene expression analyses based on a mRNA immune signature was used to identify samples with relative low amounts of tumor infiltrating lymphocytes (TILs), which were previously found to disturb tumor copy number and LOH (loss of heterozygosity) profiling. We specifically compared the genomic and gene expression profiles of CHEK2*1100delC breast cancers (n = 14) with BRCAX (familial non-BRCA1/BRCA2/CHEK2*1100delC mutated) breast cancers (n = 34) of the luminal intrinsic subtypes for which both SNP-array and gene expression data is available. High amounts of TILs were found in a relatively small number of luminal breast cancers as compared to breast cancers of the basal-like subtype. As expected, these samples mostly have very few copy number aberrations and no detectable regions of LOH. By unsupervised hierarchical clustering of copy number data we observed a great degree of heterogeneity amongst the CHEK2*1100delC breast cancers, comparable to the BRCAX breast cancers. Furthermore, copy number aberrations were mostly seen at low frequencies in both the CHEK2*1100delC and BRCAX group of breast cancers. However, supervised class comparison identified copy number loss of chromosomal arm 1p to be associated with CHEK2*1100delC status. In conclusion, in contrast to basal-like BRCA1 mutated breast cancers, no apparent specific somatic copy number aberration (CNA) profile for CHEK2*1100delC breast cancers was found. With the possible exception of copy number loss of chromosomal arm 1p in a subset of tumors, which might be involved in CHEK2 tumorigenesis. This difference in CNAs profiles might be explained by the need for BRCA1-deficient tumor cells to acquire survival factors, by for example specific copy number aberrations, to expand. Such factors may not be needed for breast tumors with a defect in a non-essential gene such as CHEK2.

Gene expression profiling of choline-deprived neural precursor cells isolated from mouse brain.

PubMed

Niculescu, Mihai D; Craciunescu, Corneliu N; Zeisel, Steven H

2005-04-04

Choline is an essential nutrient and an important methyl donor. Choline deficiency alters fetal development of the hippocampus in rodents and these changes are associated with decreased memory function lasting throughout life. Also, choline deficiency alters global and gene-specific DNA methylation in several models. This gene expression profiling study describes changes in cortical neural precursor cells from embryonic day 14 mice, after 48 h of exposure to a choline-deficient medium. Using Significance Analysis of Microarrays, we found the expression of 1003 genes to be significantly changed (from a total of 16,000 total genes spotted on the array), with a false discovery rate below 5%. A total of 846 genes were overexpressed while 157 were underexpressed. Classification by gene ontology revealed that 331 of these genes modulate cell proliferation, apoptosis, neuronal and glial differentiation, methyl metabolism, and calcium-binding protein classes. Twenty-seven genes that had changed expression have previously been reported to be regulated by promoter or intron methylation. These findings support our previous work suggesting that choline deficiency decreases the proliferation of neural precursors and possibly increases premature neuronal differentiation and apoptosis.

Somatic mutations affect key pathways in lung adenocarcinoma

PubMed Central

Ding, Li; Getz, Gad; Wheeler, David A.; Mardis, Elaine R.; McLellan, Michael D.; Cibulskis, Kristian; Sougnez, Carrie; Greulich, Heidi; Muzny, Donna M.; Morgan, Margaret B.; Fulton, Lucinda; Fulton, Robert S.; Zhang, Qunyuan; Wendl, Michael C.; Lawrence, Michael S.; Larson, David E.; Chen, Ken; Dooling, David J.; Sabo, Aniko; Hawes, Alicia C.; Shen, Hua; Jhangiani, Shalini N.; Lewis, Lora R.; Hall, Otis; Zhu, Yiming; Mathew, Tittu; Ren, Yanru; Yao, Jiqiang; Scherer, Steven E.; Clerc, Kerstin; Metcalf, Ginger A.; Ng, Brian; Milosavljevic, Aleksandar; Gonzalez-Garay, Manuel L.; Osborne, John R.; Meyer, Rick; Shi, Xiaoqi; Tang, Yuzhu; Koboldt, Daniel C.; Lin, Ling; Abbott, Rachel; Miner, Tracie L.; Pohl, Craig; Fewell, Ginger; Haipek, Carrie; Schmidt, Heather; Dunford-Shore, Brian H.; Kraja, Aldi; Crosby, Seth D.; Sawyer, Christopher S.; Vickery, Tammi; Sander, Sacha; Robinson, Jody; Winckler, Wendy; Baldwin, Jennifer; Chirieac, Lucian R.; Dutt, Amit; Fennell, Tim; Hanna, Megan; Johnson, Bruce E.; Onofrio, Robert C.; Thomas, Roman K.; Tonon, Giovanni; Weir, Barbara A.; Zhao, Xiaojun; Ziaugra, Liuda; Zody, Michael C.; Giordano, Thomas; Orringer, Mark B.; Roth, Jack A.; Spitz, Margaret R.; Wistuba, Ignacio I.; Ozenberger, Bradley; Good, Peter J.; Chang, Andrew C.; Beer, David G.; Watson, Mark A.; Ladanyi, Marc; Broderick, Stephen; Yoshizawa, Akihiko; Travis, William D.; Pao, William; Province, Michael A.; Weinstock, George M.; Varmus, Harold E.; Gabriel, Stacey B.; Lander, Eric S.; Gibbs, Richard A.; Meyerson, Matthew; Wilson, Richard K.

2009-01-01

Determining the genetic basis of cancer requires comprehensive analyses of large collections of histopathologically well-classified primary tumours. Here we report the results of a collaborative study to discover somatic mutations in 188 human lung adenocarcinomas. DNA sequencing of 623 genes with known or potential relationships to cancer revealed more than 1,000 somatic mutations across the samples. Our analysis identified 26 genes that are mutated at significantly high frequencies and thus are probably involved in carcinogenesis. The frequently mutated genes include tyrosine kinases, among them the EGFR homologue ERBB4; multiple ephrin receptor genes, notably EPHA3; vascular endothelial growth factor receptor KDR; and NTRK genes. These data provide evidence of somatic mutations in primary lung adenocarcinoma for several tumour suppressor genes involved in other cancers—including NF1, APC, RB1 and ATM—and for sequence changes in PTPRD as well as the frequently deleted gene LRP1B. The observed mutational profiles correlate with clinical features, smoking status and DNA repair defects. These results are reinforced by data integration including single nucleotide polymorphism array and gene expression array. Our findings shed further light on several important signalling pathways involved in lung adenocarcinoma, and suggest new molecular targets for treatment. PMID:18948947

Temporal patterns in the transcriptomic response of rainbow trout, Oncorhynchus mykiss, to crude oil.

PubMed

Hook, Sharon E; Lampi, Mark A; Febbo, Eric J; Ward, Jeff A; Parkerton, Thomas F

2010-09-01

Time is often not characterized as a variable in ecotoxicogenomic studies. In this study, temporal changes in gene expression were determined during exposure to crude oil and a subsequent recovery period. Juvenile rainbow trout, Oncorhynchus mykiss, were exposed for 96 h to the water accommodated fractions of 0.4, 2 or 10 mgl(-1) crude oil loadings. Following 96 h of exposure, fish were transferred to recovery tanks. Gill and liver samples were collected after 24 and 96 h of exposure, and after 96 h of recovery for RNA extraction and microarray analysis. Fluorescently labeled cDNA was hybridized against matched controls, using salmonid cDNA arrays. Each exposure scenario generated unique patterns of altered gene expression. More genes responded to crude oil in the gill than in the liver. In the gill, 1137 genes had altered expression at 24 h, 2003 genes had altered expression levels at 96 h of exposure, yet by 96 h of recovery, no genes were significantly altered in expression. In the liver at 10 mgl(-1), only five genes were changed at 24 h, yet 192 genes had altered expression after 96 h recovery. At 2 mgl(-1) in the liver, many genes had altered regulation at all three time points. The 0.4 mgl(-1) loading also showed 289 genes upregulated at 24 h after exposure. The Gene Ontology terms associated with altered expression in the liver suggested that the processes of protein synthesis, xenobiotic metabolism, and oxidoreductase activity were altered. The concentration-responsive expression profile of cytochrome P450 1A, a biomarker for oil exposure, did not predict the majority of gene expression profiles in any tissue or dose, since direct relationships with dose were not observed for most genes. While the genes and their associated functions agree with known modes of toxic action for crude oil, the gene lists obtained do not match our previously published work, presumably due to array analysis procedures. These results demonstrate that changes in gene expression with time and dose may be complicated, and should be characterized in controlled laboratory settings before attempts are made to interpret responses in field-collected organisms. Further, processes for analyzing microarray data need to be developed such that standardized gene lists are developed, or that analysis does not rely on lists of significantly altered genes before arrays can be further evaluated as a monitoring tool. Crown Copyright 2010. Published by Elsevier B.V. All rights reserved.

Tumor Touch Imprints as Source for Whole Genome Analysis of Neuroblastoma Tumors

PubMed Central

Brunner, Clemens; Brunner-Herglotz, Bettina; Ziegler, Andrea; Frech, Christian; Amann, Gabriele; Ladenstein, Ruth; Ambros, Inge M.; Ambros, Peter F.

2016-01-01

Introduction Tumor touch imprints (TTIs) are routinely used for the molecular diagnosis of neuroblastomas by interphase fluorescence in-situ hybridization (I-FISH). However, in order to facilitate a comprehensive, up-to-date molecular diagnosis of neuroblastomas and to identify new markers to refine risk and therapy stratification methods, whole genome approaches are needed. We examined the applicability of an ultra-high density SNP array platform that identifies copy number changes of varying sizes down to a few exons for the detection of genomic changes in tumor DNA extracted from TTIs. Material and Methods DNAs were extracted from TTIs of 46 neuroblastoma and 4 other pediatric tumors. The DNAs were analyzed on the Cytoscan HD SNP array platform to evaluate numerical and structural genomic aberrations. The quality of the data obtained from TTIs was compared to that from randomly chosen fresh or fresh frozen solid tumors (n = 212) and I-FISH validation was performed. Results SNP array profiles were obtained from 48 (out of 50) TTI DNAs of which 47 showed genomic aberrations. The high marker density allowed for single gene analysis, e.g. loss of nine exons in the ATRX gene and the visualization of chromothripsis. Data quality was comparable to fresh or fresh frozen tumor SNP profiles. SNP array results were confirmed by I-FISH. Conclusion TTIs are an excellent source for SNP array processing with the advantage of simple handling, distribution and storage of tumor tissue on glass slides. The minimal amount of tumor tissue needed to analyze whole genomes makes TTIs an economic surrogate source in the molecular diagnostic work up of tumor samples. PMID:27560999

Integrating Microarray Analysis and the Soybean Genome to Understand the Soybean's Iron Deficiency Response

USDA-ARS?s Scientific Manuscript database

Transcriptional profiles of soybean (Glycine max, L. Merr) near isogenic lines Clark (PI548553, iron efficient) and IsoClark (PI547430, iron inefficient) were analyzed and compared using the Affymetrix® GeneChip® Soybean Genome Array. A comparison of plants grown under Fe-sufficient and Fe-limited ...

A focused microarray approach to functional glycomics: transcriptional regulation of the glycome.

PubMed

Comelli, Elena M; Head, Steven R; Gilmartin, Tim; Whisenant, Thomas; Haslam, Stuart M; North, Simon J; Wong, Nyet-Kui; Kudo, Takashi; Narimatsu, Hisashi; Esko, Jeffrey D; Drickamer, Kurt; Dell, Anne; Paulson, James C

2006-02-01

Glycosylation is the most common posttranslational modification of proteins, yet genes relevant to the synthesis of glycan structures and function are incompletely represented and poorly annotated on the commercially available arrays. To fill the need for expression analysis of such genes, we employed the Affymetrix technology to develop a focused and highly annotated glycogene-chip representing human and murine glycogenes, including glycosyltransferases, nucleotide sugar transporters, glycosidases, proteoglycans, and glycan-binding proteins. In this report, the array has been used to generate glycogene-expression profiles of nine murine tissues. Global analysis with a hierarchical clustering algorithm reveals that expression profiles in immune tissues (thymus [THY], spleen [SPL], lymph node, and bone marrow [BM]) are more closely related, relative to those of nonimmune tissues (kidney [KID], liver [LIV], brain [BRN], and testes [TES]). Of the biosynthetic enzymes, those responsible for synthesis of the core regions of N- and O-linked oligosaccharides are ubiquitously expressed, whereas glycosyltransferases that elaborate terminal structures are expressed in a highly tissue-specific manner, accounting for tissue and ultimately cell-type-specific glycosylation. Comparison of gene expression profiles with matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) profiling of N-linked oligosaccharides suggested that the alpha1-3 fucosyltransferase 9, Fut9, is the enzyme responsible for terminal fucosylation in KID and BRN, a finding validated by analysis of Fut9 knockout mice. Two families of glycan-binding proteins, C-type lectins and Siglecs, are predominately expressed in the immune tissues, consistent with their emerging functions in both innate and acquired immunity. The glycogene chip reported in this study is available to the scientific community through the Consortium for Functional Glycomics (CFG) (http://www.functionalglycomics.org).

Arabidopsis Gene Family Profiler (aGFP)--user-oriented transcriptomic database with easy-to-use graphic interface.

PubMed

Dupl'áková, Nikoleta; Renák, David; Hovanec, Patrik; Honysová, Barbora; Twell, David; Honys, David

2007-07-23

Microarray technologies now belong to the standard functional genomics toolbox and have undergone massive development leading to increased genome coverage, accuracy and reliability. The number of experiments exploiting microarray technology has markedly increased in recent years. In parallel with the rapid accumulation of transcriptomic data, on-line analysis tools are being introduced to simplify their use. Global statistical data analysis methods contribute to the development of overall concepts about gene expression patterns and to query and compose working hypotheses. More recently, these applications are being supplemented with more specialized products offering visualization and specific data mining tools. We present a curated gene family-oriented gene expression database, Arabidopsis Gene Family Profiler (aGFP; http://agfp.ueb.cas.cz), which gives the user access to a large collection of normalised Affymetrix ATH1 microarray datasets. The database currently contains NASC Array and AtGenExpress transcriptomic datasets for various tissues at different developmental stages of wild type plants gathered from nearly 350 gene chips. The Arabidopsis GFP database has been designed as an easy-to-use tool for users needing an easily accessible resource for expression data of single genes, pre-defined gene families or custom gene sets, with the further possibility of keyword search. Arabidopsis Gene Family Profiler presents a user-friendly web interface using both graphic and text output. Data are stored at the MySQL server and individual queries are created in PHP script. The most distinguishable features of Arabidopsis Gene Family Profiler database are: 1) the presentation of normalized datasets (Affymetrix MAS algorithm and calculation of model-based gene-expression values based on the Perfect Match-only model); 2) the choice between two different normalization algorithms (Affymetrix MAS4 or MAS5 algorithms); 3) an intuitive interface; 4) an interactive "virtual plant" visualizing the spatial and developmental expression profiles of both gene families and individual genes. Arabidopsis GFP gives users the possibility to analyze current Arabidopsis developmental transcriptomic data starting with simple global queries that can be expanded and further refined to visualize comparative and highly selective gene expression profiles.

N-(3-Benzoylphenyl)-1H-Indole-2-Carboxamide decreases triglyceride levels by downregulation of Apoc3 gene expression in acute hyperlipidemic rat model.

PubMed

Hamadneh, Lama; Al-Essa, Luay; Hikmat, Suhair; Al-Qirim, Tariq; Abu Sheikha, Ghassan; Al-Hiari, Yusuf; Azmy, Nisrin; Shattat, Ghassan

2017-07-01

Hyperlipidemia is a known cause of coronary vascular diseases, which is a major cause of death in many parts of the world. Targeting several pathways that lead to increase in lipid profiles is of great potential to control diseases. 1H-indole-2-carboxamide derivatives were tested for their hypolipidemic activity at the molecular level in comparison with bezafibrate. The gene expression profiles of lipoprotein signaling and cholesterol metabolism and fatty acid metabolism PCR arrays were determined in rats with acute hyperlipidemia induced by Triton WR1339. Lipid profiles of serum from treated rats showed significant hypolipidemic effect by the compounds. Several genes of potential interest were reported to be overexpressed by Triton WR1339 including Apoc3, Apob, Hmgcs2, Apoa1, Apoe, Apof, acsl1, and Decr1. Most of the overexpressed genes were downregulated by N-(3-Benzoylphenyl)-1H-Indole-2-Carboxamide with significant decreases in Apoc3, Apob, Acaa2, Acsl1, and Slc247a5 gene expression levels. N-(4-Benzoylphenyl)-1H-Indole-2-Carboxamide and bezafibrate did not significantly affect the gene expression levels which were increased with acute hyperlipidemia induced by Triton WR1339. In conclusion, gene expression profiling identified the possible mechanism in which Triton WR1339 induces its acute hyperlipidemic effect which was reversed by the use of N-(3-Benzoylphenyl)-1H-Indole-2-Carboxamide.

IL-17A Mediates a Selective Gene Expression Profile in Asthmatic Human Airway Smooth Muscle Cells

PubMed Central

Dragon, Stéphane; Hirst, Stuart J.; Lee, Tak H.

2014-01-01

Airway smooth muscle (ASM) cells are thought to contribute to the pathogenesis of allergic asthma by orchestrating and perpetuating airway inflammation and remodeling responses. In this study, we evaluated the IL-17RA signal transduction and gene expression profile in ASM cells from subjects with mild asthma and healthy individuals. Human primary ASM cells were treated with IL-17A and probed by the Affymetrix GeneChip array, and gene targets were validated by real-time quantitative RT-PCR. Genomic analysis underlined the proinflammatory nature of IL-17A, as multiple NF-κB regulatory factors and chemokines were induced in ASM cells. Transcriptional regulators consisting of primary response genes were overrepresented and displayed dynamic expression profiles. IL-17A poorly enhanced IL-1β or IL-22 gene responses in ASM cells from both subjects with mild asthma and healthy donors. Interestingly, protein modifications to the NF-κB regulatory network were not observed after IL-17A stimulation, although oscillations in IκBε expression were detected. ASM cells from subjects with mild asthma up-regulated more genes with greater overall variability in response to IL-17A than from healthy donors. Finally, in response to IL-17A, ASM cells displayed rapid activation of the extracellular signal–regulated kinase/ribosomal S6 kinase signaling pathway and increased nuclear levels of phosphorylated extracellular signal–regulated kinase. Taken together, our results suggest that IL-17A mediated modest gene expression response, which, in cooperation with the NF-κB signaling network, may regulate the gene expression profile in ASM cells. PMID:24393021

Gene expression profiles in rainbow trout, Onchorynchus mykiss, exposed to a simple chemical mixture.

PubMed

Hook, Sharon E; Skillman, Ann D; Gopalan, Banu; Small, Jack A; Schultz, Irvin R

2008-03-01

Among proposed uses for microarrays in environmental toxiciology is the identification of key contributors to toxicity within a mixture. However, it remains uncertain whether the transcriptomic profiles resulting from exposure to a mixture have patterns of altered gene expression that contain identifiable contributions from each toxicant component. We exposed isogenic rainbow trout Onchorynchus mykiss, to sublethal levels of ethynylestradiol, 2,2,4,4-tetrabromodiphenyl ether, and chromium VI or to a mixture of all three toxicants Fluorescently labeled complementary DNA (cDNA) were generated and hybridized against a commercially available Salmonid array spotted with 16,000 cDNAs. Data were analyzed using analysis of variance (p<0.05) with a Benjamani-Hochberg multiple test correction (Genespring [Agilent] software package) to identify up and downregulated genes. Gene clustering patterns that can be used as "expression signatures" were determined using hierarchical cluster analysis. The gene ontology terms associated with significantly altered genes were also used to identify functional groups that were associated with toxicant exposure. Cross-ontological analytics approach was used to assign functional annotations to genes with "unknown" function. Our analysis indicates that transcriptomic profiles resulting from the mixture exposure resemble those of the individual contaminant exposures, but are not a simple additive list. However, patterns of altered genes representative of each component of the mixture are clearly discernible, and the functional classes of genes altered represent the individual components of the mixture. These findings indicate that the use of microarrays to identify transcriptomic profiles may aid in the identification of key stressors within a chemical mixture, ultimately improving environmental assessment.

CGI: Java Software for Mapping and Visualizing Data from Array-based Comparative Genomic Hybridization and Expression Profiling

PubMed Central

Gu, Joyce Xiuweu-Xu; Wei, Michael Yang; Rao, Pulivarthi H.; Lau, Ching C.; Behl, Sanjiv; Man, Tsz-Kwong

2007-01-01

With the increasing application of various genomic technologies in biomedical research, there is a need to integrate these data to correlate candidate genes/regions that are identified by different genomic platforms. Although there are tools that can analyze data from individual platforms, essential software for integration of genomic data is still lacking. Here, we present a novel Java-based program called CGI (Cytogenetics-Genomics Integrator) that matches the BAC clones from array-based comparative genomic hybridization (aCGH) to genes from RNA expression profiling datasets. The matching is computed via a fast, backend MySQL database containing UCSC Genome Browser annotations. This program also provides an easy-to-use graphical user interface for visualizing and summarizing the correlation of DNA copy number changes and RNA expression patterns from a set of experiments. In addition, CGI uses a Java applet to display the copy number values of a specific BAC clone in aCGH experiments side by side with the expression levels of genes that are mapped back to that BAC clone from the microarray experiments. The CGI program is built on top of extensible, reusable graphic components specifically designed for biologists. It is cross-platform compatible and the source code is freely available under the General Public License. PMID:19936083

CGI: Java software for mapping and visualizing data from array-based comparative genomic hybridization and expression profiling.

PubMed

Gu, Joyce Xiuweu-Xu; Wei, Michael Yang; Rao, Pulivarthi H; Lau, Ching C; Behl, Sanjiv; Man, Tsz-Kwong

2007-10-06

With the increasing application of various genomic technologies in biomedical research, there is a need to integrate these data to correlate candidate genes/regions that are identified by different genomic platforms. Although there are tools that can analyze data from individual platforms, essential software for integration of genomic data is still lacking. Here, we present a novel Java-based program called CGI (Cytogenetics-Genomics Integrator) that matches the BAC clones from array-based comparative genomic hybridization (aCGH) to genes from RNA expression profiling datasets. The matching is computed via a fast, backend MySQL database containing UCSC Genome Browser annotations. This program also provides an easy-to-use graphical user interface for visualizing and summarizing the correlation of DNA copy number changes and RNA expression patterns from a set of experiments. In addition, CGI uses a Java applet to display the copy number values of a specific BAC clone in aCGH experiments side by side with the expression levels of genes that are mapped back to that BAC clone from the microarray experiments. The CGI program is built on top of extensible, reusable graphic components specifically designed for biologists. It is cross-platform compatible and the source code is freely available under the General Public License.

Effects of soluble and particulate Cr(VI) on genome-wide DNA methylation in human B lymphoblastoid cells.

PubMed

Lou, Jianlin; Wang, Yu; Chen, Junqiang; Ju, Li; Yu, Min; Jiang, Zhaoqiang; Feng, Lingfang; Jin, Lingzhi; Zhang, Xing

2015-10-01

Several previous studies highlighted the potential epigenetic effects of Cr(VI), especially DNA methylation. However, few studies have compared the effects of Cr(VI) on DNA methylation profiles between soluble and particulate chromate in vitro. Accordingly, Illumina Infinium Human Methylation 450K BeadChip array was used to analyze DNA methylation profiles of human B lymphoblastoid cells exposed to potassium dichromate or lead chromate, and the cell viability was also studied. Array based DNA methylation analysis showed that the impacts of Cr(VI) on DNA methylation were limited, only about 40 differentially methylated CpG sites, with an overlap of 15CpG sites, were induced by both potassium dichromate and lead chromate. The results of mRNA expression showed that after Cr(VI) treatment, mRNA expression changes of four genes (TBL1Y, FZD5, IKZF2, and KIAA1949) were consistent with their DNA methylation alteration, but DNA methylation changes of other six genes did not correlate with mRNA expression. In conclusion, both of soluble and particulate Cr(VI) could induce a small amount of differentially methylated sites in human B lymphoblastoid cells, and the correlations between DNA methylation changes and mRNA expression varied between different genes. Copyright © 2015 Elsevier B.V. All rights reserved.

Synthetic Genetic Arrays: Automation of Yeast Genetics.

PubMed

Kuzmin, Elena; Costanzo, Michael; Andrews, Brenda; Boone, Charles

2016-04-01

Genome-sequencing efforts have led to great strides in the annotation of protein-coding genes and other genomic elements. The current challenge is to understand the functional role of each gene and how genes work together to modulate cellular processes. Genetic interactions define phenotypic relationships between genes and reveal the functional organization of a cell. Synthetic genetic array (SGA) methodology automates yeast genetics and enables large-scale and systematic mapping of genetic interaction networks in the budding yeast,Saccharomyces cerevisiae SGA facilitates construction of an output array of double mutants from an input array of single mutants through a series of replica pinning steps. Subsequent analysis of genetic interactions from SGA-derived mutants relies on accurate quantification of colony size, which serves as a proxy for fitness. Since its development, SGA has given rise to a variety of other experimental approaches for functional profiling of the yeast genome and has been applied in a multitude of other contexts, such as genome-wide screens for synthetic dosage lethality and integration with high-content screening for systematic assessment of morphology defects. SGA-like strategies can also be implemented similarly in a number of other cell types and organisms, includingSchizosaccharomyces pombe,Escherichia coli, Caenorhabditis elegans, and human cancer cell lines. The genetic networks emerging from these studies not only generate functional wiring diagrams but may also play a key role in our understanding of the complex relationship between genotype and phenotype. © 2016 Cold Spring Harbor Laboratory Press.

Clofibrate-induced gene expression changes in rat liver: a cross-laboratory analysis using membrane cDNA arrays.

PubMed Central

Baker, Valerie A; Harries, Helen M; Waring, Jeff F; Duggan, Colette M; Ni, Hong A; Jolly, Robert A; Yoon, Lawrence W; De Souza, Angus T; Schmid, Judith E; Brown, Roger H; Ulrich, Roger G; Rockett, John C

2004-01-01

Microarrays have the potential to significantly impact our ability to identify toxic hazards by the identification of mechanistically relevant markers of toxicity. To be useful for risk assessment, however, microarray data must be challenged to determine reliability and interlaboratory reproducibility. As part of a series of studies conducted by the International Life Sciences Institute Health and Environmental Science Institute Technical Committee on the Application of Genomics to Mechanism-Based Risk Assessment, the biological response in rats to the hepatotoxin clofibrate was investigated. Animals were treated with high (250 mg/kg/day) or low (25 mg/kg/day) doses for 1, 3, or 7 days in two laboratories. Clinical chemistry parameters were measured, livers removed for histopathological assessment, and gene expression analysis was conducted using cDNA arrays. Expression changes in genes involved in fatty acid metabolism (e.g., acyl-CoA oxidase), cell proliferation (e.g., topoisomerase II-Alpha), and fatty acid oxidation (e.g., cytochrome P450 4A1), consistent with the mechanism of clofibrate hepatotoxicity, were detected. Observed differences in gene expression levels correlated with the level of biological response induced in the two in vivo studies. Generally, there was a high level of concordance between the gene expression profiles generated from pooled and individual RNA samples. Quantitative real-time polymerase chain reaction was used to confirm modulations for a number of peroxisome proliferator marker genes. Though the results indicate some variability in the quantitative nature of the microarray data, this appears due largely to differences in experimental and data analysis procedures used within each laboratory. In summary, this study demonstrates the potential for gene expression profiling to identify toxic hazards by the identification of mechanistically relevant markers of toxicity. PMID:15033592

Comparative analysis of gene expression profiles of OPN signaling pathway in four kinds of liver diseases.

PubMed

Wang, Gaiping; Chen, Shasha; Zhao, Congcong; Li, Xiaofang; Zhao, Weiming; Yang, Jing; Chang, Cuifang; Xu, Cunshuan

2016-09-01

To explore the relevance of OPN signalling pathway to the occurrence and development of nonalcoholic fatty liver disease (NAFLD), liver cirrhosis (LC), hepatic cancer (HC) and acute hepatic failure (AHF) at transcriptional level, Rat Genome 230 2.0 Array was used to detect expression profiles of OPN signalling pathway-related genes in four kinds of liver diseases. The results showed that 23, 33, 59 and 74 genes were significantly changed in the above four kinds of liver diseases, respectively. H-clustering analysis showed that the expression profiles of OPN signalling-related genes were notably different in four kinds of liver diseases. Subsequently, a total of above-mentioned 147 genes were categorized into four clusters by k-means according to the similarity of gene expression, and expression analysis systematic explorer (EASE) functional enrichment analysis revealed that OPN signalling pathway-related genes were involved in cell adhesion and migration, cell proliferation, apoptosis, stress and inflammatory reaction, etc. Finally, ingenuity pathway analysis (IPA) software was used to predict the functions of OPN signalling-related genes, and the results indicated that the activities of ROS production, cell adhesion and migration, cell proliferation were remarkably increased, while that of apoptosis, stress and inflammatory reaction were reduced in four kinds of liver diseases. In summary, the above physiological activities changed more obviously in LC, HC and AHF than in NAFLD.

Differentially expressed genes in nonsmall cell lung cancer: expression profiling of cancer-related genes in squamous cell lung cancer.

PubMed

Kettunen, Eeva; Anttila, Sisko; Seppänen, Jouni K; Karjalainen, Antti; Edgren, Henrik; Lindström, Irmeli; Salovaara, Reijo; Nissén, Anna-Maria; Salo, Jarmo; Mattson, Karin; Hollmén, Jaakko; Knuutila, Sakari; Wikman, Harriet

2004-03-01

The expression patterns of cancer-related genes in 13 cases of squamous cell lung cancer (SCC) were characterized and compared with those in normal lung tissue and 13 adenocarcinomas (AC), the other major type of nonsmall cell lung cancer (NSCLC). cDNA array was used to screen the gene expression levels and the array results were verified using a real-time reverse-transcriptase-polymerase chain reaction (RT-PCR). Thirty-nine percent of the 25 most upregulated and the 25 most downregulated genes were common to SCC and AC. Of these genes, DSP, HMGA1 (alias HMGIY), TIMP1, MIF, CCNB1, TN, MMP11, and MMP12 were upregulated and COPEB (alias CPBP), TYROBP, BENE, BMPR2, SOCS3, TIMP3, CAV1, and CAV2 were downregulated. The expression levels of several genes from distinct protein families (cytokeratins and hemidesmosomal proteins) were markedly increased in SCC compared with AC and normal lung. In addition, several genes, overexpressed in SCC, such as HMGA1, CDK4, IGFBP3, MMP9, MMP11, MMP12, and MMP14, fell into distinct chromosomal loci, which we have detected as gained regions on the basis of comparative genomic hybridization data. Our study revealed new candidate genes involved in NSCLC.

Effects of the EVCAM chemical validation library on differentiation using marker gene expression in lmouse embryonic stem cells

EPA Science Inventory

The adherent cell differentiation and cytotoxicity (ACDC) assay was used to profile the effects of the ECVAM EST validation chemical library (19 compounds) on J1 mouse embryonic stem cells (mESC). PCR-based TaqMan Low Density Arrays (TLDA) provided a high-content assessment of al...

Construction and application of a bovine immune-endocrine cDNA microarray.

PubMed

Tao, Wenjing; Mallard, Bonnie; Karrow, Niel; Bridle, Byram

2004-09-01

A variety of commercial DNA arrays specific for humans and rodents are widely available; however, microarrays containing well-characterized genes to study pathway-specific gene expression are not as accessible for domestic animals, such as cattle, sheep and pigs. Therefore, a small-scale application-targeted bovine immune-endocrine cDNA array was developed to evaluate genetic pathways involved in the immune-endocrine axis of cattle during periods of altered homeostasis provoked by physiological or environmental stressors, such as infection, vaccination or disease. For this purpose, 167 cDNA sequences corresponding to immune, endocrine and inflammatory response genes were collected and categorized. Positive controls included 5 housekeeping genes (glyceraldehydes-3-phosphate dehydrogenase, hypoxanthine phosphoribosyltransferase, ribosomal protein L19, beta-actin, beta2-microglobulin) and bovine genomic DNA. Negative controls were a bacterial gene (Rhodococcus equi 17-kDa virulence-associated protein) and a partial sequence of the plasmid pACYC177. In addition, RNA extracted from un-stimulated, as well as superantigen (Staphylococcus aureus enterotoxin-A, S. aureus Cowan Pansorbin Cells) and mitogen-stimulated (LPS, ConA) bovine blood leukocytes was mixed, reverse transcribed and PCR amplified using gene-specific primers. The endocrine-associated genes were amplified from cDNA derived from un-stimulated bovine hypothalamus, pituitary, adrenal and thyroid gland tissues. The array was constructed in 4 repeating grids of 180 duplicated spots by coupling the PCR amplified 213-630 bp gene fragments onto poly-l-lysine coated glass slides. The bovine immune-endocrine arrays were standardized and preliminary gene expression profiles generated using Cy3 and Cy5 labelled cDNA from un-stimulated and ConA (5 microg/ml) stimulated PBMC of 4 healthy Holstein cows (2-4 replicate arrays/cow) in a time course study. Mononuclear cell-derived cytokine and chemokine (IL-2, IL-1alpha, TNFalpha, IFN-gamma, TGFbeta-1, MCP-1, MCP-2 and MIP-3alpha) mRNA exhibited a repeatable and consistently low expression in un-stimulated cells and at least a two-fold increased expression following 6 and 24 h ConA stimulation as compared to 0 h un-stimulated controls. In contrast, expression of antigen presenting molecules, MHC-DR, MHC-DQ and MHC-DY, were consistently at least two-fold lower following 6 and 24 h ConA stimulation. The only endocrine gene with differential expression following ConA stimulation was prolactin. Additionally, due to the high level of genetic homology between ovine, swine and bovine genes, RNA similarly acquired from sheep and pigs was evaluated and similar gene expression patterns were noted. These data demonstrate that this application-targeted array containing a set of well characterized genes can be used to determine the relative gene expression corresponding to immune-endocrine responses of cattle and related species, sheep and pigs.

Proof of Concept Study to Assess Fetal Gene Expression in Amniotic Fluid by NanoArray PCR

PubMed Central

Massingham, Lauren J.; Johnson, Kirby L.; Bianchi, Diana W.; Pei, Shermin; Peter, Inga; Cowan, Janet M.; Tantravahi, Umadevi; Morrison, Tom B.

2011-01-01

Microarray analysis of cell-free RNA in amniotic fluid (AF) supernatant has revealed differential fetal gene expression as a function of gestational age and karyotype. Once informative genes are identified, research moves to a more focused platform such as quantitative reverse transcriptase-PCR. Standardized NanoArray PCR (SNAP) is a recently developed gene profiling technology that enables the measurement of transcripts from samples containing reduced quantities or degraded nucleic acids. We used a previously developed SNAP gene panel as proof of concept to determine whether fetal functional gene expression could be ascertained from AF supernatant. RNA was extracted and converted to cDNA from 19 AF supernatant samples of euploid fetuses between 15 to 20 weeks of gestation, and transcript abundance of 21 genes was measured. Statistically significant differences in expression, as a function of advancing gestational age, were observed for 5 of 21 genes. ANXA5, GUSB, and PPIA showed decreasing gene expression over time, whereas CASC3 and ZNF264 showed increasing gene expression over time. Statistically significantly increased expression of MTOR and STAT2 was seen in female compared with male fetuses. This study demonstrates the feasibility of focused fetal gene expression analysis using SNAP technology. In the future, this technique could be optimized to examine specific genes instrumental in fetal organ system function, which could be a useful addition to prenatal care. PMID:21827969

Gene Expression Profiling of Multiple Leiomyomata Uteri and Matched Normal Tissue from a Single Patient

PubMed Central

Dimitrova, Irina K.; Richer, Jennifer K.; Rudolph, Michael C.; Spoelstra, Nicole S.; Reno, Elaine M.; Medina, Theresa M.; Bradford, Andrew P.

2009-01-01

Objective To identify differentially expressed genes between fibroid and adjacent normal myometrium in an identical hormonal and genetic background. Design Array analysis of 3 leiomyomata and matched adjacent normal myometrium in a single patient. Setting University of Colorado Hospital. Patient(s) A single female undergoing medically indicated hysterectomy for symptomatic fibroids. Interventions(s) mRNA isolation and microarray analysis, reverse-transcriptase polymerase chain reaction, western blotting and immunohistochemistry. Main Outcome Measure(s) Changes in mRNA and protein levels in leiomyomata and matched normal myometrium. Result(s) Expression of 197 genes was increased and 619 decreased, significantly by at least 2 fold, in leiomyomata relative to normal myometrium. Expression profiles between tumors were similar and normal myometrial samples showed minimal variation. Changes in, and variation of, expression of selected genes were confirmed in additional normal and leiomyoma samples from multiple patients. Conclusion(s) Analysis of multiple tumors from a single patient confirmed changes in expression of genes described in previous, apparently disparate, studies and identified novel targets. Gene expression profiles in leiomyomata are consistent with increased activation of mitogenic pathways and inhibition of apoptosis. Down-regulation of genes implicated in invasion and metastasis, of cancers, was observed in fibroids. This expression pattern may underlie the benign nature of uterine leiomyomata and may aid in the differential diagnosis of leiomyosarcoma. PMID:18672237

Toward a comprehensive and systematic methylome signature in colorectal cancers.

PubMed

Ashktorab, Hassan; Rahi, Hamed; Wansley, Daniel; Varma, Sudhir; Shokrani, Babak; Lee, Edward; Daremipouran, Mohammad; Laiyemo, Adeyinka; Goel, Ajay; Carethers, John M; Brim, Hassan

2013-08-01

CpG Island Methylator Phenotype (CIMP) is one of the underlying mechanisms in colorectal cancer (CRC). This study aimed to define a methylome signature in CRC through a methylation microarray analysis and a compilation of promising CIMP markers from the literature. Illumina HumanMethylation27 (IHM27) array data was generated and analyzed based on statistical differences in methylation data (1st approach) or based on overall differences in methylation percentages using lower 95% CI (2nd approach). Pyrosequencing was performed for the validation of nine genes. A meta-analysis was used to identify CIMP and non-CIMP markers that were hypermethylated in CRC but did not yet make it to the CIMP genes' list. Our 1st approach for array data analysis demonstrated the limitations in selecting genes for further validation, highlighting the need for the 2nd bioinformatics approach to adequately select genes with differential aberrant methylation. A more comprehensive list, which included non-CIMP genes, such as APC, EVL, CD109, PTEN, TWIST1, DCC, PTPRD, SFRP1, ICAM5, RASSF1A, EYA4, 30ST2, LAMA1, KCNQ5, ADHEF1, and TFPI2, was established. Array data are useful to categorize and cluster colonic lesions based on their global methylation profiles; however, its usefulness in identifying robust methylation markers is limited and rely on the data analysis method. We have identified 16 non-CIMP-panel genes for which we provide rationale for inclusion in a more comprehensive characterization of CIMP+ CRCs. The identification of a definitive list for methylome specific genes in CRC will contribute to better clinical management of CRC patients.

Gene expression profiles in liver of mouse after chronic exposure to drinking water.

PubMed

Wu, Bing; Zhang, Yan; Zhao, Dayong; Zhang, Xuxiang; Kong, Zhiming; Cheng, Shupei

2009-10-01

cDNA micorarray approach was applied to hepatic transcriptional profile analysis in male mouse (Mus musculus, ICR) to assess the potential health effects of drinking water in Nanjing, China. Mice were treated with continuous exposure to drinking water for 90 days. Hepatic gene expression was analyzed with Affymetrix Mouse Genome 430A 2.0 arrays, and pathway analysis was carried out by Molecule Annotation System 2.0 and KEGG pathway database. A total of 836 genes were found to be significantly altered (1.5-fold, P < or = 0.05), including 294 up-regulated genes and 542 down-regulated genes. According to biological pathway analysis, drinking water exposure resulted in aberration of gene expression and biological pathways linked to xenobiotic metabolism, signal transduction, cell cycle and oxidative stress response. Further, deregulation of several genes associated with carcinogenesis or tumor progression including Ccnd1, Egfr, Map2k3, Mcm2, Orc2l and Smad2 was observed. Although transcription changes in identified genes are unlikely to be used as a sole indicator of adverse health effects, the results of this study could enhance our understanding of early toxic effects of drinking water exposure and support future studies on drinking water safety.

Identification of Mouse Serum miRNA Endogenous References by Global Gene Expression Profiles

PubMed Central

Mi, Qing-Sheng; Weiland, Matthew; Qi, Rui-Qun; Gao, Xing-Hua; Poisson, Laila M.; Zhou, Li

2012-01-01

MicroRNAs (miRNAs) are recently discovered small non-coding RNAs and can serve as serum biomarkers for disease diagnosis and prognoses. Lack of reliable serum miRNA endogenous references for normalization in miRNA gene expression makes single miRNA assays inaccurate. Using TaqMan® real-time PCR miRNA arrays with a global gene expression normalization strategy, we have analyzed serum miRNA expression profiles of 20 female mice of NOD/ShiLtJ (n = 8), NOR/LtJ (n = 6), and C57BL/6J (n = 6) at different ages and disease conditions. We identified five miRNAs, miR-146a, miR-16, miR-195, miR-30e and miR-744, to be stably expressed in all strains, which could serve as mouse serum miRNA endogenous references for single assay experiments. PMID:22348064

Blood cell gene expression associated with cellular stress defense is modulated by antioxidant-rich food in a randomised controlled clinical trial of male smokers.

PubMed

Bøhn, Siv K; Myhrstad, Mari C; Thoresen, Magne; Holden, Marit; Karlsen, Anette; Tunheim, Siv Haugen; Erlund, Iris; Svendsen, Mette; Seljeflot, Ingebjørg; Moskaug, Jan O; Duttaroy, Asim K; Laake, Petter; Arnesen, Harald; Tonstad, Serena; Collins, Andrew; Drevon, Christan A; Blomhoff, Rune

2010-09-16

Plant-based diets rich in fruit and vegetables can prevent development of several chronic age-related diseases. However, the mechanisms behind this protective effect are not elucidated. We have tested the hypothesis that intake of antioxidant-rich foods can affect groups of genes associated with cellular stress defence in human blood cells. NCT00520819 http://clinicaltrials.gov. In an 8-week dietary intervention study, 102 healthy male smokers were randomised to either a diet rich in various antioxidant-rich foods, a kiwifruit diet (three kiwifruits/d added to the regular diet) or a control group. Blood cell gene expression profiles were obtained from 10 randomly selected individuals of each group. Diet-induced changes on gene expression were compared to controls using a novel application of the gene set enrichment analysis (GSEA) on transcription profiles obtained using Affymetrix HG-U133-Plus 2.0 whole genome arrays. Changes were observed in the blood cell gene expression profiles in both intervention groups when compared to the control group. Groups of genes involved in regulation of cellular stress defence, such as DNA repair, apoptosis and hypoxia, were significantly upregulated (GSEA, FDR q-values < 5%) by both diets compared to the control group. Genes with common regulatory motifs for aryl hydrocarbon receptor (AhR) and AhR nuclear translocator (AhR/ARNT) were upregulated by both interventions (FDR q-values < 5%). Plasma antioxidant biomarkers (polyphenols/carotenoids) increased in both groups. The observed changes in the blood cell gene expression profiles suggest that the beneficial effects of a plant-based diet on human health may be mediated through optimization of defence processes.

Metabolomic and Transcriptomic Comparison of Solid-State and Submerged Fermentation of Penicillium expansum KACC 40815

PubMed Central

Park, Hye Min; Singh, Digar; Lee, Choong Hwan

2016-01-01

Penicillium spp. are known to harbor a wide array of secondary metabolites with cryptic bioactivities. However, the metabolomics of these species is not well-understood in terms of different fermentation models and conditions. The present study involved metabolomics profiling and transcriptomic analysis of Penicillium expansum 40815 under solid-state fermentation (SSF) and submerged fermentation (SmF). Metabolite profiling was carried out using ultra-performance liquid chromatography quadruple time-of-flight mass spectrometry with multivariate analysis, followed by transcriptomic analyses of differentially expressed genes. In principal component analysis, the metabolite profiling data was studied under different experimental sets, including SSF and SmF. The significantly different metabolites such as polyketide metabolites (agonodepside B, rotiorin, verrucosidin, and ochrephilone) and corresponding gene transcripts (polyketide synthase, aromatic prenyltransferase, and terpenoid synthase) were primarily detected under SmF conditions. In contrast, the meroterpenoid compounds (andrastin A and C) and their genes transcripts were exclusively detected under SSF conditions. We demonstrated that the metabolite production and its corresponding gene expression levels in P. expansum 40815 were significantly influenced by the varying growth parameters and the immediate environment. This study further provides a foundation to produce specific metabolites by regulating fermentation conditions. PMID:26863302

Metabolomic and Transcriptomic Comparison of Solid-State and Submerged Fermentation of Penicillium expansum KACC 40815.

PubMed

Kim, Hyang Yeon; Heo, Do Yeon; Park, Hye Min; Singh, Digar; Lee, Choong Hwan

2016-01-01

Penicillium spp. are known to harbor a wide array of secondary metabolites with cryptic bioactivities. However, the metabolomics of these species is not well-understood in terms of different fermentation models and conditions. The present study involved metabolomics profiling and transcriptomic analysis of Penicillium expansum 40815 under solid-state fermentation (SSF) and submerged fermentation (SmF). Metabolite profiling was carried out using ultra-performance liquid chromatography quadruple time-of-flight mass spectrometry with multivariate analysis, followed by transcriptomic analyses of differentially expressed genes. In principal component analysis, the metabolite profiling data was studied under different experimental sets, including SSF and SmF. The significantly different metabolites such as polyketide metabolites (agonodepside B, rotiorin, verrucosidin, and ochrephilone) and corresponding gene transcripts (polyketide synthase, aromatic prenyltransferase, and terpenoid synthase) were primarily detected under SmF conditions. In contrast, the meroterpenoid compounds (andrastin A and C) and their genes transcripts were exclusively detected under SSF conditions. We demonstrated that the metabolite production and its corresponding gene expression levels in P. expansum 40815 were significantly influenced by the varying growth parameters and the immediate environment. This study further provides a foundation to produce specific metabolites by regulating fermentation conditions.

A combined analysis of genome-wide expression profiling of bipolar disorder in human prefrontal cortex.

PubMed

Wang, Jinglu; Qu, Susu; Wang, Weixiao; Guo, Liyuan; Zhang, Kunlin; Chang, Suhua; Wang, Jing

2016-11-01

Numbers of gene expression profiling studies of bipolar disorder have been published. Besides different array chips and tissues, variety of the data processes in different cohorts aggravated the inconsistency of results of these genome-wide gene expression profiling studies. By searching the gene expression databases, we obtained six data sets for prefrontal cortex (PFC) of bipolar disorder with raw data and combinable platforms. We used standardized pre-processing and quality control procedures to analyze each data set separately and then combined them into a large gene expression matrix with 101 bipolar disorder subjects and 106 controls. A standard linear mixed-effects model was used to calculate the differentially expressed genes (DEGs). Multiple levels of sensitivity analyses and cross validation with genetic data were conducted. Functional and network analyses were carried out on basis of the DEGs. In the result, we identified 198 unique differentially expressed genes in the PFC of bipolar disorder and control. Among them, 115 DEGs were robust to at least three leave-one-out tests or different pre-processing methods; 51 DEGs were validated with genetic association signals. Pathway enrichment analysis showed these DEGs were related with regulation of neurological system, cell death and apoptosis, and several basic binding processes. Protein-protein interaction network further identified one key hub gene. We have contributed the most comprehensive integrated analysis of bipolar disorder expression profiling studies in PFC to date. The DEGs, especially those with multiple validations, may denote a common signature of bipolar disorder and contribute to the pathogenesis of disease. Copyright © 2016 Elsevier Ltd. All rights reserved.

Molecular profiling of single circulating tumor cells from lung cancer patients.

PubMed

Park, Seung-Min; Wong, Dawson J; Ooi, Chin Chun; Kurtz, David M; Vermesh, Ophir; Aalipour, Amin; Suh, Susie; Pian, Kelsey L; Chabon, Jacob J; Lee, Sang Hun; Jamali, Mehran; Say, Carmen; Carter, Justin N; Lee, Luke P; Kuschner, Ware G; Schwartz, Erich J; Shrager, Joseph B; Neal, Joel W; Wakelee, Heather A; Diehn, Maximilian; Nair, Viswam S; Wang, Shan X; Gambhir, Sanjiv S

2016-12-27

Circulating tumor cells (CTCs) are established cancer biomarkers for the "liquid biopsy" of tumors. Molecular analysis of single CTCs, which recapitulate primary and metastatic tumor biology, remains challenging because current platforms have limited throughput, are expensive, and are not easily translatable to the clinic. Here, we report a massively parallel, multigene-profiling nanoplatform to compartmentalize and analyze hundreds of single CTCs. After high-efficiency magnetic collection of CTC from blood, a single-cell nanowell array performs CTC mutation profiling using modular gene panels. Using this approach, we demonstrated multigene expression profiling of individual CTCs from non-small-cell lung cancer (NSCLC) patients with remarkable sensitivity. Thus, we report a high-throughput, multiplexed strategy for single-cell mutation profiling of individual lung cancer CTCs toward minimally invasive cancer therapy prediction and disease monitoring.

Analysis of cellular response by exposure to acute or chronic radiation in human lymphoblastoid TK-6 cells

NASA Astrophysics Data System (ADS)

Ohnishi, T.; Yasumoto, J.; Takahashi, A.; Ohnishi, K.

To clarify the biological effects of low-dose rate radiation on human health for long-term stay in space, we analyzed the induction of apoptosis and apoptosis-related gene expression after irradiation with different dose-rate in human lymphoblastoid TK-6 cells harboring wild-type p53 gene. We irradiated TK-6 cells by X-ray at 1.5 Gy (1 Gy/min) and then sampled at 25 hr after culturing. We also irradiated by gamma-ray at 1.5 Gy (1 mGy/min) and then sampled immediately or 25 hr after irradiation. For DNA ladder analysis, we extracted DNA from these samples and electrophoresed with 2% agarose gel. In addition, we extracted mRNA from these samples for DNA-array analysis. mRNA from non-irradiated cells was used as a control. After labeling the cDNA against mRNA with [α -33P]-dCTP and hybridizing onto DNA array (Human Apoptosis Expression Array, R&D Systems), we scanned the profiles of the spots by a phosphorimager (BAS5000, FUJI FILM) and calculated using a NIH Image program. The data of each DNA-array were normalized with eight kinds of house keeping genes. We analyzed the expression level of apoptosis-related genes such as p53-related, Bcl-2 family, Caspase family and Fas-related genes. DNA ladders were obviously detected in the cells exposed to a high dose-rate radiation. We detected the induction of the gene expression of apoptosis-promotive genes. In contrast, almost no apoptosis was observed in the cells exposed to the chronic radiation at a low dose-rate. In addition, we detected the induction of the gene expression of apoptosis-suppressive genes as compared with apoptosis promotive-genes immediately after chronic irradiation. These results lead the importance of biological meaning of exposure to radiation at low dose-rate from an aspect of carcinogenesis. Finally, the effects of chronic irradiation become a highly important issue in space radiation biology for human health.

Fluorescence-based bioassays for the detection and evaluation of food materials.

PubMed

Nishi, Kentaro; Isobe, Shin-Ichiro; Zhu, Yun; Kiyama, Ryoiti

2015-10-13

We summarize here the recent progress in fluorescence-based bioassays for the detection and evaluation of food materials by focusing on fluorescent dyes used in bioassays and applications of these assays for food safety, quality and efficacy. Fluorescent dyes have been used in various bioassays, such as biosensing, cell assay, energy transfer-based assay, probing, protein/immunological assay and microarray/biochip assay. Among the arrays used in microarray/biochip assay, fluorescence-based microarrays/biochips, such as antibody/protein microarrays, bead/suspension arrays, capillary/sensor arrays, DNA microarrays/polymerase chain reaction (PCR)-based arrays, glycan/lectin arrays, immunoassay/enzyme-linked immunosorbent assay (ELISA)-based arrays, microfluidic chips and tissue arrays, have been developed and used for the assessment of allergy/poisoning/toxicity, contamination and efficacy/mechanism, and quality control/safety. DNA microarray assays have been used widely for food safety and quality as well as searches for active components. DNA microarray-based gene expression profiling may be useful for such purposes due to its advantages in the evaluation of pathway-based intracellular signaling in response to food materials.

Fluorescence-Based Bioassays for the Detection and Evaluation of Food Materials

PubMed Central

Nishi, Kentaro; Isobe, Shin-Ichiro; Zhu, Yun; Kiyama, Ryoiti

2015-01-01

We summarize here the recent progress in fluorescence-based bioassays for the detection and evaluation of food materials by focusing on fluorescent dyes used in bioassays and applications of these assays for food safety, quality and efficacy. Fluorescent dyes have been used in various bioassays, such as biosensing, cell assay, energy transfer-based assay, probing, protein/immunological assay and microarray/biochip assay. Among the arrays used in microarray/biochip assay, fluorescence-based microarrays/biochips, such as antibody/protein microarrays, bead/suspension arrays, capillary/sensor arrays, DNA microarrays/polymerase chain reaction (PCR)-based arrays, glycan/lectin arrays, immunoassay/enzyme-linked immunosorbent assay (ELISA)-based arrays, microfluidic chips and tissue arrays, have been developed and used for the assessment of allergy/poisoning/toxicity, contamination and efficacy/mechanism, and quality control/safety. DNA microarray assays have been used widely for food safety and quality as well as searches for active components. DNA microarray-based gene expression profiling may be useful for such purposes due to its advantages in the evaluation of pathway-based intracellular signaling in response to food materials. PMID:26473869

Transcript Profiling of Two Alfalfa Genotypes with Contrasting Cell Wall Composition in Stems Using a Cross-Species Platform: Optimizing Analysis by Masking Biased Probes

USDA-ARS?s Scientific Manuscript database

The stem cell walls of alfalfa [Medicago sativa (L.) ssp. sativa] genotype 252 have high cellulose and lignin concentrations, while stem cell walls of genotype 1283 have low cellulose and lignin concentrations. The GeneChip® Medicago Genome Array, developed for Medicago truncatula, is a suitable pla...

Expression Profile of DNA Damage Signaling Genes in Proton Exposed Mouse Brain

NASA Astrophysics Data System (ADS)

Ramesh, Govindarajan; Wu, Honglu

Exposure of living systems to radiation results in a wide assortment of lesions, the most signif-icant of is damage to genomic DNA which induce several cellular functions such as cell cycle arrest, repair, apoptosis etc. The radiation induced DNA damage investigation is one of the im-portant area in biology, but still the information available regarding the effects of proton is very limited. In this report, we investigated the differential gene expression pattern of DNA damage signaling genes particularly, damaged DNA binding, repair, cell cycle arrest, checkpoints and apoptosis using quantitative real-time RT-PCR array in proton exposed mouse brain tissues. The expression profiles showed significant changes in DNA damage related genes in 2Gy proton exposed mouse brain tissues as compared with control brain tissues. Furthermore, we also show that significantly increased levels of apoptotic related genes, caspase-3 and 8 activities in these cells, suggesting that in addition to differential expression of DNA damage genes, the alteration of apoptosis related genes may also contribute to the radiation induced DNA damage followed by programmed cell death. In summary, our findings suggest that proton exposed brain tissue undergo severe DNA damage which in turn destabilize the chromatin stability.

Gene-expression profiles of epithelial cells treated with EMD in vitro: analysis using complementary DNA arrays.

PubMed

Kapferer, I; Schmidt, S; Gstir, R; Durstberger, G; Huber, L A; Vietor, I

2011-02-01

During surgical periodontal treatment, EMD is topically applied in order to facilitate regeneration of the periodontal ligament, acellular cementum and alveolar bone. Suppresion of epithelial down-growth is essential for successful periodontal regeneration; however, the underlying mechanisms of how EMD influences epithelial wound healing are poorly understood. In the present study, the effects of EMD on gene-expression profiling in an epithelial cell line (HSC-2) model were investigated. Gene-expression modifications, determined using a comparative genome-wide expression-profiling strategy, were independently validated by quantitative real-time RT-PCR. Additionally, cell cycle, cell growth and in vitro wound-healing assays were conducted. A set of 43 EMD-regulated genes was defined, which may be responsible for the reduced epithelial down-growth upon EMD application. Gene ontology analysis revealed genes that could be attributed to pathways of locomotion, developmental processes and associated processes such as regulation of cell size and cell growth. Additionally, eight regulated genes have previously been reported to take part in the process of epithelial-to-mesenchymal transition. Several independent experimental assays revealed significant inhibition of cell migration, growth and cell cycle by EMD. The set of EMD-regulated genes identified in this study offers the opportunity to clarify mechanisms underlying the effects of EMD on epithelial cells. Reduced epithelial repopulation of the dental root upon periodontal surgery may be the consequence of reduced migration and cell growth, as well as epithelial-to-mesenchymal transition. © 2010 John Wiley & Sons A/S.

Copy number variations of genes involved in stress responses reflect the redox state and DNA damage in brewing yeasts.

PubMed

Adamczyk, Jagoda; Deregowska, Anna; Skoneczny, Marek; Skoneczna, Adrianna; Natkanska, Urszula; Kwiatkowska, Aleksandra; Rawska, Ewa; Potocki, Leszek; Kuna, Ewelina; Panek, Anita; Lewinska, Anna; Wnuk, Maciej

2016-09-01

The yeast strains of the Saccharomyces sensu stricto complex involved in beer production are a heterogeneous group whose genetic and genomic features are not adequately determined. Thus, the aim of the present study was to provide a genetic characterization of selected group of commercially available brewing yeasts both ale top-fermenting and lager bottom-fermenting strains. Molecular karyotyping revealed that the diversity of chromosome patterns and four strains with the most accented genetic variabilities were selected and subjected to genome-wide array-based comparative genomic hybridization (array-CGH) analysis. The differences in the gene copy number were found in five functional gene categories: (1) maltose metabolism and transport, (2) response to toxin, (3) siderophore transport, (4) cellular aldehyde metabolic process, and (5) L-iditol 2-dehydrogenase activity (p < 0.05). In the Saflager W-34/70 strain (Fermentis) with the most affected array-CGH profile, loss of aryl-alcohol dehydrogenase (AAD) gene dosage correlated with an imbalanced redox state, oxidative DNA damage and breaks, lower levels of nucleolar proteins Nop1 and Fob1, and diminished tolerance to fermentation-associated stress stimuli compared to other strains. We suggest that compromised stress response may not only promote oxidant-based changes in the nucleolus state that may affect fermentation performance but also provide novel directions for future strain improvement.

Divergent evolution of arrested development in the dauer stage of Caenorhabditis elegans and the infective stage of Heterodera glycines

PubMed Central

Elling, Axel A; Mitreva, Makedonka; Recknor, Justin; Gai, Xiaowu; Martin, John; Maier, Thomas R; McDermott, Jeffrey P; Hewezi, Tarek; McK Bird, David; Davis, Eric L; Hussey, Richard S; Nettleton, Dan; McCarter, James P; Baum, Thomas J

2007-01-01

Background The soybean cyst nematode Heterodera glycines is the most important parasite in soybean production worldwide. A comprehensive analysis of large-scale gene expression changes throughout the development of plant-parasitic nematodes has been lacking to date. Results We report an extensive genomic analysis of H. glycines, beginning with the generation of 20,100 expressed sequence tags (ESTs). In-depth analysis of these ESTs plus approximately 1,900 previously published sequences predicted 6,860 unique H. glycines genes and allowed a classification by function using InterProScan. Expression profiling of all 6,860 genes throughout the H. glycines life cycle was undertaken using the Affymetrix Soybean Genome Array GeneChip. Our data sets and results represent a comprehensive resource for molecular studies of H. glycines. Demonstrating the power of this resource, we were able to address whether arrested development in the Caenorhabditis elegans dauer larva and the H. glycines infective second-stage juvenile (J2) exhibits shared gene expression profiles. We determined that the gene expression profiles associated with the C. elegans dauer pathway are not uniformly conserved in H. glycines and that the expression profiles of genes for metabolic enzymes of C. elegans dauer larvae and H. glycines infective J2 are dissimilar. Conclusion Our results indicate that hallmark gene expression patterns and metabolism features are not shared in the developmentally arrested life stages of C. elegans and H. glycines, suggesting that developmental arrest in these two nematode species has undergone more divergent evolution than previously thought and pointing to the need for detailed genomic analyses of individual parasite species. PMID:17919324

The database of chromosome imbalance regions and genes resided in lung cancer from Asian and Caucasian identified by array-comparative genomic hybridization

PubMed Central

2012-01-01

Background Cancer-related genes show racial differences. Therefore, identification and characterization of DNA copy number alteration regions in different racial groups helps to dissect the mechanism of tumorigenesis. Methods Array-comparative genomic hybridization (array-CGH) was analyzed for DNA copy number profile in 40 Asian and 20 Caucasian lung cancer patients. Three methods including MetaCore analysis for disease and pathway correlations, concordance analysis between array-CGH database and the expression array database, and literature search for copy number variation genes were performed to select novel lung cancer candidate genes. Four candidate oncogenes were validated for DNA copy number and mRNA and protein expression by quantitative polymerase chain reaction (qPCR), chromogenic in situ hybridization (CISH), reverse transcriptase-qPCR (RT-qPCR), and immunohistochemistry (IHC) in more patients. Results We identified 20 chromosomal imbalance regions harboring 459 genes for Caucasian and 17 regions containing 476 genes for Asian lung cancer patients. Seven common chromosomal imbalance regions harboring 117 genes, included gain on 3p13-14, 6p22.1, 9q21.13, 13q14.1, and 17p13.3; and loss on 3p22.2-22.3 and 13q13.3 were found both in Asian and Caucasian patients. Gene validation for four genes including ARHGAP19 (10q24.1) functioning in Rho activity control, FRAT2 (10q24.1) involved in Wnt signaling, PAFAH1B1 (17p13.3) functioning in motility control, and ZNF322A (6p22.1) involved in MAPK signaling was performed using qPCR and RT-qPCR. Mean gene dosage and mRNA expression level of the four candidate genes in tumor tissues were significantly higher than the corresponding normal tissues (P<0.001~P=0.06). In addition, CISH analysis of patients indicated that copy number amplification indeed occurred for ARHGAP19 and ZNF322A genes in lung cancer patients. IHC analysis of paraffin blocks from Asian Caucasian patients demonstrated that the frequency of PAFAH1B1 protein overexpression was 68% in Asian and 70% in Caucasian. Conclusions Our study provides an invaluable database revealing common and differential imbalance regions at specific chromosomes among Asian and Caucasian lung cancer patients. Four validation methods confirmed our database, which would help in further studies on the mechanism of lung tumorigenesis. PMID:22691236

Feasibility of the AML profiler (Skyline™ Array) for patient risk stratification in a multicentre trial: a preliminary comparison with the conventional approach.

PubMed

Nomdedéu, Josep F; Puigdecanet, Eulalia; Bussaglia, Elena; Hernández, Juan José; Carricondo, Maite; Estivill, Camino; Martí-Tutusaus, Josep Maria; Tormo, Mar; Zamora, Lurdes; Serrano, Elena; Perea, Granada; de Llano, Maria Paz Queipo; García, Antoni; Sánchez-Ortega, Isabel; Ribera, Josep Maria; Nonell, Lara; Aventin, Anna; Solé, Francesc; Brunet, Maria Salut; Sierra, Jorge

2017-12-01

Deoxyribonucleic acid microarrays allow researchers to measure mRNA levels of thousands of genes in a single experiment and could be useful for diagnostic purposes in patients with acute myeloid leukaemia (AML). We assessed the feasibility of the AML profiler (Skyline™ Array) in genetic stratification of patients with de novo AML and compared the results with those obtained using the standard cytogenetic and molecular approach. Diagnostic bone marrow from 31 consecutive de novo AML cases was used to test MLL-PTD, FLT3-ITD and TKD, NPM1 and CEBPAdm mutations. Purified RNA was used to assess RUNX1-RUNX1T1, PML-RARα and CBFβ-MYH11 rearrangements. RNA remnants underwent gene expression profiling analysis using the AML profiler, which detects chromosomal aberrations: t(8;21), t(15;17), inv(16), mutations (CEBPAdm, ABD-NPM1) and BAALC and EVI1 expression. Thirty cases were successfully analysed with both methods. Five cases had FLT3-ITD. In one case, a t(8;21) was correctly detected by both methods. Four cases had inv(16); in one, the RNA quality was unsatisfactory and it was not hybridized, and in the other three, the AML profiler detected the genetic lesion - this being a rare type I translocation in one case. Two cases with acute promyelocytic leukaemia were diagnosed by both methods. Results for NPM1 mutations were concordant in all but two cases (2/11, non-ABD mutations). Analysis of costs and turnaround times showed that the AML profiler was no more expensive than the conventional molecular approach. These results suggest that the AML profiler could be useful in multicentre trials to rapidly identify patients with AML with a good prognosis. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

Integrated analysis of copy number alteration and RNA expression profiles of cancer using a high-resolution whole-genome oligonucleotide array.

PubMed

Jung, Seung-Hyun; Shin, Seung-Hun; Yim, Seon-Hee; Choi, Hye-Sun; Lee, Sug-Hyung; Chung, Yeun-Jun

2009-07-31

Recently, microarray-based comparative genomic hybridization (array-CGH) has emerged as a very efficient technology with higher resolution for the genome-wide identification of copy number alterations (CNA). Although CNAs are thought to affect gene expression, there is no platform currently available for the integrated CNA-expression analysis. To achieve high-resolution copy number analysis integrated with expression profiles, we established human 30k oligoarray-based genome-wide copy number analysis system and explored the applicability of this system for integrated genome and transcriptome analysis using MDA-MB-231 cell line. We compared the CNAs detected by the oligoarray with those detected by the 3k BAC array for validation. The oligoarray identified the single copy difference more accurately and sensitively than the BAC array. Seventeen CNAs detected by both platforms in MDA-MB-231 such as gains of 5p15.33-13.1, 8q11.22-8q21.13, 17p11.2, and losses of 1p32.3, 8p23.3-8p11.21, and 9p21 were consistently identified in previous studies on breast cancer. There were 122 other small CNAs (mean size 1.79 mb) that were detected by oligoarray only, not by BAC-array. We performed genomic qPCR targeting 7 CNA regions, detected by oligoarray only, and one non-CNA region to validate the oligoarray CNA detection. All qPCR results were consistent with the oligoarray-CGH results. When we explored the possibility of combined interpretation of both DNA copy number and RNA expression profiles, mean DNA copy number and RNA expression levels showed a significant correlation. In conclusion, this 30k oligoarray-CGH system can be a reasonable choice for analyzing whole genome CNAs and RNA expression profiles at a lower cost.

Gene expression profile of steroid-induced necrosis of femoral head of rats.

PubMed

Tong, Peijian; Wu, Chengliang; Jin, Hongting; Mao, Qiang; Yu, Nanze; Holz, Jonathan D; Shan, Letian; Liu, Hui; Xiao, Luwei

2011-10-01

The key to treating steroid-induced necrosis of femoral heads (SINFH) is early diagnosis. Dramatic improvements in diagnosis could be made if the pathogenesis of SINFH was more fully understood; however, the underlying mechanism of this disease is currently unknown. To explore the potential mechanism of SINFH, we performed gene array analysis on a rat model of the disease and compare the expression profile with that of normal rats. A quantitative RT-PCR and immunohistochemistry (IHC) assays were used to confirm the microarray results. Compared to the control group, 190 genes in the experimental group were differentially expressed, with 52 up-regulated and 138 down-regulated. Of these genes, 102 are known (deposited in GenBank), while 88 of them are unknown. The known genes can be divided into several families according to their biological functions, such as oxidative stress, apoptosis, signal transduction, angiogenesis, extracellular matrix, lipid metabolism, and transcription related genes. The results of quantitative RT-PCR and IHC were consistent with gene chip results. Our findings indicate that many genes involved in diverse signaling pathways were differentially expressed between SINFH rats and normal rats. Furthermore, our findings suggest that the development of SINFH is a complicated and dynamic process affected by multiple factors and signaling pathways and regulated by various genes.

The changes of gene expression profiling between segmental vitiligo, generalized vitiligo and healthy individual.

PubMed

Wang, Ping; Li, Yong; Nie, Huiqiong; Zhang, Xiaoyan; Shao, Qiongyan; Hou, Xiuli; Xu, Wen; Hong, Weisong; Xu, Aie

2016-10-01

Vitiligo is a common acquired depigmentation skin disease characterized by loss or dysfunction of melanocytes within the skin lesion, but its pathologenesis is far from lucid. The gene expression profiling of segmental vitiligo (SV) and generalized vitiligo (GV) need further investigation. To better understanding the common and distinct factors, especially in the view of gene expression profile, which were involved in the diseases development and maintenance of segmental vitiligo (SV) and generalized vitiligo (GV). Peripheral bloods were collected from SV, GV and healthy individual (HI), followed by leukocytes separation and total RNA extraction. The high-throughput whole genome expression microarrays were used to assay the gene expression profiles between HI, SV and GV. Bioinformatics tools were employed to annotated the biological function of differently expressed genes. Quantitative PCR assay was used to validate the gene expression of array. Compared to HI, 239 over-expressed genes and 175 down-expressed genes detected in SV, 688 over-expressed genes and 560 down-expressed genes were found in GV, following the criteria of log2 (fold change)≥0.585 and P value<0.05. In these differently expressed genes, 60 over-expressed genes and 60 down-expressed genes had similar tendency in SV and GV. Compared to SV, 223 genes were up regulated and 129 genes were down regulated in GV. In the SV with HI as control, the differently expressed genes were mainly involved in the adaptive immune response, cytokine-cytokine receptor interaction, chemokine signaling, focal adhesion and sphingolipid metabolism. The differently expressed genes between GV and HI were mainly involved in the innate immune, autophagy, apoptosis, melanocyte biology, ubiquitin mediated proteolysis and tyrosine metabolism, which was different from SV. While the differently expressed genes between SV and GV were mainly involved in the metabolism pathway of purine, pyrimidine, glycolysis and sphingolipid. Above results suggested that they not only shared part bio-process and signal pathway, but more important, they utilized different biological mechanism in their pathogenesis and maintenance. Our results provide a comprehensive view on the gene expression profiling change between SV and GV especially in the side of leukocytes, and may facilitate the future study on their molecular mechanism and theraputic targets. Copyright © 2016 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.

Adaptation of the Biolog Phenotype MicroArrayTM Technology to Profile the Obligate Anaerobe Geobacter metallireducens

DOE Office of Scientific and Technical Information (OSTI.GOV)

Joyner, Dominique; Fortney, Julian; Chakraborty, Romy

2010-05-17

The Biolog OmniLog? Phenotype MicroArray (PM) plate technology was successfully adapted to generate a select phenotypic profile of the strict anaerobe Geobacter metallireducens (G.m.). The profile generated for G.m. provides insight into the chemical sensitivity of the organism as well as some of its metabolic capabilities when grown with a basal medium containing acetate and Fe(III). The PM technology was developed for aerobic organisms. The reduction of a tetrazolium dye by the test organism represents metabolic activity on the array which is detected and measured by the OmniLog(R) system. We have previously adapted the technology for the anaerobic sulfate reducingmore » bacterium Desulfovibrio vulgaris. In this work, we have taken the technology a step further by adapting it for the iron reducing obligate anaerobe Geobacter metallireducens. In an osmotic stress microarray it was determined that the organism has higher sensitivity to impermeable solutes 3-6percent KCl and 2-5percent NaNO3 that result in osmotic stress by osmosis to the cell than to permeable non-ionic solutes represented by 5-20percent ethylene glycol and 2-3percent urea. The osmotic stress microarray also includes an array of osmoprotectants and precursor molecules that were screened to identify substrates that would provide osmotic protection to NaCl stress. None of the substrates tested conferred resistance to elevated concentrations of salt. Verification studies in which G.m. was grown in defined medium amended with 100mM NaCl (MIC) and the common osmoprotectants betaine, glycine and proline supported the PM findings. Further verification was done by analysis of transcriptomic profiles of G.m. grown under 100mM NaCl stress that revealed up-regulation of genes related to degradation rather than accumulation of the above-mentioned osmoprotectants. The phenotypic profile, supported by additional analysis indicates that the accumulation of these osmoprotectants as a response to salt stress does not occur in G.m. and response to stress must occur by other mechanisms. The Phenotype MicroArray technology can be reliably used as a rapid screening tool for characterization in anaerobic microbial ecology.« less

Prospective technical validation and assessment of intra-tumour heterogeneity of a low density array hypoxia gene profile in head and neck squamous cell carcinoma.

PubMed

Betts, Guy N J; Eustace, Amanda; Patiar, Shalini; Valentine, Helen R; Irlam, Joely; Ramachandran, Anassuya; Merve, Ashirwad; Homer, Jarrod J; Möller-Levet, Carla; Buffa, Francesca M; Hall, Gillian; Miller, Crispin J; Harris, Adrian L; West, Catharine M L

2013-01-01

Tumour hypoxia is associated with a poor prognosis in head and neck squamous cell carcinoma (HNSCC), however there is no accepted method for assessing hypoxia clinically. We aimed to conduct a technical validation of a hypoxia gene expression signature using the TaqMan Low Density Array (TLDA) platform to investigate if this approach reliably identified hypoxic tumours. Tumour samples (n=201) from 80 HNSCC patients were collected prospectively from two centres. Fifty-three patients received pimonidazole prior to surgery. TaqMan Low Density Array-Hypoxia Scores (TLDA-HS) were obtained by quantitative real-time PCR (qPCR) using a 25-gene signature and customised TLDA cards. Assay performance was assessed as coefficient of variation (CoV). The assay was sensitive with linear reaction efficiencies across a 4 log(10) range of inputted cDNA (0.001-10 ng/μl). Intra- (CoV=6.9%) and inter- (CoV=2.0%) assay reproducibility were excellent. Intra-tumour heterogeneity was lower for TLDA-HS (23.2%) than for pimonidazole (67.2%) or single gene measurements of CA9 (62.2%), VEGFA (45.0%) or HIG2 (39.4%). TLDA-HS in HNSCC cell lines increased with decreasing pO(2). TLDA-HS correlated with Affymetrix U133 Plus 2.0 microarray HS (p<0.01) and positive pimonidazole scores (p=0.005). Gene expression measurements of hypoxia using a 25-gene signature and TLDA cards are sensitive, reproducible and associated with lower intra-tumour heterogeneity than assaying individual genes or pimonidazole binding. The approach is suitable for further assessment of prognostic and predictive capability in clinical trial material. Copyright © 2012 Elsevier Ltd. All rights reserved.

Use of human antigen presenting cell gene array profiling to examine the effect of human T-cell leukemia virus type 1 Tax on primary human dendritic cells.

PubMed

Ahuja, Jaya; Kampani, Karan; Datta, Suman; Wigdahl, Brian; Flaig, Katherine E; Jain, Pooja

2006-02-01

Human T-cell leukemia virus type 1 (HTLV-1) is etiologically linked to adult T-cell leukemia and a progressive demyelinating disorder termed HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). One of the most striking features of the immune response in HAM/TSP centers on the expansion of HTLV-1-specific CD8(+) cytotoxic T lymphocyte (CTL) compartment in the peripheral blood and cerebrospinal fluid. More than 90% of the HTLV-1-specific CTLs are directed against the viral Tax (11-19) peptide implying that Tax is available for immune recognition by antigen presenting cells, such as dendritic cells (DCs). DCs obtained from HAM/TSP patients have been shown to be infected with HTLV-1 and exhibit rapid maturation. Therefore, we hypothesized that presentation of Tax peptides by activated DCs to naIve CD8(+) T cells may play an important role in the induction of a Tax-specific CTL response and neurologic dysfunction. In this study, a pathway-specific antigen presenting cell gene array was used to study transcriptional changes induced by exposure of monocyte-derived DCs to extracellular HTLV-1 Tax protein. Approximately 100 genes were differentially expressed including genes encoding toll-like receptors, cell surface receptors, proteins involved in antigen uptake and presentation and adhesion molecules. The differential regulation of chemokines and cytokines characteristic of functional DC activation was also observed by the gene array analyses. Furthermore, the expression pattern of signal transduction genes was also significantly altered. These results have suggested that Tax-mediated DC gene regulation might play a critical role in cellular activation and the mechanisms resulting in HTLV-1-induced disease.

The expression of native and cultured human retinal pigment epithelial cells grown in different culture conditions.

PubMed

Tian, J; Ishibashi, K; Honda, S; Boylan, S A; Hjelmeland, L M; Handa, J T

2005-11-01

To determine the transcriptional proximity of retinal pigment epithelium (RPE) cells grown under different culture conditions and native RPE. ARPE-19 cells were grown under five conditions in 10% CO(2): "subconfluent" in DMEM/F12+10% FBS, "confluent" in serum and serum withdrawn, and "differentiated" for 2.5 months in serum and serum withdrawn medium. Native RPE was laser microdissected. Total RNA was extracted, reverse transcribed, and radiolabelled probes were hybridised to an array containing 5,353 genes. Arrays were evaluated by hierarchical cluster analysis and significance analysis of microarrays. 78% of genes were expressed by native RPE while 45.3--47.7% were expressed by ARPE-19 cells, depending on culture condition. While the most abundant genes were expressed by native and cultured cells, significant differences in low abundance genes were seen. Hierarchical cluster analysis showed that confluent and differentiated, serum withdrawn cultures clustered closest to native RPE, and that serum segregated cultured cells from native RPE. The number of differentially expressed genes and their function, and profile of expressed and unexpressed genes, demonstrate differences between native and cultured cells. While ARPE-19 cells have significant value for studying RPE behaviour, investigators must be aware of how culture conditions can influence the mRNA phenotype of the cell.

The Long Noncoding RNA Landscape of the Mouse Eye.

PubMed

Chen, Weiwei; Yang, Shuai; Zhou, Zhonglou; Zhao, Xiaoting; Zhong, Jiayun; Reinach, Peter S; Yan, Dongsheng

2017-12-01

Long noncoding RNAs (lncRNAs) are important regulators of diverse biological functions. However, an extensive in-depth analysis of their expression profile and function in mammalian eyes is still lacking. Here we describe comprehensive landscapes of stage-dependent and tissue-specific lncRNA expression in the mouse eye. Affymetrix transcriptome array profiled lncRNA signatures from six different ocular tissue subsets (i.e., cornea, lens, retina, RPE, choroid, and sclera) in newborn and 8-week-old mice. Quantitative RT-PCR analysis validated array findings. Cis analyses and Gene Ontology (GO) annotation of protein-coding genes adjacent to signature lncRNA loci clarified potential lncRNA roles in maintaining tissue identity and regulating eye maturation during the aforementioned phase. In newborn and 8-week-old mice, we identified 47,332 protein-coding and noncoding gene transcripts. LncRNAs comprise 19,313 of these transcripts annotated in public data banks. During this maturation phase of these six different tissue subsets, more than 1000 lncRNAs expression levels underwent ≥2-fold changes. qRT-PCR analysis confirmed part of the gene microarray analysis results. K-means clustering identified 910 lncRNAs in the P0 groups and 686 lncRNAs in the postnatal 8-week-old groups, suggesting distinct tissue-specific lncRNA clusters. GO analysis of protein-coding genes proximal to lncRNA signatures resolved close correlations with their tissue-specific functional maturation between P0 and 8 weeks of age in the 6 tissue subsets. Characterizating maturational changes in lncRNA expression patterns as well as tissue-specific lncRNA signatures in six ocular tissues suggest important contributions made by lncRNA to the control of developmental processes in the mouse eye.

Identification of interactive gene networks: a novel approach in gene array profiling of myometrial events during guinea pig pregnancy.

PubMed

Mason, Clifford W; Swaan, Peter W; Weiner, Carl P

2006-06-01

The transition from myometrial quiescence to activation is poorly understood, and the analysis of array data is limited by the available data mining tools. We applied functional analysis and logical operations along regulatory gene networks to identify molecular processes and pathways underlying quiescence and activation. We analyzed some 18,400 transcripts and variants in guinea pig myometrium at stages corresponding to quiescence and activation, and compared them to the nonpregnant (control) counterpart using a functional mapping tool, MetaCore (GeneGo, St Joseph, MI) to identify novel gene networks composed of biological pathways during mid (MP) and late (LP) pregnancy. Genes altered during quiescence and or activation were identified following gene specific comparisons with myometrium from nonpregnant animals, and then linked to curated pathways and formulated networks. The MP and LP networks were subtracted from each other to identify unique genomic events during those periods. For example, changes 2-fold or greater in genes mediating protein biosynthesis, programmed cell death, microtubule polymerization, and microtubule based movement were noted during the transition to LP. We describe a novel approach combining microarrays and genetic data to identify networks associated with normal myometrial events. The resulting insights help identify potential biomarkers and permit future targeted investigations of these pathways or networks to confirm or refute their importance.

Transcription profiling provides insights into gene pathways involved in horn and scurs development in cattle

PubMed Central

2010-01-01

Background Two types of horns are evident in cattle - fixed horns attached to the skull and a variation called scurs, which refers to small loosely attached horns. Cattle lacking horns are referred to as polled. Although both the Poll and Scurs loci have been mapped to BTA1 and 19 respectively, the underlying genetic basis of these phenotypes is unknown, and so far, no candidate genes regulating these developmental processes have been described. This study is the first reported attempt at transcript profiling to identify genes and pathways contributing to horn and scurs development in Brahman cattle, relative to polled counterparts. Results Expression patterns in polled, horned and scurs tissues were obtained using the Agilent 44 k bovine array. The most notable feature when comparing transcriptional profiles of developing horn tissues against polled was the down regulation of genes coding for elements of the cadherin junction as well as those involved in epidermal development. We hypothesize this as a key event involved in keratinocyte migration and subsequent horn development. In the polled-scurs comparison, the most prevalent differentially expressed transcripts code for genes involved in extracellular matrix remodelling, which were up regulated in scurs tissues relative to polled. Conclusion For this first time we describe networks of genes involved in horn and scurs development. Interestingly, we did not observe differential expression in any of the genes present on the fine mapped region of BTA1 known to contain the Poll locus. PMID:20537189

Comparison of gene expression responses to hypoxia in viviparous (Xiphophorus) and oviparous (Oryzias) fishes using a medaka microarray.

PubMed

Boswell, Mikki G; Wells, Melissa C; Kirk, Lyndsey M; Ju, Zhenlin; Zhang, Ziping; Booth, Rachell E; Walter, Ronald B

2009-03-01

Gene expression profiling using DNA microarray technology is a useful tool for assessing gene transcript level responses after an organism is exposed to environmental stress. Herein, we detail results from studies using an 8 k medaka (Oryzias latipes) microarray to assess modulated gene expression patterns upon hypoxia exposure of the live-bearing aquaria fish, Xiphophorus maculatus. To assess the reproducibility and reliability of using the medaka array in cross-genus hybridization, a two-factor ANOVA analysis of gene expression was employed. The data show the tissue source of the RNA used for array hybridization contributed more to the observed response of modulated gene targets than did the species source of the RNA. In addition, hierarchical clustering via heat map analyses of groupings of tissues and species (Xiphophorus and medaka) suggests that hypoxia induced similar responses in the same tissues from these two diverse aquatic model organisms. Our Xiphophorus results indicate 206 brain, 37 liver, and 925 gill gene targets exhibit hypoxia induced expression changes. Analysis of the Xiphophorus data to determine those features exhibiting a significant (p<0.05)+/-3 fold change produced only two gene targets within brain tissue and 80 features within gill tissue. Of these 82 characterized features, 39 were identified via homology searching (cut-off E-value of 1 x 10(-5)) and placed into one or more biological process gene ontology groups. Among these 39 genes, metabolic energy changes and manipulation was the most affected biological pathway (13 genes).

