title: Selective estrogen receptor-beta agonists repress transcription of proinflammatory

status: Public on Apr 12 2008

city: San Francisco

first: Dale

last: Leitman

address: 513 Parnassus Ave.

department: Ob/Gyn

institute: University of California, San Francisco

summary:
In addition to their role in the development and function of the reproductive system, estrogens have significant anti-inflammatory properties. Although both estrogen receptors (ERs) can mediate anti-inflammatory actions, ERbeta is a more desirable therapeutic target because ERalpha mediates the proliferative effects of estrogens on the mammary gland and uterus. In fact, selective ERbeta agonists have beneficial effects in preclinical models involving inflammation without causing growth-promoting effects on the uterus or mammary gland. However, their mechanism of action is unclear. The purpose of this study was to use microarray analysis to determine whether ERbeta-selective compounds produce their anti-inflammatory effects by repressing transcription of proinflammatory genes. We identified 49 genes that were activated by TNF-alpha in human osteosarcoma U2OS cells expressing ERbeta. Estradiol treatment significantly reduced the activation by TNF-alpha on 18 genes via ERbeta or ERalpha. Most repressed genes were inflammatory genes, such as TNF-alpha, IL-6, and CSF2. Three ERbeta-selective compounds, ERB-041, WAY-202196, and WAY-214156, repressed the expression of these and other inflammatory genes. ERB-041 was the most ERbeta-selective compound, whereas WAY-202196 and WAY-214156 were the most potent. The ERbeta-selective compounds repressed inflammatory genes by recruiting the coactivator, SRC-2. ERB-041 also repressed cytokine genes in PBMCs, demonstrating that ERbeta-selective estrogens have anti-inflammatory properties in immune cells. Our study suggests that the anti-inflammatory effects of ERB-041 and other ERbeta-selective estrogens in animal models are due to transcriptional repression of proinflammatory genes. These compounds might represent a new class of drugs to treat inflammatory disorders.
Keywords: estrogen receptor, gene regulation, TNF

overall_design: Expression of ER in the U2OS-ERß cells was induced with doxycycline. The cells were treated in the absence or presence of 10 nM E2 for 18 h followed by incubation with TNF-{alpha} for 1 h.  Total RNA were extracted, followed by cRNA labelling, and hybridization of fragmented samples to the U95Av2 arrays (n = 3 for untreated, n = 4 for TNF-{alpha}-treated, n = 4 for TNF-{alpha}- and E2-treated samples). The data were analyzed using the Microarray Suite Version 5.0 with the default parameters. The comparative data generated for each treated group were analyzed further in Microsoft Excel. TNF-{alpha}-induced genes were selected for further analysis only if they had a 1.2 signal log ratio mean value (2.3-fold change) and were statistically significant (p < 0.05) in at least three separate experiments.

first: Aleksandra

last: Cvoro

first: Deirdre

last: Tatomer

first: Meng-Kian

last: Tee

first: Tatjana

last: Zogovic

first: Heather

last: Harris


title: Expression data from Arabidopsis thaliana (Ler) rosette leaves treated with paraquat (methyl viologen)

status: Public on Feb 11 2008

city: Zurich

first: Christophe

last: Laloi

address: Universitaetstrasse 2

department: Institute of Plant Science

country: Switzerland

institute: ETH Zurich

summary:
We used microarrays to detail Arabidopsis gene expression in response to paraquat, a herbicide that acts as a terminal oxidant of photosystem I that in the light leads to the enhanced generation of superoxide and hydrogen peroxide inside plastids. Within a few hours after paraquat treatment changes in nuclear gene expression occur. Distinct sets of genes were activated that were different from those induced by another reactive oxygen species, singlet oxygen.
Keywords: Time course

overall_design: Arabidopsis thaliana rosette leaves were harvested 1, 2, and 4 h after spraying either with a solution of 20 microM paraquat (methyl viologen, Sigma) in 0.1% Tween or with Tween alone for RNA extraction and hybridization on Affymetrix ATH1 microarrays. Plants were grown on soil for 3 weeks under continuous light at 90 mmol. m-2 . s-1. For each sample, the rosette leaves of five to six 3-week-old plants (before they start bolting) were collected for RNA extraction. Total RNAs from two separate biological experiments were pooled for the preparation of cDNA and the subsequent synthesis of biotin-labeled complementary RNA as recommended by Affymetrix.

first: Klaus

last: Apel


title: Higher order transcriptional regulation conferred by the bountiful gain of function mutant

status: Public on Jan 31 2008

city: Loughborough

last: Nottingham Arabidopsis Stock Centre (NASC)

address: Sutton Bonington Campus

department: School of Biosciences, University of Nottingham

country: United Kingdom

summary:
In recent years our group has been constructing an activation tag library based on the En-I transposon system which resulted in the identification of novel Arabidopsis mutants (Marsch Martinez et al 2002; manuscript accepted for publication in Plant Phys). Among them the bountiful (bou) mutant showed dominant alterations in leaf size and morphology, delayed flowering, vertically oriented siliques and higher yield. Sequence analysis of the genomic region flanking the transposon insertion in combination with expression analysis indicated that the mutant phenotype observed was presumably caused by over-expression of a gene encoding a yet uncharacterised DNA binding protein. In particular over-expression in the activation tagged mutant seems to cause ectopic expression of the BOU gene in all vegetative and reproductive tissues while the endogenous pattern of expression appeared to be restricted only to root tissue. Over-expression lines in which the BOU ORF was driven by the 35S promoter displayed the bountiful phenotype confirming that the activation tagged phenotype was indeed caused by activation of the BOU gene. Protein sequence analysis indicates a putative role for BOU as a chromatin remodelling factor which in association with the expression pattern suggests a possible involvement of BOU in high-hierarchy order of regulation of gene expression. Hence microarrays could be very useful for the identification of downstream interacting factors or target geneswhich will help us to gain insights towards unravelling the biological role of the BOU gene. The specific interaction to flowering time genes will be studied independently using RT-PCR to reveal the relationship to other genes in the regulatory pathway. As experimental setup we intend to compare gene expression between bountiful mutant and wild-type arabidopsis plants using rosette leaf tissue as source for RNA. Though the BOU gene is endogenously expressed in a root-specific mannerits ectopic expression in the bountiful mutant phenotipically affects the whole plant including leaves.

Experimenter name: Raffaella Greco
Experimenter department: Pereira Lab
Experimenter institute: Plant Research International
Experimenter address: Business Unit Genomics
Plant Research International
Postbus 16
Wageningen
Experimenter zip/postal_code: 6700 AA
Experimenter country: The Netherlands
Keywords: genetic_modification_design

overall_design: 2 samples were used in this experiment

first: Raffaella

last: Greco

first: Andy

last: Pereira

first: Zoe

last: Emmerson

first: Beatrice

last: Schildknecht


title: Low_Mo_Arabidopsis_mapping_MOT1

status: Public on Jan 15 2008

city: St. Louis

first: Ivan

last: Baxter

state: MO

address: 975 North Warson Rd

department: PLant Genetics Research Unit

country: USA

institute: USDA-ARS

summary:
ABSTRACT
Molybdenum (Mo) is an essential micronutrient for plants, serving as a cofactor for enzymes involved in nitrate assimilation, sulfite detoxification, abscisic acid biosynthesis and purine degradation.  Here we show that natural variation in shoot Mo content across 92 Arabidopsis thaliana accessions is controlled by variation in a mitochondrially localized transporter (Molybdenum Transporter 1 -MOT1) that belongs to the sulfate transporter superfamily. A deletion in the MOT1 promoter is strongly associated with low shoot Mo, occurring in seven of the accessions with the lowest shoot content of Mo. Consistent with the low Mo phenotype, MOT1 expression in low Mo accessions is reduced. Reciprocal grafting experiments demonstrate that the roots of Ler-0 are responsible for the low Mo accumulation in shoot, and GUS localization demonstrates that MOT1 is expressed strongly in the roots. MOT1 contains an N-terminal mitochondrial targeting sequence, and expression of MOT1 tagged with GFP in protoplasts, and transgenic plants, establishes the mitochondrial localization of this protein. Furthermore, expression of MOT1 specifically enhances Mo accumulation in yeast by 5-fold, consistent with MOT1 functioning as a molybdate transporter.  This work provides the first molecular insight into the processes that regulate Mo accumulation in plants, and shows that novel loci can be detected by association mapping.
Keywords: genomic hybridization bulked segregant analysis

overall_design: Hybridizations from a set of Bulk Segregant analysis. An F2 population from 14501 in the col-0 background crossed to Ler-0 was analyzed. This series contains the 3 hybs from Ler used to identify Single Feature Polymorphisms, the 3 hybs of Col-0 they were compared to, and 1 hyb for each pool from the BSA mapping(High Mo pool, low Mo Pool).


title: Anti-tumor Activity of Histone Deacetylase Inhibitors in Non-Small Cell Lung Cancer Cells

status: Public on Jan 08 2008

city: Tokyo

first: Akihiko

last: Gemma

address: Bumkyo-ku

country: Japan

institute: Nippon Medical School

summary:
In order to ascertain the potential for histone deacetylase (HDAC) inhibitor-based treatment in non-small cell lung cancer (NSCLC), we analyzed the anti-tumour effects of Trichostatin A (TSA) and suberoylanilide hydroxamic acid (vorinostat) in a panel of 16 NSCLC cell lines via MTT assay.  TSA and vorinostat both displayed strong anti-tumor activities in a proportion of NSCLC cell lines, and suggesting the need for the use of predictive markers to select patients receiving this treatment.  There was a strong correlation between the responsiveness to TSA and vorinostat (P < 0.0001).
To identify a molecular model of sensitivity to HDAC inhibitor treatment in NSCLC, we conducted a gene expression profiling study using cDNA arrays on the same set of cell lines and related the cytotoxic activity of TSA to corresponding gene expression pattern using a modified NCI program.  In addition, pathway analysis was performed with Pathway Architect software.  We used nine genes, which were identified by gene-drug sensitivity correlation and pathway analysis, to build a support vector machine (SVM) algorithm model by which sensitive cell lines were distinguished from resistant cell lines.  The prediction performance of the SVM model was validated by an additional seven cell lines, resulting in a prediction value of 100% in respect to determining response to TSA.  Our results suggested that [1] HDAC inhibitors may be promising anticancer drugs to NSCLC, and [2] the nine gene classifer is useful in predicting drug sensitivity to HDAC inhibitors and may contribute to achieving individualized therapy for NSCLC patients.

