title: Microarray Analysis of Baboon neonates consuming long-chain polyunsaturated fatty acid formula

city: Ithaca

first: Kumar

last: Kothapalli

state: NY

address: Savagae Hall

institute: Cornell University

summary:
Docosahexaenoic acid (DHA, 22:6n-3) and arachidonic acid (ARA, 20:4n-6) are the major long chain polyunsaturated fatty acids (LCPUFA) of the central nervous system. We examined the alterations in transcriptional profiles in neonate baboon cerebral cortex brain tissue using high-density Affymetrix oligonucleotide arrays ("U133 plus 2.0"). Twelve neonates were divided into 3 groups: Control (C, no DHA-ARA); 1× LCPUFA (L, 0.33%DHA-0.67%ARA); 3× LCPUFA (L3, 1.00%DHA-0.67%ARA). Significance analysis (P < 0.05) of a total of >54,000 probe sets (ps) identified changes in expression levels of 1108 ps, representing 2.05% of the total ps on the oligoarray, with most ps showing <2-fold change. Comparing L and C, 534 ps were upregulated with supplementation, and 574 ps were downregulated.  For the L3/C comparisons, 666 ps were upregulated and 442 ps were downregulated.  Characterization of differentially regulated genes by gene ontology assign them to diverse biological functions, notably lipid metabolism, ion channel and transport, G-protein and signal transduction, development, visual perception, membrane function, apoptosis, cytoskeleton, peptidases, cell cycle, stress response, kinases and phosphatases, transcription regulation, receptors, and ubiquitin, with about 400 transcripts having no defined function.  Of differentially expressed genes involved in lipid metabolism, PLA2G6, a phospholipase recently associated with infantile neuroaxonal dystrophy, was downregulated in both LCPUFA groups.  ELOVL5, a PUFA elongase, was the only enzyme in the LCPUFA biosynthetic pathway that was differentially expressed.  Mitochondrial fatty acid carrier, CPT2, was among several genes associated with mitochondrial fatty acid oxidation to be downregulated, while the mitochondrial proton carrier, UCP2, was upregulated.  PUFA-regulated receptor, Retinoic acid receptor alpha (RARA) was repressed in both groups, while HNF4A, a PUFA regulated factor, was downregulated in L3/C.  Genes involved in neural development, TIMM8A, NRG1, SEMA3D and NUMB, were upregulated while HES1 and SIM1 showed decreased expression. LUM, EML2, TIMP3 and TTC8 which have roles in visual perception were upregulated and TIA1, a silencer of COX2 gene translation is upregulated in L3/C. Ingenuity network analysis identified a top network associated with nervous system development and function with EGFR as the outstanding interaction partner. In this network EGFR interacts with genes involved in neural or visual perception, TIMP3, NRG1, ADAM17, EDG7 and FGF7. Quantitative real time-PCR analysis on selected genes confirmed the array results. The L and L3 groups have DHA and ARA content within the ranges observed in human and baboon breastmilk, and thus these data indicate that LCPUFA concentrations within the normal range of human breastmilk induce global changes in gene expression across numerous functions including transcripts involved in neural development, visual perception, etc.
Keywords: dietary dose response

overall_design: Twelve baboon neonates born spontaneously around 182 days gestation were randomized into 3 groups (n=4 per group).  They were fed for 12 weeks on one of three formulas: C: Control (no DHA-ARA); L: 1× LCPUFA (0.33%DHA-0.67%ARA); L3: 3× LCPUFA (1.00%DHA-0.67%ARA).

first: Joshua

middle: C

last: Anthony

first: Bruce

last: Pan

first: Andrea

last: Hsieh

first: Peter

middle: W

last: Nathanielsz

first: James

middle: T

last: Brenna


title: A genomic screen for activators of the antioxidant response element

status: Public on Feb 22 2007

city: San Diego

first: John

middle: R

last: Walker

state: CA

address: 10675 John Jay Hopkins

country: USA

institute: Genomics Institute of the Novartis Research Foundation

summary:
The antioxidant response element (ARE) is a cis-acting regulatory enhancer element found in the 5’ flanking region of many phase II detoxification enzymes.  Upregulation of ARE-dependent target genes is known to have neuroprotective effects; yet, the mechanism of activation is largely unknown.  By screening an arrayed collection of approximately 15,000 full-length expression cDNAs in the human neuroblastoma cell line IMR-32 with an ARE-luciferase reporter, we have identified several cDNAs not previously associated with ARE activation.  A subset of cDNAs, including sequestosome 1 (SQSTM1) and dipeptidylpeptidase III (DPP3), activated the ARE in primary mouse-derived cortical neurons.  Overexpression of SQSTM1 and DPP3 in IMR-32 cells stimulated NRF2 nuclear translocation and led to increased levels of NAD(P)H:quinone oxidoreductase 1 (NQO1), a protein which is transcriptionally regulated by the ARE.  When transfected into IMR-32 neuroblastoma cells that were depleted of transcription factor NRF2 by RNA interference, SQSTM1 and DPP3 were unable to activate the ARE or induce NQO1 expression, indicating that the ARE activation upon ectopic expression of these cDNAs is mediated by NRF2.  Studies with pharmacological inhibitors indicated that 1-phosphatidylinositol 3-kinase (PI3K) and protein kinase C (PKC) signaling are also essential for activity.  Lastly, overexpression of these cDNAs conferred partial resistance to hydrogen peroxide induced toxicity, consistent with the induction of antioxidant and phase II detoxification enzymes which can protect from oxidative stress.  This work and other such studies may provide mechanisms for activating the ARE in the absence of general oxidative stress, and a novel therapeutic approach to degenerative diseases and aging.
Keywords: overexpression of different antioxidant response element activators

overall_design: Samples were taken from duplicate transfections of different antioxidant response element activators

first: Yanxia

last: Liu

first: Jonathan

last: Kern

first: Jeffrey

middle: A

last: Johnson

middle: G

last: Schultz

first: Hendrik

last: Luesch


title: Pilocytic astrocytoma

status: Public on Mar 14 2007

city: Saint Louis

first: Mukesh

middle: K

last: Sharma

state: MO

address: 4566 Scott Avenue, room 306

department: Neurology

institute: Washington University School of Medicine in Saint Louis

summary:
Pilocytic astrocytomas (PAs) are the most common glioma in children. While many PAs are slow growing or clinically indolent, others exhibit more aggressive features with tumor recurrence and death. In order to identify genetic signatures that might predict PA clinical behavior, we performed gene expression profiling on 41 primary PAs arising sporadically and in patients with neurofibromatosis type 1 (NF1). While no expression signature was found that could discriminate clinically-aggressive or recurrent tumors from more indolent cases, PAs arising in patients with NF1 did exhibit a unique gene expression pattern. In addition, we identified a gene expression signature that stratified PAs by location (supratentorial versus infratentorial).
Keywords: Human, WHO grade I, Brain tumor, Pilocytic astrocytoma, NF1

overall_design:
41 pilocytic astrocytoma samples were analyzed.
For additional metadata details, see Web Link below.