Impact of Collection and Storage of Lung Tumor Tissue on Whole Genome Expression Profiling

PubMed Central

Freidin, Maxim B.; Bhudia, Neesa; Lim, Eric; Nicholson, Andrew G.; Cookson, William O.; Moffatt, Miriam F.

2012-01-01

Gene expression profiling could assist in revealing biomarkers of lung cancer prognosis and progression. The handling of biological samples may strongly influence global gene expression, a fact that has not been addressed in many studies. We sought to investigate the changes in gene expression that may occur as a result of sample processing time and conditions. Using Illumina Human WG-6 arrays, we quantified gene expression in lung carcinoma samples from six patients obtained at chest opening before and immediately after lung resection with storage in RNAlater [T1a(CO) and T1b(LR)], after receipt of the sample for histopathology, placed in RNAlater [T2a(HP)]; snap frozen [T2b(HP.SF)]; or snap frozen and stored for 1 week [T2c(HP.SFA)], as well as formalin-fixed, paraffin-embedded (FFPE) block samples. Sampling immediately after resection closely represented the tissue obtained in situ, with only 1% of genes differing more than twofold [T1a(CO) versus T1b(LR)]. Delaying tissue harvest for an average of 30 minutes from the operating theater had a significant impact on gene expression, with approximately 25% of genes differing between T1a(CO) and T2a(HP). Many genes previously identified as lung cancer biomarkers were altered during this period. Examination of FFPE specimens showed minimal correlation with fresh samples. This study shows that tissue collection immediately after lung resection with conservation in RNAlater is an optimal strategy for gene expression profiling. PMID:22240448

Gene expression profile of endoscopically active and inactive ulcerative colitis: preliminary data.

PubMed

Ţieranu, Cristian George; Dobre, Maria; Mănuc, Teodora Ecaterina; Milanesi, Elena; Pleşea, Iancu Emil; Popa, Caterina; Mănuc, Mircea; Ţieranu, Ioana; Preda, Carmen Monica; Diculescu, Mihai Mircea; Ionescu, Elena Mirela; Becheanu, Gabriel

2017-01-01

Multiple cytokines and chemokines related to immune response, apoptosis and inflammation have been identified as molecules implicated in ulcerative colitis (UC) pathogenesis. The aim of this study was to identify the differences at gene expression level of a panel of candidate genes in mucosa from patients with active UC (UCA), patients in remission (UCR), and normal controls. Eleven individuals were enrolled in the study: eight UC patients (four with active lesions, four with mucosal healing) and three controls without inflammatory bowel disease (IBD) seen on endoscopy. All the individuals underwent mucosal biopsy during colonoscopy. Gene expression profile was evaluated by polymerase chain reaction (PCR) array, investigating 84 genes implicated in apoptosis, inflammation, immune response, cellular adhesion, tissue remodeling and mucous secretion. Seventeen and three genes out of 84 were found significantly differentially expressed in UCA and UCR compared to controls, respectively. In particular, REG1A and CHI3L1 genes reported an up-regulation in UCA with a fold difference above 200. In UCR patients, the levels of CASP1, LYZ and ISG15 were different compared to controls. However, since a significant up-regulation of both CASP1 and LYZ was observed also in the UCA group, only ISG15 levels remained associated to the remission state. ISG15, that plays a key role in the innate immune response, seemed to be specifically associated to the UC remission state. These preliminary data represent a starting point for defining the gene profile of UC in different stages in Romanian population. Identification of genes implicated in UC pathogenesis could be useful to select new therapeutic targets.

GST-PRIME: an algorithm for genome-wide primer design.

PubMed

Leister, Dario; Varotto, Claudio

2007-01-01

The profiling of mRNA expression based on DNA arrays has become a powerful tool to study genome-wide transcription of genes in a number of organisms. GST-PRIME is a software package created to facilitate large-scale primer design for the amplification of probes to be immobilized on arrays for transcriptome analyses, even though it can be also applied in low-throughput approaches. GST-PRIME allows highly efficient, direct amplification of gene-sequence tags (GSTs) from genomic DNA (gDNA), starting from annotated genome or transcript sequences. GST-PRIME provides a customer-friendly platform for automatic primer design, and despite the relative simplicity of the algorithm, experimental tests in the model plant species Arabidopsis thaliana confirmed the reliability of the software. This chapter describes the algorithm used for primer design, its input and output files, and the installation of the standalone package and its use.

Leaving out control groups: an internal contrast analysis of gene expression profiles in atrial fibrillation patients--a systems biology approach to clinical categorization.

PubMed

Vanhoutte, Kurt; de Asmundis, Carlo; Francesconi, Anna; Figysl, Jurgen; Steurs, Griet; Boussy, Tim; Roos, Markus; Mueller, Andreas; Massimo, Lucio; Paparella, Gaetano; Van Caelenberg, Kristien; Chierchia, Gian Battista; Sarkozy, Andrea; Terradellas, Pedro Brugada Y; Zizi, Martin

2009-01-01

Atrial fibrillation (AF) is a frequent chronic dysrythmia with an incidence that increases with age (>40). Because of its medical and socio-economic impacts it is expected to become an increasing burden on most health care systems. AF is a multi-factorial disease for which the identification of subtypes is warranted. Novel approaches based on the broad concepts of systems biology may overcome the blurred notion of normal and pathological phenotype, which is inherent to high throughput molecular arrays analysis. Here we apply an internal contrast algorithm on AF patient data with an analytical focus on potential entry pathways into the disease. We used a RMA (Robust Multichip Average) normalized Affymetrix micro-array data set from 10 AF patients (geo_accession #GSE2240). Four series of probes were selected based on physiopathogenic links with AF entryways: apoptosis (remodeling), MAP kinase (cell remodeling), OXPHOS (ability to sustain hemodynamic workload) and glycolysis (ischemia). Annotated probe lists were polled with Bioconductor packages in R (version 2.7.1). Genetic profile contrasts were analysed with hierarchical clustering and principal component analysis. The analysis revealed distinct patient groups for all probe sets. A substantial part (54% till 67%) of the variance is explained in the first 2 principal components. Genes in PC1/2 with high discriminatory value were selected and analyzed in detail. We aim for reliable molecular stratification of AF. We show that stratification is possible based on physiologically relevant gene sets. Genes with high contrast value are likely to give pathophysiological insight into permanent AF subtypes.

Organogenic nodule development in hop (Humulus lupulus L.): Transcript and metabolic responses

PubMed Central

Fortes, Ana M; Santos, Filipa; Choi, Young H; Silva, Marta S; Figueiredo, Andreia; Sousa, Lisete; Pessoa, Fernando; Santos, Bartolomeu A; Sebastiana, Mónica; Palme, Klaus; Malhó, Rui; Verpoorte, Rob; Pais, Maria S

2008-01-01

Background Hop (Humulus lupulus L.) is an economically important plant forming organogenic nodules which can be used for genetic transformation and micropropagation. We are interested in the mechanisms underlying reprogramming of cells through stress and hormone treatments. Results An integrated molecular and metabolomic approach was used to investigate global gene expression and metabolic responses during development of hop's organogenic nodules. Transcript profiling using a 3,324-cDNA clone array revealed differential regulation of 133 unigenes, classified into 11 functional categories. Several pathways seem to be determinant in organogenic nodule formation, namely defense and stress response, sugar and lipid metabolism, synthesis of secondary metabolites and hormone signaling. Metabolic profiling using 1H NMR spectroscopy associated to two-dimensional techniques showed the importance of metabolites related to oxidative stress response, lipid and sugar metabolism and secondary metabolism in organogenic nodule formation. Conclusion The expression profile of genes pivotal for energy metabolism, together with metabolites profile, suggested that these morphogenic structures gain energy through a heterotrophic, transport-dependent and sugar-degrading anaerobic metabolism. Polyamines and auxins are likely to be involved in the regulation of expression of many genes related to organogenic nodule formation. These results represent substantial progress toward a better understanding of this complex developmental program and reveal novel information regarding morphogenesis in plants. PMID:18823540

Expression Profiles of TGF-β and TLR Pathways in Porphyromonas gingivalis and Prevotella intermedia Challenged Osteoblasts.

PubMed

Aydin, Kubra; Ekinci, Fatma Yesim; Korachi, May

2015-04-01

The presence of certain oral pathogens at implant sites can hinder the osseointegration process. However, it is unclear how and by what microorganisms it happens. This study investigated whether the presence of oral pathogens of Porphyromonas gingivalis and Prevotella intermedia individually, play a role in the failure of bone formation by determining the expression profiles of Transforming Growth Factor Beta (TGF-β/Bone Morphogenic Protein (BMP) and Toll-Like Receptor (TLR) pathways in challenged osteoblasts. Cell viability of P. gingivalis and P. intermedia challenged osteoblasts were determined by WST assay. Changes in osteoblast morphology and inhibition of mineralization were observed by Scanning Electron Microscopy (SEM) and Von Kossa staining, respectively. Expression of TGF-β and TLR pathway genes on challenged cells were identified by RT profiler array. Both P. gingivalis and P. intermedia challenges resulted in reduced viability and mineralization of osteoblasts. Viability was reduced to 56.8% (P. gingivalis) and 52.75% (P. intermedia) at 1000 multiplicity. Amongst 48 genes examined, expressions of BMPER, SMAD1, IL8 and NFRKB were found to be highly upregulated by both bacterial challenges (Fold Change > 4). P. gingivalis and P. intermedia could play a role in implant failure by changing the expression profiles of genes related to bone formation and resorption.

Gene expression patterns during somatic embryo development and germination in maize Hi II callus cultures.

PubMed

Che, Ping; Love, Tanzy M; Frame, Bronwyn R; Wang, Kan; Carriquiry, Alicia L; Howell, Stephen H

2006-09-01

Gene expression patterns were profiled during somatic embryogenesis in a regeneration-proficient maize hybrid line, Hi II, in an effort to identify genes that might be used as developmental markers or targets to optimize regeneration steps for recovering maize plants from tissue culture. Gene expression profiles were generated from embryogenic calli induced to undergo embryo maturation and germination. Over 1,000 genes in the 12,060 element arrays showed significant time variation during somatic embryo development. A substantial number of genes were downregulated during embryo maturation, largely histone and ribosomal protein genes, which may result from a slowdown in cell proliferation and growth during embryo maturation. The expression of these genes dramatically recovered at germination. Other genes up-regulated during embryo maturation included genes encoding hydrolytic enzymes (nucleases, glucosidases and proteases) and a few storage genes (an alpha-zein and caleosin), which are good candidates for developmental marker genes. Germination is accompanied by the up-regulation of a number of stress response and membrane transporter genes, and, as expected, greening is associated with the up-regulation of many genes encoding photosynthetic and chloroplast components. Thus, some, but not all genes typically associated with zygotic embryogenesis are significantly up or down-regulated during somatic embryogenesis in Hi II maize line regeneration. Although many genes varied in expression throughout somatic embryo development in this study, no statistically significant gene expression changes were detected between total embryogenic callus and callus enriched for transition stage somatic embryos.

Expression profiles of differentially regulated genes during the early stages of apple flower infection with Erwinia amylovora

PubMed Central

Sarowar, Sujon; Zhao, Youfu; Soria-Guerra, Ruth Elena; Ali, Shahjahan; Zheng, Danman; Wang, Dongping; Korban, Schuyler S.

2011-01-01

To identify genes involved in the response to the fire blight pathogen Erwinia amylovora in apple (Malus×domestica), expression profiles were investigated using an apple oligo (70-mer) array representing 40, 000 genes. Blossoms of a fire blight-susceptible apple cultivar Gala were collected from trees growing in the orchard, placed on a tray in the laboratory, and spray-inoculated with a suspension of E. amylovora at a concentration of 108 cfu ml−1. Uninoculated detached flowers served as controls at each time point. Expression profiles were captured at three different time points post-inoculation at 2, 8, and 24 h, together with those at 0 h (uninoculated). A total of about 3500 genes were found to be significantly modulated in response to at least one of the three time points. Among those, a total of 770, 855, and 1002 genes were up-regulated, by 2-fold, at 2, 8, and 24 h following inoculation, respectively; while, 748, 1024, and 1455 genes were down-regulated, by 2-fold, at 2, 8, and 24 h following inoculation, respectively. Over the three time points post-inoculation, 365 genes were commonly up-regulated and 374 genes were commonly down-regulated. Both sets of genes were classified based on their functional categories. The majority of up-regulated genes were involved in metabolism, signal transduction, signalling, transport, and stress response. A number of transcripts encoding proteins/enzymes known to be up-regulated under particular biotic and abiotic stress were also up-regulated following E. amylovora treatment. Those up- or down-regulated genes encode transcription factors, signaling components, defense-related, transporter, and metabolism, all of which have been associated with disease responses in Arabidopsis and rice, suggesting similar response pathways are involved in apple blossoms. PMID:21725032

Effects of strenuous exercise on Th1/Th2 gene expression from human peripheral blood mononuclear cells of marathon participants.

PubMed

Xiang, Lianbin; Rehm, Kristina E; Marshall, Gailen D

2014-08-01

Physical stressors, such as strenuous exercise, can have numerous effects on the human body including the immune system. The aim of this study was to evaluate the gene expression profile of Th1/Th2 cytokines and related transcription factor genes in order to investigate possible immune imbalances before and after a marathon. Blood samples were collected from 16 normal volunteers 24-48 h before and one week after completing a marathon race. Gene expression of Th1 and Th2 related cytokines from human peripheral blood mononuclear cells (PBMC) was analyzed using Human Th1-Th2-Th3 RT(2) Profiler PCR Array and qRT-PCR that measured the transcript levels of 84 genes related to T cell activation. We found that PBMC express a characteristic Th2-like gene profile one week post-marathon compared to pre-marathon. The majority of genes up-regulated one week post-marathon such as IL-4, GATA3, and CCR4 were Th2 associated. For Th1-related genes, CXCR3 and IRF1 were up-regulated one week post-marathon. There was a trend of down-regulation of two Th1 related genes, T-bet and STAT1. Th3-related gene expression patterns did not change in the study. The ratios of both IFN-γ/IL-4 and T-bet/GATA3 gene expressions were significantly lower one week after marathon. These findings suggest that a Th1/Th2 immune imbalance persisted at least 1 week after completion of a marathon which offers a mechanistic rationale for the increased risk of upper respiratory tract infections often reported after strenuous exercise. Copyright © 2014 Elsevier Ltd. All rights reserved.

Gene expression pattern recognition algorithm inferences to classify samples exposed to chemical agents

NASA Astrophysics Data System (ADS)

Bushel, Pierre R.; Bennett, Lee; Hamadeh, Hisham; Green, James; Ableson, Alan; Misener, Steve; Paules, Richard; Afshari, Cynthia

2002-06-01

We present an analysis of pattern recognition procedures used to predict the classes of samples exposed to pharmacologic agents by comparing gene expression patterns from samples treated with two classes of compounds. Rat liver mRNA samples following exposure for 24 hours with phenobarbital or peroxisome proliferators were analyzed using a 1700 rat cDNA microarray platform. Sets of genes that were consistently differentially expressed in the rat liver samples following treatment were stored in the MicroArray Project System (MAPS) database. MAPS identified 238 genes in common that possessed a low probability (P < 0.01) of being randomly detected as differentially expressed at the 95% confidence level. Hierarchical cluster analysis on the 238 genes clustered specific gene expression profiles that separated samples based on exposure to a particular class of compound.

Transcript profiling of Wilms tumors reveals connections to kidney morphogenesis and expression patterns associated with anaplasia.

PubMed

Li, Wenliang; Kessler, Patricia; Williams, Bryan R G

2005-01-13

Anaplasia (unfavorable histology) is associated with therapy resistance and poor prognosis of Wilms tumor, but the molecular basis for this phenotype is unclear. Here, we used a cDNA array with 9240 clones relevant to cancer biology and/or kidney development to examine the expression profiles of 54 Wilms tumors, five normal kidneys and fetal kidney. By linking genes differentially expressed between fetal kidney and Wilms tumors to kidney morphogenesis, we found that genes expressed at a higher level in Wilms tumors tend to be expressed more in uninduced metanephrogenic mesenchyme or blastema than in their differentiated structures. Conversely, genes expressed at a lower level in Wilms tumors tend to be expressed less in uninduced metanephrogenic mesenchyme or blastema. We also identified 97 clones representing 76 Unigenes or unclustered ESTs that clearly separate anaplastic Wilms tumors from tumors with favorable histology. Genes in this set provide insight into the nature of the abnormal nuclear morphology of anaplastic tumors and may facilitate identification of molecular targets to improve their responsiveness to treatment.

Functional Genomics Using the Saccharomyces cerevisiae Yeast Deletion Collections.

PubMed

Nislow, Corey; Wong, Lai Hong; Lee, Amy Huei-Yi; Giaever, Guri

2016-09-01

Constructed by a consortium of 16 laboratories, the Saccharomyces genome-wide deletion collections have, for the past decade, provided a powerful, rapid, and inexpensive approach for functional profiling of the yeast genome. Loss-of-function deletion mutants were systematically created using a polymerase chain reaction (PCR)-based gene deletion strategy to generate a start-to-stop codon replacement of each open reading frame by homologous recombination. Each strain carries two molecular barcodes that serve as unique strain identifiers, enabling their growth to be analyzed in parallel and the fitness contribution of each gene to be quantitatively assessed by hybridization to high-density oligonucleotide arrays or through the use of next-generation sequencing technologies. Functional profiling of the deletion collections, using either strain-by-strain or parallel assays, provides an unbiased approach to systematically survey the yeast genome. The Saccharomyces yeast deletion collections have proved immensely powerful in contributing to the understanding of gene function, including functional relationships between genes and genetic pathways in response to diverse genetic and environmental perturbations. © 2016 Cold Spring Harbor Laboratory Press.

Heterologous oligonucleotide microarrays for transcriptomics in a non-model species; a proof-of-concept study of drought stress in Musa

PubMed Central

Davey, Mark W; Graham, Neil S; Vanholme, Bartel; Swennen, Rony; May, Sean T; Keulemans, Johan

2009-01-01

Background 'Systems-wide' approaches such as microarray RNA-profiling are ideally suited to the study of the complex overlapping responses of plants to biotic and abiotic stresses. However, commercial microarrays are only available for a limited number of plant species and development costs are so substantial as to be prohibitive for most research groups. Here we evaluate the use of cross-hybridisation to Affymetrix oligonucleotide GeneChip® microarrays to profile the response of the banana (Musa spp.) leaf transcriptome to drought stress using a genomic DNA (gDNA)-based probe-selection strategy to improve the efficiency of detection of differentially expressed Musa transcripts. Results Following cross-hybridisation of Musa gDNA to the Rice GeneChip® Genome Array, ~33,700 gene-specific probe-sets had a sufficiently high degree of homology to be retained for transcriptomic analyses. In a proof-of-concept approach, pooled RNA representing a single biological replicate of control and drought stressed leaves of the Musa cultivar 'Cachaco' were hybridised to the Affymetrix Rice Genome Array. A total of 2,910 Musa gene homologues with a >2-fold difference in expression levels were subsequently identified. These drought-responsive transcripts included many functional classes associated with plant biotic and abiotic stress responses, as well as a range of regulatory genes known to be involved in coordinating abiotic stress responses. This latter group included members of the ERF, DREB, MYB, bZIP and bHLH transcription factor families. Fifty-two of these drought-sensitive Musa transcripts were homologous to genes underlying QTLs for drought and cold tolerance in rice, including in 2 instances QTLs associated with a single underlying gene. The list of drought-responsive transcripts also included genes identified in publicly-available comparative transcriptomics experiments. Conclusion Our results demonstrate that despite the general paucity of nucleotide sequence data in Musa and only distant phylogenetic relations to rice, gDNA probe-based cross-hybridisation to the Rice GeneChip® is a highly promising strategy to study complex biological responses and illustrates the potential of such strategies for gene discovery in non-model species. PMID:19758430

Methylation profiling identifies 2 groups of gliomas according to their tumorigenesis.

PubMed

Laffaire, Julien; Everhard, Sibille; Idbaih, Ahmed; Crinière, Emmanuelle; Marie, Yannick; de Reyniès, Aurelien; Schiappa, Renaud; Mokhtari, Karima; Hoang-Xuan, Khê; Sanson, Marc; Delattre, Jean-Yves; Thillet, Joëlle; Ducray, François

2011-01-01

Extensive genomic and gene expression studies have been performed in gliomas, but the epigenetic alterations that characterize different subtypes of gliomas remain largely unknown. Here, we analyzed the methylation patterns of 807 genes (1536 CpGs) in a series of 33 low-grade gliomas (LGGs), 36 glioblastomas (GBMs), 8 paired initial and recurrent gliomas, and 9 controls. This analysis was performed with Illumina's Golden Gate Bead methylation arrays and was correlated with clinical, histological, genomic, gene expression, and genotyping data, including IDH1 mutations. Unsupervised hierarchical clustering resulted in 2 groups of gliomas: a group corresponding to de novo GBMs and a group consisting of LGGs, recurrent anaplastic gliomas, and secondary GBMs. When compared with de novo GBMs and controls, this latter group was characterized by a very high frequency of IDH1 mutations and by a hypermethylated profile similar to the recently described glioma CpG island methylator phenotype. MGMT methylation was more frequent in this group. Among the LGG cluster, 1p19q codeleted LGG displayed a distinct methylation profile. A study of paired initial and recurrent gliomas demonstrated that methylation profiles were remarkably stable across glioma evolution, even during anaplastic transformation, suggesting that epigenetic alterations occur early during gliomagenesis. Using the Cancer Genome Atlas data set, we demonstrated that GBM samples that had an LGG-like hypermethylated profile had a high rate of IDH1 mutations and a better outcome. Finally, we identified several hypermethylated and downregulated genes that may be associated with LGG and GBM oncogenesis, LGG oncogenesis, 1p19q codeleted LGG oncogenesis, and GBM oncogenesis.

Methylation profiling identifies 2 groups of gliomas according to their tumorigenesis

PubMed Central

Laffaire, Julien; Everhard, Sibille; Idbaih, Ahmed; Crinière, Emmanuelle; Marie, Yannick; de Reyniès, Aurelien; Schiappa, Renaud; Mokhtari, Karima; Hoang-Xuan, Khê; Sanson, Marc; Delattre, Jean-Yves; Thillet, Joëlle; Ducray, François

2011-01-01

Extensive genomic and gene expression studies have been performed in gliomas, but the epigenetic alterations that characterize different subtypes of gliomas remain largely unknown. Here, we analyzed the methylation patterns of 807 genes (1536 CpGs) in a series of 33 low-grade gliomas (LGGs), 36 glioblastomas (GBMs), 8 paired initial and recurrent gliomas, and 9 controls. This analysis was performed with Illumina's Golden Gate Bead methylation arrays and was correlated with clinical, histological, genomic, gene expression, and genotyping data, including IDH1 mutations. Unsupervised hierarchical clustering resulted in 2 groups of gliomas: a group corresponding to de novo GBMs and a group consisting of LGGs, recurrent anaplastic gliomas, and secondary GBMs. When compared with de novo GBMs and controls, this latter group was characterized by a very high frequency of IDH1 mutations and by a hypermethylated profile similar to the recently described glioma CpG island methylator phenotype. MGMT methylation was more frequent in this group. Among the LGG cluster, 1p19q codeleted LGG displayed a distinct methylation profile. A study of paired initial and recurrent gliomas demonstrated that methylation profiles were remarkably stable across glioma evolution, even during anaplastic transformation, suggesting that epigenetic alterations occur early during gliomagenesis. Using the Cancer Genome Atlas data set, we demonstrated that GBM samples that had an LGG-like hypermethylated profile had a high rate of IDH1 mutations and a better outcome. Finally, we identified several hypermethylated and downregulated genes that may be associated with LGG and GBM oncogenesis, LGG oncogenesis, 1p19q codeleted LGG oncogenesis, and GBM oncogenesis. PMID:20926426

Aneuploidy-dependent massive deregulation of the cellular transcriptome and apparent divergence of the Wnt/beta-catenin signaling pathway in human rectal carcinomas.

PubMed

Grade, Marian; Ghadimi, B Michael; Varma, Sudhir; Simon, Richard; Wangsa, Danny; Barenboim-Stapleton, Linda; Liersch, Torsten; Becker, Heinz; Ried, Thomas; Difilippantonio, Michael J

2006-01-01

To identify genetic alterations underlying rectal carcinogenesis, we used global gene expression profiling of a series of 17 locally advanced rectal adenocarcinomas and 20 normal rectal mucosa biopsies on oligonucleotide arrays. A total of 351 genes were differentially expressed (P < 1.0e-7) between normal rectal mucosa and rectal carcinomas, 77 genes had a >5-fold difference, and 85 genes always had at least a 2-fold change in all of the matched samples. Twelve genes satisfied all three of these criteria. Altered expression of genes such as PTGS2 (COX-2), WNT1, TGFB1, VEGF, and MYC was confirmed, whereas our data for other genes, like PPARD and LEF1, were inconsistent with previous reports. In addition, we found deregulated expression of many genes whose involvement in rectal carcinogenesis has not been reported. By mapping the genomic imbalances in the tumors using comparative genomic hybridization, we could show that DNA copy number gains of recurrently aneuploid chromosome arms 7p, 8q, 13q, 18q, 20p, and 20q correlated significantly with their average chromosome arm expression profile. Taken together, our results show that both the high-level, significant transcriptional deregulation of specific genes and general modification of the average transcriptional activity of genes residing on aneuploid chromosomes coexist in rectal adenocarcinomas.

Aneuploidy-Dependent Massive Deregulation of the Cellular Transcriptome and Apparent Divergence of the Wnt/β-catenin Signaling Pathway in Human Rectal Carcinomas

PubMed Central

Grade, Marian; Ghadimi, B. Michael; Varma, Sudhir; Simon, Richard; Wangsa, Danny; Barenboim-Stapleton, Linda; Liersch, Torsten; Becker, Heinz; Ried, Thomas; Difilippantonio, Michael J.

2016-01-01

To identify genetic alterations underlying rectal carcinogenesis, we used global gene expression profiling of a series of 17 locally advanced rectal adenocarcinomas and 20 normal rectal mucosa biopsies on oligonucleotide arrays. A total of 351 genes were differentially expressed (P < 1.0e–7) between normal rectal mucosa and rectal carcinomas, 77 genes had a >5-fold difference, and 85 genes always had at least a 2-fold change in all of the matched samples. Twelve genes satisfied all three of these criteria. Altered expression of genes such as PTGS2 (COX-2), WNT1, TGFB1, VEGF, and MYC was confirmed, whereas our data for other genes, like PPARD and LEF1, were inconsistent with previous reports. In addition, we found deregulated expression of many genes whose involvement in rectal carcinogenesis has not been reported. By mapping the genomic imbalances in the tumors using comparative genomic hybridization, we could show that DNA copy number gains of recurrently aneuploid chromosome arms 7p, 8q, 13q, 18q, 20p, and 20q correlated significantly with their average chromosome arm expression profile. Taken together, our results show that both the high-level, significant transcriptional deregulation of specific genes and general modification of the average transcriptional activity of genes residing on aneuploid chromosomes coexist in rectal adenocarcinomas. PMID:16397240

Tissue-Specific Transcriptomic Profiling of Sorghum propinquum using a Rice Genome Array

PubMed Central

Zhang, Ting; Zhao, Xiuqin; Huang, Liyu; Liu, Xiaoyue; Zong, Ying; Zhu, Linghua; Yang, Daichang; Fu, Binying

2013-01-01

Sorghum (Sorghum bicolor) is one of the world's most important cereal crops. S. propinquum is a perennial wild relative of S. bicolor with well-developed rhizomes. Functional genomics analysis of S. propinquum, especially with respect to molecular mechanisms related to rhizome growth and development, can contribute to the development of more sustainable grain, forage, and bioenergy cropping systems. In this study, we used a whole rice genome oligonucleotide microarray to obtain tissue-specific gene expression profiles of S. propinquum with special emphasis on rhizome development. A total of 548 tissue-enriched genes were detected, including 31 and 114 unique genes that were expressed predominantly in the rhizome tips (RT) and internodes (RI), respectively. Further GO analysis indicated that the functions of these tissue-enriched genes corresponded to their characteristic biological processes. A few distinct cis-elements, including ABA-responsive RY repeat CATGCA, sugar-repressive TTATCC, and GA-responsive TAACAA, were found to be prevalent in RT-enriched genes, implying an important role in rhizome growth and development. Comprehensive comparative analysis of these rhizome-enriched genes and rhizome-specific genes previously identified in Oryza longistaminata and S. propinquum indicated that phytohormones, including ABA, GA, and SA, are key regulators of gene expression during rhizome development. Co-localization of rhizome-enriched genes with rhizome-related QTLs in rice and sorghum generated functional candidates for future cloning of genes associated with rhizome growth and development. PMID:23536906

Clustered, regularly interspaced short palindromic repeat (CRISPR) diversity and virulence factor distribution in avian Escherichia coli.

PubMed

Fu, Qiang; Su, Zhixin; Cheng, Yuqiang; Wang, Zhaofei; Li, Shiyu; Wang, Heng'an; Sun, Jianhe; Yan, Yaxian

In order to investigate the diverse characteristics of clustered, regularly interspaced short palindromic repeat (CRISPR) arrays and the distribution of virulence factor genes in avian Escherichia coli, 80 E. coli isolates obtained from chickens with avian pathogenic E. coli (APEC) or avian fecal commensal E. coli (AFEC) were identified. Using the multiplex polymerase chain reaction (PCR), five genes were subjected to phylogenetic typing and examined for CRISPR arrays to study genetic relatedness among the strains. The strains were further analyzed for CRISPR loci and virulence factor genes to determine a possible association between their CRISPR elements and their potential virulence. The strains were divided into five phylogenetic groups: A, B1, B2, D and E. It was confirmed that two types of CRISPR arrays, CRISPR1 and CRISPR2, which contain up to 246 distinct spacers, were amplified in most of the strains. Further classification of the isolates was achieved by sorting them into nine CRISPR clusters based on their spacer profiles, which indicates a candidate typing method for E. coli. Several significant differences in invasion-associated gene distribution were found between the APEC isolates and the AFEC isolates. Our results identified the distribution of 11 virulence genes and CRISPR diversity in 80 strains. It was demonstrated that, with the exception of iucD and aslA, there was no sharp demarcation in the gene distribution between the pathogenic (APEC) and commensal (AFEC) strains, while the total number of indicated CRISPR spacers may have a positive correlation with the potential pathogenicity of the E. coli isolates. Copyright © 2016. Published by Elsevier Masson SAS.

TARGET Research Goals

Cancer.gov

TARGET researchers use various sequencing and array-based methods to examine the genomes, transcriptomes, and for some diseases epigenomes of select childhood cancers. This “multi-omic” approach generates a comprehensive profile of molecular alterations for each cancer type. Alterations are changes in DNA or RNA, such as rearrangements in chromosome structure or variations in gene expression, respectively. Through computational analyses and assays to validate biological function, TARGET researchers predict which alterations disrupt the function of a gene or pathway and promote cancer growth, progression, and/or survival. Researchers identify candidate therapeutic targets and/or prognostic markers from the cancer-associated alterations.

Genome wide profiling in oral squamous cell carcinoma identifies a four genetic marker signature of prognostic significance

PubMed Central

Vincent-Chong, Vui King; Salahshourifar, Iman; Woo, Kar Mun; Anwar, Arif; Razali, Rozaimi; Gudimella, Ranganath; Rahman, Zainal Ariff Abdul; Ismail, Siti Mazlipah; Kallarakkal, Thomas George; Ramanathan, Anand; Wan Mustafa, Wan Mahadzir; Abraham, Mannil Thomas; Tay, Keng Kiong; Zain, Rosnah Binti

2017-01-01

Background Cancers of the oral cavity are primarily oral squamous cell carcinomas (OSCCs). Many of the OSCCs present at late stages with an exceptionally poor prognosis. A probable limitation in management of patients with OSCC lies in the insufficient knowledge pertaining to the linkage between copy number alterations in OSCC and oral tumourigenesis thereby resulting in an inability to deliver targeted therapy. Objectives The current study aimed to identify copy number alterations (CNAs) in OSCC using array comparative genomic hybridization (array CGH) and to correlate the CNAs with clinico-pathologic parameters and clinical outcomes. Materials and methods Using array CGH, genome-wide profiling was performed on 75 OSCCs. Selected genes that were harboured in the frequently amplified and deleted regions were validated using quantitative polymerase chain reaction (qPCR). Thereafter, pathway and network functional analysis were carried out using Ingenuity Pathway Analysis (IPA) software. Results Multiple chromosomal regions including 3q, 5p, 7p, 8q, 9p, 10p, 11q were frequently amplified, while 3p and 8p chromosomal regions were frequently deleted. These findings were in confirmation with our previous study using ultra-dense array CGH. In addition, amplification of 8q, 11q, 7p and 9p and deletion of 8p chromosomal regions showed a significant correlation with clinico-pathologic parameters such as the size of the tumour, metastatic lymph nodes and pathological staging. Co-amplification of 7p, 8q, 9p and 11q regions that harbored amplified genes namely CCND1, EGFR, TPM2 and LRP12 respectively, when combined, continues to be an independent prognostic factor in OSCC. Conclusion Amplification of 3q, 5p, 7p, 8q, 9p, 10p, 11q and deletion of 3p and 8p chromosomal regions were recurrent among OSCC patients. Co-alteration of 7p, 8q, 9p and 11q was found to be associated with clinico-pathologic parameters and poor survival. These regions contain genes that play critical roles in tumourigenesis pathways. PMID:28384287

Expression Profiling of Transcriptome and Its Associated Disease Risk in Yang Deficiency Constitution of Healthy Subjects

PubMed Central

Yu, Ruoxi; Yang, Yin; Han, Yuanyuan; Hou, Pengwei; Li, Yingshuai; Li, Siqi

2016-01-01

Objectives. Differences among healthy subjects and associated disease risks are of substantial interest in clinical medicine. According to the theory of “constitution-disease correlation” in traditional Chinese medicine, we try to find out if there is any connection between intolerance of cold in Yang deficiency constitution and molecular evidence and if there is any gene expression basis in specific disorders. Methods. Peripheral blood mononuclear cells were collected from Chinese Han individuals with Yang deficiency constitution (n = 20) and balanced constitution (n = 8) (aged 18–28) and global gene expression profiles were determined between them using the Affymetrix HG-U133 Plus 2.0 array. Results. The results showed that when the fold change was ≥1.2 and q ≤ 0.05, 909 genes were upregulated in the Yang deficiency constitution, while 1189 genes were downregulated. According to our research differential genes found in Yang deficiency constitution were usually related to lower immunity, metabolic disorders, and cancer tendency. Conclusion. Gene expression disturbance exists in Yang deficiency constitution, which corresponds to the concept of constitution and gene classification. It also suggests people with Yang deficiency constitution are susceptible to autoimmune diseases, enteritis, arthritis, metabolism disorders, and cancer, which provides molecular evidence for the theory of “constitution-disease correlation.” PMID:28484499

Coordinated transcriptional regulation patterns associated with infertility phenotypes in men

PubMed Central

Ellis, Peter J I; Furlong, Robert A; Conner, Sarah J; Kirkman‐Brown, Jackson; Afnan, Masoud; Barratt, Christopher; Griffin, Darren K; Affara, Nabeel A

2007-01-01

Introduction Microarray gene‐expression profiling is a powerful tool for global analysis of the transcriptional consequences of disease phenotypes. Understanding the genetic correlates of particular pathological states is important for more accurate diagnosis and screening of patients, and thus for suggesting appropriate avenues of treatment. As yet, there has been little research describing gene‐expression profiling of infertile and subfertile men, and thus the underlying transcriptional events involved in loss of spermatogenesis remain unclear. Here we present the results of an initial screen of 33 patients with differing spermatogenic phenotypes. Methods Oligonucleotide array expression profiling was performed on testis biopsies for 33 patients presenting for testicular sperm extraction. Significantly regulated genes were selected using a mixed model analysis of variance. Principle components analysis and hierarchical clustering were used to interpret the resulting dataset with reference to the patient history, clinical findings and histological composition of the biopsies. Results Striking patterns of coordinated gene expression were found. The most significant contains multiple germ cell‐specific genes and corresponds to the degree of successful spermatogenesis in each patient, whereas a second pattern corresponds to inflammatory activity within the testis. Smaller‐scale patterns were also observed, relating to unique features of the individual biopsies. PMID:17496197

An alternative method to amplify RNA without loss of signal conservation for expression analysis with a proteinase DNA microarray in the ArrayTube format.

PubMed

Schüler, Susann; Wenz, Ingrid; Wiederanders, B; Slickers, P; Ehricht, R

2006-06-12

Recent developments in DNA microarray technology led to a variety of open and closed devices and systems including high and low density microarrays for high-throughput screening applications as well as microarrays of lower density for specific diagnostic purposes. Beside predefined microarrays for specific applications manufacturers offer the production of custom-designed microarrays adapted to customers' wishes. Array based assays demand complex procedures including several steps for sample preparation (RNA extraction, amplification and sample labelling), hybridization and detection, thus leading to a high variability between several approaches and resulting in the necessity of extensive standardization and normalization procedures. In the present work a custom designed human proteinase DNA microarray of lower density in ArrayTube format was established. This highly economic open platform only requires standard laboratory equipment and allows the study of the molecular regulation of cell behaviour by proteinases. We established a procedure for sample preparation and hybridization and verified the array based gene expression profile by quantitative real-time PCR (QRT-PCR). Moreover, we compared the results with the well established Affymetrix microarray. By application of standard labelling procedures with e.g. Klenow fragment exo-, single primer amplification (SPA) or In Vitro Transcription (IVT) we noticed a loss of signal conservation for some genes. To overcome this problem we developed a protocol in accordance with the SPA protocol, in which we included target specific primers designed individually for each spotted oligomer. Here we present a complete array based assay in which only the specific transcripts of interest are amplified in parallel and in a linear manner. The array represents a proof of principle which can be adapted to other species as well. As the designed protocol for amplifying mRNA starts from as little as 100 ng total RNA, it presents an alternative method for detecting even low expressed genes by microarray experiments in a highly reproducible and sensitive manner. Preservation of signal integrity is demonstrated out by QRT-PCR measurements. The little amounts of total RNA necessary for the analyses make this method applicable for investigations with limited material as in clinical samples from, for example, organ or tumour biopsies. Those are arguments in favour of the high potential of our assay compared to established procedures for amplification within the field of diagnostic expression profiling. Nevertheless, the screening character of microarray data must be mentioned, and independent methods should verify the results.

Two-pass imputation algorithm for missing value estimation in gene expression time series.

PubMed

Tsiporkova, Elena; Boeva, Veselka

2007-10-01

Gene expression microarray experiments frequently generate datasets with multiple values missing. However, most of the analysis, mining, and classification methods for gene expression data require a complete matrix of gene array values. Therefore, the accurate estimation of missing values in such datasets has been recognized as an important issue, and several imputation algorithms have already been proposed to the biological community. Most of these approaches, however, are not particularly suitable for time series expression profiles. In view of this, we propose a novel imputation algorithm, which is specially suited for the estimation of missing values in gene expression time series data. The algorithm utilizes Dynamic Time Warping (DTW) distance in order to measure the similarity between time expression profiles, and subsequently selects for each gene expression profile with missing values a dedicated set of candidate profiles for estimation. Three different DTW-based imputation (DTWimpute) algorithms have been considered: position-wise, neighborhood-wise, and two-pass imputation. These have initially been prototyped in Perl, and their accuracy has been evaluated on yeast expression time series data using several different parameter settings. The experiments have shown that the two-pass algorithm consistently outperforms, in particular for datasets with a higher level of missing entries, the neighborhood-wise and the position-wise algorithms. The performance of the two-pass DTWimpute algorithm has further been benchmarked against the weighted K-Nearest Neighbors algorithm, which is widely used in the biological community; the former algorithm has appeared superior to the latter one. Motivated by these findings, indicating clearly the added value of the DTW techniques for missing value estimation in time series data, we have built an optimized C++ implementation of the two-pass DTWimpute algorithm. The software also provides for a choice between three different initial rough imputation methods.

Identification of molecular pathways affected by pterostilbene, a natural dimethylether analog of resveratrol

PubMed Central

Pan, Zhiqiang; Agarwal, Ameeta K; Xu, Tao; Feng, Qin; Baerson, Scott R; Duke, Stephen O; Rimando, Agnes M

2008-01-01

Background Pterostilbene, a naturally occurring phenolic compound produced by agronomically important plant genera such as Vitis and Vacciunium, is a phytoalexin exhibiting potent antifungal activity. Additionally, recent studies have demonstrated several important pharmacological properties associated with pterostilbene. Despite this, a systematic study of the effects of pterostilbene on eukaryotic cells at the molecular level has not been previously reported. Thus, the aim of the present study was to identify the cellular pathways affected by pterostilbene by performing transcript profiling studies, employing the model yeast Saccharomyces cerevisiae. Methods S. cerevisiae strain S288C was exposed to pterostilbene at the IC50 concentration (70 μM) for one generation (3 h). Transcript profiling experiments were performed on three biological replicate samples using the Affymetrix GeneChip Yeast Genome S98 Array. The data were analyzed using the statistical methods available in the GeneSifter microarray data analysis system. To validate the results, eleven differentially expressed genes were further examined by quantitative real-time RT-PCR, and S. cerevisiae mutant strains with deletions in these genes were analyzed for altered sensitivity to pterostilbene. Results Transcript profiling studies revealed that pterostilbene exposure significantly down-regulated the expression of genes involved in methionine metabolism, while the expression of genes involved in mitochondrial functions, drug detoxification, and transcription factor activity were significantly up-regulated. Additional analyses revealed that a large number of genes involved in lipid metabolism were also affected by pterostilbene treatment. Conclusion Using transcript profiling, we have identified the cellular pathways targeted by pterostilbene, an analog of resveratrol. The observed response in lipid metabolism genes is consistent with its known hypolipidemic properties, and the induction of mitochondrial genes is consistent with its demonstrated role in apoptosis in human cancer cell lines. Furthermore, our data show that pterostilbene has a significant effect on methionine metabolism, a previously unreported effect for this compound. PMID:18366703

Transcriptional network inference from functional similarity and expression data: a global supervised approach.

PubMed

Ambroise, Jérôme; Robert, Annie; Macq, Benoit; Gala, Jean-Luc

2012-01-06

An important challenge in system biology is the inference of biological networks from postgenomic data. Among these biological networks, a gene transcriptional regulatory network focuses on interactions existing between transcription factors (TFs) and and their corresponding target genes. A large number of reverse engineering algorithms were proposed to infer such networks from gene expression profiles, but most current methods have relatively low predictive performances. In this paper, we introduce the novel TNIFSED method (Transcriptional Network Inference from Functional Similarity and Expression Data), that infers a transcriptional network from the integration of correlations and partial correlations of gene expression profiles and gene functional similarities through a supervised classifier. In the current work, TNIFSED was applied to predict the transcriptional network in Escherichia coli and in Saccharomyces cerevisiae, using datasets of 445 and 170 affymetrix arrays, respectively. Using the area under the curve of the receiver operating characteristics and the F-measure as indicators, we showed the predictive performance of TNIFSED to be better than unsupervised state-of-the-art methods. TNIFSED performed slightly worse than the supervised SIRENE algorithm for the target genes identification of the TF having a wide range of yet identified target genes but better for TF having only few identified target genes. Our results indicate that TNIFSED is complementary to the SIRENE algorithm, and particularly suitable to discover target genes of "orphan" TFs.

Expression Profiling-Based Identification of CO2-Responsive Genes Regulated by CCM1 Controlling a Carbon-Concentrating Mechanism in Chlamydomonas reinhardtii1

PubMed Central

Miura, Kenji; Yamano, Takashi; Yoshioka, Satoshi; Kohinata, Tsutomu; Inoue, Yoshihiro; Taniguchi, Fumiya; Asamizu, Erika; Nakamura, Yasukazu; Tabata, Satoshi; Yamato, Katsuyuki T.; Ohyama, Kanji; Fukuzawa, Hideya

2004-01-01

Photosynthetic acclimation to CO2-limiting stress is associated with control of genetic and physiological responses through a signal transduction pathway, followed by integrated monitoring of the environmental changes. Although several CO2-responsive genes have been previously isolated, genome-wide analysis has not been applied to the isolation of CO2-responsive genes that may function as part of a carbon-concentrating mechanism (CCM) in photosynthetic eukaryotes. By comparing expression profiles of cells grown under CO2-rich conditions with those of cells grown under CO2-limiting conditions using a cDNA membrane array containing 10,368 expressed sequence tags, 51 low-CO2 inducible genes and 32 genes repressed by low CO2 whose mRNA levels were changed more than 2.5-fold in Chlamydomonas reinhardtii Dangeard were detected. The fact that the induction of almost all low-CO2 inducible genes was impaired in the ccm1 mutant suggests that CCM1 is a master regulator of CCM through putative low-CO2 signal transduction pathways. Among low-CO2 inducible genes, two novel genes, LciA and LciB, were identified, which may be involved in inorganic carbon transport. Possible functions of low-CO2 inducible and/or CCM1-regulated genes are discussed in relation to the CCM. PMID:15235119

A Dynamic Bronchial Airway Gene Expression Signature of Chronic Obstructive Pulmonary Disease and Lung Function Impairment

PubMed Central

Steiling, Katrina; van den Berge, Maarten; Hijazi, Kahkeshan; Florido, Roberta; Campbell, Joshua; Liu, Gang; Xiao, Ji; Zhang, Xiaohui; Duclos, Grant; Drizik, Eduard; Si, Huiqing; Perdomo, Catalina; Dumont, Charles; Coxson, Harvey O.; Alekseyev, Yuriy O.; Sin, Don; Pare, Peter; Hogg, James C.; McWilliams, Annette; Hiemstra, Pieter S.; Sterk, Peter J.; Timens, Wim; Chang, Jeffrey T.; Sebastiani, Paola; O’Connor, George T.; Bild, Andrea H.; Postma, Dirkje S.; Lam, Stephen

2013-01-01

Rationale: Molecular phenotyping of chronic obstructive pulmonary disease (COPD) has been impeded in part by the difficulty in obtaining lung tissue samples from individuals with impaired lung function. Objectives: We sought to determine whether COPD-associated processes are reflected in gene expression profiles of bronchial airway epithelial cells obtained by bronchoscopy. Methods: Gene expression profiling of bronchial brushings obtained from 238 current and former smokers with and without COPD was performed using Affymetrix Human Gene 1.0 ST Arrays. Measurements and Main Results: We identified 98 genes whose expression levels were associated with COPD status, FEV1% predicted, and FEV1/FVC. In silico analysis identified activating transcription factor 4 (ATF4) as a potential transcriptional regulator of genes with COPD-associated airway expression, and ATF4 overexpression in airway epithelial cells in vitro recapitulates COPD-associated gene expression changes. Genes with COPD-associated expression in the bronchial airway epithelium had similarly altered expression profiles in prior studies performed on small-airway epithelium and lung parenchyma, suggesting that transcriptomic alterations in the bronchial airway epithelium reflect molecular events found at more distal sites of disease activity. Many of the airway COPD-associated gene expression changes revert toward baseline after therapy with the inhaled corticosteroid fluticasone in independent cohorts. Conclusions: Our findings demonstrate a molecular field of injury throughout the bronchial airway of active and former smokers with COPD that may be driven in part by ATF4 and is modifiable with therapy. Bronchial airway epithelium may ultimately serve as a relatively accessible tissue in which to measure biomarkers of disease activity for guiding clinical management of COPD. PMID:23471465

A Sorghum bicolor expression atlas reveals dynamic genotype-specific expression profiles for vegetative tissues of grain, sweet and bioenergy sorghums.

PubMed

Shakoor, Nadia; Nair, Ramesh; Crasta, Oswald; Morris, Geoffrey; Feltus, Alex; Kresovich, Stephen

2014-01-23

Effective improvement in sorghum crop development necessitates a genomics-based approach to identify functional genes and QTLs. Sequenced in 2009, a comprehensive annotation of the sorghum genome and the development of functional genomics resources is key to enable the discovery and deployment of regulatory and metabolic genes and gene networks for crop improvement. This study utilizes the first commercially available whole-transcriptome sorghum microarray (Sorgh-WTa520972F) to identify tissue and genotype-specific expression patterns for all identified Sorghum bicolor exons and UTRs. The genechip contains 1,026,373 probes covering 149,182 exons (27,577 genes) across the Sorghum bicolor nuclear, chloroplast, and mitochondrial genomes. Specific probesets were also included for putative non-coding RNAs that may play a role in gene regulation (e.g., microRNAs), and confirmed functional small RNAs in related species (maize and sugarcane) were also included in our array design. We generated expression data for 78 samples with a combination of four different tissue types (shoot, root, leaf and stem), two dissected stem tissues (pith and rind) and six diverse genotypes, which included 6 public sorghum lines (R159, Atlas, Fremont, PI152611, AR2400 and PI455230) representing grain, sweet, forage, and high biomass ideotypes. Here we present a summary of the microarray dataset, including analysis of tissue-specific gene expression profiles and associated expression profiles of relevant metabolic pathways. With an aim to enable identification and functional characterization of genes in sorghum, this expression atlas presents a new and valuable resource to the research community.

A Sorghum bicolor expression atlas reveals dynamic genotype-specific expression profiles for vegetative tissues of grain, sweet and bioenergy sorghums

PubMed Central

2014-01-01

Background Effective improvement in sorghum crop development necessitates a genomics-based approach to identify functional genes and QTLs. Sequenced in 2009, a comprehensive annotation of the sorghum genome and the development of functional genomics resources is key to enable the discovery and deployment of regulatory and metabolic genes and gene networks for crop improvement. Results This study utilizes the first commercially available whole-transcriptome sorghum microarray (Sorgh-WTa520972F) to identify tissue and genotype-specific expression patterns for all identified Sorghum bicolor exons and UTRs. The genechip contains 1,026,373 probes covering 149,182 exons (27,577 genes) across the Sorghum bicolor nuclear, chloroplast, and mitochondrial genomes. Specific probesets were also included for putative non-coding RNAs that may play a role in gene regulation (e.g., microRNAs), and confirmed functional small RNAs in related species (maize and sugarcane) were also included in our array design. We generated expression data for 78 samples with a combination of four different tissue types (shoot, root, leaf and stem), two dissected stem tissues (pith and rind) and six diverse genotypes, which included 6 public sorghum lines (R159, Atlas, Fremont, PI152611, AR2400 and PI455230) representing grain, sweet, forage, and high biomass ideotypes. Conclusions Here we present a summary of the microarray dataset, including analysis of tissue-specific gene expression profiles and associated expression profiles of relevant metabolic pathways. With an aim to enable identification and functional characterization of genes in sorghum, this expression atlas presents a new and valuable resource to the research community. PMID:24456189

Unique gene expression profiles of donor-matched human retinal and choroidal vascular endothelial cells.

PubMed

Smith, Justine R; Choi, Dongseok; Chipps, Timothy J; Pan, Yuzhen; Zamora, David O; Davies, Michael H; Babra, Bobby; Powers, Michael R; Planck, Stephen R; Rosenbaum, James T

2007-06-01

Consistent with clinical observations that posterior uveitis frequently involves the retinal vasculature and recent recognition of vascular heterogeneity, the hypothesis for this study was that retinal vascular endothelium was a cell population of unique molecular phenotype. Donor-matched cultures of primary retinal and choroidal endothelial cells from six human cadavers were incubated with either Toxoplasma gondii tachyzoites (10:1, parasites per cell) or Escherichia coli lipopolysaccharide (100 ng/mL); control cultures were simultaneously incubated with medium. Gene expression profiling of endothelial cells was performed using oligonucleotide arrays containing probes designed to detect 8746 human transcripts. After normalization, differential gene expression was assessed by the significance analysis of microarrays, with the false-discovery rate set at 5%. For selected genes, differences in the level of expression between retinal and choroidal cells were evaluated by real-time RT-PCR. Graphic descriptive analysis demonstrated a strong correlation between gene expression of unstimulated retinal and choroidal endothelial cells, but also highlighted distinctly different patterns of expression that were greater than differences noted between donors or between unstimulated and stimulated cells. Overall, 779 (8.9%) of 8746 transcripts were differentially represented. Of note, the 330 transcripts that were present at higher levels in retinal cells included a larger percentage of transcripts encoding molecules involved in the immune response. Differential gene expression was confirmed for 12 transcripts by RT-PCR. Retinal and choroidal vascular endothelial cells display distinctive gene expression profiles. The findings suggest the possibility of treating posterior uveitis by targeting specific interactions between the retinal endothelial cell and an infiltrating leukocyte.

Microarray profiling of progesterone-regulated endometrial genes during the rhesus monkey secretory phase

PubMed Central

Ace, Christopher I; Okulicz, William C

2004-01-01

Background In the endometrium the steroid hormone progesterone (P), acting through its nuclear receptors, regulates the expression of specific target genes and gene networks required for endometrial maturation. Proper endometrial maturation is considered a requirement for embryo implantation. Endometrial receptivity is a complex process that is spatially and temporally restricted and the identity of genes that regulate receptivity has been pursued by a number of investigators. Methods In this study we have used high density oligonucleotide microarrays to screen for changes in mRNA transcript levels between normal proliferative and adequate secretory phases in Rhesus monkey artificial menstrual cycles. Biotinylated cRNA was prepared from day 13 and days 21–23 of the reproductive cycle and transcript levels were compared by hybridization to Affymetrix HG-U95A arrays. Results Of ~12,000 genes profiled, we identified 108 genes that were significantly regulated during the shift from a proliferative to an adequate secretory endometrium. Of these genes, 39 were up-regulated at days 21–23 versus day 13, and 69 were down-regulated. Genes up-regulated in P-dominant tissue included: secretoglobin (uteroglobin), histone 2A, polo-like kinase (PLK), spermidine/spermine acetyltransferase 2 (SAT2), secretory leukocyte protease inhibitor (SLPI) and metallothionein 1G (MT1G), all of which have been previously documented as elevated in the Rhesus monkey or human endometrium during the secretory phase. Genes down-regulated included: transforming growth factor beta-induced (TGFBI or BIGH3), matrix metalloproteinase 11 (stromelysin 3), proenkephalin (PENK), cysteine/glycine-rich protein 2 (CSRP2), collagen type VII alpha 1 (COL7A1), secreted frizzled-related protein 4 (SFRP4), progesterone receptor membrane component 1 (PGRMC1), chemokine (C-X-C) ligand 12 (CXCL12) and biglycan (BGN). In addition, many novel/unknown genes were also identified. Validation of array data was performed by semi-quantitative RT-PCR of two selected up-regulated genes using temporal (cycle day specific) endometrial cDNA populations. This approach confirmed up-regulation of WAP four-disulfide core domain 2 (WFDC2) and SLPI during the expected window of receptivity. Conclusion The identification of P-regulated genes and gene pathways in the primate endometrium is expected to be an important first step in elucidating the cellular processes necessary for the development of a receptive environment for implantation. PMID:15239838

Leaving out control groups: an internal contrast analysis of gene expression profiles in atrial fibrillation patients ‐ A systems biology approach to clinical categorization

PubMed Central

Vanhoutte, Kurt; de Asmundis, Carlo; Francesconi, Anna; Figys1, Jurgen; Steurs, Griet; Boussy, Tim; Roos, Markus; Mueller, Andreas; Massimo, Lucio; Paparella, Gaetano; Van Caelenberg, Kristien; Chierchia, Gian Battista; Sarkozy, Andrea; Y Terradellas, Pedro Brugada; Zizi, Martin

2009-01-01

Atrial fibrillation (AF) is a frequent chronic dysrythmia with an incidence that increases with age (>40). Because of its medical and socio-economic impacts it is expected to become an increasing burden on most health care systems. AF is a multi-factorial disease for which the identification of subtypes is warranted. Novel approaches based on the broad concepts of systems biology may overcome the blurred notion of normal and pathological phenotype, which is inherent to high throughput molecular arrays analysis. Here we apply an internal contrast algorithm on AF patient data with an analytical focus on potential entry pathways into the disease. We used a RMA (Robust Multichip Average) normalized Affymetrix micro-array data set from 10 AF patients (geo_accession #GSE2240). Four series of probes were selected based on physiopathogenic links with AF entryways: apoptosis (remodeling), MAP kinase (cell remodeling), OXPHOS (ability to sustain hemodynamic workload) and glycolysis (ischemia). Annotated probe lists were polled with Bioconductor packages in R (version 2.7.1). Genetic profile contrasts were analysed with hierarchical clustering and principal component analysis. The analysis revealed distinct patient groups for all probe sets. A substantial part (54% till 67%) of the variance is explained in the first 2 principal components. Genes in PC1/2 with high discriminatory value were selected and analyzed in detail. We aim for reliable molecular stratification of AF. We show that stratification is possible based on physiologically relevant gene sets. Genes with high contrast value are likely to give pathophysiological insight into permanent AF subtypes. PMID:19255648

Organic trace mineral levels in the first 96-h post-hatch impact growth performance and intestinal gene expression in broiler chicks.

PubMed

Brennan, K M; Samuel, R S; Graugnard, T Ao; Xiao, R; Cantor, A H; Pescatore, A J

2013-12-01

Alterations in nutrient intake in the avian neonatal posthatch period can impact development, performance, and metabolism in adulthood. Very little is known about how mineral levels during the post-hatch period affect or “program” gene expression patterns later in life. The objective of this study was to determine the effect of post-hatch (0 to 96 h) dietary mineral supplementation on performance, tissue mineral content, and intestinal gene expression profiles in 21-day-old broiler chicks. One-day-old chicks were randomly assigned to one of two treatment groups consisting of N (organic Zn, Cu, and Mn provided at 100 % of recommendations (National Research Council 1994)) and/or L (organic Zn, Cu, and Mn provided at 20 % of recommendations (National Research Council 1994)) diets fed in two intervals (days 1–4, days 5–21) as follows: (1)N–Lor (2)L–L. Performance parameters did not differ between treatments except that body weight gain was greater (P < 0.05) in L–L birds than N–L birds over the experimental period. Bone mineral content was similar for both treatments at day 21. Intestinal gene expression profiling was examined using the Affymetrix GeneChip Chicken genome array. Ingenuity pathway analysis revealed differences in gene expression profiles between N and L treatments at day 5. At day 21, profiles were unique between N–L and L–L, suggesting that the diet fed until day 4 had an impact on gene expression patterns at day 21 even when birds were fed the same diets day 5–day 21. In this study, we demonstrated that diets fed for the 96 h post-hatch had long-term effects on gene expression, providing unique information as to why post-hatch diets are so important for the longterm bird health and productivity.

Peripheral blood mononuclear cell gene expression profile in obese boys who followed a moderate energy-restricted diet: differences between high and low responders at baseline and after the intervention.

PubMed

Rendo-Urteaga, Tara; García-Calzón, Sonia; González-Muniesa, Pedro; Milagro, Fermín I; Chueca, María; Oyarzabal, Mirentxu; Azcona-Sanjulián, M Cristina; Martínez, J Alfredo; Marti, Amelia

2015-01-28

The present study analyses the gene expression profile of peripheral blood mononuclear cells (PBMC) from obese boys. The aims of the present study were to identify baseline differences between low responders (LR) and high responders (HR) after 10 weeks of a moderate energy-restricted dietary intervention, and to compare the gene expression profile between the baseline and the endpoint of the nutritional intervention. Spanish obese boys (age 10-14 years) were advised to follow a 10-week moderate energy-restricted diet. Participants were classified into two groups based on the association between the response to the nutritional intervention and the changes in BMI standard deviation score (BMI-SDS): HR group (n 6), who had a more decreased BMI-SDS; LR group (n 6), who either maintained or had an even increased BMI-SDS. The expression of 28,869 genes was analysed in PBMC from both groups at baseline and after the nutritional intervention, using the Affymetrix Human Gene 1.1 ST 24-Array plate microarray. At baseline, the HR group showed a lower expression of inflammation and immune response-related pathways, which suggests that the LR group could have a more developed pro-inflammatory phenotype. Concomitantly, LEPR and SIRPB1 genes were highly expressed in the LR group, indicating a tendency towards an impaired immune response and leptin resistance. Moreover, the moderate energy-restricted diet was able to down-regulate the inflammatory 'mitogen-activated protein kinase signalling pathway' in the HR group, as well as some inflammatory genes (AREG and TNFAIP3). The present study confirms that changes in the gene expression profile of PBMC in obese boys may help to understand the weight-loss response. However, further research is required to confirm these findings.

Conjunctival transcriptome profiling of Solomon Islanders with active trachoma in the absence of Chlamydia trachomatis infection.

PubMed

Vasileva, Hristina; Butcher, Robert; Pickering, Harry; Sokana, Oliver; Jack, Kelvin; Solomon, Anthony W; Holland, Martin J; Roberts, Chrissy H

2018-02-21

Clinical signs of active (inflammatory) trachoma are found in many children in the Solomon Islands, but the majority of these individuals have no serological evidence of previous infection with Chlamydia trachomatis. In Temotu and Rennell and Bellona provinces, ocular infections with C. trachomatis were seldom detected among children with active trachoma; a similar lack of association was seen between active trachoma and other common bacterial and viral causes of follicular conjunctivitis. Here, we set out to characterise patterns of gene expression at the conjunctivae of children in these provinces with and without clinical signs of trachomatous inflammation-follicular (TF) and C. trachomatis infection. Purified RNA from children with and without active trachoma was run on Affymetrix GeneChip Human Transcriptome Array 2.0 microarrays. Profiles were compared between individuals with ocular C. trachomatis infection and TF (group DI; n = 6), individuals with TF but no C. trachomatis infection (group D; n = 7), and individuals without TF or C. trachomatis infection (group N; n = 7). Differential gene expression and gene set enrichment for pathway membership were assessed. Conjunctival gene expression profiles were more similar within-group than between-group. Principal components analysis indicated that the first and second principal components combined explained almost 50% of the variance in the dataset. When comparing the DI group to the N group, genes involved in T-cell proliferation, B-cell signalling and CD8+ T cell signalling pathways were differentially regulated. When comparing the DI group to the D group, CD8+ T-cell regulation, interferon-gamma and IL17 production pathways were enriched. Genes involved in RNA transcription and translation pathways were upregulated when comparing the D group to the N group. Gene expression profiles in children in the Solomon Islands indicate immune responses consistent with bacterial infection when TF and C. trachomatis infection are concurrent. The transcriptomes of children with TF but without identified infection were not consistent with allergic or viral conjunctivitis.

Comprehensive analysis of alternative splicing and functionality in neuronal differentiation of P19 cells.

PubMed

Suzuki, Hitoshi; Osaki, Ken; Sano, Kaori; Alam, A H M Khurshid; Nakamura, Yuichiro; Ishigaki, Yasuhito; Kawahara, Kozo; Tsukahara, Toshifumi

2011-02-18

Alternative splicing, which produces multiple mRNAs from a single gene, occurs in most human genes and contributes to protein diversity. Many alternative isoforms are expressed in a spatio-temporal manner, and function in diverse processes, including in the neural system. The purpose of the present study was to comprehensively investigate neural-splicing using P19 cells. GeneChip Exon Array analysis was performed using total RNAs purified from cells during neuronal cell differentiation. To efficiently and readily extract the alternative exon candidates, 9 filtering conditions were prepared, yielding 262 candidate exons (236 genes). Semiquantitative RT-PCR results in 30 randomly selected candidates suggested that 87% of the candidates were differentially alternatively spliced in neuronal cells compared to undifferentiated cells. Gene ontology and pathway analyses suggested that many of the candidate genes were associated with neural events. Together with 66 genes whose functions in neural cells or organs were reported previously, 47 candidate genes were found to be linked to 189 events in the gene-level profile of neural differentiation. By text-mining for the alternative isoform, distinct functions of the isoforms of 9 candidate genes indicated by the result of Exon Array were confirmed. Alternative exons were successfully extracted. Results from the informatics analyses suggested that neural events were primarily governed by genes whose expression was increased and whose transcripts were differentially alternatively spliced in the neuronal cells. In addition to known functions in neural cells or organs, the uninvestigated alternative splicing events of 11 genes among 47 candidate genes suggested that cell cycle events are also potentially important. These genes may help researchers to differentiate the roles of alternative splicing in cell differentiation and cell proliferation.

Mitochondrial Gene Expression Profiles and Metabolic Pathways in the Amygdala Associated with Exaggerated Fear in an Animal Model of PTSD.

PubMed

Li, He; Li, Xin; Smerin, Stanley E; Zhang, Lei; Jia, Min; Xing, Guoqiang; Su, Yan A; Wen, Jillian; Benedek, David; Ursano, Robert

2014-01-01

The metabolic mechanisms underlying the development of exaggerated fear in post-traumatic stress disorder (PTSD) are not well defined. In the present study, alteration in the expression of genes associated with mitochondrial function in the amygdala of an animal model of PTSD was determined. Amygdala tissue samples were excised from 10 non-stressed control rats and 10 stressed rats, 14 days post-stress treatment. Total RNA was isolated, cDNA was synthesized, and gene expression levels were determined using a cDNA microarray. During the development of the exaggerated fear associated with PTSD, 48 genes were found to be significantly upregulated and 37 were significantly downregulated in the amygdala complex based on stringent criteria (p < 0.01). Ingenuity pathway analysis revealed up- or downregulation in the amygdala complex of four signaling networks - one associated with inflammatory and apoptotic pathways, one with immune mediators and metabolism, one with transcriptional factors, and one with chromatin remodeling. Thus, informatics of a neuronal gene array allowed us to determine the expression profile of mitochondrial genes in the amygdala complex of an animal model of PTSD. The result is a further understanding of the metabolic and neuronal signaling mechanisms associated with delayed and exaggerated fear.

Early indicators of exposure to biological threat agents using host gene profiles in peripheral blood mononuclear cells

PubMed Central

Das, Rina; Hammamieh, Rasha; Neill, Roger; Ludwig, George V; Eker, Steven; Lincoln, Patrick; Ramamoorthy, Preveen; Dhokalia, Apsara; Mani, Sachin; Mendis, Chanaka; Cummings, Christiano; Kearney, Brian; Royaee, Atabak; Huang, Xiao-Zhe; Paranavitana, Chrysanthi; Smith, Leonard; Peel, Sheila; Kanesa-Thasan, Niranjan; Hoover, David; Lindler, Luther E; Yang, David; Henchal, Erik; Jett, Marti

2008-01-01

Background Effective prophylaxis and treatment for infections caused by biological threat agents (BTA) rely upon early diagnosis and rapid initiation of therapy. Most methods for identifying pathogens in body fluids and tissues require that the pathogen proliferate to detectable and dangerous levels, thereby delaying diagnosis and treatment, especially during the prelatent stages when symptoms for most BTA are indistinguishable flu-like signs. Methods To detect exposures to the various pathogens more rapidly, especially during these early stages, we evaluated a suite of host responses to biological threat agents using global gene expression profiling on complementary DNA arrays. Results We found that certain gene expression patterns were unique to each pathogen and that other gene changes occurred in response to multiple agents, perhaps relating to the eventual course of illness. Nonhuman primates were exposed to some pathogens and the in vitro and in vivo findings were compared. We found major gene expression changes at the earliest times tested post exposure to aerosolized B. anthracis spores and 30 min post exposure to a bacterial toxin. Conclusion Host gene expression patterns have the potential to serve as diagnostic markers or predict the course of impending illness and may lead to new stage-appropriate therapeutic strategies to ameliorate the devastating effects of exposure to biothreat agents. PMID:18667072

Comparative expression profiling of Nicotiana benthamiana leaves systemically infected with three fruit tree viruses.

PubMed

Dardick, Christopher

2007-08-01

Plant viruses cause a wide array of disease symptoms and cytopathic effects. Although some of these changes are virus specific, many appear to be common even among diverse viruses. Currently, little is known about the underlying molecular determinants. To identify gene expression changes that are concomitant with virus symptoms, we performed comparative expression profiling experiments on Nicotiana benthamiana leaves infected with one of three different fruit tree viruses that produce distinct symptoms: Plum pox potyvirus (PPV; leaf distortion and mosaic), Tomato ringspot nepovirus (ToRSV; tissue necrosis and general chlorosis), and Prunus necrotic ringspot ilarvirus (PNRSV; subtle chlorotic mottling). The numbers of statistically significant genes identified were consistent with the severity of the observed symptoms: 1,082 (ToRSV), 744 (PPV), and 89 (PNRSV). In all, 56% of the gene expression changes found in PPV-infected leaves also were altered by ToRSV, 87% of which changed in the same direction. Both PPV- and ToRSV-infected leaves showed widespread repression of genes associated with plastid functions. PPV uniquely induced the expression of large numbers of cytosolic ribosomal genes whereas ToRSV repressed the expression of plastidic ribosomal genes. How these and other observed expression changes might be associated with symptom development are discussed.

Transcriptome Profiling of In-Vivo Produced Bovine Pre-implantation Embryos Using Two-color Microarray Platform.

PubMed

Salehi, Reza; Tsoi, Stephen C M; Colazo, Marcos G; Ambrose, Divakar J; Robert, Claude; Dyck, Michael K

2017-01-30

Early embryonic loss is a large contributor to infertility in cattle. Moreover, bovine becomes an interesting model to study human preimplantation embryo development due to their similar developmental process. Although genetic factors are known to affect early embryonic development, the discovery of such factors has been a serious challenge. Microarray technology allows quantitative measurement and gene expression profiling of transcript levels on a genome-wide basis. One of the main decisions that have to be made when planning a microarray experiment is whether to use a one- or two-color approach. Two-color design increases technical replication, minimizes variability, improves sensitivity and accuracy as well as allows having loop designs, defining the common reference samples. Although microarray is a powerful biological tool, there are potential pitfalls that can attenuate its power. Hence, in this technical paper we demonstrate an optimized protocol for RNA extraction, amplification, labeling, hybridization of the labeled amplified RNA to the array, array scanning and data analysis using the two-color analysis strategy.

Alteration of gene expression profiles in skeletal muscle of rats exposed to microgravity during a spaceflight

NASA Technical Reports Server (NTRS)

Taylor, Wayne E.; Bhasin, Shalender; Lalani, Rukhsana; Datta, Anuj; Gonzalez-Cadavid, Nestor F.

2002-01-01

To clarify the mechanism of skeletal muscle wasting during spaceflights, we investigated whether intramuscular gene expression profiles are affected, by using DNA microarray methods. Male rats sent on the 17-day NASA STS-90 Neurolab spaceflight were sacrificed 24 hours after return to earth (MG group). Ground control rats were maintained for 17 days in flight-simulated cages (CS group). Spaceflight induced a 19% and 23% loss of tibialis anterior and gastrocnemius muscle mass, respectively, as compared to ground controls. Muscle RNA was analyzed by the Clontech Atlas DNA expression array in four rats, with two MG/ CS pairs for the tibialis anterior, and one pair for the gastrocnemius. Alterations in gene expression were verified for selected genes by reverse-transcription PCR. In both muscles of MG rats, mRNAs for 12 genes were up-regulated by over 2-fold, and 38 were down-regulated compared to controls. There was inhibition of genes for cell proliferation and growth factor cascades, including cell cycle genes and signal transduction proteins, such as p21 Cip1, retinoblastoma (Rb), cyclins G1/S, -E and -D3, MAP kinase 3, MAD3, and ras related protein RAB2. These data indicate that following exposure to microgravity, there is downregulation of genes involved in regulation of muscle satellite cell replication.

Genome-wide transcriptional profiling of human glioblastoma cells in response to ITE treatment.

PubMed

Kang, Bo; Zhou, Yanwen; Zheng, Min; Wang, Ying-Jie

2015-09-01

A ligand-activated transcription factor aryl hydrocarbon receptor (AhR) is recently revealed to play a key role in embryogenesis and tumorigenesis (Feng et al. [1], Safe et al. [2]) and 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE) (Song et al. [3]) is an endogenous AhR ligand that possesses anti-tumor activity. In order to gain insights into how ITE acts via the AhR in embryogenesis and tumorigenesis, we analyzed the genome-wide transcriptional profiles of the following three groups of cells: the human glioblastoma U87 parental cells, U87 tumor sphere cells treated with vehicle (DMSO) and U87 tumor sphere cells treated with ITE. Here, we provide the details of the sample gathering strategy and show the quality controls and the analyses associated with our gene array data deposited into the Gene Expression Omnibus (GEO) under the accession code of GSE67986.

Expression Profiles of TGF-β and TLR Pathways in Porphyromonas gingivalis and Prevotella intermedia Challenged Osteoblasts

PubMed Central

Aydin, Kubra; Ekinci, Fatma Yesim; Korachi, May

2015-01-01

Background: The presence of certain oral pathogens at implant sites can hinder the osseointegration process. However, it is unclear how and by what microorganisms it happens. Objectives: This study investigated whether the presence of oral pathogens of Porphyromonas gingivalis and Prevotella intermedia individually, play a role in the failure of bone formation by determining the expression profiles of Transforming Growth Factor Beta (TGF-β/Bone Morphogenic Protein (BMP) and Toll-Like Receptor (TLR) pathways in challenged osteoblasts. Materials and Methods: Cell viability of P. gingivalis and P. intermedia challenged osteoblasts were determined by WST assay. Changes in osteoblast morphology and inhibition of mineralization were observed by Scanning Electron Microscopy (SEM) and Von Kossa staining, respectively. Expression of TGF-β and TLR pathway genes on challenged cells were identified by RT profiler array. Both P. gingivalis and P. intermedia challenges resulted in reduced viability and mineralization of osteoblasts. Results: Viability was reduced to 56.8% (P. gingivalis) and 52.75% (P. intermedia) at 1000 multiplicity. Amongst 48 genes examined, expressions of BMPER, SMAD1, IL8 and NFRKB were found to be highly upregulated by both bacterial challenges (Fold Change > 4). Conclusions: P. gingivalis and P. intermedia could play a role in implant failure by changing the expression profiles of genes related to bone formation and resorption. PMID:26034550

Profiling of Cytokines Secreted by Conventional Aqueous Outflow Pathway Endothelial Cells Activated In Vitro and Ex Vivo With Laser Irradiation

PubMed Central

Alvarado, Jorge A.; Chau, Phuonglan; Wu, Jianfeng; Juster, Richard; Shifera, Amde Selassie; Geske, Michael

2015-01-01

Purpose To profile which cytokine genes are differentially expressed (DE) as up- or downregulated by cultured human trabecular meshwork (TMEs) and Schlemm's canal endothelial cells (SCEs) after three experimental treatments consisting of selective laser trabeculoplasty (SLT) irradiation, exposure to media conditioned either by SLT-irradiated TMEs (TME-cm) or by SCEs (SCE-cm). Also, to profile which cytokines are upregulated ex vivo in SLT-irradiated human conventional aqueous outflow pathway (CAOP) tissues. Methods After each treatment, Affymetrix microarray assays were used to detect upregulated and downregulated genes for cytokines and their receptors in TMEs and SCEs. ELISA and protein antibody arrays were used to detect upregulated cytokines secreted in SLT-irradiated CAOP tissues ex vivo. Results The SLT irradiation upregulated numerous cytokine genes in TMEs, but only a few in SCEs. Exposure to TME- and SCE-cm induced SCEs to upregulate many more cytokine genes than TMEs. Selective laser trabeculoplasty irradiation and exposure to TME-cm downregulated several cytokine genes in TMEs but none in SCEs. Selective laser trabeculoplasty irradiation induced one upregulated and three downregulated cytokine-receptor genes in TMEs but none in SCEs. Exposure to TME-cm induced upregulation of one and downregulation of another receptor gene in TMEs, whereas two unique cytokine-receptor genes were upregulated in SCEs. Cytokine protein expression analysis showed that at least eight cytokines were upregulated in SLT-irradiated human CAOP tissues in situ/ex vivo. Conclusions This study has helped us identify a cytokine signaling pathway and to consider newly identified mechanisms regulating aqueous outflow that may lay the foundation for the future development of cytokine-based glaucoma therapies. PMID:26529044

Profiling of Cytokines Secreted by Conventional Aqueous Outflow Pathway Endothelial Cells Activated In Vitro and Ex Vivo With Laser Irradiation.

PubMed

Alvarado, Jorge A; Chau, Phuonglan; Wu, Jianfeng; Juster, Richard; Shifera, Amde Selassie; Geske, Michael

2015-11-01

To profile which cytokine genes are differentially expressed (DE) as up- or downregulated by cultured human trabecular meshwork (TMEs) and Schlemm's canal endothelial cells (SCEs) after three experimental treatments consisting of selective laser trabeculoplasty (SLT) irradiation, exposure to media conditioned either by SLT-irradiated TMEs (TME-cm) or by SCEs (SCE-cm). Also, to profile which cytokines are upregulated ex vivo in SLT-irradiated human conventional aqueous outflow pathway (CAOP) tissues. After each treatment, Affymetrix microarray assays were used to detect upregulated and downregulated genes for cytokines and their receptors in TMEs and SCEs. ELISA and protein antibody arrays were used to detect upregulated cytokines secreted in SLT-irradiated CAOP tissues ex vivo. The SLT irradiation upregulated numerous cytokine genes in TMEs, but only a few in SCEs. Exposure to TME- and SCE-cm induced SCEs to upregulate many more cytokine genes than TMEs. Selective laser trabeculoplasty irradiation and exposure to TME-cm downregulated several cytokine genes in TMEs but none in SCEs. Selective laser trabeculoplasty irradiation induced one upregulated and three downregulated cytokine-receptor genes in TMEs but none in SCEs. Exposure to TME-cm induced upregulation of one and downregulation of another receptor gene in TMEs, whereas two unique cytokine-receptor genes were upregulated in SCEs. Cytokine protein expression analysis showed that at least eight cytokines were upregulated in SLT-irradiated human CAOP tissues in situ/ex vivo. This study has helped us identify a cytokine signaling pathway and to consider newly identified mechanisms regulating aqueous outflow that may lay the foundation for the future development of cytokine-based glaucoma therapies.

Gene expression profiling of acute myeloid leukemia samples from adult patients with AML-M1 and -M2 through boutique microarrays, real-time PCR and droplet digital PCR.

PubMed

Handschuh, Luiza; Kaźmierczak, Maciej; Milewski, Marek C; Góralski, Michał; Łuczak, Magdalena; Wojtaszewska, Marzena; Uszczyńska-Ratajczak, Barbara; Lewandowski, Krzysztof; Komarnicki, Mieczysław; Figlerowicz, Marek

2018-03-01

Acute myeloid leukemia (AML) is the most common and severe form of acute leukemia diagnosed in adults. Owing to its heterogeneity, AML is divided into classes associated with different treatment outcomes and specific gene expression profiles. Based on previous studies on AML, in this study, we designed and generated an AML-array containing 900 oligonucleotide probes complementary to human genes implicated in hematopoietic cell differentiation and maturation, proliferation, apoptosis and leukemic transformation. The AML-array was used to hybridize 118 samples from 33 patients with AML of the M1 and M2 subtypes of the French-American‑British (FAB) classification and 15 healthy volunteers (HV). Rigorous analysis of the microarray data revealed that 83 genes were differentially expressed between the patients with AML and the HV, including genes not yet discussed in the context of AML pathogenesis. The most overexpressed genes in AML were STMN1, KITLG, CDK6, MCM5, KRAS, CEBPA, MYC, ANGPT1, SRGN, RPLP0, ENO1 and SET, whereas the most underexpressed genes were IFITM1, LTB, FCN1, BIRC3, LYZ, ADD3, S100A9, FCER1G, PTRPE, CD74 and TMSB4X. The overexpression of the CPA3 gene was specific for AML with mutated NPM1 and FLT3. Although the microarray-based method was insufficient to differentiate between any other AML subgroups, quantitative PCR approaches enabled us to identify 3 genes (ANXA3, S100A9 and WT1) whose expression can be used to discriminate between the 2 studied AML FAB subtypes. The expression levels of the ANXA3 and S100A9 genes were increased, whereas those of WT1 were decreased in the AML-M2 compared to the AML-M1 group. We also examined the association between the STMN1, CAT and ABL1 genes, and the FLT3 and NPM1 mutation status. FLT3+/NPM1- AML was associated with the highest expression of STMN1, and ABL1 was upregulated in FLT3+ AML and CAT in FLT3- AML, irrespectively of the NPM1 mutation status. Moreover, our results indicated that CAT and WT1 gene expression levels correlated with the response to therapy. CAT expression was highest in patients who remained longer under complete remission, whereas WT1 expression increased with treatment resistance. On the whole, this study demonstrates that the AML-array can potentially serve as a first-line screening tool, and may be helpful for the diagnosis of AML, whereas the differentiation between AML subgroups can be more successfully performed with PCR-based analysis of a few marker genes.

Single cell transcriptomic analysis of prostate cancer cells.

PubMed

Welty, Christopher J; Coleman, Ilsa; Coleman, Roger; Lakely, Bryce; Xia, Jing; Chen, Shu; Gulati, Roman; Larson, Sandy R; Lange, Paul H; Montgomery, Bruce; Nelson, Peter S; Vessella, Robert L; Morrissey, Colm

2013-02-16

The ability to interrogate circulating tumor cells (CTC) and disseminated tumor cells (DTC) is restricted by the small number detected and isolated (typically <10). To determine if a commercially available technology could provide a transcriptomic profile of a single prostate cancer (PCa) cell, we clonally selected and cultured a single passage of cell cycle synchronized C4-2B PCa cells. Ten sets of single, 5-, or 10-cells were isolated using a micromanipulator under direct visualization with an inverted microscope. Additionally, two groups of 10 individual DTC, each isolated from bone marrow of 2 patients with metastatic PCa were obtained. RNA was amplified using the WT-Ovation™ One-Direct Amplification System. The amplified material was hybridized on a 44K Whole Human Gene Expression Microarray. A high stringency threshold, a mean Alexa Fluor® 3 signal intensity above 300, was used for gene detection. Relative expression levels were validated for select genes using real-time PCR (RT-qPCR). Using this approach, 22,410, 20,423, and 17,009 probes were positive on the arrays from 10-cell pools, 5-cell pools, and single-cells, respectively. The sensitivity and specificity of gene detection on the single-cell analyses were 0.739 and 0.972 respectively when compared to 10-cell pools, and 0.814 and 0.979 respectively when compared to 5-cell pools, demonstrating a low false positive rate. Among 10,000 randomly selected pairs of genes, the Pearson correlation coefficient was 0.875 between the single-cell and 5-cell pools and 0.783 between the single-cell and 10-cell pools. As expected, abundant transcripts in the 5- and 10-cell samples were detected by RT-qPCR in the single-cell isolates, while lower abundance messages were not. Using the same stringency, 16,039 probes were positive on the patient single-cell arrays. Cluster analysis showed that all 10 DTC grouped together within each patient. A transcriptomic profile can be reliably obtained from a single cell using commercially available technology. As expected, fewer amplified genes are detected from a single-cell sample than from pooled-cell samples, however this method can be used to reliably obtain a transcriptomic profile from DTC isolated from the bone marrow of patients with PCa.