training sample set:
GSM94303 PC9
GSM94304 PC7
GSM94305 PC14
GSM94306 A549
GSM94308 LK2
GSM94313 RERF LC-KJ
GSM94314 RERF LC-MS
GSM94315 RERF-LC-AI
GSM94316 PC-1
GSM94317 PC-3
GSM94319 PC-10
GSM94323 ABC-1
GSM94324 EBC-1
GSM94325 LC2/ad
GSM94328 SQ-5
GSM94329 QG-56

test sample set:
GSM94307 LU65
GSM94326 LC1/sq
GSM94327 LC-1F
GSM254967 LCOK
GSM254968 LCD
GSM254969 H1650
GSM254970 H1975
Keywords: HDAC inhibitor, NSCLC, cDNA array, drug sensitivity test, pathway analysis

overall_design: gene expression analyis in lung cancer cell lines


title: Key Regulatory Molecules of Cartilage Destruction in Rheumatoid Arthritis: An in vitro Study

status: Public on Jan 04 2008

city: Berlin

first: Thomas

last: Häupl

address: Charitéplatz 1

department: Rheumatology

country: Germany

institute: Charité

summary:
We have studied the expression profile of 3D cultured human chondrocytes that were stimulated with supernatant of synovial fibroblasts derived from a RA patient (RASF=HSE cell line) and from a normal donor (NDSF=K4IM cell line), respectively. For this purpose, passage 2 human chondrocytes were cultured for 14 days in alginate beads and subsequently stimulated for 48 hours with supernatant of RASF and NDSF. Baseline expression was determined of unstimulated chondrocytes. Differential genome-wide microarray analysis of RASF and NDSF stimulated chondrocytes disclosed a distinct expression profile related to cartilage destruction  involving marker genes of inflammation (COX-2), NF-kappa B signaling pathway (TLR2), cytokines/chemokines and receptors (CXCL1-3, CCL20, CXCL8, CXCR4, IL-6, IL-1beta), matrix degradation (MMP-10, MMP-12) and suppressed matrix synthesis (COMP). Thus, transcriptome profiling of RASF and NDSF stimulated chondrocytes revealed a disturbed catabolic-anabolic homeostasis of chondrocyte function. This study provides a comprehensive insight into the molecular regulatory processes induced in human chondrocytes during RA-related cartilage destruction.
Keywords: dose response

overall_design: To reveal the RA-related chondrocyte gene expression profile, genome-wide microarray analysis of RASF stimulated human chondrocytes compared to NDSF stimulation was performed. Unstimulated chondrocytes were analyzed for baseline gene expression. For microarray analysis of RASF and NDSF stimulated and unstimulated chondrocytes, respectively, two RNA pools were analyzed, each pool consisting of equal amounts of RNA from three different donors.

first: Kristin

last: Andreas

first: Carsten

last: Lübke

first: Tilo

last: Dehne

first: Lars

last: Morawietz

first: Jochen

last: Ringe

first: Christian

last: Kaps

first: Michael

last: Sittinger


title: TRAIL Is a Novel Antiviral Protein against Dengue Virus

status: Public on Jan 05 2008

city: Belmont

first: Katherine

middle: J

last: Martin

state: MA

address: 61 Carleton Rd

institute: BioArray Consulting

summary:
Dengue fever is an important tropical illness for which there is currently no virus-specific treatment. To shed light on mechanisms involved in the cellular response to dengue virus (DV), we assessed gene expression changes, using Affymetrix GeneChips (HG-U133A), of infected primary human cells and identified changes common to all cells. The common response genes included a set of 23 genes significantly induced upon DV infection of human umbilical vein endothelial cells (HUVECs), dendritic cells (DCs), monocytes, and B cells (analysis of variance, P < 0.05). Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), one of the common response genes, was identified as a key link between type I and type II interferon response genes. We found that DV induces TRAIL expression in immune cells and HUVECs at the mRNA and protein levels. The induction of TRAIL expression by DV was found to be dependent on an intact type I interferon signaling pathway. A significant increase in DV RNA accumulation was observed in anti-TRAIL antibody-treated monocytes, B cells, and HUVECs, and, conversely, a decrease in DV RNA was seen in recombinant TRAIL-treated monocytes. Furthermore, recombinant TRAIL inhibited DV titers in DV-infected DCs by an apoptosis-independent mechanism. These data suggest that TRAIL plays an important role in the antiviral response to DV infection and is a candidate for antiviral interventions against DV.
We used Affymetrix microarrays to study the response of human host cells to dengue virus (DV).
Keywords: virus infection of multiple cell types

overall_design: For three human cell types, RNA was extracted and hybridized on Affymetrix microarrays. We compared a total of 10 samples. Five were infected in vitro for 48 hours with DV, including HUVECs (n=2), monocytes (n=2), and B-cells (n=1). Five were mock-infected controls of the same cell types and numbers. From these samples, were identified 23 genes that were induced by DV infection in all of the cell types.

first: Rajas

middle: V

last: Warke

first: Kris

last: Giaya

first: Sunil

middle: K

last: Shaw

first: Alan

middle: L

last: Rothman

first: Irene

last: Bosch


title: Identification of genes responsive to ultrasound in human leukemia U937 cells

status: Public on Jan 23 2008

city: Toyama

first: Yoshiaki

last: Tabuchi

state: Toyama

address: 2630 Sugitani

department: Life Science Research Center

institute: University of Toyama

summary:
Here, the effects of non-thermal low intensity pulsed ultrasound (LIPUS) on the gene expression in human lymphoma U937 cells were investigated using by an Affymetrix GeneChip system. Six hours after LIPUS treatment (0.3 W/cm2 for 1 min), apoptosis (14±3.8%, mean±SD) with minimal cell lysis was observed. At 3 h post-treatment, LIPUS down-regulated 193 genes and up-regulated 201 genes by >1.5-fold.
Keywords: ultrasound, gene expression, Human lymphoma U937 cell

overall_design: U937 cells, a human lymphoma cell line, were treated with low intensity pulsed ultrasound (0.3 W/cm2 for 1 min) and followed by incubation for 3 h at 37°C. Non-treated cells were served as control. Total RNA samples were prepared from the cells. Gene expression was analyzed by an Affymetrix GeneChip® system with Human Expression Array U133A which was spotted with 22,283 probe sets. Sample preparation for array hybridization was carried out as described in the manufacture’s instructions.

first: Ichiro

last: Takasaki

first: Takashi

last: Kondo


title: Early transcriptional responses induced by benzo(a)pyrene diol epoxide in human amnion epithelial FL cells

status: Public on Jan 01 2008

city: Hangzhou

first: Lu

last: XiangYun

address: 388 Yuhangtang Road

country: China

institute: Zhejiang University School of Medicine

summary:
Genotoxic agents cause cellular DNA damage and stress responses, including transcriptional changes. Here we focused on the early transcriptional responses of human cells to benzo(a)pyrene diol epoxide (BPDE), which causes bulky DNA adduct damage. Human amnion epithelial FL cells were exposed to three doses of BPDE (5, 50, and 500 nM) and the vehicle control DMSO, and differential gene expression profiles were obtained 4 h after exposure using oligonucleotide microarrays followed by validation with quantitative real-time RT-PCR.
Compared with a few genes affected by the low and medium-dose exposure, extensive and robust changes in gene expression were induced by the high-dose BPDE. We found that the expression of cell cycle-regulators, signaling molecules and transcription factors were significantly altered and important signaling pathways related to cell survival or apoptosis were affected by BPDE. Several genes and related regulatory pathways that were previously not known to be responsive to this genotoxic agent have now been implicated, which helps to draw the whole picture of how cells respond to environmental chemical exposure via transcriptional regulation.
Keywords: Dose response

overall_design: Human amnion epithelial FL cells were exposed to vehicle control (dimethyl sulfoxide) and increasing doses (5, 50, 500 nM) of anti-benzo(a)pyrene diol epoxide (anti-BPDE), respectively. The transcriptomes of the three treatments were compared to that of the control, respectively, to test the hypothesis that a characterized differential expression profile would be generated by exposure to various doses of this genotoxic agent.

first: Xiangyun

last: Lu

first: Jimin

last: Shao

first: Hongjuan

last: Li

first: Yingnian

last: Yu


title: Identification of genes responsive to mild hyperthermia in human leukemia U937 cells

status: Public on Jan 03 2008

summary:
The rationale underlying hyperthermia is the fact that temperatures over 42.5˚C are highly cytotoxic to tumor cells. On the other hand, although mild hyperthermia at a range from 39 to 41˚C alone did not induce cytotoxicity in tumor cells, mild hyperthermia is reported to show a synergism with radiotherapy and anti-cancer drugs. Here, the effects of mild hyperthermia (41˚C for 30 min) on the gene expression in human lymphoma U937 cells were investigated using by an Affymetrix GeneChip system. Although the cells treated with the mild hyperthermia did not induce apoptosis, a significant increase in protein levels of heat shock proteins, Hsp40 and Hsp70, was observed following activation of heat shock factor-1. At 3 h post-treatment, 938 probe sets that were differentially expressed by >1.5-fold were identified.
Keywords: mild hyperthermia, gene expression, Human lymphoma U937 cell

overall_design: U937 cells, a human lymphoma cell line, were treated with mild hyperthermia (41˚C for 30 min) and followed by incubation for 3 h at 37°C. Non-treated cells were served as control. Total RNA samples were prepared from the cells. Gene expression was analyzed by an Affymetrix GeneChip® system with Human Expression Array U133A which was spotted with 22,283 probe sets. Sample preparation for array hybridization was carried out as described in the manufacture’s instructions.


title: Gene expression in wild-type and transgenic plants overexpressing rice OsTOP6A1 gene

status: Public on Dec 26 2007

city: New Delhi

first: Mukesh

last: Jain

state: New Delhi

address: New Mehrauli Road

country: India

institute: Jawaharlal Nehru University

summary:
10-day-old wild-type and homozygous transgenic Arabidopsis seedlings (overexpressing rice topoisomerase 6 subunit A1; OsTOP6A1) grown under normal growth conditions were used for total RNA isolation. The 5 micrograms of each total RNA sample was processed for microarray analysis according to Affymetrix protocol.
Keywords: Arabidopsis, wild-type and transgenic seedlings, differential gene expression

overall_design: The experiment involves the study of differential gene expression in transgenic Arabidopsis seedlings overexpressing rice topoisomerase 6 subunit A1 (OsTOP6A1) cDNA as compared to wild-type.Three biological replicates for each sample were used for microarray analysis.

first: Akhilesh

last: Tyagi

first: Jitendra

middle: P

last: Khurana


title: Cell-specific nitrogen responses in the Arabidopsis root

status: Public on Dec 19 2007

city: New York

first: Kenneth

middle: David

last: Birnbaum

state: NY

address: 12 Waverly Place

department: Biology

institute: New York University

summary:
The organs of multicellular species are comprised of cell types that must function together to perform specific tasks. One critical organ function is responding to internal or external change but little is known about how responses are tailored to specific cell types or coordinated among them on a global level. Here we use cellular profiling of five Arabidopsis root cell types in response to a limiting resource, nitrogen, to uncover a vast and predominantly cell-specific response that was largely undetectable using traditional methods. These methods reveal a new class of cell-specific nitrogen responses. As a proof-of-principle, we dissected one cell-specific response circuit that mediates nitrogen-induced changes in root branching from pericycle cells. Thus, cellular response profiling links gene modules to discrete functions in specific cell types.
Keywords: cell type comparison, comparative genomic hybridization, genetic modification

overall_design: The whole experiment was carried out in triplicate with 84 chips in total (28 experiments).  The nitrogen response in 5 FACS-sorted root cell types (LRC, lateral root cap; EpiC, epidermis and cortex; EndoP, endodermis and pericycle; Peri, pericycle; Stele, stele), protoplasts and Col-0 whole roots was assayed using microarrays.  5mM KNO3 was used as a nitrogen treatment for 2 hours.  5mM KCl was used as a control treatment for the same length of time.  1mM MSX in combination with 5mM KNO3 (or with 5mM KCl as a control) was used to block the assimilation of KNO3, and 5mM Gln added to restore it in order to seperate KNO3 effects from assimilated-nitrogen effects.  5mM KNO3 was either maintained in the protoplast-generating solution (continuous treat) or not (transitory treat) to test how to best preserve the cellular nitrogen-response during the 1 hr protoplast generating procedure.  As controls we used protoplasts, whole roots frozen directly after the 2hr treatment or 3.5hr treatment, or whole roots incubated in protoplast-generating solution (minus enzymes; transitory or continuous treated) and then frozen.  After FACS-sorting or mock FACS-sorting cells were frozen.