first: David

middle: B

last: Mansur

first: Guido

last: Reifenberger

first: Arie

last: Perry

last: Leonard

first: Kenneth

middle: D

last: Aldape

first: Meredith

last: Albin

first: Ryan

middle: J

last: Emnett

first: Simon

last: Loeser

first: Mark

last: Watson

first: Rakesh

last: Nagarajan

middle: H

last: Gutmann


title: Molecular Marker for predicting development of cancer in ulcerative colitis

status: Public on Feb 28 2007

city: Bunkyo-ku

first: Toshiaki

last: Watanabe

state: Tokyo

department: Department of Surgical Oncology

country: Japan

institute: University of Tokyo

summary:
Samples were taken from either surgically resected specimens or during surveillance colonoscopic examination. The expression profiles were determined using Affymetrix Human Genome U133 Plus 2.0 arrays. Comparison between the sample groups allow to identify a set of discriminating genes that can be used for molecular markers for predicting development of cancer and/or dysplasia in ulcerative colitis, and to characterize potential diagnostic markers in UC-associated neoplasm.
Keywords: repeat

overall_design: 43 UC-NonCa, 10 UC-Ca, 60 sporadic-Ca and 6 UC-associated Ca were analyzed

first: Takashi

last: Kobunai

first: Etsuko

last: Toda

first: Yoshinori

last: Okayama

first: Toshinori

last: Oka


title: Treatment of cultured human aortic endothelial cells with oligomeric procyanidins extracts

status: Public on Feb 17 2007

city: LONDON

first: Roger

last: Corder

address: Charterhouse Square

department: William Harvey Research Institute

country: United Kingdom

institute: Queen Mary, University of London

summary:
Human aortic endothelial cells were grown in culture until confluent. In three experiments using cells derived from three separate donors confluent cultures were incubated for 6 h with contol medium, or medium containing either extracts of oligomeric procyanidins from cranberry juice or red wine, or a procyanidin-rich grape seed extract. At the end of the 6 h treatment period conditioned media samples were retained for immunoassay of secreted peptides and proteins, and RNA was extracted for microarray analysis.
Keywords: Comparative analysis of procyandins on vascular endothelial gene expression

overall_design: Each experiment used cells from one donor. Treatment conditions were: control medium, cranberry extract (CRE), grape seed extract (GSE), and red wine extract (RWE).

last: Pothecary

first: Delphine

middle: M

last: Lees

first: Noorafza

middle: Q

last: Khan

first: Elizabeth

last: Wood

first: Joyce

middle: L

last: Wong


title: Comparative analysis of cellular mRNA incorporation into MLV and HIV1 virus-like particles

status: Public on Mar 08 2007

city: Baltimore

first: Catherine

last: Scollick

state: MD

address: 615 N. Wolfe St.

institute: Johns Hopkins University School of Public Health

summary:
We hypothesize that cellular mRNAs are incorporated nonselectively into retrovirus particles
Keywords: comparative expression profiling

overall_design: This study utilizes microarray analysis to test these hypotheses. Transfected cells were grown, RNA was isolated and used for global gene expression profiling (Affymetrix Human U133 Plus 2.0 array). Statistically significant gene expression was determined as a minimum 6 counts of 9 pairwise comparisons, minimum 1.5-fold change, and p < 0.05. Further, Absolute | FC - FC SEM | >= 1.5.

first: Samuel

last: Rulli

last: Hibbert

first: Jane

last: Mirro

first: Thoru

last: Pederson

first: Shyam

last: Biswal

first: Alan

last: Rein


title: CD133+ and CD133- CSC lines in primary human GBM

status: Public on Mar 06 2007

city: Regensburg

first: Christoph

middle: P.

last: Beier

address: Universitatsstrasse 84

country: Germany

institute: University of Regensburg

summary:
We cultured tumor cells from 22 GBM under medium conditions favoring the growth of neural stem cells. 11 out of 15 primary GBM contained a significant CD133+ subpopulation that comprised cells showing all hallmarks of neural stem cells. Cell lines derived from these CD133+ GBM showed a neurosphere-like, non-adherent growth pattern. In contrast, 4 out of 15 cell lines derived from primary GBM grew adherent in vitro and were driven by CD133- tumor cells that fulfilled stem cell criteria. In vivo, these GBM were characterized by a significantly lower proliferation index but similar GFAP staining as compared to CD133+ GBM. Gene arrays from 2x3 representative cells lines are given.
Keywords: Cancer stem cell, CD133, glioblastoma

overall_design: Human glioblastoma cells cultured in DMEM supplemented with EGF, FGF, LIF, B27.

middle: P

first: Dagmar

first: Ulrich

last: Bogdahn

first: Peter

last: Oefner


title: Gene expression time-course in the human skin during elicitation of allergic contact dermatitis

status: Public on Apr 03 2007

city: Copenhagen

first: Jorgen

last: Olsen

address: Blegdamsvej 3

department: Department of Cellular and Molecular Medicine

country: Denmark

institute: University of Copenhagen

summary:
Genes involved in the inflammatory response resulting in allergic contact dermatitis (ACD) are only partly known. In this study, we introduce the use of high density oligonucleotide arrays for gene expression profiling in human skin during the elicitation of ACD. Skin biopsies from normal and nickel-exposed skin were obtained from 7 nickel-allergic patients and 5 non-allergic controls at four different time points during elicitation of eczema: 0h, 7h, 48h and 96h. Each gene expression profile was analysed by hybridization to high density oligonucletide arrays.
Cluster analysis of 74 genes found to be differentially expressed in the patients over time revealed that the patient samples may be categorised into two groups: An early time point group (0h and 7h) and a late time point group (48h and 96h). Compared to the early time points, late time point skin samples were characterised by the up-regulation of inflammatory molecules including genes involved in the class I antigen presenting pathway and genes involved in lymphocyte adhesion and motility
Correspondence analysis including both patients and controls revealed three distinct groups: i) the control group, ii) the early time point patient group (0h and 7h) and iii) the late time point patient group (48h and 96h).
Keywords: time course, allergic response

overall_design:
For the microarray study, 7 nickel-allergic patients and 5 non-allergic controls were recruited. All subjects were female. The skin covering the upper nates was exposed to nickel delivered during a patch test. Skin biopsies were taken to generate a time-series. Skin was exposed to nickel for 0h, 7h, 48h and 96h.
All nickel allergic patients reacted with visible eczama at the 48h and 96h time-points. No eczema was vissible at the 0h or 7h time-points. Control subjects did not show visible eczema at any time-point.