Identification of diagnostic markers in colorectal cancer via integrative epigenomics and genomics data

PubMed Central

KOK-SIN, TEOW; MOKHTAR, NORFILZA MOHD; HASSAN, NUR ZARINA ALI; SAGAP, ISMAIL; ROSE, ISA MOHAMED; HARUN, ROSLAN; JAMAL, RAHMAN

2015-01-01

Apart from genetic mutations, epigenetic alteration is a common phenomenon that contributes to neoplastic transformation in colorectal cancer. Transcriptional silencing of tumor-suppressor genes without changes in the DNA sequence is explained by the existence of promoter hypermethylation. To test this hypothesis, we integrated the epigenome and transcriptome data from a similar set of colorectal tissue samples. Methylation profiling was performed using the Illumina InfiniumHumanMethylation27 BeadChip on 55 paired cancer and adjacent normal epithelial cells. Fifteen of the 55 paired tissues were used for gene expression profiling using the Affymetrix GeneChip Human Gene 1.0 ST array. Validation was carried out on 150 colorectal tissues using the methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) technique. PCA and supervised hierarchical clustering in the two microarray datasets showed good separation between cancer and normal samples. Significant genes from the two analyses were obtained based on a ≥2-fold change and a false discovery rate (FDR) P-value of <0.05. We identified 1,081 differentially hypermethylated CpG sites and 36 hypomethylated CpG sites. We also found 709 upregulated and 699 downregulated genes from the gene expression profiling. A comparison of the two datasets revealed 32 overlapping genes with 27 being hypermethylated with downregulated expression and 4 hypermethylated with upregulated expression. One gene was found to be hypomethylated and downregulated. The most enriched molecular pathway identified was cell adhesion molecules that involved 4 overlapped genes, JAM2, NCAM1, ITGA8 and CNTN1. In the present study, we successfully identified a group of genes that showed methylation and gene expression changes in well-defined colorectal cancer tissues with high purity. The integrated analysis gives additional insight regarding the regulation of colorectal cancer-associated genes and their underlying mechanisms that contribute to colorectal carcinogenesis. PMID:25997610

Sex-specific patterns and deregulation of endocrine pathways in the gene expression profiles of Bangladeshi adults exposed to arsenic contaminated drinking water

DOE Office of Scientific and Technical Information (OSTI.GOV)

Muñoz, Alexandra; Chervona, Yana; Hall, Megan

Arsenic contamination of drinking water occurs globally and is associated with numerous diseases including skin, lung and bladder cancers, and cardiovascular disease. Recent research indicates that arsenic may be an endocrine disruptor. This study was conducted to evaluate the nature of gene expression changes among males and females exposed to arsenic contaminated water in Bangladesh at high and low doses. Twenty-nine (55% male) Bangladeshi adults with water arsenic exposure ranging from 50 to 1000 μg/L were selected from the Folic Acid Creatinine Trial. RNA was extracted from peripheral blood mononuclear cells for gene expression profiling using Affymetrix 1.0 ST arrays.more » Differentially expressed genes were assessed between high and low exposure groups for males and females separately and findings were validated using quantitative real-time PCR. There were 534 and 645 differentially expressed genes (p < 0.05) in the peripheral blood mononuclear cells of males and females, respectively, when high and low water arsenic exposure groups were compared. Only 43 genes overlapped between the two sexes, with 29 changing in opposite directions. Despite the difference in gene sets both males and females exhibited common biological changes including deregulation of 17β-hydroxysteroid dehydrogenase enzymes, deregulation of genes downstream of Sp1 (specificity protein 1) transcription factor, and prediction of estrogen receptor alpha as a key hub in cardiovascular networks. Arsenic-exposed adults exhibit sex-specific gene expression profiles that implicate involvement of the endocrine system. Due to arsenic's possible role as an endocrine disruptor, exposure thresholds for arsenic may require different parameters for males and females. - Highlights: • Males and females exhibit unique gene expression changes in response to arsenic. • Only 23 genes are common among the differentially expressed genes for the sexes. • Male and female gene lists exhibit common biological implications. • Both sexes exhibit deregulation of cardiovascular and endocrine pathways.« less

A Practical Platform for Blood Biomarker Study by Using Global Gene Expression Profiling of Peripheral Whole Blood

PubMed Central

Schmid, Patrick; Yao, Hui; Galdzicki, Michal; Berger, Bonnie; Wu, Erxi; Kohane, Isaac S.

2009-01-01

Background Although microarray technology has become the most common method for studying global gene expression, a plethora of technical factors across the experiment contribute to the variable of genome gene expression profiling using peripheral whole blood. A practical platform needs to be established in order to obtain reliable and reproducible data to meet clinical requirements for biomarker study. Methods and Findings We applied peripheral whole blood samples with globin reduction and performed genome-wide transcriptome analysis using Illumina BeadChips. Real-time PCR was subsequently used to evaluate the quality of array data and elucidate the mode in which hemoglobin interferes in gene expression profiling. We demonstrated that, when applied in the context of standard microarray processing procedures, globin reduction results in a consistent and significant increase in the quality of beadarray data. When compared to their pre-globin reduction counterparts, post-globin reduction samples show improved detection statistics, lowered variance and increased sensitivity. More importantly, gender gene separation is remarkably clearer in post-globin reduction samples than in pre-globin reduction samples. Our study suggests that the poor data obtained from pre-globin reduction samples is the result of the high concentration of hemoglobin derived from red blood cells either interfering with target mRNA binding or giving the pseudo binding background signal. Conclusion We therefore recommend the combination of performing globin mRNA reduction in peripheral whole blood samples and hybridizing on Illumina BeadChips as the practical approach for biomarker study. PMID:19381341

Identification of genetic aberrations on chromosome 22 outside the NF2 locus in schwannomatosis and neurofibromatosis type 2.

PubMed

Buckley, Patrick G; Mantripragada, Kiran K; Díaz de Ståhl, Teresita; Piotrowski, Arkadiusz; Hansson, Caisa M; Kiss, Hajnalka; Vetrie, David; Ernberg, Ingemar T; Nordenskjöld, Magnus; Bolund, Lars; Sainio, Markku; Rouleau, Guy A; Niimura, Michihito; Wallace, Andrew J; Evans, D Gareth R; Grigelionis, Gintautas; Menzel, Uwe; Dumanski, Jan P

2005-12-01

Schwannomatosis is characterized by multiple peripheral and cranial nerve schwannomas that occur in the absence of bilateral 8th cranial nerve schwannomas. The latter is the main diagnostic criterion of neurofibromatosis type 2 (NF2), which is a related but distinct disorder. The genetic factors underlying the differences between schwannomatosis and NF2 are poorly understood, although available evidence implicates chromosome 22 as the primary location of the gene(s) of interest. To investigate this, we comprehensively profiled the DNA copy number in samples from sporadic and familial schwannomatosis, NF2, and a large cohort of normal controls. Using a tiling-path chromosome 22 genomic array, we identified two candidate regions of copy number variation, which were further characterized by a PCR-based array with higher resolution. The latter approach allows the detection of minute alterations in total genomic DNA, with as little as 1.5 kb per measurement point of nonredundant sequence on the array. In DNA derived from peripheral blood from a schwannomatosis patient and a sporadic schwannoma sample, we detected rearrangements of the immunoglobulin lambda (IGL) locus, which is unlikely to be due to a B-cell specific somatic recombination of IGL. Analysis of normal controls indicated that these IGL rearrangements were restricted to schwannomatosis/schwannoma samples. In the second candidate region spanning GSTT1 and CABIN1 genes, we observed a frequent copy number polymorphism at the GSTT1 locus. We further describe missense mutations in the CABIN1 gene that are specific to samples from schwannomatosis and NF2 and make this gene a plausible candidate for contributing to the pathogenesis of these disorders. Copyright 2005 Wiley-Liss, Inc.

Omic personality: implications of stable transcript and methylation profiles for personalized medicine.

PubMed

Tabassum, Rubina; Sivadas, Ambily; Agrawal, Vartika; Tian, Haozheng; Arafat, Dalia; Gibson, Greg

2015-08-13

Personalized medicine is predicated on the notion that individual biochemical and genomic profiles are relatively constant in times of good health and to some extent predictive of disease or therapeutic response. We report a pilot study quantifying gene expression and methylation profile consistency over time, addressing the reasons for individual uniqueness, and its relation to N = 1 phenotypes. Whole blood samples from four African American women, four Caucasian women, and four Caucasian men drawn from the Atlanta Center for Health Discovery and Well Being study at three successive 6-month intervals were profiled by RNA-Seq, miRNA-Seq, and Illumina Methylation 450 K arrays. Standard regression approaches were used to evaluate the proportion of variance for each type of omic measure among individuals, and to quantify correlations among measures and with clinical attributes related to wellness. Longitudinal omic profiles were in general highly consistent over time, with an average of 67 % variance in transcript abundance, 42 % in CpG methylation level (but 88 % for the most differentiated CpG per gene), and 50 % in miRNA abundance among individuals, which are all comparable to 74 % variance among individuals for 74 clinical traits. One third of the variance could be attributed to differential blood cell type abundance, which was also fairly stable over time, and a lesser amount to expression quantitative trait loci (eQTL) effects. Seven conserved axes of covariance that capture diverse aspects of immune function explained over half of the variance. These axes also explained a considerable proportion of individually extreme transcript abundance, namely approximately 100 genes that were significantly up-regulated or down-regulated in each person and were in some cases enriched for relevant gene activities that plausibly associate with clinical attributes. A similar fraction of genes had individually divergent methylation levels, but these did not overlap with the transcripts, and fewer than 20 % of genes had significantly correlated methylation and gene expression. People express an "omic personality" consisting of peripheral blood transcriptional and epigenetic profiles that are constant over the course of a year and reflect various types of immune activity. Baseline genomic profiles can provide a window into the molecular basis of traits that might be useful for explaining medical conditions or guiding personalized health decisions.

Follicular lymphomas with and without translocation t(14;18) differ in gene expression profiles and genetic alterations

PubMed Central

Leich, Ellen; Salaverria, Itziar; Bea, Silvia; Zettl, Andreas; Wright, George; Moreno, Victor; Gascoyne, Randy D.; Chan, Wing-Chung; Braziel, Rita M.; Rimsza, Lisa M.; Weisenburger, Dennis D.; Delabie, Jan; Jaffe, Elaine S.; Lister, Andrew; Fitzgibbon, Jude; Staudt, Louis M.; Hartmann, Elena M.; Mueller-Hermelink, Hans-Konrad; Campo, Elias; Ott, German

2009-01-01

Follicular lymphoma (FL) is genetically characterized by the presence of the t(14;18)(q32;q21) chromosomal translocation in approximately 90% of cases. In contrast to FL carrying the t(14;18), their t(14;18)-negative counterparts are less well studied about their immunohistochemical, genetic, molecular, and clinical features. Within a previously published series of 184 FLs grades 1 to 3A with available gene expression data, we identified 17 FLs lacking the t(14;18). Comparative genomic hybridization and high-resolution single nucleotide polymorphism (SNP) array profiling showed that gains/amplifications of the BCL2 gene locus in 18q were restricted to the t(14;18)-positive FL subgroup. A comparison of gene expression profiles showed an enrichment of germinal center B cell–associated signatures in t(14;18)-positive FL, whereas activated B cell–like, NFκB, proliferation, and bystander cell signatures were enriched in t(14;18)-negative FL. These findings were confirmed by immunohistochemistry in an independent validation series of 84 FLs, in which 32% of t(14;18)-negative FLs showed weak or absent CD10 expression and 91% an increased Ki67 proliferation rate. Although overall survival did not differ between FL with and without t(14;18), our findings suggest distinct molecular features of t(14;18)-negative FL. PMID:19471018

Gene expression profiling defined pathways correlated with fibroblast cell proliferation induced by Opisthorchis viverrini excretory/secretory product.

PubMed

Thuwajit, Chanitra; Thuwajit, Peti; Uchida, Kazuhiko; Daorueang, Daoyot; Kaewkes, Sasithorn; Wongkham, Sopit; Miwa, Masanao

2006-06-14

To investigate the mechanism of fibroblast cell proliferation stimulated by the Opisthorchis viverrini excretory/secretory (ES) product. NIH-3T3, mouse fibroblast cells were treated with O. viverrini ES product by non-contact co-cultured with the adult parasites. Total RNA from NIH-3T3 treated and untreated with O. viverrini was extracted, reverse transcribed and hybridized with the mouse 15K complementary DNA (cDNA) array. The result was analyzed by ArrayVision version 5 and GeneSpring version 5 softwares. After normalization, the ratios of gene expression of parasite treated to untreated NIH-3T3 cells of 2-and more-fold upregulated was defined as the differentially expressed genes. The expression levels of the signal transduction genes were validated by semi-quantitative SYBR-based real-time RT-PCR. Among a total of 15,000 genes/ESTs, 239 genes with established cell proliferation-related function were 2 fold-and more-up-regulated by O. viverrini ES product compared to those in cells without exposure to the parasitic product. These genes were classified into groups including energy and metabolism, signal transduction, protein synthesis and translation, matrix and structural protein, transcription control, cell cycle and DNA replication. Moreover, the expressions of serine-threonine kinase receptor, receptor tyrosine kinase and collagen production-related genes were up-regulated by O. viverrini ES product. The expression level of signal transduction genes; pkC, pdgfr alpha, jak 1, eps 8, tgf beta 1i4, strap and h ras measured by real-time RT-PCR confirmed their expression levels to those obtained from cDNA array. However, only the up-regulated expression of pkC, eps 8 and tgfbeta 1i4 which are the downstream signaling molecules of either epidermal growth factor (EGF) or transforming growth factor-beta (TGF-beta) showed statistical significance (P < 0.05). O. viverrini ES product stimulates the significant changes of gene expression in several functional categories and these mainly include transcripts related to cell proliferation. The TGF-beta and EGF signal transduction pathways are indicated as the possible pathways of O. viverrini-driven cell proliferation.

Modular assembly of transposable element arrays by microsatellite targeting in the guayule and rice genomes.

PubMed

Valdes Franco, José A; Wang, Yi; Huo, Naxin; Ponciano, Grisel; Colvin, Howard A; McMahan, Colleen M; Gu, Yong Q; Belknap, William R

2018-04-19

Guayule (Parthenium argentatum A. Gray) is a rubber-producing desert shrub native to Mexico and the United States. Guayule represents an alternative to Hevea brasiliensis as a source for commercial natural rubber. The efficient application of modern molecular/genetic tools to guayule improvement requires characterization of its genome. The 1.6 Gb guayule genome was sequenced, assembled and annotated. The final 1.5 Gb assembly, while fragmented (N 50  = 22 kb), maps > 95% of the shotgun reads and is essentially complete. Approximately 40,000 transcribed, protein encoding genes were annotated on the assembly. Further characterization of this genome revealed 15 families of small, microsatellite-associated, transposable elements (TEs) with unexpected chromosomal distribution profiles. These SaTar (Satellite Targeted) elements, which are non-autonomous Mu-like elements (MULEs), were frequently observed in multimeric linear arrays of unrelated individual elements within which no individual element is interrupted by another. This uniformly non-nested TE multimer architecture has not been previously described in either eukaryotic or prokaryotic genomes. Five families of similarly distributed non-autonomous MULEs (microsatellite associated, modularly assembled) were characterized in the rice genome. Families of TEs with similar structures and distribution profiles were identified in sorghum and citrus. The sequencing and assembly of the guayule genome provides a foundation for application of current crop improvement technologies to this plant. In addition, characterization of this genome revealed SaTar elements with distribution profiles unique among TEs. Satar targeting appears based on an alternative MULE recombination mechanism with the potential to impact gene evolution.

Emerging Use of Gene Expression Microarrays in Plant Physiology

DOE PAGES

Wullschleger, Stan D.; Difazio, Stephen P.

2003-01-01

Microarrays have become an important technology for the global analysis of gene expression in humans, animals, plants, and microbes. Implemented in the context of a well-designed experiment, cDNA and oligonucleotide arrays can provide highthroughput, simultaneous analysis of transcript abundance for hundreds, if not thousands, of genes. However, despite widespread acceptance, the use of microarrays as a tool to better understand processes of interest to the plant physiologist is still being explored. To help illustrate current uses of microarrays in the plant sciences, several case studies that we believe demonstrate the emerging application of gene expression arrays in plant physiology weremore » selected from among the many posters and presentations at the 2003 Plant and Animal Genome XI Conference. Based on this survey, microarrays are being used to assess gene expression in plants exposed to the experimental manipulation of air temperature, soil water content and aluminium concentration in the root zone. Analysis often includes characterizing transcript profiles for multiple post-treatment sampling periods and categorizing genes with common patterns of response using hierarchical clustering techniques. In addition, microarrays are also providing insights into developmental changes in gene expression associated with fibre and root elongation in cotton and maize, respectively. Technical and analytical limitations of microarrays are discussed and projects attempting to advance areas of microarray design and data analysis are highlighted. Finally, although much work remains, we conclude that microarrays are a valuable tool for the plant physiologist interested in the characterization and identification of individual genes and gene families with potential application in the fields of agriculture, horticulture and forestry.« less

Expression profiling of the solute carrier gene family in chicken intestine from the late embryonic to early post-hatch stages.

PubMed

Li, H; Gilbert, E R; Zhang, Y; Crasta, O; Emmerson, D; Webb, K E; Wong, E A

2008-08-01

Intestinal development during late embryogenesis and early post-hatch has a long-term influence on digestive and absorptive capacity in chickens. The objective of this research was to obtain a global view of intestinal solute carrier (SLC) gene family member expression from late embryogenesis until 2 weeks post-hatch with a focus on SLC genes involved in uptake of sugars and amino acids. Small intestine samples from male chicks were collected on embryonic days 18 (E18) and 20 (E20), day of hatch and days 1, 3, 7 and 14 post-hatch. The expression profiles of 162 SLC genes belonging to 41 SLC families were determined using Affymetrix chicken genome microarrays. The majority of SLC genes showed little or no difference in level of expression during E18-D14. A number of well-known intestinal transporters were upregulated between E18 and D14 including the amino acid transporters rBAT, y(+)LAT-2 and EAAT3, the peptide transporter PepT1 and the sugar transporters SGLT1, GLUT2 and GLUT5. The amino acid transporters CAT-1 and CAT-2 were downregulated. In addition, several glucose and amino acid transporters that are novel to our understanding of nutrient absorption in the chicken intestine were discovered through the arrays (SGLT6, SNAT1, SNAT2 and AST). These results represent a comprehensive characterization of the expression profiles of the SLC family of genes at different stages of development in the chicken intestine and lay the ground work for future nutritional studies.

Male reproductive development: gene expression profiling of maize anther and pollen ontogeny

PubMed Central

Ma, Jiong; Skibbe, David S; Fernandes, John; Walbot, Virginia

2008-01-01

Background During flowering, central anther cells switch from mitosis to meiosis, ultimately forming pollen containing haploid sperm. Four rings of surrounding somatic cells differentiate to support first meiosis and later pollen dispersal. Synchronous development of many anthers per tassel and within each anther facilitates dissection of carefully staged maize anthers for transcriptome profiling. Results Global gene expression profiles of 7 stages representing 29 days of anther development are analyzed using a 44 K oligonucleotide array querying approximately 80% of maize protein-coding genes. Mature haploid pollen containing just two cell types expresses 10,000 transcripts. Anthers contain 5 major cell types and express >24,000 transcript types: each anther stage expresses approximately 10,000 constitutive and approximately 10,000 or more transcripts restricted to one or a few stages. The lowest complexity is present during meiosis. Large suites of stage-specific and co-expressed genes are identified through Gene Ontology and clustering analyses as functional classes for pre-meiotic, meiotic, and post-meiotic anther development. MADS box and zinc finger transcription factors with constitutive and stage-limited expression are identified. Conclusions We propose that the extensive gene expression of anther cells and pollen represents the key test of maize genome fitness, permitting strong selection against deleterious alleles in diploid anthers and haploid pollen. Because flowering plants show a substantial bias for male-sterile compared to female-sterile mutations, we propose that this fitness test is general. Because both somatic and germinal cells are transcriptionally quiescent during meiosis, we hypothesize that successful completion of meiosis is required to trigger maturation of anther somatic cells. PMID:19099579

EFEMP1 as a novel DNA methylation marker for prostate cancer: array-based DNA methylation and expression profiling.

PubMed

Kim, Yong-June; Yoon, Hyung-Yoon; Kim, Seon-Kyu; Kim, Young-Won; Kim, Eun-Jung; Kim, Isaac Yi; Kim, Wun-Jae

2011-07-01

Abnormal DNA methylation is associated with many human cancers. The aim of the present study was to identify novel methylation markers in prostate cancer (PCa) by microarray analysis and to test whether these markers could discriminate normal and PCa cells. Microarray-based DNA methylation and gene expression profiling was carried out using a panel of PCa cell lines and a control normal prostate cell line. The methylation status of candidate genes in prostate cell lines was confirmed by real-time reverse transcriptase-PCR, bisulfite sequencing analysis, and treatment with a demethylation agent. DNA methylation and gene expression analysis in 203 human prostate specimens, including 106 PCa and 97 benign prostate hyperplasia (BPH), were carried out. Further validation using microarray gene expression data from the Gene Expression Omnibus (GEO) was carried out. Epidermal growth factor-containing fibulin-like extracellular matrix protein 1 (EFEMP1) was identified as a lead candidate methylation marker for PCa. The gene expression level of EFEMP1 was significantly higher in tissue samples from patients with BPH than in those with PCa (P < 0.001). The sensitivity and specificity of EFEMP1 methylation status in discriminating between PCa and BPH reached 95.3% (101 of 106) and 86.6% (84 of 97), respectively. From the GEO data set, we confirmed that the expression level of EFEMP1 was significantly different between PCa and BPH. Genome-wide characterization of DNA methylation profiles enabled the identification of EFEMP1 aberrant methylation patterns in PCa. EFEMP1 might be a useful indicator for the detection of PCa.

Transcriptional profiling of Medicago truncatula meristematic root cells

PubMed Central

Holmes, Peta; Goffard, Nicolas; Weiller, Georg F; Rolfe, Barry G; Imin, Nijat

2008-01-01

Background The root apical meristem of crop and model legume Medicago truncatula is a significantly different stem cell system to that of the widely studied model plant species Arabidopsis thaliana. In this study we used the Affymetrix Medicago GeneChip® to compare the transcriptomes of meristem and non-meristematic root to identify root meristem specific candidate genes. Results Using mRNA from root meristem and non-meristem we were able to identify 324 and 363 transcripts differentially expressed from the two regions. With bioinformatics tools developed to functionally annotate the Medicago genome array we could identify significant changes in metabolism, signalling and the differentially expression of 55 transcription factors in meristematic and non-meristematic roots. Conclusion This is the first comprehensive analysis of M. truncatula root meristem cells using this genome array. This data will facilitate the mapping of regulatory and metabolic networks involved in the open root meristem of M. truncatula and provides candidates for functional analysis. PMID:18302802

An integrated workflow for analysis of ChIP-chip data.

PubMed

Weigelt, Karin; Moehle, Christoph; Stempfl, Thomas; Weber, Bernhard; Langmann, Thomas

2008-08-01

Although ChIP-chip is a powerful tool for genome-wide discovery of transcription factor target genes, the steps involving raw data analysis, identification of promoters, and correlation with binding sites are still laborious processes. Therefore, we report an integrated workflow for the analysis of promoter tiling arrays with the Genomatix ChipInspector system. We compare this tool with open-source software packages to identify PU.1 regulated genes in mouse macrophages. Our results suggest that ChipInspector data analysis, comparative genomics for binding site prediction, and pathway/network modeling significantly facilitate and enhance whole-genome promoter profiling to reveal in vivo sites of transcription factor-DNA interactions.

Human Adaptation Genetic Response Suites: Toward New Interventions and Countermeasures for Spaceflight

NASA Technical Reports Server (NTRS)

Sundaresan, A.; Pellis, N. R.

2005-01-01

Genetic response suites in human lymphocytes in response to microgravity are important to identify and further study in order to augment human physiological adaptation to novel environments. Emerging technologies, such as DNA micro array profiling, have the potential to identify novel genes that are involved in mediating adaptation to these environments. These genes may prove to be therapeutically valuable as new targets for countermeasures, or as predictive biomarkers of response to these new environments. Human lymphocytes cultured in lg and microgravity analog culture were analyzed for their differential gene expression response. Different groups of genes related to the immune response, cardiovascular system and stress response were then analyzed. Analysis of cells from multiple donors reveals a small shared set that are likely to be essential to adaptation. These three groups focus on human adaptation to new environments. The shared set contains genes related to T cell activation, immune response and stress response to analog microgravity.

THE USE OF GENE ARRAYS TO DETERMINE TEMPORAL GENE INDUCTION IN SHEEPSHEAD MINNOWS EXPOSED TO E2

EPA Science Inventory

Gene arrays provide a means to study differential gene expression in fish exposed to environmental estrogens by providing a "snapshot" of the genes expressed at a given time. Such array data may also uncover previously unknown biochemical pathways affected by estrogenic compounds...

Identification and validation of biomarkers of IgV(H) mutation status in chronic lymphocytic leukemia using microfluidics quantitative real-time polymerase chain reaction technology.

PubMed

Abruzzo, Lynne V; Barron, Lynn L; Anderson, Keith; Newman, Rachel J; Wierda, William G; O'brien, Susan; Ferrajoli, Alessandra; Luthra, Madan; Talwalkar, Sameer; Luthra, Rajyalakshmi; Jones, Dan; Keating, Michael J; Coombes, Kevin R

2007-09-01

To develop a model incorporating relevant prognostic biomarkers for untreated chronic lymphocytic leukemia patients, we re-analyzed the raw data from four published gene expression profiling studies. We selected 88 candidate biomarkers linked to immunoglobulin heavy-chain variable region gene (IgV(H)) mutation status and produced a reliable and reproducible microfluidics quantitative real-time polymerase chain reaction array. We applied this array to a training set of 29 purified samples from previously untreated patients. In an unsupervised analysis, the samples clustered into two groups. Using a cutoff point of 2% homology to the germline IgV(H) sequence, one group contained all 14 IgV(H)-unmutated samples; the other contained all 15 mutated samples. We confirmed the differential expression of 37 of the candidate biomarkers using two-sample t-tests. Next, we constructed 16 different models to predict IgV(H) mutation status and evaluated their performance on an independent test set of 20 new samples. Nine models correctly classified 11 of 11 IgV(H)-mutated cases and eight of nine IgV(H)-unmutated cases, with some models using three to seven genes. Thus, we can classify cases with 95% accuracy based on the expression of as few as three genes.

Gingival transcriptome patterns during induction and resolution of experimental gingivitis in humans.

PubMed

Offenbacher, Steven; Barros, Silvana P; Paquette, David W; Winston, J Leslie; Biesbrock, Aaron R; Thomason, Ryan G; Gibb, Roger D; Fulmer, Andy W; Tiesman, Jay P; Juhlin, Kenton D; Wang, Shuo L; Reichling, Tim D; Chen, Ker-Sang; Ho, Begonia

2009-12-01

To our knowledge, changes in the patterns of whole-transcriptome gene expression that occur during the induction and resolution of experimental gingivitis in humans were not previously explored using bioinformatic tools. Gingival biopsy samples collected from 14 subjects during a 28-day stent-induced experimental gingivitis model, followed by treatment, and resolution at days 28 through 35 were analyzed using gene-expression arrays. Biopsy samples were collected at different sites within each subject at baseline (day 0), at the peak of gingivitis (day 28), and at resolution (day 35) and processed using whole-transcriptome gene-expression arrays. Gene-expression data were analyzed to identify biologic themes and pathways associated with changes in gene-expression profiles that occur during the induction and resolution of experimental gingivitis using bioinformatic tools. During disease induction and resolution, the dominant expression pathway was the immune response, with 131 immune response genes significantly up- or downregulated during induction, during resolution, or during both at P <0.05. During induction, there was significant transient increase in the expression of inflammatory and oxidative stress mediators, including interleukin (IL)-1 alpha (IL1A), IL-1 beta (IL1B), IL8, RANTES, colony stimulating factor 3 (CSF3), and superoxide dismutase 2 (SOD2), and a decreased expression of IP10, interferon inducible T-cell alpha chemoattractant (ITAC), matrix metalloproteinase 10 (MMP10), and beta 4 defensin (DEFB4). These genes reversed expression patterns upon resolution in parallel with the reversal of gingival inflammation. A relatively small subset (11.9%) of the immune response genes analyzed by array was transiently activated in response to biofilm overgrowth, suggesting a degree of specificity in the transcriptome-expression response. The fact that this same subset demonstrates a reversal in expression patterns during clinical resolution implicates these genes as being critical for maintaining tissue homeostasis at the biofilm-gingival interface. In addition to the immune response pathway as the dominant response theme, new candidate genes and pathways were identified as being selectively modulated in experimental gingivitis, including neural processes, epithelial defenses, angiogenesis, and wound healing.

Branches of the NF-κB signaling pathway regulate proliferation of oval cells in rat liver regeneration.

PubMed

Zhao, W M; Qin, Y L; Niu, Z P; Chang, C F; Yang, J; Li, M H; Zhou, Y; Xu, C S

2016-03-24

The NF-kB (nuclear factor kB) pathway is involved in the proliferation of many cell types. To explore the mechanism of the NF-kB signaling pathway underlying the oval cell proliferation during rat liver regeneration, the Rat Genome 230 2.0 Array was used to detect expression changes of NF-kB signaling pathway-related genes in oval cells. The results revealed that the expression levels of many genes in the NF-kB pathway were significantly changed. This included 48 known genes and 16 homologous genes, as well as 370 genes and 85 homologous genes related to cell proliferation. To further understand the biological significance of these changes, an expression profile function was used to analyze the potential biological processes. The results showed that the NF-kB pathway promoted oval cell proliferation mainly through three signaling branches; the tumor necrosis factor alpha branch (TNF-a pathway), the growth factor branch, and the chemokine branch. An integrated statistics method was used to define the key genes in the NF-kB pathway. Seven genes were identified to play vital roles in the NF-kB pathway. To confirm these results, the protein content, including two key genes (TNF and FGF11) and two non-key genes (CCL2 and TNFRSF12A), were analyzed using two-dimensional gel electrophoresis and MALDI-TOF/TOF mass spectrometry. The results were generally consistent with those of the array data. To conclude, three branches and seven key genes were involved in the NF-kB signaling pathway that regulates oval cell proliferation during rat liver regeneration.

Insights into the Sigma-1 receptor chaperone’s cellular functions: a microarray report

PubMed Central

Tsai, Shang-Yi; Rothman, Richard Kyle; Su, Tsung-Ping

2013-01-01

We previously demonstrated that Sig-1Rs are critical regulators in neuronal morphogenesis and development via the regulation of oxidative stress and mitochondrial functions. In the present study, we sought to identify pathways and genes that are affected by Sig-1R. Gene expression profiles were examined in rat hippocampal neurons that had been cultured for18 days in vitro (DIV). The cells were transduced with AAV siRNA targeting Sig-1R on DIV 10 for 7 days, followed by gene expression analysis using a rat genome cDNA array. The gene array results indicated that Sig-1R knockdown hampered cellular functions including steroid biogenesis, protein ubiquitination, actin cytoskeleton network, and Nrf-2 mediated oxidative stress. Many of the cellular components important for actin polymerization and synapse plasticity, including F-actin capping protein and neurofilaments, were significantly changed in AAV-siSig-1R neurons. Further, cytochrome c was reduced in AAV-Sig-1R neurons whereas free-radical generating enzymes including cytochrome p450 and cytochrome b-245 were increased. The microarray results also suggest that Sig-1Rs may regulate genes that are involved in the pathogenesis of many CNS diseases including Alzheimer’s disease and Parkinson’s disease. These data further confirmed that Sig-1Rs play critical roles in the CNS and thus these findings may aid in future development of therapeutic treatments targeting neurodegenerative disorders. PMID:21905129

A Sorghum bicolor expression atlas reveals dynamic genotype-specific expression profiles for vegetative tissues of grain, sweet and bioenergy sorghums

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shakoor, N; Nair, R; Crasta, O

2014-01-23

Background: Effective improvement in sorghum crop development necessitates a genomics-based approach to identify functional genes and QTLs. Sequenced in 2009, a comprehensive annotation of the sorghum genome and the development of functional genomics resources is key to enable the discovery and deployment of regulatory and metabolic genes and gene networks for crop improvement. Results: This study utilizes the first commercially available whole-transcriptome sorghum microarray (Sorgh-WTa520972F) to identify tissue and genotype-specific expression patterns for all identified Sorghum bicolor exons and UTRs. The genechip contains 1,026,373 probes covering 149,182 exons (27,577 genes) across the Sorghum bicolor nuclear, chloroplast, and mitochondrial genomes. Specificmore » probesets were also included for putative non-coding RNAs that may play a role in gene regulation (e. g., microRNAs), and confirmed functional small RNAs in related species (maize and sugarcane) were also included in our array design. We generated expression data for 78 samples with a combination of four different tissue types (shoot, root, leaf and stem), two dissected stem tissues (pith and rind) and six diverse genotypes, which included 6 public sorghum lines (R159, Atlas, Fremont, PI152611, AR2400 and PI455230) representing grain, sweet, forage, and high biomass ideotypes. Conclusions: Here we present a summary of the microarray dataset, including analysis of tissue-specific gene expression profiles and associated expression profiles of relevant metabolic pathways. With an aim to enable identification and functional characterization of genes in sorghum, this expression atlas presents a new and valuable resource to the research community.« less

Transcriptional Profile of Brain Injury in Hypothermic Circulatory Arrest and Cardiopulmonary Bypass

PubMed Central

Allen, Jeremiah G.; Weiss, Eric S.; Wilson, Mary Ann; Arnaoutakis, George J.; Blue, Mary E.; Talbot, C. Conover; Jie, Chunfa; Lange, Mary S.; Troncoso, Juan C.; Johnston, Michael V.; Baumgartner, William A.

2011-01-01

Background Little is known about the molecular mechanisms of neurologic complications after hypothermic circulatory arrest (HCA) with cardiopulmonary bypass (CPB). Canine genome sequencing allows profiling of genomic changes after HCA and CPB alone. We hypothesize that gene regulation will increase with increased severity of injury. Methods Dogs underwent 2-hour HCA at 18°C (n = 10), 1-hour HCA (n = 8), or 2-hour CPB at 32°C alone (n = 8). In each group, half were sacrificed at 8 hours and half at 24 hours after treatment. After neurologic scoring, brains were harvested for genomic analysis. Hippocampal RNA isolates were analyzed using canine oligonucleotide expression arrays containing 42,028 probes. Results Consistent with prior work, dogs that underwent 2-hour HCA experienced severe neurologic injury. One hour of HCA caused intermediate clinical damage. Cardiopulmonary bypass alone yielded normal clinical scores. Cardiopulmonary bypass, 1-hour HCA, and 2-hour HCA groups historically demonstrated increasing degrees of histopathologic damage (previously published). Exploratory analysis revealed differences in significantly regulated genes (false discovery rate < 10%, absolute fold change ≥ 1.2), with increases in differential gene expression with injury severity. At 8 hours and 24 hours after insult, 2-hour HCA dogs had 502 and 1,057 genes regulated, respectively; 1-hour HCA dogs had 179 and 56 genes regulated; and CPB alone dogs had 5 and 0 genes regulated. Conclusions Our genomic profile of canine brains after HCA and CPB revealed 1-hour and 2-hour HCA induced markedly increased gene regulation, in contrast to the minimal effect of CPB alone. This adds to the body of neurologic literature supporting the safety of CPB alone and the minimal effect of CPB on a normal brain, while illuminating genomic results of both. PMID:20494057

The genomic response of a human uterine endometrial adenocarcinoma cell line to 17alpha-ethynyl estradiol.

PubMed

Naciff, Jorge M; Khambatta, Zubin S; Thomason, Ryan G; Carr, Gregory J; Tiesman, Jay P; Singleton, David W; Khan, Sohaib A; Daston, George P

2009-01-01

We have determined the gene expression profile induced by 17 alpha-ethynyl estradiol (EE) in Ishikawa cells, a human uterine-derived estrogen-sensitive cell line, at various doses (1 pM, 100 pM, 10 nM, and 1 microM) and time points (8, 24, and 48 h). The transcript profiles were compared between treatment groups and controls (vehicle-treated) using high-density oligonucleotide arrays to determine the expression level of approximately 38,500 human genes. By trend analysis, we determined that the expression of 2560 genes was modified by exposure to EE in a dose- and time-dependent manner (p

Gene expression profile of patients with Mayer-Rokitansky-Küster-Hauser syndrome: new insights into the potential role of developmental pathways.

PubMed

Nodale, Cristina; Ceccarelli, Simona; Giuliano, Mariateresa; Cammarota, Marcella; D'Amici, Sirio; Vescarelli, Enrica; Maffucci, Diana; Bellati, Filippo; Panici, Pierluigi Benedetti; Romano, Ferdinando; Angeloni, Antonio; Marchese, Cinzia

2014-01-01

Mayer-Rokitansky-Küster-Hauser syndrome (MRKHS) is a rare disease characterized by congenital aplasia of uterus and vagina. Although many studies have investigated several candidate genes, up to now none of them seem to be responsible for the aetiology of the syndrome. In our study, we identified differences in gene expression profile of in vitro cultured vaginal tissue of MRHKS patients using whole-genome microarray analysis. A group of eight out of sixteen MRKHS patients that underwent reconstruction of neovagina with an autologous in vitro cultured vaginal tissue were subjected to microarray analysis and compared with five healthy controls. Results obtained by array were confirmed by qRT-PCR and further extended to other eight MRKHS patients. Gene profiling of MRKHS patients delineated 275 differentially expressed genes, of which 133 downregulated and 142 upregulated. We selected six deregulated genes (MUC1, HOXC8, HOXB2, HOXB5, JAG1 and DLL1) on the basis of their fold change, their differential expression in most patients and their relevant role in embryological development. All patients showed upregulation of MUC1, while HOXB2 and HOXB5 were downregulated, as well as Notch ligands JAG1 and DLL1 in the majority of them. Interestingly, HOXC8 was significantly upregulated in 47% of patients, with a differential expression only in MRKHS type I patients. Taken together, our results highlighted the dysregulation of developmental genes, thus suggesting a potential alteration of networks involved in the formation of the female reproductive tract and providing a useful clue for understanding the pathophysiology of MRKHS.

Gene Expression Profile of Patients with Mayer-Rokitansky-Küster-Hauser Syndrome: New Insights into the Potential Role of Developmental Pathways

PubMed Central

Giuliano, Mariateresa; Cammarota, Marcella; D’Amici, Sirio; Vescarelli, Enrica; Maffucci, Diana; Bellati, Filippo; Panici, Pierluigi Benedetti; Romano, Ferdinando; Angeloni, Antonio; Marchese, Cinzia

2014-01-01

Mayer-Rokitansky-Küster-Hauser syndrome (MRKHS) is a rare disease characterized by congenital aplasia of uterus and vagina. Although many studies have investigated several candidate genes, up to now none of them seem to be responsible for the aetiology of the syndrome. In our study, we identified differences in gene expression profile of in vitro cultured vaginal tissue of MRHKS patients using whole-genome microarray analysis. A group of eight out of sixteen MRKHS patients that underwent reconstruction of neovagina with an autologous in vitro cultured vaginal tissue were subjected to microarray analysis and compared with five healthy controls. Results obtained by array were confirmed by qRT-PCR and further extended to other eight MRKHS patients. Gene profiling of MRKHS patients delineated 275 differentially expressed genes, of which 133 downregulated and 142 upregulated. We selected six deregulated genes (MUC1, HOXC8, HOXB2, HOXB5, JAG1 and DLL1) on the basis of their fold change, their differential expression in most patients and their relevant role in embryological development. All patients showed upregulation of MUC1, while HOXB2 and HOXB5 were downregulated, as well as Notch ligands JAG1 and DLL1 in the majority of them. Interestingly, HOXC8 was significantly upregulated in 47% of patients, with a differential expression only in MRKHS type I patients. Taken together, our results highlighted the dysregulation of developmental genes, thus suggesting a potential alteration of networks involved in the formation of the female reproductive tract and providing a useful clue for understanding the pathophysiology of MRKHS. PMID:24608967

Genomic profiling of 766 cancer-related genes in archived esophageal normal and carcinoma tissues.

PubMed

Chen, Jing; Guo, Liping; Peiffer, Daniel A; Zhou, Lixin; Chan, Owen Tsan Mo; Bibikova, Marina; Wickham-Garcia, Eliza; Lu, Shih-Hsin; Zhan, Qimin; Wang-Rodriguez, Jessica; Jiang, Wei; Fan, Jian-Bing

2008-05-15

We employed the BeadArraytrade mark technology to perform a genetic analysis in 33 formalin-fixed, paraffin-embedded (FFPE) human esophageal carcinomas, mostly squamous-cell-carcinoma (ESCC), and their adjacent normal tissues. A total of 1,432 single nucleotide polymorphisms (SNPs) derived from 766 cancer-related genes were genotyped with partially degraded genomic DNAs isolated from these samples. This directly targeted genomic profiling identified not only previously reported somatic gene amplifications (e.g., CCND1) and deletions (e.g., CDKN2A and CDKN2B) but also novel genomic aberrations. Among these novel targets, the most frequently deleted genomic regions were chromosome 3p (including tumor suppressor genes FANCD2 and CTNNB1) and chromosome 5 (including tumor suppressor gene APC). The most frequently amplified genomic region was chromosome 3q (containing DVL3, MLF1, ABCC5, BCL6, AGTR1 and known oncogenes TNK2, TNFSF10, FGF12). The chromosome 3p deletion and 3q amplification occurred coincidently in nearly all of the affected cases, suggesting a molecular mechanism for the generation of somatic chromosomal aberrations. We also detected significant differences in germline allele frequency between the esophageal cohort of our study and normal control samples from the International HapMap Project for 10 genes (CSF1, KIAA1804, IL2, PMS2, IRF7, FLT3, NTRK2, MAP3K9, ERBB2 and PRKAR1A), suggesting that they might play roles in esophageal cancer susceptibility and/or development. Taken together, our results demonstrated the utility of the BeadArray technology for high-throughput genetic analysis in FFPE tumor tissues and provided a detailed genetic profiling of cancer-related genes in human esophageal cancer. (c) 2008 Wiley-Liss, Inc.

Detection of growth hormone doping by gene expression profiling of peripheral blood.

PubMed

Mitchell, Christopher J; Nelson, Anne E; Cowley, Mark J; Kaplan, Warren; Stone, Glenn; Sutton, Selina K; Lau, Amie; Lee, Carol M Y; Ho, Ken K Y

2009-12-01

GH abuse is a significant problem in many sports, and there is currently no robust test that allows detection of doping beyond a short window after administration. Our objective was to evaluate gene expression profiling in peripheral blood leukocytes in-vivo as a test for GH doping in humans. Seven men and thirteen women were administered GH, 2 mg/d sc for 8 wk. Blood was collected at baseline and at 8 wk. RNA was extracted from the white cell fraction. Microarray analysis was undertaken using Agilent 44K G4112F arrays using a two-color design. Quantitative RT-PCR using TaqMan gene expression assays was performed for validation of selected differentially expressed genes. GH induced an approximately 2-fold increase in circulating IGF-I that was maintained throughout the 8 wk of the study. GH induced significant changes in gene expression with 353 in women and 41 in men detected with a false discovery rate of less than 5%. None of the differentially expressed genes were common between men and women. The maximal changes were a doubling for up-regulated or halving for down-regulated genes, similar in magnitude to the variation between individuals. Quantitative RT-PCR for seven target genes showed good concordance between microarray and quantitative PCR data in women but not in men. Gene expression analysis of peripheral blood leukocytes is unlikely to be a viable approach for the detection of GH doping.

Epigenetic regulation of depot-specific gene expression in adipose tissue.

PubMed

Gehrke, Sandra; Brueckner, Bodo; Schepky, Andreas; Klein, Johannes; Iwen, Alexander; Bosch, Thomas C G; Wenck, Horst; Winnefeld, Marc; Hagemann, Sabine

2013-01-01

In humans, adipose tissue is distributed in subcutaneous abdominal and subcutaneous gluteal depots that comprise a variety of functional differences. Whereas energy storage in gluteal adipose tissue has been shown to mediate a protective effect, an increase of abdominal adipose tissue is associated with metabolic disorders. However, the molecular basis of depot-specific characteristics is not completely understood yet. Using array-based analyses of transcription profiles, we identified a specific set of genes that was differentially expressed between subcutaneous abdominal and gluteal adipose tissue. To investigate the role of epigenetic regulation in depot-specific gene expression, we additionally analyzed genome-wide DNA methylation patterns in abdominal and gluteal depots. By combining both data sets, we identified a highly significant set of depot-specifically expressed genes that appear to be epigenetically regulated. Interestingly, the majority of these genes form part of the homeobox gene family. Moreover, genes involved in fatty acid metabolism were also differentially expressed. Therefore we suppose that changes in gene expression profiles might account for depot-specific differences in lipid composition. Indeed, triglycerides and fatty acids of abdominal adipose tissue were more saturated compared to triglycerides and fatty acids in gluteal adipose tissue. Taken together, our results uncover clear differences between abdominal and gluteal adipose tissue on the gene expression and DNA methylation level as well as in fatty acid composition. Therefore, a detailed molecular characterization of adipose tissue depots will be essential to develop new treatment strategies for metabolic syndrome associated complications.

Novel POC1A mutation in primordial dwarfism reveals new insights for centriole biogenesis.

PubMed

Koparir, Asuman; Karatas, Omer F; Yuceturk, Betul; Yuksel, Bayram; Bayrak, Ali O; Gerdan, Omer F; Sagiroglu, Mahmut S; Gezdirici, Alper; Kirimtay, Koray; Selcuk, Ece; Karabay, Arzu; Creighton, Chad J; Yuksel, Adnan; Ozen, Mustafa

2015-10-01

POC1A encodes a WD repeat protein localizing to centrioles and spindle poles and is associated with short stature, onychodysplasia, facial dysmorphism and hypotrichosis (SOFT) syndrome. These main features are related to the defect in cell proliferation of chondrocytes in growth plate. In the current study, we aimed at identifying the molecular basis of two patients with primordial dwarfism (PD) in a single family through utilization of whole-exome sequencing. A novel homozygous p.T120A missense mutation was detected in POC1A in both patients, a known causative gene of SOFT syndrome, and confirmed using Sanger sequencing. To test the pathogenicity of the detected mutation, primary fibroblast cultures obtained from the patients and a control individual were used. For evaluating the global gene expression profile of cells carrying p.T120A mutation in POC1A, we performed the gene expression array and compared their expression profiles to those of control fibroblast cells. The gene expression array analysis showed that 4800 transcript probes were significantly deregulated in cells with p.T120A mutation in comparison to the control. GO term association results showed that deregulated genes are mostly involved in the extracellular matrix and cytoskeleton. Furthermore, the p.T120A missense mutation in POC1A caused the formation of abnormal mitotic spindle structure, including supernumerary centrosomes, and changes in POC1A were accompanied by alterations in another centrosome-associated WD repeat protein p80-katanin. As a result, we identified a novel mutation in POC1A of patients with PD and showed that this mutation causes the formation of multiple numbers of centrioles and multipolar spindles with abnormal chromosome arrangement. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Differential Expression of CHL1 Gene during Development of Major Human Cancers

PubMed Central

Senchenko, Vera N.; Krasnov, George S.; Dmitriev, Alexey A.; Kudryavtseva, Anna V.; Anedchenko, Ekaterina A.; Braga, Eleonora A.; Pronina, Irina V.; Kondratieva, Tatiana T.; Ivanov, Sergey V.; Zabarovsky, Eugene R.; Lerman, Michael I.

2011-01-01

Background CHL1 gene (also known as CALL) on 3p26.3 encodes a one-pass trans-membrane cell adhesion molecule (CAM). Previously CAMs of this type, including L1, were shown to be involved in cancer growth and metastasis. Methodology/Principal Findings We used Clontech Cancer Profiling Arrays (19 different types of cancers, 395 samples) to analyze expression of the CHL1 gene. The results were further validated by RT-qPCR for breast, renal and lung cancer. Cancer Profiling Arrays revealed differential expression of the gene: down-regulation/silencing in a majority of primary tumors and up-regulation associated with invasive/metastatic growth. Frequent down-regulation (>40% of cases) was detected in 11 types of cancer (breast, kidney, rectum, colon, thyroid, stomach, skin, small intestine, bladder, vulva and pancreatic cancer) and frequent up-regulation (>40% of cases) – in 5 types (lung, ovary, uterus, liver and trachea) of cancer. Using real-time quantitative PCR (RT-qPCR) we found that CHL1 expression was decreased in 61% of breast, 60% of lung, 87% of clear cell and 89% papillary renal cancer specimens (P<0.03 for all the cases). There was a higher frequency of CHL1 mRNA decrease in lung squamous cell carcinoma compared to adenocarcinoma (81% vs. 38%, P = 0.02) without association with tumor progression. Conclusions/Significance Our results suggested that CHL1 is involved in the development of different human cancers. Initially, during the primary tumor growth CHL1 could act as a putative tumor suppressor and is silenced to facilitate in situ tumor growth for 11 cancer types. We also suggested that re-expression of the gene on the edge of tumor mass might promote local invasive growth and enable further metastatic spread in ovary, colon and breast cancer. Our data also supported the role of CHL1 as a potentially novel specific biomarker in the early pathogenesis of two major histological types of renal cancer. PMID:21408220

Transcript and metabolite analysis in Trincadeira cultivar reveals novel information regarding the dynamics of grape ripening.

PubMed

Fortes, Ana M; Agudelo-Romero, Patricia; Silva, Marta S; Ali, Kashif; Sousa, Lisete; Maltese, Federica; Choi, Young H; Grimplet, Jerome; Martinez-Zapater, José M; Verpoorte, Robert; Pais, Maria S

2011-11-02

Grapes (Vitis vinifera L.) are economically the most important fruit crop worldwide. However, the complexity of molecular and biochemical events that lead to the onset of ripening of nonclimacteric fruits is not fully understood which is further complicated in grapes due to seasonal and cultivar specific variation. The Portuguese wine variety Trincadeira gives rise to high quality wines but presents extremely irregular berry ripening among seasons probably due to high susceptibility to abiotic and biotic stresses. Ripening of Trincadeira grapes was studied taking into account the transcriptional and metabolic profilings complemented with biochemical data. The mRNA expression profiles of four time points spanning developmental stages from pea size green berries, through véraison and mature berries (EL 32, EL 34, EL 35 and EL 36) and in two seasons (2007 and 2008) were compared using the Affymetrix GrapeGen® genome array containing 23096 probesets corresponding to 18726 unique sequences. Over 50% of these probesets were significantly differentially expressed (1.5 fold) between at least two developmental stages. A common set of modulated transcripts corresponding to 5877 unigenes indicates the activation of common pathways between years despite the irregular development of Trincadeira grapes. These unigenes were assigned to the functional categories of "metabolism", "development", "cellular process", "diverse/miscellanenous functions", "regulation overview", "response to stimulus, stress", "signaling", "transport overview", "xenoprotein, transposable element" and "unknown". Quantitative RT-PCR validated microarrays results being carried out for eight selected genes and five developmental stages (EL 32, EL 34, EL 35, EL 36 and EL 38). Metabolic profiling using 1H NMR spectroscopy associated to two-dimensional techniques showed the importance of metabolites related to oxidative stress response, amino acid and sugar metabolism as well as secondary metabolism. These results were integrated with transcriptional profiling obtained using genome array to provide new information regarding the network of events leading to grape ripening. Altogether the data obtained provides the most extensive survey obtained so far for gene expression and metabolites accumulated during grape ripening. Moreover, it highlighted information obtained in a poorly known variety exhibiting particular characteristics that may be cultivar specific or dependent upon climatic conditions. Several genes were identified that had not been previously reported in the context of grape ripening namely genes involved in carbohydrate and amino acid metabolisms as well as in growth regulators; metabolism, epigenetic factors and signaling pathways. Some of these genes were annotated as receptors, transcription factors, and kinases and constitute good candidates for functional analysis in order to establish a model for ripening control of a non-climacteric fruit.

EPConDB: a web resource for gene expression related to pancreatic development, beta-cell function and diabetes.

PubMed

Mazzarelli, Joan M; Brestelli, John; Gorski, Regina K; Liu, Junmin; Manduchi, Elisabetta; Pinney, Deborah F; Schug, Jonathan; White, Peter; Kaestner, Klaus H; Stoeckert, Christian J

2007-01-01

EPConDB (http://www.cbil.upenn.edu/EPConDB) is a public web site that supports research in diabetes, pancreatic development and beta-cell function by providing information about genes expressed in cells of the pancreas. EPConDB displays expression profiles for individual genes and information about transcripts, promoter elements and transcription factor binding sites. Gene expression results are obtained from studies examining tissue expression, pancreatic development and growth, differentiation of insulin-producing cells, islet or beta-cell injury, and genetic models of impaired beta-cell function. The expression datasets are derived using different microarray platforms, including the BCBC PancChips and Affymetrix gene expression arrays. Other datasets include semi-quantitative RT-PCR and MPSS expression studies. For selected microarray studies, lists of differentially expressed genes, derived from PaGE analysis, are displayed on the site. EPConDB provides database queries and tools to examine the relationship between a gene, its transcriptional regulation, protein function and expression in pancreatic tissues.

A gene expression signature of confinement in peripheral blood of red wolves (Canis rufus).

PubMed

Kennerly, Erin; Ballmann, Anne; Martin, Stanton; Wolfinger, Russ; Gregory, Simon; Stoskopf, Michael; Gibson, Greg

2008-06-01

The stresses that animals experience as a result of modification of their ecological circumstances induce physiological changes that leave a signature in profiles of gene expression. We illustrate this concept in a comparison of free range and confined North American red wolves (Canis rufus). Transcription profiling of peripheral blood samples from 13 red wolf individuals in the Alligator River region of North Carolina revealed a strong signal of differentiation. Four hundred eighty-two out of 2980 transcripts detected on Illumina HumanRef8 oligonucleotide bead arrays were found to differentiate free range and confined wolves at a false discovery rate of 12.8% and P < 0.05. Over-representation of genes in focal adhesion, insulin signalling, proteasomal, and tryptophan metabolism pathways suggests the activation of pro-inflammatory and stress responses in confined animals. Consequently, characterization of differential transcript abundance in an accessible tissue such as peripheral blood identifies biomarkers that could be useful in animal management practices and for evaluating the impact of habitat changes on population health, particularly as attention turns to the impact of climate change on physiology and in turn species distributions.

Plants, plant pathogens, and microgravity--a deadly trio.

PubMed

Leach, J E; Ryba-White, M; Sun, Q; Wu, C J; Hilaire, E; Gartner, C; Nedukha, O; Kordyum, E; Keck, M; Leung, H; Guikema, J A

2001-06-01

Plants grown in spaceflight conditions are more susceptible to colonization by plant pathogens. The underlying causes for this enhanced susceptibility are not known. Possibly the formation of structural barriers and the activation of plant defense response components are impaired in spaceflight conditions. Either condition would result from altered gene expression of the plant. Because of the tools available, past studies focused on a few physiological responses or biochemical pathways. With recent advances in genomics research, new tools, including microarray technologies, are available to examine the global impact of growth in the spacecraft on the plant's gene expression profile. In ground-based studies, we have developed cDNA subtraction libraries of rice that are enriched for genes induced during pathogen infection and the defense response. Arrays of these genes are being used to dissect plant defense response pathways in a model system involving wild-type rice plants and lesion mimic mutants. The lesion mimic mutants are ideal experimental tools because they erratically develop defense response-like lesions in the absence of pathogens. The gene expression profiles from these ground-based studies will provide the molecular basis for understanding the biochemical and physiological impacts of spaceflight on plant growth, development and disease defense responses. This, in turn, will allow the development of strategies to manage plant disease for life in the space environment.

The placental immune milieu is characterized by a Th2- and anti-inflammatory transcription profile, regardless of maternal allergy, and associates with neonatal immunity.

PubMed

Abelius, Martina S; Janefjord, Camilla; Ernerudh, Jan; Berg, Göran; Matthiesen, Leif; Duchén, Karel; Nilsson, Lennart J; Jenmalm, Maria C

2015-05-01

How maternal allergy affects the systemic and local immunological environment during pregnancy and the immune development of the offspring is unclear. Expression of 40 genes was quantified by PCR arrays in placenta, peripheral blood mononuclear cells (PBMC), and cord blood mononuclear cells (CBMC) from 7 allergic and 12 non-allergic women and their offspring. Placental gene expression was dominated by a Th2-/anti-inflammatory profile, irrespectively of maternal allergy, as compared to gene expression in PBMC. p35 expression in placenta correlated with fetal Tbx21 (ρ = -0.88, P < 0.001) and IL-5 expression in PBMC with fetal galectin1 (ρ = 0.91, P < 0.001). Increased expression of Th2-associated CCL22 in CBMC preceded allergy development. Gene expression locally and systemically during pregnancy was partly associated with the offspring's gene expression, possibly indicating that the immunological milieu is important for fetal immune development. Maternal allergy was not associated with an enhanced Th2 immunity in placenta or PBMC, while a marked prenatal Th2 skewing, shown as increased CCL22 mRNA expression, might contribute to postnatal allergy development. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Plants, plant pathogens, and microgravity--a deadly trio

NASA Technical Reports Server (NTRS)

Leach, J. E.; Ryba-White, M.; Sun, Q.; Wu, C. J.; Hilaire, E.; Gartner, C.; Nedukha, O.; Kordyum, E.; Keck, M.; Leung, H.;

Microarray Analysis Gene Expression Profiles in Laryngeal Muscle After Recurrent Laryngeal Nerve Injury.

PubMed

Bijangi-Vishehsaraei, Khadijeh; Blum, Kevin; Zhang, Hongji; Safa, Ahmad R; Halum, Stacey L

2016-03-01

The pathophysiology of recurrent laryngeal nerve (RLN) transection injury is rare in that it is characteristically followed by a high degree of spontaneous reinnervation, with reinnervation of the laryngeal adductor complex (AC) preceding that of the abducting posterior cricoarytenoid (PCA) muscle. Here, we aim to elucidate the differentially expressed myogenic factors following RLN injury that may be at least partially responsible for the spontaneous reinnervation. F344 male rats underwent RLN injury (n = 12) or sham surgery (n = 12). One week after RLN injury, larynges were harvested following euthanasia. The mRNA was extracted from PCA and AC muscles bilaterally, and microarray analysis was performed using a full rat genome array. Microarray analysis of denervated AC and PCA muscles demonstrated dramatic differences in gene expression profiles, with 205 individual probes that were differentially expressed between the denervated AC and PCA muscles and only 14 genes with similar expression patterns. The differential expression patterns of the AC and PCA suggest different mechanisms of reinnervation. The PCA showed the gene patterns of Wallerian degeneration, while the AC expressed the gene patterns of reinnervation by adjacent axonal sprouting. This finding may reveal important therapeutic targets applicable to RLN and other peripheral nerve injuries. © The Author(s) 2015.

Kidney Transplant Rejection and Tissue Injury by Gene Profiling of Biopsies and Peripheral Blood Lymphocytes

PubMed Central

Flechner, Stuart M.; Kurian, Sunil M.; Head, Steven R.; Sharp, Starlette M.; Whisenant, Thomas C.; Zhang, Jie; Chismar, Jeffrey D.; Horvath, Steve; Mondala, Tony; Gilmartin, Timothy; Cook, Daniel J.; Kay, Steven A.; Walker, John R.; Salomon, Daniel R.

2007-01-01

A major challenge for kidney transplantation is balancing the need for immunosuppression to prevent rejection, while minimizing drug-induced toxicities. We used DNA microarrays (HG-U95Av2 GeneChips, Affymetrix) to determine gene expression profiles for kidney biopsies and peripheral blood lymphocytes (PBLs) in transplant patients including normal donor kidneys, well-functioning transplants without rejection, kidneys undergoing acute rejection, and transplants with renal dysfunction without rejection. We developed a data analysis schema based on expression signal determination, class comparison and prediction, hierarchical clustering, statistical power analysis and real-time quantitative PCR validation. We identified distinct gene expression signatures for both biopsies and PBLs that correlated significantly with each of the different classes of transplant patients. This is the most complete report to date using commercial arrays to identify unique expression signatures in transplant biopsies distinguishing acute rejection, acute dysfunction without rejection and well-functioning transplants with no rejection history. We demonstrate for the first time the successful application of high density DNA chip analysis of PBL as a diagnostic tool for transplantation. The significance of these results, if validated in a multicenter prospective trial, would be the establishment of a metric based on gene expression signatures for monitoring the immune status and immunosuppression of transplanted patients. PMID:15307835

Target genes discovery through copy number alteration analysis in human hepatocellular carcinoma.

PubMed

Gu, De-Leung; Chen, Yen-Hsieh; Shih, Jou-Ho; Lin, Chi-Hung; Jou, Yuh-Shan; Chen, Chian-Feng

2013-12-21

High-throughput short-read sequencing of exomes and whole cancer genomes in multiple human hepatocellular carcinoma (HCC) cohorts confirmed previously identified frequently mutated somatic genes, such as TP53, CTNNB1 and AXIN1, and identified several novel genes with moderate mutation frequencies, including ARID1A, ARID2, MLL, MLL2, MLL3, MLL4, IRF2, ATM, CDKN2A, FGF19, PIK3CA, RPS6KA3, JAK1, KEAP1, NFE2L2, C16orf62, LEPR, RAC2, and IL6ST. Functional classification of these mutated genes suggested that alterations in pathways participating in chromatin remodeling, Wnt/β-catenin signaling, JAK/STAT signaling, and oxidative stress play critical roles in HCC tumorigenesis. Nevertheless, because there are few druggable genes used in HCC therapy, the identification of new therapeutic targets through integrated genomic approaches remains an important task. Because a large amount of HCC genomic data genotyped by high density single nucleotide polymorphism arrays is deposited in the public domain, copy number alteration (CNA) analyses of these arrays is a cost-effective way to reveal target genes through profiling of recurrent and overlapping amplicons, homozygous deletions and potentially unbalanced chromosomal translocations accumulated during HCC progression. Moreover, integration of CNAs with other high-throughput genomic data, such as aberrantly coding transcriptomes and non-coding gene expression in human HCC tissues and rodent HCC models, provides lines of evidence that can be used to facilitate the identification of novel HCC target genes with the potential of improving the survival of HCC patients.

MicroArray Facility: a laboratory information management system with extended support for Nylon based technologies.

PubMed

Honoré, Paul; Granjeaud, Samuel; Tagett, Rebecca; Deraco, Stéphane; Beaudoing, Emmanuel; Rougemont, Jacques; Debono, Stéphane; Hingamp, Pascal

2006-09-20

High throughput gene expression profiling (GEP) is becoming a routine technique in life science laboratories. With experimental designs that repeatedly span thousands of genes and hundreds of samples, relying on a dedicated database infrastructure is no longer an option.GEP technology is a fast moving target, with new approaches constantly broadening the field diversity. This technology heterogeneity, compounded by the informatics complexity of GEP databases, means that software developments have so far focused on mainstream techniques, leaving less typical yet established techniques such as Nylon microarrays at best partially supported. MAF (MicroArray Facility) is the laboratory database system we have developed for managing the design, production and hybridization of spotted microarrays. Although it can support the widely used glass microarrays and oligo-chips, MAF was designed with the specific idiosyncrasies of Nylon based microarrays in mind. Notably single channel radioactive probes, microarray stripping and reuse, vector control hybridizations and spike-in controls are all natively supported by the software suite. MicroArray Facility is MIAME supportive and dynamically provides feedback on missing annotations to help users estimate effective MIAME compliance. Genomic data such as clone identifiers and gene symbols are also directly annotated by MAF software using standard public resources. The MAGE-ML data format is implemented for full data export. Journalized database operations (audit tracking), data anonymization, material traceability and user/project level confidentiality policies are also managed by MAF. MicroArray Facility is a complete data management system for microarray producers and end-users. Particular care has been devoted to adequately model Nylon based microarrays. The MAF system, developed and implemented in both private and academic environments, has proved a robust solution for shared facilities and industry service providers alike.

MicroArray Facility: a laboratory information management system with extended support for Nylon based technologies

PubMed Central

Honoré, Paul; Granjeaud, Samuel; Tagett, Rebecca; Deraco, Stéphane; Beaudoing, Emmanuel; Rougemont, Jacques; Debono, Stéphane; Hingamp, Pascal

2006-01-01

Background High throughput gene expression profiling (GEP) is becoming a routine technique in life science laboratories. With experimental designs that repeatedly span thousands of genes and hundreds of samples, relying on a dedicated database infrastructure is no longer an option. GEP technology is a fast moving target, with new approaches constantly broadening the field diversity. This technology heterogeneity, compounded by the informatics complexity of GEP databases, means that software developments have so far focused on mainstream techniques, leaving less typical yet established techniques such as Nylon microarrays at best partially supported. Results MAF (MicroArray Facility) is the laboratory database system we have developed for managing the design, production and hybridization of spotted microarrays. Although it can support the widely used glass microarrays and oligo-chips, MAF was designed with the specific idiosyncrasies of Nylon based microarrays in mind. Notably single channel radioactive probes, microarray stripping and reuse, vector control hybridizations and spike-in controls are all natively supported by the software suite. MicroArray Facility is MIAME supportive and dynamically provides feedback on missing annotations to help users estimate effective MIAME compliance. Genomic data such as clone identifiers and gene symbols are also directly annotated by MAF software using standard public resources. The MAGE-ML data format is implemented for full data export. Journalized database operations (audit tracking), data anonymization, material traceability and user/project level confidentiality policies are also managed by MAF. Conclusion MicroArray Facility is a complete data management system for microarray producers and end-users. Particular care has been devoted to adequately model Nylon based microarrays. The MAF system, developed and implemented in both private and academic environments, has proved a robust solution for shared facilities and industry service providers alike. PMID:16987406

Spatial analysis of biomineralization associated gene expression from the mantle organ of the pearl oyster Pinctada maxima

PubMed Central

2011-01-01

Background Biomineralization is a process encompassing all mineral containing tissues produced within an organism. One of the most dynamic examples of this process is the formation of the mollusk shell, comprising a variety of crystal phases and microstructures. The organic component incorporated within the shell is said to dictate this architecture. However general understanding of how this process is achieved remains ambiguous. The mantle is a conserved organ involved in shell formation throughout molluscs. Specifically the mantle is thought to be responsible for secreting the protein component of the shell. This study employs molecular approaches to determine the spatial expression of genes within the mantle tissue to further the elucidation of the shell biomineralization. Results A microarray platform was custom generated (PmaxArray 1.0) from the pearl oyster Pinctada maxima. PmaxArray 1.0 consists of 4992 expressed sequence tags (ESTs) originating from mantle tissue. This microarray was used to analyze the spatial expression of ESTs throughout the mantle organ. The mantle was dissected into five discrete regions and analyzed for differential gene expression with PmaxArray 1.0. Over 2000 ESTs were determined to be differentially expressed among the tissue sections, identifying five major expression regions. In situ hybridization validated and further localized the expression for a subset of these ESTs. Comparative sequence similarity analysis of these ESTs revealed a number of the transcripts were novel while others showed significant sequence similarities to previously characterized shell related genes. Conclusions This investigation has mapped the spatial distribution for over 2000 ESTs present on PmaxArray 1.0 with reference to specific locations of the mantle. Expression profile clusters have indicated at least five unique functioning zones in the mantle. Three of these zones are likely involved in shell related activities including formation of nacre, periostracum and calcitic prismatic microstructure. A number of novel and known transcripts have been identified from these clusters. The development of PmaxArray 1.0, and the spatial map of its ESTs expression in the mantle has begun characterizing the molecular mechanisms linking the organics and inorganics of the molluscan shell. PMID:21936921

Overexpression of miR-9 in mast cells is associated with invasive behavior and spontaneous metastasis

PubMed Central

2014-01-01

Background While microRNA (miRNA) expression is known to be altered in a variety of human malignancies contributing to cancer development and progression, the potential role of miRNA dysregulation in malignant mast cell disease has not been previously explored. The purpose of this study was to investigate the potential contribution of miRNA dysregulation to the biology of canine mast cell tumors (MCTs), a well-established spontaneous model of malignant mast cell disease. Methods We evaluated the miRNA expression profiles from biologically low-grade and biologically high-grade primary canine MCTs using real-time PCR-based TaqMan Low Density miRNA Arrays and performed real-time PCR to evaluate miR-9 expression in primary canine MCTs, malignant mast cell lines, and normal bone marrow-derived mast cells (BMMCs). Mouse mast cell lines and BMMCs were transduced with empty or pre-miR-9 expressing lentiviral constructs and cell proliferation, caspase 3/7 activity, and invasion were assessed. Transcriptional profiling of cells overexpressing miR-9 was performed using Affymetrix GeneChip Mouse Gene 2.0 ST arrays and real-time PCR was performed to validate changes in mRNA expression. Results Our data demonstrate that unique miRNA expression profiles correlate with the biological behavior of primary canine MCTs and that miR-9 expression is increased in biologically high grade canine MCTs and malignant cell lines compared to biologically low grade tumors and normal canine BMMCs. In transformed mouse malignant mast cell lines expressing either wild-type (C57) or activating (P815) KIT mutations and mouse BMMCs, miR-9 overexpression significantly enhanced invasion but had no effect on cell proliferation or apoptosis. Transcriptional profiling of normal mouse BMMCs and P815 cells possessing enforced miR-9 expression demonstrated dysregulation of several genes, including upregulation of CMA1, a protease involved in activation of matrix metalloproteases and extracellular matrix remodeling. Conclusions Our findings demonstrate that unique miRNA expression profiles correlate with the biological behavior of canine MCTs. Furthermore, dysregulation of miR-9 is associated with MCT metastasis potentially through the induction of an invasive phenotype, identifying a potentially novel pathway for therapeutic intervention. PMID:24517413

Gene Transcription Profile of the Detached Retina (An AOS Thesis)

PubMed Central

Zacks, David N.

2009-01-01

Purpose: Separation of the neurosensory retina from the retinal pigment epithelium (RPE) yields many morphologic and functional consequences, including death of the photoreceptor cells, Müller cell hypertrophy, and inner retinal rewiring. Many of these changes are due to the separation-induced activation of specific genes. In this work, we define the gene transcription profile within the retina as a function of time after detachment. We also define the early activation of kinases that might be responsible for the detachment-induced changes in gene transcription. Methods: Separation of the retina from the RPE was induced in Brown-Norway rats by the injection of 1% hyaluronic acid into the subretinal space. Retinas were harvested at 1, 7, and 28 days after separation. Gene transcription profiles for each time point were determined using the Affymetrix Rat 230A gene microarray chip. Transcription levels in detached retinas were compared to those of nondetached retinas with the BRB-ArrayTools Version 3.6.0 using a random variance analysis of variance (ANOVA) model. Confirmation of the significant transcriptional changes for a subset of the genes was performed using microfluidic quantitative real-time polymerase chain reaction (qRT-PCR) assays. Kinase activation was explored using Western blot analysis to look for early phosphorylation of any of the 3 main families of mitogen-activated protein kinases (MAPK): the p38 family, the Janus kinase family, and the p42/p44 family. Results: Retinas separated from the RPE showed extensive alterations in their gene transcription profile. Many of these changes were initiated as early as 1 day after separation, with significant increases by 7 days. ANOVA analysis defined 144 genes that had significantly altered transcription levels as a function of time after separation when setting a false discovery rate at ≤0.1. Confirmatory RT-PCR was performed on 51 of these 144 genes. Differential transcription detected on the microarray chip was confirmed by qRT-PCR for all 51 genes. Western blot analysis showed that the p42/p44 family of MAPK was phosphorylated within 2 hours of retinal-RPE separation. This phosphorylation was detachment-induced and could be inhibited by specific inhibitors of MAPK phosphorylation. Conclusions: Separation of the retina from the RPE induces significant alteration in the gene transcription profile within the retina. These profiles are not static, but change as a function of time after detachment. These gene transcription changes are preceded by the activation of the p42/p44 family of MAPK. This altered transcription may serve as the basis for many of the morphologic, biochemical, and functional changes seen within the detached retina. PMID:20126507

Transcriptomic assessment of resistance to effects of an aryl hydrocarbon receptor (AHR) agonist in embryos of Atlantic killifish (Fundulus heteroclitus) from a marine Superfund site.

PubMed

Oleksiak, Marjorie F; Karchner, Sibel I; Jenny, Matthew J; Franks, Diana G; Welch, David B Mark; Hahn, Mark E

2011-05-24

Populations of Atlantic killifish (Fundulus heteroclitus) have evolved resistance to the embryotoxic effects of polychlorinated biphenyls (PCBs) and other halogenated and nonhalogenated aromatic hydrocarbons that act through an aryl hydrocarbon receptor (AHR)-dependent signaling pathway. The resistance is accompanied by reduced sensitivity to induction of cytochrome P450 1A (CYP1A), a widely used biomarker of aromatic hydrocarbon exposure and effect, but whether the reduced sensitivity is specific to CYP1A or reflects a genome-wide reduction in responsiveness to all AHR-mediated changes in gene expression is unknown. We compared gene expression profiles and the response to 3,3',4,4',5-pentachlorobiphenyl (PCB-126) exposure in embryos (5 and 10 dpf) and larvae (15 dpf) from F. heteroclitus populations inhabiting the New Bedford Harbor, Massachusetts (NBH) Superfund site (PCB-resistant) and a reference site, Scorton Creek, Massachusetts (SC; PCB-sensitive). Analysis using a 7,000-gene cDNA array revealed striking differences in responsiveness to PCB-126 between the populations; the differences occur at all three stages examined. There was a sizeable set of PCB-responsive genes in the sensitive SC population, a much smaller set of PCB-responsive genes in NBH fish, and few similarities in PCB-responsive genes between the two populations. Most of the array results were confirmed, and additional PCB-regulated genes identified, by RNA-Seq (deep pyrosequencing). The results suggest that NBH fish possess a gene regulatory defect that is not specific to one target gene such as CYP1A but rather lies in a regulatory pathway that controls the transcriptional response of multiple genes to PCB exposure. The results are consistent with genome-wide disruption of AHR-dependent signaling in NBH fish.

Biological mechanism analysis of acute renal allograft rejection: integrated of mRNA and microRNA expression profiles.

PubMed

Huang, Shi-Ming; Zhao, Xia; Zhao, Xue-Mei; Wang, Xiao-Ying; Li, Shan-Shan; Zhu, Yu-Hui

2014-01-01

Renal transplantation is the preferred method for most patients with end-stage renal disease, however, acute renal allograft rejection is still a major risk factor for recipients leading to renal injury. To improve the early diagnosis and treatment of acute rejection, study on the molecular mechanism of it is urgent. MicroRNA (miRNA) expression profile and mRNA expression profile of acute renal allograft rejection and well-functioning allograft downloaded from ArrayExpress database were applied to identify differentially expressed (DE) miRNAs and DE mRNAs. DE miRNAs targets were predicted by combining five algorithm. By overlapping the DE mRNAs and DE miRNAs targets, common genes were obtained. Differentially co-expressed genes (DCGs) were identified by differential co-expression profile (DCp) and differential co-expression enrichment (DCe) methods in Differentially Co-expressed Genes and Links (DCGL) package. Then, co-expression network of DCGs and the cluster analysis were performed. Functional enrichment analysis for DCGs was undergone. A total of 1270 miRNA targets were predicted and 698 DE mRNAs were obtained. While overlapping miRNA targets and DE mRNAs, 59 common genes were gained. We obtained 103 DCGs and 5 transcription factors (TFs) based on regulatory impact factors (RIF), then built the regulation network of miRNA targets and DE mRNAs. By clustering the co-expression network, 5 modules were obtained. Thereinto, module 1 had the highest degree and module 2 showed the most number of DCGs and common genes. TF CEBPB and several common genes, such as RXRA, BASP1 and AKAP10, were mapped on the co-expression network. C1R showed the highest degree in the network. These genes might be associated with human acute renal allograft rejection. We conducted biological analysis on integration of DE mRNA and DE miRNA in acute renal allograft rejection, displayed gene expression patterns and screened out genes and TFs that may be related to acute renal allograft rejection.

Biological mechanism analysis of acute renal allograft rejection: integrated of mRNA and microRNA expression profiles

PubMed Central

Huang, Shi-Ming; Zhao, Xia; Zhao, Xue-Mei; Wang, Xiao-Ying; Li, Shan-Shan; Zhu, Yu-Hui

2014-01-01

Objectives: Renal transplantation is the preferred method for most patients with end-stage renal disease, however, acute renal allograft rejection is still a major risk factor for recipients leading to renal injury. To improve the early diagnosis and treatment of acute rejection, study on the molecular mechanism of it is urgent. Methods: MicroRNA (miRNA) expression profile and mRNA expression profile of acute renal allograft rejection and well-functioning allograft downloaded from ArrayExpress database were applied to identify differentially expressed (DE) miRNAs and DE mRNAs. DE miRNAs targets were predicted by combining five algorithm. By overlapping the DE mRNAs and DE miRNAs targets, common genes were obtained. Differentially co-expressed genes (DCGs) were identified by differential co-expression profile (DCp) and differential co-expression enrichment (DCe) methods in Differentially Co-expressed Genes and Links (DCGL) package. Then, co-expression network of DCGs and the cluster analysis were performed. Functional enrichment analysis for DCGs was undergone. Results: A total of 1270 miRNA targets were predicted and 698 DE mRNAs were obtained. While overlapping miRNA targets and DE mRNAs, 59 common genes were gained. We obtained 103 DCGs and 5 transcription factors (TFs) based on regulatory impact factors (RIF), then built the regulation network of miRNA targets and DE mRNAs. By clustering the co-expression network, 5 modules were obtained. Thereinto, module 1 had the highest degree and module 2 showed the most number of DCGs and common genes. TF CEBPB and several common genes, such as RXRA, BASP1 and AKAP10, were mapped on the co-expression network. C1R showed the highest degree in the network. These genes might be associated with human acute renal allograft rejection. Conclusions: We conducted biological analysis on integration of DE mRNA and DE miRNA in acute renal allograft rejection, displayed gene expression patterns and screened out genes and TFs that may be related to acute renal allograft rejection. PMID:25664019

Expression signature as a biomarker for prenatal diagnosis of trisomy 21.

PubMed

Volk, Marija; Maver, Aleš; Lovrečić, Luca; Juvan, Peter; Peterlin, Borut

2013-01-01

A universal biomarker panel with the potential to predict high-risk pregnancies or adverse pregnancy outcome does not exist. Transcriptome analysis is a powerful tool to capture differentially expressed genes (DEG), which can be used as biomarker-diagnostic-predictive tool for various conditions in prenatal setting. In search of biomarker set for predicting high-risk pregnancies, we performed global expression profiling to find DEG in Ts21. Subsequently, we performed targeted validation and diagnostic performance evaluation on a larger group of case and control samples. Initially, transcriptomic profiles of 10 cultivated amniocyte samples with Ts21 and 9 with normal euploid constitution were determined using expression microarrays. Datasets from Ts21 transcriptomic studies from GEO repository were incorporated. DEG were discovered using linear regression modelling and validated using RT-PCR quantification on an independent sample of 16 cases with Ts21 and 32 controls. The classification performance of Ts21 status based on expression profiling was performed using supervised machine learning algorithm and evaluated using a leave-one-out cross validation approach. Global gene expression profiling has revealed significant expression changes between normal and Ts21 samples, which in combination with data from previously performed Ts21 transcriptomic studies, were used to generate a multi-gene biomarker for Ts21, comprising of 9 gene expression profiles. In addition to biomarker's high performance in discriminating samples from global expression profiling, we were also able to show its discriminatory performance on a larger sample set 2, validated using RT-PCR experiment (AUC=0.97), while its performance on data from previously published studies reached discriminatory AUC values of 1.00. Our results show that transcriptomic changes might potentially be used to discriminate trisomy of chromosome 21 in the prenatal setting. As expressional alterations reflect both, causal and reactive cellular mechanisms, transcriptomic changes may thus have future potential in the diagnosis of a wide array of heterogeneous diseases that result from genetic disturbances.

At-TAX: a whole genome tiling array resource for developmental expression analysis and transcript identification in Arabidopsis thaliana

PubMed Central

Laubinger, Sascha; Zeller, Georg; Henz, Stefan R; Sachsenberg, Timo; Widmer, Christian K; Naouar, Naïra; Vuylsteke, Marnik; Schölkopf, Bernhard; Rätsch, Gunnar; Weigel, Detlef

2008-01-01

Gene expression maps for model organisms, including Arabidopsis thaliana, have typically been created using gene-centric expression arrays. Here, we describe a comprehensive expression atlas, Arabidopsis thaliana Tiling Array Express (At-TAX), which is based on whole-genome tiling arrays. We demonstrate that tiling arrays are accurate tools for gene expression analysis and identified more than 1,000 unannotated transcribed regions. Visualizations of gene expression estimates, transcribed regions, and tiling probe measurements are accessible online at the At-TAX homepage. PMID:18613972

Epigenetic repression of HOXB cluster in oral cancer cell lines.

PubMed

Xavier, Flávia Caló Aquino; Destro, Maria Fernanda de Souza Setubal; Duarte, Carina Magalhães Esteves; Nunes, Fabio Daumas

2014-08-01

Aberrant DNA methylation is a fundamental transcriptional control mechanism in carcinogenesis. The expression of homeobox genes is usually controlled by an epigenetic mechanism, such as the methylation of CpG islands in the promoter region. The aim of this study was to describe the differential methylation pattern of HOX genes in oral squamous cell carcinoma (OSCC) cell lines and transcript status in a group of hypermethylated and hypomethylated genes. Quantitative analysis of DNA methylation was performed on two OSCC cell lines (SCC4 and SCC9) using a method denominated Human Homeobox Genes EpiTect Methyl qPCR Arrays, which allowed fast, precise methylation detection of 24 HOX specific genes without bisulfite conversion. Methylation greater than 50% was detected in HOXA11, HOXA6, HOXA7, HOXA9, HOXB1, HOXB2, HOXB3, HOXB4, HOXB5, HOXB6, HOXC8 and HOXD10. Both cell lines demonstrated similar hypermethylation status for eight HOX genes. A similar pattern of promoter hypermethylation and hypomethylation was demonstrated for the HOXB cluster and HOXA cluster, respectively. Moreover, the hypermethylation profile of the HOXB cluster, especially HOXB4, was correlated with decreased transcript expression, which was restored following treatment with 5-aza-2'-deoxycytidine. The homeobox methylation profile in OSCC cell lines is consistent with an epigenetic biomarker. Copyright © 2014 Elsevier Ltd. All rights reserved.

Genomic profiling of rice sperm cell transcripts reveals conserved and distinct elements in the flowering plant male germ lineage.