middle: D

first: Gloria

middle: M

last: Coruzzi

first: Miriam

last: Gifford

first: Alexis

last: Dean

first: Rodrigo

middle: A

last: Gutierrez


title: Microarray Analysis of Arabidopsis Genome Response to Aluminum Stress

status: Public on Dec 31 2007

city: Memphis

first: Shirlean

middle: B

last: Goodwin

state: TN

address: 3774 Walker Ave

institute: University of Memphis

summary:
To better understand the mechanisms involved in aluminum (Al) toxicity and tolerance in plants, microarray technology was used to evaluate changes in gene expression in Arabidopsis thaliana under Al stress. With the use of Affymetrix Arabidopsis ATH1 Genechip, a comparison of RNA expression profiles was made between control and Al-treated Arabidopsis seedlings. A total of 256 genes were identified as Al-responsive. Ninety-four genes were shown to be up-regulated and 162 were down-regulated; comprising 1.1% of the 24,000 Arabidopsis genes. Real-time RT-PCR was used to confirm the microarray data. RNAs associated with known protein functions were classified into categories. The analysis showed that a large number of transcription factors and several putative signaling components were up-regulated by aluminum. Chloroplast structural and photosynthetic genes were, in general, down-regulated. A number of previously identified Al-responsive genes showed similar expression pattern changes in this study. For example, GST, Auxin-regulated, Peroxidase, and Chitinase, were found to be up-regulated by Al-stress, whereas, genes such as Wali 3 and Wali 4, were down-regulated. We also identified several up-regulated genes involved in vacuolar signaling, sorting and docking. Three genes were also up-regulated by Al-stress, Ras GTP-binding protein, ABC-cassette binding, and the AtELP1 receptor genes, have previously been documented as responsive to drought and/or oxidative stress and may play important roles the detoxification of Al ions by transportation and storage into root vacuoles. Ultrastructural changes in the roots tips cells of Arabidopsis were evaluated using TEM and EDXMA with SEM. Results using combined TEM and SEM-EDXMA showed that Al accumulattion in the root tip of Arabidopsis was detectable. The coupling these techniques can be useful for measuring Al in situ in cellular and sub cellular section of plant roots.
Keywords: Stress response

overall_design: 2 biological replications for a total of 4 Affymetrix GeneChips, control and Al-treated, total RNA was extracted from 100 pooled Arabidopsis seedlings for each GeneChip.

first: Thomas

middle: R

last: Sutter


title: Gene expression profiling in wear-particle induced and infectious endoprosthesis loosening

first: Lars

last: Morawietz

address: Chariteplatz 1

department: Institute for Pathology

institute: Charite University Hospital

summary:
The aim of the study was to identify markers for the early diagnosis of endoprosthesis loosening, for the differentiation between wear-particle induced and septic loosening, as well as to gather new insights into the pathogenesis.
824 genes were differentially expressed with a fold change greater than 2. Among these were Chitinase 1, CD52, Calpain 3, Apolipoprotein, CD18, Lysyl oxidase, Cathepsin D, E-Cadherin, VE-Cadherin, Nidogen, Angiopoietin 1 and Thrombospondin 2, and the differential expression levels were validated by RT-PCR. The chitinase activity was significantly higher in the blood from patients with wear-particle induced prosthesis loosening (p=0.001). However, using chitinase activity as a marker for early diagnosis, it has a specificity of 83% and a sensitivity of only 52%, due to a high variability both in the disease and in the control group.
Keywords: Tissue type comparison, disease state analysis, search for new biomarkers

overall_design: Gene expression profiles were generated from 5 periprosthetic membranes of wear-particle induced and 5 of infectious (septic) type using Affymetrix HG U133A oligonucleotide microarrays. The results of selected differentially expressed genes were validated by RT-PCR (n=30). The enzyme activity and the genotype of chitinase-1 were assessed in serum samples from 313 consecutive patients hospitalized for endoprosthesis loosening (n=54) or for other reasons, serving as control subjects (n=259).

first: Ralf-Juergen

last: Kuban

first: Ute

last: Ungethuem

first: Joerg

middle: H

last: Schroeder

first: Christian

last: Kaps

first: Andreas

last: Weimann

first: Veit

last: Krenn


title: Expression profiling of the Parkinsonian Brain

city: London

first: Linda

last: Moran

address: Fulham Palace Road

department: Neuropathology

institute: Imperial College London

summary:
Affymetrix HG_U133 array sets (A and B chips) were used to determine the whole genome transcription profile of clinically documented and neuropathologically confirmed cases of sporadic Parkinson's disease as well as controls.
Keywords: disease-state analysis

overall_design: Post mortem brain tissue samples from Parkinson's disease and control cases (substantia nigra, split into medial and lateral portions, and frontal cortex) were prepared for this study. In total, 47 individual tissue samples were analyzed using one A and one B GeneChip per sample, i.e. 15 samples of medial parkinsonian SN, 9 samples of lateral parkinsonian SN, 8 medial nigra control samples and 7 lateral nigra control samples. Lateral and medial nigra samples were from the same cases. In addition, frontal cerebral cortex was analyzed in five of the PD cases and in three of the controls (Neurogenetics 2006 7: 1–11; Acta Neuropathologica (Berlin) 2007 113: 253-263

first: Manuel

last: Graeber


title: Expression data of Drosophila 3rd instar larval wing discs taken from strains selected for wing shape.

status: Public on Jan 02 2008

city: San Diego

first: Ralph

last: Greenspan

state: CA

address: 10640 John Jay Hopkins Dr.

institute: The Neurosciences Institute

summary:
We measured gene expression across the whole genome in a panel of lines selected for a wing shape trait (angular offset).  The lines were created in separate experiments, originating from two widely separated populations, and including multiple replicates of one population, but all were created using the same selection regime and trait.  Here we evaluate the data with two objectives: 1) to identify candidate wing shape genes for future testing and validation, and 2) to assess variation among lines in the outcome of identical selection regimes
We used microarrays to detail the global programme of gene expression underlying the phenotypic differences among strains selected for wing shape angular offset.
Keywords: strain comparison

overall_design: Drosophila wing discs from each of 12 selected lines were dissected at 6.2 hr prior to pupariation for RNA extraction and hybridization on Affymetrix microarrays.

first: K

middle: E

last: Weber

first: R

last: Greenspan.


title: Expression data from human breast cancer cells (MCF-7) coexpressing ERalpha and Erbeta, treated with phytoestrogens

status: Public on Feb 07 2008

city: Santa Monica

first: Edmund

middle: C

last: Chang

state: California

address: 2225 Colorado Ave

institute: Kite Pharma

summary:
We used microarrays to detail the global transcriptional response mediated by ERalpha or ERbeta to the phytoestrogen genistein in the MCF-7 human breast cancer cell model.
Keywords: ligand response over time course

overall_design: MCF-7 human breast cancer cells expressing endogenouse Estrogen Receptor Alpha (ERalpha) were infected with adenovirus carrying either estrogen receptor beta (AdERb) or no insert (Ad) at multiplicity of infection (moi) of 20.  Cells were then treated with either vehicle control (veh), 6nM 17beta-estradiol (E2), 6nM genistein (LG), 300nM genistein (HG), 300nM S-Equol (EQ), HG+3uM ICI182,780 (IG), EQ+3uM ICI 182,780(IE) for additional periods of 4h or 24hr before RNA extraction and hybridization on Affymetrix microarrays. We sought to determine if genistein and S-Equol, phytoestrogens selective for the ERbeta can elicit transcriptional response distinctive from those mediated by the ERalpha.

first: Benita

middle: S

last: Katzenellenbogen

first: Tze

last: Charn

first: John

first: SungHee

last: Park

first: William

middle: G

last: Helferich

first: Barry

last: Komm


title: Experiment to identify downstream targets of Arabidopsis REVOLUTA (HDZIPIII) transcription factor.

status: Public on Dec 05 2007

city: Palo Alto

first: M. Kathryn

last: Barton

address: 260 Panama Street

department: Dept. of Plant Biology

institute: Carnegie Institution

summary:
The establishment of ad/abaxial polarity is a fundamental event in plant development.  It is critical for correct polar development of the leaf (the upper portion of the leaf is chloroplast rich and optimized for light capture while the lower portion is optimized for gas exchange) and for creating an environment that allows the formation of new meristems (centers of stem cell growth).  Class III homeodomain-leucine zipper (HD-ZIPIII) proteins are conserved plant proteins that act as potent regulators of ad/abaxial polarity.   HD-ZIPIII protein activity promotes the development of upper (adaxial) leaf fates and meristem formation; in its absence lower (abaxial) leaf fates develop and meristems fail to form.   A network of regulatory factors controls the establishment of ad/abaxial leaf fates.  However, this network is incomplete and how these factors control one another is poorly understood.  Here we report a new family of plant genes, the LITTLE ZIPPER (ZPR) genes (consisting principally of a stretch of leucine zipper similar to the leucine zipper in HD-ZIPIII proteins) that are transcriptionally up-regulated by HD-ZIPIII activity.  Furthermore, we show that the ZPR proteins interact with and repress HD-ZIPIII activity, thus forming a negative feedback loop.  Our results suggest that HD-ZIPIII proteins exist in active homodimers and, together with the ZPR proteins, in inactive heterodimers.   The newly discovered HD-ZIPIII/ZPR regulatory module would not only serve to dampen the effect of fluctuations in HD-ZIPIII protein levels but more importantly would provide a point of regulation - control over the ratio of inactive heterodimers to active homodimers - that could be influenced by other components of the pathway.  For instance, the binding of a small hydrophobic molecule to the conserved (yet little understood) START domain present in the HD-ZIPIII proteins may influence the type of dimer formed.
Published in:
Wenkel, S., Emery, J., Hou, B.-H., Evans, M.M.S. and M.K. Barton, 2007, A Feedback Regulatory Module Formed by LITTLE ZIPPER and HD-ZIPIII Genes.  Plant Cell 2007
Keywords: screen for downstream genes, transcription factor induction

overall_design: One of the five Arabidopsis HD-ZIPIII proteins, REVOLUTA, was placed under glucocorticoid control by fusing the glucocorticoid receptor (GR) domain to its amino terminus.   This gene fusion was placed under the control of the highly and constitutively expressed viral CaMV35S promoter.  Since the HD-ZIPIII genes are controlled by microRNAs, it was also necessary to introduce mutations that disrupted the microRNA complementary sequence.  Transgenic Arabidopsis seedlings carrying this construct (GR-REV) as well as wild-type seedlings were grown in liquid culture for ten days.  At this point, one flask each of wt and transgenic seedlings was treated with carrier alone (ethanol) and one flask each of wt and transgenic seedlings was treated with dexamethasone.  RNA was harvested from these samples after one hour, labeled and hybridized to Affymetrix microarrays.