RNA was extracted from the skin biopsies and 34 biopsies yielded RNA of sufficient quantity and quality for micro-array analysis.

first: Malene

last: Pedersen

first: Lone

last: Skov

first: Torkil

last: Menné

first: Jeanne

last: Duus

first: Jørgen


title: Analysis of microdissected invasive lobular and ductal breast carcinomas in relation to normal ductal and lobular cells

status: Public on Mar 16 2007

city: Olomouc

first: Jan

last: Bouchal

address: Hnevotinska 3

department: Department of Clinical and Molecular Pathology

country: Czech Republic

institute: Palacky University

summary:
The aim of our study was to identify gene expression profiles of ductal and lobular carcinomas in relation to normal ductal and lobular cells. We examined ten mastectomy specimens from postmenopausal breast cancer patients. Ductal and lobular tumor and normal cells were microdissected from cryosections. Fifty nanograms of total RNA were amplified and labeled by PCR and in vitro transcription. GCOS pairwise comparison algorithm and rank products have identified multiple genes that are differentially expressed in comparisons between ductal and lobular tumor and normal cell types. The results suggest that these genes are involved in epithelial-mesenchymal transition, TGFbeta and Wnt signaling. These changes are present in both tumor types but appear to be more prominent in lobular carcinomas.
Keywords: cell type comparison

overall_design: Ten surgical specimens obtained by mastectomy from postmenopausal patients with invasive ductal (IDC) and lobular breast (ILC) carcinomas were investigated. Of these, 5 were IDCs and 5 were ILCs. Tumor and normal tissues from the same mammary gland were identified by an experienced pathologist, snap-frozen in liquid nitrogen and stored at -80ºC for further analysis. Microdissection, RNA isolation, amplification, labeling and microarray analysis are described in sample definitions. Samples from particular cell types (normal ductal, normal lobular, tumor ductal, tumor lobular - 10, 10, 5, 5 samples, respectively) were considered as biological replicates and were compared in between.

first: Gulisa

last: Turashvili

first: Karl

last: Baumforth

first: Wenbin

last: Wei

first: Marta

last: Dziechciarkova

first: Jiri

last: Ehrmann

last: Klein

first: Eduard

last: Fridman

first: Jozef

last: Skarda

first: Marian

last: Hajduch

first: Paul

last: Murray

first: Zdenek

last: Kolar


title: Expression data from hsr containing human embryonic stem cell line, H14, and parent H14  human embryonic stem cell line.

status: Public on Feb 10 2007

city: Sheffield

first: Duncan

last: Baker

address: Western Bank

department: Department of Biomedical Science

institute: University of Sheffield

summary:
Homogenous staining regions (hsr) are cytogenetic representations of a gene amplification. They are found exclusively in tumour cells. We found a hsr in the human embryonic stem cell (hESC) line H14, this is the first report of a hsr in hESCs. FISH and CGH studies showed that the hsr was derived from chromosome 17p11.2. The gene expression analysis studies were performed to identify the genes upregulated due to the hsr by comparison the the hsr-contianing H14 cells with a karyotypically normal parent H14 cell line.
Keywords: karyotypically abnormal hESC vs karyotypically normal hESC.

overall_design: H14 HSR positive embryonic stem cells and H14 HSR negative cells were hybridised to Affymetrix U133 Plus 2.0 expression arrays. Replicates were also run using H14 positive and H14 negative cells from different passages.

first: Peter

last: Andrews

first: Neil

last: Harrison

last: Heath

first: Edna

last: Maltby

first: Kath

last: Smith

first: Hazel

last: Holden

first: Pamela

last: Shaw

first: Harry

last: Moore


title: Synthesis and Anticancer Properties of Water-Soluble Zinc Ionophore: Cell Culture and Xenograft Model

status: Public on Feb 07 2007

city: Los Angeles

first: Joseph

middle: Gerard

last: Hacia

address: 2250 Alcazar Street, IGM 261

department: Biochemistry and Molecular Biology

institute: University of Southern California

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

overall_design: Refer to individual Series


title: Identification of genes that are linked with optineurin expression

status: Public on Feb 02 2007

city: Tuebingen

first: Nicole

last: Weisschuh

address: Roentgenweg 11

institute: University Eye Hospital

summary:
This study aimed to identify genes that are linked with optineurin expression using a combined siRNA-microarray approach
Keywords: siRNA treatment study

overall_design:
Samples a01, a02 and a03 were treated with siRNA specific for optineurin mRNA
Samples b01, b02 and b03 were treated with water (mock experiment)

first: Marcel

middle: V

last: Alavi

first: Michael

last: Bonin

first: Bernd

last: Wissinger


title: Genes regulated by vitamin D in bronchial smooth muscle cells

status: Public on Feb 20 2007

city: Quebec

first: Yohan

last: Bossé

state: Quebec

address: 2725, chemin Sainte-Foy

country: Canada

institute: Laval University

summary:
Studied gene regulation in bronchial smooth muscle cells following vitamin D stimulation.
Keywords: comparison

overall_design: Human primary bronchial smooth muscle cells were from the same patient in all hybridization. Cells were treated for 24 hours with 100 nM of 1alpha,25-Dihydroxyvitamin D3 or with the same concentration of vehicle (ethanol at 0.05%). The experiment was done in triplicates (a total of 6 samples were analyzed).