PubMed

Russell, Scott D; Gou, Xiaoping; Wong, Chui E; Wang, Xinkun; Yuan, Tong; Wei, Xiaoping; Bhalla, Prem L; Singh, Mohan B

2012-08-01

Genomic assay of sperm cell RNA provides insight into functional control, modes of regulation, and contributions of male gametes to double fertilization. Sperm cells of rice (Oryza sativa) were isolated from field-grown, disease-free plants and RNA was processed for use with the full-genome Affymetrix microarray. Comparison with Gene Expression Omnibus (GEO) reference arrays confirmed expressionally distinct gene profiles. A total of 10,732 distinct gene sequences were detected in sperm cells, of which 1668 were not expressed in pollen or seedlings. Pathways enriched in male germ cells included ubiquitin-mediated pathways, pathways involved in chromatin modeling including histones, histone modification and nonhistone epigenetic modification, and pathways related to RNAi and gene silencing. Genome-wide expression patterns in angiosperm sperm cells indicate common and divergent themes in the male germline that appear to be largely self-regulating through highly up-regulated chromatin modification pathways. A core of highly conserved genes appear common to all sperm cells, but evidence is still emerging that another class of genes have diverged in expression between monocots and dicots since their divergence. Sperm cell transcripts present at fusion may be transmitted through plasmogamy during double fertilization to effect immediate post-fertilization expression of early embryo and (or) endosperm development. © 2012 The Authors. New Phytologist © 2012 New Phytologist Trust.

Analysis of ripening-related gene expression in papaya using an Arabidopsis-based microarray

PubMed Central

2012-01-01

Background Papaya (Carica papaya L.) is a commercially important crop that produces climacteric fruits with a soft and sweet pulp that contain a wide range of health promoting phytochemicals. Despite its importance, little is known about transcriptional modifications during papaya fruit ripening and their control. In this study we report the analysis of ripe papaya transcriptome by using a cross-species (XSpecies) microarray technique based on the phylogenetic proximity between papaya and Arabidopsis thaliana. Results Papaya transcriptome analyses resulted in the identification of 414 ripening-related genes with some having their expression validated by qPCR. The transcription profile was compared with that from ripening tomato and grape. There were many similarities between papaya and tomato especially with respect to the expression of genes encoding proteins involved in primary metabolism, regulation of transcription, biotic and abiotic stress and cell wall metabolism. XSpecies microarray data indicated that transcription factors (TFs) of the MADS-box, NAC and AP2/ERF gene families were involved in the control of papaya ripening and revealed that cell wall-related gene expression in papaya had similarities to the expression profiles seen in Arabidopsis during hypocotyl development. Conclusion The cross-species array experiment identified a ripening-related set of genes in papaya allowing the comparison of transcription control between papaya and other fruit bearing taxa during the ripening process. PMID:23256600

Microarray-based genomic profiling reveals novel genomic aberrations in follicular lymphoma which associate with patient survival and gene expression status.

PubMed

Schwaenen, Carsten; Viardot, Andreas; Berger, Hilmar; Barth, Thomas F E; Bentink, Stefan; Döhner, Hartmut; Enz, Martina; Feller, Alfred C; Hansmann, Martin-Leo; Hummel, Michael; Kestler, Hans A; Klapper, Wolfram; Kreuz, Markus; Lenze, Dido; Loeffler, Markus; Möller, Peter; Müller-Hermelink, Hans-Konrad; Ott, German; Rosolowski, Maciej; Rosenwald, Andreas; Ruf, Sandra; Siebert, Reiner; Spang, Rainer; Stein, Harald; Truemper, Lorenz; Lichter, Peter; Bentz, Martin; Wessendorf, Swen

2009-01-01

Follicular lymphoma (FL) is characterized by a large number of chromosomal aberrations. However, their exact genomic extension and involved target genes remain to be determined. For this purpose, we used array-based intermediate-high resolution genomic profiling in combination with Affymetrix gene expression analysis. Tumor specimens from 128 FL patients were analyzed for the presence of genomic aberrations and the results were correlated to clinical data sets and mRNA expression levels. In 114 (89%) of the 128 analyzed cases, a total of 688 genomic aberrations (384 gains/amplifications and 304 losses) were detected. Frequent genomic aberrations were: -1p36 (18%), +2p15 (24%), -3q (14%), -6q (25%), +7p (19%), +7q (23%), +8q (14%), -9p (16%), -11q (15%), +12q (20%), -13q (11%), -17p (16%), +18p (18%), and +18q (28%). Critical segments of these imbalances were delineated to genomic fragments with a minimum size down to 0.2 Mb. By comparison of these with mRNA gene expression data, putative candidate genes were identified. Moreover, we found that deletions affecting the tumor suppressor gene CDKN2A/B on 9p21 were detected in nontransformed FL grade I-II. For this aberration as well as for -6q25 and -6q26, an association with inferior survival was observed.

Correction of Spatial Bias in Oligonucleotide Array Data

PubMed Central

Lemieux, Sébastien

2013-01-01

Background. Oligonucleotide microarrays allow for high-throughput gene expression profiling assays. The technology relies on the fundamental assumption that observed hybridization signal intensities (HSIs) for each intended target, on average, correlate with their target's true concentration in the sample. However, systematic, nonbiological variation from several sources undermines this hypothesis. Background hybridization signal has been previously identified as one such important source, one manifestation of which appears in the form of spatial autocorrelation. Results. We propose an algorithm, pyn, for the elimination of spatial autocorrelation in HSIs, exploiting the duality of desirable mutual information shared by probes in a common probe set and undesirable mutual information shared by spatially proximate probes. We show that this correction procedure reduces spatial autocorrelation in HSIs; increases HSI reproducibility across replicate arrays; increases differentially expressed gene detection power; and performs better than previously published methods. Conclusions. The proposed algorithm increases both precision and accuracy, while requiring virtually no changes to users' current analysis pipelines: the correction consists merely of a transformation of raw HSIs (e.g., CEL files for Affymetrix arrays). A free, open-source implementation is provided as an R package, compatible with standard Bioconductor tools. The approach may also be tailored to other platform types and other sources of bias. PMID:23573083

Differential expression of hepatic genes with embryonic exposure to an environmentally relevant PCB mixture in Japanese quail (Coturnix japonica).

PubMed

Bohannon, Meredith E; Porter, Tom E; Lavoie, Emma T; Ottinger, Mary Ann

2018-06-22

The upper Hudson River was contaminated with polychlorinated biphenyls (PCB) Aroclor mixtures from the 1940s until the late 1970s. Several well-established biomarkers, such as induction of hepatic cytochrome P450 monooxygenases, were used to measure exposure to PCBs and similar contaminants in birds. In the present study, Japanese quail eggs were injected with a PCB mixture based upon a congener profile found in spotted sandpiper eggs at the upper Hudson River and subsequently, RNA was extracted from hatchling liver tissue for hybridization to a customized chicken cDNA microarray. Nominal concentrations of the mixture used for microarray hybridization were 0, 6, 12, or 49 μg/g egg. Hepatic gene expression profiles were analyzed using cluster and pathway analyses. Results showed potentially useful biomarkers of both exposure and effect attributed to PCB mixture. Biorag and Ingenuity Pathway Analysis® analyses revealed differentially expressed genes including those involved in glycolysis, xenobiotic metabolism, replication, protein degradation, and tumor regulation. These genes included cytochrome P450 1A5 (CYP1A5), cytochrome b5 (CYB5), NADH-cytochrome b5 reductase, glutathione S-transferase (GSTA), fructose bisphosphate aldolase (ALDOB), glycogen phosphorylase, carbonic anhydrase, and DNA topoisomerase II. CYP1A5, CYB5, GSTA, and ALDOB were chosen for quantitative real-time polymerase chain reaction confirmation, as these genes exhibited a clear dose response on the array. Data demonstrated that an initial transcriptional profile associated with an environmentally relevant PCB mixture in Japanese quail occurred.

DNA methylation profiling of esophageal adenocarcinoma using Methylation Ligation-dependent Macroarray (MLM).

PubMed

Guilleret, Isabelle; Losi, Lorena; Chelbi, Sonia T; Fonda, Sergio; Bougel, Stéphanie; Saponaro, Sara; Gozzi, Gaia; Alberti, Loredana; Braunschweig, Richard; Benhattar, Jean

2016-10-14

Most types of cancer cells are characterized by aberrant methylation of promoter genes. In this study, we described a rapid, reproducible, and relatively inexpensive approach allowing the detection of multiple human methylated promoter genes from many tissue samples, without the need of bisulfite conversion. The Methylation Ligation-dependent Macroarray (MLM), an array-based analysis, was designed in order to measure methylation levels of 58 genes previously described as putative biomarkers of cancer. The performance of the design was proven by screening the methylation profile of DNA from esophageal cell lines, as well as microdissected formalin-fixed and paraffin-embedded (FFPE) tissues from esophageal adenocarcinoma (EAC). Using the MLM approach, we identified 32 (55%) hypermethylated promoters in EAC, and not or rarely methylated in normal tissues. Among them, 21promoters were found aberrantly methylated in more than half of tumors. Moreover, seven of them (ADAMTS18, APC, DKK2, FOXL2, GPX3, TIMP3 and WIF1) were found aberrantly methylated in all or almost all the tumor samples, suggesting an important role for these genes in EAC. In addition, dysregulation of the Wnt pathway with hypermethylation of several Wnt antagonist genes was frequently observed. MLM revealed a homogeneous pattern of methylation for a majority of tumors which were associated with an advanced stage at presentation and a poor prognosis. Interestingly, the few tumors presenting less methylation changes had a lower pathological stage. In conclusion, this study demonstrated the feasibility and accuracy of MLM for DNA methylation profiling of FFPE tissue samples. Copyright © 2016 Elsevier Inc. All rights reserved.

Assessment of Normal Variability in Peripheral Blood Gene Expression

DOE PAGES

Campbell, Catherine; Vernon, Suzanne D.; Karem, Kevin L.; ...

2002-01-01

Peripheral blood is representative of many systemic processes and is an ideal sample for expression profiling of diseases that have no known or accessible lesion. Peripheral blood is a complex mixture of cell types and some differences in peripheral blood gene expression may reflect the timing of sample collection rather than an underlying disease process. For this reason, it is important to assess study design factors that may cause variability in gene expression not related to what is being analyzed. Variation in the gene expression of circulating peripheral blood mononuclear cells (PBMCs) from three healthy volunteers sampled three times onemore » day each week for one month was examined for 1,176 genes printed on filter arrays. Less than 1% of the genes showed any variation in expression that was related to the time of collection, and none of the changes were noted in more than one individual. These results suggest that observed variation was due to experimental variability.« less

[Chiparray-based identification of gene expression in HUVECs treated with low frequency electric fields].

PubMed

Ulrich, D; Ulrich, F; Silny, J; Unglaub, F; Pallua, N

2006-06-01

After high-voltage electric injury, patients often show progressive tissue necrosis and thrombosis of blood vessels even remote from the entry and exit sites of electrical current. Recently, we were able to demonstrate IN VIVO and VITRO the release of several prothrombotic factors. In this study, we report on IN VITRO studies performed to characterize gene expression profiles using a DNA-microarray in HUVECs (human umbilical vein endothelial cells) exposed to low frequency electrical current. HUVECs were plated and grown to confluence in a culture chamber. They were exposed to 25 periods of 50 Hz sinusoidal waves. The periods had field strength of 60 V/cm and duration of 100 ms. Periods were interrupted by 10-second intervals to prevent significant joule heating. Control HUVECs were treated identically except that no electric field was applied. Samples from control and treated cells were taken after six and 24 hours. A PIQOR Immunology Array (Milteny Biotech) containing 1076 cDNAs was used for gene expression analysis. Hybridization of Cy3- and Cy5-labelled samples, image capture, and signal quantification of hybridized arrays were performed. Local background was subtracted from the signal to obtain the net signal intensity and the ratio of Cy5/Cy3. The ratios were normalized to the median of all ratios and the mean of the ratios of four corresponding spots was computed. More than two-fold increases or decreases of the gene expression were regarded as relevant. A total of 413 genes (1s + s) respectively 345 genes (2s + s) could be detected. The results obtained display a distinct expression pattern of up-regulated genes known to be important for hemostasis (e.g. UPA, UPAR, ECE1, PAFAH1B1, PGT, INOS, ENOS, TPA, ICAM1, VCAM1, PAI1, PAI2, VWF, PTGDR, F3, THBD), which was most evident after 24 hours. This expression profile might lead to a hypercoagulated state. Furthermore, the expression of genes involved in angiogenesis was reduced whereas the expression of those involved in platelet formation was increased. Our results indicate that low frequency electrical fields induce a distinct signature of differential gene expression in exposed HUVECs. This might explain the clinical observation of thrombosis and progressive tissue necrosis after electrical injury.

Microarray analysis of peripheral blood lymphocytes from ALS patients and the SAFE detection of the KEGG ALS pathway

PubMed Central

2011-01-01

Background Sporadic amyotrophic lateral sclerosis (sALS) is a motor neuron disease with poorly understood etiology. Results of gene expression profiling studies of whole blood from ALS patients have not been validated and are difficult to relate to ALS pathogenesis because gene expression profiles depend on the relative abundance of the different cell types present in whole blood. We conducted microarray analyses using Agilent Human Whole Genome 4 × 44k Arrays on a more homogeneous cell population, namely purified peripheral blood lymphocytes (PBLs), from ALS patients and healthy controls to identify molecular signatures possibly relevant to ALS pathogenesis. Methods Differentially expressed genes were determined by LIMMA (Linear Models for MicroArray) and SAM (Significance Analysis of Microarrays) analyses. The SAFE (Significance Analysis of Function and Expression) procedure was used to identify molecular pathway perturbations. Proteasome inhibition assays were conducted on cultured peripheral blood mononuclear cells (PBMCs) from ALS patients to confirm alteration of the Ubiquitin/Proteasome System (UPS). Results For the first time, using SAFE in a global gene ontology analysis (gene set size 5-100), we show significant perturbation of the KEGG (Kyoto Encyclopedia of Genes and Genomes) ALS pathway of motor neuron degeneration in PBLs from ALS patients. This was the only KEGG disease pathway significantly upregulated among 25, and contributing genes, including SOD1, represented 54% of the encoded proteins or protein complexes of the KEGG ALS pathway. Further SAFE analysis, including gene set sizes >100, showed that only neurodegenerative diseases (4 out of 34 disease pathways) including ALS were significantly upregulated. Changes in UBR2 expression correlated inversely with time since onset of disease and directly with ALSFRS-R, implying that UBR2 was increased early in the course of ALS. Cultured PBMCs from ALS patients accumulated more ubiquitinated proteins than PBMCs from healthy controls in a serum-dependent manner confirming changes in this pathway. Conclusions Our study indicates that PBLs from sALS patients are strong responders to systemic signals or local signals acquired by cell trafficking, representing changes in gene expression similar to those present in brain and spinal cord of sALS patients. PBLs may provide a useful means to study ALS pathogenesis. PMID:22027401

Microelectronic electroporation array

NASA Astrophysics Data System (ADS)

Johnson, Lee J.; Shaffer, Kara J.; Skeath, Perry; Perkins, Frank K.; Pancrazio, Joseph; Scribner, Dean

2004-06-01

Gene Array technology has allowed for the study of gene binding by creating thousands of potential binding sites on a single device. A limitation of the current technology is that the effects of the gene and the gene-derived proteins cannot be studied in situ the same way, thousand site cell arrays are not readily available. We propose a new device structure to study the effects of gene modification on cells. This new array technology uses electroporation to target specific areas within a cell culture for transfection of genes. Electroporation arrays will allow high throughput analysis of gene effects on a given cell's response to a stress or a genes ability to restore normal cell function in disease modeling cells. Fluorescent imaging of dye labeled indicator molecules or cell viability will provide results indicating the most effective genes. The electroporation array consists of a microelectronic circuit, ancillary electronics, protecting electrode surface for cell culturing and a perfusion system for gene or drug delivery. The advantages of the current device are that there are 3200 sites for electroporation, all or any subsets of the electrodes can be activated. The cells are held in place by the electrode material. This technology could also be applied to high throughput screening of cell impermeant drugs.

Effects of chronic morphine and morphine withdrawal on gene expression in rat peripheral blood mononuclear cells.

PubMed

Desjardins, Stephane; Belkai, Emilie; Crete, Dominique; Cordonnier, Laurie; Scherrmann, Jean-Michel; Noble, Florence; Marie-Claire, Cynthia

2008-12-01

Chronic morphine treatment alters gene expression in brain structures. There are increasing evidences showing a correlation, in gene expression modulation, between blood cells and brain in psychological troubles. To test whether gene expression regulation in blood cells could be found in drug addiction, we investigated gene expression profiles in peripheral blood mononuclear (PBMC) cells of saline and morphine-treated rats. In rats chronically treated with morphine, the behavioral signs of spontaneous withdrawal were observed and a withdrawal score was determined. This score enabled to select the time points at which the animals displayed the mildest and strongest withdrawal signs (12 h and 36 h after the last injection). Oligonucleotide arrays were used to assess differential gene expression in the PBMCs and quantitative real-time RT-PCR to validate the modulation of several candidate genes 12 h and 36 h after the last injection. Among the 812 differentially expressed candidates, several genes (Adcy5, Htr2a) and pathways (Map kinases, G-proteins, integrins) have already been described as modulated in the brain of morphine-treated rats. Sixteen out of the twenty-four tested candidates were validated at 12 h, some of them showed a sustained modulation at 36 h while for most of them the modulation evolved as the withdrawal score increased. This study suggests similarities between the gene expression profile in PBMCs and brain of morphine-treated rats. Thus, the searching of correlations between the severity of the withdrawal and the PBMCs gene expression pattern by transcriptional analysis of blood cells could be promising for the study of the mechanisms of addiction.

Genome-scale analysis and comparison of gene expression profiles in developing and germinated pollen in Oryza sativa

PubMed Central

2010-01-01

Background Pollen development from the microspore involves a series of coordinated cellular events, and the resulting mature pollen has a specialized function to quickly germinate, produce a polar-growth pollen tube derived from the vegetative cell, and deliver two sperm cells into the embryo sac for double fertilization. The gene expression profiles of developing and germinated pollen have been characterised by use of the eudicot model plant Arabidopsis. Rice, one of the most important cereal crops, has been used as an excellent monocot model. A comprehensive analysis of transcriptome profiles of developing and germinated pollen in rice is important to understand the conserved and diverse mechanism underlying pollen development and germination in eudicots and monocots. Results We used Affymetrix GeneChip® Rice Genome Array to comprehensively analyzed the dynamic changes in the transcriptomes of rice pollen at five sequential developmental stages from microspores to germinated pollen. Among the 51,279 transcripts on the array, we found 25,062 pollen-preferential transcripts, among which 2,203 were development stage-enriched. The diversity of transcripts decreased greatly from microspores to mature and germinated pollen, whereas the number of stage-enriched transcripts displayed a "U-type" change, with the lowest at the bicellular pollen stage; and a transition of overrepresented stage-enriched transcript groups associated with different functional categories, which indicates a shift in gene expression program at the bicellular pollen stage. About 54% of the now-annotated rice F-box protein genes were expressed preferentially in pollen. The transcriptome profile of germinated pollen was significantly and positively correlated with that of mature pollen. Analysis of expression profiles and coexpressed features of the pollen-preferential transcripts related to cell cycle, transcription, the ubiquitin/26S proteasome system, phytohormone signalling, the kinase system and defense/stress response revealed five expression patterns, which are compatible with changes in major cellular events during pollen development and germination. A comparison of pollen transcriptomes between rice and Arabidopsis revealed that 56.6% of the rice pollen preferential genes had homologs in Arabidopsis genome, but 63.4% of these homologs were expressed, with a small proportion being expressed preferentially, in Arabidopsis pollen. Rice and Arabidopsis pollen had non-conservative transcription factors each. Conclusions Our results demonstrated that rice pollen expressed a set of reduced but specific transcripts in comparison with vegetative tissues, and the number of stage-enriched transcripts displayed a "U-type" change during pollen development, with the lowest at the bicellular pollen stage. These features are conserved in rice and Arabidopsis. The shift in gene expression program at the bicellular pollen stage may be important to the transition from earlier cell division to later pollen maturity. Pollen at maturity pre-synthesized transcripts needed for germination and early pollen tube growth. The transcription regulation associated with pollen development would have divergence between the two species. Our results also provide novel insights into the molecular program and key components of the regulatory network regulating pollen development and germination. PMID:20507633

Theobroma cacao L. pathogenesis-related gene tandem array members show diverse expression dynamics in response to pathogen colonization.

PubMed

Fister, Andrew S; Mejia, Luis C; Zhang, Yufan; Herre, Edward Allen; Maximova, Siela N; Guiltinan, Mark J

2016-05-17

The pathogenesis-related (PR) group of proteins are operationally defined as polypeptides that increase in concentration in plant tissues upon contact with a pathogen. To date, 17 classes of highly divergent proteins have been described that act through multiple mechanisms of pathogen resistance. Characterizing these families in cacao, an economically important tree crop, and comparing the families to those in other species, is an important step in understanding cacao's immune response. Using publically available resources, all members of the 17 recognized pathogenesis-related gene families in the genome of Theobroma cacao were identified and annotated resulting in a set of ~350 members in both published cacao genomes. Approximately 50 % of these genes are organized in tandem arrays scattered throughout the genome. This feature was observed in five additional plant taxa (three dicots and two monocots), suggesting that tandem duplication has played an important role in the evolution of the PR genes in higher plants. Expression profiling captured the dynamics and complexity of PR genes expression at basal levels and after induction by two cacao pathogens (the oomycete, Phytophthora palmivora, and the fungus, Colletotrichum theobromicola), identifying specific genes within families that are more responsive to pathogen challenge. Subsequent qRT-PCR validated the induction of several PR-1, PR-3, PR-4, and PR-10 family members, with greater than 1000 fold induction detected for specific genes. We describe candidate genes that are likely to be involved in cacao's defense against Phytophthora and Colletotrichum infection and could be potentially useful for marker-assisted selection for breeding of disease resistant cacao varieties. The data presented here, along with existing cacao-omics resources, will enable targeted functional genetic screening of defense genes likely to play critical functions in cacao's defense against its pathogens.

Altered intragenic DNA methylation of HOOK2 gene in adipose tissue from individuals with obesity and type 2 diabetes

PubMed Central

Fernández-Bayón, Gustavo; Morales-Sánchez, Paula; Sanz, Lourdes; Turienzo, Estrella; González, Juan José; Martinez-Faedo, Ceferino; Suarez-Gutiérrez, Lorena; Ares, Jessica; Díaz-Naya, Lucia; Martin-Nieto, Alicia; Fernández-Morera, Juan L.; Fraga, Mario F.

2017-01-01

Aims/Hypothesis Failure in glucose response to insulin is a common pathology associated with obesity. In this study, we analyzed the genome wide DNA methylation profile of visceral adipose tissue (VAT) samples in a population of individuals with obesity and assessed whether differential methylation profiles are associated with the presence of type 2 diabetes (T2D). Methods More than 485,000 CpG genome sites from VAT samples from women with obesity undergoing gastric bypass (n = 18), and classified as suffering from type 2 diabetes (T2D) or not (no type 2 diabetes, NT2D), were analyzed using DNA methylation arrays. Results We found significant differential methylation between T2D and NT2D samples in 24 CpGs that map with sixteen genes, one of which, HOOK2, demonstrated a significant correlation between differentially hypermethylated regions on the gene body and the presence of type 2 diabetes. This was validated by pyrosequencing in a population of 91 samples from both males and females with obesity. Furthermore, when these results were analyzed by gender, female T2D samples were found hypermethylated at the cg04657146-region and the cg 11738485-region of HOOK2 gene, whilst, interestingly, male samples were found hypomethylated in this latter region. Conclusion The differential methylation profile of the HOOK2 gene in individuals with T2D and obesity might be related to the attendant T2D, but further studies are required to identify the potential role of HOOK2 gene in T2D disease. The finding of gender differences in T2D methylation of HOOK2 also warrants further investigation. PMID:29228058

Coherent and incoherent combination of Gaussian beams employing lens array distributed on the spherical chamber.

PubMed

Huang, Zhihua; Wei, Xiaofeng; Li, Mingzhong; Wang, Jianjun; Lin, Honghuan; Xu, Dangpeng; Deng, Ying; Zhang, Rui

2012-04-01

Coherent and incoherent combination of Gaussian beams employing a lens array distributed on the spherical chamber is theoretically analyzed. The output field of each source in the array is coupled through an individual optical system whose local optical axis coincides with the radial direction of the chamber. The resulting intensity profile near the origin is derived. The intensity profile and power in the bucket on the target for rectangular and hexagonal arrangement are numerically calculated. The influences of the center-to-center separation and the ring number of the focusing lens array are given. The synthetic intensity profile of incoherent combination changes little for a lens array scale much smaller than the chamber size. In contrast, the synthetic intensity profile of coherent combination shows an interference pattern with a sharp central peak and sidelobes.

Gene expression profiles in the rat central nervous system induced by JP-8 jet fuel vapor exposure.

PubMed

Lin, Baochuan; Ritchie, Glenn D; Rossi, John; Pancrazio, Joseph J

2004-06-17

Jet propulsion fuel-8 (JP-8) is the predominant fuel for military land vehicles and aircraft used in the US and NATO. Occupational exposure to jet fuel in military personnel has raised concern for the health risk associated with such exposure in the Department of Defense. Clinical studies of humans chronically exposed to jet fuel have suggested both neurotoxicity and neurobehavioral deficits. We utilized rat neurobiology U34 array to measure gene expression changes in whole brain tissue of rats exposed repeatedly to JP-8, under conditions that simulated possible occupational exposure (6 h/day for 91 days) to JP-8 vapor at 250, 500, and 1000 mg/m(3), respectively. Our studies revealed that the gene expression changes of exposure groups can be divided into two main categories according to their functions: (1). neurotransmitter signaling pathways; and (2). stress response. The implications of these gene expression changes are discussed.

Array-Based Comparative Genomic Hybridization Analysis Reveals Chromosomal Copy Number Aberrations Associated with Clinical Outcome in Canine Diffuse Large B-Cell Lymphoma

PubMed Central

Bresolin, Silvia; Marconato, Laura; Comazzi, Stefano; Te Kronnie, Geertruy; Aresu, Luca

2014-01-01

Canine Diffuse Large B-cell Lymphoma (cDLBCL) is an aggressive cancer with variable clinical response. Despite recent attempts by gene expression profiling to identify the dog as a potential animal model for human DLBCL, this tumor remains biologically heterogeneous with no prognostic biomarkers to predict prognosis. The aim of this work was to identify copy number aberrations (CNAs) by high-resolution array comparative genomic hybridization (aCGH) in 12 dogs with newly diagnosed DLBCL. In a subset of these dogs, the genetic profiles at the end of therapy and at relapse were also assessed. In primary DLBCLs, 90 different genomic imbalances were counted, consisting of 46 gains and 44 losses. Two gains in chr13 were significantly correlated with clinical stage. In addition, specific regions of gains and losses were significantly associated to duration of remission. In primary DLBCLs, individual variability was found, however 14 recurrent CNAs (>30%) were identified. Losses involving IGK, IGL and IGH were always found, and gains along the length of chr13 and chr31 were often observed (>41%). In these segments, MYC, LDHB, HSF1, KIT and PDGFRα are annotated. At the end of therapy, dogs in remission showed four new CNAs, whereas three new CNAs were observed in dogs at relapse compared with the previous profiles. One ex novo CNA, involving TCR, was present in dogs in remission after therapy, possibly induced by the autologous vaccine. Overall, aCGH identified small CNAs associated with outcome, which, along with future expression studies, may reveal target genes relevant to cDLBCL. PMID:25372838

Arachidonic and oleic acid exert distinct effects on the DNA methylome

PubMed Central

Silva-Martínez, Guillermo A.; Rodríguez-Ríos, Dalia; Alvarado-Caudillo, Yolanda; Vaquero, Alejandro; Esteller, Manel; Carmona, F. Javier; Moran, Sebastian; Nielsen, Finn C.; Wickström-Lindholm, Marie; Wrobel, Katarzyna; Wrobel, Kazimierz; Barbosa-Sabanero, Gloria; Zaina, Silvio; Lund, Gertrud

2016-01-01

ABSTRACT Abnormal fatty acid metabolism and availability are landmarks of metabolic diseases, which in turn are associated with aberrant DNA methylation profiles. To understand the role of fatty acids in disease epigenetics, we sought DNA methylation profiles specifically induced by arachidonic (AA) or oleic acid (OA) in cultured cells and compared those with published profiles of normal and diseased tissues. THP-1 monocytes were stimulated with AA or OA and analyzed using Infinium HumanMethylation450 BeadChip (Illumina) and Human Exon 1.0 ST array (Affymetrix). Data were corroborated in mouse embryonic fibroblasts. Comparisons with publicly available data were conducted by standard bioinformatics. AA and OA elicited a complex response marked by a general DNA hypermethylation and hypomethylation in the 1–200 μM range, respectively, with a maximal differential response at the 100 μM dose. The divergent response to AA and OA was prominent within the gene body of target genes, where it correlated positively with transcription. AA-induced DNA methylation profiles were similar to the corresponding profiles described for palmitic acid, atherosclerosis, diabetes, obesity, and autism, but relatively dissimilar from OA-induced profiles. Furthermore, human atherosclerosis grade-associated DNA methylation profiles were significantly enriched in AA-induced profiles. Biochemical evidence pointed to β-oxidation, PPAR-α, and sirtuin 1 as important mediators of AA-induced DNA methylation changes. In conclusion, AA and OA exert distinct effects on the DNA methylome. The observation that AA may contribute to shape the epigenome of important metabolic diseases, supports and expands current diet-based therapeutic and preventive efforts. PMID:27088456

MALDI-TOF mass spectrometry for quantitative gene expression analysis of acid responses in Staphylococcus aureus.

PubMed

Rode, Tone Mari; Berget, Ingunn; Langsrud, Solveig; Møretrø, Trond; Holck, Askild

2009-07-01

Microorganisms are constantly exposed to new and altered growth conditions, and respond by changing gene expression patterns. Several methods for studying gene expression exist. During the last decade, the analysis of microarrays has been one of the most common approaches applied for large scale gene expression studies. A relatively new method for gene expression analysis is MassARRAY, which combines real competitive-PCR and MALDI-TOF (matrix-assisted laser desorption/ionization time-of-flight) mass spectrometry. In contrast to microarray methods, MassARRAY technology is suitable for analysing a larger number of samples, though for a smaller set of genes. In this study we compare the results from MassARRAY with microarrays on gene expression responses of Staphylococcus aureus exposed to acid stress at pH 4.5. RNA isolated from the same stress experiments was analysed using both the MassARRAY and the microarray methods. The MassARRAY and microarray methods showed good correlation. Both MassARRAY and microarray estimated somewhat lower fold changes compared with quantitative real-time PCR (qRT-PCR). The results confirmed the up-regulation of the urease genes in acidic environments, and also indicated the importance of metal ion regulation. This study shows that the MassARRAY technology is suitable for gene expression analysis in prokaryotes, and has advantages when a set of genes is being analysed for an organism exposed to many different environmental conditions.

Transcriptome complexity in cardiac development and diseases--an expanding universe between genome and phenome.

PubMed

Gao, Chen; Wang, Yibin

2014-01-01

With the advancement of transcriptome profiling by micro-arrays and high-throughput RNA-sequencing, transcriptome complexity and its dynamics are revealed at different levels in cardiovascular development and diseases. In this review, we will highlight the recent progress in our knowledge of cardiovascular transcriptome complexity contributed by RNA splicing, RNA editing and noncoding RNAs. The emerging importance of many of these previously under-explored aspects of gene regulation in cardiovascular development and pathology will be discussed.

Genome-wide high-resolution aCGH analysis of gestational choriocarcinomas.

PubMed

Poaty, Henriette; Coullin, Philippe; Peko, Jean Félix; Dessen, Philippe; Diatta, Ange Lucien; Valent, Alexander; Leguern, Eric; Prévot, Sophie; Gombé-Mbalawa, Charles; Candelier, Jean-Jacques; Picard, Jean-Yves; Bernheim, Alain

2012-01-01

Eleven samples of DNA from choriocarcinomas were studied by high resolution CGH-array 244 K. They were studied after histopathological confirmation of the diagnosis, of the androgenic etiology and after a microsatellite marker analysis confirming the absence of contamination of tumor DNA from maternal DNA. Three cell lines, BeWo, JAR, JEG were also studied by this high resolution pangenomic technique. According to aCGH analysis, the de novo choriocarcinomas exhibited simple chromosomal rearrangements or normal profiles. The cell lines showed various and complex chromosomal aberrations. 23 Minimal Critical Regions were defined that allowed us to list the genes that were potentially implicated. Among them, unusually high numbers of microRNA clusters and imprinted genes were observed.

Genome-wide profiling of gene expression in the epididymis of alpha-chlorohydrin-induced infertile rats using an oligonucleotide microarray

PubMed Central

2010-01-01

Background As one of the chlorinated antifertility compounds, alpha-chlorohydrin (ACH) can inhibit glyceraldehyde-3-phosphate dehydrogenase (G3PDH) activity in epididymal sperm and affect sperm energy metabolism, maturation and fertilization, eventually leading to male infertility. Further studies demonstrated that the inhibitory effect of ACH on G3PDH is not only confined to epididymal sperm but also to the epididymis. Moreover, little investigation on gene expression changes in the epididymis after ACH treatment has been conducted. Therefore, gene expression studies may indicate new epididymal targets related to sperm maturation and fertility through the analysis of ACH-treated infertile animals. Methods Rats were treated with ACH for ten consecutive days, and then each male rat copulated with two female rats in proestrus. Then sperm maturation and other fertility parameters were analyzed. Furthermore, we identified epididymal-specific genes that are associated with fertility between control and ACH groups using an Affymetrix Rat 230 2.0 oligo-microarray. Finally, we performed RT-PCR analysis for several differentially expressed genes to validate the alteration in gene expression observed by oligonucleotide microarray. Results Among all the differentially expressed genes, we analyzed and screened the down-regulated genes associated with metabolism processes, which are considered the major targets of ACH action. Simultaneously, the genes that were up-regulated by chlorohydrin were detected. The genes that negatively regulate sperm maturation and fertility include apoptosis and immune-related genes and have not been reported previously. The overall results of PCR analysis for selected genes were consistent with the array data. Conclusions In this study, we have described the genome-wide profiles of gene expression in the epididymides of infertile rats induced by ACH, which could become potential epididymal specific targets for male contraception and infertility treatment. PMID:20409345

Genome-wide profiling of gene expression in the epididymis of alpha-chlorohydrin-induced infertile rats using an oligonucleotide microarray.

PubMed

Xie, Shuwu; Zhu, Yan; Ma, Li; Lu, Yingying; Zhou, Jieyun; Gui, Youlun; Cao, Lin

2010-04-22

As one of the chlorinated antifertility compounds, alpha-chlorohydrin (ACH) can inhibit glyceraldehyde-3-phosphate dehydrogenase (G3PDH) activity in epididymal sperm and affect sperm energy metabolism, maturation and fertilization, eventually leading to male infertility. Further studies demonstrated that the inhibitory effect of ACH on G3PDH is not only confined to epididymal sperm but also to the epididymis. Moreover, little investigation on gene expression changes in the epididymis after ACH treatment has been conducted. Therefore, gene expression studies may indicate new epididymal targets related to sperm maturation and fertility through the analysis of ACH-treated infertile animals. Rats were treated with ACH for ten consecutive days, and then each male rat copulated with two female rats in proestrus. Then sperm maturation and other fertility parameters were analyzed. Furthermore, we identified epididymal-specific genes that are associated with fertility between control and ACH groups using an Affymetrix Rat 230 2.0 oligo-microarray. Finally, we performed RT-PCR analysis for several differentially expressed genes to validate the alteration in gene expression observed by oligonucleotide microarray. Among all the differentially expressed genes, we analyzed and screened the down-regulated genes associated with metabolism processes, which are considered the major targets of ACH action. Simultaneously, the genes that were up-regulated by chlorohydrin were detected. The genes that negatively regulate sperm maturation and fertility include apoptosis and immune-related genes and have not been reported previously. The overall results of PCR analysis for selected genes were consistent with the array data. In this study, we have described the genome-wide profiles of gene expression in the epididymides of infertile rats induced by ACH, which could become potential epididymal specific targets for male contraception and infertility treatment.

Integrated genomic classification of melanocytic tumors of the central nervous system using mutation analysis, copy number alterations and DNA methylation profiling.

PubMed

Griewank, Klaus; Koelsche, Christian; van de Nes, Johannes A P; Schrimpf, Daniel; Gessi, Marco; Möller, Inga; Sucker, Antje; Scolyer, Richard A; Buckland, Michael E; Murali, Rajmohan; Pietsch, Torsten; von Deimling, Andreas; Schadendorf, Dirk

2018-06-11

In the central nervous system, distinguishing primary leptomeningeal melanocytic tumors from melanoma metastases and predicting their biological behavior solely using histopathologic criteria can be challenging. We aimed to assess the diagnostic and prognostic value of integrated molecular analysis. Targeted next-generation-sequencing, array-based genome-wide methylation analysis and BAP1 immunohistochemistry was performed on the largest cohort of central nervous system melanocytic tumors analyzed to date, incl. 47 primary tumors of the central nervous system, 16 uveal melanomas. 13 cutaneous melanoma metastasis and 2 blue nevus-like melanomas. Gene mutation, DNA-methylation and copy-number profiles were correlated with clinicopathological features. Combining mutation, copy-number and DNA-methylation profiles clearly distinguished cutaneous melanoma metastases from other melanocytic tumors. Primary leptomeningeal melanocytic tumors, uveal melanomas and blue nevus-like melanoma showed common DNA-methylation, copy-number alteration and gene mutation signatures. Notably, tumors demonstrating chromosome 3 monosomy and BAP1 alterations formed a homogeneous subset within this group. Integrated molecular profiling aids in distinguishing primary from metastatic melanocytic tumors of the central nervous system. Primary leptomeningeal melanocytic tumors, uveal melanoma and blue nevus-like melanoma share molecular similarity with chromosome 3 and BAP1 alterations markers of poor prognosis. Copyright ©2018, American Association for Cancer Research.

Novel Insights into DNA Methylation Features in Spermatozoa: Stability and Peculiarities

PubMed Central

Sayols, Sergi; Chianese, Chiara; Giachini, Claudia; Heyn, Holger; Esteller, Manel

2012-01-01

Data about the entire sperm DNA methylome are limited to two sperm donors whereas studies dealing with a greater number of subjects focused only on a few genes or were based on low resolution arrays. This implies that information about what we can consider as a normal sperm DNA methylome and whether it is stable among different normozoospermic individuals is still missing. The definition of the DNA methylation profile of normozoospermic men, the entity of inter-individual variability and the epigenetic characterization of quality-fractioned sperm subpopulations in the same subject (intra-individual variability) are relevant for a better understanding of pathological conditions. We addressed these questions by using the high resolution Infinium 450K methylation array and compared normal sperm DNA methylomes against somatic and cancer cells. Our study, based on the largest number of subjects (n = 8) ever considered for such a large number of CpGs (n = 487,517), provided clear evidence for i) a highly conserved DNA methylation profile among normozoospermic subjects; ii) a stable sperm DNA methylation pattern in different quality-fractioned sperm populations of the same individual. The latter finding is particularly relevant if we consider that different quality fractioned sperm subpopulations show differences in their structural features, metabolic and genomic profiles. We demonstrate, for the first time, that DNA methylation in normozoospermic men remains highly uniform regardless the quality of sperm subpopulations. In addition, our analysis provided both confirmatory and novel data concerning the sperm DNA methylome, including its peculiar features in respect to somatic and cancer cells. Our description about a highly polarized sperm DNA methylation profile, the clearly distinct genomic and functional organization of hypo- versus hypermethylated loci as well as the association of histone-enriched hypomethylated loci with embryonic development, which we now extended also to hypomethylated piRNAs-linked genes, provides solid basis for future basic and clinical research. PMID:23071498

Transcriptome Profiling of Lotus japonicus Roots During Arbuscular Mycorrhiza Development and Comparison with that of Nodulation

PubMed Central

Deguchi, Yuichi; Banba, Mari; Shimoda, Yoshikazu; Chechetka, Svetlana A.; Suzuri, Ryota; Okusako, Yasuhiro; Ooki, Yasuhiro; Toyokura, Koichi; Suzuki, Akihiro; Uchiumi, Toshiki; Higashi, Shiro; Abe, Mikiko; Kouchi, Hiroshi; Izui, Katsura; Hata, Shingo

2007-01-01

Abstract To better understand the molecular responses of plants to arbuscular mycorrhizal (AM) fungi, we analyzed the differential gene expression patterns of Lotus japonicus, a model legume, with the aid of a large-scale cDNA macroarray. Experiments were carried out considering the effects of contaminating microorganisms in the soil inoculants. When the colonization by AM fungi, i.e. Glomus mosseae and Gigaspora margarita, was well established, four cysteine protease genes were induced. In situ hybridization revealed that these cysteine protease genes were specifically expressed in arbuscule-containing inner cortical cells of AM roots. On the other hand, phenylpropanoid biosynthesis-related genes for phenylalanine ammonia-lyase (PAL), chalcone synthase, etc. were repressed in the later stage, although they were moderately up-regulated on the initial association with the AM fungus. Real-time RT–PCR experiments supported the array experiments. To further confirm the characteristic expression, a PAL promoter was fused with a reporter gene and introduced into L. japonicus, and then the transformants were grown with a commercial inoculum of G. mosseae. The reporter activity was augmented throughout the roots due to the presence of contaminating microorganisms in the inoculum. Interestingly, G. mosseae only colonized where the reporter activity was low. Comparison of the transcriptome profiles of AM roots and nitrogen-fixing root nodules formed with Mesorhizobium loti indicated that the PAL genes and other phenylpropanoid biosynthesis-related genes were similarly repressed in the two organs. PMID:17634281

Therapeutic effects of Euphorbia Pekinensis and Glycyrrhiza glabra on Hepatocellular Carcinoma Ascites Partially Via Regulating the Frk-Arhgdib-Inpp5d-Avpr2-Aqp4 Signal Axis

NASA Astrophysics Data System (ADS)

Zhang, Yanqiong; Yan, Chen; Li, Yuting; Mao, Xia; Tao, Weiwei; Tang, Yuping; Lin, Ya; Guo, Qiuyan; Duan, Jingao; Lin, Na

2017-02-01

To clarify unknown rationalities of herbaceous compatibility of Euphorbia Pekinensis (DJ) and Glycyrrhiza glabra (GC) acting on hepatocellular carcinoma (HCC) ascites, peritoneum transcriptomics profiling of 15 subjects, including normal control (Con), HCC ascites mouse model (Mod), DJ-alone, DJ/GC-synergy and DJ/GC-antagonism treatment groups were performed on OneArray platform, followed by differentially expressed genes (DEGs) screening. DEGs between Mod and Con groups were considered as HCC ascites-related genes, and those among different drug treatment and Mod groups were identified as DJ/GC-combination-related genes. Then, an interaction network of HCC ascites-related gene-DJ/GC combination-related gene-known therapeutic target gene for ascites was constructed. Based on nodes’ degree, closeness, betweenness and k-coreness, the Frk-Arhgdib-Inpp5d-Avpr2-Aqp4 axis with highly network topological importance was demonstrated to be a candidate target of DJ/GC combination acting on HCC ascites. Importantly, both qPCR and western blot analyses verified this regulatory effects based on HCC ascites mice in vivo and M-1 collecting duct cells in vitro. Collectively, different combination designs of DJ and GC may lead to synergistic or antagonistic effects on HCC ascites partially via regulating the Frk-Arhgdib-Inpp5d-Avpr2-Aqp4 axis, implying that global gene expression profiling combined with network analysis can offer an effective way to understand pharmacological mechanisms of traditional Chinese medicine prescriptions.

GeneChip{sup {trademark}} screening assay for cystic fibrosis mutations

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cronn, M.T.; Miyada, C.G.; Fucini, R.V.

1994-09-01

GeneChip{sup {trademark}} assays are based on high density, carefully designed arrays of short oligonucleotide probes (13-16 bases) built directly on derivatized silica substrates. DNA target sequence analysis is achieved by hybridizing fluorescently labeled amplification products to these arrays. Fluorescent hybridization signals located within the probe array are translated into target sequence information using the known probe sequence at each array feature. The mutation screening assay for cystic fibrosis includes sets of oligonucleotide probes designed to detect numerous different mutations that have been described in 14 exons and one intron of the CFTR gene. Each mutation site is addressed by amore » sub-array of at least 40 probe sequences, half designed to detect the wild type gene sequence and half designed to detect the reported mutant sequence. Hybridization with homozygous mutant, homozygous wild type or heterozygous targets results in distinctive hybridization patterns within a sub-array, permitting specific discrimination of each mutation. The GeneChip probe arrays are very small (approximately 1 cm{sup 2}). There miniature size coupled with their high information content make GeneChip probe arrays a useful and practical means for providing CF mutation analysis in a clinical setting.« less

Thyroid paraganglioma. Report of 3 cases and description of an immunohistochemical profile useful in the differential diagnosis with medullary thyroid carcinoma, based on complementary DNA array results.

PubMed

Castelblanco, Esmeralda; Gallel, Pilar; Ros, Susana; Gatius, Sonia; Valls, Joan; De-Cubas, Aguirre A; Maliszewska, Agnieszka; Yebra-Pimentel, M Teresa; Menarguez, Javier; Gamallo, Carlos; Opocher, Giuseppe; Robledo, Mercedes; Matias-Guiu, Xavier

2012-07-01

Thyroid paraganglioma is a rare disorder that sometimes poses problems in differential diagnosis with medullary thyroid carcinoma. So far, differential diagnosis is solved with the help of some markers that are frequently expressed in medullary thyroid carcinoma (thyroid transcription factor 1, calcitonin, and carcinoembryonic antigen). However, some of these markers are not absolutely specific of medullary thyroid carcinoma and may be expressed in other tumors. Here we report 3 new cases of thyroid paraganglioma and describe our strategy to design a diagnostic immunohistochemical battery. First, we performed a comparative analysis of the expression profile of head and neck paragangliomas and medullary thyroid carcinoma, obtained after complementary DNA array analysis of 2 series of fresh-frozen samples of paragangliomas and medullary thyroid carcinoma, respectively. Seven biomarkers showing differential expression were selected (nicotinamide adenine dinucleotide dehydrogenase 1 alpha subcomplex, 4-like 2, NDUFA4L2; cytochrome c oxidase subunit IV isoform 2; vesicular monoamine transporter 2; calcitonin gene-related protein/calcitonin; carcinoembryonic antigen; and thyroid transcription factor 1) for immunohistochemical analysis. Two tissue microarrays were constructed from 2 different series of paraffin-embedded samples of paragangliomas and medullary thyroid carcinoma. We provide a classifying rule for differential diagnosis that combines negativity or low staining for calcitonin gene-related protein (histologic score, <10) or calcitonin (histologic score, <50) together with positivity of any of NADH dehydrogenase 1 alpha subcomplex, 4-like 2; cytochrome c oxidase subunit IV isoform 2; or vesicular monoamine transporter 2 to predict paragangliomas, showing a prediction error of 0%. Finally, the immunohistochemical battery was checked in paraffin-embedded blocks from 4 examples of thyroid paraganglioma (1 previously reported case and 3 new cases), showing also a prediction error of 0%. Our results suggest that the comparative expression profile, obtained by complementary DNA arrays, seems to be a good tool to design immunohistochemical batteries used in differential diagnosis. Copyright © 2012 Elsevier Inc. All rights reserved.

Changes in the transcriptional profile in response to overexpression of the osteopontin-c splice isoform in ovarian (OvCar-3) and prostate (PC-3) cancer cell lines

PubMed Central

2014-01-01

Background Especially in human tumor cells, the osteopontin (OPN) primary transcript is subject to alternative splicing, generating three isoforms termed OPNa, OPNb and OPNc. We previously demonstrated that the OPNc splice variant activates several aspects of the progression of ovarian and prostate cancers. The goal of the present study was to develop cell line models to determine the impact of OPNc overexpression on main cancer signaling pathways and thus obtain insights into the mechanisms of OPNc pro-tumorigenic roles. Methods Human ovarian and prostate cancer cell lines, OvCar-3 and PC-3 cells, respectively, were stably transfected to overexpress OPNc. Transcriptomic profiling was performed on these cells and compared to controls, to identify OPNc overexpression-dependent changes in gene expression levels and pathways by qRT-PCR analyses. Results Among 84 genes tested by using a multiplex real-time PCR Cancer Pathway Array approach, 34 and 16, respectively, were differentially expressed between OvCar-3 and PC-3 OPNc-overexpressing cells in relation to control clones. Differentially expressed genes are included in all main hallmarks of cancer, and several interacting proteins have been identified using an interactome network analysis. Based on marked up-regulation of Vegfa transcript in response to OPNc overexpression, we partially validated the array data by demonstrating that conditioned medium (CM) secreted from OvCar-3 and PC-3 OPNc-overexpressing cells significantly induced endothelial cell adhesion, proliferation and migration, compared to CM secreted from control cells. Conclusions Overall, the present study elucidated transcriptional changes of OvCar-3 and PC-3 cancer cell lines in response to OPNc overexpression, which provides an assessment for predicting the molecular mechanisms by which this splice variant promotes tumor progression features. PMID:24928374

Partial Least Squares Based Gene Expression Analysis in EBV- Positive and EBV-Negative Posttransplant Lymphoproliferative Disorders.

PubMed

Wu, Sa; Zhang, Xin; Li, Zhi-Ming; Shi, Yan-Xia; Huang, Jia-Jia; Xia, Yi; Yang, Hang; Jiang, Wen-Qi

2013-01-01

Post-transplant lymphoproliferative disorder (PTLD) is a common complication of therapeutic immunosuppression after organ transplantation. Gene expression profile facilitates the identification of biological difference between Epstein-Barr virus (EBV) positive and negative PTLDs. Previous studies mainly implemented variance/regression analysis without considering unaccounted array specific factors. The aim of this study is to investigate the gene expression difference between EBV positive and negative PTLDs through partial least squares (PLS) based analysis. With a microarray data set from the Gene Expression Omnibus database, we performed PLS based analysis. We acquired 1188 differentially expressed genes. Pathway and Gene Ontology enrichment analysis identified significantly over-representation of dysregulated genes in immune response and cancer related biological processes. Network analysis identified three hub genes with degrees higher than 15, including CREBBP, ATXN1, and PML. Proteins encoded by CREBBP and PML have been reported to be interact with EBV before. Our findings shed light on expression distinction of EBV positive and negative PTLDs with the hope to offer theoretical support for future therapeutic study.

Evolutionary characterization and transcript profiling of β-tubulin genes in flax (Linum usitatissimum L.) during plant development.

PubMed

Gavazzi, Floriana; Pigna, Gaia; Braglia, Luca; Gianì, Silvia; Breviario, Diego; Morello, Laura

2017-12-08

Microtubules, polymerized from alpha and beta-tubulin monomers, play a fundamental role in plant morphogenesis, determining the cell division plane, the direction of cell expansion and the deposition of cell wall material. During polarized pollen tube elongation, microtubules serve as tracks for vesicular transport and deposition of proteins/lipids at the tip membrane. Such functions are controlled by cortical microtubule arrays. Aim of this study was to first characterize the flax β-tubulin family by sequence and phylogenetic analysis and to investigate differential expression of β-tubulin genes possibly related to fibre elongation and to flower development. We report the cloning and characterization of the complete flax β-tubulin gene family: exon-intron organization, duplicated gene comparison, phylogenetic analysis and expression pattern during stem and hypocotyl elongation and during flower development. Sequence analysis of the fourteen expressed β-tubulin genes revealed that the recent whole genome duplication of the flax genome was followed by massive retention of duplicated tubulin genes. Expression analysis showed that β-tubulin mRNA profiles gradually changed along with phloem fibre development in both the stem and hypocotyl. In flowers, changes in relative tubulin transcript levels took place at anthesis in anthers, but not in carpels. Phylogenetic analysis supports the origin of extant plant β-tubulin genes from four ancestral genes pre-dating angiosperm separation. Expression analysis suggests that particular tubulin subpopulations are more suitable to sustain different microtubule functions such as cell elongation, cell wall thickening or pollen tube growth. Tubulin genes possibly related to different microtubule functions were identified as candidate for more detailed studies.

Genome-wide transcriptional profiling of human glioblastoma cells in response to ITE treatment

PubMed Central

Kang, Bo; Zhou, Yanwen; Zheng, Min; Wang, Ying-Jie

2015-01-01

A ligand-activated transcription factor aryl hydrocarbon receptor (AhR) is recently revealed to play a key role in embryogenesis and tumorigenesis (Feng et al. [1], Safe et al. [2]) and 2-(1′H-indole-3′-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE) (Song et al. [3]) is an endogenous AhR ligand that possesses anti-tumor activity. In order to gain insights into how ITE acts via the AhR in embryogenesis and tumorigenesis, we analyzed the genome-wide transcriptional profiles of the following three groups of cells: the human glioblastoma U87 parental cells, U87 tumor sphere cells treated with vehicle (DMSO) and U87 tumor sphere cells treated with ITE. Here, we provide the details of the sample gathering strategy and show the quality controls and the analyses associated with our gene array data deposited into the Gene Expression Omnibus (GEO) under the accession code of GSE67986. PMID:26484269

SBR-Blood: systems biology repository for hematopoietic cells.

PubMed

Lichtenberg, Jens; Heuston, Elisabeth F; Mishra, Tejaswini; Keller, Cheryl A; Hardison, Ross C; Bodine, David M

2016-01-04

Extensive research into hematopoiesis (the development of blood cells) over several decades has generated large sets of expression and epigenetic profiles in multiple human and mouse blood cell types. However, there is no single location to analyze how gene regulatory processes lead to different mature blood cells. We have developed a new database framework called hematopoietic Systems Biology Repository (SBR-Blood), available online at http://sbrblood.nhgri.nih.gov, which allows user-initiated analyses for cell type correlations or gene-specific behavior during differentiation using publicly available datasets for array- and sequencing-based platforms from mouse hematopoietic cells. SBR-Blood organizes information by both cell identity and by hematopoietic lineage. The validity and usability of SBR-Blood has been established through the reproduction of workflows relevant to expression data, DNA methylation, histone modifications and transcription factor occupancy profiles. Published by Oxford University Press on behalf of Nucleic Acids Research 2015. This work is written by (a) US Government employee(s) and is in the public domain in the US.

Effective formation method for an aspherical microlens array based on an aperiodic moving mask during exposure.

PubMed

Shi, Lifang; Du, Chunlei; Dong, Xiaochun; Deng, Qiling; Luo, Xiangang

2007-12-01

An aperiodic mask design method for fabricating a microlens array with an aspherical profile is proposed. The nonlinear relationship between exposure doses and lens profile is considered, and the select criteria of quantization interval and fabrication range of the method are given. The mask function of a quadrangle microlens array with a hyperboloid profile used in the infrared was constructed by using this method. The microlens array can be effectively fabricated during a one time exposure process using the mask. Reactive ion etching was carried out to transfer the structure into the substrate of germanium. The measurement results indicate that the roughness is less than 10 nm (pv), and the profile error is less than 40 nm (rms).

Statistical Analysis of Microarray Data with Replicated Spots: A Case Study with Synechococcus WH8102

PubMed Central

Thomas, E. V.; Phillippy, K. H.; Brahamsha, B.; Haaland, D. M.; Timlin, J. A.; Elbourne, L. D. H.; Palenik, B.; Paulsen, I. T.

2009-01-01

Until recently microarray experiments often involved relatively few arrays with only a single representation of each gene on each array. A complete genome microarray with multiple spots per gene (spread out spatially across the array) was developed in order to compare the gene expression of a marine cyanobacterium and a knockout mutant strain in a defined artificial seawater medium. Statistical methods were developed for analysis in the special situation of this case study where there is gene replication within an array and where relatively few arrays are used, which can be the case with current array technology. Due in part to the replication within an array, it was possible to detect very small changes in the levels of expression between the wild type and mutant strains. One interesting biological outcome of this experiment is the indication of the extent to which the phosphorus regulatory system of this cyanobacterium affects the expression of multiple genes beyond those strictly involved in phosphorus acquisition. PMID:19404483

Statistical Analysis of Microarray Data with Replicated Spots: A Case Study with Synechococcus WH8102

DOE PAGES

Thomas, E. V.; Phillippy, K. H.; Brahamsha, B.; ...

2009-01-01

Until recently microarray experiments often involved relatively few arrays with only a single representation of each gene on each array. A complete genome microarray with multiple spots per gene (spread out spatially across the array) was developed in order to compare the gene expression of a marine cyanobacterium and a knockout mutant strain in a defined artificial seawater medium. Statistical methods were developed for analysis in the special situation of this case study where there is gene replication within an array and where relatively few arrays are used, which can be the case with current array technology. Due in partmore » to the replication within an array, it was possible to detect very small changes in the levels of expression between the wild type and mutant strains. One interesting biological outcome of this experiment is the indication of the extent to which the phosphorus regulatory system of this cyanobacterium affects the expression of multiple genes beyond those strictly involved in phosphorus acquisition.« less

Gene expression profiling during asexual development of the late blight pathogen Phytophthora infestans reveals a highly dynamic transcriptome.

PubMed

Judelson, Howard S; Ah-Fong, Audrey M V; Aux, George; Avrova, Anna O; Bruce, Catherine; Cakir, Cahid; da Cunha, Luis; Grenville-Briggs, Laura; Latijnhouwers, Maita; Ligterink, Wilco; Meijer, Harold J G; Roberts, Samuel; Thurber, Carrie S; Whisson, Stephen C; Birch, Paul R J; Govers, Francine; Kamoun, Sophien; van West, Pieter; Windass, John

2008-04-01

Much of the pathogenic success of Phytophthora infestans, the potato and tomato late blight agent, relies on its ability to generate from mycelia large amounts of sporangia, which release zoospores that encyst and form infection structures. To better understand these stages, Affymetrix GeneChips based on 15,650 unigenes were designed and used to profile the life cycle. Approximately half of P. infestans genes were found to exhibit significant differential expression between developmental transitions, with approximately (1)/(10) being stage-specific and most changes occurring during zoosporogenesis. Quantitative reverse-transcription polymerase chain reaction assays confirmed the robustness of the array results and showed that similar patterns of differential expression were obtained regardless of whether hyphae were from laboratory media or infected tomato. Differentially expressed genes encode potential cellular regulators, especially protein kinases; metabolic enzymes such as those involved in glycolysis, gluconeogenesis, or the biosynthesis of amino acids or lipids; regulators of DNA synthesis; structural proteins, including predicted flagellar proteins; and pathogenicity factors, including cell-wall-degrading enzymes, RXLR effector proteins, and enzymes protecting against plant defense responses. Curiously, some stage-specific transcripts do not appear to encode functional proteins. These findings reveal many new aspects of oomycete biology, as well as potential targets for crop protection chemicals.

Gene expression profiles reveal that DCN, DIO1, and DIO2 are underexpressed in benign and malignant thyroid tumors.

PubMed

Arnaldi, L A T; Borra, R C; Maciel, R M B; Cerutti, J M

2005-03-01

To investigate the molecular events involved in the pathogenesis and/or progression of thyroid tumors, we compared the gene expression profiles of three thyroid carcinoma cell lines, which represent major tumor subtypes of thyroid cancer and normal thyroid tissue. Using cDNA array methodology, we investigated the expression of 1807 open reading frame expressed sequence tags (ORESTES), selected from head and neck tumor libraries generated through the Brazilian Human Cancer Project-LICR/FAPESP. We found that 505 transcripts were differentially expressed in the thyroid carcinoma cell lines. Using a more stringent criterion, transcripts underexpressed or overexpressed more than fivefold in 1 of 3 or 3 of 3 carcinoma cell lines, a list of 55 ESTs were detected. Five candidate genes were further validated by quantitative polymerase chain reaction (qPCR) in an independent set of 52 thyroid tumors and 22 matched normal thyroid tissues. DCN was found underexpressed in a high percentage of the follicular thyroid adenomas, follicular thyroid carcinomas, and follicular variant of papillary thyroid carcinomas. DIO1 and DIO2 were underexpressed in nearly all papillary thyroid carcinomas. These genes not only could help to better define a tumor signature for thyroid tumors, but may, in part, also become useful as potential targets for thyroid tumor treatment.

TET2 Mutations Are Associated with Specific 5-Methylcytosine and 5-Hydroxymethylcytosine Profiles in Patients with Chronic Myelomonocytic Leukemia

PubMed Central

Pérez, Cristina; Martínez-Calle, Nicolas; Martín-Subero, José Ignacio; Segura, Victor; Delabesse, Eric; Fernandez-Mercado, Marta; Garate, Leire; Alvarez, Sara; Rifon, José; Varea, Sara; Boultwood, Jacqueline; Wainscoat, James S.; Cigudosa, Juan Cruz; Calasanz, María José; Cross, Nicholas C. P.

2012-01-01

Chronic myelomonocytic leukemia (CMML) has recently been associated with a high incidence of diverse mutations in genes such as TET2 or EZH2 that are implicated in epigenetic mechanisms. We have performed genome-wide DNA methylation arrays and mutational analysis of TET2, IDH1, IDH2, EZH2 and JAK2 in a group of 24 patients with CMML. 249 genes were differentially methylated between CMML patients and controls. Using Ingenuity pathway analysis, we identified enrichment in a gene network centered around PLC, JNK and ERK suggesting that these pathways, whose deregulation has beenrecently described in CMML, are affected by epigenetic mechanisms. Mutations of TET2, JAK2 and EZH2 were found in 15 patients (65%), 4 patients (17%) and 1 patient (4%) respectively while no mutations in the IDH1 and IDH2 genes were identified. Interestingly, patients with wild type TET2 clustered separately from patients with TET2 mutations, showed a higher degree of hypermethylation and were associated with higher risk karyotypes. Our results demonstrate the presence of aberrant DNA methylation in CMML and identifies TET2 mutant CMML as a biologically distinct disease subtype with a different epigenetic profile. PMID:22328940

Molecular Classifiers for Acute Kidney Transplant Rejection in Peripheral Blood by Whole Genome Gene Expression Profiling

PubMed Central

Kurian, S. M.; Williams, A. N.; Gelbart, T.; Campbell, D.; Mondala, T. S.; Head, S. R.; Horvath, S.; Gaber, L.; Thompson, R.; Whisenant, T.; Lin, W.; Langfelder, P.; Robison, E. H.; Schaffer, R. L.; Fisher, J. S.; Friedewald, J.; Flechner, S. M.; Chan, L. K.; Wiseman, A. C.; Shidban, H.; Mendez, R.; Heilman, R.; Abecassis, M. M.; Marsh, C. L.; Salomon, D. R.

2015-01-01

There are no minimally invasive diagnostic metrics for acute kidney transplant rejection (AR), especially in the setting of the common confounding diagnosis, acute dysfunction with no rejection (ADNR). Thus, though kidney transplant biopsies remain the gold standard, they are invasive, have substantial risks, sampling error issues and significant costs and are not suitable for serial monitoring. Global gene expression profiles of 148 peripheral blood samples from transplant patients with excellent function and normal histology (TX; n = 46), AR (n = 63) and ADNR (n = 39), from two independent cohorts were analyzed with DNA microarrays. We applied a new normalization tool, frozen robust multi-array analysis, particularly suitable for clinical diagnostics, multiple prediction tools to discover, refine and validate robust molecular classifiers and we tested a novel one-by-one analysis strategy to model the real clinical application of this test. Multiple three-way classifier tools identified 200 highest value probesets with sensitivity, specificity, positive predictive value, negative predictive value and area under the curve for the validation cohort ranging from 82% to 100%, 76% to 95%, 76% to 95%, 79% to 100%, 84% to 100% and 0.817 to 0.968, respectively. We conclude that peripheral blood gene expression profiling can be used as a minimally invasive tool to accurately reveal TX, AR and ADNR in the setting of acute kidney transplant dysfunction. PMID:24725967

Hepatic CD36 downregulation parallels steatosis improvement in morbidly obese undergoing bariatric surgery.

PubMed

Pardina, E; Ferrer, R; Rossell, J; Ricart-Jané, D; Méndez-Lara, K A; Baena-Fustegueras, J A; Lecube, A; Julve, J; Peinado-Onsurbe, J

2017-09-01

The notion that hepatic expression of genes involved in lipid metabolism is altered in obese patients is relatively new and its relationship with hepatic steatosis and cardiometabolic alterations remains unclear. We assessed the impact of Roux-en-Y gastric bypass surgery (RYGB) on the expression profile of genes related to metabolic syndrome in liver biopsies from morbidly obese individuals using a custom-made, focused cDNA microarray, and assessed the relationship between the expression profile and hepatic steatosis regression. Plasma and liver samples were obtained from patients at baseline and 12 months after surgery. Samples were assayed for chemical and gene expression analyses, as appropriate. Gene expression profiles were assessed using custom-made, focused TaqMan low-density array cards. RYGB-induced weight loss produced a favorable reduction in fat deposits, insulin resistance (estimated by homeostasis model assessment of insulin resistance (HOMA-IR)), and plasma and hepatic lipid levels. Compared with the baseline values, the gene expression levels of key targets of lipid metabolism were significantly altered: CD36 was significantly downregulated (-40%; P=0.001), whereas APOB (+27%; P=0.032) and SCARB1 (+37%; P=0.040) were upregulated in response to surgery-induced weight reduction. We also observed a favorable reduction in the expression of the PAI1 gene (-80%; P=0.007) and a significant increase in the expression of the PPARA (+60%; P=0.014) and PPARGC1 genes (+36%; P=0.015). Notably, the relative fold decrease in the expression of the CD36 gene was directly associated with a concomitant reduction in the cholesterol (Spearman's r=0.92; P=0.001) and phospholipid (Spearman's r=0.76; P=0.04) contents in this tissue. For the first time, RYGB-induced weight loss was shown to promote a favorable downregulation of CD36 expression, which was proportional to a favorable reduction in the hepatic cholesterol and phospholipid contents in our morbidly obese subjects following surgery.

Gene profiling, biomarkers and pathways characterizing HCV-related hepatocellular carcinoma

PubMed Central

De Giorgi, Valeria; Monaco, Alessandro; Worchech, Andrea; Tornesello, MariaLina; Izzo, Francesco; Buonaguro, Luigi; Marincola, Francesco M; Wang, Ena; Buonaguro, Franco M

2009-01-01

Background Hepatitis C virus (HCV) infection is a major cause of hepatocellular carcinoma (HCC) worldwide. The molecular mechanisms of HCV-induced hepatocarcinogenesis are not yet fully elucidated. Besides indirect effects as tissue inflammation and regeneration, a more direct oncogenic activity of HCV can be postulated leading to an altered expression of cellular genes by early HCV viral proteins. In the present study, a comparison of gene expression patterns has been performed by microarray analysis on liver biopsies from HCV-positive HCC patients and HCV-negative controls. Methods Gene expression profiling of liver tissues has been performed using a high-density microarray containing 36'000 oligos, representing 90% of the human genes. Samples were obtained from 14 patients affected by HCV-related HCC and 7 HCV-negative non-liver-cancer patients, enrolled at INT in Naples. Transcriptional profiles identified in liver biopsies from HCC nodules and paired non-adjacent non-HCC liver tissue of the same HCV-positive patients were compared to those from HCV-negative controls by the Cluster program. The pathway analysis was performed using the BRB-Array- Tools based on the "Ingenuity System Database". Significance threshold of t-test was set at 0.001. Results Significant differences were found between the expression patterns of several genes falling into different metabolic and inflammation/immunity pathways in HCV-related HCC tissues as well as the non-HCC counterpart compared to normal liver tissues. Only few genes were found differentially expressed between HCV-related HCC tissues and paired non-HCC counterpart. Conclusion In this study, informative data on the global gene expression pattern of HCV-related HCC and non-HCC counterpart, as well as on their difference with the one observed in normal liver tissues have been obtained. These results may lead to the identification of specific biomarkers relevant to develop tools for detection, diagnosis, and classification of HCV-related HCC. PMID:19821982

Identification of Chemical-Genetic Interactions via Parallel Analysis of Barcoded Yeast Strains.

PubMed

Suresh, Sundari; Schlecht, Ulrich; Xu, Weihong; Miranda, Molly; Davis, Ronald W; Nislow, Corey; Giaever, Guri; St Onge, Robert P

2016-09-01

The Yeast Knockout Collection is a complete set of gene deletion strains for the budding yeast, Saccharomyces cerevisiae In each strain, one of approximately 6000 open-reading frames is replaced with a dominant selectable marker flanked by two DNA barcodes. These barcodes, which are unique to each gene, allow the growth of thousands of strains to be individually measured from a single pooled culture. The collection, and other resources that followed, has ushered in a new era in chemical biology, enabling unbiased and systematic identification of chemical-genetic interactions (CGIs) with remarkable ease. CGIs link bioactive compounds to biological processes, and hence can reveal the mechanism of action of growth-inhibitory compounds in vivo, including those of antifungal, antibiotic, and anticancer drugs. The chemogenomic profiling method described here measures the sensitivity induced in yeast heterozygous and homozygous deletion strains in the presence of a chemical inhibitor of growth (termed haploinsufficiency profiling and homozygous profiling, respectively, or HIPHOP). The protocol is both scalable and amenable to automation. After competitive growth of yeast knockout collection cultures, with and without chemical inhibitors, CGIs can be identified and quantified using either array- or sequencing-based approaches as described here. © 2016 Cold Spring Harbor Laboratory Press.

SNP array profiling of childhood adrenocortical tumors reveals distinct pathways of tumorigenesis and highlights candidate driver genes.

PubMed

Letouzé, Eric; Rosati, Roberto; Komechen, Heloisa; Doghman, Mabrouka; Marisa, Laetitia; Flück, Christa; de Krijger, Ronald R; van Noesel, Max M; Mas, Jean-Christophe; Pianovski, Mara A D; Zambetti, Gerard P; Figueiredo, Bonald C; Lalli, Enzo

2012-07-01

Childhood adrenocortical tumors (ACT) are rare malignancies, except in southern Brazil, where a higher incidence rate is associated to a high frequency of the founder R337H TP53 mutation. To date, copy number alterations in these tumors have only been analyzed by low-resolution comparative genomic hybridization. We analyzed an international series of 25 childhood ACT using high-resolution single nucleotide polymorphism arrays to: 1) detect focal copy number alterations highlighting candidate driver genes; and 2) compare genetic alterations between Brazilian patients carrying the R337H TP53 mutation and non-Brazilian patients. We identified 16 significantly recurrent chromosomal alterations (q-value < 0.05), the most frequent being -4q34, +9q33-q34, +19p, loss of heterozygosity (LOH) of chromosome 17 and 11p15. Focal amplifications and homozygous deletions comprising well-known oncogenes (MYC, MDM2, PDGFRA, KIT, MCL1, BCL2L1) and tumor suppressors (TP53, RB1, RPH3AL) were identified. In addition, eight focal deletions were detected at 4q34, defining a sharp peak region around the noncoding RNA LINC00290 gene. Although non-Brazilian tumors with a mutated TP53 were similar to Brazilian tumors, those with a wild-type TP53 displayed distinct genomic profiles, with significantly fewer rearrangements (P = 0.019). In particular, three alterations (LOH of chromosome 17, +9q33-q34, and -4q34) were significantly more frequent in TP53-mutated samples. Finally, two of four TP53 wild-type tumors displayed as sole rearrangement a copy-neutral LOH of the imprinted region at 11p15, supporting a major role for this region in ACT development. Our findings highlight potential driver genes and cellular pathways implicated in childhood ACT and demonstrate the existence of different oncogenic routes in this pathology.

Altered gene expression changes in Arabidopsis leaf tissues and protoplasts in response to Plum pox virus infection

PubMed Central

Babu, Mohan; Griffiths, Jonathan S; Huang, Tyng-Shyan; Wang, Aiming

2008-01-01

Background Virus infection induces the activation and suppression of global gene expression in the host. Profiling gene expression changes in the host may provide insights into the molecular mechanisms that underlie host physiological and phenotypic responses to virus infection. In this study, the Arabidopsis Affymetrix ATH1 array was used to assess global gene expression changes in Arabidopsis thaliana plants infected with Plum pox virus (PPV). To identify early genes in response to PPV infection, an Arabidopsis synchronized single-cell transformation system was developed. Arabidopsis protoplasts were transfected with a PPV infectious clone and global gene expression changes in the transfected protoplasts were profiled. Results Microarray analysis of PPV-infected Arabidopsis leaf tissues identified 2013 and 1457 genes that were significantly (Q ≤ 0.05) up- (≥ 2.5 fold) and downregulated (≤ -2.5 fold), respectively. Genes associated with soluble sugar, starch and amino acid, intracellular membrane/membrane-bound organelles, chloroplast, and protein fate were upregulated, while genes related to development/storage proteins, protein synthesis and translation, and cell wall-associated components were downregulated. These gene expression changes were associated with PPV infection and symptom development. Further transcriptional profiling of protoplasts transfected with a PPV infectious clone revealed the upregulation of defence and cellular signalling genes as early as 6 hours post transfection. A cross sequence comparison analysis of genes differentially regulated by PPV-infected Arabidopsis leaves against uniEST sequences derived from PPV-infected leaves of Prunus persica, a natural host of PPV, identified orthologs related to defence, metabolism and protein synthesis. The cross comparison of genes differentially regulated by PPV infection and by the infections of other positive sense RNA viruses revealed a common set of 416 genes. These identified genes, particularly the early responsive genes, may be critical in virus infection. Conclusion Gene expression changes in PPV-infected Arabidopsis are the molecular basis of stress and defence-like responses, PPV pathogenesis and symptom development. The differentially regulated genes, particularly the early responsive genes, and a common set of genes regulated by infections of PPV and other positive sense RNA viruses identified in this study are candidates suitable for further functional characterization to shed lights on molecular virus-host interactions. PMID:18613973

Microarray analysis of laser capture microdissected-anulus cells from the human intervertebral disc.

PubMed

Gruber, Helen E; Mougeot, Jean-Luc; Hoelscher, Gretchen; Ingram, Jane A; Hanley, Edward N

2007-05-15

Five Thompson Grade I/II discs (Group 1), 7 Grade III discs (Group 2), and 3 Grade IV discs (Group IV) were studied here in a project approved by the authors' Human Subjects Institutional Review Board. Our objective was to use laser capture microdissection (LCM) to harvest cells from the human anulus and to derive gene expression profiles using microarray analysis. Appropriate gene expression is essential in the intervertebral disc for maintenance of extracellular matrix (ECM), ECM remodeling, and maintenance of a viable disc cell population. During disc degeneration, cell numbers drop, making gene expression studies challenging. LCM was used to harvest cells from paraffin-embedded sections of human anulus tissue. Gene profiling used Affymetrix GeneChip Human X3P arrays. ANOVA and SAM permutation analysis were applied to dCHIP normalized, filtered, and log-transformed gene expression data ( approximately 33,500 probes), and data analyzed to identify genes that were significantly differentially expressed between the 3 groups. We identified 47 genes that were significantly differentially expressed between the 3 groups (P < 0.001 and lowest q values). Compared with the healthiest discs (Grade I/II), 13 genes were up-regulated and 19 down-regulated in both the Grade III and the Grade IV discs. Genes with biologic significance regulated during degeneration involved cell senescence, low cell division rates, hypoxia-related genes, heat-shock protein 70 interacting protein, neuropilin 2, and interleukin-23p19 (interleukin-12 family). Results expand our understanding of disc aging and degeneration and show that LCM is a valuable technique that can be used to collect mRNA amounts adequate for microarray analysis from the sparse cell population of the human anulus.

CLL Cells Respond to B-Cell Receptor Stimulation with a MicroRNA/mRNA Signature Associated with MYC Activation and Cell Cycle Progression

PubMed Central

Pede, Valerie; Rombout, Ans; Vermeire, Jolien; Naessens, Evelien; Mestdagh, Pieter; Robberecht, Nore; Vanderstraeten, Hanne; Van Roy, Nadine; Vandesompele, Jo; Speleman, Frank; Philippé, Jan; Verhasselt, Bruno

2013-01-01

Chronic lymphocytic leukemia (CLL) is a disease with variable clinical outcome. Several prognostic factors such as the immunoglobulin heavy chain variable genes (IGHV) mutation status are linked to the B-cell receptor (BCR) complex, supporting a role for triggering the BCR in vivo in the pathogenesis. The miRNA profile upon stimulation and correlation with IGHV mutation status is however unknown. To evaluate the transcriptional response of peripheral blood CLL cells upon BCR stimulation in vitro, miRNA and mRNA expression was measured using hybridization arrays and qPCR. We found both IGHV mutated and unmutated CLL cells to respond with increased expression of MYC and other genes associated with BCR activation, and a phenotype of cell cycle progression. Genome-wide expression studies showed hsa-miR-132-3p/hsa-miR-212 miRNA cluster induction associated with a set of downregulated genes, enriched for genes modulated by BCR activation and amplified by Myc. We conclude that BCR triggering of CLL cells induces a transcriptional response of genes associated with BCR activation, enhanced cell cycle entry and progression and suggest that part of the transcriptional profiles linked to IGHV mutation status observed in isolated peripheral blood are not cell intrinsic but rather secondary to in vivo BCR stimulation. PMID:23560086

Coexposure of mice to trovafloxacin and lipopolysaccharide, a model of idiosyncratic hepatotoxicity, results in a unique gene expression profile and interferon gamma-dependent liver injury.

PubMed

Shaw, Patrick J; Ditewig, Amy C; Waring, Jeffrey F; Liguori, Michael J; Blomme, Eric A; Ganey, Patricia E; Roth, Robert A

2009-01-01

The antibiotic trovafloxacin (TVX) has caused severe idiosyncratic hepatotoxicity in people, whereas levofloxacin (LVX) has not. Mice cotreated with TVX and lipopolysaccharide (LPS), but not with LVX and LPS, develop severe hepatocellular necrosis. Mice were treated with TVX and/or LPS, and hepatic gene expression changes were measured before liver injury using gene array. Hepatic gene expression profiles from mice treated with TVX/LPS clustered differently from those treated with LPS or TVX alone. Several of the probe sets expressed differently in TVX/LPS-treated mice were involved in interferon (IFN) signaling and the janus kinase/signal transducers and activators of transcription (JAK/STAT) pathway. A time course of plasma concentrations of IFN-gamma and interleukin (IL)-18, which directly induces IFN-gamma production, revealed that both cytokines were selectively increased in TVX/LPS-treated mice. Both IL-18(-/-) and IFN-gamma(-/-) mice were significantly protected from TVX/LPS-induced liver injury. In addition, IFN-gamma(-/-) mice had decreased plasma concentrations of tumor necrosis factor-alpha, IL-18, and IL-1beta when compared to wild-type mice. In conclusion, the altered expression of genes involved in IFN signaling in TVX/LPS-treated mice led to the finding that IL-18 and IFN-gamma play a critical role in TVX/LPS-induced liver injury.

Systematic evaluation of RNA quality, microarray data reliability and pathway analysis in fresh, fresh frozen and formalin-fixed paraffin-embedded tissue samples.

PubMed

Wimmer, Isabella; Tröscher, Anna R; Brunner, Florian; Rubino, Stephen J; Bien, Christian G; Weiner, Howard L; Lassmann, Hans; Bauer, Jan

2018-04-20

Formalin-fixed paraffin-embedded (FFPE) tissues are valuable resources commonly used in pathology. However, formalin fixation modifies nucleic acids challenging the isolation of high-quality RNA for genetic profiling. Here, we assessed feasibility and reliability of microarray studies analysing transcriptome data from fresh, fresh-frozen (FF) and FFPE tissues. We show that reproducible microarray data can be generated from only 2 ng FFPE-derived RNA. For RNA quality assessment, fragment size distribution (DV200) and qPCR proved most suitable. During RNA isolation, extending tissue lysis time to 10 hours reduced high-molecular-weight species, while additional incubation at 70 °C markedly increased RNA yields. Since FF- and FFPE-derived microarrays constitute different data entities, we used indirect measures to investigate gene signal variation and relative gene expression. Whole-genome analyses revealed high concordance rates, while reviewing on single-genes basis showed higher data variation in FFPE than FF arrays. Using an experimental model, gene set enrichment analysis (GSEA) of FFPE-derived microarrays and fresh tissue-derived RNA-Seq datasets yielded similarly affected pathways confirming the applicability of FFPE tissue in global gene expression analysis. Our study provides a workflow comprising RNA isolation, quality assessment and microarray profiling using minimal RNA input, thus enabling hypothesis-generating pathway analyses from limited amounts of precious, pathologically significant FFPE tissues.

Human adaptation genetic response suites: Toward new interventions and countermeasures for spaceflight

NASA Astrophysics Data System (ADS)

Sundaresan, A.; Pellis, N. R.

2005-08-01

Genetic response suites in human lymphocytes in response to microgravity are important to identify and further study in order to augment human physiological adaptation to novel environments. Emerging technologies, such as DNA micro array profiling, have the potential to identify novel genes that are involved in mediating adaptation to these environments. These genes may prove to be therapeutically valuable as new targets for countermeasures, or as predictive biomarkers of response to these new environments. Human lymphocytes cultured in 1g and microgravity analog culture were analyzed for their differential gene expression response. Different groups of genes related to the immune response, cardiovascular system and stress response were then analyzed. Analysis of cells from multiple donors reveals a small shared set that are likely to be essential to adaptation. These three groups focus on human adaptation to new environments. The shared set contains genes related to T cell activation, immune response and stress response to analog microgravity.

Effects of simulated microgravity on gene expression and biological phenotypes of a single generation Caenorhabditis elegans cultured on 2 different media

NASA Astrophysics Data System (ADS)

Tee, Ling Fei; Neoh, Hui-min; Then, Sue Mian; Murad, Nor Azian; Asillam, Mohd Fairos; Hashim, Mohd Helmy; Nathan, Sheila; Jamal, Rahman

2017-11-01

Studies of multigenerational Caenorhabditis elegans exposed to long-term spaceflight have revealed expression changes of genes involved in longevity, DNA repair, and locomotion. However, results from spaceflight experiments are difficult to reproduce as space missions are costly and opportunities are rather limited for researchers. In addition, multigenerational cultures of C. elegans used in previous studies contribute to mixture of gene expression profiles from both larvae and adult worms, which were recently reported to be different. Usage of different culture media during microgravity simulation experiments might also give rise to differences in the gene expression and biological phenotypes of the worms. In this study, we investigated the effects of simulated microgravity on the gene expression and biological phenotype profiles of a single generation of C. elegans worms cultured on 2 different culture media. A desktop Random Positioning Machine (RPM) was used to simulate microgravity on the worms for approximately 52 to 54 h. Gene expression profile was analysed using the Affymetrix GeneChip® C. elegans 1.0 ST Array. Only one gene (R01H2.2) was found to be downregulated in nematode growth medium (NGM)-cultured worms exposed to simulated microgravity. On the other hand, eight genes were differentially expressed for C. elegans Maintenance Medium (CeMM)-cultured worms in microgravity; six were upregulated, while two were downregulated. Five of the upregulated genes (C07E3.15, C34H3.21, C32D5.16, F35H8.9 and C34F11.17) encode non-coding RNAs. In terms of biological phenotype, we observed that microgravity-simulated worms experienced minimal changes in terms of lifespan, locomotion and reproductive capabilities in comparison with the ground controls. Taking it all together, simulated microgravity on a single generation of C. elegans did not confer major changes to their gene expression and biological phenotype. Nevertheless, exposure of the worms to microgravity lead to higher expression of non-coding RNA genes, which may play an epigenetic role in the worms during longer terms of microgravity exposure.