first: Kathryn

first: Stephan

last: Wenkel

last: Emery

first: Bi-Huei

last: Hou

first: Khar-Wai

last: Lye

first: Matthew

last: Evans


title: Expression data from flowers of two isogenic lines J10AB and J03AB at different developmental stages

status: Public on Dec 07 2007

city: Wuhan

first: hua

middle: cai

last: dong

state: Hubei

address: Xudong Second Road No2

department: Genomics and Molecular Biology

institute: Institute of Oil Crops Research of CAAS

summary:
Dominant genic male sterility (DGMS) is a valuable system for hybrid seed production and improvement of plant populations, but the molecular mechanism of DGMS is not well understood. To facilitate the analysis of mechanism underlying DGMS, we compared gene expression profiles between fertile and sterile flowers of two pair of Brassica napus isogenic lines: J10AB and J03AB which only differ in sterility-related locus and fertility restore-related locus respectively by using Arabidopsis thaliana ATH-1 microarray.
Gene expression profiling revealed that 360 genes differed in expression between the two lines and the number of differently expressed genes increased with progressing developmental stages. Cluster analysis result indicates that most important gene expression changes occur from PMC (anther less than 1mm) to uninucleate stage (anther in 3mm). There are 182 differential expressed genes detected between fertile plants and sterile plants at same stage.  Some important pathways related to fertility control by KEGG analysis, such as glycolysis/gluconeogenesis pathway, flavonoid biosynthesis pathway and fatty acid metabolism pathway. Another abortion usually associated with suppression of gene expression of these pathways.
Keywords: Time course analysis

overall_design: The Homozygous DGMS two-type line J10AB was compared with heterozygous two-type line J03AB at three different stages of development: tetrad stage, uninucleate stage and trinucleate stage (Flower buds <1, 1-3 and >3mm, respectively) which were confirmed by cytology observation. All these flower buds were collected by hand. For each stage, two biological repetitions and two experimental repetitions were used, resulting in four hybridizations per developmental stage. Gene expression was compared between lines at different stages of development, as well as between developmental stages of fertile plants and sterile plants of each line.

first: Caihua

last: Dong

first: Shengwu

last: Hu

first: Lei

last: Shi

first: Jin

last: Zhou

first: Yunfang

last: Fan

first: Shengyi

last: Liu


title: Gene expression abnormalities in histologically normal breast epithelium of breast cancer patients

status: Public on Dec 10 2007

city: Boston

first: Marc

middle: E.

last: Lenburg

address: 715 Albany Street, E613B

department: Genetics and Genomics

institute: Boston University School of Medicine

summary:
Normal-appearing epithelium of cancer patients can harbor occult genetic abnormalities. Data comprehensively comparing gene expression between histologically normal breast epithelium of breast cancer patients and cancer-free controls are limited. The present study compares global gene expression between these groups. We performed microarrays using RNA from microdissected histologically normal terminal ductal-lobular units (TDLU) from 2 groups: (i) cancer normal (CN) (TDLUs adjacent to untreated ER1 breast cancers (n = 14)) and (ii) reduction mammoplasty (RM) (TDLUs of age-matched women without breast disease (n = 15)). Cyber-T identi?ed differentially expressed genes. Quantitative RT-PCR (qRT-PCR), immunohistochemistry (IHC), and comparison to independent microarray data including 6 carcinomas in situ (CIS), validated the results. Gene ontology (GO), UniProt and published literature evaluated gene function. About 127 probesets, corresponding to 105 genes, were differentially expressed between CN and RM (p < 0.0009, corresponding to FDR <0.10). 104/127 (82%) probesets were also differentially expressed between CIS and RM, nearly always (102/104 (98%)) in the same direction as in CN vs. RM. Two-thirds of the 105 genes were implicated previously in carcinogenesis. Overrepresented functional groups included transcription, G-protein coupled and chemokine receptor activity, the MAPK cascade and immediate early genes. Most genes in these categories were under-expressed in CN vs. RM. We conclude that global gene expression abnormalities exist in normal epithelium of breast cancer patients and are also present in early cancers. Thus, cancer-related pathways may be perturbed in normal epithelium. These abnormalities could be markers of disease risk, occult disease, or the tissue’s response to an existing tumor.
Keywords: disease state analysis

overall_design: 29 samples from histologically normal microdissected breast epithelium are included in this series. 14 samples are from epithelium adjacent to a breast tumor, 15 samples were obtained from patients undergoing reduction mammoplasty without apparent breast cancer

first: Anusri

last: Tripathi

first: Chialin

last: King

first: Antonio

last: de la Morenas

first: Kristina

last: Perry

first: Bohdana

last: Burke

first: Gregory

last: Antoine

first: Erwin

last: Hirsch

first: Maureen

last: Kavanah

first: Jane

last: Mendez

first: Michael

last: Stone

first: Norman

last: Gerry

first: Carol

last: Rosenberg


status: Public on Dec 01 2007

city: Gothenburg

first: Lars

last: Palmqvist

address: Sahlgrenska University Hospital

department: Institute of Biomedicine

country: Sweden

institute: University of Gothenburg

summary:
Physical factors can have major influences on the proliferation and differentiation fate of hematopoietic stem cells in culture. Recently, we demonstrated that cord blood CD34+ cells undergo accelerated and increased megakaryocyte differentiation when incubated at 39°C. In this study, we investigated in detail the impacts of mild hyperthermia on the kinetics of megakaryocyte differentiation, maturation, polyploidization and cell viability. The qualitative and quantitative effects on Mk differentiation were found to be rapidly induced, and optimization of the culture length at 39°C led to greater Mk yields. Mild hyperthermia did not promote endomitosis of cord blood-derived Mk, but rather led to a small reduction in the proportion of polyploid Mk. Moreover, it had little impact on viability but induced a significant shift in the proportion of cells undergoing apoptosis at 37°C to necrosis at 39°C. Finally, our results suggest that the effects on megakaryocyte differentiation could be the consequences of aberrant expression of key megakaryocyte transcription factors induced by mild hyperthermia.
Keywords: Effect of mild hyperthermia on Mk differentiation

overall_design: To define potential differences between megakaryocytes (Mk) derived at 37°C or 39°C, we compared their gene expression repertoire by micro-gene chip technology (Affymetrix GeneChip HG133A). Two independent experiments were carried out on Mk at 37 or 39°C. CD41a+ Mk issued from CB CD34+ cells were cultured for 7-days with TPO at 100 ng/ml and purified by positive magnetic selection using a CD41a-FITC monoclonal antibody (Immunotech, Marseille, France) and MACS columns according to manufacturer’s instructions (Miltenyi, Auburn, CA, USA) with a purity of >95%.

first: Nicolas

last: Pineault

first: Jean-François

last: Boucher

first: Marie-Pierre

last: Cayer

first: Lucie

last: Boyer

first: Réal

last: Lemieux

first: Chantal

last: Proulx


title: Gene expression profiling and correlation with outcome in clinical trials of the proteasome inhibitor bortezomib

status: Public on Dec 06 2007

city: Cambridge

first: George

last: Mulligan

address: 35 Landsdowne Stree

department: Clinical Development

institute: Millennium Pharmaceuticals

summary:
The aims of this study were to assess the feasibility of prospective pharmacogenomics research in multicenter international clinical trials of bortezomib in multiple myeloma and to develop predictive classifiers of response and survival with bortezomib. Patients with relapsed myeloma enrolled in phase 2 and phase 3 clinical trials of bortezomib and consented to genomic analyses of pretreatment tumor samples. Bone marrow aspirates were subject to a negative-selection procedure to enrich for tumor cells, and these samples were used for gene expression profiling using DNA microarrays. Data quality and correlations with trial outcomes were assessed by multiple groups. Gene expression in this dataset was consistent with data published from a single-center study of newly diagnosed multiple myeloma. Response and survival classifiers were developed and shown to be significantly associated with outcome via testing on independent data. The survival classifier improved on the risk stratification provided by the International Staging System. Predictive models and biologic correlates of response show some specificity for bortezomib rather than dexamethasone. Informative gene expression data and genomic classifiers that predict clinical outcome can be derived from prospective clinical trials of new anticancer agents.
Keywords: Gene expression profiling; correlation with outcome in clinical trials of the proteasome inhibitor bortezomib

overall_design:
Purified myeloma samples were collected prior to enrolment in clinical trials of bortezomib (PS-341). Samples were subject to replicate gene expression profiling using the Affymetrix 133A/B microarray. Data was normalized in MAS5.0 and the median of replicates is reported. Data was normalized to a Ttimmed mean of 15o and is NOT log transformed.  Various patient parameters are reported as well as response, TTP and survival upon treatment with bortezomib or dexamethasone.

*** The authors have not submitted the CEL files for GSE9782.  Thus, GSE9782 is incomplete.

first: Barb

last: Bryant


title: Genome-wide transcriptome analysis of Arabidopsis and siz1-3 response to drought stress

first: Rafael

last: Catala

address: 1230 York ave

department: Plan Molecular Biology

institute: The Rockefeller University

summary:
Post-translational modifications of proteins by Small Ubiquitin-like Modifiers (SUMOs) regulate protein degradation and localization, protein-protein interaction, and transcriptional activity. SUMO E3 ligase functions are executed by SIZ1/SIZ2 in yeast and PIAS family members in human. The Arabidopsis genome contains only one gene, SIZ1, that is orthologous to yeast SIZ1/SIZ2. Here, we show that the Arabidopsis SIZ1 interacts with SUM1 and SCE1a, the SUMO E2 conjugating enzyme. Compared to WT, the null mutant siz-1-3 is smaller in statue because of reduced expression of gene involved in brassinosteroid biosynthesis and signalling. Drought stress induces the accumulation of SUMO-protein conjugates, which is in part dependent on SIZ1 but not on ABA. Mutant plants of siz1-3 have significant lower tolerance to drought stress. Genome wide expression analysis identified about 2,000 Arabidopsis genes that are responsive to drought, and SIZ1 mediates the induction of 600 of these genes by a pathway independent of DREB2A and ABA. SIZ1-dependent, drought-responsive genes include those encoding enzymes of the anthocyanin synthesis pathway and jasmonate response. From these results, we conclude that SIZ1 regulates Arabidopsis development and plays a role in drought stress response probably through the control of gene expression.
Keywords: stress response

overall_design: 12 samples that includes 3 replicates of Col 0 under control conditions, 3 replicates of Col 0 expossed to 2 hr of dehydration, 3 replicates of siz1-3 under control conditions and 3 replicates of siz1-3 expossed to 2 hr of dehydration.


title: Identification of putative targets of AP3/PI

status: Public on Nov 28 2007

city: New Haven

first: Chloe

last: Mara

state: CT

address: 160 Prospect Street

department: MCDB

institute: Yale University

summary:
To identify genes regulated by AP3/PI, we carried out microarray experiments using an Arabidopsis whole genome GeneChip array (ATH1 GeneChip, Affymetrix, Santa Clara, CA) in conjunction with an inducible AP3-GR system. For these experiments, we used 35S::AP3-GR transgenic plants in a 35S::PI, ap3-3 null mutant background for various dex or mock treatments. RNA was extracted from inflorescences at 0 and 4 hours after dex or a mock treatment and used as probes for our microarray experiments. Three biological replicates of each were hybridized to Affymetrix ATH1 arrays. We used the Affymetrix Microarray Suite software (MAS) to identify genes whose expression profiles changed only after dex-treatment and are likely targets of AP3/PI.
Keywords: time course

overall_design: For these experiments, we used 35S::AP3-GR transgenic plants in a 35S::PI, ap3-3 null mutant background for various DEX or MOCK treatments. RNA was extracted from inflorescences at 0 and 4 hours after DEX or a MOCK treatment and used as probes for our microarray experiments. Three biological replicates of each were hybridized to Affymetrix ATH1 arrays. We used the Affymetrix Microarray Suite software (MAS 5.0) to identify genes whose expression profiles changed only after DEX-treatment and are likely targets of AP3/PI.