first: Karim

last: Maghni

first: Thomas

last: Hudson


title: Gene expression in asbestos exposed lung cells

status: Public on Mar 05 2007

city: Solna

first: Penny

last: Nymark

state: Stockholm

address: Nobels väg 13

department: Institute of Environmental Medicine

country: Sweden

institute: Karolinska Institutet

summary:
Asbestos has been shown to cause chromosomal damage and DNA aberrations. The fiber is associated with many different lung diseases such as asbestosis, malignant mesothelioma, and lung cancer, but the disease-related processes are still largely unknown. Our aim was to identify specific gene expression profiles by using Affymetrix arrays, in human cell lines A549, Beas-2B, and MeT5A exposed to asbestos in a time-dependent manner. The hybridization data was analyzed using an algorithm specifically designed for clustering short time series expression data, a canonical correlation analysis (CCA) for identifying correlations between the cell lines, and a Gene Ontology (GO) analysis method for the identification of enriched differentially expressed biological processes.
Keywords: time course, gene expression, cell line, asbestos

overall_design: Samples from three cell lines were collected from asbestos exposed and non-exposed controls at 1h, 6h, 24h, 48h and 7days (not all time-points for each cell lines). Replicates were done for one exposed sample of each cell line.

last: Lindholm

first: Mikko

last: Korpela

first: Leo

last: Lahti

first: Salla

last: Ruosaari

first: Samuel

last: Kaski

first: Jaakko

last: Hollmen

first: Sisko

last: Anttila

first: Vuokko

last: Kinnula

first: Sakari

last: Knuutila


title: Synthesis and Anticancer Properties of Water-Soluble Zinc Ionophores 2

first: Joseph

last: Hacia

summary:
We have demonstrated that water-soluble zinc ionophores can be administered to mice at relatively high doses and inhibit the growth of A549 lung cancer cells grown in xenograft models.  Gene expression profiles of tumor specimens harvested from mice four hours after treatment confirmed that the activation of stress responsive genes occurs in vivo.  These findings lead us to propose that the pharmacologic delivery of zinc to tumors using water solubilized ionophores is a potential approach to cancer therapy.
Keywords: Dose response

overall_design: 1.25 million A549 cells were injected subcutaneously/intramuscularly into the right hind flank of 6 week old CD-1 nude mice that had been irradiated with 4 Gy of total body irradiation from a 137Cs radiation source one day prior to tumor implantation.  When the average size of tumors reached approximately 100 mm3, mice were randomized by tumor size to treatment groups, typically containing 6-8 mice per group.  Tumor and body weight measurements were performed three times per week.  Tumor volume was calculated using the equation V (mm3) = a x b2/2, where a is the largest diameter and b is the smallest diameter.  No significant body weight loss was observed.  To perform gene expression profiling, mice were treated intravenously with one dose (100 μmol/kg) PCI-5002, PCI-5003, or control vehicle (4 mice per group) when the average A549 tumor size reached 500 mm3.  After four hours, tumors were harvested and snap frozen immediately on dry ice.  Tumor tissue was homogenized, and total RNA was isolated and subjected to analysis using Human Genome U133 Plus 2.0 Arrays as described above.

first: Darren

last: Magda

first: Jonathan

last: Sessler


title: Expression data from a paraquat time course experiment in wild type and SoxR deficient strains

city: Amherst

first: Jeffrey

last: Blanchard

state: MA

address: 203 Morrill Science Center IVN

department: Biology

institute: University of Massachusetts Amherst

summary:
SoxR and SoxS constitute an intracellular signal response system that rapidly detects changes in superoxide levels and modulates gene expression in E. coli.
A time series microarray design was used to identify co-regulated SoxRS dependent and independent genes affected by superoxide
Keywords: time course

overall_design: To determine the immediate transcriptional response of E. coli to superoxide, we conducted a time series assay of mRNA levels immediately following the addition of paraquat.  The wild type strain MG1655 and a strain with a precise deletion in soxR was grown in EZ Rich Defined Medium, a modification of Neidhardt’s defined rich medium.  When cultures reached an OD600=0.5, 250µM paraquat (PQ) was added, a concentration that triggers the SoxRS transcriptional cascade, but only has a small effect on exponential growth rate in rich media. Samples were taken immediately before exposure to PQ, and every 2 minutes after exposure, for 10 minutes.

first: Wei-Yun

last: Chen

first: Erin

last: Conlon

first: Pablo

last: Pomposiello


title: Synthesis and Anticancer Properties of Water-Soluble Zinc Ionophores 1

summary:
We have shown that water solubilized versions of a zinc ionophore increase intracellular concentrations of free zinc and have antiproliferative activity in exponential phase A549 lung cancer cultures. The gene expression profiles of A549 lung cancer cultures treated with the lead compound PCI-5002 reveal the activation of stress response pathways.  Medium supplementation with zinc (25 μM) led to activation of additional oxidative stress response as well as apoptotic pathways.  We propose that the pharmacologic delivery of zinc to tumors using water solubilized ionophores is a potential approach to cancer therapy.
Keywords: Dose response

overall_design: A549 human lung cancer cells (1 x 105 cells per T-25 flask in 7 mL complete RPMI 1640 medium) were seeded 8 days prior to treatment of non-cycling plateau phase cultures with drug.  At 4 hours prior to RNA isolation, sapphyrin PCI-5002 (10 μM final concentration), ZnOAc2 (25 μM final concentration), the combination, or control (5% mannitol) solution was added to the cultures.  Each experiment was performed in triplicate.  After incubation, all cultures were washed twice with HBSS supplemented with 0.5% BSA and total RNA was isolated and subjected to analysis on Human Genome U133 Plus 2.0 Arrays (Affymetrix, Santa Clara, CA), as described.  ArrayAssist software (Stratagene/Affymetrix) and the RMA (Robust Microarray Analysis) algorithm were used to generate scaled gene expression values.


title: Endothelial cell culture with Chapel Hill Ultrafine particle

status: Public on Jan 26 2007

city: RTP

first: Edward

last: Karoly

state: NC

address: MD 58D

department: HSD

institute: U.S. EPA

summary:
Epidemiology studies have linked exposure to pollutant particles to increased cardiovascular mortality and morbidity, however, the mechanism remains unknown.  In this study, we hypothesized that the ultrafine fraction of ambient pollutant particles would cause endothelial cells dysfunction. We profiled gene expression of human pulmonary artery endothelial cells (HPAEC) exposed to ultrafine Chapel Hill particles (UFP) (100μg/ml) or vehicle for 4h with Affymetrix HG U133 Plus 2.0 chips (N = 4 each).  Using an unpaired t-test (p <0.01, 5% false discovery rate) we found 426 unique genes to be differentially expressed with 320 upregulated genes and 106 downregulated genes.  Among these genes, we noted upregulation of genes related to coagulation-inflammation circuitry including tissue factor (F3), coagulation factor II receptor-like 2 (F2RL2, PAR3), interleukin (IL)-6 and IL-8. Upregulation of these genes were independently confirmed by RT-PCR and/or protein release. Genes related to the CXC chemokine family that have been implicated in the pathogenesis of vascular disease were upregulated, including MCP-1 (2.60 fold), IL-8 (2.47 fold), CXCL1 (1.41 fold), CXCL2 (1.95 fold), CXCL3 (2.28 fold) and CXCR4 (1.30 fold). In addition, genes related to clotting independent signaling of F3 were also differentially expressed, including FOS, JUN and NFKBIA.  Treatment of HPAEC with UFP for 16 hours increased the release of IL6 and IL8 by 1.9-fold and 1.8-fold respectively.  Pretreatment of HPAEC with a blocking antibody against F3 attenuated IL6 and IL8 release by 30% and 70% respectively.  Thus using gene profiling, we uncovered that UFP may induce vascular endothelial cells to express genes related to clotting and angiogenesis. These results provide a novel hypothesis that PM may cause cardiovascular adverse health effects via induction of tissue factor in vascular endothelial cells which then triggers clotting dependent and independent downstream signaling.
Keywords: particle treatment