DNA-Sequence Based Typing of the Cronobacter Genus Using MLST, CRISPR-cas Array and Capsular Profiling

PubMed Central

Ogrodzki, Pauline; Forsythe, Stephen J.

2017-01-01

The Cronobacter genus is composed of seven species, within which a number of pathovars have been described. The most notable infections by Cronobacter spp. are of infants through the consumption of contaminated infant formula. The description of the genus has greatly improved in recent years through DNA sequencing techniques, and this has led to a robust means of identification. However some species are highly clonal and this limits the ability to discriminate between unrelated strains by some methods of genotyping. This article updates the application of three genotyping methods across the Cronobacter genus. The three genotyping methods were multilocus sequence typing (MLST), capsular profiling of the K-antigen and colanic acid (CA) biosynthesis regions, and CRISPR-cas array profiling. A total of 1654 MLST profiled and 286 whole genome sequenced strains, available by open access at the PubMLST Cronobacter database, were used this analysis. The predominance of C. sakazakii and C. malonaticus in clinical infections was confirmed. The majority of clinical strains being in the C. sakazakii clonal complexes (CC) 1 and 4, sequence types (ST) 8 and 12 and C. malonaticus ST7. The capsular profile K2:CA2, previously proposed as being strongly associated with C. sakazakii and C. malonaticus isolates from severe neonatal infections, was also found in C. turicensis, C. dublinensis and C. universalis. The majority of CRISPR-cas types across the genus was the I-E (Ecoli) type. Some strains of C. dublinensis and C. muytjensii encoded the I-F (Ypseudo) type, and others lacked the cas gene loci. The significance of the expanding profiling will be of benefit to researchers as well as governmental and industrial risk assessors. PMID:29033918

Whole Genome Expression in Peripheral-Blood Samples of Workers Professionally Exposed to Polycyclic Aromatic Hydrocarbons

PubMed Central

Wu, Ming-Tsang; Lee, Tzu-Chi; Su, Hung-Ju; Huang, Jie-Len; Peng, Chiung-Yu; Wang, Weihsin; Chou, Ting-Yu; Lin, Ming-Yen; Lin, Wen-Yi; Huang, Chia-Tsuan; Pan, Chih-Hong; Ho, Chi-Kung

2011-01-01

This study aims to examine global gene expression profiles before and after the work-shift among coke-oven workers (COW). COW work six consecutive days and then take two days off. Two blood and urine samples in each worker were collected before starting to work after two-days off and end-of-shift in the sixth-day work in 2009. Altered gene expressions (ratio of gene expression levels between end-of-shift and pre-shift work) were performed by Human OneArray expression system which probes ∼30,000-transcription expression profiling of human genes. Sixteen workers, all men, were enrolled in this study. Median urinary 1-hydroxypyrene (1OHP) levels (μmole/mole creatinine) in end-of-shift work were significantly higher than those in pre-shift work (2.58 vs. 0.29, p = 0.0002). Among the 20,341 genes which passed experimental quality control, 26 gene expression changes, 7 positive and 19 negative, were highly correlated with across-the-shift urinary 1OHP levels (end-of-shift – pre-shift 1OHP) (p-value < 0.001). The high and low exposure groups of across-the-shift urinary 1OHP levels dichotomized in ∼2.00 μmole/mole creatinine were able to be distinguished by these 26 genes. Some of them are known to be involved in apoptosis, chromosome stability/DNA repair, cell cycle control/tumor suppressor, cell adhesion, development/spermatogenesis, immune function, and neuronal cell function. These findings in COW will be an ideal model to study the relationship of PAHs exposure with acute changes of gene expressions. PMID:21854004

Biasogram: Visualization of Confounding Technical Bias in Gene Expression Data

PubMed Central

Krzystanek, Marcin; Szallasi, Zoltan; Eklund, Aron C.

2013-01-01

Gene expression profiles of clinical cohorts can be used to identify genes that are correlated with a clinical variable of interest such as patient outcome or response to a particular drug. However, expression measurements are susceptible to technical bias caused by variation in extraneous factors such as RNA quality and array hybridization conditions. If such technical bias is correlated with the clinical variable of interest, the likelihood of identifying false positive genes is increased. Here we describe a method to visualize an expression matrix as a projection of all genes onto a plane defined by a clinical variable and a technical nuisance variable. The resulting plot indicates the extent to which each gene is correlated with the clinical variable or the technical variable. We demonstrate this method by applying it to three clinical trial microarray data sets, one of which identified genes that may have been driven by a confounding technical variable. This approach can be used as a quality control step to identify data sets that are likely to yield false positive results. PMID:23613961

Toll like receptors gene expression of human keratinocytes cultured of severe burn injury.

PubMed

Cornick, Sarita Mac; Noronha, Silvana Aparecida Alves Corrêa de; Noronha, Samuel Marcos Ribeiro de; Cezillo, Marcus V B; Ferreira, Lydia Masako; Gragnani, Alfredo

2014-01-01

To evaluate the expression profile of genes related to Toll Like Receptors (TLR) pathways of human Primary Epidermal keratinocytes of patients with severe burns. After obtaining viable fragments of skin with and without burning, culture hKEP was initiated by the enzymatic method using Dispase (Sigma-Aldrich). These cells were treated with Trizol(r) (Life Technologies) for extraction of total RNA. This was quantified and analyzed for purity for obtaining cDNA for the analysis of gene expression using specific TLR pathways PCR Arrays plates (SA Biosciences). After the analysis of gene expression we found that 21% of these genes were differentially expressed, of which 100% were repressed or hyporegulated. Among these, the following genes (fold decrease): HSPA1A (-58), HRAS (-36), MAP2K3 (-23), TOLLIP (-23), RELA (-18), FOS (-16), and TLR1 (-6.0). This study contributes to the understanding of the molecular mechanisms related to TLR pathways and underlying wound infection caused by the burn. Furthermore, it may provide new strategies to restore normal expression of these genes and thereby change the healing process and improve clinical outcome.

Three-Dimensional mRNA Measurements Reveal Minimal Regional Heterogeneity in Esophageal Squamous Cell Carcinoma

PubMed Central

Yan, Wusheng; Shih, Joanna; Rodriguez-Canales, Jaime; Tangrea, Michael A.; Player, Audrey; Diao, Lixia; Hu, Nan; Goldstein, Alisa M.; Wang, Jing; Taylor, Philip R.; Lippman, Scott M.; Wistuba, Ignacio I.; Emmert-Buck, Michael R.; Erickson, Heidi S.

2014-01-01

The classic tumor clonal evolution theory postulates that cancers change over time to produce unique molecular subclones within a parent neoplasm, presumably including regional differences in gene expression. More recently, however, this notion has been challenged by studies showing that tumors maintain a relatively stable transcript profile. To examine these competing hypotheses, we microdissected discrete subregions containing approximately 3000 to 8000 cells (500 to 1500 μm in diameter) from ex vivo esophageal squamous cell carcinoma (ESCC) specimens and analyzed transcriptomes throughout three-dimensional tumor space. Overall mRNA profiles were highly similar in all 59 intratumor comparisons, in distinct contrast to the markedly different global expression patterns observed in other dissected cell populations. For example, normal esophageal basal cells contained 1918 and 624 differentially expressed genes at a greater than twofold level (95% confidence level of <5% false positives), compared with normal differentiated esophageal cells and ESCC, respectively. In contrast, intratumor regions had only zero to four gene changes at a greater than twofold level, with most tumor comparisons showing none. The present data indicate that, when analyzed using a standard array-based method at this level of histological resolution, ESCC contains little regional mRNA heterogeneity. PMID:23219752

Compression dynamics of quasi-spherical wire arrays with different linear mass profiles

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mitrofanov, K. N., E-mail: mitrofan@triniti.ru; Aleksandrov, V. V.; Gritsuk, A. N.

Results of experimental studies of the implosion of quasi-spherical wire (or metalized fiber) arrays are presented. The goal of the experiments was to achieve synchronous three-dimensional compression of the plasma produced in different regions of a quasi-spherical array into its geometrical center. To search for optimal synchronization conditions, quasi-spherical arrays with different initial profiles of the linear mass were used. The following dependences of the linear mass on the poloidal angle were used: m{sub l}(θ) ∝ sin{sup –1}θ and m{sub l}(θ) ∝ sin{sup –2}θ. The compression dynamics of such arrays was compared with that of quasi-spherical arrays without linear massmore » profiling, m{sub l}(θ) = const. To verify the experimental data, the spatiotemporal dynamics of plasma compression in quasi-spherical arrays was studied using various diagnostics. The experiments on three-dimensional implosion of quasi-spherical arrays made it possible to study how the frozen-in magnetic field of the discharge current penetrates into the array. By measuring the magnetic field in the plasma of a quasi-spherical array, information is obtained on the processes of plasma production and formation of plasma flows from the wire/fiber regions with and without an additionally deposited mass. It is found that penetration of the magnetic flux depends on the initial linear mass profile m{sub l}(θ) of the quasi-spherical array. From space-resolved spectral measurements and frame imaging of plasma X-ray emission, information is obtained on the dimensions and shape of the X-ray source formed during the implosion of a quasi-spherical array. The intensity of this source is estimated and compared with that of the Z-pinch formed during the implosion of a cylindrical array.« less

Unique inflammatory RNA profiles of microglia in Creutzfeldt-Jakob disease

NASA Astrophysics Data System (ADS)

Baker, Christopher A.; Manuelidis, Laura

2003-01-01

Previous studies in Creutzfeldt-Jakob disease (CJD) have shown that myeloid cells in the periphery as well as derivative microglial cells in the brain are infectious. Microglia can show an activated phenotype before prion protein (PrP) pathology is detectable in brain, and isolated infectious microglia contain very little PrP. To find whether a set of inflammatory genes are significantly induced or suppressed with infection, we analyzed RNA from isolated microglia with relevant cDNA arrays, and identified 30 transcripts not previously examined in any transmissible spongiform encephalopathy. This CJD expression profile contrasted with that of uninfected microglia exposed to prototypic inflammatory stimuli such as lipopolysaccharide and IFN-, as well as PrP amyloid. These findings underscore inflammatory pathways evoked by the infectious agent in brain. Transcript profiles unique for CJD microglia and other myeloid cells provide opportunities for more sensitive preclinical diagnoses of infectious and noninfectious neurodegenerative diseases.

Molecular classification and molecular forecasting of breast cancer: ready for clinical application?

PubMed

Brenton, James D; Carey, Lisa A; Ahmed, Ahmed Ashour; Caldas, Carlos

2005-10-10

Profiling breast cancer with expression arrays has become common, and it has been suggested that the results from early studies will lead to understanding of the molecular differences between clinical cases and allow individualization of care. We critically review two main applications of expression profiling; studies unraveling novel breast cancer classifications and those that aim to identify novel markers for prediction of clinical outcome. Breast cancer may now be subclassified into luminal, basal, and HER2 subtypes with distinct differences in prognosis and response to therapy. However, profiling studies to identify predictive markers have suffered from methodologic problems that prevent general application of their results. Future work will need to reanalyze existing microarray data sets to identify more representative sets of candidate genes for use as prognostic signatures and will need to take into account the new knowledge of molecular subtypes of breast cancer when assessing predictive effects.

Transcript and metabolite analysis in Trincadeira cultivar reveals novel information regarding the dynamics of grape ripening

PubMed Central

2011-01-01

Background Grapes (Vitis vinifera L.) are economically the most important fruit crop worldwide. However, the complexity of molecular and biochemical events that lead to the onset of ripening of nonclimacteric fruits is not fully understood which is further complicated in grapes due to seasonal and cultivar specific variation. The Portuguese wine variety Trincadeira gives rise to high quality wines but presents extremely irregular berry ripening among seasons probably due to high susceptibility to abiotic and biotic stresses. Results Ripening of Trincadeira grapes was studied taking into account the transcriptional and metabolic profilings complemented with biochemical data. The mRNA expression profiles of four time points spanning developmental stages from pea size green berries, through véraison and mature berries (EL 32, EL 34, EL 35 and EL 36) and in two seasons (2007 and 2008) were compared using the Affymetrix GrapeGen® genome array containing 23096 probesets corresponding to 18726 unique sequences. Over 50% of these probesets were significantly differentially expressed (1.5 fold) between at least two developmental stages. A common set of modulated transcripts corresponding to 5877 unigenes indicates the activation of common pathways between years despite the irregular development of Trincadeira grapes. These unigenes were assigned to the functional categories of "metabolism", "development", "cellular process", "diverse/miscellanenous functions", "regulation overview", "response to stimulus, stress", "signaling", "transport overview", "xenoprotein, transposable element" and "unknown". Quantitative RT-PCR validated microarrays results being carried out for eight selected genes and five developmental stages (EL 32, EL 34, EL 35, EL 36 and EL 38). Metabolic profiling using 1H NMR spectroscopy associated to two-dimensional techniques showed the importance of metabolites related to oxidative stress response, amino acid and sugar metabolism as well as secondary metabolism. These results were integrated with transcriptional profiling obtained using genome array to provide new information regarding the network of events leading to grape ripening. Conclusions Altogether the data obtained provides the most extensive survey obtained so far for gene expression and metabolites accumulated during grape ripening. Moreover, it highlighted information obtained in a poorly known variety exhibiting particular characteristics that may be cultivar specific or dependent upon climatic conditions. Several genes were identified that had not been previously reported in the context of grape ripening namely genes involved in carbohydrate and amino acid metabolisms as well as in growth regulators; metabolism, epigenetic factors and signaling pathways. Some of these genes were annotated as receptors, transcription factors, and kinases and constitute good candidates for functional analysis in order to establish a model for ripening control of a non-climacteric fruit. PMID:22047180

Parallel mRNA, proteomics and miRNA expression analysis in cell line models of the intestine.

PubMed

O'Sullivan, Finbarr; Keenan, Joanne; Aherne, Sinead; O'Neill, Fiona; Clarke, Colin; Henry, Michael; Meleady, Paula; Breen, Laura; Barron, Niall; Clynes, Martin; Horgan, Karina; Doolan, Padraig; Murphy, Richard

2017-11-07

To identify miRNA-regulated proteins differentially expressed between Caco2 and HT-29: two principal cell line models of the intestine. Exponentially growing Caco-2 and HT-29 cells were harvested and prepared for mRNA, miRNA and proteomic profiling. mRNA microarray profiling analysis was carried out using the Affymetrix GeneChip Human Gene 1.0 ST array. miRNA microarray profiling analysis was carried out using the Affymetrix Genechip miRNA 3.0 array. Quantitative Label-free LC-MS/MS proteomic analysis was performed using a Dionex Ultimate 3000 RSLCnano system coupled to a hybrid linear ion trap/Orbitrap mass spectrometer. Peptide identities were validated in Proteome Discoverer 2.1 and were subsequently imported into Progenesis QI software for further analysis. Hierarchical cluster analysis for all three parallel datasets (miRNA, proteomics, mRNA) was conducted in the R software environment using the Euclidean distance measure and Ward's clustering algorithm. The prediction of miRNA and oppositely correlated protein/mRNA interactions was performed using TargetScan 6.1. GO biological process, molecular function and cellular component enrichment analysis was carried out for the DE miRNA, protein and mRNA lists via the Pathway Studio 11.3 Web interface using their Mammalian database. Differential expression (DE) profiling comparing the intestinal cell lines HT-29 and Caco-2 identified 1795 Genes, 168 Proteins and 160 miRNAs as DE between the two cell lines. At the gene level, 1084 genes were upregulated and 711 were downregulated in the Caco-2 cell line relative to the HT-29 cell line. At the protein level, 57 proteins were found to be upregulated and 111 downregulated in the Caco-2 cell line relative to the HT-29 cell line. Finally, at the miRNAs level, 104 were upregulated and 56 downregulated in the Caco-2 cell line relative to the HT-29 cell line. Gene ontology (GO) analysis of the DE mRNA identified cell adhesion, migration and ECM organization, cellular lipid and cholesterol metabolic processes, small molecule transport and a range of responses to external stimuli, while similar analysis of the DE protein list identified gene expression/transcription, epigenetic mechanisms, DNA replication, differentiation and translation ontology categories. The DE protein and gene lists were found to share 15 biological processes including for example epithelial cell differentiation [ P value ≤ 1.81613E-08 (protein list); P ≤ 0.000434311 (gene list)] and actin filament bundle assembly [ P value ≤ 0.001582797 (protein list); P ≤ 0.002733714 (gene list)]. Analysis was conducted on the three data streams acquired in parallel to identify targets undergoing potential miRNA translational repression identified 34 proteins, whose respective mRNAs were detected but no change in expression was observed. Of these 34 proteins, 27 proteins downregulated in the Caco-2 cell line relative to the HT-29 cell line and predicted to be targeted by 19 unique anti-correlated/upregulated microRNAs and 7 proteins upregulated in the Caco-2 cell line relative to the HT-29 cell line and predicted to be targeted by 15 unique anti-correlated/downregulated microRNAs. This first study providing "tri-omics" analysis of the principal intestinal cell line models Caco-2 and HT-29 has identified 34 proteins potentially undergoing miRNA translational repression.

Parallel mRNA, proteomics and miRNA expression analysis in cell line models of the intestine

PubMed Central

O’Sullivan, Finbarr; Keenan, Joanne; Aherne, Sinead; O’Neill, Fiona; Clarke, Colin; Henry, Michael; Meleady, Paula; Breen, Laura; Barron, Niall; Clynes, Martin; Horgan, Karina; Doolan, Padraig; Murphy, Richard

2017-01-01

AIM To identify miRNA-regulated proteins differentially expressed between Caco2 and HT-29: two principal cell line models of the intestine. METHODS Exponentially growing Caco-2 and HT-29 cells were harvested and prepared for mRNA, miRNA and proteomic profiling. mRNA microarray profiling analysis was carried out using the Affymetrix GeneChip Human Gene 1.0 ST array. miRNA microarray profiling analysis was carried out using the Affymetrix Genechip miRNA 3.0 array. Quantitative Label-free LC-MS/MS proteomic analysis was performed using a Dionex Ultimate 3000 RSLCnano system coupled to a hybrid linear ion trap/Orbitrap mass spectrometer. Peptide identities were validated in Proteome Discoverer 2.1 and were subsequently imported into Progenesis QI software for further analysis. Hierarchical cluster analysis for all three parallel datasets (miRNA, proteomics, mRNA) was conducted in the R software environment using the Euclidean distance measure and Ward’s clustering algorithm. The prediction of miRNA and oppositely correlated protein/mRNA interactions was performed using TargetScan 6.1. GO biological process, molecular function and cellular component enrichment analysis was carried out for the DE miRNA, protein and mRNA lists via the Pathway Studio 11.3 Web interface using their Mammalian database. RESULTS Differential expression (DE) profiling comparing the intestinal cell lines HT-29 and Caco-2 identified 1795 Genes, 168 Proteins and 160 miRNAs as DE between the two cell lines. At the gene level, 1084 genes were upregulated and 711 were downregulated in the Caco-2 cell line relative to the HT-29 cell line. At the protein level, 57 proteins were found to be upregulated and 111 downregulated in the Caco-2 cell line relative to the HT-29 cell line. Finally, at the miRNAs level, 104 were upregulated and 56 downregulated in the Caco-2 cell line relative to the HT-29 cell line. Gene ontology (GO) analysis of the DE mRNA identified cell adhesion, migration and ECM organization, cellular lipid and cholesterol metabolic processes, small molecule transport and a range of responses to external stimuli, while similar analysis of the DE protein list identified gene expression/transcription, epigenetic mechanisms, DNA replication, differentiation and translation ontology categories. The DE protein and gene lists were found to share 15 biological processes including for example epithelial cell differentiation [P value ≤ 1.81613E-08 (protein list); P ≤ 0.000434311 (gene list)] and actin filament bundle assembly [P value ≤ 0.001582797 (protein list); P ≤ 0.002733714 (gene list)]. Analysis was conducted on the three data streams acquired in parallel to identify targets undergoing potential miRNA translational repression identified 34 proteins, whose respective mRNAs were detected but no change in expression was observed. Of these 34 proteins, 27 proteins downregulated in the Caco-2 cell line relative to the HT-29 cell line and predicted to be targeted by 19 unique anti-correlated/upregulated microRNAs and 7 proteins upregulated in the Caco-2 cell line relative to the HT-29 cell line and predicted to be targeted by 15 unique anti-correlated/downregulated microRNAs. CONCLUSION This first study providing “tri-omics” analysis of the principal intestinal cell line models Caco-2 and HT-29 has identified 34 proteins potentially undergoing miRNA translational repression. PMID:29151691

Analysis of genomic aberrations and gene expression profiling identifies novel lesions and pathways in myeloproliferative neoplasms

PubMed Central

Rice, K L; Lin, X; Wolniak, K; Ebert, B L; Berkofsky-Fessler, W; Buzzai, M; Sun, Y; Xi, C; Elkin, P; Levine, R; Golub, T; Gilliland, D G; Crispino, J D; Licht, J D; Zhang, W

2011-01-01

Polycythemia vera (PV), essential thrombocythemia and primary myelofibrosis, are myeloproliferative neoplasms (MPNs) with distinct clinical features and are associated with the JAK2V617F mutation. To identify genomic anomalies involved in the pathogenesis of these disorders, we profiled 87 MPN patients using Affymetrix 250K single-nucleotide polymorphism (SNP) arrays. Aberrations affecting chr9 were the most frequently observed and included 9pLOH (n=16), trisomy 9 (n=6) and amplifications of 9p13.3–23.3 (n=1), 9q33.1–34.13 (n=1) and 9q34.13 (n=6). Patients with trisomy 9 were associated with elevated JAK2V617F mutant allele burden, suggesting that gain of chr9 represents an alternative mechanism for increasing JAK2V617F dosage. Gene expression profiling of patients with and without chr9 abnormalities (+9, 9pLOH), identified genes potentially involved in disease pathogenesis including JAK2, STAT5B and MAPK14. We also observed recurrent gains of 1p36.31–36.33 (n=6), 17q21.2–q21.31 (n=5) and 17q25.1–25.3 (n=5) and deletions affecting 18p11.31–11.32 (n=8). Combined SNP and gene expression analysis identified aberrations affecting components of a non-canonical PRC2 complex (EZH1, SUZ12 and JARID2) and genes comprising a ‘HSC signature' (MLLT3, SMARCA2 and PBX1). We show that NFIB, which is amplified in 7/87 MPN patients and upregulated in PV CD34+ cells, protects cells from apoptosis induced by cytokine withdrawal. PMID:22829077

Genomic profiling of human penile carcinoma predicts worse prognosis and survival.

PubMed

Busso-Lopes, Ariane F; Marchi, Fábio A; Kuasne, Hellen; Scapulatempo-Neto, Cristovam; Trindade-Filho, José Carlos S; de Jesus, Carlos Márcio N; Lopes, Ademar; Guimarães, Gustavo C; Rogatto, Silvia R

2015-02-01

The molecular mechanisms underlying penile carcinoma are still poorly understood, and the detection of genetic markers would be of great benefit for these patients. In this study, we assessed the genomic profile aiming at identifying potential prognostic biomarkers in penile carcinoma. Globally, 46 penile carcinoma samples were considered to evaluate DNA copy-number alterations via array comparative genomic hybridization (aCGH) combined with human papillomavirus (HPV) genotyping. Specific genes were investigated by using qPCR, FISH, and RT-qPCR. Genomic alterations mapped at 3p and 8p were related to worse prognostic features, including advanced T and clinical stage, recurrence and death from the disease. Losses of 3p21.1-p14.3 and gains of 3q25.31-q29 were associated with reduced cancer-specific and disease-free survival. Genomic alterations detected for chromosome 3 (LAMP3, PPARG, TNFSF10 genes) and 8 (DLC1) were evaluated by qPCR. DLC1 and PPARG losses were associated with poor prognosis characteristics. Losses of DLC1 were an independent risk factor for recurrence on multivariate analysis. The gene-expression analysis showed downexpression of DLC1 and PPARG and overexpression of LAMP3 and TNFSF10 genes. Chromosome Y losses and MYC gene (8q24) gains were confirmed by FISH. HPV infection was detected in 34.8% of the samples, and 19 differential genomic regions were obtained related to viral status. At first time, we described recurrent copy-number alterations and its potential prognostic value in penile carcinomas. We also showed a specific genomic profile according to HPV infection, supporting the hypothesis that penile tumors present distinct etiologies according to virus status. ©2014 American Association for Cancer Research.

Gene expression signatures differentiate ovarian/peritoneal serous carcinoma from breast carcinoma in effusions

PubMed Central

Davidson, Ben; Stavnes, Helene Tuft; Holth, Arild; Chen, Xu; Yang, Yanqin; Shih, Ie-Ming; Wang, Tian-Li

2011-01-01

Abstract Ovarian/primary peritoneal carcinoma and breast carcinoma are the gynaecological cancers that most frequently involve the serosal cavities. With the objective of improving on the limited diagnostic panel currently available for the differential diagnosis of these two malignancies, as well as to define tumour-specific biological targets, we compared their global gene expression patterns. Gene expression profiles of 10 serous ovarian/peritoneal and eight ductal breast carcinoma effusions were analysed using the HumanRef-8 BeadChip from Illumina. Differentially expressed candidate genes were validated using quantitative real-time PCR and immunohistochemistry. Unsupervised hierarchical clustering using all 54,675 genes in the array separated ovarian from breast carcinoma samples. We identified 288 unique probes that were significantly differentially expressed in the two cancers by greater than 3.5-fold, of which 81 and 207 were overexpressed in breast and ovarian/peritoneal carcinoma, respectively. SAM analysis identified 1078 differentially expressed probes with false discovery rate less than 0.05. Genes overexpressed in breast carcinoma included TFF1, TFF3, FOXA1, CA12, GATA3, SDC1, PITX1, TH, EHFD1, EFEMP1, TOB1 and KLF2. Genes overexpressed in ovarian/peritoneal carcinoma included SPON1, RBP1, MFGE8, TM4SF12, MMP7, KLK5/6/7, FOLR1/3, PAX8, APOL2 and NRCAM. The differential expression of 14 genes was validated by quantitative real-time PCR, and differences in 5 gene products were confirmed by immunohistochemistry. Expression profiling distinguishes ovarian/peritoneal carcinoma from breast carcinoma and identifies genes that are differentially expressed in these two tumour types. The molecular signatures unique to these cancers may facilitate their differential diagnosis and may provide a molecular basis for therapeutic target discovery. PMID:20132413

Within and between Whorls: Comparative Transcriptional Profiling of Aquilegia and Arabidopsis

PubMed Central

Voelckel, Claudia; Borevitz, Justin O.; Kramer, Elena M.; Hodges, Scott A.

2010-01-01

Background The genus Aquilegia is an emerging model system in plant evolutionary biology predominantly because of its wide variation in floral traits and associated floral ecology. The anatomy of the Aquilegia flower is also very distinct. There are two whorls of petaloid organs, the outer whorl of sepals and the second whorl of petals that form nectar spurs, as well as a recently evolved fifth whorl of staminodia inserted between stamens and carpels. Methodology/Principal Findings We designed an oligonucleotide microarray based on EST sequences from a mixed tissue, normalized cDNA library of an A. formosa x A. pubescens F2 population representing 17,246 unigenes. We then used this array to analyze floral gene expression in late pre-anthesis stage floral organs from a natural A. formosa population. In particular, we tested for gene expression patterns specific to each floral whorl and to combinations of whorls that correspond to traditional and modified ABC model groupings. Similar analyses were performed on gene expression data of Arabidopsis thaliana whorls previously obtained using the Ath1 gene chips (data available through The Arabidopsis Information Resource). Conclusions/Significance Our comparative gene expression analyses suggest that 1) petaloid sepals and petals of A. formosa share gene expression patterns more than either have organ-specific patterns, 2) petals of A. formosa and A. thaliana may be independently derived, 3) staminodia express B and C genes similar to stamens but the staminodium genetic program has also converged on aspects of the carpel program and 4) staminodia have unique up-regulation of regulatory genes and genes that have been implicated with defense against microbial infection and herbivory. Our study also highlights the value of comparative gene expression profiling and the Aquilegia microarray in particular for the study of floral evolution and ecology. PMID:20352114

A global analysis of protein expression profiles in Sinorhizobium meliloti: discovery of new genes for nodule occupancy and stress adaptation.

PubMed

Djordjevic, Michael A; Chen, Han Cai; Natera, Siria; Van Noorden, Giel; Menzel, Christian; Taylor, Scott; Renard, Clotilde; Geiger, Otto; Weiller, Georg F

2003-06-01

A proteomic examination of Sinorhizobium meliloti strain 1021 was undertaken using a combination of 2-D gel electrophoresis, peptide mass fingerprinting, and bioinformatics. Our goal was to identify (i) putative symbiosis- or nutrient-stress-specific proteins, (ii) the biochemical pathways active under different conditions, (iii) potential new genes, and (iv) the extent of posttranslational modifications of S. meliloti proteins. In total, we identified the protein products of 810 genes (13.1% of the genome's coding capacity). The 810 genes generated 1,180 gene products, with chromosomal genes accounting for 78% of the gene products identified (18.8% of the chromosome's coding capacity). The activity of 53 metabolic pathways was inferred from bioinformatic analysis of proteins with assigned Enzyme Commission numbers. Of the remaining proteins that did not encode enzymes, ABC-type transporters composed 12.7% and regulatory proteins 3.4% of the total. Proteins with up to seven transmembrane domains were identified in membrane preparations. A total of 27 putative nodule-specific proteins and 35 nutrient-stress-specific proteins were identified and used as a basis to define genes and describe processes occurring in S. meliloti cells in nodules and under stress. Several nodule proteins from the plant host were present in the nodule bacteria preparations. We also identified seven potentially novel proteins not predicted from the DNA sequence. Post-translational modifications such as N-terminal processing could be inferred from the data. The posttranslational addition of UMP to the key regulator of nitrogen metabolism, PII, was demonstrated. This work demonstrates the utility of combining mass spectrometry with protein arraying or separation techniques to identify candidate genes involved in important biological processes and niche occupations that may be intransigent to other methods of gene expression profiling.

Potential roles of DNA methylation in the initiation and establishment of replicative senescence revealed by array-based methylome and transcriptome analyses

PubMed Central

Sakaki, Mizuho; Ebihara, Yukiko; Okamura, Kohji; Nakabayashi, Kazuhiko; Igarashi, Arisa; Matsumoto, Kenji; Hata, Kenichiro; Kobayashi, Yoshiro

2017-01-01

Cellular senescence is classified into two groups: replicative and premature senescence. Gene expression and epigenetic changes are reported to differ between these two groups and cell types. Normal human diploid fibroblast TIG-3 cells have often been used in cellular senescence research; however, their epigenetic profiles are still not fully understood. To elucidate how cellular senescence is epigenetically regulated in TIG-3 cells, we analyzed the gene expression and DNA methylation profiles of three types of senescent cells, namely, replicatively senescent, ras-induced senescent (RIS), and non-permissive temperature-induced senescent SVts8 cells, using gene expression and DNA methylation microarrays. The expression of genes involved in the cell cycle and immune response was commonly either down- or up-regulated in the three types of senescent cells, respectively. The altered DNA methylation patterns were observed in replicatively senescent cells, but not in prematurely senescent cells. Interestingly, hypomethylated CpG sites detected on non-CpG island regions (“open sea”) were enriched in immune response-related genes that had non-CpG island promoters. The integrated analysis of gene expression and methylation in replicatively senescent cells demonstrated that differentially expressed 867 genes, including cell cycle- and immune response-related genes, were associated with DNA methylation changes in CpG sites close to the transcription start sites (TSSs). Furthermore, several miRNAs regulated in part through DNA methylation were found to affect the expression of their targeted genes. Taken together, these results indicate that the epigenetic changes of DNA methylation regulate the expression of a certain portion of genes and partly contribute to the introduction and establishment of replicative senescence. PMID:28158250

Transcriptional expression of type-I interferon response genes and stability of housekeeping genes in the human endometrium and endometriosis.

PubMed

Vestergaard, Anna L; Knudsen, Ulla B; Munk, Torben; Rosbach, Hanne; Martensen, Pia M

2011-04-01

Endometriosis is a painful chronic female disease defined by the presence of endometrial tissue implants in ectopic (Ec) locations. The pathogenesis is much debated, and type-I interferons (IFNs) could be involved. The expression of genes of the type-I IFN response were profiled by a specific PCR array of RNA obtained from Ec and eutopic (Eu) endometrium collected from nine endometriosis patients and nine healthy control women. Transcriptional expression levels of selected IFN-regulated and housekeeping genes (HKGs) were investigated by real-time quantitative reverse transcriptase PCR (qRT-PCR). Stably expressed HKGs for valid normalization of transcriptional studies of endometrium and endometriosis have not yet been published. Here, seven HKGs were evaluated for stability using the GeNorm and NormFinder software. A normalization factor based on HMBS, TBP and YWHAZ expression was suitable for normalization of qRT-PCR studies of Eu versus Ec endometrium. In the endometrial cell lines HEC1A, HEC1B, Ishikawa and RL95-2, HMBS and HPRT1 were the most stably expressed. The IFN-specific PCR array indicated significantly different expression of the genes BST2, COL16A1, HOXB2 and ISG20 between the endometrial tissue types. However, by correctly normalized qRT-PCR, levels of BST2, COL16A1 and the highly type-I IFN-stimulated genes ISG12A and 6-16 displayed insignificant variations. Conversely, HOXB2 and ISG20 transcriptions were significantly reduced in endometriosis lesions compared with endometrium from endometriosis patients and healthy controls. In conclusion, appropriate HKGs for normalization of qRT-PCR studies of endometrium and endometriosis have been identified here. Abolished expression of ISG20 and HOX genes could be important in endometriosis.

Genome-Wide High-Resolution aCGH Analysis of Gestational Choriocarcinomas

PubMed Central

Poaty, Henriette; Coullin, Philippe; Peko, Jean Félix; Dessen, Philippe; Diatta, Ange Lucien; Valent, Alexander; Leguern, Eric; Prévot, Sophie; Gombé-Mbalawa, Charles; Candelier, Jean-Jacques; Picard, Jean-Yves; Bernheim, Alain

2012-01-01

Eleven samples of DNA from choriocarcinomas were studied by high resolution CGH-array 244 K. They were studied after histopathological confirmation of the diagnosis, of the androgenic etiology and after a microsatellite marker analysis confirming the absence of contamination of tumor DNA from maternal DNA. Three cell lines, BeWo, JAR, JEG were also studied by this high resolution pangenomic technique. According to aCGH analysis, the de novo choriocarcinomas exhibited simple chromosomal rearrangements or normal profiles. The cell lines showed various and complex chromosomal aberrations. 23 Minimal Critical Regions were defined that allowed us to list the genes that were potentially implicated. Among them, unusually high numbers of microRNA clusters and imprinted genes were observed. PMID:22253721

Biological and metabolic response in STS-135 space-flown mouse skin.

PubMed

Mao, X W; Pecaut, M J; Stodieck, L S; Ferguson, V L; Bateman, T A; Bouxsein, M L; Gridley, D S

2014-08-01

There is evidence that space flight condition-induced biological damage is associated with increased oxidative stress and extracellular matrix (ECM) remodeling. To explore possible mechanisms, changes in gene expression profiles implicated in oxidative stress and in ECM remodeling in mouse skin were examined after space flight. The metabolic effects of space flight in skin tissues were also characterized. Space Shuttle Atlantis (STS-135) was launched at the Kennedy Space Center on a 13-day mission. Female C57BL/6 mice were flown in the STS-135 using animal enclosure modules (AEMs). Within 3-5 h after landing, the mice were euthanized and skin samples were harvested for gene array analysis and metabolic biochemical assays. Many genes responsible for regulating production and metabolism of reactive oxygen species (ROS) were significantly (p < 0.05) altered in the flight group, with fold changes >1.5 compared to AEM control. For ECM profile, several genes encoding matrix and metalloproteinases involved in ECM remodeling were significantly up-/down-regulated following space flight. To characterize the metabolic effects of space flight, global biochemical profiles were evaluated. Of 332 named biochemicals, 19 differed significantly (p < 0.05) between space flight skin samples and AEM ground controls, with 12 up-regulated and 7 down-regulated including altered amino acid, carbohydrate metabolism, cell signaling, and transmethylation pathways. Collectively, the data demonstrated that space flight condition leads to a shift in biological and metabolic homeostasis as the consequence of increased regulation in cellular antioxidants, ROS production, and tissue remodeling. This indicates that astronauts may be at increased risk for pathophysiologic damage or carcinogenesis in cutaneous tissue.

Detection of doublecortin domain-containing 2 (DCDC2), a new candidate tumor suppressor gene of hepatocellular carcinoma, by triple combination array analysis

PubMed Central

2013-01-01

Background To detect genes correlated with hepatocellular carcinoma (HCC), we developed a triple combination array consisting of methylation array, gene expression array and single nucleotide polymorphism (SNP) array analysis. Methods A surgical specimen obtained from a 68-year-old female HCC patient was analyzed by triple combination array, which identified doublecortin domain-containing 2 (DCDC2) as a candidate tumor suppressor gene of HCC. Subsequently, samples from 48 HCC patients were evaluated for their DCDC2 methylation and expression status using methylation specific PCR (MSP) and semi-quantitative reverse transcriptase (RT) PCR, respectively. Then, we investigated the relationship between clinicopathological factors and methylation status of DCDC2. Results DCDC2 was revealed to be hypermethylated (methylation value 0.846, range 0–1.0) in cancer tissue, compared with adjacent normal tissue (0.212) by methylation array in the 68-year-old female patient. Expression array showed decreased expression of DCDC2 in cancerous tissue. SNP array showed that the copy number of chromosome 6p22.1, in which DCDC2 resides, was normal. MSP revealed hypermethylation of the promoter region of DCDC2 in 41 of the tumor samples. DCDC2 expression was significantly decreased in the cases with methylation (P = 0.048). Furthermore, the methylated cases revealed worse prognosis for overall survival than unmethylated cases (P = 0.048). Conclusions The present study indicates that triple combination array is an effective method to detect novel genes related to HCC. We propose that DCDC2 is a tumor suppressor gene of HCC. PMID:24034596

Genomic profiling of plasma cell disorders in a clinical setting: integration of microarray and FISH, after CD138 selection of bone marrow

PubMed Central

Berry, Nadine Kaye; Bain, Nicole L; Enjeti, Anoop K; Rowlings, Philip

2014-01-01

Aim To evaluate the role of whole genome comparative genomic hybridisation microarray (array-CGH) in detecting genomic imbalances as compared to conventional karyotype (GTG-analysis) or myeloma specific fluorescence in situ hybridisation (FISH) panel in a diagnostic setting for plasma cell dyscrasia (PCD). Methods A myeloma-specific interphase FISH (i-FISH) panel was carried out on CD138 PC-enriched bone marrow (BM) from 20 patients having BM biopsies for evaluation of PCD. Whole genome array-CGH was performed on reference (control) and neoplastic (test patient) genomic DNA extracted from CD138 PC-enriched BM and analysed. Results Comparison of techniques demonstrated a much higher detection rate of genomic imbalances using array-CGH. Genomic imbalances were detected in 1, 19 and 20 patients using GTG-analysis, i-FISH and array-CGH, respectively. Genomic rearrangements were detected in one patient using GTG-analysis and seven patients using i-FISH, while none were detected using array-CGH. I-FISH was the most sensitive method for detecting gene rearrangements and GTG-analysis was the least sensitive method overall. All copy number aberrations observed in GTG-analysis were detected using array-CGH and i-FISH. Conclusions We show that array-CGH performed on CD138-enriched PCs significantly improves the detection of clinically relevant and possibly novel genomic abnormalities in PCD, and thus could be considered as a standard diagnostic technique in combination with IGH rearrangement i-FISH. PMID:23969274

Genomic profiling of plasma cell disorders in a clinical setting: integration of microarray and FISH, after CD138 selection of bone marrow.

PubMed

Berry, Nadine Kaye; Bain, Nicole L; Enjeti, Anoop K; Rowlings, Philip

2014-01-01

To evaluate the role of whole genome comparative genomic hybridisation microarray (array-CGH) in detecting genomic imbalances as compared to conventional karyotype (GTG-analysis) or myeloma specific fluorescence in situ hybridisation (FISH) panel in a diagnostic setting for plasma cell dyscrasia (PCD). A myeloma-specific interphase FISH (i-FISH) panel was carried out on CD138 PC-enriched bone marrow (BM) from 20 patients having BM biopsies for evaluation of PCD. Whole genome array-CGH was performed on reference (control) and neoplastic (test patient) genomic DNA extracted from CD138 PC-enriched BM and analysed. Comparison of techniques demonstrated a much higher detection rate of genomic imbalances using array-CGH. Genomic imbalances were detected in 1, 19 and 20 patients using GTG-analysis, i-FISH and array-CGH, respectively. Genomic rearrangements were detected in one patient using GTG-analysis and seven patients using i-FISH, while none were detected using array-CGH. I-FISH was the most sensitive method for detecting gene rearrangements and GTG-analysis was the least sensitive method overall. All copy number aberrations observed in GTG-analysis were detected using array-CGH and i-FISH. We show that array-CGH performed on CD138-enriched PCs significantly improves the detection of clinically relevant and possibly novel genomic abnormalities in PCD, and thus could be considered as a standard diagnostic technique in combination with IGH rearrangement i-FISH.

ChIP-chip.

PubMed

Kim, Tae Hoon; Dekker, Job

2018-05-01

ChIP-chip can be used to analyze protein-DNA interactions in a region-wide and genome-wide manner. DNA microarrays contain PCR products or oligonucleotide probes that are designed to represent genomic sequences. Identification of genomic sites that interact with a specific protein is based on competitive hybridization of the ChIP-enriched DNA and the input DNA to DNA microarrays. The ChIP-chip protocol can be divided into two main sections: Amplification of ChIP DNA and hybridization of ChIP DNA to arrays. A large amount of DNA is required to hybridize to DNA arrays, and hybridization to a set of multiple commercial arrays that represent the entire human genome requires two rounds of PCR amplifications. The relative hybridization intensity of ChIP DNA and that of the input DNA is used to determine whether the probe sequence is a potential site of protein-DNA interaction. Resolution of actual genomic sites bound by the protein is dependent on the size of the chromatin and on the genomic distance between the probes on the array. As with expression profiling using gene chips, ChIP-chip experiments require multiple replicates for reliable statistical measure of protein-DNA interactions. © 2018 Cold Spring Harbor Laboratory Press.

Detection of pathogenic copy number variants in children with idiopathic intellectual disability using 500 K SNP array genomic hybridization

PubMed Central

2009-01-01

Background Array genomic hybridization is being used clinically to detect pathogenic copy number variants in children with intellectual disability and other birth defects. However, there is no agreement regarding the kind of array, the distribution of probes across the genome, or the resolution that is most appropriate for clinical use. Results We performed 500 K Affymetrix GeneChip® array genomic hybridization in 100 idiopathic intellectual disability trios, each comprised of a child with intellectual disability of unknown cause and both unaffected parents. We found pathogenic genomic imbalance in 16 of these 100 individuals with idiopathic intellectual disability. In comparison, we had found pathogenic genomic imbalance in 11 of 100 children with idiopathic intellectual disability in a previous cohort who had been studied by 100 K GeneChip® array genomic hybridization. Among 54 intellectual disability trios selected from the previous cohort who were re-tested with 500 K GeneChip® array genomic hybridization, we identified all 10 previously-detected pathogenic genomic alterations and at least one additional pathogenic copy number variant that had not been detected with 100 K GeneChip® array genomic hybridization. Many benign copy number variants, including one that was de novo, were also detected with 500 K array genomic hybridization, but it was possible to distinguish the benign and pathogenic copy number variants with confidence in all but 3 (1.9%) of the 154 intellectual disability trios studied. Conclusion Affymetrix GeneChip® 500 K array genomic hybridization detected pathogenic genomic imbalance in 10 of 10 patients with idiopathic developmental disability in whom 100 K GeneChip® array genomic hybridization had found genomic imbalance, 1 of 44 patients in whom 100 K GeneChip® array genomic hybridization had found no abnormality, and 16 of 100 patients who had not previously been tested. Effective clinical interpretation of these studies requires considerable skill and experience. PMID:19917086

Therapeutic effects of Euphorbia Pekinensis and Glycyrrhiza glabra on Hepatocellular Carcinoma Ascites Partially Via Regulating the Frk-Arhgdib-Inpp5d-Avpr2-Aqp4 Signal Axis

PubMed Central

Zhang, Yanqiong; Yan, Chen; Li, Yuting; Mao, Xia; Tao, Weiwei; Tang, Yuping; Lin, Ya; Guo, Qiuyan; Duan, Jingao; Lin, Na

2017-01-01

To clarify unknown rationalities of herbaceous compatibility of Euphorbia Pekinensis (DJ) and Glycyrrhiza glabra (GC) acting on hepatocellular carcinoma (HCC) ascites, peritoneum transcriptomics profiling of 15 subjects, including normal control (Con), HCC ascites mouse model (Mod), DJ-alone, DJ/GC-synergy and DJ/GC-antagonism treatment groups were performed on OneArray platform, followed by differentially expressed genes (DEGs) screening. DEGs between Mod and Con groups were considered as HCC ascites-related genes, and those among different drug treatment and Mod groups were identified as DJ/GC-combination-related genes. Then, an interaction network of HCC ascites-related gene-DJ/GC combination-related gene-known therapeutic target gene for ascites was constructed. Based on nodes’ degree, closeness, betweenness and k-coreness, the Frk-Arhgdib-Inpp5d-Avpr2-Aqp4 axis with highly network topological importance was demonstrated to be a candidate target of DJ/GC combination acting on HCC ascites. Importantly, both qPCR and western blot analyses verified this regulatory effects based on HCC ascites mice in vivo and M-1 collecting duct cells in vitro. Collectively, different combination designs of DJ and GC may lead to synergistic or antagonistic effects on HCC ascites partially via regulating the Frk-Arhgdib-Inpp5d-Avpr2-Aqp4 axis, implying that global gene expression profiling combined with network analysis can offer an effective way to understand pharmacological mechanisms of traditional Chinese medicine prescriptions. PMID:28165501

Lysophosphatidic acid induces expression of genes in human oral keratinocytes involved in wound healing.

PubMed

Thorlakson, Hong Huynh; Engen, Stian Andre; Schreurs, Olav; Schenck, Karl; Blix, Inger Johanne Schytte

2017-08-01

Epithelial cells participate in wound healing by covering wounds, but also as important mediators of wound healing processes. Topical application of the phospholipid growth factor lysophosphatidic acid (LPA) accelerates dermal wound healing and we hypothesized that LPA can play a role in human oral wound healing through its effects on human oral keratinocytes (HOK). HOK were isolated from gingival biopsies and exposed to LPA. The LPA receptor profile, signal transduction pathways, gene expression and secretion of selected cytokines were analyzed. HOK expressed the receptors LPA 1 , LPA 5 and LPA 6 and LPA activated the ERK1/2, JNK and p38 intracellular pathways, substantiated by secretion of IL-6 and IL-8. The early (2h) and intermediate (6h) gene expression profiles of HOK after LPA treatment showed a wide array of regulated genes. The majority of the strongest upregulated genes were related to chemotaxis and inflammation, and became downregulated after 6h. At 6h, genes coding for factors involved in extracellular matrix remodeling and re-epithelialization became highly expressed. IL-36γ, not earlier known to be regulated by LPA, was strongly transcribed and translated but not secreted. After stimulation with LPA, HOK responded by regulating factors and genes that are essential in wound healing processes. As LPA is found in saliva and is released by activated cells after wounding, our results indicate that LPA has a favorable physiological role in oral wound healing. This may further point towards a beneficial role for application of LPA on oral surgical or chronic wounds. Copyright © 2017 Elsevier Ltd. All rights reserved.

Aspirin Enhances Osteogenic Potential of Periodontal Ligament Stem Cells (PDLSCs) and Modulates the Expression Profile of Growth Factor-Associated Genes in PDLSCs.

PubMed

Abd Rahman, Fazliny; Mohd Ali, Johari; Abdullah, Mariam; Abu Kasim, Noor Hayaty; Musa, Sabri

2016-07-01

This study investigates the effects of aspirin (ASA) on the proliferative capacity, osteogenic potential, and expression of growth factor-associated genes in periodontal ligament stem cells (PDLSCs). Mesenchymal stem cells (MSCs) from PDL tissue were isolated from human premolars (n = 3). The MSCs' identity was confirmed by immunophenotyping and trilineage differentiation assays. Cell proliferation activity was assessed through 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Polymerase chain reaction array was used to profile the expression of 84 growth factor-associated genes. Pathway analysis was used to identify the biologic functions and canonic pathways activated by ASA treatment. The osteogenic potential was evaluated through mineralization assay. ASA at 1,000 μM enhances osteogenic potential of PDLSCs. Using a fold change (FC) of 2.0 as a threshold value, the gene expression analyses indicated that 19 genes were differentially expressed, which includes 12 upregulated and seven downregulated genes. Fibroblast growth factor 9 (FGF9), vascular endothelial growth factor A (VEGFA), interleukin-2, bone morphogenetic protein-10, VEGFC, and 2 (FGF2) were markedly upregulated (FC range, 6 to 15), whereas pleotropin, FGF5, brain-derived neurotrophic factor, and Dickkopf WNT signaling pathway inhibitor 1 were markedly downregulated (FC 32). Of the 84 growth factor-associated genes screened, 35 showed high cycle threshold values (≥35). ASA modulates the expression of growth factor-associated genes and enhances osteogenic potential in PDLSCs. ASA upregulated the expression of genes that could activate biologic functions and canonic pathways related to cell proliferation, human embryonic stem cell pluripotency, tissue regeneration, and differentiation. These findings suggest that ASA enhances PDLSC function and may be useful in regenerative dentistry applications, particularly in the areas of periodontal health and regeneration.

Gene expression profiling of Japanese psoriatic skin reveals an increased activity in molecular stress and immune response signals.

PubMed

Kulski, Jerzy K; Kenworthy, William; Bellgard, Matthew; Taplin, Ross; Okamoto, Koichi; Oka, Akira; Mabuchi, Tomotaka; Ozawa, Akira; Tamiya, Gen; Inoko, Hidetoshi

2005-12-01

Gene expression profiling was performed on biopsies of affected and unaffected psoriatic skin and normal skin from seven Japanese patients to obtain insights into the pathways that control this disease. HUG95A Affymetrix DNA chips that contained oligonucleotide arrays of approximately 12,000 well-characterized human genes were used in the study. The statistical analysis of the Affymetrix data, based on the ranking of the Student t-test statistic, revealed a complex regulation of molecular stress and immune gene responses. The majority of the 266 induced genes in affected and unaffected psoriatic skin were involved with interferon mediation, immunity, cell adhesion, cytoskeleton restructuring, protein trafficking and degradation, RNA regulation and degradation, signalling transduction, apoptosis and atypical epidermal cellular proliferation and differentiation. The disturbances in the normal protein degradation equilibrium of skin were reflected by the significant increase in the gene expression of various protease inhibitors and proteinases, including the induced components of the ATP/ubiquitin-dependent non-lysosomal proteolytic pathway that is involved with peptide processing and presentation to T cells. Some of the up-regulated genes, such as TGM1, IVL, FABP5, CSTA and SPRR, are well-known psoriatic markers involved in atypical epidermal cellular organization and differentiation. In the comparison between the affected and unaffected psoriatic skin, the transcription factor JUNB was found at the top of the statistical rankings for the up-regulated genes in affected skin, suggesting that it has an important but as yet undefined role in psoriasis. Our gene expression data and analysis suggest that psoriasis is a chronic interferon- and T-cell-mediated immune disease of the skin where the imbalance in epidermal cellular structure, growth and differentiation arises from the molecular antiviral stress signals initiating inappropriate immune responses.

Advances in autism genetics: on the threshold of a new neurobiology

PubMed Central

Abrahams, Brett S.; Geschwind, Daniel H.

2009-01-01

Autism is a heterogeneous syndrome defined by impairments in three core domains: social interaction, language and range of interests. Recent work has led to the identification of several autism susceptibility genes and an increased appreciation of the contribution of de novo and inherited copy number variation. Promising strategies are also being applied to identify common genetic risk variants. Systems biology approaches, including array-based expression profiling, are poised to provide additional insights into this group of disorders, in which heterogeneity, both genetic and phenotypic, is emerging as a dominant theme. PMID:18414403

Molecular Profiles for Lung Cancer Pathogenesis and Detection in US Veterans

DTIC Science & Technology

2011-10-01

raw image files that were subsequently converted to probe set data. Raw expression data was analyzed using the BRB-ArrayTools v .4.1.0 developed by Dr...epithelium of cigarette smokers. Am J Respir Cell Mol Biol 2003;29:331-343 14. Spira A, Beane J, Shah V , et al. Effects of cigarette smoke on the...human airway epithelial cell transcriptome. Proc Natl Acad Sci U S A 2004;101:10143-10148 15. Spira A, Beane JE, Shah V , et al. Airway epithelial gene

Spiral biasing adaptor for use in Si drift detectors and Si drift detector arrays

DOEpatents

Li, Zheng; Chen, Wei

2016-07-05

A drift detector array, preferably a silicon drift detector (SDD) array, that uses a low current biasing adaptor is disclosed. The biasing adaptor is customizable for any desired geometry of the drift detector single cell with minimum drift time of carriers. The biasing adaptor has spiral shaped ion-implants that generate the desired voltage profile. The biasing adaptor can be processed on the same wafer as the drift detector array and only one biasing adaptor chip/side is needed for one drift detector array to generate the voltage profiles on the front side and back side of the detector array.

Transcriptomic assessment of resistance to effects of an aryl hydrocarbon receptor (AHR) agonist in embryos of Atlantic killifish (Fundulus heteroclitus) from a marine Superfund site

PubMed Central

2011-01-01

Background Populations of Atlantic killifish (Fundulus heteroclitus) have evolved resistance to the embryotoxic effects of polychlorinated biphenyls (PCBs) and other halogenated and nonhalogenated aromatic hydrocarbons that act through an aryl hydrocarbon receptor (AHR)-dependent signaling pathway. The resistance is accompanied by reduced sensitivity to induction of cytochrome P450 1A (CYP1A), a widely used biomarker of aromatic hydrocarbon exposure and effect, but whether the reduced sensitivity is specific to CYP1A or reflects a genome-wide reduction in responsiveness to all AHR-mediated changes in gene expression is unknown. We compared gene expression profiles and the response to 3,3',4,4',5-pentachlorobiphenyl (PCB-126) exposure in embryos (5 and 10 dpf) and larvae (15 dpf) from F. heteroclitus populations inhabiting the New Bedford Harbor, Massachusetts (NBH) Superfund site (PCB-resistant) and a reference site, Scorton Creek, Massachusetts (SC; PCB-sensitive). Results Analysis using a 7,000-gene cDNA array revealed striking differences in responsiveness to PCB-126 between the populations; the differences occur at all three stages examined. There was a sizeable set of PCB-responsive genes in the sensitive SC population, a much smaller set of PCB-responsive genes in NBH fish, and few similarities in PCB-responsive genes between the two populations. Most of the array results were confirmed, and additional PCB-regulated genes identified, by RNA-Seq (deep pyrosequencing). Conclusions The results suggest that NBH fish possess a gene regulatory defect that is not specific to one target gene such as CYP1A but rather lies in a regulatory pathway that controls the transcriptional response of multiple genes to PCB exposure. The results are consistent with genome-wide disruption of AHR-dependent signaling in NBH fish. PMID:21609454

Improved intra-array and interarray normalization of peptide microarray phosphorylation for phosphorylome and kinome profiling by rational selection of relevant spots

PubMed Central

Scholma, Jetse; Fuhler, Gwenny M.; Joore, Jos; Hulsman, Marc; Schivo, Stefano; List, Alan F.; Reinders, Marcel J. T.; Peppelenbosch, Maikel P.; Post, Janine N.

2016-01-01

Massive parallel analysis using array technology has become the mainstay for analysis of genomes and transcriptomes. Analogously, the predominance of phosphorylation as a regulator of cellular metabolism has fostered the development of peptide arrays of kinase consensus substrates that allow the charting of cellular phosphorylation events (often called kinome profiling). However, whereas the bioinformatical framework for expression array analysis is well-developed, no advanced analysis tools are yet available for kinome profiling. Especially intra-array and interarray normalization of peptide array phosphorylation remain problematic, due to the absence of “housekeeping” kinases and the obvious fallacy of the assumption that different experimental conditions should exhibit equal amounts of kinase activity. Here we describe the development of analysis tools that reliably quantify phosphorylation of peptide arrays and that allow normalization of the signals obtained. We provide a method for intraslide gradient correction and spot quality control. We describe a novel interarray normalization procedure, named repetitive signal enhancement, RSE, which provides a mathematical approach to limit the false negative results occuring with the use of other normalization procedures. Using in silico and biological experiments we show that employing such protocols yields superior insight into cellular physiology as compared to classical analysis tools for kinome profiling. PMID:27225531

Altered DNA methylation profile in Norwegian patients with Autoimmune Addison's Disease.

PubMed

Bjanesoy, Trine E; Andreassen, Bettina Kulle; Bratland, Eirik; Reiner, Andrew; Islam, Shahinul; Husebye, Eystein S; Bakke, Marit

2014-06-01

Autoimmune Addison's Disease (AAD) is an endocrine and immunological disease of uncertain pathogenesis resulting from the immune system's destruction of the hormone producing cells of the adrenal cortex. The underlying molecular mechanisms are largely unknown, but it is commonly accepted that a combination of genetic susceptibility and environmental impact is critical. In the present study, we identified multiple hypomethylated gene promoter regions in patients with isolated AAD using DNA isolated from CD4+ T cells. The identified differentially methylated regions were distributed evenly across the 10.5-kb-promoter regions covered by the array, and a substantial number localized to promoters of genes involved in immune regulation and autoimmunity. This study reveals a hypomethylated status in CD4+ T cells from AAD patients and indicates differential methylation of promoters of key genes involved in immune responses. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

Characterization of transformation related genes in oral cancer cells.

PubMed

Chang, D D; Park, N H; Denny, C T; Nelson, S F; Pe, M

1998-04-16

A cDNA representational difference analysis (cDNA-RDA) and an arrayed filter technique were used to characterize transformation-related genes in oral cancer. From an initial comparison of normal oral epithelial cells and a human papilloma virus (HPV)-immortalized oral epithelial cell line, we obtained 384 differentially expressed gene fragments and arrayed them on a filter. Two hundred and twelve redundant clones were identified by three rounds of back hybridization. Sequence analysis of the remaining clones revealed 99 unique clones corresponding to 69 genes. The expression of these transformation related gene fragments in three nontumorigenic HPV-immortalized oral epithelial cell lines and three oral cancer cell lines were simultaneously monitored using a cDNA array hybridization. Although there was a considerable cell line-to-cell line variability in the expression of these clones, a reliable prediction of their expression could be made from the cDNA array hybridization. Our study demonstrates the utility of combining cDNA-RDA and arrayed filters in high-throughput gene expression difference analysis. The differentially expressed genes identified in this study should be informative in studying oral epithelial cell carcinogenesis.

TEMPORAL GENE INDUCTION PATTERNS IN SHEEPSHEAD MINNOWS EXPOSED TO 17-ESTRADIOL

EPA Science Inventory

Gene arrays provide a powerful method to examine changes in gene expression in fish due to chemical exposures in the environment. In this study, we expanded an existing gene array for sheepshead minnows (Cyprinodon variegatus) (SHM) and used it to examine temporal changes in gene...

Interference with ethylene perception at receptor level sheds light on auxin and transcriptional circuits associated with the climacteric ripening of apple fruit (Malus x domestica Borkh.).

PubMed

Tadiello, Alice; Longhi, Sara; Moretto, Marco; Ferrarini, Alberto; Tononi, Paola; Farneti, Brian; Busatto, Nicola; Vrhovsek, Urska; Molin, Alessandra Dal; Avanzato, Carla; Biasioli, Franco; Cappellin, Luca; Scholz, Matthias; Velasco, Riccardo; Trainotti, Livio; Delledonne, Massimo; Costa, Fabrizio

2016-12-01

Apple (Malus x domestica Borkh.) is a model species for studying the metabolic changes that occur at the onset of ripening in fruit crops, and the physiological mechanisms that are governed by the hormone ethylene. In this study, to dissect the climacteric interplay in apple, a multidisciplinary approach was employed. To this end, a comprehensive analysis of gene expression together with the investigation of several physiological entities (texture, volatilome and content of polyphenolic compounds) was performed throughout fruit development and ripening. The transcriptomic profiling was conducted with two microarray platforms: a dedicated custom array (iRIPE) and a whole genome array specifically enriched with ripening-related genes for apple (WGAA). The transcriptomic and phenotypic changes following the application of 1-methylcyclopropene (1-MCP), an ethylene inhibitor leading to important modifications in overall fruit physiology, were also highlighted. The integrative comparative network analysis showed both negative and positive correlations between ripening-related transcripts and the accumulation of specific metabolites or texture components. The ripening distortion caused by the inhibition of ethylene perception, in addition to affecting the ethylene pathway, stimulated the de-repression of auxin-related genes, transcription factors and photosynthetic genes. Overall, the comprehensive repertoire of results obtained here advances the elucidation of the multi-layered climacteric mechanism of fruit ripening, thus suggesting a possible transcriptional circuit governed by hormones and transcription factors. © 2016 The Authors The Plant Journal © 2016 John Wiley & Sons Ltd.

Genome-wide analysis of alternative splicing during dendritic cell response to a bacterial challenge.

PubMed

Rodrigues, Raquel; Grosso, Ana Rita; Moita, Luís

2013-01-01

The immune system relies on the plasticity of its components to produce appropriate responses to frequent environmental challenges. Dendritic cells (DCs) are critical initiators of innate immunity and orchestrate the later and more specific adaptive immunity. The generation of diversity in transcriptional programs is central for effective immune responses. Alternative splicing is widely considered a key generator of transcriptional and proteomic complexity, but its role has been rarely addressed systematically in immune cells. Here we used splicing-sensitive arrays to assess genome-wide gene- and exon-level expression profiles in human DCs in response to a bacterial challenge. We find widespread alternative splicing events and splicing factor transcriptional signatures induced by an E. coli challenge to human DCs. Alternative splicing acts in concert with transcriptional modulation, but these two mechanisms of gene regulation affect primarily distinct functional gene groups. Alternative splicing is likely to have an important role in DC immunobiology because it affects genes known to be involved in DC development, endocytosis, antigen presentation and cell cycle arrest.

NCBI GEO: mining millions of expression profiles--database and tools.

PubMed

Barrett, Tanya; Suzek, Tugba O; Troup, Dennis B; Wilhite, Stephen E; Ngau, Wing-Chi; Ledoux, Pierre; Rudnev, Dmitry; Lash, Alex E; Fujibuchi, Wataru; Edgar, Ron

2005-01-01

The Gene Expression Omnibus (GEO) at the National Center for Biotechnology Information (NCBI) is the largest fully public repository for high-throughput molecular abundance data, primarily gene expression data. The database has a flexible and open design that allows the submission, storage and retrieval of many data types. These data include microarray-based experiments measuring the abundance of mRNA, genomic DNA and protein molecules, as well as non-array-based technologies such as serial analysis of gene expression (SAGE) and mass spectrometry proteomic technology. GEO currently holds over 30,000 submissions representing approximately half a billion individual molecular abundance measurements, for over 100 organisms. Here, we describe recent database developments that facilitate effective mining and visualization of these data. Features are provided to examine data from both experiment- and gene-centric perspectives using user-friendly Web-based interfaces accessible to those without computational or microarray-related analytical expertise. The GEO database is publicly accessible through the World Wide Web at http://www.ncbi.nlm.nih.gov/geo.

Gene expression profiling to identify the toxicities and potentially relevant human disease outcomes associated with environmental heavy metal exposure.

PubMed

Korashy, Hesham M; Attafi, Ibraheem M; Famulski, Konrad S; Bakheet, Saleh A; Hafez, Mohammed M; Alsaad, Abdulaziz M S; Al-Ghadeer, Abdul Rahman M

2017-02-01

Heavy metals are the most commonly encountered toxic substances that increase susceptibility to various diseases after prolonged exposure. We have previously shown that healthy volunteers living near a mining area had significant contamination with heavy metals associated with significant changes in the expression of some detoxifying genes, xenobiotic metabolizing enzymes, and DNA repair genes. However, alterations of most of the molecular target genes associated with diseases are still unknown. Thus, the aims of this study were to (a) evaluate the gene expression profile and (b) identify the toxicities and potentially relevant human disease outcomes associated with long-term human exposure to environmental heavy metals in mining area using microarray analysis. For this purpose, 40 healthy male volunteers who were residents of a heavy metal-polluted area (Mahd Al-Dhahab city, Saudi Arabia) and 20 healthy male volunteers who were residents of a non-heavy metal-polluted area were included in the study. Total RNA was isolated from whole blood using PAXgene Blood RNA tubes and then reversed transcribed and hybridized to the gene array using the Affymetrix U219 GeneChip. Microarray analysis showed about 2129 genes were identified and differentially altered, among which a shared set of 425 genes was differentially expressed in the heavy metal-exposed groups. Ingenuity pathway analysis revealed that the most altered gene-regulated diseases in heavy metal-exposed groups included hematological and developmental disorders and mostly renal and urological diseases. Quantitative real-time polymerase chain reaction closely matched the microarray data for some genes tested. Importantly, changes in gene-related diseases were attributed to alterations in the genes encoded for protein synthesis. Renal and urological diseases were the diseases that were most frequently associated with the heavy metal-exposed group. Therefore, there is a need for further studies to validate these genes, which could be used as early biomarkers to prevent renal injury. Copyright © 2016 Elsevier Ltd. All rights reserved.

Global gene expression and systems biology analysis of bovine monocyte-derived macrophages in response to in vitro challenge with Mycobacterium bovis.

PubMed

Magee, David A; Taraktsoglou, Maria; Killick, Kate E; Nalpas, Nicolas C; Browne, John A; Park, Stephen D E; Conlon, Kevin M; Lynn, David J; Hokamp, Karsten; Gordon, Stephen V; Gormley, Eamonn; MacHugh, David E

2012-01-01

Mycobacterium bovis, the causative agent of bovine tuberculosis, is a major cause of mortality in global cattle populations. Macrophages are among the first cell types to encounter M. bovis following exposure and the response elicited by these cells is pivotal in determining the outcome of infection. Here, a functional genomics approach was undertaken to investigate global gene expression profiles in bovine monocyte-derived macrophages (MDM) purified from seven age-matched non-related females, in response to in vitro challenge with M. bovis (multiplicity of infection 2:1). Total cellular RNA was extracted from non-challenged control and M. bovis-challenged MDM for all animals at intervals of 2 hours, 6 hours and 24 hours post-challenge and prepared for global gene expression analysis using the Affymetrix® GeneChip® Bovine Genome Array. Comparison of M. bovis-challenged MDM gene expression profiles with those from the non-challenged MDM controls at each time point identified 3,064 differentially expressed genes 2 hours post-challenge, with 4,451 and 5,267 differentially expressed genes detected at the 6 hour and 24 hour time points, respectively (adjusted P-value threshold ≤ 0.05). Notably, the number of downregulated genes exceeded the number of upregulated genes in the M. bovis-challenged MDM across all time points; however, the fold-change in expression for the upregulated genes was markedly higher than that for the downregulated genes. Systems analysis revealed enrichment for genes involved in: (1) the inflammatory response; (2) cell signalling pathways, including Toll-like receptors and intracellular pathogen recognition receptors; and (3) apoptosis. The increased number of downregulated genes is consistent with previous studies showing that M. bovis infection is associated with the repression of host gene expression. The results also support roles for MyD88-independent signalling and intracellular PRRs in mediating the host response to M. bovis.

Genome-Wide Mapping of Cystitis Due to Streptococcus agalactiae and Escherichia coli in Mice Identifies a Unique Bladder Transcriptome That Signifies Pathogen-Specific Antimicrobial Defense against Urinary Tract Infection

PubMed Central

Tan, Chee K.; Carey, Alison J.; Cui, Xiangqin; Webb, Richard I.; Ipe, Deepak; Crowley, Michael; Cripps, Allan W.; Benjamin, William H.; Ulett, Kimberly B.; Schembri, Mark A.

2012-01-01

The most common causes of urinary tract infections (UTIs) are Gram-negative pathogens such as Escherichia coli; however, Gram-positive organisms, including Streptococcus agalactiae, or group B streptococcus (GBS), also cause UTI. In GBS infection, UTI progresses to cystitis once the bacteria colonize the bladder, but the host responses triggered in the bladder immediately following infection are largely unknown. Here, we used genome-wide expression profiling to map the bladder transcriptome of GBS UTI in mice infected transurethrally with uropathogenic GBS that was cultured from a 35-year-old women with cystitis. RNA from bladders was applied to Affymetrix Gene-1.0ST microarrays; quantitative reverse transcriptase PCR (qRT-PCR) was used to analyze selected gene responses identified in array data sets. A surprisingly small significant-gene list of 172 genes was identified at 24 h; this compared to 2,507 genes identified in a side-by-side comparison with uropathogenic E. coli (UPEC). No genes exhibited significantly altered expression at 2 h in GBS-infected mice according to arrays despite high bladder bacterial loads at this early time point. The absence of a marked early host response to GBS juxtaposed with broad-based bladder responses activated by UPEC at 2 h. Bioinformatics analyses, including integrative system-level network mapping, revealed multiple activated biological pathways in the GBS bladder transcriptome that regulate leukocyte activation, inflammation, apoptosis, and cytokine-chemokine biosynthesis. These findings define a novel, minimalistic type of bladder host response triggered by GBS UTI, which comprises collective antimicrobial pathways that differ dramatically from those activated by UPEC. Overall, this study emphasizes the unique nature of bladder immune activation mechanisms triggered by distinct uropathogens. PMID:22733575

ArrayVigil: a methodology for statistical comparison of gene signatures using segregated-one-tailed (SOT) Wilcoxon's signed-rank test.

PubMed

Khan, Haseeb Ahmad

2005-01-28

Due to versatile diagnostic and prognostic fidelity molecular signatures or fingerprints are anticipated as the most powerful tools for cancer management in the near future. Notwithstanding the experimental advancements in microarray technology, methods for analyzing either whole arrays or gene signatures have not been firmly established. Recently, an algorithm, ArraySolver has been reported by Khan for two-group comparison of microarray gene expression data using two-tailed Wilcoxon signed-rank test. Most of the molecular signatures are composed of two sets of genes (hybrid signatures) wherein up-regulation of one set and down-regulation of the other set collectively define the purpose of a gene signature. Since the direction of a selected gene's expression (positive or negative) with respect to a particular disease condition is known, application of one-tailed statistics could be a more relevant choice. A novel method, ArrayVigil, is described for comparing hybrid signatures using segregated-one-tailed (SOT) Wilcoxon signed-rank test and the results compared with integrated-two-tailed (ITT) procedures (SPSS and ArraySolver). ArrayVigil resulted in lower P values than those obtained from ITT statistics while comparing real data from four signatures.

Differential Gene Expression Profiling of Functionally and Developmentally Distinct Human Prostate Epithelial Populations

PubMed Central

Liu, Haibo; Cadaneanu, Radu M; Lai, Kevin; Zhang, Baohui; Huo, Lihong; An, Dong Sun; Li, Xinmin; Lewis, Michael S; Garraway, Isla P

2015-01-01

BACKGROUND Human fetal prostate buds appear in the 10th gestational week as solid cords, which branch and form lumens in response to androgen 1. Previous in vivo analysis of prostate epithelia isolated from benign prostatectomy specimens indicated that Epcam+CD44−CD49fHi basal cells possess efficient tubule initiation capability relative to other subpopulations 2. Stromal interactions and branching morphogenesis displayed by adult tubule-initiating cells (TIC) are reminiscent of fetal prostate development. In the current study, we evaluated in vivo tubule initiation by human fetal prostate cells and determined expression profiles of fetal and adult epithelial subpopulations in an effort to identify pathways used by TIC. METHODS Immunostaining and FACS analysis based on Epcam, CD44, and CD49f expression demonstrated the majority (99.9%) of fetal prostate epithelial cells (FC) were Epcam+CD44− with variable levels of CD49f expression. Fetal populations isolated via cell sorting were implanted into immunocompromised mice. Total RNA isolation from Epcam+CD44−CD49fHi FC, adult Epcam+CD44−CD49fHi TIC, Epcam+CD44+CD49fHi basal cells (BC), and Epcam+CD44−CD49fLo luminal cells (LC) was performed, followed by microarray analysis of 19 samples using the Affymetrix Gene Chip Human U133 Plus 2.0 Array. Data was analyzed using Partek Genomics Suite Version 6.4. Genes selected showed >2-fold difference in expression and P < 5.00E-2. Results were validated with RT-PCR. RESULTS Grafts retrieved from Epcam+CD44− fetal cell implants displayed tubule formation with differentiation into basal and luminal compartments, while only stromal outgrowths were recovered from Epcam- fetal cell implants. Hierarchical clustering revealed four distinct groups determined by antigenic profile (TIC, BC, LC) and developmental stage (FC). TIC and BC displayed basal gene expression profiles, while LC expressed secretory genes. FC had a unique profile with the most similarities to adult TIC. Functional, network, and canonical pathway identification using Ingenuity Pathway Analysis Version 7.6 compiled genes with the highest differential expression (TIC relative to BC or LC). Many of these genes were found to be significantly associated with prostate tumorigenesis. CONCLUSIONS Our results demonstrate clustering gene expression profiles of FC and adult TIC. Pathways associated with TIC are known to be deregulated in cancer, suggesting a cell-of-origin role for TIC versus re-emergence of pathways common to these cells in tumorigenesis. Prostate 75: 764–776, 2015. © The Authors. The Prostate, published by Wiley Periodicals, Inc. PMID:25663004

Differential gene expression profiling of functionally and developmentally distinct human prostate epithelial populations.

PubMed

Liu, Haibo; Cadaneanu, Radu M; Lai, Kevin; Zhang, Baohui; Huo, Lihong; An, Dong Sun; Li, Xinmin; Lewis, Michael S; Garraway, Isla P

2015-05-01

Human fetal prostate buds appear in the 10th gestational week as solid cords, which branch and form lumens in response to androgen 1. Previous in vivo analysis of prostate epithelia isolated from benign prostatectomy specimens indicated that Epcam⁺ CD44⁻ CD49f(Hi) basal cells possess efficient tubule initiation capability relative to other subpopulations 2. Stromal interactions and branching morphogenesis displayed by adult tubule-initiating cells (TIC) are reminiscent of fetal prostate development. In the current study, we evaluated in vivo tubule initiation by human fetal prostate cells and determined expression profiles of fetal and adult epithelial subpopulations in an effort to identify pathways used by TIC. Immunostaining and FACS analysis based on Epcam, CD44, and CD49f expression demonstrated the majority (99.9%) of fetal prostate epithelial cells (FC) were Epcam⁺ CD44⁻ with variable levels of CD49f expression. Fetal populations isolated via cell sorting were implanted into immunocompromised mice. Total RNA isolation from Epcam⁺ CD44⁻ CD49f(Hi) FC, adult Epcam⁺ CD44⁻ CD49f(Hi) TIC, Epcam⁺ CD44⁺ CD49f(Hi) basal cells (BC), and Epcam⁺ CD44⁻ CD49f(Lo) luminal cells (LC) was performed, followed by microarray analysis of 19 samples using the Affymetrix Gene Chip Human U133 Plus 2.0 Array. Data was analyzed using Partek Genomics Suite Version 6.4. Genes selected showed >2-fold difference in expression and P < 5.00E-2. Results were validated with RT-PCR. Grafts retrieved from Epcam⁺ CD44⁻ fetal cell implants displayed tubule formation with differentiation into basal and luminal compartments, while only stromal outgrowths were recovered from Epcam- fetal cell implants. Hierarchical clustering revealed four distinct groups determined by antigenic profile (TIC, BC, LC) and developmental stage (FC). TIC and BC displayed basal gene expression profiles, while LC expressed secretory genes. FC had a unique profile with the most similarities to adult TIC. Functional, network, and canonical pathway identification using Ingenuity Pathway Analysis Version 7.6 compiled genes with the highest differential expression (TIC relative to BC or LC). Many of these genes were found to be significantly associated with prostate tumorigenesis. Our results demonstrate clustering gene expression profiles of FC and adult TIC. Pathways associated with TIC are known to be deregulated in cancer, suggesting a cell-of-origin role for TIC versus re-emergence of pathways common to these cells in tumorigenesis. © 2015 The Authors. The Prostate, published by Wiley Periodicals, Inc.

Isoflavone supplement composition and equol producer status affect gene expression in adipose tissue: a double-blind, randomized, placebo-controlled crossover trial in postmenopausal women.

PubMed

van der Velpen, Vera; Geelen, Anouk; Hollman, Peter C H; Schouten, Evert G; van 't Veer, Pieter; Afman, Lydia A

2014-11-01

Isoflavone supplements, consumed by women experiencing menopausal symptoms, are suggested to have positive effects on menopause-related adiposity and cardiovascular disease risk profile, but discussions about their safety are still ongoing. The objective was to study the effects of an 8-wk consumption of 2 different isoflavone supplements compared with placebo on whole-genome gene expression in the adipose tissue of postmenopausal women. This double-blind, randomized, placebo-controlled crossover intervention consisted of 2 substudies, one with a low-genistein (LG) supplement (56% daidzein + daidzin, 16% genistein + genistin, and 28% glycitein + glycitin) and the other with a high-genistein (HG) supplement (49% daidzein + daidzin, 41% genistein + genistin, and 10% glycitein + glycitin). Both supplements provided ∼ 100 mg isoflavones/d (aglycone equivalents). After the 8-wk isoflavone and placebo period, whole-genome arrays were performed in subcutaneous adipose tissue of postmenopausal women (n = 26 after LG, n = 31 after HG). Participants were randomized by equol-producing phenotype, and data analysis was performed per substudy for equol producers and nonproducers separately. Gene set enrichment analysis showed downregulation of expression of energy metabolism-related genes after LG supplementation (n = 24) in both equol-producing phenotypes and oppositely regulated expression for equol producers (down) and nonproducers (up) after HG supplementation (n = 31). Expression of inflammation-related genes was upregulated in equol producers but downregulated in nonproducers, independent of supplement type. Only 4.4-7.0% of the genes with significantly changed expression were estrogen responsive. Body weight, adipocyte size, and plasma lipid profile were not affected by isoflavone supplementation. Effects of isoflavones on adipose tissue gene expression were influenced by supplement composition and equol-producing phenotype, whereas estrogen-responsive effects were lacking. LG isoflavone supplementation resulted in a caloric restriction-like gene expression profile for both producer phenotypes and pointed toward a potential beneficial effect, whereas both supplements induced anti-inflammatory gene expression in equol producers. The study was registered at clinicaltrials.gov as NCT01556737. © 2014 American Society for Nutrition.

Identification of suitable genes contributes to lung adenocarcinoma clustering by multiple meta-analysis methods.

PubMed

Yang, Ze-Hui; Zheng, Rui; Gao, Yuan; Zhang, Qiang

2016-09-01

With the widespread application of high-throughput technology, numerous meta-analysis methods have been proposed for differential expression profiling across multiple studies. We identified the suitable differentially expressed (DE) genes that contributed to lung adenocarcinoma (ADC) clustering based on seven popular multiple meta-analysis methods. Seven microarray expression profiles of ADC and normal controls were extracted from the ArrayExpress database. The Bioconductor was used to perform the data preliminary preprocessing. Then, DE genes across multiple studies were identified. Hierarchical clustering was applied to compare the classification performance for microarray data samples. The classification efficiency was compared based on accuracy, sensitivity and specificity. Across seven datasets, 573 ADC cases and 222 normal controls were collected. After filtering out unexpressed and noninformative genes, 3688 genes were remained for further analysis. The classification efficiency analysis showed that DE genes identified by sum of ranks method separated ADC from normal controls with the best accuracy, sensitivity and specificity of 0.953, 0.969 and 0.932, respectively. The gene set with the highest classification accuracy mainly participated in the regulation of response to external stimulus (P = 7.97E-04), cyclic nucleotide-mediated signaling (P = 0.01), regulation of cell morphogenesis (P = 0.01) and regulation of cell proliferation (P = 0.01). Evaluation of DE genes identified by different meta-analysis methods in classification efficiency provided a new perspective to the choice of the suitable method in a given application. Varying meta-analysis methods always present varying abilities, so synthetic consideration should be taken when providing meta-analysis methods for particular research. © 2015 John Wiley & Sons Ltd.

Epigenetic hereditary transcription profiles III, evidence for an epigenetic network resulting in gender, tissue and age-specific variation in overall transcription

PubMed Central

Simons, Johannes WIM

2009-01-01

Background We have previously shown that deviations from the average transcription profile of a group of functionally related genes are not only heritable, but also demonstrate specific patterns associated with age, gender and differentiation, thereby implicating genome-wide nuclear programming as the cause. To determine whether these results could be reproduced, a different micro-array database (obtained from two types of muscle tissue, derived from 81 human donors aged between 16 to 89 years) was studied. Results This new database also revealed the existence of age, gender and tissue-specific features in a small group of functionally related genes. In order to further analyze this phenomenon, a method was developed for quantifying the contribution of different factors to the variability in gene expression, and for generating a database limited to residual values reflecting constitutional differences between individuals. These constitutional differences, presumably epigenetic in origin, contribute to about 50% of the observed residual variance which is connected with a network of interrelated changes in gene expression with some genes displaying a decrease or increase in residual variation with age. Conclusion Epigenetic variation in gene expression without a clear concomitant relation to gene function appears to be a widespread phenomenon. This variation is connected with interactions between genes, is gender and tissue specific and is related to cellular aging. This finding, together with the method developed for analysis, might contribute to the elucidation of the role of nuclear programming in differentiation, aging and carcinogenesis Reviewers This article was reviewed by Thiago M. Venancio (nominated by Aravind Iyer), Hua Li (nominated by Arcady Mushegian) and Arcady Mushegian and J.P.de Magelhaes (nominated by G. Church). PMID:19796384

Gene expression profiling in Ishikawa cells: A fingerprint for estrogen active compounds

DOE Office of Scientific and Technical Information (OSTI.GOV)

Boehme, Kathleen; Simon, Stephanie; Mueller, Stefan O.

2009-04-01

Several anthropogenous and naturally occurring substances, referred to as estrogen active compounds (EACs), are able to interfere with hormone and in particular estrogen receptor signaling. EACs can either cause adverse health effects in humans and wildlife populations or have beneficial effects on estrogen-dependent diseases. The aim of this study was to examine global gene expression profiles in estrogen receptor (ER)-proficient Ishikawa plus and ER-deficient Ishikawa minus endometrial cancer cells treated with selected well-known EACs (Diethylstilbestrol, Genistein, Zearalenone, Resveratrol, Bisphenol A and o,p'-DDT). We also investigated the effect of the pure antiestrogen ICI 182,780 (ICI) on the expression patterns caused bymore » these compounds. Transcript levels were quantified 24 h after compound treatment using Illumina BeadChip Arrays. We identified 87 genes with similar expression changes in response to all EAC treatments in Ishikawa plus. ICI lowered the magnitude or reversed the expression of these genes, indicating ER dependent regulation. Apart from estrogenic gene regulation, Bisphenol A, o,p'-DDT, Zearalenone, Genistein and Resveratrol displayed similarities to ICI in their expression patterns, suggesting mixed estrogenic/antiestrogenic properties. In particular, the predominant antiestrogenic expression response of Resveratrol could be clearly distinguished from the other test compounds, indicating a distinct mechanism of action. Divergent gene expression patterns of the phytoestrogens, as well as weaker estrogenic gene expression regulation determined for the anthropogenous chemicals Bisphenol A and o,p'-DDT, warrants a careful assessment of potential detrimental and/or beneficial effects of EACs. The characteristic expression fingerprints and the identified subset of putative marker genes can be used for screening chemicals with an unknown mode of action and for predicting their potential to exert endocrine disrupting effects.« less

Transcriptomic profiling of human peritumoral neocortex tissues revealed genes possibly involved in tumor-induced epilepsy.