first: Vivian

last: Irish


title: Effects of sex and age on skeletal muscle gene expression in normal men and women

status: Public on Nov 24 2007

city: Rochester

first: Stephen

last: Welle

address: 601 Elmwood Avenue

institute: University of Rochester

summary:
Muscle biopsies taken from vastus lateralis muscle of 15 men and 15 women after 3 days of standardized diet and activity to examine effects of sex and age
Subjects were either young adults (7 men and 7 women, 20-29 yr old) or older (8 men and 8 women, 65-75 yr old)
Affymetrix U133A and U133B arrays were scanned both before (S1) and after (S2) antibody enhancement. This file has U133A S1 data.
Effects of age were computed for men and women separately with an alternative method previously and data submitted as GSE362 (men) and GSE674 (women).
Keywords: sex differences, age-related differences

overall_design: Gene expressions profiles were generated for each individual for a total of 30 profiles (15 male, 15 female, 14 younger, 16 older)


title: Detection of genes differentially expressed in radioresistant and radiosensitive tumors before and after irradiation

city: Chicago

first: Nikolai

middle: N.

last: Khodarev

state: IL

address: 5841 South Maryland Avenue

department: Radiation and Cellular Oncology

institute: The University of Chicago

summary:
Nu61, a radiation-resistant human tumor xenograft, was selected from a parental radiosensitive tumor SCC-61 by eight serial cycles of passage in athymic nude mice and in vivo irradiation.
Obtained tumors were profiled using Affymetrix U133A arrays. Most abundant gene pattern associated with radioresistant phenotype was presented by IFN-inducible, Stat1-dependent pathway
Keywords: Pair-wise comparison of radiosensitive vs radioresistant tumors; time course of irradation response

overall_design: Obtained tumors were established as xenografts in nude mice. RNA were purified from each xenograft, normalized by concentration and used for hybridization with each array. Three arrays were used for each group (un-treated nu61 and SCC61 and irradiated nu61 and SCC61 5 and 24 hours post-IR). Each group contained from 5 animals; tumors were randomized by size and three tumors from each group were used for profiling

middle: N

last: Beckett

first: Edwardine

last: Labay

first: Thomas

last: Darga

first: Helena

middle: J

last: Mauceri

first: Elena

middle: V

last: Efimova

first: Sean

last: Pitroda

first: Andy

last: Minn

first: Bernard

last: Roizman

first: Ralph

last: Weichselbaum


title: Interferon response of radioresistant and radiosensitive human head&neck tumor cell lines

summary:
Nu61, a radiation-resistant human tumor xenograft, was selected from a parental radiosensitive tumor SCC-61 by eight serial cycles of passage in athymic nude mice and in vivo irradiation.
Obtained tumors xenografts were profiled in two independent experiments using Affymetrix U133A arrays. Most abundant gene pattern associated with radioresistant phenotype was presented by IFN-inducible, Stat1-dependent pathway. In these experiments we detected genes responding to IFN alpha, beta and gamma in nu61 and SCC61
Keywords: Pair-wise comparison of un-treated and treated tumor cell lines

overall_design: Cell lines derived from nu61 and SCC61 were grown in vitro and treated by the mixture of IFNalpha, IFNbeta and IFN gamma (100,100 and 5ng/ml respectively). 5 hours after administration of IFNs cells were lysed, RNA collected and used for hybridization with U133A chips to detect genes responding to IFN treatment in nu61 and SCC61

first: Michael


title: Detection of genes differentially expressed in radioresistant tumors

summary:
Nu61, a radiation-resistant human tumor xenograft, was selected from a parental radiosensitive tumor SCC-61 by eight serial cycles of passage in athymic nude mice and in vivo irradiation.
Obtained tumors were profiled using Affymetrix U133A arrays. Most abundant gene pattern associated with radioresistant phenotype was presented by IFN-inducible, Stat1-dependent pathway
Keywords: Pair-wise comparison of radiosensitive vs radioresistant tumors

overall_design: Obtained tumors were established as xenografts in nude mice. RNA were purified from each xenograft, normalized by concentration and three samples of RNA from each group were pooled for hybridization with each array. Three arrays were used for each group.


title: Radioresistant and radiosensitive tumors and cell lines

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Hypoxia

status: Public on Dec 11 2007

city: Toronto

first: Sara

last: Arab

state: ONT

address: 610 University Avenue

department: Cardiology

country: Canada

institute: Toronto General Hospital

summary:
Systemic arterial smooth muscle cells are exposed to a broad range of oxygen concentrations under physiological conditions.  Hypoxia can modulate the proliferative response of smooth muscle cells leading to speculation about its role in vasculogenesis, vascular remodelling and the pathogenesis of arterial disease.  The effect of hypoxia has been inconsistent, however, with both enhanced proliferation and growth arrest reported.   Nevertheless, these reports support an important effect of hypoxia on smooth muscle cell proliferation and, given its physiological and clinical relevance, this requires clarification.  We posited that variation in O2 concentration, within the range that exists in vivo, may have different effects on the proliferation and survival of vascular smooth muscle cells.
Keywords: expression profiles of hypoxia (1% and 3%) in HASMC

overall_design: Human aortic smooth muscle cells (HASMC) were propagated to passage 6 in SMGM-2 medium reached 80% confluence, the media was changed and the cells were incubated for a further 16 hrs or 48 hrs under either normoxic or hypoxic conditions (1%  and 3%O2 ).

first: Julie

last: Basuray

last: Ward

first: Peter

last: Liu


title: Genetic Endothelial Systems Biology of Sickle Stroke Risk

status: Public on Dec 18 2007

city: Minneapolis

first: Robert

middle: P.

last: Hebbel

state: MN

address: MMC 480, 420 Delaware St. SE

department: Medicine

institute: University of Minnesota

summary:
Genetic differences in endothelial biology could underlie development of phenotypic heterogeneity amongst individuals afflicted with vascular diseases. We obtained BOEC (blood outgrowth endothelial cells) from 20 subjects with sickle cell anemia (age 4-19) shown to be either at-risk (n=11) or not-at-risk (n=9) for ischemic stroke due to, respectively, having or not having occlusive disease at the Circle of Willis (CoW).
Gene expression profiling identified no significant single gene differences between the two groups, as expected. However, analysis of Biological Systems Scores, using gene sets that were pre-determined to survey each of nine biological systems, showed that only changes in inflammation signaling are characteristic of the at-risk subjects, as supported by multiple statistical approaches
Keywords: disease status analysis

overall_design: We obtained BOEC (blood outgrowth endothelial cells) from 20 subjects with sickle cell anemia (age 4-19) shown to be either at-risk (n=11) or not-at-risk (n=9) for ischemic stroke due to, respectively, having or not having occlusive disease at the Circle of Willis (CoW). To allow power calculations to be done, we performed microarray analysis on BOEC from 27 normal subjects of diverse ages. Gene expression profilings were obtained by using Affymetrix U133A chips

first: Liming

last: Milbauer

first: Peng

last: Wei

first: Judy

last: Enenstein

first: Aixiang

last: Jiang

first: Cheryl

last: Hillery

first: J

middle: P

last: Scott

last: Nelson

first: Vidya

last: Bodempudi

first: James

last: Topper

first: Ruey-Bing

last: Yang

first: Betsy

last: Hirsch

first: Wei

last: Pan


title: Early gibberellin responses and DELLA protein direct targets in Arabidopsis

status: Public on Nov 21 2007

city: Durham

first: Rodolfo

last: Zentella

state: NC

address: Box 90338, Science Dr

institute: Duke University

first: Zhong-Lin

last: Zhang

first: Mehea

last: Park

first: Stephen

last: Thomas

first: Akira

last: Endo

first: Kohji

last: Murase

first: Christine

middle: M

last: Fleet

first: Yusuke

last: Jikumaru

first: Eiji

last: Nambara

first: Yuji

last: Kamiya

first: Tai-ping

last: Sun


title: Early gibberellin responses in Arabidopsis

summary:
The aim is to identify early gibberellin responsive genes in a gibberellin deficient strain such as ga1-3. Such genes are likely regulated by DELLA proteins which are master gibberellin repressors. DELLA proteins are rapidly degraded after gibberellin treatment, but their direct target genes still need to be elucidated.
Keywords: hormone response, gibberellin treatment

overall_design: A set of 4 biological replicates was generated for each treament. Arabidopsis seedlings were treated with water or 2 uM GA4 and whole shoots collected after 1h. A comparison of water vs. GA4 treated samples should render a list of early gibberellin responsive genes.


title: Letrozole (Femara) early response to treatment

first: John

last: Walker

address: 10675 John Jay Hopkins

institute: Genomics Institute of the Novartis Research Foundation

summary:
In the present investigation, we have exploited the opportunity provided by neoadjuvant treatment of a group of postmenopausal women with large operable or locally advanced breast cancer (in which therapy is given with the primary tumour remaining within the breast) to take sequential biopsies of the same cancers before and after 10-14 days treatment with letrozole.  RNA extracted from the biopsies has been subjected to Affymetrix microarray analysis and the data from paired biopsies interrogated to discover genes whose expression is most influenced by oestrogen deprivation.
Keywords: time course

overall_design: biopsies were taken from the same subjects both pretreatment and after 10-14 days Letrozol, 2.5 mg/day, oral

first: William

last: Miller

first: Alexey

last: Larionov

first: Lorna

last: Renshaw

first: Thomas

last: Anderson

first: Sharon

last: White

first: Garret

last: Hampton

first: Steven

last: Ho

first: Andreas

last: Krause

first: Dean

last: Evans

first: J

last: Dixon


title: Phenotypic and transcriptional response to selection for alcohol sensitivity in Drosophila melanogaster

status: Public on Nov 20 2007

city: Raleigh

first: Tatiana

last: Morozova

address: campus box 7617

department: zoology

institute: NCSU

summary:
Alcoholism is a complex disorder determined by interactions between genetic and environmental risk factors. Drosophila represents a powerful model system to dissect the genetic architecture of alcohol sensitivity, as large numbers of flies can readily be reared in defined genetic backgrounds and under controlled environmental conditions. Furthermore, flies exposed to ethanol undergo physiological and behavioral changes that resemble human alcohol intoxication, including loss of postural control, sedation, and development of tolerance.
We performed artificial selection for alcohol sensitivity for 25 generations and created duplicate selection lines that are either highly sensitive or resistant to ethanol exposure along with unselected control lines. We used whole genome expression analysis to identify 1,678 probe sets with different expression levels between the divergent lines, pooled across replicates, at a false discovery rate of q < 0.001. We assessed to what extent genes with altered transcriptional regulation might be causally associated with ethanol sensitivity by measuring alcohol sensitivity of 37 co-isogenic P-element insertional mutations in 35 candidate genes, and found that 32 of these mutants differed in sensitivity to ethanol exposure from their co-isogenic controls. Furthermore, 23 of these novel genes have human orthologues.
Combining whole genome expression profiling with selection for genetically divergent lines is an effective approach for identifying candidate genes that affect complex traits, such as alcohol sensitivity. Because of evolutionary conservation of function, it is likely that human orthologues of genes affecting alcohol sensitivity in Drosophila may contribute to alcohol-associated phenotypes in humans.
Keywords: artificial selection, whole genome expression profiling

overall_design:
Starting with flies from Raleigh natural population (see material and methods) we performed artificial selection for alcohol sensitivity for 35 generation.  In each generations  we scored 60 males and females, separately, from each line (resistant, sensitive, and control)using inebriometer, and the 20 highest-scoring flies from the resistant lines and the 20 lowest-scoring flies from the sensitive lines were selected as parents for the next generation. Control line flies were scored each generation and 20 random flies were used as parents.
At generation 25, two replicates of 15 three-five day old virgin males and females were collected from each selection line. Total RNA was extracted from the 24 samples . Biotinylated cRNA probes were hybridized to high density oligonucleotide microarrays (Affymetrix, Inc. Drosophila GeneChip 2.0) and visualized with a streptavidin-phycoerythrin conjugate, as described in the Affymetrix GeneChip Expression Analysis Technical Manual (2000), using internal references for quantification. The quantitative estimate of expression of each probe set is the Signal (Sig) metric, as described in the Affymetrix Microarray Suite, Version 5.0.