overall_design: Human pulmonary artery endothelial cell cultures were treated with Chapel Hill Ultrafine particles or with vehicle control for 4h.  4 bological replicates each for treatment (100ug/ml) and control.  8 affy chips total.

first: Yuh-Chin

last: Huang


title: Immature dendritic cells under hypoxic condition

status: Public on Jan 27 2007

city: Genoa

first: Paolo

last: Fardin

address: Largo G. Gaslini 5

department: Molecular Biology

country: Italy

institute: Istiuto Giannina Gaslini

summary:
Dendritic cells (DCs) are professional antigen-presenting cells whose activity is intrinsically linked to the microenvironment. Hypoxia is a condition of low oxygen tension occurring in inflammatory tissues that creates a special microenvironment conditioning cell physiology. We studied the effects of hypoxia on the differentiation of human monocytes into DCs. Immature DCs were differentiated in vitro from human monocytes under normoxic (iDCs) or hypoxic (Hi-DCs) conditions and the gene expression profile was determined. Hi-DCs expressed novel hypoxia-inducible genes and were characterized by up-regulation of pathways associated with cell movement/migration.
Keywords: stress response, cell differentiation

overall_design: The expression profile of immature dendritic cells under normoxia vs hypoxia was studied. Three healthy donors were used as biologial replicates.

first: Annamaria

last: Ricciardi

first: Angela

last: Elia

first: Paola

last: Cappello

first: Maura

last: Puppo

first: Cristina

last: Vanni

first: Alessandra

last: Eva

first: David

last: Munroe

first: Xiaolin

last: Wu

first: Mirella

last: Giovarelli

first: Luigi

last: Varesio


title: Gene expression profile of human dendritic cells after infection with A. fumigatus

status: Public on Feb 08 2007

city: Würzburg

first: Juergen

last: Loeffler

address: Josef-Schneider-Str. 2

department: Medizinische Klinik & Poliklinik II

institute: Universitätsklinikum Würzburg

summary:
DCs are localized under the mucosa of the lungs and the gastrointestinal tract, and therefore come into close contact with A. fumigatus germ tubes during early steps of infection as soon as fungi become invasive. For a more detailed insight into differentially regulated genes, whole genome microarray analysis was performed.
On average, unstimulated DCs showed expression of about 13.500 genes (35% of all genes spotted on the chip). After 6h co-cultivation of DCs and live A. fumigatus germ tubes, 590 genes showed a 4fold altered gene expression, if normalizing data with VSN before. Therein, a wide range of immune response genes, including genes encoding for cytokines (IL-1α, IL-1β, IL-6, IL-8, IL-10, IL-12p40, TNF-α), chemokines (CCL5, CCL20, CXCL10), immunorelevant receptors (TLR2, TLR4, PTX-3), as well as costimulatory molecules (CD40, CD80, CD83, CD86) were differentially regulated.
Keywords: dendritic cell, A. fumigatus germ tubes, infection for 6h, affymetrix whole genome microarray analysis

overall_design: Human DCs were generated from two independent, healthy blood donors. 5 x 106 DCs were either cultivated without fungi or together with 5 x 106 A. fumigatus germ tubes for 6h until isolation of total RNA.

first: Markus

last: Mezger

first: Susanne

last: Kneitz

first: Jürgen

last: Löffler


title: Gene expression analysis of IOSE cells treated with TGFb1, a time course study

status: Public on Jan 09 2007

city: Columbus

first: Sandya

last: Liyanarachchi

state: OH

address: Rm 830, 460, W 12th Avenue

department: Human Genetics

institute: OSU

summary:
Unlike ovarian cancer, normal ovarian epithelium response to TGFb1 induced growth inhibition. This time course study tried to idenify genes that showed changes after additionof TGFb1 in immortalized ovarian surface epithelial cells (IOSE) which is derived from normal ovarian epithelial cells
Keywords: Time-course

overall_design: IOSE cells were treated with TGFb1 for 0, 3, 6, and 12hrs. Cells were harvested for total RNA extraction. Affymetrix expression microarray was performed to analyse gene expression changes.

first: Huaxia

last: Qin

first: Michael

middle: W

last: Chan

first: Kenneth

last: Nephew

first: Ramana

last: Davuluri

first: Tim


title: Gene expression of growing antler in Cervus elaphus

status: Public on Jan 19 2007

city: Madrid

first: Manuel

last: Nieto-Díaz

state: Madrid

address: Avda. Doctor Arce, 37

department: Neural Plasticity

country: Spain

institute: Instituto de Neurobiología ramón y Cajal (CSIC)

summary:
The aim of the study is to characterize the gene expression of the growing antlers during their process of regeneration and fast growth
Keywords: tissue type and state comparison

overall_design: Gene profiles of RNA samples from antler tip, base (pedicle) and frontal bone. Samples were harvested during spring, at the period of maximum antler growth (60 days after casting the previous antlers). And additional sample (DEER4) was harvested at the end of the growing season (90 days after the casting).