PubMed

Niesen, Charles E; Xu, Jun; Fan, Xuemo; Li, Xiaojin; Wheeler, Christopher J; Mamelak, Adam N; Wang, Charles

2013-01-01

The molecular mechanism underlying tumor-induced epileptogenesis is poorly understood. Alterations in the peritumoral microenvironment are believed to play a significant role in inducing epileptogenesis. We hypothesize that the change of gene expression in brain peritumoral tissues may contribute to the increased neuronal excitability and epileptogenesis. To identify the genes possibly involved in tumor-induced epilepsy, a genome-wide gene expression profiling was conducted using Affymetrix HG U133 plus 2.0 arrays and RNAs derived from formalin-fixed paraffin embedded (FFPE) peritumoral cortex tissue slides from 5-seizure vs. 5-non-seizure low grade brain tumor patients. We identified many differentially expressed genes (DEGs). Seven dysregulated genes (i.e., C1QB, CALCRL, CCR1, KAL1, SLC1A2, SSTR1 and TYRO3) were validated by qRT-PCR, which showed a high concordance. Principal Component Analysis (PCA) showed that epilepsy subjects were clustered together tightly (except one sample) and were clearly separated from the non-epilepsy subjects. Molecular functional categorization showed that significant portions of the DEGs functioned as receptor activity, molecular binding including enzyme binding and transcription factor binding. Pathway analysis showed these DEGs were mainly enriched in focal adhesion, ECM-receptor interaction, and cell adhesion molecules pathways. In conclusion, our study showed that dysregulation of gene expression in the peritumoral tissues may be one of the major mechanisms of brain tumor induced-epilepsy. However, due to the small sample size of the present study, further validation study is needed. A deeper characterization on the dysregulated genes involved in brain tumor-induced epilepsy may shed some light on the management of epilepsy due to brain tumors.

Overexpression of the S100A2 protein as a prognostic marker for patients with stage II and III colorectal cancer

PubMed Central

MASUDA, TAIKI; ISHIKAWA, TOSHIAKI; MOGUSHI, KAORU; OKAZAKI, SATOSHI; ISHIGURO, MEGUMI; IIDA, SATORU; MIZUSHIMA, HIROSHI; TANAKA, HIROSHI; UETAKE, HIROYUKI; SUGIHARA, KENICHI

2016-01-01

We aimed to identify a novel prognostic biomarker related to recurrence in stage II and III colorectal cancer (CRC) patients. Stage II and III CRC tissue mRNA expression was profiled using an Affymetrix Gene Chip, and copy number profiles of 125 patients were generated using an Affymetrix 250K Sty array. Genes showing both upregulated expression and copy number gains in cases involving recurrence were extracted as candidate biomarkers. The protein expression of the candidate gene was assessed using immunohistochemical staining of tissue from 161 patients. The relationship between protein expression and clinicopathological features was also examined. We identified 9 candidate genes related to recurrence of stage II and III CRC, whose mRNA expression was significantly higher in CRC than in normal tissue. Of these proteins, the S100 calcium-binding protein A2 (S100A2) has been observed in several human cancers. S100A2 protein overexpression in CRC cells was associated with significantly worse overall survival and relapse-free survival, indicating that S100A2 is an independent risk factor for stage II and III CRC recurrence. S100A2 overexpression in cancer cells could be a biomarker of poor prognosis in stage II and III CRC recurrence and a target for treatment of this disease. PMID:26783118

Signaling Pathways Involved in Lunar Dust Induced Cytotoxicity

NASA Technical Reports Server (NTRS)

Zhang, Ye; Lam, Chiu-Wing; Scully, Robert R.; Williams, Kyle; Zalesak, Selina; Wu, Honglu; James, John T.

2014-01-01

The Moon's surface is covered by a layer of fine, reactive dust. Lunar dust contain about 1-2% of very fine dust (< 3 micron), that is respirable. The habitable area of any lunar landing vehicle and outpost would inevitably be contaminated with lunar dust that could pose a health risk. The purpose of the study is to evaluate the toxicity of Apollo moon dust in rodents to assess the health risk of dust exposures to humans. One of the particular interests in the study is to evaluate dust-induced changes of the expression of fibrosis-related genes, and to identify specific signaling pathways involved in lunar dust-induced toxicity. F344 rats were exposed for 4 weeks (6h/d; 5d/wk) in nose-only inhalation chambers to concentrations of 0 (control air), 2.1, 6.1, 21, and 61 mg/m(exp 3) of lunar dust. Five rats per group were euthanized 1 day, 1 week, 1 month, and 3 months after the last inhalation exposure. The total RNAs were isolated from the blood or lung tissue after being lavaged, using the Qigen RNeasy kit. The Rat Fibrosis RT2 Profile PCR Array was used to profile the expression of 84 genes relevant to fibrosis. The genes with significant expression changes are identified and the gene expression data were further analyzed using IPA pathway analysis tool to determine the signaling pathways with significant changes.

Global characterization of copy number variants in epilepsy patients from whole genome sequencing

PubMed Central

Meloche, Caroline; Andrade, Danielle M.; Lafreniere, Ron G.; Gravel, Micheline; Spiegelman, Dan; Dionne-Laporte, Alexandre; Boelman, Cyrus; Hamdan, Fadi F.; Michaud, Jacques L.; Rouleau, Guy; Minassian, Berge A.; Bourque, Guillaume; Cossette, Patrick

2018-01-01

Epilepsy will affect nearly 3% of people at some point during their lifetime. Previous copy number variants (CNVs) studies of epilepsy have used array-based technology and were restricted to the detection of large or exonic events. In contrast, whole-genome sequencing (WGS) has the potential to more comprehensively profile CNVs but existing analytic methods suffer from limited accuracy. We show that this is in part due to the non-uniformity of read coverage, even after intra-sample normalization. To improve on this, we developed PopSV, an algorithm that uses multiple samples to control for technical variation and enables the robust detection of CNVs. Using WGS and PopSV, we performed a comprehensive characterization of CNVs in 198 individuals affected with epilepsy and 301 controls. For both large and small variants, we found an enrichment of rare exonic events in epilepsy patients, especially in genes with predicted loss-of-function intolerance. Notably, this genome-wide survey also revealed an enrichment of rare non-coding CNVs near previously known epilepsy genes. This enrichment was strongest for non-coding CNVs located within 100 Kbp of an epilepsy gene and in regions associated with changes in the gene expression, such as expression QTLs or DNase I hypersensitive sites. Finally, we report on 21 potentially damaging events that could be associated with known or new candidate epilepsy genes. Our results suggest that comprehensive sequence-based profiling of CNVs could help explain a larger fraction of epilepsy cases. PMID:29649218

Arabidopsis gene expression patterns during spaceflight

NASA Astrophysics Data System (ADS)

Paul, A.-L.; Ferl, R. J.

The exposure of Arabidopsis thaliana (Arabidopsis) plants to spaceflight environments resulted in the differential expression of hundreds of genes. A 5 day mission on orbiter Columbia in 1999 (STS-93) carried transgenic Arabidopsis plants engineered with a transgene composed of the alcohol dehydrogenase (Adh) gene promoter linked to the β -Glucuronidase (GUS) reporter gene. The plants were used to evaluate the effects of spaceflight on two fronts. First, expression patterns visualized with the Adh/GUS transgene were used to address specifically the possibility that spaceflight induces a hypoxic stress response, and to assess whether any spaceflight response was similar to control terrestrial hypoxia-induced gene expression patterns. (Paul et al., Plant Physiol. 2001, 126:613). Second, genome-wide patterns of native gene expression were evaluated utilizing the Affymetrix ATH1 GeneChip? array of 8,000 Arabidopsis genes. As a control for the veracity of the array analyses, a selection of genes identified with the arrays was further characterized with quantitative Real-Time RT PCR (ABI - TaqmanTM). Comparison of the patterns of expression for arrays of hybridized with RNA isolated from plants exposed to spaceflight compared to the control arrays revealed hundreds of genes that were differentially expressed in response to spaceflight, yet most genes that are hallmarks of hypoxic stress were unaffected. These results will be discussed in light of current models for plant responses to the spaceflight environment, and with regard to potential future flight opportunities.

Generation of miniaturized planar ecombinant antibody arrays using a microcantilever-based printer

NASA Astrophysics Data System (ADS)

Petersson, Linn; Berthet Duroure, Nathalie; Auger, Angèle; Dexlin-Mellby, Linda; Borrebaeck, Carl AK; Ait Ikhlef, Ali; Wingren, Christer

2014-07-01

Miniaturized (Ø 10 μm), multiplexed (>5-plex), and high-density (>100 000 spots cm-2) antibody arrays will play a key role in generating protein expression profiles in health and disease. However, producing such antibody arrays is challenging, and it is the type and range of available spotters which set the stage. This pilot study explored the use of a novel microspotting tool, BioplumeTM—consisting of an array of micromachined silicon cantilevers with integrated microfluidic channels—to produce miniaturized, multiplexed, and high-density planar recombinant antibody arrays for protein expression profiling which targets crude, directly labelled serum. The results demonstrated that 16-plex recombinant antibody arrays could be produced—based on miniaturized spot features (78.5 um2, Ø 10 μm) at a 7-125-times increased spot density (250 000 spots cm-2), interfaced with a fluorescent-based read-out. This prototype platform was found to display adequate reproducibility (spot-to-spot) and an assay sensitivity in the pM range. The feasibility of the array platform for serum protein profiling was outlined.

Improved Protein Arrays for Quantitative Systems Analysis of the Dynamics of Signaling Pathway Interactions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang, Chin-Rang

Astronauts and workers in nuclear plants who repeatedly exposed to low doses of ionizing radiation (IR, <10 cGy) are likely to incur specific changes in signal transduction and gene expression in various tissues of their body. Remarkable advances in high throughput genomics and proteomics technologies enable researchers to broaden their focus from examining single gene/protein kinetics to better understanding global gene/protein expression profiling and biological pathway analyses, namely Systems Biology. An ultimate goal of systems biology is to develop dynamic mathematical models of interacting biological systems capable of simulating living systems in a computer. This Glue Grant is to complementmore » Dr. Boothman’s existing DOE grant (No. DE-FG02-06ER64186) entitled “The IGF1/IGF-1R-MAPK-Secretory Clusterin (sCLU) Pathway: Mediator of a Low Dose IR-Inducible Bystander Effect” to develop sensitive and quantitative proteomic technology that suitable for low dose radiobiology researches. An improved version of quantitative protein array platform utilizing linear Quantum dot signaling for systematically measuring protein levels and phosphorylation states for systems biology modeling is presented. The signals are amplified by a confocal laser Quantum dot scanner resulting in ~1000-fold more sensitivity than traditional Western blots and show the good linearity that is impossible for the signals of HRP-amplification. Therefore this improved protein array technology is suitable to detect weak responses of low dose radiation. Software is developed to facilitate the quantitative readout of signaling network activities. Kinetics of EGFRvIII mutant signaling was analyzed to quantify cross-talks between EGFR and other signaling pathways.« less

Functional identification of genes responsible for the biosynthesis of 1-methoxy-indol-3-ylmethyl-glucosinolate in Brassica rapa ssp. chinensis

PubMed Central

2014-01-01

Background Brassica vegetables contain a class of secondary metabolites, the glucosinolates (GS), whose specific degradation products determine the characteristic flavor and smell. While some of the respective degradation products of particular GS are recognized as health promoting substances for humans, recent studies also show evidence that namely the 1-methoxy-indol-3-ylmethyl GS might be deleterious by forming characteristic DNA adducts. Therefore, a deeper knowledge of aspects involved in the biosynthesis of indole GS is crucial to design vegetables with an improved secondary metabolite profile. Results Initially the leafy Brassica vegetable pak choi (Brassica rapa ssp. chinensis) was established as suitable tool to elicit very high concentrations of 1-methoxy-indol-3-ylmethyl GS by application of methyl jasmonate. Differentially expressed candidate genes were discovered in a comparative microarray analysis using the 2 × 104 K format Brassica Array and compared to available gene expression data from the Arabidopsis AtGenExpress effort. Arabidopsis knock out mutants of the respective candidate gene homologs were subjected to a comprehensive examination of their GS profiles and confirmed the exclusive involvement of polypeptide 4 of the cytochrome P450 monooxygenase subfamily CYP81F in 1-methoxy-indol-3-ylmethyl GS biosynthesis. Functional characterization of the two identified isoforms coding for CYP81F4 in the Brassica rapa genome was performed using expression analysis and heterologous complementation of the respective Arabidopsis mutant. Conclusions Specific differences discovered in a comparative microarray and glucosinolate profiling analysis enables the functional attribution of Brassica rapa ssp. chinensis genes coding for polypeptide 4 of the cytochrome P450 monooxygenase subfamily CYP81F to their metabolic role in indole glucosinolate biosynthesis. These new identified Brassica genes will enable the development of genetic tools for breeding vegetables with improved GS composition in the near future. PMID:24886080

Projection-based estimation and nonuniformity correction of sensitivity profiles in phased-array surface coils.

PubMed

Yun, Sungdae; Kyriakos, Walid E; Chung, Jun-Young; Han, Yeji; Yoo, Seung-Schik; Park, Hyunwook

2007-03-01

To develop a novel approach for calculating the accurate sensitivity profiles of phased-array coils, resulting in correction of nonuniform intensity in parallel MRI. The proposed intensity-correction method estimates the accurate sensitivity profile of each channel of the phased-array coil. The sensitivity profile is estimated by fitting a nonlinear curve to every projection view through the imaged object. The nonlinear curve-fitting efficiently obtains the low-frequency sensitivity profile by eliminating the high-frequency image contents. Filtered back-projection (FBP) is then used to compute the estimates of the sensitivity profile of each channel. The method was applied to both phantom and brain images acquired from the phased-array coil. Intensity-corrected images from the proposed method had more uniform intensity than those obtained by the commonly used sum-of-squares (SOS) approach. With the use of the proposed correction method, the intensity variation was reduced to 6.1% from 13.1% of the SOS. When the proposed approach was applied to the computation of the sensitivity maps during sensitivity encoding (SENSE) reconstruction, it outperformed the SOS approach in terms of the reconstructed image uniformity. The proposed method is more effective at correcting the intensity nonuniformity of phased-array surface-coil images than the conventional SOS method. In addition, the method was shown to be resilient to noise and was successfully applied for image reconstruction in parallel imaging.

Luteolin and apigenin activate the Oct-4/Sox2 signal via NFATc1 in human periodontal ligament cells.

PubMed

Liu, Lu; Peng, Zhengjun; Huang, Haoquan; Xu, Zhezhen; Wei, Xi

2016-10-01

Identifying small molecules to activate the Oct-4/Sox2-derived pluripotency network represents a hopeful and safe method to pluripotency without genetic manipulation. Luteolin and apigenin, two major bioactive flavonoids, enhance reprogramming efficiency and increase expression of Oct-4/Sox2/c-Myc, albeit the detailed mechanism regulating pluripotency in dental-derived cells remains unknown. In the present study, to elucidate the effect of luteolin/apigenin on pluripotency of periodontal ligament cells (PDLCs) through interaction with downstream signals, we examined cell cycle, proliferation, apoptosis, expression of Oct-4/Sox2/c-Myc, and multilineage differentiation of PDLCs with luteolin/apigenin treatment. Moreover, we profiled the differentially expressed pluripotency genes by PCR arrays. Our results demonstrated that luteolin/apigenin restrained cell proliferation, increased apoptosis, and arrested PDLCs in G2/M and S phase. Luteolin and apigenin activated expression of Oct-4, Sox2, and c-Myc in a time- and dose-dependent pattern, and repressed lineage-specific differentiation. PCR arrays profiled multiple signals in PDLCs with luteolin/apigenin treatment, among which NFATc1 was the major upregulated gene. Notably, blocking of the NFATc1 signal with INCA-6 significantly decreased mRNA and protein expression of Oct-4, Sox2, and c-Myc in PDLCs with luteolin/apigenin treatment, indicating that NFATc1 may act as an upstream modulator of Oct-4/Sox2 signal. Taken together, this study showed that luteolin and apigenin effectively maintain pluripotency of PDLCs through activation of Oct-4/Sox2 signal via NFATc1. © 2016 International Federation for Cell Biology.

High-resolution array comparative genomic hybridization (aCGH) identifies copy number alterations in diffuse large B-cell lymphoma that predict response to immuno-chemotherapy

PubMed Central

Kreisel, F.; Kulkarni, S.; Kerns, R. T.; Hassan, A.; Deshmukh, H.; Nagarajan, R.; Frater, J. L.; Cashen, A.

2013-01-01

Despite recent attempts at sub-categorization, including gene expression profiling into prognostically different groups of “germinal center B-cell type” and “activated B-cell type”, diffuse large B-cell lymphoma (DLBCL) remains a biologically heterogenous tumor with no clear prognostic biomarkers to guide therapy. Whole genome, high resolution array comparative genomic hybridization (aCGH) was performed on 4 cases of chemoresistant DLBCL and 4 cases of chemo-responsive DLBCL to identify genetic differences which may correlate with response to R-CHOP therapy. Array CGH analysis identified 7 DNA copy number alteration (CNA) regions exclusive to the chemoresistant group, consisting of amplifications at 1p36.13, 1q42.3, 3p21.31, 7q11.23, and 16p13.3, and loss at 9p21.3, and 14p21.31. Copy number loss of the tumor suppressor genes CDKN2A (p16, p14) and CDKN2B (p15) at 9p21.3 was validated by fluorescence in situ hybridization and immunohistochemistry as independent techniques. In the chemo-sensitive group, 12 CNAs were detected consisting of segment gains on 1p36.11, 1p36.22, 2q11.2, 8q24.3, 12p13.33, and 22q13.2 and segment loss on 6p21.32. RUNX3, a tumor suppressor gene located on 1p36.11 and MTHFR, which encodes for the enzyme methylenetetrahydrofolate reductase, located on 1p36.22 are the only known genes in this group associated with lymphoma. Whole genome aCGH analysis has detected copy number alterations exclusive to either chemoresistant or chemo-responsive DLBCL that may represent consistent clonal changes predictive for prognosis and outcome of chemotherapy. PMID:21504712

Genome-scale approaches to the epigenetics of common human disease

PubMed Central

2011-01-01

Traditionally, the pathology of human disease has been focused on microscopic examination of affected tissues, chemical and biochemical analysis of biopsy samples, other available samples of convenience, such as blood, and noninvasive or invasive imaging of varying complexity, in order to classify disease and illuminate its mechanistic basis. The molecular age has complemented this armamentarium with gene expression arrays and selective analysis of individual genes. However, we are entering a new era of epigenomic profiling, i.e., genome-scale analysis of cell-heritable nonsequence genetic change, such as DNA methylation. The epigenome offers access to stable measurements of cellular state and to biobanked material for large-scale epidemiological studies. Some of these genome-scale technologies are beginning to be applied to create the new field of epigenetic epidemiology. PMID:19844740

Normal gene expression in male F344 rat nasal transitional and respiratory epithelium.

PubMed

Hester, Susan D; Benavides, Gina B; Sartor, Maureen; Yoon, Lawrence; Wolf, Douglas C; Morgan, Kevin T

2002-02-20

The nasal epithelium is an important target site for chemically-induced toxicity and carcinogenicity in rodents. Gene expression profiles were determined in order to provide normal baseline data for nasal transitional/respiratory epithelium from healthy rats. Cells lining the rat nasal passages were collected and gene expression analysis was performed using Clontech cDNA Rat Atlas 1.2 arrays (1185 genes). The percentages of genes within specific average expression ranges were 4.2% at 45,000-1000, 14.8% at 1000-200, 25.0% at 200-68, and 56.0% below 68. Nine out of a subset of ten genes were confirmed for relative signal intensity using quantitative real-time RT-PCR. The most highly expressed genes included those involved in phase I (e.g. cytochrome P450s) and phase II (e.g. glutathione S-transferases) xenobiotic metabolism, bioenergetics (e.g. cytochrome oxidase), osmotic balance (e.g. Na(+)/K(+) ATPase) and epithelial ionic homeostasis (e.g. ion channels). Such baseline data will contribute to further understanding the normal physiology of these cells and facilitate the interpretation of responses by the nasal epithelial cells to xenobiotic treatment or disease.

Innate and adaptive immunity gene expression of human keratinocytes cultured of severe burn injury.

PubMed

Noronha, Silvana Aparecida Alves Corrêa de; Noronha, Samuel Marcos Ribeiro de; Lanziani, Larissa Elias; Ferreira, Lydia Masako; Gragnani, Alfredo

2014-01-01

Evaluate the expression profile of genes related to Innate and Adaptive Immune System (IAIS) of human Primary Epidermal keratinocytes (hPEKP) of patients with severe burns. After obtaining viable fragments of skin with and without burning, culture hKEP was initiated by the enzymatic method using Dispase (Sigma-Aldrich). These cells were treated with Trizol(r) (Life Technologies) for extraction of total RNA. This was quantified and analyzed for purity for obtaining cDNA for the analysis of gene expression using specific IAIS PCR Arrays plates (SA Biosciences). After the analysis of gene expression we found that 63% of these genes were differentially expressed, of which 77% were repressed and 23% were hyper-regulated. Among these, the following genes (fold increase or decrease): IL8 (41), IL6 (32), TNF (-92), HLA-E (-86), LYS (-74), CCR6 (- 73), CD86 (-41) and HLA-A (-35). This study contributes to the understanding of the molecular mechanisms underlying wound infection caused by the burn. Furthermore, it may provide new strategies to restore normal expression of these genes and thereby change the healing process and improve clinical outcome.

Major carcinogenic pathways identified by gene expression analysis of peritoneal mesotheliomas following chemical treatment in F344 rats

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Yongbaek; Thai-Vu Ton; De Angelo, Anthony B.

2006-07-15

This study was performed to characterize the gene expression profile and to identify the major carcinogenic pathways involved in rat peritoneal mesothelioma (RPM) formation following treatment of Fischer 344 rats with o-nitrotoluene (o-NT) or bromochloracetic acid (BCA). Oligo arrays, with over 20,000 target genes, were used to evaluate o-NT- and BCA-induced RPMs, when compared to a non-transformed mesothelial cell line (Fred-PE). Analysis using Ingenuity Pathway Analysis software revealed 169 cancer-related genes that were categorized into binding activity, growth and proliferation, cell cycle progression, apoptosis, and invasion and metastasis. The microarray data were validated by positive correlation with quantitative real-time RT-PCRmore » on 16 selected genes including igf1, tgfb3 and nov. Important carcinogenic pathways involved in RPM formation included insulin-like growth factor 1 (IGF-1), p38 MAPkinase, Wnt/{beta}-catenin and integrin signaling pathways. This study demonstrated that mesotheliomas in rats exposed to o-NT- and BCA were similar to mesotheliomas in humans, at least at the cellular and molecular level.« less

WebaCGH: an interactive online tool for the analysis and display of array comparative genomic hybridisation data.

PubMed

Frankenberger, Casey; Wu, Xiaolin; Harmon, Jerry; Church, Deanna; Gangi, Lisa M; Munroe, David J; Urzúa, Ulises

2006-01-01

Gene copy number variations occur both in normal cells and in numerous pathologies including cancer and developmental diseases. Array comparative genomic hybridisation (aCGH) is an emerging technology that allows detection of chromosomal gains and losses in a high-resolution format. When aCGH is performed on cDNA and oligonucleotide microarrays, the impact of DNA copy number on gene transcription profiles may be directly compared. We have created an online software tool, WebaCGH, that functions to (i) upload aCGH and gene transcription results from multiple experiments; (ii) identify significant aberrant regions using a local Z-score threshold in user-selected chromosomal segments subjected to smoothing with moving averages; and (iii) display results in a graphical format with full genome and individual chromosome views. In the individual chromosome display, data can be zoomed in/out in both dimensions (i.e. ratio and physical location) and plotted features can have 'mouse over' linking to outside databases to identify loci of interest. Uploaded data can be stored indefinitely for subsequent retrieval and analysis. WebaCGH was created as a Java-based web application using the open-source database MySQL. WebaCGH is freely accessible at http://129.43.22.27/WebaCGH/welcome.htm Xiaolin Wu (forestwu@mail.nih.gov) or Ulises Urzúa (uurzua@med.uchile.cl).

Gene expression abnormalities in histologically normal breast epithelium from patients with luminal type of breast cancer.

PubMed

Zubor, Pavol; Hatok, Jozef; Moricova, Petra; Kajo, Karol; Kapustova, Ivana; Mendelova, Andrea; Racay, Peter; Danko, Jan

2015-05-01

The gene expression profile of breast cancer has been described as a great breakthrough on the way to comprehend differences in cancer origin, behavior and therapy. However, gene expression profile in histologically normal epithelium (HNEpi) which could harbor genetic abnormalities predisposing breast tissue to develop malignancy was minor scope for scientists in the past. Thus, we aimed to analyze gene expressions in HNEpi and breast cancer tissue (BCTis) in order to establish its value as potential diagnostic marker for cancer development. We evaluated a panel of disease-specific genes in luminal type (A/B) of breast cancer and tumor surrounding HNEpi by qRT-PCR Array in 32 microdissected samples. There was 20.2 and 2.4% deregulation rate in genes with at least 2-fold or 5-fold over-expression between luminal (A/B) type breast carcinomas and tumor surrounding HNEpi, respectively. The high-grade luminal carcinomas showed higher number of deregulated genes compared to low-grade cases (50.6 vs. 23.8% with at least 2-fold deregulation rate). The main overexpressed genes in HNEpi were KLK5, SCGB1D2, GSN, EGFR and NGFR. The significant differences in gene expression between BCTis and HNEpi samples were revealed for BAG1, C3, CCNA2, CD44, FGF1, FOSL1, ID2, IL6R, NGFB, NGFR, PAPPA, PLAU, SERPINB5, THBS1 and TP53 gene (p < 0.05) and BCL2L2, CTSB, ITGB4, JUN, KIT, KLF5, SCGB1D2, SCGB2A1, SERPINE1 (p < 0.01), and EGFR, GABRP, GSN, MAP2K7 and THBS2 (p < 0.001), and GSN, KLK5 (p < 0.0001). The ontological gene analyses revealed high deregulations in gene group directly associated with breast cancer prognosis and origin.

LS-CAP: an algorithm for identifying cytogenetic aberrations in hepatocellular carcinoma using microarray data.

PubMed

He, Xianmin; Wei, Qing; Sun, Meiqian; Fu, Xuping; Fan, Sichang; Li, Yao

2006-05-01

Biological techniques such as Array-Comparative genomic hybridization (CGH), fluorescent in situ hybridization (FISH) and affymetrix single nucleotide pleomorphism (SNP) array have been used to detect cytogenetic aberrations. However, on genomic scale, these techniques are labor intensive and time consuming. Comparative genomic microarray analysis (CGMA) has been used to identify cytogenetic changes in hepatocellular carcinoma (HCC) using gene expression microarray data. However, CGMA algorithm can not give precise localization of aberrations, fails to identify small cytogenetic changes, and exhibits false negatives and positives. Locally un-weighted smoothing cytogenetic aberrations prediction (LS-CAP) based on local smoothing and binomial distribution can be expected to address these problems. LS-CAP algorithm was built and used on HCC microarray profiles. Eighteen cytogenetic abnormalities were identified, among them 5 were reported previously, and 12 were proven by CGH studies. LS-CAP effectively reduced the false negatives and positives, and precisely located small fragments with cytogenetic aberrations.

Analysis of microRNA and gene expression profiling in triazole fungicide-treated HepG2 cell line.

PubMed

An, Yu Ri; Kim, Seung Jun; Oh, Moon-Ju; Kim, Hyun-Mi; Shim, Il-Seob; Kim, Pil-Je; Choi, Kyunghee; Hwang, Seung Yong

2013-01-07

MicroRNA (miRNA) plays an important role in various diseases and in cellular and molecular responses to toxicants. In the present study, we investigated differential expression of miRNAs in response to three triazole fungicides (myclobutanil, propiconazole, and triadimefon). The human hepatoma cell line (HepG2) was treated with the above triazoles for 3 h or 48 h. miRNA-based microarray experiments were carried out using the Agilent human miRNA v13 array. At early exposure (3h), six miRNAs were differentially expressed and at late exposure (48 h), three miRNAs were significantly expressed. Overall, this study provides an array of potential biomarkers for the above triazole fungicides. Furthermore, these miRNAs induced by triazoles could be the foundation for the development of a miRNA-based toxic biomarker library that can predict environmental toxicity. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Increased expression of resistin in ectopic endometrial tissue of women with endometriosis.

PubMed

Oh, Yoon Kyung; Ha, Young Ran; Yi, Kyong Wook; Park, Hyun Tae; Shin, Jung-Ho; Kim, Tak; Hur, Jun-Young

2017-11-01

Inflammation is a key process in the establishment and progression of endometriosis. Resistin, an adipocytokine, has biological properties linked to immunologic functions, but its role in endometriosis is unclear. Resistin gene expression was examined in eutopic and ectopic endometrial tissues from women with (n=25) or without (n=25) endometriosis. Resistin mRNA and protein levels were determined in endometrial tissue using quantitative real-time reverse transcription PCR and Western blotting, following adipokine profiling arrays. Resistin protein was detected in human endometrial tissues using an adipokine array test. Resistin mRNA and protein levels were significantly higher in ectopic endometrial tissue of patients with endometriosis than in normal eutopic endometrial tissue. Our results indicate that resistin is differentially expressed in endometrial tissues from women with endometriosis and imply a role for resistin in endometriosis-associated pelvic inflammation. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Profiling and bioinformatics analyses reveal differential circular RNA expression in radioresistant esophageal cancer cells.

PubMed

Su, Huafang; Lin, Fuqiang; Deng, Xia; Shen, Lanxiao; Fang, Ya; Fei, Zhenghua; Zhao, Lihao; Zhang, Xuebang; Pan, Huanle; Xie, Deyao; Jin, Xiance; Xie, Congying

2016-07-28

Acquired radioresistance during radiotherapy is considered as the most important reason for local tumor recurrence or treatment failure. Circular RNAs (circRNAs) have recently been identified as microRNA sponges and involve in various biological processes. The purpose of this study is to investigate the role of circRNAs in the radioresistance of esophageal cancer. Total RNA was isolated from human parental cell line KYSE-150 and self-established radioresistant esophageal cancer cell line KYSE-150R, and hybridized to Arraystar Human circRNA Array. Quantitative real-time PCR was used to confirm the circRNA expression profiles obtained from the microarray data. Bioinformatic tools including gene ontology (GO) analysis, KEGG pathway analysis and network analysis were done for further assessment. Among the detected candidate 3752 circRNA genes, significant upregulation of 57 circRNAs and downregulation of 17 circRNAs in human radioresistant esophageal cancer cell line KYSE-150R were observed compared with the parental cell line KYSE-150 (fold change ≥2.0 and P < 0.05). There were 9 out of these candidate circRNAs were validated by real-time PCR. GO analysis revealed that numerous target genes, including most microRNAs were involved in the biological processes. There were more than 400 target genes enrichment on Wnt signaling pathway. CircRNA_001059 and circRNA_000167 were the two largest nodes in circRNA/microRNA co-expression network. Our study revealed a comprehensive expression and functional profile of differentially expressed circRNAs in radioresistant esophageal cancer cells, indicating possible involvement of these dysregulated circRNAs in the development of radiation resistance.

Transcript and protein profiling identifies signaling, growth arrest, apoptosis, and NF-κB survival signatures following GNRH receptor activation

PubMed Central

Meyer, Colette; Sims, Andrew H; Morgan, Kevin; Harrison, Beth; Muir, Morwenna; Bai, Jianing; Faratian, Dana; Millar, Robert P; Langdon, Simon P

2013-01-01

GNRH significantly inhibits proliferation of a proportion of cancer cell lines by activating GNRH receptor (GNRHR)-G protein signaling. Therefore, manipulation of GNRHR signaling may have an under-utilized role in treating certain breast and ovarian cancers. However, the precise signaling pathways necessary for the effect and the features of cellular responses remain poorly defined. We used transcriptomic and proteomic profiling approaches to characterize the effects of GNRHR activation in sensitive cells (HEK293-GNRHR, SCL60) in vitro and in vivo, compared to unresponsive HEK293. Analyses of gene expression demonstrated a dynamic response to the GNRH superagonist Triptorelin. Early and mid-phase changes (0.5–1.0 h) comprised mainly transcription factors. Later changes (8–24 h) included a GNRH target gene, CGA, and up- or downregulation of transcripts encoding signaling and cell division machinery. Pathway analysis identified altered MAPK and cell cycle pathways, consistent with occurrence of G2/M arrest and apoptosis. Nuclear factor kappa B (NF-κB) pathway gene transcripts were differentially expressed between control and Triptorelin-treated SCL60 cultures. Reverse-phase protein and phospho-proteomic array analyses profiled responses in cultured cells and SCL60 xenografts in vivo during Triptorelin anti-proliferation. Increased phosphorylated NF-κB (p65) occurred in SCL60 in vitro, and p-NF-κB and IκBϵ were higher in treated xenografts than controls after 4 days Triptorelin. NF-κB inhibition enhanced the anti-proliferative effect of Triptorelin in SCL60 cultures. This study reveals details of pathways interacting with intense GNRHR signaling, identifies potential anti-proliferative target genes, and implicates the NF-κB survival pathway as a node for enhancing GNRH agonist-induced anti-proliferation. PMID:23202794

Facilitated beam-walking recovery during acute phase by kynurenic acid treatment in a rat model of photochemically induced thrombosis causing focal cerebral ischemia.

PubMed

Abo, Masahiro; Yamauchi, Hideki; Suzuki, Masahiko; Sakuma, Mio; Urashima, Mitsuyoshi

We previously demonstrated the presence of activated areas in the non-injured contralateral sensorimotor cortex in addition to the ipsilateral sensorimotor cortex of the area surrounding a brain infarction, using a rat model of focal photochemically induced thrombosis (PIT) and functional magnetic resonance imaging. Using this model, we next applied gene expression profiling to screen key molecules upregulated in the activated area. RNA was extracted from the ipsilateral and contralateral sensorimotor cortex to the focal brain infarction and from the sham controlled cortex, and hybridized to gene-expression profiling arrays containing 1,322 neurology-related genes. Results showed that glycine receptors were upregulated in both the ipsilateral and contralateral cortex to the focal ischemic lesion. To prove the preclinical significance of upregulated glycine receptors, kynurenic acid, an endogenous antagonist to glycine receptors on neuronal cells, was administered intrathecally. As a result, the kynurenic acid significantly improved behavioral recovery within 10 days from paralysis induced by the focal PIT (p < 0.0001), as evaluated with beam walking. These results suggest that intrathecal administration of a glycine receptor antagonist may facilitate behavioral recovery during the acute phase after brain infarction. Copyright (c) 2006 S. Karger AG, Basel.

Apoptosis Induction and Gene Expression Profile Alterations of Cutaneous T-Cell Lymphoma Cells following Their Exposure to Bortezomib and Methotrexate

PubMed Central

Kontsioti, Frieda; Konsta, Eugene; Vikentiou, Miriam; Spathis, Aris; Papageorgiou, Sotiris; Vasilatou, Diamantina; Gkontopoulos, Konstantinos; Mpazani, Efthimia; Karakitsos, Petros; Rigopoulos, Dimitrios; Dimitriadis, George

2017-01-01

Mycosis fungoides (MF) and its leukemic variant Sézary syndrome (SS) comprise the majority of CTCL, a heterogenous group of non-Hodgkins lymphomas involving the skin. The CTCL’s resistance to chemotherapy and the lack of full understanding of their pathogenesis request further investigation. With the view of a more targeted therapy, we evaluated in vitro the effectiveness of bortezomib and methotrexate, as well as their combination in CTCL cell lines, regarding apoptosis induction. Our data are of clinical value and indicate that the bortezomib/methotrexate combinational therapy has an inferior impact on the apoptosis of CTCL compared to monotherapy, with bortezomib presenting as the most efficient treatment option for SS and methotrexate for MF. Using PCR arrays technology, we also investigated the alterations in the expression profile of genes related to DNA repair pathways in CTCL cell lines after treatment with bortezomib or methotrexate. We found that both agents, but mostly bortezomib, significantly deregulate a large number of genes in SS and MF cell lines, suggesting another pathway through which these agents could induce apoptosis in CTCL. Finally, we show that SS and MF respond differently to treatment, verifying their distinct nature and further emphasizing the need for discrete treatment approaches. PMID:28107479

A 19-SNP coronary heart disease gene score profile in subjects with type 2 diabetes: the coronary heart disease risk in type 2 diabetes (CoRDia study) study baseline characteristics.

PubMed

Beaney, Katherine E; Ward, Claire E; Bappa, Dauda A S; McGale, Nadine; Davies, Anna K; Hirani, Shashivadan P; Li, KaWah; Howard, Philip; Vance, Dwaine R; Crockard, Martin A; Lamont, John V; Newman, Stanton; Humphries, Steve E

2016-10-03

The coronary risk in diabetes (CoRDia) trial (n = 211) compares the effectiveness of usual diabetes care with a self-management intervention (SMI), with and without personalised risk information (including genetics), on clinical and behavioural outcomes. Here we present an assessment of randomisation, the cardiac risk genotyping assay, and the genetic characteristics of the recruits. Ten-year coronary heart disease (CHD) risk was calculated using the UKPDS score. Genetic CHD risk was determined by genotyping 19 single nucleotide polymorphisms (SNPs) using Randox's Cardiac Risk Prediction Array and calculating a gene score (GS). Accuracy of the array was assessed by genotyping a subset of pre-genotyped samples (n = 185). Overall, 10-year CHD risk ranged from 2-72 % but did not differ between the randomisation groups (p = 0.13). The array results were 99.8 % concordant with the pre-determined genotypes. The GS did not differ between the Caucasian participants in the CoRDia SMI plus risk group (n = 66) (p = 0.80) and a sample of UK healthy men (n = 1360). The GS was also associated with LDL-cholesterol (p = 0.05) and family history (p = 0.03) in a sample of UK healthy men (n = 1360). CHD risk is high in this group of T2D subjects. The risk array is an accurate genotyping assay, and is suitable for estimating an individual's genetic CHD risk. Trial registration This study has been registered at ClinicalTrials.gov; registration identifier NCT01891786.

An integration of genome-wide association study and gene expression profiling to prioritize the discovery of novel susceptibility Loci for osteoporosis-related traits.

PubMed

Hsu, Yi-Hsiang; Zillikens, M Carola; Wilson, Scott G; Farber, Charles R; Demissie, Serkalem; Soranzo, Nicole; Bianchi, Estelle N; Grundberg, Elin; Liang, Liming; Richards, J Brent; Estrada, Karol; Zhou, Yanhua; van Nas, Atila; Moffatt, Miriam F; Zhai, Guangju; Hofman, Albert; van Meurs, Joyce B; Pols, Huibert A P; Price, Roger I; Nilsson, Olle; Pastinen, Tomi; Cupples, L Adrienne; Lusis, Aldons J; Schadt, Eric E; Ferrari, Serge; Uitterlinden, André G; Rivadeneira, Fernando; Spector, Timothy D; Karasik, David; Kiel, Douglas P

2010-06-10

Osteoporosis is a complex disorder and commonly leads to fractures in elderly persons. Genome-wide association studies (GWAS) have become an unbiased approach to identify variations in the genome that potentially affect health. However, the genetic variants identified so far only explain a small proportion of the heritability for complex traits. Due to the modest genetic effect size and inadequate power, true association signals may not be revealed based on a stringent genome-wide significance threshold. Here, we take advantage of SNP and transcript arrays and integrate GWAS and expression signature profiling relevant to the skeletal system in cellular and animal models to prioritize the discovery of novel candidate genes for osteoporosis-related traits, including bone mineral density (BMD) at the lumbar spine (LS) and femoral neck (FN), as well as geometric indices of the hip (femoral neck-shaft angle, NSA; femoral neck length, NL; and narrow-neck width, NW). A two-stage meta-analysis of GWAS from 7,633 Caucasian women and 3,657 men, revealed three novel loci associated with osteoporosis-related traits, including chromosome 1p13.2 (RAP1A, p = 3.6x10(-8)), 2q11.2 (TBC1D8), and 18q11.2 (OSBPL1A), and confirmed a previously reported region near TNFRSF11B/OPG gene. We also prioritized 16 suggestive genome-wide significant candidate genes based on their potential involvement in skeletal metabolism. Among them, 3 candidate genes were associated with BMD in women. Notably, 2 out of these 3 genes (GPR177, p = 2.6x10(-13); SOX6, p = 6.4x10(-10)) associated with BMD in women have been successfully replicated in a large-scale meta-analysis of BMD, but none of the non-prioritized candidates (associated with BMD) did. Our results support the concept of our prioritization strategy. In the absence of direct biological support for identified genes, we highlighted the efficiency of subsequent functional characterization using publicly available expression profiling relevant to the skeletal system in cellular or whole animal models to prioritize candidate genes for further functional validation.

A microfabricated low-profile wideband antenna array for terahertz communications.

PubMed

Luk, K M; Zhou, S F; Li, Y J; Wu, F; Ng, K B; Chan, C H; Pang, S W

2017-04-28

While terahertz communications are considered to be the future solutions for the increasing demands on bandwidth, terahertz equivalents of radio frequency front-end components have not been realized. It remains challenging to achieve wideband, low profile antenna arrays with highly directive beams of radiation. Here, based on the complementary antenna approach, a wideband 2 × 2 cavity-backed slot antenna array with a corrugated surface is proposed. The approach is based on a unidirectional antenna with a cardiac radiation pattern and stable frequency characteristics that is achieved by integrating a series-resonant electric dipole with a parallel-resonant magnetic dipole. In this design, the slots work as magnetic dipoles while the corrugated surface radiates as an array of electric dipoles. The proposed antenna is realized at 1 THz operating frequency by stacking multiple metallized layers using the microfabrication technology. S-parameter measurements of this terahertz low-profile metallic antenna array demonstrate high efficiency at terahertz frequencies. Fractional bandwidth and gain are measured to be 26% and 14 dBi which are consistent with the simulated results. The proposed antenna can be used as the building block for larger antenna arrays with more directive beams, paving the way to develop high gain low-profile antennas for future communication needs.

Gene Expression Profiling in Limb-Girdle Muscular Dystrophy 2A

PubMed Central

Sáenz, Amets; Azpitarte, Margarita; Armañanzas, Rubén; Leturcq, France; Alzualde, Ainhoa; Inza, Iñaki; García-Bragado, Federico; De la Herran, Gaspar; Corcuera, Julián; Cabello, Ana; Navarro, Carmen; De la Torre, Carolina; Gallardo, Eduard; Illa, Isabel; de Munain, Adolfo López

2008-01-01

Limb-girdle muscular dystrophy type 2A (LGMD2A) is a recessive genetic disorder caused by mutations in calpain 3 (CAPN3). Calpain 3 plays different roles in muscular cells, but little is known about its functions or in vivo substrates. The aim of this study was to identify the genes showing an altered expression in LGMD2A patients and the possible pathways they are implicated in. Ten muscle samples from LGMD2A patients with in which molecular diagnosis was ascertained were investigated using array technology to analyze gene expression profiling as compared to ten normal muscle samples. Upregulated genes were mostly those related to extracellular matrix (different collagens), cell adhesion (fibronectin), muscle development (myosins and melusin) and signal transduction. It is therefore suggested that different proteins located or participating in the costameric region are implicated in processes regulated by calpain 3 during skeletal muscle development. Genes participating in the ubiquitin proteasome degradation pathway were found to be deregulated in LGMD2A patients, suggesting that regulation of this pathway may be under the control of calpain 3 activity. As frizzled-related protein (FRZB) is upregulated in LGMD2A muscle samples, it could be hypothesized that β-catenin regulation is also altered at the Wnt signaling pathway, leading to an incorrect myogenesis. Conversely, expression of most transcription factor genes was downregulated (MYC, FOS and EGR1). Finally, the upregulation of IL-32 and immunoglobulin genes may induce the eosinophil chemoattraction explaining the inflammatory findings observed in presymptomatic stages. The obtained results try to shed some light on identification of novel therapeutic targets for limb-girdle muscular dystrophies. PMID:19015733

Differential Gene Expression in Gonadotropin-Releasing Hormone Neurons of Male and Metestrous Female Mice.

PubMed

Vastagh, Csaba; Rodolosse, Annie; Solymosi, Norbert; Farkas, Imre; Auer, Herbert; Sárvári, Miklós; Liposits, Zsolt

2015-01-01

Gonadotropin-releasing hormone (GnRH) neurons play a pivotal role in the regulation of the hypothalamic-pituitary gonadal axis in a sex-specific manner. We hypothesized that the differences seen in reproductive functions of males and females are associated with a sexually dimorphic gene expression profile of GnRH neurons. We compared the transcriptome of GnRH neurons obtained from intact metestrous female and male GnRH-green fluorescent protein transgenic mice. About 1,500 individual GnRH neurons from each sex were sampled with laser capture microdissection followed by whole-transcriptome amplification for gene expression profiling. Under stringent selection criteria (fold change >1.6, adjusted p value 0.01), Affymetrix Mouse Genome 430 PM array analysis identified 543 differentially expressed genes. Sexual dimorphism was most apparent in gene clusters associated with synaptic communication, signal transduction, cell adhesion, vesicular transport and cell metabolism. To validate microarray results, 57 genes were selected, and 91% of their differential expression was confirmed by real-time PCR. Similarly, 88% of microarray results were confirmed with PCR from independent samples obtained by patch pipette harvesting and pooling of 30 GnRH neurons from each sex. We found significant differences in the expression of genes involved in vesicle priming and docking (Syt1, Cplx1), GABAergic (Gabra3, Gabrb3, Gabrg2) and glutamatergic (Gria1, Grin1, Slc17a6) neurotransmission, peptide signaling (Sstr3, Npr2, Cxcr4) and the regulation of intracellular ion homeostasis (Cacna1, Cacnb1, Cacng5, Kcnq2, Kcnc1). The striking sexual dimorphism of the GnRH neuron transcriptome we report here contributes to a better understanding of the differences in cellular mechanisms of GnRH neurons in the two sexes. © 2015 S. Karger AG, Basel.

Messenger RNA profiling of human platelets by microarray hybridization.

PubMed

Bugert, Peter; Dugrillon, Alex; Günaydin, Ayse; Eichler, Hermann; Klüter, Harald

2003-10-01

Platelets are generally believed to be inactive in terms of de novo protein synthesis. On the other hand, the presence of ribosomes and mRNA molecules is well established. Many studies have used reverse transcriptase (RT) -PCR for detection of gene transcripts in platelets. As RT-PCR is a very sensitive method, any leukocyte contamination of platelet preparations can lead to false results. We performed three filtration procedures to minimize leukocyte contamination of pooled buffy-coat platelet concentrates prior to RNA isolation. Furthermore, by applying a genomic PCR approach with 50 amplification cycles we demonstrated that nucleated cells were not detectable. Microarray hybridization was used to analyze 9,850 individual human genes in RNA from purified platelets. In total we identified 1,526 (15.5%) positive genes. The data were confirmed in six individual experiments each performed on a PC pooled from four individual blood donations. Genes specific for nucleated blood cells such as CD4, CD83 and others were negative and verified the purity of PC. Overrepresentation of positive genes was found in the functional categories of glycoproteins/integrins (22.6% vs. 15.5%, p=0.029) and receptors (20.7% vs. 15.5%, p<0.001). Gene transcripts encoding RANTES, GRO-alpha, MIP-1alpha, MIP-1beta, and others were found at high levels of signal intensity and confirmed literature data. This work provides a mRNA profile of human platelets and a complete list of results can be downloaded from the website of our institute www.ma.uni-heidelberg.de/inst/iti/plt_array.xls. The knowledge about gene transcripts may have an impact on the characterization of novel proteins and their functions in platelets.

Measurement of indicator genes using global complementary DNA (cDNA) amplification, by polyadenylic acid reverse transcriptase polymerase chain reaction (poly A RT-PCR): A feasibility study using paired samples from tissue and ductal juice in patients undergoing pancreatoduodenectomy.

PubMed

Sanyal, Sudip; Siriwardena, Ajith K; Byers, Richard

2018-06-01

The aim of this study is to compare gene expression profiles in RNA isolated from pancreatic ductal juice with the RNA expression profiles of the same genes from matched intra-operative tissue samples from pancreatic tumours. Intra-operative sampling of pancreatic juice and collection of matched tissue samples was undertaken in patients undergoing pancreatoduodenectomy for clinically suspected pancreatic cancer and a precursor lesion, main-duct intraductal papillary mucinous neoplasm. RNA was isolated and Poly A PCR was used to globally amplify the RNA. Real-time polymerase chain reaction (RT-PCR) was used to measure expression levels of 17 genes selected from microarray studies. Spearman's rank correlation test was used to examine the relationship of gene expression between pancreatic juice and tissue. The study was approved by Regional Ethics Committee. Mesothelin (MSLN) showed significant correlation (p < 0.008) in expression levels between paired pancreatic juice and tissue samples in pancreas cancer. In intraductal papillary mucinous neoplasms (IPMN), Matrix Metalloproteinase 7 (MMP7), showed significant correlation (p < 0.01) in the expression levels between paired pancreatic juice and tissue samples. This study confirms that RNA analysis of paired pancreatic juice and tissue samples and establishment of cDNA using poly A PCR is technically feasible. Application of the technique to non-invasively obtained pancreatic juice during endoscopic assessment of tumours and the use of gene arrays of cancer indicator genes are the next steps in development of this technique. Copyright © 2018 IAP and EPC. Published by Elsevier B.V. All rights reserved.

Overcoming chemotherapy resistance of ovarian cancer cells by liposomal cisplatin: molecular mechanisms unveiled by gene expression profiling.

PubMed

Koch, Martin; Krieger, Michaela L; Stölting, Daniel; Brenner, Norbert; Beier, Manfred; Jaehde, Ulrich; Wiese, Michael; Royer, Hans-Dieter; Bendas, Gerd

2013-04-15

Previously we reported that liposomal cisplatin (CDDP) overcomes CDDP resistance of ovarian A2780cis cancer cells (Krieger et al., Int. J. Pharm. 389, 2010, 10-17). Here we find that the cytotoxic activity of liposomal CDDP is not associated with detectable DNA platination in resistant ovarian cancer cells. This suggests that the mode of action of liposomal CDDP is different from the free drug. To gain insight into mechanisms of liposomal CDDP activity, we performed a transcriptome analysis of untreated A2780cis cells, and A2780cis cells in response to exposure with IC50 values of free or liposomal CDDP. A process network analysis of upregulated genes showed that liposomal CDDP induced a highly different gene expression profile in comparison to the free drug. p53 was identified as a key player directing transcriptional responses to free or liposomal CDDP. The free drug induced expression of essential genes of the intrinsic (mitochondrial) apoptosis pathway (BAX, BID, CASP9) most likely through p38MAPK activation. In contrast, liposomal CDDP induced expression of genes from DNA damage pathways and several genes of the extrinsic pathway of apoptosis (TNFRSF10B-DR5, CD70-TNFSF7). It thus appears that liposomal CDDP overcomes CDDP resistance by inducing DNA damage and in consequence programmed cell death by the extrinsic pathway. Predictions from gene expression data with respect to apoptosis activation were confirmed at the protein level by an apoptosis antibody array. This sheds new light on liposomal drug carrier approaches in cancer and suggests liposomal CDDP as promising strategy for the treatment of CDDP resistant ovarian carcinomas. Copyright © 2013 Elsevier Inc. All rights reserved.

Defining the gene expression signature of rhabdomyosarcoma by meta-analysis

PubMed Central

Romualdi, Chiara; De Pittà, Cristiano; Tombolan, Lucia; Bortoluzzi, Stefania; Sartori, Francesca; Rosolen, Angelo; Lanfranchi, Gerolamo

2006-01-01

Background Rhabdomyosarcoma is a highly malignant soft tissue sarcoma in childhood and arises as a consequence of regulatory disruption of the growth and differentiation pathways of myogenic precursor cells. The pathogenic pathways involved in this tumor are mostly unknown and therefore a better characterization of RMS gene expression profile would represent a considerable advance. The availability of publicly available gene expression datasets have opened up new challenges especially for the integration of data generated by different research groups and different array platforms with the purpose of obtaining new insights on the biological process investigated. Results In this work we performed a meta-analysis on four microarray and two SAGE datasets of gene expression data on RMS in order to evaluate the degree of agreement of the biological results obtained by these different studies and to identify common regulatory pathways that could be responsible of tumor growth. Regulatory pathways and biological processes significantly enriched has been investigated and a list of differentially meta-profiles have been identified as possible candidate of aggressiveness of RMS. Conclusion Our results point to a general down regulation of the energy production pathways, suggesting a hypoxic physiology for RMS cells. This result agrees with the high malignancy of RMS and with its resistance to most of the therapeutic treatments. In this context, different isoforms of the ANT gene have been consistently identified for the first time as differentially expressed in RMS. This gene is involved in anti-apoptotic processes when cells grow in low oxygen conditions. These new insights in the biological processes responsible of RMS growth and development demonstrate the effective advantage of the use of integrated analysis of gene expression studies. PMID:17090319

Altered Cytokine Gene Expression in Peripheral Blood Monocytes across the Menstrual Cycle in Primary Dysmenorrhea: A Case-Control Study

PubMed Central

Ma, Hongyue; Hong, Min; Duan, Jinao; Liu, Pei; Fan, Xinsheng; Shang, Erxin; Su, Shulan; Guo, Jianming; Qian, Dawei; Tang, Yuping

2013-01-01

Primary dysmenorrhea is one of the most common gynecological complaints in young women, but potential peripheral immunologic features underlying this condition remain undefined. In this paper, we compared 84 common cytokine gene expression profiles of peripheral blood mononuclear cells (PBMCs) from six primary dysmenorrheic young women and three unaffected controls on the seventh day before (secretory phase), and the first (menstrual phase) and the fifth (regenerative phase) days of menstruation, using a real-time PCR array assay combined with pattern recognition and gene function annotation methods. Comparisons between dysmenorrhea and normal control groups identified 11 (nine increased and two decreased), 14 (five increased and nine decreased), and 15 (seven increased and eight decreased) genes with ≥2-fold difference in expression (P<0.05) in the three phases of menstruation, respectively. In the menstrual phase, genes encoding pro-inflammatory cytokines (IL1B, TNF, IL6, and IL8) were up-regulated, and genes encoding TGF-β superfamily members (BMP4, BMP6, GDF5, GDF11, LEFTY2, NODAL, and MSTN) were down-regulated. Functional annotation revealed an excessive inflammatory response and insufficient TGF-β superfamily member signals with anti-inflammatory consequences, which may directly contribute to menstrual pain. In the secretory and regenerative phases, increased expression of pro-inflammatory cytokines and decreased expression of growth factors were also observed. These factors may be involved in the regulation of decidualization, endometrium breakdown and repair, and indirectly exacerbate primary dysmenorrhea. This first study of cytokine gene expression profiles in PBMCs from young primary dysmenorrheic women demonstrates a shift in the balance between expression patterns of pro-inflammatory cytokines and TGF-β superfamily members across the whole menstrual cycle, underlying the peripheral immunologic features of primary dysmenorrhea. PMID:23390521

Challenging the Cancer Molecular Stratification Dogma: Intratumoral Heterogeneity Undermines Consensus Molecular Subtypes and Potential Diagnostic Value in Colorectal Cancer.

PubMed

Dunne, Philip D; McArt, Darragh G; Bradley, Conor A; O'Reilly, Paul G; Barrett, Helen L; Cummins, Robert; O'Grady, Tony; Arthur, Ken; Loughrey, Maurice B; Allen, Wendy L; McDade, Simon S; Waugh, David J; Hamilton, Peter W; Longley, Daniel B; Kay, Elaine W; Johnston, Patrick G; Lawler, Mark; Salto-Tellez, Manuel; Van Schaeybroeck, Sandra

2016-08-15

A number of independent gene expression profiling studies have identified transcriptional subtypes in colorectal cancer with potential diagnostic utility, culminating in publication of a colorectal cancer Consensus Molecular Subtype classification. The worst prognostic subtype has been defined by genes associated with stem-like biology. Recently, it has been shown that the majority of genes associated with this poor prognostic group are stromal derived. We investigated the potential for tumor misclassification into multiple diagnostic subgroups based on tumoral region sampled. We performed multiregion tissue RNA extraction/transcriptomic analysis using colorectal-specific arrays on invasive front, central tumor, and lymph node regions selected from tissue samples from 25 colorectal cancer patients. We identified a consensus 30-gene list, which represents the intratumoral heterogeneity within a cohort of primary colorectal cancer tumors. Using a series of online datasets, we showed that this gene list displays prognostic potential HR = 2.914 (confidence interval 0.9286-9.162) in stage II/III colorectal cancer patients, but in addition, we demonstrated that these genes are stromal derived, challenging the assumption that poor prognosis tumors with stem-like biology have undergone a widespread epithelial-mesenchymal transition. Most importantly, we showed that patients can be simultaneously classified into multiple diagnostically relevant subgroups based purely on the tumoral region analyzed. Gene expression profiles derived from the nonmalignant stromal region can influence assignment of colorectal cancer transcriptional subtypes, questioning the current molecular classification dogma and highlighting the need to consider pathology sampling region and degree of stromal infiltration when employing transcription-based classifiers to underpin clinical decision making in colorectal cancer. Clin Cancer Res; 22(16); 4095-104. ©2016 AACRSee related commentary by Morris and Kopetz, p. 3989. ©2016 American Association for Cancer Research.

Genome-wide profiling of chromosomal alterations in renal cell carcinoma using high-density single nucleotide polymorphism arrays

PubMed Central

Chen, Meng; Ye, Yuanqing; Yang, Hushan; Tamboli, Pheroze; Matin, Surena; Tannir, Nizar M.; Wood, Christopher G.; Gu, Jian; Wu, Xifeng

2009-01-01

Purpose The identification of genetic aberrations may help understand the mechanisms of tumorigenesis and has important implications in diagnosis, prognosis, and treatment. Methods We applied Illumina's 317K high-density SNP-arrays to profile chromosomal aberrations in clear cell renal cell carcinoma (ccRCC) from 80 patients and analyzed the association of LOH/amplification events with clinicopathological characteristics and telomere length. Results The most common loss of heterozygosity (LOH) were 3p (69 cases) including 38 whole 3p arm losses, 30 large fragment LOH (spanning 3p21-36), and 1 interstitial LOH (spanning 3p12-14, 3p21-22, 3p24.1-24.2, and 3p24.3), followed by chromosome losses at 8p12-pter, 6q23.3-27, 14q24.1-qter, 9q32-qter, 10q22.3-qter, 9p13.3-pter, 4q28.3-qter, and 13q12.1-21.1. We also found several smallest overlapping regions of LOH that contained tumor suppressor genes. One smallest LOH in 8p12 had a size of 0.29 Mb and only contained one gene (NRG1). The most frequent chromosome gains were at 5q (32 cases), including 10 whole 5q amplification, 21 large amplifications encompassing 5q32-ter, and 1 focal amplification in 5q35.3 (0.42 Mb). The other common chromosome gains were 1q25.1-qter, 7q21.13-qter, 8q24.12-qter, and whole 7p arm. Significant associations of LOH at 9p, 9q, 14q, and 18q were observed with higher nuclear grade. Significant associations with tumor stage were observed for LOH at 14q, 18p, and 21q. Finally, we found that tumors with LOH at 2q, 6p, 6q, 9p, 9q, and 17p had significantly shorter telomere length than those without LOH. Conclusion This is the first study to use Illumina's SNP-CGH array that provides a close estimate of the size and frequency of chromosome LOH and amplifications of ccRCC. The identified regions and genes may become diagnostic and prognostic biomarkers as well as potential targets of therapy. PMID:19521957

Evidence for Alteration of Gene Regulatory Networks through MicroRNAs of the HIV-infected brain: novel analysis of retrospective cases.

PubMed

Tatro, Erick T; Scott, Erick R; Nguyen, Timothy B; Salaria, Shahid; Banerjee, Sugato; Moore, David J; Masliah, Eliezer; Achim, Cristian L; Everall, Ian P

2010-04-26

HIV infection disturbs the central nervous system (CNS) through inflammation and glial activation. Evidence suggests roles for microRNA (miRNA) in host defense and neuronal homeostasis, though little is known about miRNAs' role in HIV CNS infection. MiRNAs are non-coding RNAs that regulate gene translation through post-transcriptional mechanisms. Messenger-RNA profiling alone is insufficient to elucidate the dynamic dance of molecular expression of the genome. We sought to clarify RNA alterations in the frontal cortex (FC) of HIV-infected individuals and those concurrently infected and diagnosed with major depressive disorder (MDD). This report is the first published study of large-scale miRNA profiling from human HIV-infected FC. The goals of this study were to: 1. Identify changes in miRNA expression that occurred in the frontal cortex (FC) of HIV individuals, 2. Determine whether miRNA expression profiles of the FC could differentiate HIV from HIV/MDD, and 3. Adapt a method to meaningfully integrate gene expression data and miRNA expression data in clinical samples. We isolated RNA from the FC (n = 3) of three separate groups (uninfected controls, HIV, and HIV/MDD) and then pooled the RNA within each group for use in large-scale miRNA profiling. RNA from HIV and HIV/MDD patients (n = 4 per group) were also used for non-pooled mRNA analysis on Affymetrix U133 Plus 2.0 arrays. We then utilized a method for integrating the two datasets in a Target Bias Analysis. We found miRNAs of three types: A) Those with many dysregulated mRNA targets of less stringent statistical significance, B) Fewer dysregulated target-genes of highly stringent statistical significance, and C) unclear bias. In HIV/MDD, more miRNAs were downregulated than in HIV alone. Specific miRNA families at targeted chromosomal loci were dysregulated. The dysregulated miRNAs clustered on Chromosomes 14, 17, 19, and X. A small subset of dysregulated genes had many 3' untranslated region (3'UTR) target-sites for dysregulated miRNAs. We provide evidence that certain miRNAs serve as key elements in gene regulatory networks in HIV-infected FC and may be implicated in neurobehavioral disorder. Finally, our data indicates that some genes may serve as hubs of miRNA activity.

Gene expression profiling of histologically normal breast tissue in females with human epidermal growth factor receptor 2‑positive breast cancer.

PubMed

Zubor, Pavol; Hatok, Jozef; Moricova, Petra; Kapustova, Ivana; Kajo, Karol; Mendelova, Andrea; Sivonova, Monika Kmetova; Danko, Jan

2015-02-01

Gene expression profile‑based taxonomy of breast cancer (BC) has been described as a significant breakthrough in comprehending the differences in the origin and behavior of cancer to allow individually tailored therapeutic approaches. In line with this, we hypothesized that the gene expression profile of histologically normal epithelium (HNEpi) could harbor certain genetic abnormalities predisposing breast tissue cells to develop human epidermal growth factor receptor 2 (HER2)‑positive BC. Thus, the aim of the present study was to assess gene expression in normal and BC tissue (BCTis) from patients with BC in order to establish its value as a potential diagnostic marker for cancer development. An array study evaluating a panel of 84 pathway‑ and disease‑specific genes in HER2‑positive BC and tumor‑adjacent HNEpi was performed using quantitative polymerase chain reaction in 12 patients using microdissected samples from frozen tissue. Common prognostic and predictive parameters of BC were assessed by immunohistochemistry and in situ hybridization. In the BCTis and HNEpi samples of 12 HER2‑positive subjects with BC, the expression of 2,016 genes was assessed. A total of 39.3% of genes were deregulated at a minimal two‑fold deregulation rate and 10.7% at a five‑fold deregulation rate in samples of HNEpi or BCTis. Significant differences in gene expression between BCTis and HNEpi samples were revealed for BCL2L2, CD44, CTSD, EGFR, ERBB2, ITGA6, NGFB, RPL27, SCBG2A1 and SCGB1D2 genes (P<0.05), as well as GSN, KIT, KLK5, SERPINB5 and STC2 genes (P<0.01). Insignificant differences (P<0.07) were observed for CCNA1, CLU, DLC1, GABRP and IL6 genes. The ontological gene analyses revealed that the majority of the deregulated genes in the HNEpi samples were part of the functional gene group directly associated with BC origin and prognosis. Functional analysis showed that the most frequent gene deregulations occurred in genes associated with apoptosis and cell cycle regulation in BCTis samples, and with angiogenesis, regulation of the cell cycle and transcriptional activity in HNEpi samples. The molecular profiling of HNEpi breast tissue revealed gene expression abnormalities that may represent potential markers of increased risk for HER2‑positive malignant transformation of breast tissue, and may be able to be employed as predictors of prognosis.

MobilomeFINDER: web-based tools for in silico and experimental discovery of bacterial genomic islands

PubMed Central

Ou, Hong-Yu; He, Xinyi; Harrison, Ewan M.; Kulasekara, Bridget R.; Thani, Ali Bin; Kadioglu, Aras; Lory, Stephen; Hinton, Jay C. D.; Barer, Michael R.; Rajakumar, Kumar

2007-01-01

MobilomeFINDER (http://mml.sjtu.edu.cn/MobilomeFINDER) is an interactive online tool that facilitates bacterial genomic island or ‘mobile genome’ (mobilome) discovery; it integrates the ArrayOme and tRNAcc software packages. ArrayOme utilizes a microarray-derived comparative genomic hybridization input data set to generate ‘inferred contigs’ produced by merging adjacent genes classified as ‘present’. Collectively these ‘fragments’ represent a hypothetical ‘microarray-visualized genome (MVG)’. ArrayOme permits recognition of discordances between physical genome and MVG sizes, thereby enabling identification of strains rich in microarray-elusive novel genes. Individual tRNAcc tools facilitate automated identification of genomic islands by comparative analysis of the contents and contexts of tRNA sites and other integration hotspots in closely related sequenced genomes. Accessory tools facilitate design of hotspot-flanking primers for in silico and/or wet-science-based interrogation of cognate loci in unsequenced strains and analysis of islands for features suggestive of foreign origins; island-specific and genome-contextual features are tabulated and represented in schematic and graphical forms. To date we have used MobilomeFINDER to analyse several Enterobacteriaceae, Pseudomonas aeruginosa and Streptococcus suis genomes. MobilomeFINDER enables high-throughput island identification and characterization through increased exploitation of emerging sequence data and PCR-based profiling of unsequenced test strains; subsequent targeted yeast recombination-based capture permits full-length sequencing and detailed functional studies of novel genomic islands. PMID:17537813

Statistical identification of gene association by CID in application of constructing ER regulatory network

PubMed Central

Liu, Li-Yu D; Chen, Chien-Yu; Chen, Mei-Ju M; Tsai, Ming-Shian; Lee, Cho-Han S; Phang, Tzu L; Chang, Li-Yun; Kuo, Wen-Hung; Hwa, Hsiao-Lin; Lien, Huang-Chun; Jung, Shih-Ming; Lin, Yi-Shing; Chang, King-Jen; Hsieh, Fon-Jou

2009-01-01

Background A variety of high-throughput techniques are now available for constructing comprehensive gene regulatory networks in systems biology. In this study, we report a new statistical approach for facilitating in silico inference of regulatory network structure. The new measure of association, coefficient of intrinsic dependence (CID), is model-free and can be applied to both continuous and categorical distributions. When given two variables X and Y, CID answers whether Y is dependent on X by examining the conditional distribution of Y given X. In this paper, we apply CID to analyze the regulatory relationships between transcription factors (TFs) (X) and their downstream genes (Y) based on clinical data. More specifically, we use estrogen receptor α (ERα) as the variable X, and the analyses are based on 48 clinical breast cancer gene expression arrays (48A). Results The analytical utility of CID was evaluated in comparison with four commonly used statistical methods, Galton-Pearson's correlation coefficient (GPCC), Student's t-test (STT), coefficient of determination (CoD), and mutual information (MI). When being compared to GPCC, CoD, and MI, CID reveals its preferential ability to discover the regulatory association where distribution of the mRNA expression levels on X and Y does not fit linear models. On the other hand, when CID is used to measure the association of a continuous variable (Y) against a discrete variable (X), it shows similar performance as compared to STT, and appears to outperform CoD and MI. In addition, this study established a two-layer transcriptional regulatory network to exemplify the usage of CID, in combination with GPCC, in deciphering gene networks based on gene expression profiles from patient arrays. Conclusion CID is shown to provide useful information for identifying associations between genes and transcription factors of interest in patient arrays. When coupled with the relationships detected by GPCC, the association predicted by CID are applicable to the construction of transcriptional regulatory networks. This study shows how information from different data sources and learning algorithms can be integrated to investigate whether relevant regulatory mechanisms identified in cell models can also be partially re-identified in clinical samples of breast cancers. Availability the implementation of CID in R codes can be freely downloaded from . PMID:19292896

Assessment of DNA methylation profiling and copy number variation as indications of clonal relationship in ipsilateral and contralateral breast cancers to distinguish recurrent breast cancer from a second primary tumour.

PubMed

Huang, Katie T; Mikeska, Thomas; Li, Jason; Takano, Elena A; Millar, Ewan K A; Graham, Peter H; Boyle, Samantha E; Campbell, Ian G; Speed, Terence P; Dobrovic, Alexander; Fox, Stephen B

2015-10-09

Patients with breast cancer have an increased risk of developing subsequent breast cancers. It is important to distinguish whether these tumours are de novo or recurrences of the primary tumour in order to guide the appropriate therapy. Our aim was to investigate the use of DNA methylation profiling and array comparative genomic hybridization (aCGH) to determine whether the second tumour is clonally related to the first tumour. Methylation-sensitive high-resolution melting was used to screen promoter methylation in a panel of 13 genes reported as methylated in breast cancer (RASSF1A, TWIST1, APC, WIF1, MGMT, MAL, CDH13, RARβ, BRCA1, CDH1, CDKN2A, TP73, and GSTP1) in 29 tumour pairs (16 ipsilateral and 13 contralateral). Using the methylation profile of these genes, we employed a Bayesian and an empirical statistical approach to estimate clonal relationship. Copy number alterations were analysed using aCGH on the same set of tumour pairs. There is a higher probability of the second tumour being recurrent in ipsilateral tumours compared with contralateral tumours (38 % versus 8 %; p <0.05) based on the methylation profile. Using previously reported recurrence rates as Bayesian prior probabilities, we classified 69 % of ipsilateral and 15 % of contralateral tumours as recurrent. The inferred clonal relationship results of the tumour pairs were generally concordant between methylation profiling and aCGH. Our results show that DNA methylation profiling as well as aCGH have potential as diagnostic tools in improving the clinical decisions to differentiate recurrences from a second de novo tumour.

Differential gene expression profiles of peripheral blood mononuclear cells in childhood asthma.

PubMed

Kong, Qian; Li, Wen-Jing; Huang, Hua-Rong; Zhong, Ying-Qiang; Fang, Jian-Pei

2015-05-01

Asthma is a common childhood disease with strong genetic components. This study compared whole-genome expression differences between asthmatic young children and healthy controls to identify gene signatures of childhood asthma. Total RNA extracted from peripheral blood mononuclear cells (PBMC) was subjected to microarray analysis. QRT-PCR was performed to verify the microarray results. Classification and functional characterization of differential genes were illustrated by hierarchical clustering and gene ontology analysis. Multiple logistic regression (MLR) analysis, receiver operating characteristic (ROC) curve analysis, and discriminate power were used to scan asthma-specific diagnostic markers. For fold-change>2 and p < 0.05, there were 758 named differential genes. The results of QRT-PCR confirmed successfully the array data. Hierarchical clustering divided 29 highly possible genes into seven categories and the genes in the same cluster were likely to possess similar expression patterns or functions. Gene ontology analysis presented that differential genes primarily enriched in immune response, response to stress or stimulus, and regulation of apoptosis in biological process. MLR and ROC curve analysis revealed that the combination of ADAM33, Smad7, and LIGHT possessed excellent discriminating power. The combination of ADAM33, Smad7, and LIGHT would be a reliable and useful childhood asthma model for prediction and diagnosis.

Gene expression analysis in the ventral prostate of rats exposed to vinclozolin or procymidone.

PubMed

Rosen, Mitchell B; Wilson, Vickie S; Schmid, Judith E; Gray, L Earl

2005-01-01

Vinclozolin and procymidone are antiandrogens that are thought to share a common androgen receptor (AR) mediated mechanism of action. This assessment is based primarily on morphological, AR binding, and in vitro transcriptional activation studies. Studies designed to evaluate the gene expression profiles induced by these compounds have the potential to provide further information to test this hypothesis. We have used targeted gene arrays to examine gene expression in the ventral prostate (VP) of 100-day old Sprague Dawley male rats exposed to either vinclozolin or procymidone. Animals were castrated and administered silastic implants with or without testosterone. A subset of testosterone treated animals was then dosed with 200 mg/kg of either fungicide in corn oil. Four treatment groups were used: castrated (C), testosterone (T), testosterone+vinclozolin (V), and testosterone+procymidone (P). Tissue from the VP was collected from six animals per group (3 animals per block x 2 blocks) at 20 h and at 4 days after the start of treatment. Total RNA was then isolated and gene expression analyzed using Clontech Atlas Rat 1.2 Toxicology arrays. When compared to group T, similar changes in gene expression were observed in groups C, P and V at both the 20 h and 4 day time points. After 20 h of treatment, 20 genes were similarly affected across these three treatment groups. Down-regulated genes included various molecular chaperones, the 11-kDa diazepam binding inhibitor, cyclin D1, and mitochondrial aspartate aminotransferase. Genes such as the androgen receptor, PTEN, and ERK2 were up-regulated. Three of the down-regulated genes, GRP78 (BiP), Dad1, and mitochondrial aspartate aminotransferase have been previously shown to be directly androgen regulated. Fifty-four genes were affected at 20 h, whereas, 311 genes were altered 4 days after the start of treatment. These observations, in part, may reflect regression of the VP at the later time point. These results support the hypothesis that procymidone and vinclozolin share a common mechanism or mode of action, a critical step in a cumulative risk assessment.

Integrating microarray analysis and the soybean genome to understand the soybeans iron deficiency response

PubMed Central

2009-01-01

Background Soybeans grown in the upper Midwestern United States often suffer from iron deficiency chlorosis, which results in yield loss at the end of the season. To better understand the effect of iron availability on soybean yield, we identified genes in two near isogenic lines with changes in expression patterns when plants were grown in iron sufficient and iron deficient conditions. Results Transcriptional profiles of soybean (Glycine max, L. Merr) near isogenic lines Clark (PI548553, iron efficient) and IsoClark (PI547430, iron inefficient) grown under Fe-sufficient and Fe-limited conditions were analyzed and compared using the Affymetrix® GeneChip® Soybean Genome Array. There were 835 candidate genes in the Clark (PI548553) genotype and 200 candidate genes in the IsoClark (PI547430) genotype putatively involved in soybean's iron stress response. Of these candidate genes, fifty-eight genes in the Clark genotype were identified with a genetic location within known iron efficiency QTL and 21 in the IsoClark genotype. The arrays also identified 170 single feature polymorphisms (SFPs) specific to either Clark or IsoClark. A sliding window analysis of the microarray data and the 7X genome assembly coupled with an iterative model of the data showed the candidate genes are clustered in the genome. An analysis of 5' untranslated regions in the promoter of candidate genes identified 11 conserved motifs in 248 differentially expressed genes, all from the Clark genotype, representing 129 clusters identified earlier, confirming the cluster analysis results. Conclusion These analyses have identified the first genes with expression patterns that are affected by iron stress and are located within QTL specific to iron deficiency stress. The genetic location and promoter motif analysis results support the hypothesis that the differentially expressed genes are co-regulated. The combined results of all analyses lead us to postulate iron inefficiency in soybean is a result of a mutation in a transcription factor(s), which controls the expression of genes required in inducing an iron stress response. PMID:19678937

Targeted deletion of miR-132/-212 impairs memory and alters the hippocampal transcriptome.

PubMed

Hansen, Katelin F; Sakamoto, Kensuke; Aten, Sydney; Snider, Kaitlin H; Loeser, Jacob; Hesse, Andrea M; Page, Chloe E; Pelz, Carl; Arthur, J Simon C; Impey, Soren; Obrietan, Karl

2016-02-01

miR-132 and miR-212 are structurally related microRNAs that have been found to exert powerful modulatory effects within the central nervous system (CNS). Notably, these microRNAs are tandomly processed from the same noncoding transcript, and share a common seed sequence: thus it has been difficult to assess the distinct contribution of each microRNA to gene expression within the CNS. Here, we employed a combination of conditional knockout and transgenic mouse models to examine the contribution of the miR-132/-212 gene locus to learning and memory, and then to assess the distinct effects that each microRNA has on hippocampal gene expression. Using a conditional deletion approach, we show that miR-132/-212 double-knockout mice exhibit significant cognitive deficits in spatial memory, recognition memory, and in tests of novel object recognition. Next, we utilized transgenic miR-132 and miR-212 overexpression mouse lines and the miR-132/-212 double-knockout line to explore the distinct effects of these two miRNAs on the transcriptional profile of the hippocampus. Illumina sequencing revealed that miR-132/-212 deletion increased the expression of 1138 genes; Venn analysis showed that 96 of these genes were also downregulated in mice overexpressing miR-132. Of the 58 genes that were decreased in animals overexpressing miR-212, only four of them were also increased in the knockout line. Functional gene ontology analysis of downregulated genes revealed significant enrichment of genes related to synaptic transmission, neuronal proliferation, and morphogenesis, processes known for their roles in learning, and memory formation. These data, coupled with previous studies, firmly establish a role for the miR-132/-212 gene locus as a key regulator of cognitive capacity. Further, although miR-132 and miR-212 share a seed sequence, these data indicate that these miRNAs do not exhibit strongly overlapping mRNA targeting profiles, thus indicating that these two genes may function in a complex, nonredundant manner to shape the transcriptional profile of the CNS. The dysregulation of miR-132/-212 expression could contribute to signaling mechanisms that are involved in an array of cognitive disorders. © 2016 Hansen et al.; Published by Cold Spring Harbor Laboratory Press.

Targeted deletion of miR-132/-212 impairs memory and alters the hippocampal transcriptome

PubMed Central

Hansen, Katelin F.; Sakamoto, Kensuke; Aten, Sydney; Snider, Kaitlin H.; Loeser, Jacob; Hesse, Andrea M.; Page, Chloe E.; Pelz, Carl; Arthur, J. Simon C.; Impey, Soren

2016-01-01

miR-132 and miR-212 are structurally related microRNAs that have been found to exert powerful modulatory effects within the central nervous system (CNS). Notably, these microRNAs are tandomly processed from the same noncoding transcript, and share a common seed sequence: thus it has been difficult to assess the distinct contribution of each microRNA to gene expression within the CNS. Here, we employed a combination of conditional knockout and transgenic mouse models to examine the contribution of the miR-132/-212 gene locus to learning and memory, and then to assess the distinct effects that each microRNA has on hippocampal gene expression. Using a conditional deletion approach, we show that miR-132/-212 double-knockout mice exhibit significant cognitive deficits in spatial memory, recognition memory, and in tests of novel object recognition. Next, we utilized transgenic miR-132 and miR-212 overexpression mouse lines and the miR-132/-212 double-knockout line to explore the distinct effects of these two miRNAs on the transcriptional profile of the hippocampus. Illumina sequencing revealed that miR-132/-212 deletion increased the expression of 1138 genes; Venn analysis showed that 96 of these genes were also downregulated in mice overexpressing miR-132. Of the 58 genes that were decreased in animals overexpressing miR-212, only four of them were also increased in the knockout line. Functional gene ontology analysis of downregulated genes revealed significant enrichment of genes related to synaptic transmission, neuronal proliferation, and morphogenesis, processes known for their roles in learning, and memory formation. These data, coupled with previous studies, firmly establish a role for the miR-132/-212 gene locus as a key regulator of cognitive capacity. Further, although miR-132 and miR-212 share a seed sequence, these data indicate that these miRNAs do not exhibit strongly overlapping mRNA targeting profiles, thus indicating that these two genes may function in a complex, nonredundant manner to shape the transcriptional profile of the CNS. The dysregulation of miR-132/-212 expression could contribute to signaling mechanisms that are involved in an array of cognitive disorders. PMID:26773099

Gene expression profiling and functional characterization of macrophages in response to circulatory microparticles produced during Trypanosoma cruzi infection and Chagas disease

PubMed Central

Chowdhury, Imran; Koo, Sue-jie; Gupta, Shivali; Liang, Lisa Yi; Bahar, Bojlul; Silla, Laura; Burgos, Julio Nuñez; Barrientos, Natalia; Zago, Maria Paola; Garg, Nisha Jain

2016-01-01

BACKGROUND Chronic inflammation and oxidative stress are hallmarks of chagasic cardiomyopathy (CCM). In this study, we determined if microparticles (MPs) generated during Trypanosoma cruzi (Tc) infection carry the host’s signature of inflammatory/oxidative state and provide information regarding the progression of clinical disease. METHDOS The MPs were harvested from supernatants of human PBMCs in vitro incubated with T. cruzi (control: LPS-treated), plasma of seropositive humans with clinically asymptomatic (CA) or symptomatic (CS) disease state (normal/healthy (NH) controls) and plasma of mice immunized with a protective vaccine before challenge infection (control: unvaccinated/infected). Macrophages (mφs) were incubated with MPs, and we probed the gene expression profile using the inflammatory signaling cascade and cytokine/chemokine arrays, phenotypic markers of macrophage activation by flow cytometry, cytokine profile by an ELISA and Bioplex assay, and oxidative/nitrosative stress and mitotoxicity by colorimetric and fluorometric assays. RESULTS Tc- and LPS-induced MPs stimulated proliferation, inflammatory gene expression profile and •NO release in human THP-1 mφs. LPS-MPs were more immunostimulatory than Tc-MPs. Endothelial cells, T lymphocytes and mφs were the major source of MPs shed in plasma of chagasic humans and experimentally infected mice. The CS-MPs and CA-MPs (vs. NH-MPs) elicited >2-fold increase in •NO and mitochondrial oxidative stress in THP-1 mφs; however, CS-MPs (vs. CA-MPs) elicited a more pronounced and disease-state-specific inflammatory gene expression profile (IKBKB, NR3C1, and TIRAP vs. CCR4, EGR2 and CCL3), cytokine release (IL2+IFNγ>GCSF), and surface markers of mφ activation (CD14 and CD16). The circulatory MPs of non-vaccinated/infected mice induced 7.5-fold and 40% increase in •NO and IFNγ production, respectively, while these responses were abolished when RAW264.7 mφs were incubated with circulatory MPs of vaccinated/infected mice. CONCLUSION Circulating MPs reflect in vivo levels of oxidative, nitrosative, and inflammatory state and have potential utility in evaluating disease severity and efficacy of vaccines and drug therapies against CCM. PMID:27902980

Adipose Gene Expression Profile Changes With Lung Allograft Reperfusion.