first: Robert

last: Anholt

first: Trudy

middle: F

last: Mackay


title: Early Gene Expression Profiles During Intraoperative Myocardial Ischemia-Reperfusion in Cardiac Surgery

status: Public on Nov 19 2007

first: Sara

last: Arab

summary:
Right ventricular samples were serially acquired during surgical repair of ventricular septal defect. Expression profiling revealed three patterns of gene expression: (1) increased expression above control levels within one hour of cardioplegic arrest, with further amplification during early reperfusion; (2) increased expression limited to the reperfusion phase; and (3) reduced expression during reperfusion.
Keywords: VSD Patients

overall_design:
4 replicates each:
Stage2, End-ischemia
Stage1, Immediately post-cardioplegic arrest
Stage3, After 5 min. Reperfusion


title: DELLA protein direct targets in Arabidopsis

first: Rodolfo

last: Zentella

summary:
The aim of this study is to identify early DELLA protein-responsive genes using a Dexamethasone (DEX)-inducible system. Two transgenic lines were used: one induces the expression of a dominant, gibberellin non-responsive DELLA protein (rga-delta17); the other is a control line that carries the same vector, but lacks the rga-delta17 transgene. By comparing the gene expression changes in the line that expresses the rga-delta17 protein in the presence or absence of DEX it is possible to identify putative targets of DELLA proteins. An empty vector transgenic line was included in this study to identify genes that might be regulated by the DEX inducible system that are not dependent on the DELLA protein.
Keywords: Dexamethasone treatment, gibberellin treatment, time course, transgene effect

overall_design: Seedlings of both transgenic lines were pretreated for 16 h with 2 uM GA4 to enhance gibberellin responses. Because DELLA proteins are strong signaling repressors, this pretreatment should maximize the effect of DELLA induction. Eight-day old seedlings were treated with 2 uM GA4 or a combination of 2 uM GA4 plus 10 uM DEX to induce the rga-delta17 transgene. Three biological replicas for the transgenic line that carries the DEX-inducible rga-delta17 transgene were generated at 2h and 4h. For the empty vector line, only 2 biological replicas were generated at 4h of treatment with 2 uM GA with or without 10 uM DEX.

first: Zhong-Lin

last: Zhang

first: Mehea

last: Park

first: Stephen

last: Thomas

first: Akira

last: Endo

first: Kohji

last: Murase

first: Christine

last: Fleet

first: Yusuke

last: Jikumaru

first: Eiji

last: Nambara

first: Yuji

last: Kamiya


title: Target genes of AGAMOUS during early flower development in Arabidopsis

status: Public on Nov 15 2007

city: NORWICH

last: Sablowski

state: Norfolk

address: The John Innes Centre

department: CDB

institute: JIC

summary:
Floral organs, whose identity is determined by specific combinations of homeotic genes, originate  from a group of undifferentiated cells called the floral meristem. In Arabidopsis, the homeotic gene AGAMOUS (AG) terminates meristem activity and promotes development of stamens and carpels.
To understand the program of gene expression activated by AG, we followed genome-wide expression during early stamen and carpel development.
Keywords: Developmental time course

overall_design: Described in Gomez-Mena et al, 2005, Development 132: 429-438. Briefly, synchronous development of stamens and carpels was initiated by steroid treatment of plants homozygous for the ap1-1 and cal-1 mutations and expressing a fusion between AGAMOUS and the rat glucocorticoid receptor (35S:AGGR). RNA was extracted one, three and seven days after steroid treatment; two independent steroid-treated samples and two independent untreated controls were used for each time point.


title: Identification of Transcriptional Targets Dual Functions Transcription Factor/Phosphatase Eyes Absent

status: Public on Nov 07 2007

first: Ilaria

last: Rebay

email: irebay@uchicago.edu, jcj27@mit.edu

address: 929 East 57th Street Room W340

department: Ben May Department for Cancer Research

institute: University of Chicago

summary:
Drosophila eye specification an development relies on a collection of transcription factors termed the retinal determination gene network (RDGN).  Two members of this network, Eyes absent (EYA) and Sine oculis (SO), form a transcriptional complex in which EYA provides the transactivation function while SO provides the DNA binding activity.  EYA also function as a protein tyrosine phosphatase, raising the question of whether transcriptional output is dependent or independent of phosphatase activity.  To explore this, we used microarrays together with binding site analysis, quantitative real-time PCR, chromatin immunoprecipitation, genetics, and in vivo expression analysis to identify new EYA-SO targets.  In parallel, we examined the expression profiles of tissue expressing phosphatase mutant eya and found that reducing phosphatase activity did not globally impair transcriptional output.  Among the targets identified by our analysis was the cell cycle regulatory gene, string (stg), suggesting that EYA and SO may influence cell proliferation through transcriptional regulation of stg.  Future investigation into the regulation of stg and other EYA-SO targets identified in this study will help elucidate the transcriptional circuitries whereby output from the RDGN integrates with other signaling inputs to coordinate retinal development.
Keywords: gain of function, expression comparison

overall_design: We compared gene expression across duplicate samples for controls in which eya was not overexpressed to duplicate samples in which a wildtye overexpressed eya transgene, an eya mutant transgene with the D493N mutation, which reduces phosphatase activity, and a second eya mutant transgene with the E728Q mutation, which has little is any posphatase activity.

first: Jennifer

last: Jemc


title: Differential gene expression in peripheral blood leukocytes from CHASRS particpants

status: Public on Nov 02 2007

city: Los Angeles

first: Steve

last: Cole

address: 11-934 Factor Bldg, UCLA

institute: UCLA School of Medicine

summary:
Gene expression profiling was carried out on peripheral blood leukocytes from 14 healthy older adults.  The primary research question is whether gene expression differs in individuals experiencing chronically high levels of social isolation (by UCLA Loneliness Scale) vs chronically low levels of social isolation.
Keywords: Risk prediction


title: Alterations in gene expression in Drosophila melanogaster embryo exposed 500µGy gamma rays at 22.4mGy/h

status: Public on Nov 09 2007

city: Rokkasho-mura,Kamikita-gun,

first: Keiji

last: Ogura

state: Aomori

address: Takahoko, Hacchasawa,

department: AMBIC

institute: Institute for Environmental Science

summary:
To determine what genes are affect to the reduction in mutation frequency by 500µGy gamma irradiation of Drosophila melanogaster.
The  strain is w[1118] (a loss of function mutant of compound eye pigment gene white with Canton-S background).
Keywords: time course

overall_design: 15 samples are analyzed. The control and four time course (5, 30, 60 and 90min), 3 Biological rep. (A to C).

first: K

first: J

last: Magae

first: Y

last: Kawakami

first: T

last: Koana


title: A  high resolution organ expression map reveals novel expression patterns and predicts cellular function

status: Public on Nov 05 2007

first: Siobhan

last: Brady

department: Biology

summary:
Transcriptional programs that regulate development are exquisitely controlled in space and time.  Elucidating these programs that underlie development is essential to understanding the acquisition of cell and tissue identity. We present microarray expression profiles of a high resolution set of developmental time points within a single Arabidopsis root, and a comprehensive map of nearly all root cell-types.   These cell-type specific transcriptional signatures often predict novel cellular functions.  A computational pipeline identified dominant expression patterns that demonstrate transcriptional connections between disparate cell types.  Dominant expression patterns along the root’s longitudinal axis do not strictly correlate with previously defined developmental zones, and in many cases, expression fluctuation along this axis was observed. Both robust co-regulation of gene expression and potential phasing of gene expression were identified between individual roots.  Methods that combine these two sets of profiles demonstrate transcriptionally rich and complex programs that define Arabidopsis root development in both space and time.
We used microarrays to profile expression of nearly all cell types in the Arabidopsis root, and to profile at high resolution, developmental time points along the root's longitudinal axis.
Keywords: Cell-type specific analysis using FACS, and a developmental time course.

overall_design: Microarray expression profiles of 8 new GFP marked lines with 2-3 replicates were used to augment existing microarray expression profiles of the root.  RNA isolated from 13 cross sections along a single root's longitudinal axis were also profiled by microarray analysis.  An independent root with 12 sections were used as a biological replicate.

first: David

middle: A

last: Orlando

first: Ji-Young

last: Lee

first: Jean

middle: Y

last: Wang

first: Jeremy

last: Koch

first: Jose

last: Dinneny

first: Daniel

last: Mace

first: Uwe

last: Ohler

first: Philip

last: Benfey


title: RNA polymerase is poised for activation across the genome

status: Public on Nov 11 2007

city: RTP

first: NIEHS

last: Microarray Core

address: 111 T.W. Alexander Drive

department: DIR

institute: NIEHS

type: Genome binding/occupancy profiling by genome tiling array

summary:
Regulation of gene expression is integral to the development and survival of all organisms. Transcription begins with the assembly of a pre-initiation complex at the gene promoter, followed by initiation of RNA synthesis and the transition to productive elongation. In many cases, recruitment of RNA polymerase II (Pol II) to a promoter is necessary and sufficient for activation of gene. However, there are a few notable exceptions to this paradigm, including heat shock genes and several proto-oncogenes, whose expression is attenuated by regulated stalling of polymerase elongation within the promoter-proximal region. To determine the importance of polymerase stalling for transcription regulation, we performed a genome-wide search for Drosophila genes with promoter-proximally stalled Pol II. Our data reveal that stalling is widespread, occurring at hundreds of genes that respond to stimuli and developmental signals, indicating a role for regulation of polymerase elongation in the transcriptional responses to dynamic environmental and developmental cues.
Keywords: gene expression, ChIP-chip, transcriptional regulation

overall_design:
Drosophila Schneider cells (S2) were untreated or treated with dsRNA against LacZ or NELF for 96 hours.

RNA isolation and immunoprecipitations (IPs) were performed for each of the test samples.  Each treatment group (untreated, dsRNA-LacZ, dsRNA-NELF) was cultured in duplicate.

Each RNA sample was amplified according to Affymetrix Eukaryotic One-cycle Protocol and hybridized to an Affymetrix Drosophila Genome 2.0 array, resulting in two biological replicates for each treatment group.