last: Nieto-Diaz

last: Nieto-Sampedro

first: Wolfgang

last: Pita-Thomas


title: A Genome-wide Screen for Hypermethylated Genes in Lung Cancer

status: Public on Jan 03 2007

city: South San Francisco

middle: S

last: Shames

address: 1 DNA Way

department: Oncology Biomarker Development

institute: Genentech Inc

summary:
Abstract
Background:  Promoter hypermethylation coupled with loss of heterozygosity at the same locus results in loss of gene function in many tumor cells.  The “rules” governing which genes are methylated during the pathogenesis of individual cancers, how specific methylation profiles are initially established, or what determines tumor-type specific methylation are unknown.  However, DNA methylation markers that are highly specific and sensitive for common tumors would be useful for the early detection of cancer, and those required for the malignant phenotype identify pathways important as therapeutic targets.
Methods and Findings: In an effort to identify new cancer-specific methylation markers, we employed a high throughput global expression profiling approach in lung cancer cells.  We identified 132 genes that have 5’ CpG islands, are induced from undetectable levels by 5-aza-2’-deoxycytidine (5-aza) in multiple non-small cell lung cancer cell lines, and are expressed in immortalized human bronchial epithelial cells.  As expected, these genes were also expressed in normal lung, but often not in companion primary lung cancers.  Methylation analysis of a subset (45/132) of these promoter regions in primary lung cancer (N=20) and adjacent non-malignant tissue showed that 31 genes had acquired methylation in the tumors, but did not show methylation in normal lung or lymphocytes.  We studied the eight most frequently and specifically methylated genes from our lung cancer data set in breast cancer (N=37), colon cancer (N=24), and prostate cancer (N=24) along with counterpart non-malignant tissues.  We found that seven loci were frequently methylated in both breast and lung cancers, with four showing extensive methylation in all four epithelial tumors.
Conclusions: By using a systematic biological screen we identified multiple genes that are methylated with high penetrance in primary lung, breast, colon, and prostate cancers.  The cross-tumor methylation pattern we observed for these novel markers suggests that we have identified a partial promoter hypermethylation signature for these common malignancies.  These data suggest that while tumors in different tissues vary substantially with respect to gene expression, there may be commonalities in their promoter methylation profiles that represent potential targets for early detection screening or therapeutic intervention.
Keywords: Cell line comparison

overall_design:
Drug treatment: control, 100 nM, 1 uM
Cancer vs. Normal Comparison: NSCLC vs. Normal

first: Luc

last: Girard

first: Boning

last: Gao

first: Mitsuo

last: Sato

first: Cheryl

last: Lewis

first: Narayan

last: Shivapurkar

first: Aixiang

last: Jiang

first: Charles

last: Perou

first: Young

last: Kim

first: Jonathan

last: Pollack

first: Kwun

last: Fong

first: Chi-Leung

last: Lam

first: Maria

last: Wong

first: Yu

last: Shyr

first: Rita

last: Nanda

first: Olufunmilayo

last: Olopade

first: William

last: Gerald

last: Euhus

first: Jerry

last: Shay

first: Adi

middle: F

last: Gazdar

first: John

last: Minna


title: Transcriptomics in cardiac surgery

status: Public on Jan 01 2007

city: Zurich

first: Michael

last: Zaugg

address: Raemistrasse 100

department: Anesthesiology

country: Switzerland

institute: Anesthesiology, University Hospital Zurich

summary:
Anesthetic gases elicit organ protection in patients undergoing coronary artery bypass graft (CABG) surgery.  This study aimed at identifying myocardial transcriptional phenotypes and anesthetic-induced changes in gene expression to predict cardiovascular biomarkers and cardiac function after off-pump CABG.
Keywords: cardiac surgery, anesthetics

overall_design: Patients scheduled for off-pump CABG were randomized into a group with the anesthetic gas sevoflurane (n=10) or the intravenous anesthetic propofol (n=10). Atrial samples were collected at the beginning and end of bypass surgery to determine gene expression profiles.

first: Eliana

last: Lucchinetti


title: AML genome hybridization to HGU133 Plus 2.0

status: Public on Mar 01 2007

city: Shimotsukeshi

first: Hiroyuki

last: Mano

state: Tochigi

address: 3311-1 Yakushiji

department: Division of Functional Genomics

institute: Jichi Medical University

type: Genome variation profiling by genome tiling array

summary:
CD34-positive and negative fractions were used for genomic DNA preparation, which was hybridized to HGU133 Plus 2.0 arrays.
Keywords: Disease state analysis

overall_design: Samples with a suffix "-C" are CD34-negative controls, and those with a suffix "-L" are CD34-positive leukemic fractions. A total of 23 pairs were analyzed.


title: IFN-gamma alters the response of Borrelia burgdorferi-activated endothelium to favor chronic inflammation

status: Public on Jan 05 2007

city: Stony Brook

first: Lance

middle: E

last: Palmer

address: 235 CMM

department: Center for Infectious Diseases

institute: University at Stony Brook

summary:
Borrelia burgdorferi, the agent of Lyme disease, promotes pro-inflammatory changes in endothelium that lead to the recruitment of leukocytes. The host immune response to infection results in increased levels of IFN-gamma in the serum and lesions of Lyme disease patients that correlate with greater severity of disease. Therefore, the effect of IFN-gamma on the gene expression profile of primary human endothelial cells exposed to B. burgdorferi was determined. B. burgdorferi and IFN-gamma synergistically augmented the expression of 34 genes, seven of which encode chemokines. Six of these (CCL7, CCL8, CX3CL1, CXCL9, CXCL10, and CXCL11) attract T lymphocytes, and one (CXCL2) is specific for neutrophils. Synergistic production of the attractants for T cells was confirmed at the protein level. IL-1beta, TNF-alpha, and LPS also cooperated with IFN-gamma to induce synergistic production of CXCL10 by endothelium, indicating that IFN-gamma potentiates inflammation in concert with a variety of mediators. An in vitro model of the blood vessel wall revealed that an increased number of human T lymphocytes traversed endothelium exposed to B. burgdorferi and IFN-gamma, as compared to unstimulated endothelial monolayers. In contrast, addition of IFN-gamma diminished the migration of neutrophils across B. burgdorferi-activated endothelium. IFN-gamma thus alters gene expression by endothelium exposed to B. burgdorferi in a manner that promotes recruitment of T cells and suppresses that of neutrophils. This modulation may facilitate the development of chronic inflammatory lesions in Lyme disease.
Keywords: Cell response to inflammatory stimuli

overall_design: Human umbilical vein endothelial cells (HUVEC) were stimulated with Interferon-gamma (IFN-g), Borrelia burgdorferi or both IFN-g and Borrelia or were left unstimulated.  Affymetrix HGU133 plus 2.0 slides were used in duplicate for each condition.