PubMed

Diamond, Joshua M; Arcasoy, Selim; McDonnough, Jamiela A; Sonett, Joshua R; Bacchetta, Matthew; D'Ovidio, Frank; Cantu, Edward; Bermudez, Christian A; McBurnie, Amika; Rushefski, Melanie; Kalman, Laurel H; Oyster, Michelle; D'Errico, Carly; Suzuki, Yoshikazu; Giles, Jon T; Ferrante, Anthony; Lippel, Matthew; Singh, Gopal; Lederer, David J; Christie, Jason D

2017-01-01

Obesity is a risk factor for primary graft dysfunction (PGD), a form of lung injury resulting from ischemia-reperfusion after lung transplantation, but the impact of ischemia-reperfusion on adipose tissue is unknown. We evaluated differential gene expression in thoracic visceral adipose tissue (VAT) before and after lung reperfusion. Total RNA was isolated from thoracic VAT sampled from six subjects enrolled in the Lung Transplant Body Composition study before and after allograft reperfusion and quantified using the Human Gene 2.0 ST array. Kyoto Encyclopedia of Genes and Genomes pathway analysis revealed enrichment for genes involved in complement and coagulation cascades and Jak-STAT signaling pathways. Overall, 72 genes were upregulated and 56 genes were downregulated in the postreperfusion time compared with baseline. Long pentraxin-3, a gene and plasma protein previously associated with PGD, was the most upregulated gene (19.5-fold increase, p = 0.04). Fibronectin leucine-rich transmembrane protein-3, a gene associated with cell adhesion and receptor signaling, was the most downregulated gene (4.3-fold decrease, p = 0.04). Ischemia-reperfusion has a demonstrable impact on gene expression in visceral adipose tissue in our pilot study of nonobese, non-PGD lung transplant recipients. Future evaluation will focus on differential adipose tissue gene expression and the development of PGD after transplant. © Copyright 2016 The American Society of Transplantation and the American Society of Transplant Surgeons.

Search for sarcoidosis candidate genes by integration of data from genomic, transcriptomic and proteomic studies.

PubMed

Maver, Ales; Medica, Igor; Peterlin, Borut

2009-12-01

The search for gene candidates in multifactorial diseases such as sarcoidosis can be based on the integration of linkage association data, gene expression data, and protein profile data from genomic, transcriptomic and proteomic studies, respectively. In this study we performed a literature-based search for studies reporting such data, followed by integration of collected information. Different databases were examined--Medline, HugGE Navigator, ArrayExpress and Gene Expression Omnibus (GEO). Candidate genes were defined as genes which were reported in at least 2 different types of omics studies. Genes previously investigated in sarcoidosis were excluded from further analyses. We identified 177 genes associated with sarcoidosis as potential new candidate genes. Subsequently, 9 gene candidates identified to overlap in 2 different types of studies (genomic, transcriptomic and/or proteomic) were consistently reported in at least 3 studies: SERPINB1, FABP4, S100A8, HBEGF, IL7R, LRIG1, PTPN23, DPM2 and NUP214. These genes are involved in regulation of immune response, cellular proliferation, apoptosis, inhibition of protease activity, lipid metabolism. Exact biological functions of HBEGF, LRIG1, PTPN23, DPM2 and NUP214 remain to be completely elucidated. We propose 9 candidate genes: SERPINB1, FABP4, S100A8, HBEGF, IL7R, LRIG1, PTPN23, DPM2 and NUP214, as genes with high potential for association with sarcoidosis.

Tracking Spatial and Temporal Changes in Microbial Metabolic Potential and Gene Expression Patterns Across Geochemical Gradients at Axial Seamount

NASA Astrophysics Data System (ADS)

Fortunato, C. S.; Butterfield, D. A.; Larson, B.; Algar, C. K.; Huber, J. A.

2016-12-01

Microbial communities living both near and within the subseafloor are important players in the biogeochemical cycling of the deep ocean. To better understand the metabolic and gene expression patterns of these understudied communities, we collected low-temperature diffuse fluids for metagenomic, metatranscriptomic, and geochemical analyses from Axial Seamount, an active submarine volcano off the coast of Oregon, USA in 2013-2015. In April of 2015 Axial Seamount erupted along its north rift, five months before the 2015 samples were collected. This study thus provides both spatial and temporal analysis of subseafloor microbial communities pre and post eruption. The time series for this study focused on three vents at the south end of the caldera: Anemone, Marker 33, and Marker 113. Chemistry data shows that at each vent there are different geochemical conditions and thus a potentially different microbial metabolic profile. Anemone has the most oxidizing conditions and the highest abundance and expression of sulfur oxidation genes, attributed to both SUP05 and Epsilonproteobacteria. The most reducing conditions were observed at Marker 113, the site with the lowest oxygen concentration and where methanogenesis was the dominant metabolism, with 18.5% of all annotated transcripts attributed to methanogenesis. Although individual vents were metabolically distinct, there was very little variation in the overall taxonomic and metabolic profiles of each vent across years, even after the 2015 eruption. A diffuse fluid sample taken from the North Rift Zone post eruption showed similar community taxonomy to both Anemone and Marker 33; analyses of the metabolic potential and gene expression at this site is ongoing and will act as a comparison between the communities of the time series vents and those that were closer to the eruption site. Together, these chemical and `omic datasets reveal a dynamic microbial community at each vent, taxonomically diverse and involved in a wide array of biogeochemical transformations. Results are being used to model the functional dynamics and fluxes of vent communities to more closely link microbiological productivity at hydrothermal systems to deep-sea biogeochemical processes and will be also used to inform future projects using instrumentation on the cabled array at Axial Seamount.

13. CONTROL ROOM OF GENE PUMPING STATION. CONTROL CUBICLES ARRAYED ...

Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

13. CONTROL ROOM OF GENE PUMPING STATION. CONTROL CUBICLES ARRAYED BEHIND MANAGER'S ART DECO-STYLE CONTROL DESK, WITH CONTROL CUBICLE 1 AT FAR RIGHT AND CONTROL CUBICLE 9 AT FAR LEFT. - Gene Pump Plant, South of Gene Wash Reservoir, 2 miles west of Whitsett Pump Plant, Parker Dam, San Bernardino County, CA

THE USE OF GENE ARRAYS TO MEASURE CHANGES IN GENE EXPRESSION PATTERNS IN FISH EXPOSED TO COMPOUNDS THAT MIMIC ESTROGEN

EPA Science Inventory

We have developed estrogen-sensitive gene arrays to measure changes in gene expression in sheepshead minnows and largemouth bass exposed to anthropogenic chemicals that mimic estrogen. The in vivo exposures, which realize the full physiological response in fish, result in changes...

DEVELOPMENT OF A 950-GENE DNA ARRAY FOR EXAMINING GENE EXPRESSION PATTERNS IN MOUSE TESTIS

EPA Science Inventory

Development of a 950-gene DNA array for examining gene expression patterns in mouse testis.

Rockett JC, Christopher Luft J, Brian Garges J, Krawetz SA, Hughes MR, Hee Kirn K, Oudes AJ, Dix DJ.

Reproductive Toxicology Division, National Health and Environmental Effec...

Manipulation of a two-photon state in a χ(2)-modulated nonlinear waveguide array

NASA Astrophysics Data System (ADS)

Yang, Y.; Xu, P.; Lu, L. L.; Zhu, S. N.

2014-10-01

We propose to engineer the quantum state in a high-dimensional Hilbert space by taking advantage of a χ(2)-modulated nonlinear waveguide array. By varying the pump condition and the waveguide array length, the momentum correlation between the signal and idler photons can be manipulated, exhibiting bunching, antibunching, and the evolution between these two states, which are characterized by the Schmidt number. We find the Schmidt number is dependent on a structure parameter, namely the ratio of the array length and the number of channels pumped. By designing the linear profile waveguide array, the degree of spatial entanglement shows a periodic relationship with the slope of linear profile, during which a high degree of position-bunching state is suggested. The two-photon self-focusing effect is disclosed when the χ(2) modulation in the waveguide array contains a parabolic profile, which can be designed for efficient coupling between a waveguide array and fibers. These results shed light on a feasible way to achieve desirable quantum state on a single waveguide chip by a compact engineering of χ(2) and also suggest a degree of freedom for quantum walk and other related applications.

A phase 1 trial of ABT-510 concurrent with standard chemoradiation for patients with newly diagnosed glioblastoma.

PubMed

Nabors, Louis B; Fiveash, John B; Markert, James M; Kekan, Manasi S; Gillespie, George Y; Huang, Zhi; Johnson, Martin J; Meleth, Sreelatha; Kuo, Huichien; Gladson, Candece L; Fathallah-Shaykh, Hassan M

2010-03-01

To determine the maximum tolerated dose of ABT-510, a thrombospondin-1 mimetic drug with antiangiogenic properties, when used concurrently with temozolomide and radiotherapy in patients with newly diagnosed glioblastoma. Phase 1 dose-escalation clinical trial. Comprehensive Cancer Center, University of Alabama at Birmingham. Patients A total of 23 patients with newly diagnosed, histologically verified glioblastoma enrolled between April 2005 and January 2007. Four cohorts of 3 patients each received subcutaneous ABT-510 injection at doses of 20, 50, 100, or 200 mg/d. The maximum cohort was expanded to 14 patients to obtain additional safety and gene expression data. The treatment plan included 10 weeks of induction phase (temozolomide and radiotherapy with ABT-510 for 6 weeks plus ABT-510 monotherapy for 4 weeks) followed by a maintenance phase of ABT-510 and monthly temozolomide. Patients were monitored with brain magnetic resonance imaging and laboratory testing for dose-limiting toxicities, defined as grades 3 or 4 nonhematological toxicities and grade 4 hematological toxicities. Therapy was discontinued if 14 maintenance cycles were completed, disease progression occurred, or if the patient requested withdrawal. Disease progression, survival statistics, and gene expression arrays were analyzed. There were no grade 3 or 4 dose-limiting toxicity events that appeared related to ABT-510 for the dose range of 20 to 200 mg/d. A maximum tolerated dose was not defined. Most adverse events were mild, and injection-site reactions. The median time to tumor progression was 45.9 weeks, and the median overall survival time was 64.4 weeks. Gene expression analysis using TaqMan low-density arrays identified angiogenic genes that were differentially expressed in the brains of controls compared with patients with newly diagnosed glioblastoma, and identified FGF-1 and TIE-1 as being downregulated in patients who had better clinical outcomes. ABT-510, at subcutaneous doses up to 200 mg/d, is tolerated well with concurrent temozolomide and radiotherapy in patients with newly diagnosed glioblastoma, and low-density arrays provide a useful method of exploring gene expression profiles.

Detecting novel genes with sparse arrays

PubMed Central

Haiminen, Niina; Smit, Bart; Rautio, Jari; Vitikainen, Marika; Wiebe, Marilyn; Martinez, Diego; Chee, Christine; Kunkel, Joe; Sanchez, Charles; Nelson, Mary Anne; Pakula, Tiina; Saloheimo, Markku; Penttilä, Merja; Kivioja, Teemu

2014-01-01

Species-specific genes play an important role in defining the phenotype of an organism. However, current gene prediction methods can only efficiently find genes that share features such as sequence similarity or general sequence characteristics with previously known genes. Novel sequencing methods and tiling arrays can be used to find genes without prior information and they have demonstrated that novel genes can still be found from extensively studied model organisms. Unfortunately, these methods are expensive and thus are not easily applicable, e.g., to finding genes that are expressed only in very specific conditions. We demonstrate a method for finding novel genes with sparse arrays, applying it on the 33.9 Mb genome of the filamentous fungus Trichoderma reesei. Our computational method does not require normalisations between arrays and it takes into account the multiple-testing problem typical for analysis of microarray data. In contrast to tiling arrays, that use overlapping probes, only one 25mer microarray oligonucleotide probe was used for every 100 b. Thus, only relatively little space on a microarray slide was required to cover the intergenic regions of a genome. The analysis was done as a by-product of a conventional microarray experiment with no additional costs. We found at least 23 good candidates for novel transcripts that could code for proteins and all of which were expressed at high levels. Candidate genes were found to neighbour ire1 and cre1 and many other regulatory genes. Our simple, low-cost method can easily be applied to finding novel species-specific genes without prior knowledge of their sequence properties. PMID:20691772

missMethyl: an R package for analyzing data from Illumina's HumanMethylation450 platform.

PubMed

Phipson, Belinda; Maksimovic, Jovana; Oshlack, Alicia

2016-01-15

DNA methylation is one of the most commonly studied epigenetic modifications due to its role in both disease and development. The Illumina HumanMethylation450 BeadChip is a cost-effective way to profile >450 000 CpGs across the human genome, making it a popular platform for profiling DNA methylation. Here we introduce missMethyl, an R package with a suite of tools for performing normalization, removal of unwanted variation in differential methylation analysis, differential variability testing and gene set analysis for the 450K array. missMethyl is an R package available from the Bioconductor project at www.bioconductor.org. alicia.oshlack@mcri.edu.au Supplementary data are available at Bioinformatics online. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Primary Sjögren's syndrome is characterized by distinct phenotypic and transcriptional profiles of IgD+ unswitched memory B cells.

PubMed

Roberts, Mustimbo E P; Kaminski, Denise; Jenks, Scott A; Maguire, Craig; Ching, Kathryn; Burbelo, Peter D; Iadarola, Michael J; Rosenberg, Alexander; Coca, Andreea; Anolik, Jennifer; Sanz, Iñaki

2014-09-01

The significance of distinct B cell abnormalities in primary Sjögren's syndrome (SS) remains to be established. We undertook this study to analyze the phenotype and messenger RNA (mRNA) transcript profiles of B cell subsets in patients with primary SS and to compare them with those in sicca syndrome patients and healthy controls. CD19+ B cells from 26 patients with primary SS, 27 sicca syndrome patients, and 22 healthy controls were analyzed by flow cytometry. Gene expression profiles of purified B cell subsets (from 3-5 subjects per group per test) were analyzed using Affymetrix gene arrays. Patients with primary SS had lower frequencies of CD27+IgD- switched memory B cells and CD27+IgD+ unswitched memory B cells compared with healthy controls. Unswitched memory B cell frequencies were also lower in sicca syndrome patients and correlated inversely with serologic hyperactivity in both disease states. Further, unswitched memory B cells in primary SS had lower expression of CD1c and CD21. Gene expression analysis of CD27+ memory B cells separated patients with primary SS from healthy controls and identified a subgroup of sicca syndrome patients with a primary SS-like transcript profile. Moreover, unswitched memory B cell gene expression analysis identified 187 genes differentially expressed between patients with primary SS and healthy controls. A decrease in unswitched memory B cells with serologic hyperactivity is characteristic of both established primary SS and a subgroup of sicca syndrome, which suggests the value of these B cells both as biomarkers of future disease progression and for understanding disease pathogenesis. Overall, the mRNA transcript analysis of unswitched memory B cells suggests that their activation in primary SS takes place through innate immune pathways in the context of attenuated antigen-mediated adaptive signaling. Thus, our findings provide important insight into the mechanisms and potential consequences of decreased unswitched memory B cells in primary SS. Copyright © 2014 by the American College of Rheumatology.

High-Resolution Array CGH Profiling Identifies Na/K Transporting ATPase Interacting 2 (NKAIN2) as a Predisposing Candidate Gene in Neuroblastoma

PubMed Central

Romania, Paolo; Castellano, Aurora; Surace, Cecilia; Citti, Arianna; De Ioris, Maria Antonietta; Sirleto, Pietro; De Mariano, Marilena; Longo, Luca; Boldrini, Renata; Angioni, Adriano; Locatelli, Franco; Fruci, Doriana

2013-01-01

Neuroblastoma (NB), the most common solid cancer in early childhood, usually occurs sporadically but also its familial occurance is known in 1-2% of NB patients. Germline mutations in the ALK and PHOX2B genes have been found in a subset of familial NBs. However, because some individuals harbouring mutations in these genes do not develop this tumor, additional genetic alterations appear to be required for NB pathogenesis. Herein, we studied an Italian family with three NB patients, two siblings and a first cousin, carrying an ALK germline-activating mutation R1192P, that was inherited from their unaffected mothers and with no mutations in the PHOX2B gene. A comparison between somatic and germline DNA copy number changes in the two affected siblings by a high resolution array-based Comparative Genomic Hybridization (CGH) analysis revealed a germline gain at NKAIN2 (Na/K transporting ATPase interacting 2) locus in one of the sibling, that was inherited from the parent who does not carry the ALK mutation. Surprisingly, NKAIN2 was expressed at high levels also in the affected sibling that lacks the genomic gain at this locus, clearly suggesting the existance of other regulatory mechanisms. High levels of NKAIN2 were detected in the MYCN-amplified NB cell lines and in the most aggressive NB lesions as well as in the peripheral blood of a large cohort of NB patients. Consistent with a role of NKAIN2 in NB development, NKAIN2 was down-regulated during all-trans retinoic acid differentiation in two NB cell lines. Taken together, these data indicate a potential role of NKAIN2 gene in NB growth and differentiation. PMID:24205241

High-resolution array CGH profiling identifies Na/K transporting ATPase interacting 2 (NKAIN2) as a predisposing candidate gene in neuroblastoma.

PubMed

Romania, Paolo; Castellano, Aurora; Surace, Cecilia; Citti, Arianna; De Ioris, Maria Antonietta; Sirleto, Pietro; De Mariano, Marilena; Longo, Luca; Boldrini, Renata; Angioni, Adriano; Locatelli, Franco; Fruci, Doriana

2013-01-01

Neuroblastoma (NB), the most common solid cancer in early childhood, usually occurs sporadically but also its familial occurance is known in 1-2% of NB patients. Germline mutations in the ALK and PHOX2B genes have been found in a subset of familial NBs. However, because some individuals harbouring mutations in these genes do not develop this tumor, additional genetic alterations appear to be required for NB pathogenesis. Herein, we studied an Italian family with three NB patients, two siblings and a first cousin, carrying an ALK germline-activating mutation R1192P, that was inherited from their unaffected mothers and with no mutations in the PHOX2B gene. A comparison between somatic and germline DNA copy number changes in the two affected siblings by a high resolution array-based Comparative Genomic Hybridization (CGH) analysis revealed a germline gain at NKAIN2 (Na/K transporting ATPase interacting 2) locus in one of the sibling, that was inherited from the parent who does not carry the ALK mutation. Surprisingly, NKAIN2 was expressed at high levels also in the affected sibling that lacks the genomic gain at this locus, clearly suggesting the existance of other regulatory mechanisms. High levels of NKAIN2 were detected in the MYCN-amplified NB cell lines and in the most aggressive NB lesions as well as in the peripheral blood of a large cohort of NB patients. Consistent with a role of NKAIN2 in NB development, NKAIN2 was down-regulated during all-trans retinoic acid differentiation in two NB cell lines. Taken together, these data indicate a potential role of NKAIN2 gene in NB growth and differentiation.

Transcriptome Profiling of Human FoxP3+ Regulatory T Cells

PubMed Central

Bhairavabhotla, Ravikiran; Kim, Yong C.; Glass, Deborah D.; Escobar, Thelma M.; Patel, Mira C.; Zahr, Rami; Nguyen, Cuong K.; Kilaru, Gokhul K.; Muljo, Stefan A.; Shevach, Ethan M.

2015-01-01

The major goal of this study was to perform an in depth characterization of the “gene signature” of human FoxP3+ T regulatory cells (Tregs). Highly purified Tregs and T conventional cells (Tconvs) from multiple healthy donors (HD), either freshly explanted or activated in vitro, were analyzed via RNA sequencing (RNA-seq) and gene expression changes validated using the nCounter system. Additionally, we analyzed microRNA (miRNA) expression using TaqMan low-density arrays. Our results confirm previous studies demonstrating selective gene expression of FoxP3, IKZF2, and CTLA4 in Tregs. Notably, a number of yet uncharacterized genes (RTKN2, LAYN, UTS2, CSF2RB, TRIB1, F5, CECAM4, CD70, ENC1 and NKG7) were identified and validated as being differentially expressed in human Tregs. We further characterize the functional roles of RTKN2 and LAYN by analyzing their roles in vitro human Treg suppression assays by knocking them down in Tregs and overexpressing them in Tconvs. In order to facilitate a better understanding of the human Treg gene expression signature, we have generated from our results a hypothetical interactome of genes and miRNAs in Tregs and Tconvs, PMID:26686412

Transcriptomic data analysis and differential gene expression of antioxidant pathways in king penguin juveniles (Aptenodytes patagonicus) before and after acclimatization to marine life.

PubMed

Rey, Benjamin; Dégletagne, Cyril; Duchamp, Claude

2016-12-01

In this article, we present differentially expressed gene profiles in the pectoralis muscle of wild juvenile king penguins that were either naturally acclimated to cold marine environment or experimentally immersed in cold water as compared with penguin juveniles that never experienced cold water immersion. Transcriptomic data were obtained by hybridizing penguins total cDNA on Affymetrix GeneChip Chicken Genome arrays and analyzed using maxRS algorithm , " Transcriptome analysis in non-model species: a new method for the analysis of heterologous hybridization on microarrays " (Dégletagne et al., 2010) [1] . We focused on genes involved in multiple antioxidant pathways. For better clarity, these differentially expressed genes were clustered into six functional groups according to their role in controlling redox homeostasis. The data are related to a comprehensive research study on the ontogeny of antioxidant functions in king penguins, "Hormetic response triggers multifaceted anti-oxidant strategies in immature king penguins (Aptenodytes patagonicus)" (Rey et al., 2016) [2] . The raw microarray dataset supporting the present analyses has been deposited at the Gene Expression Omnibus (GEO) repository under accessions GEO: GSE17725 and GEO: GSE82344.

Zoom‐in comparative genomic hybridisation arrays for the characterisation of variable breakpoint contiguous gene syndromes

PubMed Central

Johnston, Jennifer J; Walker, Robert L; Davis, Sean; Facio, Flavia; Turner, Joyce T; Bick, David P; Daentl, Donna L; Ellison, Jay W; Meltzer, Paul S; Biesecker, Leslie G

2007-01-01

Contiguous gene syndromes cause disorders via haploinsufficiency for adjacent genes. Some contiguous gene syndromes (CGS) have stereotypical breakpoints, but others have variable breakpoints. In CGS that have variable breakpoints, the extent of the deletions may be correlated with severity. The Greig cephalopolysyndactyly contiguous gene syndrome (GCPS‐CGS) is a multiple malformation syndrome caused by haploinsufficiency of GLI3 and adjacent genes. In addition, non‐CGS GCPS can be caused by deletions or duplications in GLI3. Although fluorescence in situ hybridisation (FISH) can identify large deletion mutations in patients with GCPS or GCPS‐CGS, it is not practical for identification of small intragenic deletions or insertions, and it is difficult to accurately characterise the extent of the large deletions using this technique. We have designed a custom comparative genomic hybridisation (CGH) array that allows identification of deletions and duplications at kilobase resolution in the vicinity of GLI3. The array averages one probe every 730 bp for a total of about 14 000 probes over 10 Mb. We have analysed 16 individuals with known or suspected deletions or duplications. In 15 of 16 individuals (14 deletions and 1 duplication), the array confirmed the prior results. In the remaining patient, the normal CGH array result was correct, and the prior assessment was a false positive quantitative polymerase chain reaction result. We conclude that high‐density CGH array analysis is more sensitive than FISH analysis for detecting deletions and provides clinically useful results on the extent of the deletion. We suggest that high‐density CGH array analysis should replace FISH analysis for assessment of deletions and duplications in patients with contiguous gene syndromes caused by variable deletions. PMID:17098889

Surface enhanced Raman gene probe and methods thereof

DOEpatents

Vo-Dinh, T.

1998-09-29

The subject invention disclosed herein is a new gene probe biosensor and methods based on surface enhanced Raman scattering (SERS) label detection. The SER gene probe biosensor comprises a support means, a SER gene probe having at least one oligonucleotide strand labeled with at least one SERS label, and a SERS active substrate disposed on the support means and having at least one of the SER gene probes adsorbed thereon. Biotargets such as bacterial and viral DNA, RNA and PNA are detected using a SER gene probe via hybridization to oligonucleotide strands complementary to the SER gene probe. The support means supporting the SERS active substrate includes a fiberoptic probe, an array of fiberoptic probes for performance of multiple assays and a waveguide microsensor array with charge-coupled devices or photodiode arrays. 18 figs.

Surface enhanced Raman gene probe and methods thereof

DOEpatents

Vo-Dinh, Tuan

1998-01-01

The subject invention disclosed herein is a new gene probe biosensor and methods thereof based on surface enhanced Raman scattering (SERS) label detection. The SER gene probe biosensor comprises a support means, a SER gene probe having at least one oligonucleotide strand labeled with at least one SERS label, and a SERS active substrate disposed on the support means and having at least one of the SER gene probes adsorbed thereon. Biotargets such as bacterial and viral DNA, RNA and PNA are detected using a SER gene probe via hybridization to oligonucleotide strands complementary to the SER gene probe. The support means supporting the SERS active substrate includes a fiberoptic probe, an array of fiberoptic probes for performance of multiple assays and a waveguide microsensor array with charge-coupled devices or photodiode arrays.

Surface enhanced Raman gene probe and methods thereof

DOEpatents

Vo-Dinh, T.

1998-02-24

The subject invention disclosed is a new gene probe biosensor and methods based on surface enhanced Raman scattering (SERS) label detection. The SER gene probe biosensor comprises a support means, a SER gene probe having at least one oligonucleotide strand labeled with at least one SERS label, and a SERS active substrate disposed on the support means and having at least one of the SER gene probes adsorbed thereon. Biotargets such as bacterial and viral DNA, RNA and PNA are detected using a SER gene probe via hybridization to oligonucleotide strands complementary to the SER gene probe. The support means includes a fiberoptic probe, an array of fiberoptic probes for performance of multiple assays and a waveguide microsensor array with charge-coupled devices or photodiode arrays. 18 figs.

Surface enhanced Raman gene probe and methods thereof

DOEpatents

Vo-Dinh, T.

1998-07-21

The subject invention disclosed is a new gene probe biosensor and methods based on surface enhanced Raman scattering (SERS) label detection. The SER gene probe biosensor comprises a support means, a SER gene probe having at least one oligonucleotide strand labeled with at least one SERS label, and a SERS active substrate disposed on the support means and having at least one of the SER gene probes adsorbed. Biotargets such as bacterial and viral DNA, RNA and PNA are detected using a SER gene probe via hybridization to oligonucleotide strands complementary to the SER gene probe. The support means supporting the SERS active substrate includes a fiberoptic probe, an array of fiberoptic probes for performance of multiple assays and a waveguide microsensor array with charge-coupled devices or photodiode arrays. 18 figs.

Incidence of antimicrobial-resistance genes and integrons in antibiotic-resistant bacteria isolated from eels and aquaculture ponds.

PubMed

Lin, Mao; Wu, Xiaomei; Yan, Qingpi; Ma, Ying; Huang, Lixing; Qin, Yingxue; Xu, Xiaojin

2016-07-07

The overuse of antimicrobials in aquaculture has promoted the selection of antimicrobial-resistant bacteria. Here we investigated the abundance of antimicrobial-resistance genes and integrons in 108 strains of antibiotic-resistant bacteria isolated from eels and aquaculture ponds in China. Conventional PCR was implemented to examine common antibiotic-resistance genes, integrons, and their gene cassette arrays. The results showed that the antibiotic-resistance genes blaTEM, tetC, sulI, aadA, floR, and qnrB were detected at high percentages, as were a number of other resistance genes. Class I integrons were present in 79.63% of the strains, and 10 out of 108 isolates carried class II integrons. Class III integrons were not detected. Three strains carried both class I and class II integrons, and 73.26% of the class I integron-positive isolates contained the qacEΔ1/sul1 gene. Fourteen types of integron cassette arrays were found among class I integron-positive isolates. A new array, dfrB4-catB3-blaOXA-10-aadA1, was discovered in this study. The gene cassette array dfrA12-orfF-aadA2 was the most widely distributed. In summary, 23 different gene cassettes encoding resistance to 8 classes of antibiotics were identified in the class I integrons, and the main cassettes contained genes encoding resistance to aminoglycosides (aad) and trimethoprim (dfr). All class II integron-positive strains had only a single gene cassette array, viz. dfrA1-catB2-sat2-aadA1. High levels of antimicrobial-resistance genes and integrons in eels and auqauculture ponds suggest that the overuse of antimicrobials should be strictly controlled and that the levels of bacterial antimicrobial-resistance genes in aquaculture should be monitored.

Entropy-based gene ranking without selection bias for the predictive classification of microarray data.

PubMed

Furlanello, Cesare; Serafini, Maria; Merler, Stefano; Jurman, Giuseppe

2003-11-06

We describe the E-RFE method for gene ranking, which is useful for the identification of markers in the predictive classification of array data. The method supports a practical modeling scheme designed to avoid the construction of classification rules based on the selection of too small gene subsets (an effect known as the selection bias, in which the estimated predictive errors are too optimistic due to testing on samples already considered in the feature selection process). With E-RFE, we speed up the recursive feature elimination (RFE) with SVM classifiers by eliminating chunks of uninteresting genes using an entropy measure of the SVM weights distribution. An optimal subset of genes is selected according to a two-strata model evaluation procedure: modeling is replicated by an external stratified-partition resampling scheme, and, within each run, an internal K-fold cross-validation is used for E-RFE ranking. Also, the optimal number of genes can be estimated according to the saturation of Zipf's law profiles. Without a decrease of classification accuracy, E-RFE allows a speed-up factor of 100 with respect to standard RFE, while improving on alternative parametric RFE reduction strategies. Thus, a process for gene selection and error estimation is made practical, ensuring control of the selection bias, and providing additional diagnostic indicators of gene importance.

Branches of NF-κb signaling pathway regulate hepatocyte proliferation in rat liver regeneration.

PubMed

Chang, C F; Zhao, W M; Mei, J X; Zhou, Y; Pan, C Y; Xu, T T; Xu, C S

2015-07-13

Previous studies have demonstrated that the nuclear factor κB (NF-κB) pathway is involved in promoting cell proliferation. To further explore the regulatory branches and their sequence in the NF-κB pathway in the promotion of hepatocyte proliferation at the transcriptional level during rat liver regeneration, Rat Genome 230 2.0 array was used to detect the expression changes of the isolated hepatocytes. We found that many genes involved in the NF-κB pathway (including 73 known genes and 19 homologous genes) and cell proliferation (including 484 genes and 104 homologous genes) were associated with liver regeneration. Expression profile function (Ep) was used to analyze the biological processes. It was revealed that the NF-κB pathway promoted hepatocyte proliferation through three branches. Several methods of integrated statistics were applied to extract and screen key genes in liver regeneration, and it indicated that eight genes may play a vital role in rat liver regeneration. To confirm the above predicted results, Ccnd1, Jun and Myc were analyzed using qRT-PCR, and the results were generally consistent with that of microarray data. It is concluded that 3 branches and 8 key genes involved in the NF-κB pathway regulate hepatocyte proliferation during rat liver regeneration.

MicroRNA signature of the human developing pancreas.

PubMed

Rosero, Samuel; Bravo-Egana, Valia; Jiang, Zhijie; Khuri, Sawsan; Tsinoremas, Nicholas; Klein, Dagmar; Sabates, Eduardo; Correa-Medina, Mayrin; Ricordi, Camillo; Domínguez-Bendala, Juan; Diez, Juan; Pastori, Ricardo L

2010-09-22

MicroRNAs are non-coding RNAs that regulate gene expression including differentiation and development by either inhibiting translation or inducing target degradation. The aim of this study is to determine the microRNA expression signature during human pancreatic development and to identify potential microRNA gene targets calculating correlations between the signature microRNAs and their corresponding mRNA targets, predicted by bioinformatics, in genome-wide RNA microarray study. The microRNA signature of human fetal pancreatic samples 10-22 weeks of gestational age (wga), was obtained by PCR-based high throughput screening with Taqman Low Density Arrays. This method led to identification of 212 microRNAs. The microRNAs were classified in 3 groups: Group number I contains 4 microRNAs with the increasing profile; II, 35 microRNAs with decreasing profile and III with 173 microRNAs, which remain unchanged. We calculated Pearson correlations between the expression profile of microRNAs and target mRNAs, predicted by TargetScan 5.1 and miRBase algorithms, using genome-wide mRNA expression data. Group I correlated with the decreasing expression of 142 target mRNAs and Group II with the increasing expression of 876 target mRNAs. Most microRNAs correlate with multiple targets, just as mRNAs are targeted by multiple microRNAs. Among the identified targets are the genes and transcription factors known to play an essential role in pancreatic development. We have determined specific groups of microRNAs in human fetal pancreas that change the degree of their expression throughout the development. A negative correlative analysis suggests an intertwined network of microRNAs and mRNAs collaborating with each other. This study provides information leading to potential two-way level of combinatorial control regulating gene expression through microRNAs targeting multiple mRNAs and, conversely, target mRNAs regulated in parallel by other microRNAs as well. This study may further the understanding of gene expression regulation in the human developing pancreas.

MicroRNA signature of the human developing pancreas

PubMed Central

2010-01-01

Background MicroRNAs are non-coding RNAs that regulate gene expression including differentiation and development by either inhibiting translation or inducing target degradation. The aim of this study is to determine the microRNA expression signature during human pancreatic development and to identify potential microRNA gene targets calculating correlations between the signature microRNAs and their corresponding mRNA targets, predicted by bioinformatics, in genome-wide RNA microarray study. Results The microRNA signature of human fetal pancreatic samples 10-22 weeks of gestational age (wga), was obtained by PCR-based high throughput screening with Taqman Low Density Arrays. This method led to identification of 212 microRNAs. The microRNAs were classified in 3 groups: Group number I contains 4 microRNAs with the increasing profile; II, 35 microRNAs with decreasing profile and III with 173 microRNAs, which remain unchanged. We calculated Pearson correlations between the expression profile of microRNAs and target mRNAs, predicted by TargetScan 5.1 and miRBase altgorithms, using genome-wide mRNA expression data. Group I correlated with the decreasing expression of 142 target mRNAs and Group II with the increasing expression of 876 target mRNAs. Most microRNAs correlate with multiple targets, just as mRNAs are targeted by multiple microRNAs. Among the identified targets are the genes and transcription factors known to play an essential role in pancreatic development. Conclusions We have determined specific groups of microRNAs in human fetal pancreas that change the degree of their expression throughout the development. A negative correlative analysis suggests an intertwined network of microRNAs and mRNAs collaborating with each other. This study provides information leading to potential two-way level of combinatorial control regulating gene expression through microRNAs targeting multiple mRNAs and, conversely, target mRNAs regulated in parallel by other microRNAs as well. This study may further the understanding of gene expression regulation in the human developing pancreas. PMID:20860821

NORMAL NASAL GENE EXPRESSION LEVELS USING CDNA ARRAY TECHNOLOGY

EPA Science Inventory

Normal Nasal Gene Expression Levels Using cDNA Array Technology.

The nasal epithelium is a target site for chemically-induced toxicity and carcinogenicity. To detect and analyze genetic events which contribute to nasal tumor development, we first defined the gene expressi...

Ketones and lactate increase cancer cell “stemness”, driving recurrence, metastasis and poor clinical outcome in breast cancer

PubMed Central

Tsirigos, Aristotelis; Lin, Zhao; Pavlides, Stephanos; Wang, Chengwang; Flomenberg, Neal; Knudsen, Erik S; Howell, Anthony; Pestell, Richard G

2011-01-01

Previously, we showed that high-energy metabolites (lactate and ketones) “fuel” tumor growth and experimental metastasis in an in vivo xenograft model, most likely by driving oxidative mitochondrial metabolism in breast cancer cells. To mechanistically understand how these metabolites affect tumor cell behavior, here we used genome-wide transcriptional profiling. Human breast cancer cells (MCF7) were cultured with lactate or ketones, and then subjected to transcriptional analysis (exon-array). Interestingly, our results show that treatment with these high-energy metabolites increases the transcriptional expression of gene profiles normally associated with “stemness”, including genes upregulated in embryonic stem (ES) cells. Similarly, we observe that lactate and ketones promote the growth of bonafide ES cells, providing functional validation. The lactate- and ketone-induced “gene signatures” were able to predict poor clinical outcome (including recurrence and metastasis) in human breast cancer patients. Taken together, our results are consistent with the idea that lactate and ketone utilization in cancer cells promotes the “cancer stem cell” phenotype, resulting in significant decreases in patient survival. One possible mechanism by which high-energy metabolites might induce stemness is by increasing the pool of Acetyl-CoA, leading to increased histone acetylation and elevated gene expression. Thus, our results mechanistically imply that clinical outcome in breast cancer could simply be determined by epigenetics and energy metabolism, rather than by the accumulation of specific “classical” gene mutations. We also suggest that high-risk cancer patients (identified by the lactate/ketone gene signatures) could be treated with new therapeutics that target oxidative mitochondrial metabolism, such as the anti-oxidant and “mitochondrial poison” metformin. Finally, we propose that this new approach to personalized cancer medicine be termed “metabolo-genomics,” which incorporates features of both (1) cell metabolism and (2) gene transcriptional profiling. This powerful new approach directly links cancer cell metabolism with clinical outcome, and suggests new therapeutic strategies for inhibiting the TCA cycle and mitochondrial oxidative phosphorylation in cancer cells. PMID:21512313

ANALYSIS OF CHANGES IN GENE EXPRESSION PATTERNS IN FISH EXPOSED TO NATURAL PHARMACEUTICAL AND ENVIRONMENTAL ESTROGENS USING GENE ARRAYS.

EPA Science Inventory

Denslow, N.D., P. Larkin, T.L. Sabo-Attwood, J. Kocerha, K.J. Kroll, M.J. Hemmer and L.C. Folmar. 2004. Analysis of Changes in Gene Expression Patterns in Fish Exposed to Natural, Pharmaceutical and Environmental Estrogens Using Gene Arrays (Abstract). Mar. Environ. Res. 58(2-5):...

Innate inflammatory gene expression profiling in potential brain-dead donors: detailed investigation of the effect of common corticosteroid therapy.

PubMed

Gholamnezhadjafari, Reza; Tajik, Nader; Falak, Reza; Aflatoonian, Reza; Dehghan, Sanaz; Rezaei, Abbas

2017-07-01

Our study aimed to assess the influence of common methylprednisolone therapy on innate inflammatory factors in potential brain-dead organ donors (BDDs). The study groups consisted of 50 potential BDDs who received 15 mg/kg/d methylprednisolone and 25 live organ donors (LDs) as control group. Innate immunity gene expression profiling was performed by RT-PCR array. Soluble serum cytokines and chemokines, complement components, heat shock protein 70 (HSP70) and high mobility group box-1 (HMGB1) were measured by ELISA. Surface expression of TLR2 and TLR4 were determined using flow cytometry. Gene expression profiling revealed up-regulation of TLRs 1, 2, 4, 5, 6, 7 and 8, MYD88, NF-κB, NF-κB1A, IRAK1, STAT3, JAK2, TNF-α, IL-1β, CD86 and CD14 in the BDD group. Remarkably, the serum levels of C-reactive protein and HSP70 were considerably higher in the BDD group. In addition, serum amounts of IL-1β, IL-6, TNF-α, HMGB1, HSP70, C3a and C5a, but not IL-8, sCD86 or monocyte chemoattractant protein-1, were significantly increased in the BDD group. Significant differences were observed in flow cytometry analysis of TLR2 and TLR4 between the two groups. In summary, common methylprednisolone therapy in BDDs did not adequately reduce systemic inflammation, which could be due to inadequate doses or inefficient impact on other inflammatory-inducing pathways, for example oxidative stress or production of damage-associated molecules.

Human cell toxicogenomic analysis of bromoacetic acid: a regulated drinking water disinfection by-product.

PubMed

Muellner, Mark G; Attene-Ramos, Matias S; Hudson, Matthew E; Wagner, Elizabeth D; Plewa, Michael J

2010-04-01

The disinfection of drinking water is a major achievement in protecting the public health. However, current disinfection methods also generate disinfection by-products (DBPs). Many DBPs are cytotoxic, genotoxic, teratogenic, and carcinogenic and represent an important class of environmentally hazardous chemicals that may carry long-term human health implications. The objective of this research was to integrate in vitro toxicology with focused toxicogenomic analysis of the regulated DBP, bromoacetic acid (BAA) and to evaluate modulation of gene expression involved in DNA damage/repair and toxic responses, with nontransformed human cells. We generated transcriptome profiles for 168 genes with 30 min and 4 hr exposure times that did not induce acute cytotoxicity. Using qRT-PCR gene arrays, the levels of 25 transcripts were modulated to a statistically significant degree in response to a 30 min treatment with BAA (16 transcripts upregulated and nine downregulated). The largest changes were observed for RAD9A and BRCA1. The majority of the altered transcript profiles are genes involved in DNA repair, especially the repair of double strand DNA breaks, and in cell cycle regulation. With 4 hr of treatment the expression of 28 genes was modulated (12 upregulated and 16 downregulated); the largest fold changes were in HMOX1 and FMO1. This work represents the first nontransformed human cell toxicogenomic study with a regulated drinking water disinfection by-product. These data implicate double strand DNA breaks as a feature of BAA exposure. Future toxicogenomic studies of DBPs will further strengthen our limited knowledge in this growing area of drinking water research. Copyright 2009 Wiley-Liss, Inc.

Gene Expression Profiling of Acute Lymphoblastic Leukemia in Children with Very Early Relapse.

PubMed

Núñez-Enríquez, Juan Carlos; Bárcenas-López, Diego Alberto; Hidalgo-Miranda, Alfredo; Jiménez-Hernández, Elva; Bekker-Méndez, Vilma Carolina; Flores-Lujano, Janet; Solis-Labastida, Karina Anastacia; Martínez-Morales, Gabriela Bibiana; Sánchez-Muñoz, Fausto; Espinoza-Hernández, Laura Eugenia; Velázquez-Aviña, Martha Margarita; Merino-Pasaye, Laura Elizabeth; García Velázquez, Alejandra Jimena; Pérez-Saldívar, María Luisa; Mojica-Espinoza, Raúl; Ramírez-Bello, Julián; Jiménez-Morales, Silvia; Mejía-Aranguré, Juan Manuel

2016-11-01

Acute lymphoblastic leukemia (ALL) is the most common childhood cancer worldwide. Mexican patients have high mortality rates, low frequency of good prognosis biomarkers (i.e., ETV6-RUNX1) and a high proportion is classified at the time of diagnosis with a high risk to relapse according to clinical features. In addition, very early relapses are more frequently observed than in other populations. The aim of the study was to identify new potential biomarkers associated with very early relapse in Mexican ALL children through transcriptome analysis. Microarray gene expression profiling on bone marrow samples of 54 pediatric ALL patients, collected at time of diagnosis and/or at relapse, was performed. Eleven patients presented relapse within the first 18 months after diagnosis. Affymetrix Human Transcriptome Array 2.0 (HTA 2.0) was used to perform gene expression analysis. Annotation and functional enrichment analyses were carried out using Gene Ontology, KEGG pathway analysis and Ingenuity Pathway Analysis tools. BLVRB, ZCCHC7, PAX5, EBF1, TMOD1 and BLNK were differentially expressed (fold-change >2.0 and p value <0.01) between relapsed and non-relapsed patients. Functional analysis of abnormally expressed genes revealed their important role in cellular processes related to the development of hematological diseases, cancer, cell death and survival and in cell-to-cell signaling interaction. Our data support previous findings showing the relevance of PAX5, EBF1 and ZCCHC7 as potential biomarkers to identify a subgroup of ALL children in high risk to relapse. Copyright © 2016 IMSS. Published by Elsevier Inc. All rights reserved.

Comparative transcriptomics indicate changes in cell wall organization and stress response in seedlings during spaceflight.

PubMed

Johnson, Christina M; Subramanian, Aswati; Pattathil, Sivakumar; Correll, Melanie J; Kiss, John Z

2017-08-21

Plants will play an important role in the future of space exploration as part of bioregenerative life support. Thus, it is important to understand the effects of microgravity and spaceflight on gene expression in plant development. We analyzed the transcriptome of Arabidopsis thaliana using the Biological Research in Canisters (BRIC) hardware during Space Shuttle mission STS-131. The bioinformatics methods used included RMA (robust multi-array average), MAS5 (Microarray Suite 5.0), and PLIER (probe logarithmic intensity error estimation). Glycome profiling was used to analyze cell wall composition in the samples. In addition, our results were compared to those of two other groups using the same hardware on the same mission (BRIC-16). In our BRIC-16 experiments, we noted expression changes in genes involved in hypoxia and heat shock responses, DNA repair, and cell wall structure between spaceflight samples compared to the ground controls. In addition, glycome profiling supported our expression analyses in that there was a difference in cell wall components between ground control and spaceflight-grown plants. Comparing our studies to those of the other BRIC-16 experiments demonstrated that, even with the same hardware and similar biological materials, differences in results in gene expression were found among these spaceflight experiments. A common theme from our BRIC-16 space experiments and those of the other two groups was the downregulation of water stress response genes in spaceflight. In addition, all three studies found differential regulation of genes associated with cell wall remodeling and stress responses between spaceflight-grown and ground control plants. © 2017 Botanical Society of America.

Knock down of GCN5 histone acetyltransferase by siRNA decreases ethanol-induced histone acetylation and affects differential expression of genes in human hepatoma cells.

PubMed

Choudhury, Mahua; Pandey, Ravi S; Clemens, Dahn L; Davis, Justin Wade; Lim, Robert W; Shukla, Shivendra D

2011-06-01

We have investigated whether Gcn5, a histone acetyltransferase (HAT), is involved in ethanol-induced acetylation of histone H3 at lysine 9 (H3AcK9) and has any effect on the gene expression. Human hepatoma HepG2 cells transfected with ethanol-metabolizing enzyme alcohol dehydrogenase 1 (VA 13 cells) were used. Knock down of Gcn5 by siRNA silencing decreased mRNA and protein levels of general control nondepressible 5 (GCN5), HAT activity, and also attenuated ethanol-induced H3AcK9 in VA13 cells. Illumina gene microarray analysis using total RNA showed 940 transcripts affected by GCN5 silencing or ethanol. Silencing caused differential expression of 891 transcripts (≥1.5-fold upregulated or downregulated). Among these, 492 transcripts were upregulated and 399 were downregulated compared with their respective controls. Using a more stringent threshold (≥2.5-fold), the array data from GCN5-silenced samples showed 57 genes differentially expressed (39 upregulated and 18 downregulated). Likewise, ethanol caused differential regulation of 57 transcripts with ≥1.5-fold change (35 gene upregulated and 22 downregulated). Further analysis showed that eight genes were differentially regulated that were common for both ethanol treatment and GCN5 silencing. Among these, SLC44A2 (a putative choline transporter) was strikingly upregulated by ethanol (three fold), and GCN5 silencing downregulated it (1.5-fold). The quantitative real-time polymerase chain reaction profile corroborated the array findings. This report demonstrates for the first time that (1) GCN5 differentially affects expression of multiple genes, (2) ethanol-induced histone H3-lysine 9 acetylation is mediated via GCN5, and (3) GCN5 is involved in ethanol-induced expression of the putative choline transporter SLC44A2. Copyright © 2011 Elsevier Inc. All rights reserved.

Genomic imbalances in esophageal carcinoma cell lines involve Wnt pathway genes.

PubMed

Brown, Jacqueline; Bothma, Hannelie; Veale, Robin; Willem, Pascale

2011-06-28

To identify molecular markers shared across South African esophageal squamous cell carcinoma (ESCC) cell lines using cytogenetics, fluorescence in situ hybridization (FISH) and single nucleotide polymorphism (SNP) array copy number analysis. We used conventional cytogenetics, FISH, and multicolor FISH to characterize the chromosomal rearrangements of five ESCC cell lines established in South Africa. The whole genome copy number profile was established from 250K SNP arrays, and data was analyzed with the CNAT 4.0 and GISTIC software. We detected common translocation breakpoints involving chromosomes 1p11-12 and 3p11.2, the latter correlated with the deletion, or interruption of the EPHA3 gene. The most significant amplifications involved the following chromosomal regions and genes: 11q13.3 (CCND1, FGF3, FGF4, FGF19, MYEOV), 8q24.21(C-MYC, FAM84B), 11q22.1-q22.3 (BIRC2, BIRC3), 5p15.2 (CTNND2), 3q11.2-q12.2 (MINA) and 18p11.32 (TYMS, YES1). The significant deletions included 1p31.2-p31.1 (CTH, GADD45α, DIRAS3), 2q22.1 (LRP1B), 3p12.1-p14.2 (FHIT), 4q22.1-q32.1 (CASP6, SMAD1), 8p23.2-q11.1 (BNIP3L) and 18q21.1-q21.2 (SMAD4, DCC). The 3p11.2 translocation breakpoint was shared across four cell lines, supporting a role for genes involved at this site, in particular, the EPHA3 gene which has previously been reported to be deleted in ESCC. The finding that a significant number of genes that were amplified (FGF3, FGF4, FGF19, CCND1 and C-MYC) or deleted (SFRP2 gene) are involved in the Wnt and fibroblast growth factor signaling pathways, suggests that these pathways may be activated in these cell lines.

Genomic imbalances in esophageal carcinoma cell lines involve Wnt pathway genes

PubMed Central

Brown, Jacqueline; Bothma, Hannelie; Veale, Robin; Willem, Pascale

2011-01-01

AIM: To identify molecular markers shared across South African esophageal squamous cell carcinoma (ESCC) cell lines using cytogenetics, fluorescence in situ hybridization (FISH) and single nucleotide polymorphism (SNP) array copy number analysis. METHODS: We used conventional cytogenetics, FISH, and multicolor FISH to characterize the chromosomal rearrangements of five ESCC cell lines established in South Africa. The whole genome copy number profile was established from 250K SNP arrays, and data was analyzed with the CNAT 4.0 and GISTIC software. RESULTS: We detected common translocation breakpoints involving chromosomes 1p11-12 and 3p11.2, the latter correlated with the deletion, or interruption of the EPHA3 gene. The most significant amplifications involved the following chromosomal regions and genes: 11q13.3 (CCND1, FGF3, FGF4, FGF19, MYEOV), 8q24.21(C-MYC, FAM84B), 11q22.1-q22.3 (BIRC2, BIRC3), 5p15.2 (CTNND2), 3q11.2-q12.2 (MINA) and 18p11.32 (TYMS, YES1). The significant deletions included 1p31.2-p31.1 (CTH, GADD45α, DIRAS3), 2q22.1 (LRP1B), 3p12.1-p14.2 (FHIT), 4q22.1-q32.1 (CASP6, SMAD1), 8p23.2-q11.1 (BNIP3L) and 18q21.1-q21.2 (SMAD4, DCC). The 3p11.2 translocation breakpoint was shared across four cell lines, supporting a role for genes involved at this site, in particular, the EPHA3 gene which has previously been reported to be deleted in ESCC. CONCLUSION: The finding that a significant number of genes that were amplified (FGF3, FGF4, FGF19, CCND1 and C-MYC) or deleted (SFRP2 gene) are involved in the Wnt and fibroblast growth factor signaling pathways, suggests that these pathways may be activated in these cell lines. PMID:21734802

Genomic analysis of lung cell lines exposures to space radiation and the effect of lunar dust on selected fibrosis gene using RT2 PCR Array

NASA Astrophysics Data System (ADS)

Yeshitla, Samrawit

In the United States (U.S.), lung cancer is the number one cause of cancer death among men and women. Previous studies on human and animal epithelial lung cells showed that ionizing radiation and certain environmental pollutants are carcinogens. The surface area of the lungs and the slow turnover rate of the epithelial cells are suggested to play a role in the vulnerability of the cells, which lead to increase in the progenitor cell of the lung. It has been proposed that these progenitor cells, when exposed to radiation undergo multiple alterations that cause the cells to become cancerous. The current thought is that the lungs contain several facultative progenitor cells that are situated throughout the lung epithelium and are regionally restricted in their regenerative capacity. In this study, normal Human Bronchial Epithelial Cells (HBECs) were immortalized through the expression of Cdk4 and hTERT and evaluated for the effects radiation using in vitro study. The HBECs retained its novel multipotent capacity in vitro and represented unrestricted progenitor cells of the adult lungs, which resemble an embryonic progenitor. Analysis of the transformed clones of human bronchial epithelial cell line, HEBC3KT exposed to Fe ions and gamma rays revealed chromosomal abnormality, which was detected with the Multi-color Fluorescent In Situ Hybridization (mFish). In Part two of this study the F344 rats exposed to lunar dust, for 4 weeks (6h/d; 5d/wk.) in nose-only inhalation chambers at concentrations of 0 (control air), 2.1, 6.8, 20.8, and 61 mg/m3 of lunar dust, were used to determine the lunar dust toxicity on the lung tissues and total RNA were prepared from the tissues and used for gene expression. Analysis of gene expression data using Ingenuity Pathway Analysis tool identified multiple pathways of which fibrosis was one of the pathways. The Rat Fibrosis RT 2 Profile PCR Array was used to profile the expression of 84 genes that are relevant to fibrosis in the lung tissue, after removing the infiltrated of the bronchoalveolar lavage (BAL) cells by lavaging. At a dose of 61mg/m 3, 29 genes showed changes and of 29 genes, nine genes, Bmp7, Cc112, Cc13, Itgb8, Serpinel, Smad6, Tgfbrl, Thbs2, and Tnf, showed significant fold change difference of up-regulation or down-regulation. The data in this study showed that iron ions and gamma rays promote chromosomal abnormality in HBEC3KT, and for the first time, the lunar dust altered gene expression of 29 genes that are relevant to fibrosis in the lung tissue.

Dynamic gene expression response to altered gravity in human T cells.

PubMed

Thiel, Cora S; Hauschild, Swantje; Huge, Andreas; Tauber, Svantje; Lauber, Beatrice A; Polzer, Jennifer; Paulsen, Katrin; Lier, Hartwin; Engelmann, Frank; Schmitz, Burkhard; Schütte, Andreas; Layer, Liliana E; Ullrich, Oliver

2017-07-12

We investigated the dynamics of immediate and initial gene expression response to different gravitational environments in human Jurkat T lymphocytic cells and compared expression profiles to identify potential gravity-regulated genes and adaptation processes. We used the Affymetrix GeneChip® Human Transcriptome Array 2.0 containing 44,699 protein coding genes and 22,829 non-protein coding genes and performed the experiments during a parabolic flight and a suborbital ballistic rocket mission to cross-validate gravity-regulated gene expression through independent research platforms and different sets of control experiments to exclude other factors than alteration of gravity. We found that gene expression in human T cells rapidly responded to altered gravity in the time frame of 20 s and 5 min. The initial response to microgravity involved mostly regulatory RNAs. We identified three gravity-regulated genes which could be cross-validated in both completely independent experiment missions: ATP6V1A/D, a vacuolar H + -ATPase (V-ATPase) responsible for acidification during bone resorption, IGHD3-3/IGHD3-10, diversity genes of the immunoglobulin heavy-chain locus participating in V(D)J recombination, and LINC00837, a long intergenic non-protein coding RNA. Due to the extensive and rapid alteration of gene expression associated with regulatory RNAs, we conclude that human cells are equipped with a robust and efficient adaptation potential when challenged with altered gravitational environments.

Dynamic gene expression changes precede dioxin-induced liver pathogenesis in medaka fish.

PubMed

Volz, David C; Hinton, David E; Law, J McHugh; Kullman, Seth W

2006-02-01

A major challenge for environmental genomics is linking gene expression to cellular toxicity and morphological alteration. Herein, we address complexities related to hepatic gene expression responses after a single injection of the aryl hydrocarbon receptor (AHR) agonist 2,3,7,8-tetrachlorodibenzo-p-dioxin (dioxin) and illustrate an initial stress response followed by cytologic and adaptive changes in the teleost fish medaka. Using a custom 175-gene array, we find that overall hepatic gene expression and histological changes are strongly dependent on dose and time. The most pronounced dioxin-induced gene expression changes occurred early and preceded morphologic alteration in the liver. Following a systematic search for putative Ah response elements (AHREs) (5'-CACGCA-3') within 2000 bp upstream of the predicted transcriptional start site, the majority (87%) of genes screened in this study did not contain an AHRE, suggesting that gene expression was not solely dependent on AHRE-mediated transcription. Moreover, in the highest dosage, we observed gene expression changes associated with adaptation that persisted for almost two weeks, including induction of a gene putatively identified as ependymin that may function in hepatic injury repair. These data suggest that the cellular response to dioxin involves both AHRE- and non-AHRE-mediated transcription, and that coupling gene expression profiling with analysis of morphologic pathogenesis is essential for establishing temporal relationships between transcriptional changes, toxicity, and adaptation to hepatic injury.

DGEM--a microarray gene expression database for primary human disease tissues.

PubMed

Xia, Yuni; Campen, Andrew; Rigsby, Dan; Guo, Ying; Feng, Xingdong; Su, Eric W; Palakal, Mathew; Li, Shuyu

2007-01-01

Gene expression patterns can reflect gene regulations in human tissues under normal or pathologic conditions. Gene expression profiling data from studies of primary human disease samples are particularly valuable since these studies often span many years in order to collect patient clinical information and achieve a large sample size. Disease-to-Gene Expression Mapper (DGEM) provides a beneficial community resource to access and analyze these data; it currently includes Affymetrix oligonucleotide array datasets for more than 40 human diseases and 1400 samples. The data are normalized to the same scale and stored in a relational database. A statistical-analysis pipeline was implemented to identify genes abnormally expressed in disease tissues or genes whose expressions are associated with clinical parameters such as cancer patient survival. Data-mining results can be queried through a web-based interface at http://dgem.dhcp.iupui.edu/. The query tool enables dynamic generation of graphs and tables that are further linked to major gene and pathway resources that connect the data to relevant biology, including Entrez Gene and Kyoto Encyclopedia of Genes and Genomes (KEGG). In summary, DGEM provides scientists and physicians a valuable tool to study disease mechanisms, to discover potential disease biomarkers for diagnosis and prognosis, and to identify novel gene targets for drug discovery. The source code is freely available for non-profit use, on request to the authors.

Transcriptional profiling of rat skeletal muscle hypertrophy under restriction of blood flow.

PubMed

Xu, Shouyu; Liu, Xueyun; Chen, Zhenhuang; Li, Gaoquan; Chen, Qin; Zhou, Guoqing; Ma, Ruijie; Yao, Xinmiao; Huang, Xiao

2016-12-15

Blood flow restriction (BFR) under low-intensity resistance training (LIRT) can produce similar effects upon muscles to that of high-intensity resistance training (HIRT) while overcoming many of the restrictions to HIRT that occurs in a clinical setting. However, the potential molecular mechanisms of BFR induced muscle hypertrophy remain largely unknown. Here, using a BFR rat model, we aim to better elucidate the mechanisms regulating muscle hypertrophy as induced by BFR and reveal possible clinical therapeutic targets for atrophy cases. We performed genome wide screening with microarray analysis to identify unique differentially expressed genes during rat muscle hypertrophy. We then successfully separated the differentially expressed genes from BRF treated soleus samples by comparing the Affymetrix rat Genome U34 2.0 array with the control. Using qRT-PCR and immunohistochemistry (IHC) we also analyzed other related differentially expressed genes. Results suggested that muscle hypertrophy induced by BFR is essentially regulated by the rate of protein turnover. Specifically, PI3K/AKT and MAPK pathways act as positive regulators in controlling protein synthesis where ubiquitin-proteasome acts as a negative regulator. This represents the first general genome wide level investigation of the gene expression profile in the rat soleus after BFR treatment. This may aid our understanding of the molecular mechanisms regulating and controlling muscle hypertrophy and provide support to the BFR strategies aiming to prevent muscle atrophy in a clinical setting. Copyright © 2016 Elsevier B.V. All rights reserved.

Characterization of a Genomic Signature of Pregnancy in the Breast

PubMed Central

Belitskaya-Lévy, Ilana; Zeleniuch-Jacquotte, Anne; Russo, Jose; Russo, Irma H.; Bordás, Pal; Åhman, Janet; Afanasyeva, Yelena; Johansson, Robert; Lenner, Per; Li, Xiaochun; de Cicco, Ricardo López; Peri, Suraj; Ross, Eric; Russo, Patricia A.; Santucci-Pereira, Julia; Sheriff, Fathima S.; Slifker, Michael; Hallmans, Göran; Toniolo, Paolo; Arslan, Alan A.

2012-01-01

The objective of the current study was to comprehensively compare the genomic profiles in the breast of parous and nulliparous postmenopausal women to identify genes that permanently change their expression following pregnancy. The study was designed as a two-phase approach. In the discovery phase, we compared breast genomic profiles of 37 parous with 18 nulliparous postmenopausal women. In the validation phase, confirmation of the genomic patterns observed in the discovery phase was sought in an independent set of 30 parous and 22 nulliparous postmenopausal women. RNA was hybridized to Affymetrix HG_U133 Plus 2.0 oligonucleotide arrays containing probes to 54,675 transcripts; scanned and the images analyzed using Affymetrix GCOS software. Surrogate variable analysis, logistic regression and significance analysis for microarrays were used to identify statistically significant differences in expression of genes. The False Discovery Rate (FDR) approach was used to control for multiple comparisons. We found that 208 genes (305 probe sets) were differentially expressed between parous and nulliparous women in both discovery and validation phases of the study at a FDR of 10% and with at least a 1.25-fold change. These genes are involved in regulation of transcription, centrosome organization, RNA splicing, cell cycle control, adhesion and differentiation. The results provide persuasive evidence that full-term pregnancy induces long-term genomic changes in the breast. The genomic signature of pregnancy could be used as an intermediate marker to assess potential chemopreventive interventions with hormones mimicking the effects of pregnancy for prevention of breast cancer. PMID:21622728

MicroRNA profiling of the murine hematopoietic system

PubMed Central

Monticelli, Silvia; Ansel, K Mark; Xiao, Changchun; Socci, Nicholas D; Krichevsky, Anna M; Thai, To-Ha; Rajewsky, Nikolaus; Marks, Debora S; Sander, Chris; Rajewsky, Klaus; Rao, Anjana; Kosik, Kenneth S

2005-01-01

Background MicroRNAs (miRNAs) are a class of recently discovered noncoding RNA genes that post-transcriptionally regulate gene expression. It is becoming clear that miRNAs play an important role in the regulation of gene expression during development. However, in mammals, expression data are principally based on whole tissue analysis and are still very incomplete. Results We used oligonucleotide arrays to analyze miRNA expression in the murine hematopoietic system. Complementary oligonucleotides capable of hybridizing to 181 miRNAs were immobilized on a membrane and probed with radiolabeled RNA derived from low molecular weight fractions of total RNA from several different hematopoietic and neuronal cells. This method allowed us to analyze cell type-specific patterns of miRNA expression and to identify miRNAs that might be important for cell lineage specification and/or cell effector functions. Conclusion This is the first report of systematic miRNA gene profiling in cells of the hematopoietic system. As expected, miRNA expression patterns were very different between hematopoietic and non-hematopoietic cells, with further subtle differences observed within the hematopoietic group. Interestingly, the most pronounced similarities were observed among fully differentiated effector cells (Th1 and Th2 lymphocytes and mast cells) and precursors at comparable stages of differentiation (double negative thymocytes and pro-B cells), suggesting that in addition to regulating the process of commitment to particular cellular lineages, miRNAs might have an important general role in the mechanism of cell differentiation and maintenance of cell identity. PMID:16086853

Gene profile in the spleen under massive partial hepatectomy using complementary DNA microarray and pathway analysis.

PubMed

Arakawa, Yusuke; Shimada, Mitsuo; Utsunomiya, Tohru; Imura, Satoru; Morine, Yuji; Ikemoto, Tetsuya; Mori, Hiroki; Kanamoto, Mami; Iwahashi, Shuichi; Saito, Yu; Takasu, Chie

2014-08-01

In general, the spleen is one of the abdominal organs connected by the portal system, and a splenectomy improves hepatic functions in the settings of partial hepatectomy (Hx) for portal hypertensive cases or living donor liver transplantation with excessive portal vein flow. Those precise mechanisms remain still unclear; therefore, we investigated the DNA expression profile in the spleen after 90% Hx in rats using complementary DNA microarray and pathway analysis. Messenger RNAs (mRNAs) were prepared from three rat spleens at each time point (0, 3, and 6 h after 90% Hx). Using the gene chip, mRNA was hybridized to Affymetrix GeneChip Rat Genome 230 2.0 Array (Affymetrix®) and pathway analysis was done with Ingenuity Pathway Analysis (IPA®). We determined the 3-h or 6-h/0-h ratio to assess the influence of Hx, and cut-off values were set at more than 2.0-fold or less than 1/2 (0.5)-fold. Chemokine activity-related genes including Cxcl1 (GRO1) and Cxcl2 (MIP-2) related pathway were upregulated in the spleen. Also, immediate early response genes including early growth response-1 (EGR1), FBJ murine osteosarcoma (FOS) and activating transcription factor 3 (ATF3) related pathway were upregulated in the spleen. We concluded that in the spleen the expression of numerous inflammatory-related genes would occur after 90% Hx. The spleen could take a harmful role and provide a negative impact during post Hx phase due to the induction of chemokine and transcription factors including GRO1 and EGR1. © 2014 Journal of Gastroenterology and Hepatology Foundation and Wiley Publishing Asia Pty Ltd.

Early bovine embryos regulate oviduct epithelial cell gene expression during in vitro co-culture.

PubMed

Schmaltz-Panneau, Barbara; Cordova, Amanda; Dhorne-Pollet, Sophie; Hennequet-Antier, Christelle; Uzbekova, Sveltlana; Martinot, Emmanuelle; Doret, Sarah; Martin, Patrice; Mermillod, Pascal; Locatelli, Yann

2014-10-01

In mammals, the oviduct may participate to the regulation of early embryo development. In vitro co-culture of early bovine embryos with bovine oviduct epithelial cells (BOEC) has been largely used to mimic the maternal environment. However, the mechanisms of BOEC action have not been clearly elucidated yet. The aim of this study was to determine the response of BOEC cultures to the presence of developing bovine embryos. A 21,581-element bovine oligonucleotide array was used compare the gene expression profiles of confluent BOEC cultured for 8 days with or without embryos. This study revealed 34 differentially expressed genes (DEG). Of these 34 genes, IFI6, ISG15, MX1, IFI27, IFI44, RSAD2, IFITM1, EPSTI1, USP18, IFIT5, and STAT1 expression increased to the greatest extent due to the presence of embryos with a major impact on antiviral and immune response. Among the mRNAs at least 25 are already described as induced by interferons. In addition, transcript levels of new candidate genes involved in the regulation of transcription, modulation of the maternal immune system and endometrial remodeling were found to be increased. We selected 7 genes and confirmed their differential expression by quantitative RT-PCR. The immunofluorescence imaging of cellular localization of STAT1 protein in BOEC showed a nuclear translocation in the presence of embryos, suggesting the activation of interferon signaling pathway. This first systematic study of BOEC transcriptome changes in response to the presence of embryos in cattle provides some evidences that these cells are able to adapt their transcriptomic profile in response to embryo signaling. Copyright © 2014 Elsevier B.V. All rights reserved.

Differential gene expression in granulosa cells from polycystic ovary syndrome patients with and without insulin resistance: identification of susceptibility gene sets through network analysis.

PubMed

Kaur, Surleen; Archer, Kellie J; Devi, M Gouri; Kriplani, Alka; Strauss, Jerome F; Singh, Rita

2012-10-01

Polycystic ovary syndrome (PCOS) is a heterogeneous, genetically complex, endocrine disorder of uncertain etiology in women. Our aim was to compare the gene expression profiles in stimulated granulosa cells of PCOS women with and without insulin resistance vs. matched controls. This study included 12 normal ovulatory women (controls), 12 women with PCOS without evidence for insulin resistance (PCOS non-IR), and 16 women with insulin resistance (PCOS-IR) undergoing in vitro fertilization. Granulosa cell gene expression profiling was accomplished using Affymetrix Human Genome-U133 arrays. Differentially expressed genes were classified according to gene ontology using ingenuity pathway analysis tools. Microarray results for selected genes were confirmed by real-time quantitative PCR. A total of 211 genes were differentially expressed in PCOS non-IR and PCOS-IR granulosa cells (fold change≥1.5; P≤0.001) vs. matched controls. Diabetes mellitus and inflammation genes were significantly increased in PCOS-IR patients. Real-time quantitative PCR confirmed higher expression of NCF2 (2.13-fold), TCF7L2 (1.92-fold), and SERPINA1 (5.35-fold). Increased expression of inflammation genes ITGAX (3.68-fold) and TAB2 (1.86-fold) was confirmed in PCOS non-IR. Different cardiometabolic disease genes were differentially expressed in the two groups. Decreased expression of CAV1 (-3.58-fold) in PCOS non-IR and SPARC (-1.88-fold) in PCOS-IR was confirmed. Differential expression of genes involved in TGF-β signaling (IGF2R, increased; and HAS2, decreased), and oxidative stress (TXNIP, increased) was confirmed in both groups. Microarray analysis demonstrated differential expression of genes linked to diabetes mellitus, inflammation, cardiovascular diseases, and infertility in the granulosa cells of PCOS women with and without insulin resistance. Because these dysregulated genes are also involved in oxidative stress, lipid metabolism, and insulin signaling, we hypothesize that these genes may be involved in follicular growth arrest and metabolic disorders associated with the different phenotypes of PCOS.

Characterization of the Cartilage DNA Methylome in Knee and Hip Osteoarthritis

PubMed Central

Rushton, Michael D; Reynard, Louise N; Barter, Matt J; Refaie, Ramsay; Rankin, Kenneth S; Young, David A; Loughlin, John

2014-01-01

Objective The aim of this study was to characterize the genome-wide DNA methylation profile of chondrocytes from knee and hip cartilage obtained from patients with osteoarthritis (OA) and hip cartilage obtained from patients with femoral neck fracture, providing the first comparison of DNA methylation between OA and non-OA hip cartilage, and between OA hip and OA knee cartilage. Methods The study was performed using the Illumina Infinium HumanMethylation450 BeadChip array, which allows the annotation of ∼480,000 CpG sites. Genome-wide methylation was assessed in chondrocyte DNA extracted from 23 hip OA patients, 73 knee OA patients, and 21 healthy hip control patients with femoral neck fracture. Results Analysis revealed that chondrocytes from the hip cartilage of OA patients and healthy controls have unique methylation profiles, with 5,322 differentially methylated loci (DMLs) identified between the 2 groups. In addition, a comparison between hip and knee OA chondrocytes revealed 5,547 DMLs between the 2 groups, including DMLs in several genes known to be involved in the pathogenesis of OA. Hip OA samples were found to cluster into 2 groups. A total of 15,239 DMLs were identified between the 2 clusters, with an enrichment of genes involved in inflammation and immunity. Similarly, we confirmed a previous report of knee OA samples that also clustered into 2 groups. Conclusion We demonstrated that global DNA methylation using a high-density array can be a powerful tool in the characterization of OA at the molecular level. Identification of pathways enriched in DMLs between OA and OA-free cartilage highlight potential etiologic mechanisms that are involved in the initiation and/or progression of the disease and that could be therapeutically targeted. PMID:24838673

Meta-analysis of Clear Cell Renal Cell Carcinoma Gene Expression Defines a Variant Subgroup and Identifies Gender Influences on Tumor Biology

PubMed Central

Brannon, A. Rose; Haake, Scott M.; Hacker, Kathryn E.; Pruthi, Raj S.; Wallen, Eric M.; Nielsen, Matthew E.; Rathmell, W. Kimryn

2011-01-01

Background Clear cell renal cell carcinoma (ccRCC) displays molecular and histologic heterogeneity. Previously described subsets of this disease, ccA and ccB, were defined based on multigene expression profiles, but it is unclear whether these subgroupings reflect the full spectrum of disease or how these molecular subtypes relate to histologic descriptions or gender. Objective Determine whether additional subtypes of ccRCC exist and whether these subtypes are related to von Hippel-Lindau (VHL) inactivation, hypoxia-inducible factor (HIF) 1 and 2 expression, tumor histology, or gender. Design, setting, and participants Six large, publicly available ccRCC gene expression databases were identified that cumulatively provided data for 480 tumors for meta-analysis via meta-array compilation. Measurements Unsupervised consensus clustering was performed on the meta-arrays. Tumors were examined for the relationship of multigene-defined consensus subtypes and expression signatures of VHL mutation and HIF status, tumor histology, and gender. Results and limitations Two dominant subsets of ccRCC were observed. However, a minor third cluster was revealed that correlated strongly with a wild type (WT) VHL expression profile and indications of variant histologies. When variant histologies were removed, ccA tumors naturally divided by gender. This technique is limited by the potential for persistent batch effect, tumor sampling bias, and restrictions of annotated information. Conclusions The ccA and ccB subsets of ccRCC are robust in meta-analysis among histologically conventional ccRCC tumors. A third group of tumors was identified that may represent a new variant of ccRCC. Within definitively clear cell tumors, gender may delineate tumors in such a way that it could have implications regarding current treatments and future drug development. PMID:22030119

Analysis of differential gene expression by bead-based fiber-optic array in growth-hormone-secreting pituitary adenomas.

PubMed

Jiang, Zhiquan; Gui, Songbo; Zhang, Yazhuo

2010-09-01

Growth-hormone-secreting pituitary adenomas (GHomas) account for approximately 20% of all pituitary neoplasms. However, the pathogenesis of GHomas remains to be elucidated. To explore the possible pathogenesis of GHomas, we used bead-based fiber-optic arrays to examine the gene expression in five GHomas and compared them to three healthy pituitaries. Four differentially expressed genes were chosen randomly for validation by quantitative real-time reverse transcription-polymerase chain reaction. We then performed pathway analysis on the identified differentially expressed genes using the Kyoto Encyclopedia of Genes and Genomes. Array analysis showed significant increases in the expression of 353 genes and 206 expressed sequence tags (ESTs) and decreases in 565 genes and 29 ESTs. Bioinformatic analysis showed that the genes HIGD1B, HOXB2, ANGPT2, HPGD and BTG2 may play an important role in the tumorigenesis and progression of GHomas. Pathway analysis showed that the wingless-type signaling pathway and extracellular-matrix receptor interactions may play a key role in the tumorigenesis and progression of GHomas. Our data suggested that there are numerous aberrantly expressed genes and pathways involved in the pathogenesis of GHomas. Bead-based fiber-optic arrays combined with pathway analysis of differentially expressed genes appear to be a valid method for investigating the pathogenesis of tumors.

Analysis of differential gene expression by bead-based fiber-optic array in growth-hormone-secreting pituitary adenomas

PubMed Central

JIANG, ZHIQUAN; GUI, SONGBO; ZHANG, YAZHUO

2010-01-01

Growth-hormone-secreting pituitary adenomas (GHomas) account for approximately 20% of all pituitary neoplasms. However, the pathogenesis of GHomas remains to be elucidated. To explore the possible pathogenesis of GHomas, we used bead-based fiber-optic arrays to examine the gene expression in five GHomas and compared them to three healthy pituitaries. Four differentially expressed genes were chosen randomly for validation by quantitative real-time reverse transcription-polymerase chain reaction. We then performed pathway analysis on the identified differentially expressed genes using the Kyoto Encyclopedia of Genes and Genomes. Array analysis showed significant increases in the expression of 353 genes and 206 expressed sequence tags (ESTs) and decreases in 565 genes and 29 ESTs. Bioinformatic analysis showed that the genes HIGD1B, HOXB2, ANGPT2, HPGD and BTG2 may play an important role in the tumorigenesis and progression of GHomas. Pathway analysis showed that the wingless-type signaling pathway and extracellular-matrix receptor interactions may play a key role in the tumorigenesis and progression of GHomas. Our data suggested that there are numerous aberrantly expressed genes and pathways involved in the pathogenesis of GHomas. Bead-based fiber-optic arrays combined with pathway analysis of differentially expressed genes appear to be a valid method for investigating the pathogenesis of tumors. PMID:22993617

Performance parameters of a liquid filled ionization chamber array

DOE Office of Scientific and Technical Information (OSTI.GOV)

Poppe, B.; Stelljes, T. S.; Looe, H. K.

2013-08-15

Purpose: In this work, the properties of the two-dimensional liquid filled ionization chamber array Octavius 1000SRS (PTW-Freiburg, Germany) for use in clinical photon-beam dosimetry are investigated.Methods: Measurements were carried out at an Elekta Synergy and Siemens Primus accelerator. For measurements of stability, linearity, and saturation effects of the 1000SRS array a Semiflex 31013 ionization chamber (PTW-Freiburg, Germany) was used as a reference. The effective point of measurement was determined by TPR measurements of the array in comparison with a Roos chamber (type 31004, PTW-Freiburg, Germany). The response of the array with varying field size and depth of measurement was evaluatedmore » using a Semiflex 31010 ionization chamber as a reference. Output factor measurements were carried out with a Semiflex 31010 ionization chamber, a diode (type 60012, PTW-Freiburg, Germany), and the detector array under investigation. The dose response function for a single detector of the array was determined by measuring 1 cm wide slit-beam dose profiles and comparing them against diode-measured profiles. Theoretical aspects of the low pass properties and of the sampling frequency of the detector array were evaluated. Dose profiles measured with the array and the diode detector were compared, and an intensity modulated radiation therapy (IMRT) field was verified using the Gamma-Index method and the visualization of line dose profiles.Results: The array showed a short and long term stability better than 0.1% and 0.2%, respectively. Fluctuations in linearity were found to be within ±0.2% for the vendor specified dose range. Saturation effects were found to be similar to those reported in other studies for liquid-filled ionization chambers. The detector's relative response varied with field size and depth of measurement, showing a small energy dependence accounting for maximum signal deviations of ±2.6% from the reference condition for the setup used. The σ-values of the Gaussian dose response function for a single detector of the array were found to be (0.72 ± 0.25) mm at 6 MV and (0.74 ± 0.25) mm at 15 MV and the corresponding low pass cutoff frequencies are 0.22 and 0.21 mm{sup −1}, respectively. For the inner 5 × 5 cm{sup 2} region and the outer 11 × 11 cm{sup 2} region of the array the Nyquist theorem is fulfilled for maximum sampling frequencies of 0.2 and 0.1 mm{sup −1}, respectively. An IMRT field verification with a Gamma-Index analysis yielded a passing rate of 95.2% for a 3 mm/3% criterion with a TPS calculation as reference.Conclusions: This study shows the applicability of the Octavius 1000SRS in modern dosimetry. Output factor and dose profile measurements illustrated the applicability of the array in small field and stereotactic dosimetry. The high spatial resolution ensures adequate measurements of dose profiles in regular and intensity modulated photon-beam fields.« less

Optimised laser microdissection of the human ocular surface epithelial regions for microarray studies

PubMed Central

2013-01-01

Background The most important challenge of performing insitu transcriptional profiling of the human ocular surface epithelial regions is obtaining samples in sufficient amounts, without contamination from adjacent tissue, as the region of interest is microscopic and closely apposed to other tissues regions. We have effectively collected ocular surface (OS) epithelial tissue samples from the Limbal Epithelial Crypt (LEC), limbus, cornea and conjunctiva of post-mortem cadaver eyes with laser microdissection (LMD) technique for gene expression studies with spotted oligonucleotide microarrays and Gene 1.0 ST arrays. Methods Human donor eyes (4 pairs for spotted oligonucleotide microarrays, 3 pairs for Gene 1.0 ST arrays) consented for research were included in this study with due ethical approval of the Nottingham Research Ethics Committee. Eye retrieval was performed within 36 hours of post-mortem period. The dissected corneoscleral buttons were immersed in OCT media and frozen in liquid nitrogen and stored at −80°C till further use. Microscopic tissue sections of interest were taken on PALM slides and stained with Toluidine Blue for laser microdissection with PALM microbeam systems. Optimisation of the laser microdissection technique was crucial for efficient and cost effective sample collection. Results The starting concentration of RNA as stipulated by the protocol of microarray platforms was taken as the cut-off concentration of RNA samples in our studies. The area of LMD tissue processed for spotted oligonucleotide microarray study ranged from 86,253 μm2 in LEC to 392,887 μm2 in LEC stroma. The RNA concentration of the LMD samples ranged from 22 to 92 pg/μl. The recommended starting concentration of the RNA samples used for Gene 1.0 ST arrays was 6 ng/5 μl. To achieve the desired RNA concentration the area of ocular surface epithelial tissue sample processed for the Gene 1.0 ST array experiments was approximately 100,0000 μm2 to 130,0000 μm2. RNA concentration of these samples ranged from 10.88 ng/12 μl to 25.8 ng/12 μl, with the RNA integrity numbers (RIN) for these samples from 3.3 to 7.9. RNA samples with RIN values below 2, that had failed to amplify satisfactorily were discarded. Conclusions The optimised protocol for sample collection and laser microdissection improved the RNA yield of the insitu ocular surface epithelial regions for effective microarray studies on spotted oligonucleotide and affymetrix platforms. PMID:24160452

Human active X-specific DNA methylation events showing stability across time and tissues

PubMed Central

Joo, Jihoon Eric; Novakovic, Boris; Cruickshank, Mark; Doyle, Lex W; Craig, Jeffrey M; Saffery, Richard

2014-01-01

The phenomenon of X chromosome inactivation in female mammals is well characterised and remains the archetypal example of dosage compensation via monoallelic expression. The temporal series of events that culminates in inactive X-specific gene silencing by DNA methylation has revealed a ‘patchwork' of gene inactivation along the chromosome, with approximately 15% of genes escaping. Such genes are therefore potentially subject to sex-specific imbalance between males and females. Aside from XIST, the non-coding RNA on the X chromosome destined to be inactivated, very little is known about the extent of loci that may be selectively silenced on the active X chromosome (Xa). Using longitudinal array-based DNA methylation profiling of two human tissues, we have identified specific and widespread active X-specific DNA methylation showing stability over time and across tissues of disparate origin. Our panel of X-chromosome loci subject to methylation on Xa reflects a potentially novel mechanism for controlling female-specific X inactivation and sex-specific dimorphisms in humans. Further work is needed to investigate these phenomena. PMID:24713664

ChIP-Chip Identifies SEC23A, CFDP1, and NSD1 as TFII-I Target Genes in Human Neural Crest Progenitor Cells.

PubMed

Makeyev, Aleksandr V; Bayarsaihan, Dashzeveg

2013-05-01

Objectives :  GTF2I and GTF2IRD1 genes located in Williams-Beuren syndrome (WBS) critical region encode TFII-I family transcription factors. The aim of this study was to map genomic sites bound by these proteins across promoter regions of developmental regulators associated with craniofacial development. Design :  Chromatin was isolated from human neural crest progenitor cells and the DNA-binding profile was generated using the human RefSeq tiling promoter ChIP-chip arrays. Results :  TFII-I transcription factors are recruited to the promoters of SEC23A, CFDP1, and NSD1 previously defined as TFII-I target genes. Moreover, our analysis revealed additional binding elements that contain E-boxes and initiator-like motifs. Conclusions :  Genome-wide promoter binding studies revealed SEC23A, CFDP1, and NSD1 linked to craniofacial or dental development as direct TFII-I targets. Developmental regulation of these genes by TFII-I factors could contribute to the WBS-specific facial dysmorphism.