Immunoprecipitations were performed either in the presence of Pol II Rpb3 or Ser2P Pol II CTD antibody or mock-immunoprecipitated with Protein-A agarose in the absence of antibody.

Each IP was paired with a genomic DNA sample obtained at the time of the experiment.   A separate labeling reaction was completed for each array type on which the IP/genomic DNA comparison was made (Agilent Dm3, Dm7 and Whole Drosophila Genome 2 chip set) and each of the two biological replicates were assayed.

first: Ginger

middle: W

last: Muse

last: Gilchrist

first: Sergei

last: Nechaev

first: Ruchir

last: Shau

first: Joel

last: Parker

first: Sherry

last: Grissom

first: Julia

last: Zeitlinger

first: Karen

last: Adelman


title: Identification of genes responding to the activity of the Arabidopsis cytochrome P450 KLUH/CYP78A5

status: Public on Dec 04 2007

city: Norwich

first: Michael

last: Lenhard

address: Colney Lane

department: Cell & Developmental Biology

institute: John Innes Centre

summary:
The Arabidopsis cytochrome P450 KLUH (KLU)/CYP78A5 promotes organ growth in a non-cell autonomous manner. To identify genes regulated by KLU activity, homozygous klu-2 mutants carrying constructs for EtOH-inducible overexpression of wild-type KLU (35S::AlcR-AlcA::KLU) or of enzymatically inactive KLU protein (35S::AlcR-AlcA::KLUmut) were induced with EtOH and sampled at 90 min and 240 min after induction for gene expression changes.
Keywords: Time course, genetic modification

overall_design: For both constructs (i.e. overexpression of wild-type and of mutant KLU protein as a control), two independently transformed lines were used, serving as biological replicates. These were sampled at 90 min and 240 min after induction. In addition, plants with the wild-type construct were also sampled immediately before induction as a control, together resulting in ten hybridizations.

first: Elena

last: Anastasiou

first: Sabine

last: Kenz

last: MacLean


title: Identification of putative Arabidopsis DEMETER target genes by GeneChip Analysis

status: Public on Nov 01 2007

first: Bob

last: Goldberg

address: 610 Charles E Young Drive East

department: Molecular, Cell and Developmental Biology

institute: University of California, Los Angeles

summary:
We used Affymetrix Arabidopsis ATH1 GeneChip to profile RNAs active in wild type columbia (glabrous) and CaMV::DME pollen and stamens.
Keywords: genetic modification

overall_design: DME is not active in wild type pollen and stamen.  DME was induced in pollen and stamen by a cauliflower mosaic virus promoter (CaMV).  RNA profiles from wild type and CaMV::DME pollen and stamens were compared using Affymetrix ATH1 GeneChip.

first: Hyunhwa

last: Ohr

first: Anhthu

middle: Q

last: Bui

first: Brandon

middle: H

last: Le

first: Robert

middle: L

last: Fischer

first: Yeonhee

last: Choi


title: Genome-wide analyses of human perisylvian cerebral cortical patterning

status: Public on Oct 31 2007

first: Brett

middle: S

last: Abrahams

address: 695 Charles Young Drive

institute: UCLA

summary:
Despite the well-established role of the frontal and posterior peri-sylvian cortices in many facets of human-cognitive specializations, including language, little is known about the developmental patterning of these regions in human brain. We performed a genome-wide analysis of human cerebral patterning during mid-gestation, a critical epoch in cortical regionalization. A total of 345 genes were identified as differentially expressed (DE) between superior temporal gyrus (STG) and the remaining cerebral cortex (CTX). GO categories representing transcription factors were enriched in STG, while cell-adhesion and extracellular matrix molecules, were enriched in the other cortical regions. Q-PCR or in situ hybridization were performed to validate differential expression in a subset of 32 genes, most of which were confirmed. LIM domain binding 1 (LDB1), which we show to be enriched in the STG, is a recently identified interactor of LIM domain only 4 (LMO4), a gene known to be involved in the asymmetric pattering of the peri-sylvian region in the developing human brain. Protocadherin 17 (PCDH17), a neuronal cell adhesion molecule, was highly enriched in focal regions of the human prefrontal cortex. Contactin Associated Protein-Like 2 (CNTNAP2), in which mutations are known to cause autism, epilepsy and language delay, showed a remarkable pattern of anterior enriched expression in cortical regions important for human higher cognition. Importantly, a similar pattern was not observed in the mouse or rat. These data highlight the importance of expression analysis of human brain and the utility of cross-species comparisons of gene expression. Genes identified here provide a foundation for understanding molecular aspects of human-cognitive specializations and disorders that disrupt them.
Keywords: Regional contrast

overall_design: Fresh frozen human mid-gestation brains were obtained from the NICHD Brain and Tissue Bank for Developmental Disorders (University of Maryland, Baltimore, MD). After separation of the two hemispheres, tissue from STG was extracted. RNA from STG and remaining CTX was hybridized on both Agilent G4110A arrays (n=8) and Affymetrix HGU133A arrays (n=17).

first: Dima

last: Tentler*

first: Julie

last: Perederiy*

middle: C

last: Oldham

first: Giovanni

last: Coppola

first: Daniel

last: Geschwind


title: DNA methyltransferase 3B (DNMT3B) mutations in ICF syndrome: gene expression analysis

city: Gainesville

first: bilian

last: jin

state: FL

address: 1601 SW Archer Road

institute: University of Florida

summary:
Genome-wide DNA methylation patterns are established and maintained by the coordinated action of three DNA methyltransferases, DNMT1, DNMT3A, and DNMT3B. DNMT3B hypomorphic germline mutations are responsible for two-thirds of Immunodeficiency, Centromere Instability, Facial Anomalies (ICF) syndrome cases. The molecular defects in transcription, DNA methylation, and chromatin structure in ICF cells remain relatively uncharacterized.
We used expressing microarrays to define the global program of gene expression to elucidate the role of DNMT3B in these processes using EBV-immortalized lymphoblastoid cell lines (LCLs) derived from ICF syndrome and normal individuals.
Keywords: disease-state analysis

overall_design: 5 Normal LCLs (GM08729, GM08728, LCL1, CM304 and CM774) and 3 ICF LCLs (4088,GM08714 and 10759) were selected  for RNA extraction and hybridization on Affymetrix UU133A/B microarrays.

first: Keith

middle: D

last: Robertson

first: Bilian

last: Jin


title: Low R:FR treatment at 16 and 22 degrees

status: Public on Oct 28 2007

city: Leicester

first: Keara

last: Franklin

address: University Road

institute: University of Leicester

summary:
The purpose of this experiment was to identify genes responding differently to a 24 h low red to far red ratio (R:FR) treatment in plants grown at 16 and 22 degrees
Keywords: Light treatment

overall_design: Microarrays were used as a gene discovery tool to identify genes showing differential regulation by low R:FR at 2 growth temperatures. Plants subject to a 24 h low R:FR light treatment were compared with control plants maintained in high R:FR. This experiment was carried out at two growth temperatures, 16oC and 22oC.

first: Garry

last: Whitelam


title: Trovafloxacin-Induced Gene Expression Changes: Comparison of Primary Human Hepatocytes to HepG2 Cells

status: Public on Oct 30 2007

city: Abbott Park

first: Michael

last: Liguori

address: 100 Abbott Park Rd

department: Cellular and Molecular Toxicology

institute: Abbott Laboratories

summary:
Primary human hepatocytes (PHH) are a main instrument in drug metabolism research and in the prediction of drug-induced phase I/II enzyme induction in humans.  The HepG2 liver-derived cell line is commonly used as a surrogate for human hepatocytes, but their use in ADME and toxicity studies can be limited because of lowered basal levels of metabolizing enzymes.  Despite their widespread use, the transcriptome of HepG2 cells compared to PHH is not well characterized.  In this study, microarray analysis was conducted to ascertain the differences and similarities in mRNA expression between HepG2 cells and human hepatocytes before and after exposure to a panel of fluoroquinolone compounds. Comparison of the naïve HepG2 cell and PHH transcriptomes revealed a substantial number of basal gene expression differences.  When HepG2 cells were dosed with a series of fluoroquinolones, trovafloxacin, which has been associated with human idiosyncratic hepatotoxicity, induced substantially more gene expression changes than the other quinolones, similar to previous observations with PHH. While TVX-treatment resulted in many gene expression differences between HepG2 cells and PHH, there were also a number of TVX-induced commonalities, including genes involved in RNA processing and mitochondrial function.   Taken together, these results provide insight for interpretation of results from drug metabolism and toxicity studies conducted with HepG2 cells in lieu of PHH, and could provide further insight into the mechanistic evaluation of TVX-induced hepatotoxicity.
Keywords: Cell Type Comparison

overall_design:
HepG2 cells were obtained from the American Type Culture Collection (Rockville, MD).  The cells were cultured in Minimum Essential Medium (Invitrogen Life Technologies, Carlsbad,CA) with 10% Fetal Bovine Serum under a humidified 5% CO2 atmosphere using T-162 plastic culture flasks.  The cells were split when they reached approximately 70-85% confluence after washing with sterile phosphate buffered saline and detachment of the cells with trypsin (Invitrogen Life Technologies, Carlsbad, CA).  The HepG2 cells were cultured in 6-well plastic plates upon exposure to quinolone compounds.  Primary human hepatocytes, obtained from In Vitro Technologies  (IVT, Baltimore, MD) in 6-well type I collagen coated plates, were cultured with 2 mL of Hepatocyte Incubation Media (IVT) at 37°C with 5% CO2 for 24 hours after receipt.

For the genomic experiments, quinolone compounds, dissolved in 0.1 N KOH (Sigma Chemical Co., St. Louis, MO), were added to the wells with fresh media at levels of 100 µM (HepG2) or 400 µM (primary human hepatocytes) for 24 hours using at least two technical replicates.  Trovafloxacin was dosed using two separate preparations of HepG2 cells and two separate donors of human hepatocytes.  Vehicle control cells were dosed with an equivalent volume of 0.1N KOH as the experimental samples.  For intracellular comparison (HepG2 cells vs PHH), naïve cells were harvested using TRIzol™ reagent (Invitrogen Life Technologies, Carlsbad, CA).

Total RNA was isolated from the TRIzol™ extracts using the standard procedure from the manufacturer.  O.D. at 260 nm determined RNA concentrations.  RNA quality was accessed using an Agilent Technologies bioanalyzer before proceeding to microarray sample preparation. Microarray analysis was performed using the standard protocol provided by Affymetrix Inc. (Santa Clara, CA) and as previously described, starting with 5 µg of total RNA (Richert et al., 2006).