first: Tarah

last: Dame

first: Barbara

last: Orenzoff

first: Martha

last: Furie


title: Simvastatin has an anti-inflammatory effect on macrophages via upregulation of Kruppel-like factor-2

status: Public on Dec 31 2006

city: Kuopio

first: Tiina

last: Tuomisto

address: Neulaniementie 2

department: Molecular Medicine

country: Finland

institute: A.I.Virtanen Institute

summary:
HMG-CoA reductase inhibitors, statins, have beneficial vascular effects beyond their cholesterol-lowering action. These pleiotropic effects include an anti-inflammatory effect on macrophages. Since macrophages play a central role in atherogenesis, we further characterized the effects on peripheral blood monocyte-macrophages (HPBM). Using Affymetrix gene chip analysis of simvastatin-treated HPBM, we found that simvastatin treatment lead to the downregulation of the expression of many proinflammatory genes including several chemokines (e.g. MCP-1, MIP-1 alpha and β, RANTES, several other CC and CXC chemokines, IL-2 receptor-β, and leukemia inhibitory factor), members of the tumor necrosis factor family (e.g. lymphotoxin beta and TRAIL), VCAM-1, ICAM-3, and tissue factor (TF). Simvastatin also modulated the expression of several transcription factors essential for the inflammatory response: simvastatin downregulated the expression of NF-kappaB relA/p65 subunit and ets-1 transcription factor, and upregulated the expression of a novel atheroprotective transcription factor, Kruppel-like factor 2 (KLF-2). The effects of simvastatin on KLF-2 and its target genes were dependent on protein prenylation, since inhibitors of protein prenylation had a similar inhibitory effect in THP-1 derived macrophages. Additionally, by lentiviral overexpression KLF-2 we showed that the effect of simvastatin on MCP-1 and TF were dependent on KLF-2. We concluded that simvastatin had a strong anti-inflammatory effect on macrophages, which includes upregulation of the atheroprotective transcription factor KLF-2. These findings further explain the beneficial pleiotropic effects of statins on cardiovascular diseases.
Keywords: time-course, response to treatment

overall_design:
Cell culture studies.
Human peripheral blood monocytes (HPBM) were isolated from buffy coats from healthy blood-donor volunteers (Finnish Red Cross, Helsinki, Finland) using Ficoll-Paque gradient centrifugation. None of the blood donors were on statin therapy.  During isolation, monocytes from 3 individuals were pooled. Adherent cells were cultured in standard medium (20% human serum, Cambrex for differentiation into macrophages. The macrophage-phenotype at day 7 after isolation was confirmed with the typical shape of macrophages and also by macrophage-immunostaining (mAB CD68, DAKO, Denmark, dilution 1:200), where macrophages presented >95% of the cell population. The study protocol has been accepted by the Ethical Committee of the University of Kuopio.

Simvastatin treatment.
Simvastatin was a generous gift from Merck & Co. The inactive lactone form of simvastatin was hydrolyzed to the corresponding β-hydroxy acid. The HPBM-macrophages were treated with statin at day 7 after the isolation. 12 hours prior to statin treatment the cell growth media were changed to serum-free media. The statin-treated (10 uM simvastatin in serum-free media) cells were collected at 12 h and 24 h for Affymetrix analyses. The toxicity of  simvastatin was assessed in cell culture: 5x concentration of simvastatin (50 uM) had no effect on cell viability.

RNA isolation.
Total RNA was isolated from the cells using Trizol reagent (Gibco BRL, USA) according to manufacturers’ instructions. The quality and amount of RNA was assessed by spectrophotometry (NanoDrop, USA) and by agarose gel electrophoresis.

Microarray analyses.
For Affymetrix analyses three separate HPBM cell isolation and simvastatin experiments were performed (HPBMs from 3 individuals at each time, total n=9). Cells were collected at 12 h and 24 h after the statin treatment. Total RNA was isolated as above. cDNAs were prepared from RNAs (5 ug of RNA) with reverse transcriptase (Superscript II primed by a poly (T) oligomer/T7 promoter). cDNAs were subsequently used as a template to make biotin-labeled cRNA with an  in vitro transcription reaction. cRNAs (15 ug) were hybridized to Affymetrix HGU133 Plus 2.0 oligonucleotide arrays, which was processed and scanned according to manufacturer’s instructions. Each array quantifies the expression of over 47 000 transcripts (including  full-length mRNA sequences and expressed sequence tags) derived from build 133 of the UniGene database (available at www.affymetrix.com).
Microarray data analysis.
Affymetrix GeneChip® Operating Software (GCOS) was used to generate .CEL files which were then converted into .DCP files using dChip (http://www.dchip.org) V1.3 software 28. The arrays were normalized to baseline array with median probe intensity, and gene expression data were generated calculating model-based expression values. In this study, genes were considered differentially expressed if they changed more than 1.5-fold (90% confidence bound of the fold change), absolute difference of signals was >100, at least 40% of the samples were called present in both groups and False Discovery Rate (FDR) was less than 1%. Hierarchical clustering was performed by dChip using Pearson correlation with a centroid-linkage method. Gene function analysis was performed by using the gene ontology mining tool GoSurfer incorporated in dChip program.

middle: T

first: Henri

last: Lumivuori

first: Emilia

last: Kansanen

first: Sanna-Kaisa

last: Hakkinen

first: Mikko

last: Turunen

first: Anthon

last: Horrevoets

first: Anna-Liisa

last: Levonen

first: Seppo

last: Yla-Herttuala


title: MCF7 inhibitor

status: Public on Dec 15 2006

city: Yokohama

first: Mariko

last: Okada

address: W518, 1-7-22, Suehiro-cho, Tsurumi-ku

institute: RIKEN RCAI

summary:
ErbB receptor ligands, epidermal growth factor (EGF) and heregulin (HRG), induce dose-dependent transient and sustained intracellular signaling, proliferation and differentiation of MCF-7 breast cancer cells, respectively. In an effort to delineate the ligand-specific cell determination mechanism, we investigated time-course gene expressions induced by EGF and HRG that induce distinct cellular phenotypes in MCF-7 cells. To analyze the effects of ligand dosage and time for the gene expression independently, we developed a statistical method for decomposing the expression profiles into the two effects. Our results indicated that signal transduction pathways devotedly convey quantitative properties of the dose-dependent activation of ErbB receptor to early transcription. The results also implied that moderate changes in the expression levels of numbers of genes, not the predominant regulation of a few specific genes, might cooperatively work at the early stage of the transcription for determining the cell fate. However, the EGF- and HRG-induced distinct signal durations resulted in the ligand-oriented biphasic induction of proteins after 20 min. The selected gene list and HRG-induced prolonged signaling suggested that transcriptional feedback to the intracellular signaling results in a graded to biphasic response in the cell determination process, and that each ErbB receptor is inextricably responsible for the control of amplitude and duration of cellular biochemical reactions.
Keywords: time course

overall_design: MCF-7 human breast cancer cells were incubated from 5min to 90min after administration of the growth hormone (heregulin (HRG)), the kinase inhibitor (U0126 (a MEK inhibitor), AG1478 (an EGFR kinase inhibitor) or both growth hormone and one of kinase inhibitor. Control was set as growth hormone/inhibitor non-treated cells. For each inhibitor, one control was used.