The Peripheral Whole Blood Transcriptome of Acute Pyelonephritis in Human Pregnancy

PubMed Central

Madan, Ichchha; Than, Nandor Gabor; Romero, Roberto; Chaemsaithong, Piya; Miranda, Jezid; Tarca, Adi L.; Bhatti, Gaurav; Draghici, Sorin; Yeo, Lami; Mazor, Moshe; Hassan, Sonia S.; Chaiworapongsa, Tinnakorn

2018-01-01

Objective Human pregnancy is characterized by activation of the innate immune response and suppression of adaptive immunity. The former is thought to provide protection against infection to the mother, and the latter, tolerance against paternal antigens expressed in fetal cells. Acute pyelonephritis is associated with an increased risk of acute respiratory distress syndrome and sepsis in pregnant (vs. nonpregnant) women. The objective of this study was to describe the gene expression profile (transcriptome) of maternal whole blood in acute pyelonephritis. Method A case-control study was conducted to include pregnant women with acute pyelonephritis (n=15) and women with a normal pregnancy (n=34). Affymetrix HG-U133 Plus 2.0 arrays (Affymetrix, Santa Clara, CA, USA) were used for gene expression profiling. A linear model was used to test the association between the presence of pyelonephritis and gene expression levels while controlling for white blood cell count and gestational age. A fold change of 1.5 was considered significant at a false discovery rate of 0.1. A subset of differentially expressed genes (n=56) was tested with real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) (cases, n=19; controls, n=59). Gene ontology and pathway analysis were applied. Results A total of 983 genes were differentially expressed in acute pyelonephritis: 457 were up-regulated and 526 were down-regulated. Significant enrichment of 300 biological processes and 63 molecular functions was found in pyelonephritis. Significantly impacted pathways in pyelonephritis included a) cytokine-cytokine receptor interaction; b) T-cell receptor signaling; c) Jak-STAT signaling; and d) complement and coagulation cascades. Of 56 genes tested by qRT-PCR, 48 (85.7%) had confirmation of differential expression. Conclusion This is the first study of the transcriptomic signature of whole blood in pregnant women with acute pyelonephritis. Acute infection during pregnancy is associated with the increased expression of genes involved in innate immunity and the decreased expression of genes involved in lymphocyte function. PMID:24293448

Missing value imputation in DNA microarrays based on conjugate gradient method.

PubMed

Dorri, Fatemeh; Azmi, Paeiz; Dorri, Faezeh

2012-02-01

Analysis of gene expression profiles needs a complete matrix of gene array values; consequently, imputation methods have been suggested. In this paper, an algorithm that is based on conjugate gradient (CG) method is proposed to estimate missing values. k-nearest neighbors of the missed entry are first selected based on absolute values of their Pearson correlation coefficient. Then a subset of genes among the k-nearest neighbors is labeled as the best similar ones. CG algorithm with this subset as its input is then used to estimate the missing values. Our proposed CG based algorithm (CGimpute) is evaluated on different data sets. The results are compared with sequential local least squares (SLLSimpute), Bayesian principle component analysis (BPCAimpute), local least squares imputation (LLSimpute), iterated local least squares imputation (ILLSimpute) and adaptive k-nearest neighbors imputation (KNNKimpute) methods. The average of normalized root mean squares error (NRMSE) and relative NRMSE in different data sets with various missing rates shows CGimpute outperforms other methods. Copyright © 2011 Elsevier Ltd. All rights reserved.

Spaceflight modulates gene expression in the whole blood of astronauts.

PubMed

Barrila, Jennifer; Ott, C Mark; LeBlanc, Carly; Mehta, Satish K; Crabbé, Aurélie; Stafford, Phillip; Pierson, Duane L; Nickerson, Cheryl A

2016-01-01

Astronauts are exposed to a unique combination of stressors during spaceflight, which leads to alterations in their physiology and potentially increases their susceptibility to disease, including infectious diseases. To evaluate the potential impact of the spaceflight environment on the regulation of molecular pathways mediating cellular stress responses, we performed a first-of-its-kind pilot study to assess spaceflight-related gene-expression changes in the whole blood of astronauts. Using an array comprised of 234 well-characterized stress-response genes, we profiled transcriptomic changes in six astronauts (four men and two women) from blood preserved before and immediately following the spaceflight. Differentially regulated transcripts included those important for DNA repair, oxidative stress, and protein folding/degradation, including HSP90AB1 , HSP27 , GPX1 , XRCC1 , BAG-1 , HHR23A , FAP48 , and C-FOS . No gender-specific differences or relationship to number of missions flown was observed. This study provides a first assessment of transcriptomic changes occurring in the whole blood of astronauts in response to spaceflight.

Spaceflight modulates gene expression in the whole blood of astronauts

PubMed Central

Barrila, Jennifer; Ott, C Mark; LeBlanc, Carly; Mehta, Satish K; Crabbé, Aurélie; Stafford, Phillip; Pierson, Duane L; Nickerson, Cheryl A

2016-01-01

Astronauts are exposed to a unique combination of stressors during spaceflight, which leads to alterations in their physiology and potentially increases their susceptibility to disease, including infectious diseases. To evaluate the potential impact of the spaceflight environment on the regulation of molecular pathways mediating cellular stress responses, we performed a first-of-its-kind pilot study to assess spaceflight-related gene-expression changes in the whole blood of astronauts. Using an array comprised of 234 well-characterized stress-response genes, we profiled transcriptomic changes in six astronauts (four men and two women) from blood preserved before and immediately following the spaceflight. Differentially regulated transcripts included those important for DNA repair, oxidative stress, and protein folding/degradation, including HSP90AB1, HSP27, GPX1, XRCC1, BAG-1, HHR23A, FAP48, and C-FOS. No gender-specific differences or relationship to number of missions flown was observed. This study provides a first assessment of transcriptomic changes occurring in the whole blood of astronauts in response to spaceflight. PMID:28725744

Comparative transcriptional profiling of two wheat genotypes, with contrasting levels of minerals in grains, shows expression differences during grain filling.

PubMed

Singh, Sudhir P; Jeet, Raja; Kumar, Jitendra; Shukla, Vishnu; Srivastava, Rakesh; Mantri, Shrikant S; Tuli, Rakesh

2014-01-01

Wheat is one of the most important cereal crops in the world. To identify the candidate genes for mineral accumulation, it is important to examine differential transcriptome between wheat genotypes, with contrasting levels of minerals in grains. A transcriptional comparison of developing grains was carried out between two wheat genotypes- Triticum aestivum Cv. WL711 (low grain mineral), and T. aestivum L. IITR26 (high grain mineral), using Affymetrix GeneChip Wheat Genome Array. The study identified a total of 580 probe sets as differentially expressed (with log2 fold change of ≥2 at p≤0.01) between the two genotypes, during grain filling. Transcripts with significant differences in induction or repression between the two genotypes included genes related to metal homeostasis, metal tolerance, lignin and flavonoid biosynthesis, amino acid and protein transport, vacuolar-sorting receptor, aquaporins, and stress responses. Meta-analysis revealed spatial and temporal signatures of a majority of the differentially regulated transcripts.

Comparative Transcriptional Profiling of Two Wheat Genotypes, with Contrasting Levels of Minerals in Grains, Shows Expression Differences during Grain Filling

PubMed Central

Singh, Sudhir P.; Jeet, Raja; Kumar, Jitendra; Shukla, Vishnu; Srivastava, Rakesh; Mantri, Shrikant S.; Tuli, Rakesh

2014-01-01

Wheat is one of the most important cereal crops in the world. To identify the candidate genes for mineral accumulation, it is important to examine differential transcriptome between wheat genotypes, with contrasting levels of minerals in grains. A transcriptional comparison of developing grains was carried out between two wheat genotypes- Triticum aestivum Cv. WL711 (low grain mineral), and T. aestivum L. IITR26 (high grain mineral), using Affymetrix GeneChip Wheat Genome Array. The study identified a total of 580 probe sets as differentially expressed (with log2 fold change of ≥2 at p≤0.01) between the two genotypes, during grain filling. Transcripts with significant differences in induction or repression between the two genotypes included genes related to metal homeostasis, metal tolerance, lignin and flavonoid biosynthesis, amino acid and protein transport, vacuolar-sorting receptor, aquaporins, and stress responses. Meta-analysis revealed spatial and temporal signatures of a majority of the differentially regulated transcripts. PMID:25364903

Experimental demonstration of a multi-wavelength distributed feedback semiconductor laser array with an equivalent chirped grating profile based on the equivalent chirp technology.

PubMed

Li, Wangzhe; Zhang, Xia; Yao, Jianping

2013-08-26

We report, to the best of our knowledge, the first realization of a multi-wavelength distributed feedback (DFB) semiconductor laser array with an equivalent chirped grating profile based on equivalent chirp technology. All the lasers in the laser array have an identical grating period with an equivalent chirped grating structure, which are realized by nonuniform sampling of the gratings. Different wavelengths are achieved by changing the sampling functions. A multi-wavelength DFB semiconductor laser array is fabricated and the lasing performance is evaluated. The results show that the equivalent chirp technology is an effective solution for monolithic integration of a multi-wavelength laser array with potential for large volume fabrication.

STUDIES OF NORMAL GENE EXPRESSION IN THE RAT NASAL EPITHELIUM USNG CDNA ARRAY TECHNOLOGY

EPA Science Inventory

Studies of Normal Gene Expression in the Rat Nasal Epithelium Using cDNA Array

The nasal epithelium is an important target site for chemically-induced toxicity and carcinogenicity .Gene expression data are being used increasingly for studies of such conditions. In or...

Perturbation of polyamine catabolism affects grape ripening of Vitis vinifera cv. Trincadeira.

PubMed

Agudelo-Romero, Patricia; Ali, Kashif; Choi, Young H; Sousa, Lisete; Verpoorte, Rob; Tiburcio, Antonio F; Fortes, Ana M

2014-01-01

Grapes are economically the most important fruit worldwide. However, the complexity of biological events that lead to ripening of nonclimacteric fruits is not fully understood, particularly the role of polyamines' catabolism. The transcriptional and metabolic profilings complemented with biochemical data were studied during ripening of Trincadeira grapes submitted to guazatine treatment, a potent inhibitor of polyamine oxidase activity. The mRNA expression profiles of one time point (EL 38) corresponding to harvest stage was compared between mock and guazatine treatments using Affymetrix GrapeGen(®) genome array. A total of 2113 probesets (1880 unigenes) were differentially expressed between these samples. Quantitative RT-PCR validated microarrays results being carried out for EL 35 (véraison berries), EL 36 (ripe berries) and EL 38 (harvest stage berries). Metabolic profiling using HPLC and (1)H NMR spectroscopy showed increase of putrescine, proline, threonine and 1-O-ethyl-β-glucoside in guazatine treated samples. Genes involved in amino acid, carbohydrate and water transport were down-regulated in guazatine treated samples suggesting that the strong dehydrated phenotype obtained in guazatine treated samples may be due to impaired transport mechanisms. Genes involved in terpenes' metabolism were differentially expressed between guazatine and mock treated samples. Altogether, results support an important role of polyamine catabolism in grape ripening namely in cell expansion and aroma development. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

Gene expression analysis of pancreatic cell lines reveals genes overexpressed in pancreatic cancer.

PubMed

Alldinger, Ingo; Dittert, Dag; Peiper, Matthias; Fusco, Alberto; Chiappetta, Gennaro; Staub, Eike; Lohr, Matthias; Jesnowski, Ralf; Baretton, Gustavo; Ockert, Detlef; Saeger, Hans-Detlev; Grützmann, Robert; Pilarsky, Christian

2005-01-01

Pancreatic cancer is one of the leading causes of cancer-related death. Using DNA gene expression analysis based on a custom made Affymetrix cancer array, we investigated the expression pattern of both primary and established pancreatic carcinoma cell lines. We analyzed the gene expression of 5 established pancreatic cancer cell lines (AsPC-1, BxPC-3, Capan-1, Capan-2 and HPAF II) and 5 primary isolates, 1 of them derived from benign pancreatic duct cells. Out of 1,540 genes which were expressed in at least 3 experiments, we found 122 genes upregulated and 18 downregulated in tumor cell lines compared to benign cells with a fold change >3. Several of the upregulated genes (like Prefoldin 5, ADAM9 and E-cadherin) have been associated with pancreatic cancer before. The other differentially regulated genes, however, play a so far unknown role in the course of human pancreatic carcinoma. By means of immunohistochemistry we could show that thymosin beta-10 (TMSB10), upregulated in tumor cell lines, is expressed in human pancreatic carcinoma, but not in non-neoplastic pancreatic tissue, suggesting a role for TMSB10 in the carcinogenesis of pancreatic carcinoma. Using gene expression profiling of pancreatic cell lines we were able to identify genes differentially expressed in pancreatic adenocarcinoma, which might contribute to pancreatic cancer development. Copyright 2005 S. Karger AG, Basel.

GeneBreak: detection of recurrent DNA copy number aberration-associated chromosomal breakpoints within genes.

PubMed

van den Broek, Evert; van Lieshout, Stef; Rausch, Christian; Ylstra, Bauke; van de Wiel, Mark A; Meijer, Gerrit A; Fijneman, Remond J A; Abeln, Sanne

2016-01-01

Development of cancer is driven by somatic alterations, including numerical and structural chromosomal aberrations. Currently, several computational methods are available and are widely applied to detect numerical copy number aberrations (CNAs) of chromosomal segments in tumor genomes. However, there is lack of computational methods that systematically detect structural chromosomal aberrations by virtue of the genomic location of CNA-associated chromosomal breaks and identify genes that appear non-randomly affected by chromosomal breakpoints across (large) series of tumor samples. 'GeneBreak' is developed to systematically identify genes recurrently affected by the genomic location of chromosomal CNA-associated breaks by a genome-wide approach, which can be applied to DNA copy number data obtained by array-Comparative Genomic Hybridization (CGH) or by (low-pass) whole genome sequencing (WGS). First, 'GeneBreak' collects the genomic locations of chromosomal CNA-associated breaks that were previously pinpointed by the segmentation algorithm that was applied to obtain CNA profiles. Next, a tailored annotation approach for breakpoint-to-gene mapping is implemented. Finally, dedicated cohort-based statistics is incorporated with correction for covariates that influence the probability to be a breakpoint gene. In addition, multiple testing correction is integrated to reveal recurrent breakpoint events. This easy-to-use algorithm, 'GeneBreak', is implemented in R ( www.cran.r-project.org ) and is available from Bioconductor ( www.bioconductor.org/packages/release/bioc/html/GeneBreak.html ).

Transcriptional modulation of some Staphylococcus aureus iron-regulated genes during growth in vitro and in a tissue cage model in vivo.

PubMed

Allard, Marianne; Moisan, Hélène; Brouillette, Eric; Gervais, Alain L; Jacques, Mario; Lacasse, Pierre; Diarra, Moussa S; Malouin, François

2006-06-01

Staphylococcus aureus can proliferate in iron-limited environments such as the mammalian host. The transcriptional profiles of 460 genes (iron-regulated, putative Fur-regulated, membrane transport, pathogenesis) obtained for S. aureus grown in iron-restricted environments in vitro and in vivo were compared in order to identify new iron-regulated genes and to evaluate their potential as possible therapeutic targets in vivo. Iron deprivation was created in vitro by 2,2-dipyridyl, and in vivo, S. aureus was grown in tissue cages implanted in mice. Bacterial RNA was obtained from each growth condition and cDNA probes were co-hybridized on DNA arrays. Thirty-six upregulated and 11 downregulated genes were commonly modulated in animals and in the low-iron medium. Real-time PCR confirmed the iron-dependent modulation of four novel genes (SACOL0161, 2170, 2369, 2431) with a Fur box motif. Some genes expressed in the dipyridyl medium were not expressed in vivo (e.g., copA, frpA, SACOL1045). Downregulated genes included an iron-storage protein gene and genes of the succinate dehydrogenase complex, reminiscent of a small RNA-dependent regulation thus far only demonstrated in Gram-negative bacteria. The expression of iron-regulated genes in distinct low-iron environments provided insight into their relative importance in vitro and in vivo and their usefulness for vaccine and drug development.

The Design and Characterization of Wideband Spline-profiled Feedhorns for Advanced Actpol

NASA Technical Reports Server (NTRS)

Simon, Sara M.; Austermann, Jason; Beall, James A.; Choi, Steve K.; Coughlin, Kevin P.; Duff, Shannon M.; Gallardo, Patricio A.; Henderson, Shawn W.; Hills, Felicity B.; Ho, Shuay-Pwu Patty;

Gene expression profiling in patients with polymyalgia rheumatica before and after symptom-abolishing glucocorticoid treatment.

PubMed

Kreiner, Frederik Flindt; Borup, Rehannah; Nielsen, Finn Cilius; Schjerling, Peter; Galbo, Henrik

2017-08-07

The pathophysiology, including the impact of gene expression, of polymyalgia rheumatica (PMR) remains elusive. We profiled the gene expression in muscle tissue in PMR patients before and after glucocorticoid treatment. Gene expression was measured using Affymetrix Human Genome U133 Plus 2.0 arrays in muscle biopsies from 8 glucocorticoid-naive patients with PMR and 10 controls before and after prednisolone-treatment for 14 days. For 14 genes, quantitative real-time PCR (qRT-PCR, n = 9 in both groups) was used to validate the microarray findings and to further investigate the expression of genes of particular interest. Prednisolone normalized erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) in PMR patients. A total of 165 putatively clinically relevant, differentially expressed genes were identified (cut-off: fold difference > ±1.2, difference of mean > 30, and p < 0.05); of these, 78 genes differed between patients and controls before treatment, 131 genes responded to treatment in a given direction only in patients, and 44 fulfilled both these criteria. In 43 of the 44 genes, treatment counteracted the initial difference. Functional clustering identified themes of biological function, including regulation of protein biosynthesis, and regulation of transcription and of extracellular matrix processes. Overall, qRT-PCR confirmed the microarray findings: Microarray-detected group differences were confirmed for 9 genes in 17 of 18 comparisons (same magnitude and direction of change); lack of group differences in microarray testing was confirmed for 5 genes in 8 of 10 comparisons. Before treatment, using qRT-PCR, expression of interleukin 6 (IL-6) was found to be 4-fold higher in patients (p < 0.05). This study identifies genes in muscle, the expression of which may impact the pathophysiology of PMR. Moreover, the study adds further evidence of the importance of IL-6 in the disease. Follow-up studies are needed to establish the exact pathophysiological relevance of the identified genes. The study was retrospectively listed on the ISRCTN registry with study ID ISRCTN69503018 and date of registration the 26th of July 2017.

CHESS (CgHExpreSS): a comprehensive analysis tool for the analysis of genomic alterations and their effects on the expression profile of the genome.

PubMed

Lee, Mikyung; Kim, Yangseok

2009-12-16

Genomic alterations frequently occur in many cancer patients and play important mechanistic roles in the pathogenesis of cancer. Furthermore, they can modify the expression level of genes due to altered copy number in the corresponding region of the chromosome. An accumulating body of evidence supports the possibility that strong genome-wide correlation exists between DNA content and gene expression. Therefore, more comprehensive analysis is needed to quantify the relationship between genomic alteration and gene expression. A well-designed bioinformatics tool is essential to perform this kind of integrative analysis. A few programs have already been introduced for integrative analysis. However, there are many limitations in their performance of comprehensive integrated analysis using published software because of limitations in implemented algorithms and visualization modules. To address this issue, we have implemented the Java-based program CHESS to allow integrative analysis of two experimental data sets: genomic alteration and genome-wide expression profile. CHESS is composed of a genomic alteration analysis module and an integrative analysis module. The genomic alteration analysis module detects genomic alteration by applying a threshold based method or SW-ARRAY algorithm and investigates whether the detected alteration is phenotype specific or not. On the other hand, the integrative analysis module measures the genomic alteration's influence on gene expression. It is divided into two separate parts. The first part calculates overall correlation between comparative genomic hybridization ratio and gene expression level by applying following three statistical methods: simple linear regression, Spearman rank correlation and Pearson's correlation. In the second part, CHESS detects the genes that are differentially expressed according to the genomic alteration pattern with three alternative statistical approaches: Student's t-test, Fisher's exact test and Chi square test. By successive operations of two modules, users can clarify how gene expression levels are affected by the phenotype specific genomic alterations. As CHESS was developed in both Java application and web environments, it can be run on a web browser or a local machine. It also supports all experimental platforms if a properly formatted text file is provided to include the chromosomal position of probes and their gene identifiers. CHESS is a user-friendly tool for investigating disease specific genomic alterations and quantitative relationships between those genomic alterations and genome-wide gene expression profiling.

Signaling events in pathogen-induced macrophage foam cell formation.

PubMed

Shaik-Dasthagirisaheb, Yazdani B; Mekasha, Samrawit; He, Xianbao; Gibson, Frank C; Ingalls, Robin R

2016-08-01

Macrophage foam cell formation is a key event in atherosclerosis. Several triggers induce low-density lipoprotein (LDL) uptake by macrophages to create foam cells, including infections with Porphyromonas gingivalis and Chlamydia pneumoniae, two pathogens that have been linked to atherosclerosis. While gene regulation during foam cell formation has been examined, comparative investigations to identify shared and specific pathogen-elicited molecular events relevant to foam cell formation are not well documented. We infected mouse bone marrow-derived macrophages with P. gingivalis or C. pneumoniae in the presence of LDL to induce foam cell formation, and examined gene expression using an atherosclerosis pathway targeted plate array. We found over 30 genes were significantly induced in response to both pathogens, including PPAR family members that are broadly important in atherosclerosis and matrix remodeling genes that may play a role in plaque development and stability. Six genes mainly involved in lipid transport were significantly downregulated. The response overall was remarkably similar and few genes were regulated in a pathogen-specific manner. Despite very divergent lifestyles, P. gingivalis and C. pneumoniae activate similar gene expression profiles during foam cell formation that may ultimately serve as targets for modulating infection-elicited foam cell burden, and progression of atherosclerosis. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Aquaculture changes the profile of antibiotic resistance and mobile genetic element associated genes in Baltic Sea sediments.

PubMed

Muziasari, Windi I; Pärnänen, Katariina; Johnson, Timothy A; Lyra, Christina; Karkman, Antti; Stedtfeld, Robert D; Tamminen, Manu; Tiedje, James M; Virta, Marko

2016-04-01

Antibiotics are commonly used in aquaculture and they can change the environmental resistome by increasing antibiotic resistance genes (ARGs). Sediment samples were collected from two fish farms located in the Northern Baltic Sea, Finland, and from a site outside the farms (control). The sediment resistome was assessed by using a highly parallel qPCR array containing 295 primer sets to detect ARGs, mobile genetic elements and the 16S rRNA gene. The fish farm resistomes were enriched in transposon and integron associated genes and in ARGs encoding resistance to antibiotics which had been used to treat fish at the farms. Aminoglycoside resistance genes were also enriched in the farm sediments despite the farms not having used aminoglycosides. In contrast, the total relative abundance values of ARGs were higher in the control sediment resistome and they were mainly genes encoding efflux pumps followed by beta-lactam resistance genes, which are found intrinsically in many bacteria. This suggests that there is a natural Baltic sediment resistome. The resistome associated with fish farms can be from native ARGs enriched by antibiotic use at the farms and/or from ARGs and mobile elements that have been introduced by fish farming. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

The 'PUCE CAFE' Project: the First 15K Coffee Microarray, a New Tool for Discovering Candidate Genes correlated to Agronomic and Quality Traits

PubMed Central

2011-01-01

Background Understanding the genetic elements that contribute to key aspects of coffee biology will have an impact on future agronomical improvements for this economically important tree. During the past years, EST collections were generated in Coffee, opening the possibility to create new tools for functional genomics. Results The "PUCE CAFE" Project, organized by the scientific consortium NESTLE/IRD/CIRAD, has developed an oligo-based microarray using 15,721 unigenes derived from published coffee EST sequences mostly obtained from different stages of fruit development and leaves in Coffea Canephora (Robusta). Hybridizations for two independent experiments served to compare global gene expression profiles in three types of tissue matter (mature beans, leaves and flowers) in C. canephora as well as in the leaves of three different coffee species (C. canephora, C. eugenoides and C. arabica). Microarray construction, statistical analyses and validation by Q-PCR analysis are presented in this study. Conclusion We have generated the first 15 K coffee array during this PUCE CAFE project, granted by Génoplante (the French consortium for plant genomics). This new tool will help study functional genomics in a wide range of experiments on various plant tissues, such as analyzing bean maturation or resistance to pathogens or drought. Furthermore, the use of this array has proven to be valid in different coffee species (diploid or tetraploid), drastically enlarging its impact for high-throughput gene expression in the community of coffee research. PMID:21208403

Nutrition metabolism plays an important role in the alternate bearing of the olive tree (Olea europaea L.).

PubMed

Turktas, Mine; Inal, Behcet; Okay, Sezer; Erkilic, Emine Gulden; Dundar, Ekrem; Hernandez, Pilar; Dorado, Gabriel; Unver, Turgay

2013-01-01

The olive tree (Olea europaea L.) is widely known for its strong tendency for alternate bearing, which severely affects the fruit yield from year to year. Microarray based gene expression analysis using RNA from olive samples (on-off years leaves and ripe-unripe fruits) are particularly useful to understand the molecular mechanisms influencing the periodicity in the olive tree. Thus, we carried out genome wide transcriptome analyses involving different organs and temporal stages of the olive tree using the NimbleGen Array containing 136,628 oligonucleotide probe sets. Cluster analyses of the genes showed that cDNAs originated from different organs could be sorted into separate groups. The nutritional control had a particularly remarkable impact on the alternate bearing of olive, as shown by the differential expression of transcripts under different temporal phases and organs. Additionally, hormonal control and flowering processes also played important roles in this phenomenon. Our analyses provide further insights into the transcript changes between "on year" and "off year" leaves along with the changes from unrpipe to ripe fruits, which shed light on the molecular mechanisms underlying the olive tree alternate bearing. These findings have important implications for the breeding and agriculture of the olive tree and other crops showing periodicity. To our knowledge, this is the first study reporting the development and use of an olive array to document the gene expression profiling associated with the alternate bearing in olive tree.

The 'PUCE CAFE' Project: the first 15K coffee microarray, a new tool for discovering candidate genes correlated to agronomic and quality traits.

PubMed

Privat, Isabelle; Bardil, Amélie; Gomez, Aureliano Bombarely; Severac, Dany; Dantec, Christelle; Fuentes, Ivanna; Mueller, Lukas; Joët, Thierry; Pot, David; Foucrier, Séverine; Dussert, Stéphane; Leroy, Thierry; Journot, Laurent; de Kochko, Alexandre; Campa, Claudine; Combes, Marie-Christine; Lashermes, Philippe; Bertrand, Benoit

2011-01-05

Understanding the genetic elements that contribute to key aspects of coffee biology will have an impact on future agronomical improvements for this economically important tree. During the past years, EST collections were generated in Coffee, opening the possibility to create new tools for functional genomics. The "PUCE CAFE" Project, organized by the scientific consortium NESTLE/IRD/CIRAD, has developed an oligo-based microarray using 15,721 unigenes derived from published coffee EST sequences mostly obtained from different stages of fruit development and leaves in Coffea Canephora (Robusta). Hybridizations for two independent experiments served to compare global gene expression profiles in three types of tissue matter (mature beans, leaves and flowers) in C. canephora as well as in the leaves of three different coffee species (C. canephora, C. eugenoides and C. arabica). Microarray construction, statistical analyses and validation by Q-PCR analysis are presented in this study. We have generated the first 15 K coffee array during this PUCE CAFE project, granted by Génoplante (the French consortium for plant genomics). This new tool will help study functional genomics in a wide range of experiments on various plant tissues, such as analyzing bean maturation or resistance to pathogens or drought. Furthermore, the use of this array has proven to be valid in different coffee species (diploid or tetraploid), drastically enlarging its impact for high-throughput gene expression in the community of coffee research.

Nutrition Metabolism Plays an Important Role in the Alternate Bearing of the Olive Tree (Olea europaea L.)

PubMed Central

Turktas, Mine; Inal, Behcet; Okay, Sezer; Erkilic, Emine Gulden; Dundar, Ekrem; Hernandez, Pilar; Dorado, Gabriel; Unver, Turgay

2013-01-01

The olive tree (Olea europaea L.) is widely known for its strong tendency for alternate bearing, which severely affects the fruit yield from year to year. Microarray based gene expression analysis using RNA from olive samples (on-off years leaves and ripe-unripe fruits) are particularly useful to understand the molecular mechanisms influencing the periodicity in the olive tree. Thus, we carried out genome wide transcriptome analyses involving different organs and temporal stages of the olive tree using the NimbleGen Array containing 136,628 oligonucleotide probe sets. Cluster analyses of the genes showed that cDNAs originated from different organs could be sorted into separate groups. The nutritional control had a particularly remarkable impact on the alternate bearing of olive, as shown by the differential expression of transcripts under different temporal phases and organs. Additionally, hormonal control and flowering processes also played important roles in this phenomenon. Our analyses provide further insights into the transcript changes between ”on year” and “off year” leaves along with the changes from unrpipe to ripe fruits, which shed light on the molecular mechanisms underlying the olive tree alternate bearing. These findings have important implications for the breeding and agriculture of the olive tree and other crops showing periodicity. To our knowledge, this is the first study reporting the development and use of an olive array to document the gene expression profiling associated with the alternate bearing in olive tree. PMID:23555820

Sustained endocrine profiles of a girl with WAGR syndrome.

PubMed

Takada, Yui; Sakai, Yasunari; Matsushita, Yuki; Ohkubo, Kazuhiro; Koga, Yuhki; Akamine, Satoshi; Torio, Michiko; Ishizaki, Yoshito; Sanefuji, Masafumi; Torisu, Hiroyuki; Shaw, Chad A; Kagami, Masayo; Hara, Toshiro; Ohga, Shouichi

2017-10-23

Wilms tumor, aniridia, genitourinary anomalies and mental retardation (WAGR) syndrome is a rare genetic disorder caused by heterozygous deletions of WT1 and PAX6 at chromosome 11p13. Deletion of BDNF is known eto be associated with hyperphagia and obesity in both humans and animal models; however, neuroendocrine and epigenetic profiles of individuals with WAGR syndrome remain to be determined. We report a 5-year-old girl with the typical phenotype of WAGR syndrome. She showed profound delays in physical growth, motor and cognitive development without signs of obesity. Array comparative genome hybridization (CGH) revealed that she carried a 14.4 Mb deletion at 11p14.3p12, encompassing the WT1, PAX6 and BDNF genes. She experienced recurrent hypoglycemic episodes at 5 years of age. Insulin tolerance and hormonal loading tests showed normal hypothalamic responses to the hypoglycemic condition and other stimulations. Methylation analysis for freshly prepared DNA from peripheral lymphocytes using the pyro-sequencing-based system showed normal patterns of methylation at known imprinting control regions. Children with WAGR syndrome may manifest profound delay in postnatal growth through unknown mechanisms. Epigenetic factors and growth-associated genes in WAGR syndrome remain to be characterized.

Transcriptome profiling of sleeping, waking, and sleep deprived adult heterozygous Aldh1L1 - eGFP-L10a mice.

PubMed

Bellesi, Michele; de Vivo, Luisa; Tononi, Giulio; Cirelli, Chiara

2015-12-01

Transcriptomic studies revealed that hundreds of mRNAs show differential expression in the brains of sleeping relative to awake rats, mice, flies, and sparrows. Although these results have offered clues regarding the molecular consequences of sleep and sleep loss, their functional significance thus far has been limited. This is probably because the previous studies pooled transcripts from all brain cells, including neurons and glia. In Bellesi et al. 2015 [1], we used the translating ribosome affinity purification technology (TRAP) and microarray analysis to obtain a genome-wide mRNA profiling of astrocytes as a function of sleep and wake. We used bacterial artificial chromosome (BAC) transgenic mice expressing eGFP tagged ribosomal protein L10a under the promoter of the Aldh1L1 gene, a highly expressed astrocytic gene. Using this approach, we could extract only the astrocytic mRNAs, and only those already committed to be translated into proteins (L10a is part of the translational machinery). Here, we report a detailed description of the protocol used in the study [1]. Array data have been submitted to NCBI GEO under accession number (GSE69079).

Correlating cellular and molecular signatures of mucosal immunity that distinguish HIV controllers from noncontrollers.

PubMed

Loke, P'ng; Favre, David; Hunt, Peter W; Leung, Jacqueline M; Kanwar, Bittoo; Martin, Jeffrey N; Deeks, Steven G; McCune, Joseph M

2010-04-15

HIV "controllers" are persons infected with human immunodeficiency virus, type I (HIV) who maintain long-term control of viremia without antiviral therapy and who usually do not develop the acquired immune deficiency syndrome (AIDS). In this study, we have correlated results from polychromatic flow cytometry and oligonucleotide expression arrays to characterize the mucosal immune responses of these subjects in relation to untreated HIV(+) persons with high viral loads and progressive disease ("noncontrollers"). Paired peripheral blood and rectosigmoid biopsies were analyzed from 9 controllers and 11 noncontrollers. Several cellular immune parameters were found to be concordant between the 2 compartments. Compared with noncontrollers, the mucosal tissues of controllers had similar levels of effector T cells and fewer regulatory T cells (Tregs). Using principal component analysis to correlate immunologic parameters with gene expression profiles, transcripts were identified that accurately distinguished between controllers and noncontrollers. Direct 2-way comparison also revealed genes that are significantly different in their expression between controllers and noncontrollers, all of which had reduced expression in controllers. In addition to providing an approach that integrates flow cytometry datasets with transcriptional profiling analysis, these results underscore the importance of the sustained inflammatory response that attends progressive HIV disease.

Integrated analysis of epigenomic and genomic changes by DNA methylation dependent mechanisms provides potential novel biomarkers for prostate cancer.

PubMed

White-Al Habeeb, Nicole M A; Ho, Linh T; Olkhov-Mitsel, Ekaterina; Kron, Ken; Pethe, Vaijayanti; Lehman, Melanie; Jovanovic, Lidija; Fleshner, Neil; van der Kwast, Theodorus; Nelson, Colleen C; Bapat, Bharati

2014-09-15

Epigenetic silencing mediated by CpG methylation is a common feature of many cancers. Characterizing aberrant DNA methylation changes associated with tumor progression may identify potential prognostic markers for prostate cancer (PCa). We treated two PCa cell lines, 22Rv1 and DU-145 with the demethylating agent 5-Aza 2'-deoxycitidine (DAC) and global methylation status was analyzed by performing methylation-sensitive restriction enzyme based differential methylation hybridization strategy followed by genome-wide CpG methylation array profiling. In addition, we examined gene expression changes using a custom microarray. Gene Set Enrichment Analysis (GSEA) identified the most significantly dysregulated pathways. In addition, we assessed methylation status of candidate genes that showed reduced CpG methylation and increased gene expression after DAC treatment, in Gleason score (GS) 8 vs. GS6 patients using three independent cohorts of patients; the publically available The Cancer Genome Atlas (TCGA) dataset, and two separate patient cohorts. Our analysis, by integrating methylation and gene expression in PCa cell lines, combined with patient tumor data, identified novel potential biomarkers for PCa patients. These markers may help elucidate the pathogenesis of PCa and represent potential prognostic markers for PCa patients.

Comparison of modeled and experimental PV array temperature profiles for accurate interpretation of module performance and degradation

NASA Astrophysics Data System (ADS)

Elwood, Teri; Simmons-Potter, Kelly

2017-08-01

Quantification of the effect of temperature on photovoltaic (PV) module efficiency is vital to the correct interpretation of PV module performance under varied environmental conditions. However, previous work has demonstrated that PV module arrays in the field are subject to significant location-based temperature variations associated with, for example, local heating/cooling and array edge effects. Such thermal non-uniformity can potentially lead to under-prediction or over-prediction of PV array performance due to an incorrect interpretation of individual module temperature de-rating. In the current work, a simulated method for modeling the thermal profile of an extended PV array has been investigated through extensive computational modeling utilizing ANSYS, a high-performance computational fluid dynamics (CFD) software tool. Using the local wind speed as an input, simulations were run to determine the velocity at particular points along modular strings corresponding to the locations of temperature sensors along strings in the field. The point velocities were utilized along with laminar flow theories in order to calculate Nusselt's number for each point. These calculations produced a heat flux profile which, when combined with local thermal and solar radiation profiles, were used as inputs in an ANSYS Thermal Transient model that generated a solar string operating temperature profile. A comparison of the data collected during field testing, and the data fabricated by ANSYS simulations, will be discussed in order to authenticate the accuracy of the model.

Identification and analysis of integrons and cassette arrays in bacterial genomes

PubMed Central

Touchon, Marie; Néron, Bertrand; Rocha, Eduardo PC

2016-01-01

Abstract Integrons recombine gene arrays and favor the spread of antibiotic resistance. Their broader roles in bacterial adaptation remain mysterious, partly due to lack of computational tools. We made a program – IntegronFinder – to identify integrons with high accuracy and sensitivity. IntegronFinder is available as a standalone program and as a web application. It searches for attC sites using covariance models, for integron-integrases using HMM profiles, and for other features (promoters, attI site) using pattern matching. We searched for integrons, integron-integrases lacking attC sites, and clusters of attC sites lacking a neighboring integron-integrase in bacterial genomes. All these elements are especially frequent in genomes of intermediate size. They are missing in some key phyla, such as α-Proteobacteria, which might reflect selection against cell lineages that acquire integrons. The similarity between attC sites is proportional to the number of cassettes in the integron, and is particularly low in clusters of attC sites lacking integron-integrases. The latter are unexpectedly abundant in genomes lacking integron-integrases or their remains, and have a large novel pool of cassettes lacking homologs in the databases. They might represent an evolutionary step between the acquisition of genes within integrons and their stabilization in the new genome. PMID:27130947

Modulation of gene expression and cell-cycle signaling pathways by the EGFR inhibitor gefitinib (Iressa) in rat urinary bladder cancer.

PubMed

Lu, Yan; Liu, Pengyuan; Van den Bergh, Francoise; Zellmer, Victoria; James, Michael; Wen, Weidong; Grubbs, Clinton J; Lubet, Ronald A; You, Ming

2012-02-01

The epidermal growth factor receptor inhibitor Iressa has shown strong preventive efficacy in the N-butyl-N-(4-hydroxybutyl)-nitrosamine (OH-BBN) model of bladder cancer in the rat. To explore its antitumor mechanism, we implemented a systems biology approach to characterize gene expression and signaling pathways in rat urinary bladder cancers treated with Iressa. Eleven bladder tumors from control rats, seven tumors from rats treated with Iressa, and seven normal bladder epithelia were profiled by the Affymetrix Rat Exon 1.0 ST Arrays. We identified 713 downregulated and 641 upregulated genes in comparing bladder tumors versus normal bladder epithelia. In addition, 178 genes were downregulated and 96 genes were upregulated when comparing control tumors versus Iressa-treated tumors. Two coexpression modules that were significantly correlated with tumor status and treatment status were identified [r = 0.70, P = 2.80 × 10(-15) (bladder tumor vs. normal bladder epithelium) and r = 0.63, P = 2.00 × 10(-42) (Iressa-treated tumor vs. control tumor), respectively]. Both tumor module and treatment module were enriched for genes involved in cell-cycle processes. Twenty-four and twenty-one highly connected hub genes likely to be key drivers in cell cycle were identified in the tumor module and treatment module, respectively. Analysis of microRNA genes on the array chips showed that tumor module and treatment module were significantly associated with expression levels of let-7c (r = 0.54, P = 3.70 × 10(-8) and r = 0.73, P = 1.50 × 10(-65), respectively). These results suggest that let-7c downregulation and its regulated cell-cycle pathway may play an integral role in governing bladder tumor suppression or collaborative oncogenesis and that Iressa exhibits its preventive efficacy on bladder tumorigenesis by upregulating let-7 and inhibiting the cell cycle. Cell culture study confirmed that the increased expression of let-7c decreases Iressa-treated bladder tumor cell growth. The identified hub genes may also serve as pharmacodynamic or efficacy biomarkers in clinical trials of chemoprevention in human bladder cancer. ©2011 AACR.

Field testing of a convergent array of acoustic Doppler profilers for high-resolution velocimetry in energetic tidal currents

DOE Office of Scientific and Technical Information (OSTI.GOV)

Harding, Samuel F.; Sellar, Brian; Richmond, Marshall C.

An array of single-beam acoustic Doppler profilers has been developed for the high resolution measurement of three-dimensional tidal flow velocities and subsequently tested in an energetic tidal site. This configuration has been developed to increase spatial resolution of velocity measurements in comparison to conventional acoustic Doppler profilers (ADPs) which characteristically use divergent acoustic beams emanating from a single instrument. This is achieved using geometrically convergent acoustic beams creating a sample volume at the focal point of 0.03 m3. Away from the focal point, the array is also able to simultaneously reconstruct three-dimensional velocity components in a profile throughout the watermore » column, and is referred to herein as a convergent-beam acoustic Doppler profiler (C-ADP). Mid-depth profiling is achieved through integration of the sensor platform with the operational commercial-scale Alstom 1MW DeepGen-IV Tidal Turbine deployed at the European Marine Energy Center, Orkney Isles, UK. This proof-of-concept paper outlines the C-ADP system configuration and comparison to measurements provided by co-installed reference instrumentation.« less

GENE ARRAY ANALYSIS OF THE VENTRAL PROSTATE IN RATS EXPOSED TO EITHER VINCLOZOLIN OR PROCYMIDONE

EPA Science Inventory

GENE ARRAY ANALYSIS OF THE VENTRAL PROSTATE IN RATS EXPOSED TO EITHER VINCLOZOLIN OR PROCYMIDONE. MB Rosen, VS Wilson, JE Schmid, and LE Gray Jr. US EPA, ORD, NHEERL, RTP, NC.

Vinclozolin (Vi) and procymidone (Pr) are antiandrogenic fungicides. While changes in gene expr...

Array data extractor (ADE): a LabVIEW program to extract and merge gene array data.

PubMed

Kurtenbach, Stefan; Kurtenbach, Sarah; Zoidl, Georg

2013-12-01

Large data sets from gene expression array studies are publicly available offering information highly valuable for research across many disciplines ranging from fundamental to clinical research. Highly advanced bioinformatics tools have been made available to researchers, but a demand for user-friendly software allowing researchers to quickly extract expression information for multiple genes from multiple studies persists. Here, we present a user-friendly LabVIEW program to automatically extract gene expression data for a list of genes from multiple normalized microarray datasets. Functionality was tested for 288 class A G protein-coupled receptors (GPCRs) and expression data from 12 studies comparing normal and diseased human hearts. Results confirmed known regulation of a beta 1 adrenergic receptor and further indicate novel research targets. Although existing software allows for complex data analyses, the LabVIEW based program presented here, "Array Data Extractor (ADE)", provides users with a tool to retrieve meaningful information from multiple normalized gene expression datasets in a fast and easy way. Further, the graphical programming language used in LabVIEW allows applying changes to the program without the need of advanced programming knowledge.

Expression profiling suggests a regulatory role of gallbladder in lipid homeostasis

PubMed Central

Yuan, Zuo-Biao; Han, Tian-Quan; Jiang, Zhao-Yan; Fei, Jian; Zhang, Yi; Qin, Jian; Tian, Zhi-Jie; Shang, Jun; Jiang, Zhi-Hong; Cai, Xing-Xing; Jiang, Yu; Zhang, Sheng-Dao; Jin, Gang

2005-01-01

AIM: To examine expression profile of gallbladder using microarray and to investigate the role of gallbladder in lipid homeostasis. METHODS: 33P-labelled cDNA derived from total RNA of gallbladder tissue was hybridized to a cDNA array representing 17000 cDNA clusters. Genes with intensities ≥2 and variation <0.33 between two samples were considered as positive signals with subtraction of background chosen from an area where no cDNA was spotted. The average gray level of two gallbladders was adopted to analyze its bioinformatics. Identified target genes were confirmed by touch-down polymerase chain reaction and sequencing. RESULTS: A total of 11 047 genes expressed in normal gallbladder, which was more than that predicted by another author, and the first 10 genes highly expressed (high gray level in hybridization image), e.g., ARPC5 (2225.88±90.46), LOC55972 (2220.32±446.51) and SLC20A2 (1865.21±98.02), were related to the function of smooth muscle contraction and material transport. Meanwhile, 149 lipid-related genes were expressed in the gallbladder, 89 of which were first identified (with gray level in hybridization image), e.g., FASN (11.42±2.62), APOD (92.61±8.90) and CYP21A2 (246.11±42.36), and they were involved in each step of lipid metabolism pathway. In addition, 19 of those 149 genes were gallstone candidate susceptibility genes (with gray level in hybridization image), e.g., HMGCR (10.98±0.31), NPC1 (34.88±12.12) and NR1H4 (16.8±0.65), which were previously thought to be expressed in the liver and/or intestine tissue only. CONCLUSION: Gallbladder expresses 11 047 genes and takes part in lipid homeostasis. PMID:15810076

Transcript Profiling Identifies Dynamic Gene Expression Patterns and an Important Role for Nrf2/Keap1 Pathway in the Developing Mouse Esophagus

PubMed Central

Li, Haiyan; Hu, Yuhui; Tevebaugh, Whitney; Yamamoto, Masayuki; Que, Jianwen; Chen, Xiaoxin

2012-01-01

Background and Aims Morphological changes during human and mouse esophageal development have been well characterized. However, changes at the molecular level in the course of esophageal morphogenesis remain unclear. This study aims to globally profile critical genes and signaling pathways during the development of mouse esophagus. By using microarray analysis this study also aims to determine how the Nrf2/Keap1 pathway regulates the morphogenesis of the esophageal epithelium. Methods Gene expression microarrays were used to survey gene expression in the esophagus at three critical phases: specification, metaplasia and maturation. The esophagi were isolated from wild-type, Nrf2−/−, Keap1−/−, or Nrf2−/−Keap1−/− embryos or young adult mice. Array data were statistically analyzed for differentially expressed genes and pathways. Histochemical and immunohistochemical staining were used to verify potential involvement of the Wnt pathway, Pparβ/δ and the PI3K/Akt pathway in the development of esophageal epithelium. Results Dynamic gene expression patterns accompanied the morphological changes of the developing esophagus at critical phases. Particularly, the Nrf2/Keap1 pathway had a baseline activity in the metaplasia phase and was further activated in the maturation phase. The Wnt pathway was active early and became inactive later in the metaplasia phase. In addition, Keap1−/− mice showed increased expression of Nrf2 downstream targets and genes involved in keratinization. Microarray and immunostaining data also suggested that esophageal hyperkeratosis in the Keap1−/− mice was due to activation of Pparβ/δ and the PI3K/Akt pathway. Conclusions Morphological changes of the esophageal epithelium are associated with dynamic changes in gene expression. Nrf2/Keap1 pathway activity is required for maturation of mouse esophageal epithelium. PMID:22567161

Molecular Characterization of Acquired Tolerance of Tumor Cells to Picropodophyllin (PPP)

PubMed Central

Hashemi, Jamileh; Worrall, Claire; Vasilcanu, Daiana; Fryknäs, Mårten; Sulaiman, Luqman; Karimi, Mohsen; Weng, Wen-Hui; Lui, Weng-Onn; Rudduck, Christina; Axelson, Magnus; Jernberg-Wiklund, Helena; Girnita, Leonard; Larsson, Olle; Larsson, Catharina

2011-01-01

Background Picropodophyllin (PPP) is a promising novel anti-neoplastic agent that efficiently kills tumor cells in vitro and causes tumor regression and increased survival in vivo. We have previously reported that PPP treatment induced moderate tolerance in two out of 10 cell lines only, and here report the acquired genomic and expression alterations associated with PPP selection over 1.5 years of treatment. Methodology/Principal Findings Copy number alterations monitored using metaphase and array-based comparative genomic hybridization analyses revealed largely overlapping alterations in parental and maximally tolerant cells. Gain/ amplification of the MYC and PVT1 loci in 8q24.21 were verified on the chromosome level. Abnormalities observed in connection to PPP treatment included regular gains and losses, as well as homozygous losses in 10q24.1-q24.2 and 12p12.3-p13.2 in one of the lines and amplification at 5q11.2 in the other. Abnormalities observed in both tolerant derivatives include amplification/gain of 5q11.2, gain of 11q12.1-q14.3 and gain of 13q33.3-qter. Using Nexus software analysis we combined the array-CGH data with data from gene expression profilings and identified genes that were altered in both inputs. A subset of genes identified as downregulated (ALDH1A3, ANXA1, TLR4 and RAB5A) or upregulated (COX6A1, NFIX, ME1, MAPK and TAP2) were validated by siRNA in the tolerant or parental cells to alter sensitivity to PPP and confirmed to alter sensitivity to PPP in further cell lines. Conclusions Long-term PPP selection lead to altered gene expression in PPP tolerant cells with increase as well as decrease of genes involved in cell death such as PTEN and BCL2. In addition, acquired genomic copy number alterations were observed that were often reflected by altered mRNA expression levels for genes in the same regions. PMID:21423728

Molecular characterization of acquired tolerance of tumor cells to picropodophyllin (PPP).

PubMed

Hashemi, Jamileh; Worrall, Claire; Vasilcanu, Daiana; Fryknäs, Mårten; Sulaiman, Luqman; Karimi, Mohsen; Weng, Wen-Hui; Lui, Weng-Onn; Rudduck, Christina; Axelson, Magnus; Jernberg-Wiklund, Helena; Girnita, Leonard; Larsson, Olle; Larsson, Catharina

2011-03-14

Picropodophyllin (PPP) is a promising novel anti-neoplastic agent that efficiently kills tumor cells in vitro and causes tumor regression and increased survival in vivo. We have previously reported that PPP treatment induced moderate tolerance in two out of 10 cell lines only, and here report the acquired genomic and expression alterations associated with PPP selection over 1.5 years of treatment. Copy number alterations monitored using metaphase and array-based comparative genomic hybridization analyses revealed largely overlapping alterations in parental and maximally tolerant cells. Gain/amplification of the MYC and PVT1 loci in 8q24.21 were verified on the chromosome level. Abnormalities observed in connection to PPP treatment included regular gains and losses, as well as homozygous losses in 10q24.1-q24.2 and 12p12.3-p13.2 in one of the lines and amplification at 5q11.2 in the other. Abnormalities observed in both tolerant derivatives include amplification/gain of 5q11.2, gain of 11q12.1-q14.3 and gain of 13q33.3-qter. Using Nexus software analysis we combined the array-CGH data with data from gene expression profilings and identified genes that were altered in both inputs. A subset of genes identified as downregulated (ALDH1A3, ANXA1, TLR4 and RAB5A) or upregulated (COX6A1, NFIX, ME1, MAPK and TAP2) were validated by siRNA in the tolerant or parental cells to alter sensitivity to PPP and confirmed to alter sensitivity to PPP in further cell lines. Long-term PPP selection lead to altered gene expression in PPP tolerant cells with increase as well as decrease of genes involved in cell death such as PTEN and BCL2. In addition, acquired genomic copy number alterations were observed that were often reflected by altered mRNA expression levels for genes in the same regions.

Underwater Application of Quantitative PCR on an Ocean Mooring

PubMed Central

Preston, Christina M.; Harris, Adeline; Ryan, John P.; Roman, Brent; Marin, Roman; Jensen, Scott; Everlove, Cheri; Birch, James; Dzenitis, John M.; Pargett, Douglas; Adachi, Masao; Turk, Kendra; Zehr, Jonathon P.; Scholin, Christopher A.

2011-01-01

The Environmental Sample Processor (ESP) is a device that allows for the underwater, autonomous application of DNA and protein probe array technologies as a means to remotely identify and quantify, in situ, marine microorganisms and substances they produce. Here, we added functionality to the ESP through the development and incorporation of a module capable of solid-phase nucleic acid extraction and quantitative PCR (qPCR). Samples collected by the instrument were homogenized in a chaotropic buffer compatible with direct detection of ribosomal RNA (rRNA) and nucleic acid purification. From a single sample, both an rRNA community profile and select gene abundances were ascertained. To illustrate this functionality, we focused on bacterioplankton commonly found along the central coast of California and that are known to vary in accordance with different oceanic conditions. DNA probe arrays targeting rRNA revealed the presence of 16S rRNA indicative of marine crenarchaea, SAR11 and marine cyanobacteria; in parallel, qPCR was used to detect 16S rRNA genes from the former two groups and the large subunit RuBisCo gene (rbcL) from Synecchococcus. The PCR-enabled ESP was deployed on a coastal mooring in Monterey Bay for 28 days during the spring-summer upwelling season. The distributions of the targeted bacterioplankon groups were as expected, with the exception of an increase in abundance of marine crenarchaea in anomalous nitrate-rich, low-salinity waters. The unexpected co-occurrence demonstrated the utility of the ESP in detecting novel events relative to previously described distributions of particular bacterioplankton groups. The ESP can easily be configured to detect and enumerate genes and gene products from a wide range of organisms. This study demonstrated for the first time that gene abundances could be assessed autonomously, underwater in near real-time and referenced against prevailing chemical, physical and bulk biological conditions. PMID:21829630

Wideband Array for C, X, and Ku-Band Applications with 5.3:1 Bandwidth

NASA Technical Reports Server (NTRS)

Novak, Markus H.; Volakis, John L.; Miranda, Felix A.

2015-01-01

Planar arrays that exploit strong intentional coupling between elements have allowed for very wide bandwidths in low-profile configurations. However, such designs also require complex impedance matching networks that must also be very compact. For many space applications, typically occurring at C-, X-, Ku-, and most recently at Ka-band, such designs require specialized and expensive fabrication techniques. To address this issue, a novel ultra-wideband array is presented, using a simplified feed network to reduce fabrication cost. The array operates from 3.5-18.5 GHz with VSWR less than 2.4 at broadside, and is of very low profile, having a total height of lambda/10 at the lowest frequency of operation. Validation is provided using a 64-element prototype array, fabricated using common Printed Circuit Board (PCB) technology. The low size, weight, and cost of this array make it attractive for space-borne applications.

Evaluation of the X-Linked High-Grade Myopia Locus (MYP1) with Cone Dysfunction and Color Vision Deficiencies

PubMed Central

Metlapally, Ravikanth; Michaelides, Michel; Bulusu, Anuradha; Li, Yi-Ju; Schwartz, Marianne; Rosenberg, Thomas; Hunt, David M.; Moore, Anthony T.; Züchner, Stephan; Rickman, Catherine Bowes; Young, Terri L.

2014-01-01

Purpose X-linked high myopia with mild cone dysfunction and color vision defects has been mapped to chromosome Xq28 (MYP1 locus). CXorf2/TEX28 is a nested, intercalated gene within the red-green opsin cone pigment gene tandem array on Xq28. The authors investigated whether TEX28 gene alterations were associated with the Xq28-linked myopia phenotype. Genomic DNA from five pedigrees (with high myopia and either protanopia or deuteranopia) that mapped to Xq28 were screened for TEX28 copy number variations (CNVs) and sequence variants. Methods To examine for CNVs, ultra-high resolution array-comparative genomic hybridization (array-CGH) assays were performed comparing the subject genomic DNA with control samples (two pairs from two pedigrees). Opsin or TEX28 gene-targeted quantitative real-time gene expression assays (comparative CT method) were performed to validate the array-CGH findings. All exons of TEX28, including intron/exon boundaries, were amplified and sequenced using standard techniques. Results Array-CGH findings revealed predicted duplications in affected patient samples. Although only three copies of TEX28 were previously reported within the opsin array, quantitative real-time analysis of the TEX28 targeted assay of affected male or carrier female individuals in these pedigrees revealed either fewer (one) or more (four or five) copies than did related and control unaffected individuals. Sequence analysis of TEX28 did not reveal any variants associated with the disease status. Conclusions CNVs have been proposed to play a role in disease inheritance and susceptibility as they affect gene dosage. TEX28 gene CNVs appear to be associated with the MYP1 X-linked myopia phenotypes. PMID:19098318

Golden Gate Assembly of CRISPR gRNA expression array for simultaneously targeting multiple genes.

PubMed

Vad-Nielsen, Johan; Lin, Lin; Bolund, Lars; Nielsen, Anders Lade; Luo, Yonglun

2016-11-01

The engineered CRISPR/Cas9 technology has developed as the most efficient and broadly used genome editing tool. However, simultaneously targeting multiple genes (or genomic loci) in the same individual cells using CRISPR/Cas9 remain one technical challenge. In this article, we have developed a Golden Gate Assembly method for the generation of CRISPR gRNA expression arrays, thus enabling simultaneous gene targeting. Using this method, the generation of CRISPR gRNA expression array can be accomplished in 2 weeks, and contains up to 30 gRNA expression cassettes. We demonstrated in the study that simultaneously targeting 10 genomic loci or simultaneously inhibition of multiple endogenous genes could be achieved using the multiplexed gRNA expression array vector in human cells. The complete set of plasmids is available through the non-profit plasmid repository Addgene.

MicroRNA profiling reveals dysregulated microRNAs and their target gene regulatory networks in cemento-ossifying fibroma.

PubMed

Pereira, Thaís Dos Santos Fontes; Brito, João Artur Ricieri; Guimarães, André Luiz Sena; Gomes, Carolina Cavaliéri; de Lacerda, Júlio Cesar Tanos; de Castro, Wagner Henriques; Coimbra, Roney Santos; Diniz, Marina Gonçalves; Gomez, Ricardo Santiago

2018-01-01

Cemento-ossifying fibroma (COF) is a benign fibro-osseous neoplasm of uncertain pathogenesis, and its treatment results in morbidity. MicroRNAs (miRNA) are small non-coding RNAs that regulate gene expression and may represent therapeutic targets. The purpose of the study was to generate a comprehensive miRNA profile of COF compared to normal bone. Additionally, the most relevant pathways and target genes of differentially expressed miRNA were investigated by in silico analysis. Nine COF and ten normal bone samples were included in the study. miRNA profiling was carried out by using TaqMan® OpenArray® Human microRNA panel containing 754 validated human miRNAs. We identified the most relevant miRNAs target genes through the leader gene approach, using STRING and Cytoscape software. Pathways enrichment analysis was performed using DIANA-miRPath. Eleven miRNAs were downregulated (hsa-miR-95-3p, hsa-miR-141-3p, hsa-miR-205-5p, hsa-miR-223-3p, hsa-miR-31-5p, hsa-miR-944, hsa-miR-200b-3p, hsa-miR-135b-5p, hsa-miR-31-3p, hsa-miR-223-5p and hsa-miR-200c-3p), and five were upregulated (hsa-miR-181a-5p, hsa-miR-181c-5p, hsa-miR-149-5p, hsa-miR-138-5p and hsa-miR-199a-3p) in COF compared to normal bone. Eighteen common target genes were predicted, and the leader genes approach identified the following genes involved in human COF: EZH2, XIAP, MET and TGFBR1. According to the biology of bone and COF, the most relevant KEGG pathways revealed by enrichment analysis were proteoglycans in cancer, miRNAs in cancer, pathways in cancer, p53-, PI3K-Akt-, FoxO- and TGF-beta signalling pathways, which were previously found to be differentially regulated in bone neoplasms, odontogenic tumours and osteogenesis. miRNA dysregulation occurs in COF, and EZH2, XIAP, MET and TGFBR1 are potential targets for functional analysis validation. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

An Integration of Genome-Wide Association Study and Gene Expression Profiling to Prioritize the Discovery of Novel Susceptibility Loci for Osteoporosis-Related Traits

PubMed Central

Demissie, Serkalem; Soranzo, Nicole; Bianchi, Estelle N.; Grundberg, Elin; Liang, Liming; Richards, J. Brent; Estrada, Karol; Zhou, Yanhua; van Nas, Atila; Moffatt, Miriam F.; Zhai, Guangju; Hofman, Albert; van Meurs, Joyce B.; Pols, Huibert A. P.; Price, Roger I.; Nilsson, Olle; Pastinen, Tomi; Cupples, L. Adrienne; Lusis, Aldons J.; Schadt, Eric E.; Ferrari, Serge; Uitterlinden, André G.

2010-01-01

Osteoporosis is a complex disorder and commonly leads to fractures in elderly persons. Genome-wide association studies (GWAS) have become an unbiased approach to identify variations in the genome that potentially affect health. However, the genetic variants identified so far only explain a small proportion of the heritability for complex traits. Due to the modest genetic effect size and inadequate power, true association signals may not be revealed based on a stringent genome-wide significance threshold. Here, we take advantage of SNP and transcript arrays and integrate GWAS and expression signature profiling relevant to the skeletal system in cellular and animal models to prioritize the discovery of novel candidate genes for osteoporosis-related traits, including bone mineral density (BMD) at the lumbar spine (LS) and femoral neck (FN), as well as geometric indices of the hip (femoral neck-shaft angle, NSA; femoral neck length, NL; and narrow-neck width, NW). A two-stage meta-analysis of GWAS from 7,633 Caucasian women and 3,657 men, revealed three novel loci associated with osteoporosis-related traits, including chromosome 1p13.2 (RAP1A, p = 3.6×10−8), 2q11.2 (TBC1D8), and 18q11.2 (OSBPL1A), and confirmed a previously reported region near TNFRSF11B/OPG gene. We also prioritized 16 suggestive genome-wide significant candidate genes based on their potential involvement in skeletal metabolism. Among them, 3 candidate genes were associated with BMD in women. Notably, 2 out of these 3 genes (GPR177, p = 2.6×10−13; SOX6, p = 6.4×10−10) associated with BMD in women have been successfully replicated in a large-scale meta-analysis of BMD, but none of the non-prioritized candidates (associated with BMD) did. Our results support the concept of our prioritization strategy. In the absence of direct biological support for identified genes, we highlighted the efficiency of subsequent functional characterization using publicly available expression profiling relevant to the skeletal system in cellular or whole animal models to prioritize candidate genes for further functional validation. PMID:20548944

Biologically relevant effects of mRNA amplification on gene expression profiles.

PubMed

van Haaften, Rachel I M; Schroen, Blanche; Janssen, Ben J A; van Erk, Arie; Debets, Jacques J M; Smeets, Hubert J M; Smits, Jos F M; van den Wijngaard, Arthur; Pinto, Yigal M; Evelo, Chris T A

2006-04-11

Gene expression microarray technology permits the analysis of global gene expression profiles. The amount of sample needed limits the use of small excision biopsies and/or needle biopsies from human or animal tissues. Linear amplification techniques have been developed to increase the amount of sample derived cDNA. These amplified samples can be hybridised on microarrays. However, little information is available whether microarrays based on amplified and unamplified material yield comparable results. In the present study we compared microarray data obtained from amplified mRNA derived from biopsies of rat cardiac left ventricle and non-amplified mRNA derived from the same organ. Biopsies were linearly amplified to acquire enough material for a microarray experiment. Both amplified and unamplified samples were hybridized to the Rat Expression Set 230 Array of Affymetrix. Analysis of the microarray data showed that unamplified material of two different left ventricles had 99.6% identical gene expression. Gene expression patterns of two biopsies obtained from the same parental organ were 96.3% identical. Similarly, gene expression pattern of two biopsies from dissimilar organs were 92.8% identical to each other.Twenty-one percent of reporters called present in parental left ventricular tissue disappeared after amplification in the biopsies. Those reporters were predominantly seen in the low intensity range. Sequence analysis showed that reporters that disappeared after amplification had a GC-content of 53.7+/-4.0%, while reporters called present in biopsy- and whole LV-samples had an average GC content of 47.8+/-5.5% (P <0.001). Those reporters were also predicted to form significantly more (0.76+/-0.07 versus 0.38+/-0.1) and longer (9.4+/-0.3 versus 8.4+/-0.4) hairpins as compared to representative control reporters present before and after amplification. This study establishes that the gene expression profile obtained after amplification of mRNA of left ventricular biopsies is representative for the whole left ventricle of the rat heart. However, specific gene transcripts present in parental tissues were undetectable in the minute left ventricular biopsies. Transcripts that were lost due to the amplification process were not randomly distributed, but had higher GC-content and hairpins in the sequence and were mainly found in the lower intensity range which includes many transcription factors from specific signalling pathways.

Biologically relevant effects of mRNA amplification on gene expression profiles

PubMed Central

van Haaften, Rachel IM; Schroen, Blanche; Janssen, Ben JA; van Erk, Arie; Debets, Jacques JM; Smeets, Hubert JM; Smits, Jos FM; van den Wijngaard, Arthur; Pinto, Yigal M; Evelo, Chris TA

2006-01-01

Background Gene expression microarray technology permits the analysis of global gene expression profiles. The amount of sample needed limits the use of small excision biopsies and/or needle biopsies from human or animal tissues. Linear amplification techniques have been developed to increase the amount of sample derived cDNA. These amplified samples can be hybridised on microarrays. However, little information is available whether microarrays based on amplified and unamplified material yield comparable results. In the present study we compared microarray data obtained from amplified mRNA derived from biopsies of rat cardiac left ventricle and non-amplified mRNA derived from the same organ. Biopsies were linearly amplified to acquire enough material for a microarray experiment. Both amplified and unamplified samples were hybridized to the Rat Expression Set 230 Array of Affymetrix. Results Analysis of the microarray data showed that unamplified material of two different left ventricles had 99.6% identical gene expression. Gene expression patterns of two biopsies obtained from the same parental organ were 96.3% identical. Similarly, gene expression pattern of two biopsies from dissimilar organs were 92.8% identical to each other. Twenty-one percent of reporters called present in parental left ventricular tissue disappeared after amplification in the biopsies. Those reporters were predominantly seen in the low intensity range. Sequence analysis showed that reporters that disappeared after amplification had a GC-content of 53.7+/-4.0%, while reporters called present in biopsy- and whole LV-samples had an average GC content of 47.8+/-5.5% (P <0.001). Those reporters were also predicted to form significantly more (0.76+/-0.07 versus 0.38+/-0.1) and longer (9.4+/-0.3 versus 8.4+/-0.4) hairpins as compared to representative control reporters present before and after amplification. Conclusion This study establishes that the gene expression profile obtained after amplification of mRNA of left ventricular biopsies is representative for the whole left ventricle of the rat heart. However, specific gene transcripts present in parental tissues were undetectable in the minute left ventricular biopsies. Transcripts that were lost due to the amplification process were not randomly distributed, but had higher GC-content and hairpins in the sequence and were mainly found in the lower intensity range which includes many transcription factors from specific signalling pathways. PMID:16608515

Characterization of canine osteosarcoma by array comparative genomic hybridization and RT-qPCR: signatures of genomic imbalance in canine osteosarcoma parallel the human counterpart.

PubMed

Angstadt, Andrea Y; Motsinger-Reif, Alison; Thomas, Rachael; Kisseberth, William C; Guillermo Couto, C; Duval, Dawn L; Nielsen, Dahlia M; Modiano, Jaime F; Breen, Matthew

2011-11-01

Osteosarcoma (OS) is the most commonly diagnosed malignant bone tumor in humans and dogs, characterized in both species by extremely complex karyotypes exhibiting high frequencies of genomic imbalance. Evaluation of genomic signatures in human OS using array comparative genomic hybridization (aCGH) has assisted in uncovering genetic mechanisms that result in disease phenotype. Previous low-resolution (10-20 Mb) aCGH analysis of canine OS identified a wide range of recurrent DNA copy number aberrations, indicating extensive genomic instability. In this study, we profiled 123 canine OS tumors by 1 Mb-resolution aCGH to generate a dataset for direct comparison with current data for human OS, concluding that several high frequency aberrations in canine and human OS are orthologous. To ensure complete coverage of gene annotation, we identified the human refseq genes that map to these orthologous aberrant dog regions and found several candidate genes warranting evaluation for OS involvement. Specifically, subsequenct FISH and qRT-PCR analysis of RUNX2, TUSC3, and PTEN indicated that expression levels correlated with genomic copy number status, showcasing RUNX2 as an OS associated gene and TUSC3 as a possible tumor suppressor candidate. Together these data demonstrate the ability of genomic comparative oncology to identify genetic abberations which may be important for OS progression. Large scale screening of genomic imbalance in canine OS further validates the use of the dog as a suitable model for human cancers, supporting the idea that dysregulation discovered in canine cancers will provide an avenue for complementary study in human counterparts. Copyright © 2011 Wiley-Liss, Inc.

Effect of human rhinovirus infection on airway epithelium tight junction protein disassembly and transepithelial permeability.

PubMed

Looi, Kevin; Troy, Niamh M; Garratt, Luke W; Iosifidis, Thomas; Bosco, Anthony; Buckley, Alysia G; Ling, Kak-Ming; Martinovich, Kelly M; Kicic-Starcevich, Elizabeth; Shaw, Nicole C; Sutanto, Erika N; Zosky, Graeme R; Rigby, Paul J; Larcombe, Alexander N; Knight, Darryl A; Kicic, Anthony; Stick, Stephen M

2016-10-11

No studies have assessed the effects of human rhinovirus (HRV) infection on epithelial tight junctions (TJs) and resultant barrier function. To correlate viral infection with TJ disassembly, epithelial barrier integrity, and function. Human airway epithelial cells were infected with HRV minor serotype 1B (HRV-1B) at various 50% tissue culture infectivity doses (TCID 50 ) over 72 hours. HRV replication was assessed by quantitative-polymerase chain reaction (qPCR) while cell viability and apoptosis were assessed by proliferation and apoptotic assays, respectively. Protein expression of claudin-1, occludin, and zonula occludens protein-1 (ZO-1) was assessed using In-Cell™ Western assays. Transepithelial permeability assays were performed to assess effects on barrier functionality. RT 2 Profiler focused qPCR arrays and pathway analysis evaluating associations between human TJ and antiviral response were performed to identify potential interactions and pathways between genes of interests. HRV-1B infection affected viability that was both time and TCID 50 dependent. Significant increases in apoptosis and viral replication post-infection correlated with viral titer. Viral infection significantly decreased claudin-1 protein expression at the lower TCID 50 , while a significant decrease in all three TJ protein expressions occurred at higher TCID 50 . Decrease in protein expression was concomitant with significant increases in epithelial permeability of fluorescein isothiocynate labeled-dextran 4 and 20 kDa. Analysis of focused qPCR arrays demonstrated a significant decrease in ZO-1 gene expression. Furthermore, network analysis between human TJ and antiviral response genes revealed possible interactions and regulation of TJ genes via interleukin (IL)-15 in response to HRV-1B infection. HRV-1B infection directly alters human airway epithelial TJ expression leading to increased epithelial permeability potentially via an antiviral response of IL-15.

Genome-wide DNA methylation profiling in infants born to gestational diabetes mellitus.

PubMed

Weng, Xiaoling; Liu, Fatao; Zhang, Hong; Kan, Mengyuan; Wang, Ting; Dong, Mingyue; Liu, Yun

2018-03-26

Offspring exposed to gestational diabetes mellitus (GDM) are at a high risk for metabolic diseases. The mechanisms behind the association between offspring exposed to GDM in utero and an increased risk of health consequences later in life remain unclear. The aim of this study was to clarify the changes in methylation levels in the foetuses of women with GDM and to explore the possible mechanisms linking maternal GDM with a high risk of metabolic diseases in offspring later in life. A genome-wide comparative methylome analysis on the umbilical cord blood of infants born to 30 women with GDM and 33 women with normal pregnancy was performed using Infinium HumanMethylation 450 BeadChip assays. A quantitative methylation analysis of 18 CpG dinucleotides was verified in the validation umbilical cord blood samples from 102 newborns exposed to GDM and 103 newborns who experienced normal pregnancy by MassARRAY EpiTYPER. A total of 4485 differentially methylated sites (DMSs), including 2150 hypermethylated sites and 2335 hypomethylated sites, with a mean β-value difference of >0.05, were identified by the 450k array. Good agreement was observed between the massarray validation data and the 450k array data (R 2 > 0.99; P < 0.0001). Thirty-seven CpGs (representing 20 genes) with a β-value difference of >0.15 between the GDM and healthy groups were identified and showed potential as clinical biomarkers for GDM. "hsa04940: Type I diabetes mellitus" was the most significant Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway, with a P-value = 3.20E-07 and 1.36E-02 in the hypermethylated and hypomethylated genepathway enrichment analyses, respectively. In the Gene Ontology (GO) pathway analyses, immune MHC-related pathways and neuron development-related pathways were significantly enriched. Our results suggest that GDM has epigenetic effects on genes that are preferentially involved in the Type I diabetes mellitus pathway, immune MHC (major histocompatibility complex)-related pathways and neuron development-related pathways, with consequences on fetal growth and development, and provide supportive evidence that DNA methylation is involved in fetal metabolic programming. Copyright © 2018. Published by Elsevier B.V.

Comparative Genome Analysis of Basidiomycete Fungi

DOE Office of Scientific and Technical Information (OSTI.GOV)

Riley, Robert; Salamov, Asaf; Morin, Emmanuelle

Fungi of the phylum Basidiomycota (basidiomycetes), make up some 37percent of the described fungi, and are important in forestry, agriculture, medicine, and bioenergy. This diverse phylum includes the mushrooms, wood rots, symbionts, and plant and animal pathogens. To better understand the diversity of phenotypes in basidiomycetes, we performed a comparative analysis of 35 basidiomycete fungi spanning the diversity of the phylum. Phylogenetic patterns of lignocellulose degrading genes suggest a continuum rather than a sharp dichotomy between the white rot and brown rot modes of wood decay. Patterns of secondary metabolic enzymes give additional insight into the broad array of phenotypesmore » found in the basidiomycetes. We suggest that the profile of an organism in lignocellulose-targeting genes can be used to predict its nutritional mode, and predict Dacryopinax sp. as a brown rot; Botryobasidium botryosum and Jaapia argillacea as white rots.« less

Construction, database integration, and application of an Oenothera EST library.

PubMed

Mrácek, Jaroslav; Greiner, Stephan; Cho, Won Kyong; Rauwolf, Uwe; Braun, Martha; Umate, Pavan; Altstätter, Johannes; Stoppel, Rhea; Mlcochová, Lada; Silber, Martina V; Volz, Stefanie M; White, Sarah; Selmeier, Renate; Rudd, Stephen; Herrmann, Reinhold G; Meurer, Jörg

2006-09-01

Coevolution of cellular genetic compartments is a fundamental aspect in eukaryotic genome evolution that becomes apparent in serious developmental disturbances after interspecific organelle exchanges. The genus Oenothera represents a unique, at present the only available, resource to study the role of the compartmentalized plant genome in diversification of populations and speciation processes. An integrated approach involving cDNA cloning, EST sequencing, and bioinformatic data mining was chosen using Oenothera elata with the genetic constitution nuclear genome AA with plastome type I. The Gene Ontology system grouped 1621 unique gene products into 17 different functional categories. Application of arrays generated from a selected fraction of ESTs revealed significantly differing expression profiles among closely related Oenothera species possessing the potential to generate fertile and incompatible plastid/nuclear hybrids (hybrid bleaching). Furthermore, the EST library provides a valuable source of PCR-based polymorphic molecular markers that are instrumental for genotyping and molecular mapping approaches.

Multipotent human stromal cells isolated from cord blood, term placenta and adult bone marrow show distinct differences in gene expression pattern

PubMed Central

Matigian, Nicholas; Brooke, Gary; Zaibak, Faten; Rossetti, Tony; Kollar, Katarina; Pelekanos, Rebecca; Heazlewood, Celena; Mackay-Sim, Alan; Wells, Christine A.; Atkinson, Kerry

2014-01-01

Multipotent mesenchymal stromal cells derived from human placenta (pMSCs), and unrestricted somatic stem cells (USSCs) derived from cord blood share many properties with human bone marrow-derived mesenchymal stromal cells (bmMSCs) and are currently in clinical trials for a wide range of clinical settings. Here we present gene expression profiles of human cord blood-derived unrestricted somatic stem cells (USSCs), human placental-derived mesenchymal stem cells (hpMSCs), and human bone marrow-derived mesenchymal stromal cells (bmMSCs), all derived from four different donors. The microarray data are available on the ArrayExpress database (www.ebi.ac.uk/arrayexpress) under accession number E-TABM-880. Additionally, the data has been integrated into a public portal, www.stemformatics.org. Our data provide a resource for understanding the differences in MSCs derived from different tissues. PMID:26484151

The chrysanthemum leaf and root transcript profiling in response to salinity stress.

PubMed

Cheng, Peilei; Gao, Jiaojiao; Feng, Yitong; Zhang, Zixin; Liu, Yanan; Fang, Weimin; Chen, Sumei; Chen, Fadi; Jiang, Jiafu

2018-06-23

RNA-Seq was applied to capture the transcriptome of the leaf and root of non-treated and salinity-treated chrysanthemum cv. 'Jinba' plants. A total of 206,868 unigenes of mean length 849 nt and of N50 length 1363 nt was identified; of these about 64% (>132,000) could be functionally assigned. Depending on the severity of the salinity stress, differential transcription was observed for genes encoding proteins involved in osmotic adjustment, in ion transport, in reactive oxygen species scavenging and in the regulation of abscisic acid (ABA) signaling. The root stress response was dominated by the up-regulation of genes involved in ion transport and homeostasis, while that of the leaf reflected the plant's effort to make osmotic adjustments and to regulate ABA signaling. An array of known transcription factors (WRKY, AP2/ERF, MYB, bHLH and NAC) were differentially transcribed. Copyright © 2018. Published by Elsevier B.V.

Development of multitissue microfluidic dynamic array for assessing changes in gene expression associated with channel catfish appetite, growth, metabolism, and intestinal health

USDA-ARS?s Scientific Manuscript database

Large-scale, gene expression methods allow for high throughput analysis of physiological pathways at a fraction of the cost of individual gene expression analysis. Systems, such as the Fluidigm quantitative PCR array described here, can provide powerful assessments of the effects of diet, environme...

Targeted capture and resequencing of 1040 genes reveal environmentally driven functional variation in grey wolves.

PubMed

Schweizer, Rena M; Robinson, Jacqueline; Harrigan, Ryan; Silva, Pedro; Galverni, Marco; Musiani, Marco; Green, Richard E; Novembre, John; Wayne, Robert K

2016-01-01

In an era of ever-increasing amounts of whole-genome sequence data for individuals and populations, the utility of traditional single nucleotide polymorphisms (SNPs) array-based genome scans is uncertain. We previously performed a SNP array-based genome scan to identify candidate genes under selection in six distinct grey wolf (Canis lupus) ecotypes. Using this information, we designed a targeted capture array for 1040 genes, including all exons and flanking regions, as well as 5000 1-kb nongenic neutral regions, and resequenced these regions in 107 wolves. Selection tests revealed striking patterns of variation within candidate genes relative to noncandidate regions and identified potentially functional variants related to local adaptation. We found 27% and 47% of candidate genes from the previous SNP array study had functional changes that were outliers in sweed and bayenv analyses, respectively. This result verifies the use of genomewide SNP surveys to tag genes that contain functional variants between populations. We highlight nonsynonymous variants in APOB, LIPG and USH2A that occur in functional domains of these proteins, and that demonstrate high correlation with precipitation seasonality and vegetation. We find Arctic and High Arctic wolf ecotypes have higher numbers of genes under selection, which highlight their conservation value and heightened threat due to climate change. This study demonstrates that combining genomewide genotyping arrays with large-scale resequencing and environmental data provides a powerful approach to discern candidate functional variants in natural populations. © 2015 John Wiley & Sons Ltd.

Digital detection of multiple minority mutants and expression levels of multiple colorectal cancer-related genes using digital-PCR coupled with bead-array.

PubMed

Huang, Huan; Li, Shuo; Sun, Lizhou; Zhou, Guohua

2015-01-01

To simultaneously analyze mutations and expression levels of multiple genes on one detection platform, we proposed a method termed "multiplex ligation-dependent probe amplification-digital amplification coupled with hydrogel bead-array" (MLPA-DABA) and applied it to diagnose colorectal cancer (CRC). CRC cells and tissues were sampled to extract nucleic acid, perform MLPA with sequence-tagged probes, perform digital emulsion polymerase chain reaction (PCR), and produce a hydrogel bead-array to immobilize beads and form a single bead layer on the array. After hybridization with fluorescent probes, the number of colored beads, which reflects the abundance of expressed genes and the mutation rate, was counted for diagnosis. Only red or green beads occurred on the chips in the mixed samples, indicating the success of single-molecule PCR. When a one-source sample was analyzed using mixed MLPA probes, beads of only one color occurred, suggesting the high specificity of the method in analyzing CRC mutation and gene expression. In gene expression analysis of a CRC tissue from one CRC patient, the mutant percentage was 3.1%, and the expression levels of CRC-related genes were much higher than those of normal tissue. The highly sensitive MLPA-DABA succeeds in the relative quantification of mutations and gene expressions of exfoliated cells in stool samples of CRC patients on the same chip platform. MLPA-DABA coupled with hydrogel bead-array is a promising method in the non-invasive diagnosis of CRC.

DiRE: identifying distant regulatory elements of co-expressed genes

PubMed Central

Gotea, Valer; Ovcharenko, Ivan

2008-01-01

Regulation of gene expression in eukaryotic genomes is established through a complex cooperative activity of proximal promoters and distant regulatory elements (REs) such as enhancers, repressors and silencers. We have developed a web server named DiRE, based on the Enhancer Identification (EI) method, for predicting distant regulatory elements in higher eukaryotic genomes, namely for determining their chromosomal location and functional characteristics. The server uses gene co-expression data, comparative genomics and profiles of transcription factor binding sites (TFBSs) to determine TFBS-association signatures that can be used for discriminating specific regulatory functions. DiRE's unique feature is its ability to detect REs outside of proximal promoter regions, as it takes advantage of the full gene locus to conduct the search. DiRE can predict common REs for any set of input genes for which the user has prior knowledge of co-expression, co-function or other biologically meaningful grouping. The server predicts function-specific REs consisting of clusters of specifically-associated TFBSs and it also scores the association of individual transcription factors (TFs) with the biological function shared by the group of input genes. Its integration with the Array2BIO server allows users to start their analysis with raw microarray expression data. The DiRE web server is freely available at http://dire.dcode.org. PMID:18487623

Metabolic and spatio-taxonomic response of uncultivated seafloor bacteria following the Deepwater Horizon oil spill

DOE Office of Scientific and Technical Information (OSTI.GOV)

Handley, K. M.; Piceno, Y. M.; Hu, P.

The release of 700 million liters of oil into the Gulf of Mexico over a few months in 2010 produced dramatic changes in the microbial ecology of the water and sediment. Here, we reconstructed the genomes of 57 widespread uncultivated bacteria from post-spill deep-sea sediments, and recovered their gene expression pattern across the seafloor. These genomes comprised a common collection of bacteria that were enriched in heavily affected sediments around the wellhead. Although rare in distal sediments, some members were still detectable at sites up to 60 km away. Many of these genomes exhibited phylogenetic clustering indicative of common traitmore » selection by the environment, and within half we identified 264 genes associated with hydrocarbon degradation. Alkane degradation ability was near ubiquitous among candidate hydrocarbon degraders, whereas just three harbored elaborate gene inventories for the degradation of alkanes and aromatic and polycyclic aromatic hydrocarbons (PAHs). Differential gene expression profiles revealed a spill-promoted microbial sulfur cycle alongside gene upregulation associated with PAH degradation. Gene expression associated with alkane degradation was widespread, although active alkane degrader identities changed along the pollution gradient. Analyses suggest that a broad metabolic capacity to respond to oil inputs exists across a large array of usually rare indigenous deep-sea bacteria.« less