Fragmented, labeled cRNA was hybridized to an Affymetrix human genome U133A array, which contains sequences corresponding to roughly 22,200 transcripts at 45°C overnight.  The arrays were washed, developed, and scanned.

first: Eric

last: Blomme

first: Jeffrey

last: Waring


title: Gene Signature for Aggression of Melanoma Metastases - Melanoma Metastasis (LeiATCC)

status: Public on Oct 02 2007

city: cambridge

first: steve

last: shen

address: 77 mass ave

institute: MIT

summary:
Metastasis is the deadliest phase of cancer progression.  Experimental models using immunodeficient mice have been used to gain insights into the mechanisms of metastasis.  We report here the identification of a “metastasis aggressiveness gene expression signature” derived using human melanoma cells selected based on their metastatic potentials in a xenotransplant metastasis model.  Comparison with expression data from human melanoma patients shows that this metastasis gene signature correlates with the aggressiveness of melanoma metastases in human patients.  Many genes encoding secreted and membrane proteins are included in the signature, suggesting the importance of tumor-microenvironment interactions during metastasis.
Keywords: disease state

overall_design: parental cell line vs xenograft model

first: Lei

last: Xu

first: Steven

last: Shen

first: Y

last: Hoshida

first: A

last: Subramanian

first: K

last: Ross

first: JP

last: Brunet

first: S

last: Wagner

last: Ramaswamy

first: J

last: Mesirov

first: R

middle: O

last: Hynes


title: Reversible and Permanent effects of Tobacco Smoke Exposure on Airway Epithelial Gene Expression

city: Boston

first: Avrum

last: Spira

address: 715 Albany Street, R304

department: Pulmonary and Critical Care Medicine

institute: Boston University Medical Center

summary:
RNA was obtained from histologically normal bronchial epithelium of never, former, and current smokers undergoing fiberoptic bronchoscopy.
Statistical analysis of the gene expression data identified gene differentially expressed between current and never smokers and classified these genes as irreversible, slowly reversible, or rapidly reversible based on their behavior in former smokers
Keywords: Disease state analysis

overall_design: Microarrays run on total RNA obtained from Bronchial Epithelium of Never Smokers (n=21), Former Smokers (n=31), and Current Smokers (n=52)

first: Jennifer

last: Beane


title: Gene Signature for Aggression of Melanoma Metastases - Melanoma Metastasis (LeiFidler)

summary:
Metastasis is the deadliest phase of cancer progression.  Experimental models using immunodeficient mice have been used to gain insights into the mechanisms of metastasis.  We report here the identification of a “metastasis aggressiveness gene expression signature” derived using human melanoma cells selected based on their metastatic potentials in a xenotransplant metastasis model.  Comparison with expression data from human melanoma patients shows that this metastasis gene signature correlates with the aggressiveness of melanoma metastases in human patients.  Many genes encoding secreted and membrane proteins are included in the signature, suggesting the importance of tumor-microenvironment interactions during metastasis.
Keywords: disease state analysis

first: L

middle: N


title: Methylmercury treatment (rand-affy-droso-366220)

status: Public on Oct 11 2007

city: Phoenix

first: Winnie

last: Liang

state: AZ

address: 445 N. Fifth Street

institute: Translational Genomics

summary:
Methylmercury (MeHg) toxicity in humans manifests deficits in neurological function.  Cases of prenatal exposure to mercury have established that the developing nervous system is most highly susceptible to perturbation by MeHg.  At a cellular level, MeHg-induced defects result from altered neuronal proliferation, migration and pathfinding.  However, the molecular targets of MeHg that give rise to these outcomes are not fully understood.  In an overall effort to identify the fundamental molecular targets of MeHg in neural development, we are investigating the effects of MeHg on gene expression and protein function in the Drosophila model.  Since the fundamental signaling pathways in development of multicellular animals have been extensively characterized in Drosophila, and demonstrate  high degree of conservation in vertebrates, we believe these data will lead us to the most pertinent pathways affected by MeHg and begin to elucidate the mechanism of MeHg neural toxicity relevant to cases of  human exposure to this prevalent environmental toxin.
Our aim is to identify fundamental signaling pathways in neural development that are targets for MeHg poisoning.
Our hypothesis is that MeHg, by direct interaction with cysteine thiol groups,  alters the function signaling pathway proteins and subsequently alters transcription of target genes in the respective pathways.  Our hypothesis is supported by our recent data demonstrating a direct action of MeHg in activating the Notch receptor pathway and upregulating target gene expression.  We anticipate a microarray analysis will elucidate additional fundamental signaling pathways where transcription is affected by this toxin.
Fertilized female flies are fed on food containing methylmercury (1-20 micromolar) or solvent control (DMSO) for a period of five days to allow for MeHg penetration in to eggs.  Flies are then transfered to a cage for embryo collection for a discrete window of time (e.g. 1hr).  Embryos are aged 16-24 hours and collected for total RNA extraction using the Trizol reagent.  Total RNA is quantitated by spectrophotometry with a Nanodrop reader.  While concentration dependent effects of MeHg are of interest, initial experiments will be conducted on samples exposed to a single concetration known to illicit effect in other bioassays (e.g. 10 micromolar).  The goal is to see which genes are up- or down-regulated with MeHg exposure, as compared to control embryos that are not exposed.  Emphasis in the analysis stage will be in identifying target genes of known signaling pathways.
Keywords: dose response

first: Matthew

last: Rand


title: Effect of Acute Physiologic Hyperinsulinemia on Gene Expression in Human Skeletal Muscle in vivo

status: Public on Sep 20 2007

city: San Antonio

first: Dawn

middle: K

last: Coletta

state: TX

address: 7703 Floyd Curl Drive

institute: UTHSCSA

summary:
This study was undertaken to test the hypothesis that short term exposure (4 hours) to physiologic hyperinsulinemia in normal, healthy subjects without a family history of diabetes would induce a low grade inflammatory response, independently of glycemic status. We performed euglycemic hyperinsulinemic (80 mU/m2/min) clamps in 12 healthy, insulin sensitive subjects with no family history of diabetes followed by biopsies of the vastus lateralis muscle taken basally and after 30 and 240 minutes of insulin infusion. Gene expression profiles were generated using Affymetrix HG-U133A arrays. No probe sets had significantly altered expression at 30 minutes of the insulin clamp, but 121 probe sets (117 upregulated and 4 downregulated) were significantly altered after 240 minutes. Hyperinsulinemia in normal, healthy human subjects increased the mRNAs for a number of inflammatory genes and transcription factors. Microarray and quantitative RT-PCR revealed the upregulation of chemokine, cc motif, ligand 2 (CCL2), CCL8, thrombomodulin (THBD), ras-related associated with diabetes (RRAD), metallothionein (MT), and serum/glucocorticoid regulated kinase (SGK), and downregulation of CITED2 (a CREB-binding protein-interacting transactivator), a known coactivator of PPAR-alpha. Interestingly, SGK and CITED2 are located at chromosome 6q23, where we previously detected strong linkage to hyperinsulinemia. A control saline infusion performed on 3 normal, healthy subjects without a family history of diabetes demonstrated that the genes altered following the euglycemic-hyperinsulinemic clamp were due to insulin and independent of biopsy removal. This study demonstrates that insulin acutely regulates the expression of genes involved in inflammation and transcription, and identifies several candidate genes/pathways for further investigation.
Keywords: Gene Expression, Skeletal Muscle, Insulin Action, Euglycemic Hyperinsulinemic Clamp, Inflammation

overall_design: Twelve subjects received a vastus lateralis muscle biopsy followed by a 180-min euglycemic, hyperinsulinemic (80 mU/m2.min) clamp. Muscle biopsies from each subject were homogenized in RNAStat solution (Tel-Test Inc., Friendswood, TX), using a Polytron homogenizer (Brinkmann Instruments Westbury, NY).  Total RNA was purified with RNeasy and DNase I treatment (Qiagen, Chatsworth, CA).  RNA was prepared for hybridization to Affymetrix (Santa Clara, CA) HG-U133A arrays according to the manufacturer’s instructions.


title: Epigenetic upregulation of B-cell inappropriate genes induces extinction of B-cell program in classical Hodgkin lymphoma

city: Berlin

first: Anke

last: Ehlers

state: Berlin

address: Hindenburgdamm 30

department: Institute of Pathology

institute: Charité Universitätsmedizin

summary:
A unique feature of the tumour cells (Hodgkin/Reed-Sternberg (HRS)) of classical Hodgkin lymphoma (cHL) is the loss of their B-cell phenotype despite their B-cell origin. Several lines of evidence suggest that epigenomic events, especially promoter DNA-methylation, are involved in this silencing of many B-cell associated genes. Here we show that DNA-demethylation alone or in conjunction with histone-acetylation is not able to reconstitute the B-cell gene expression program in cultured HRS cells. Instead, combined DNA-demethylation and histone-acetylation of B cells induce a nearly complete extinction of their B-cell expression program and a tremendous up-regulation of numerous cHL characteristic genes including key players such as Id2 known to be involved in the suppression of the B-cell phenotype. Since the up-regulation of cHL characteristic genes and the extinction of the B-cell expression program occurred simultaneously, epigenetic changes may also be responsible for the malignant transformation of cHL. The epigenetic up-regulation of cHL characteristic genes thus play - in addition to promoter DNA-hypermethylation of B-cell associated genes – a pivotal role for the reprogramming of HRS cells and explain why DNA-demethylation alone is unable to reconstitute the B-cell expression program in HRS cells.
Keywords: Epigenetic modification

overall_design:
5-aza-dC treatment:
The cHL derived cell lines L428, KMH2 and L1236 were treated with 5-aza-dC at a concentration of 5 µM for 5 days with drug and medium replacement after each 48 hours.

Combined 5-aza-dC/TSA treatment:
The Burkitt lymphoma derived cell lines (Daudi, Namalwa and Raji)were treated with 5-aza-dC at a concentration of 3 µM for 6 days. 5-aza-dC and medium was replaced at day 2 and 5. At the fifth day - in addition to 3 µM 5-aza-dC - cells were incubated for 24 hours with 625 nM TSA.

The gene expression profiles of the untreated and treated cell lines were generated in duplicates.

first: Elisabeth

last: Oker

first: Stefan

last: Bentink

first: Dido

last: Lenze

first: Harald

last: Stein

first: Michael

last: Hummel


title: Gene expression in roots and shoots of plants grown on selenate

status: Public on Oct 24 2007

city: Fort Collins

first: Douglas

last: Van Hoewyk

state: CO

address: 1521 Crestmore PL

department: Biology Department

institute: Colorado State University

summary:
Selenate is chemically similar to sulfate and can be taken up and assimilated by plants via the same transporters and enzymes.  In contrast to many other organisms, selenium (Se) has not been shown to be essential for higher plants.  In excess, Se is toxic and restricts development.  Both Se deficiency and toxicity pose problems worldwide. To obtain better insight into the effects of Se on plant metabolism and into plant mechanisms involved in Se tolerance, the transcriptome of Arabidopsis plants grown with or without selenate was studied, and Se-responsive genes identified.  Roots and shoots exhibited different Se-related changes in gene regulation and metabolism. Many genes involved in sulfur (S) uptake and assimilation were upregulated. Accordingly, Se treatment enhanced sulfate levels in plants, but the quantity of organic S metabolites decreased.  Transcripts regulating the synthesis and signaling of ethylene and jasmonic acid were also upregulated by Se.    Selenate appeared to repress plant development, as suggested by the down-regulation of genes involved in cell wall synthesis and auxin-regulated proteins.  The Se-responsive genes discovered in this study may help create plants that can better tolerate and accumulate Se, which may enhance the effectiveness of Se phytoremediation or serve as Se-fortified food.
Keywords: selenate, abiotic stress

overall_design: Arabidopsis thaliana (ecotype WS) plants were vertically grown on +/- 40 micromolar selenate MS medium. Root and shoot tissues of 10-day-old plants were seperated and  used for RNA extraction and hybridization on Affymetrix microarrays. All conditions were duplicated.

first: Doug

first: Hideki

last: Takahashi

first: Eri

last: Inoue

first: Masanori

last: Tamaoki

first: Ann

last: Hess

first: Elizabeth

last: Pilon-Smits