title: airway epithelium, large airways, pre and post-mechanical injury

status: Public on Dec 12 2006

city: New York

first: Yael

last: Strulovici-Barel

address: 1300 York Avenue

department: Department of Genetic Medicine

institute: Weill Cornell Medical College

summary:
Responses of the Human Airway Epithelium Transcriptome to In Vivo Injury
To identify genes participating in repair of the human airway epithelium following injury, we used bronchoscopy and brushing to denude the airway epithelium of healthy individuals, sequentially sampled the same region 7 and 14 days later, and assessed the recovered epithelium for relative levels of gene expression using Affymetrix high-density oligonucleotide microarrays with TaqMan PCR confirmation. Histologic assessment showed that the epithelium was denuded immediately following injury, at 7 days the epithelium was completely covered but partially de-differentiated, and by 14 days there was close to normal proportions of differentiated cells. Gene expression analysis was carried out with both the Affymetrix Microarray Suite 5.0 and Robust Multi-array Average algorithms, applying a multiple test correction to identify bona fide changes in gene expression. At day 7, there were substantial differences in the gene expression pattern compared to the resting epithelium, with a distinctive airway epithelial “repair transcriptome” of actively proliferating cells in the process of re-differentiation. The repair transcriptome at 7 days was dominated by genes encoding proteins involved in cell cycle regulation, transcription, signal transduction, metabolism and transport. Interestingly, the majority of cell cycle genes differentially expressed at day 7 belonged to the G2 and M late phases of the cell cycle, suggesting that the proliferating cells are relatively synchronized 1 wk following injury. At 14 days post-injury, the majority of the gene expression changes observed at day 7 were no longer observed, with the expression profile similar to that of resting airway epithelium. Using a class prediction algorithm, a group of 50 genes dominated by cell cycle genes, that represent a human airway epithelial “repair signature” was identified. These observations provide a baseline of the functional gene categories participating in the process of normal human airway epithelial repair that can be used in future studies of injury and repair in human airway epithelial diseases.
Keywords: response to airway injury

overall_design: comparison of gene expression in airway epithelial cells of the large airways, before and after mechanical injury caused by airway brushing

first: Adriana

last: Heguy

first: Ben-Gary

last: Harvey

first: Phil

last: Leopold

first: Igor

last: Dolgalev

first: Tina

last: Raman

first: Ronald

last: Crystal


title: 1,25 (OH)2 vitamin D3 induces expression of CCR10 and other genes

status: Public on Jan 22 2007

city: Palo Alto

first: Junliang

last: Pan

address: 3801 Miranda Avenue

department: Pathology

institute: Stanford University

summary:
Human naive T cells from peripheral blood were cultured in 24 wells coated with anti-CD3 and anti-CD28 antibodies in the presence or absence of retinoid acid, IL-12, and 1,25 (OH)2 vitamin D3.  The T cells were FACS-sorted based on expression of CD3, integrin alpha4beta7, cutaneous lymphocyte antigen (CLA) and chemokine receptor 10.  This serie includes microarray data from stimulated T cells under indicated conditions.
Keywords: Human T cell, vitamin D and A, chemokine receptor, nuclear receptor

overall_design: Total RNA was isolated from these sorted T cells using Invitrogen RNeasy mini-kit.  The quality of RNA was prechecked by Agilent BioAnalyzer.  Double-stranded cDNA was synthesized, and transcribed into Biotin-labeled cRNA, which was fragmented before hybridization.  All the procedure was performed in the Stanford University PAN facility.


title: Specific transcriptional changes in human fetus with autosomal trisomies

status: Public on Dec 01 2006

city: Tuebingen

first: Oezge

last: Altug Teber

address: Calwerstr. 7

department: Medical Genetics

institute: University of Tuebingen

summary:
Background: Among full autosomal trisomies, only trisomies of chromosome 21 (Down syndrome, DS), 18 (Edward syndrome, ES) and 13 (Patau syndrome, PS) are compatible with postnatal survival. But the mechanisms, how a supernumerary chromosome disrupts the normal development and causes specific phenotypes, are still not fully explained. As an alternative to gene dosage effects due to the trisomic chromosome, a genome-wide transcriptional dysregulation has been postulated. The aim of this study was to define the transcriptional changes in trisomy 13, 18, and 21 during early fetal development in order to define whether (1) overexpression of genes of the trisomic chromosome contributes solely to the phenotype, if (2) all genes of the trisomic chromosome are upregulated similarly and whether the ratio of gene expression is in agreement with the gene dosis, (3) whether the different trisomies behave similarly in the characteristics of transcriptional dysregulation, and (4) whether transcriptional pattern can be potentially used in prenatal diagnosis. Methods: Using oligonucleotide microarrays (Affymetrix, U133 Plus 2.0), we analyzed whole genome expression profiles representing 54.000 probe sets in cultured amniocytes (AC) and chorion villus cells (CV) from pregnancies with a normal karyotype and with trisomies of human chromosomes 21, 18 and 13. Findings: We observed a low to moderate up-regulation for a subset of genes of the trisomic chromosomes. Transcriptional level of approximately 12-13 % of the supernumerary chromosome appeared similar to the respective chromosome pair in normal karyotypes. Expression values as well as the expression patterns of genes from the trisomic chromosome can distinguish the respective trisomic samples from euploid controls. A subset of chromosome 21-genes including the DSCR1-gene involved in fetal heart development was consistently up-regulated in different tissues (AC, CV) of trisomy 21 fetuses whereas only minor changes were found for genes of all other chromosomes. In contrast, in trisomy 13 and trisomy 18 vigorous downstream transcriptional changes were found. Interpretation: Global transcriptome analysis for autosomal trisomies 13, 18, and 21 supported a combination of the two major hypotheses. As several transcriptional pathways are altered, complex regulatory mechanisms are involved in the pathogenesis of autosomal trisomies. A genome-wide transcriptional dysregulation was predominantly observed in trisomies 13 and 18, whereas a more to chromosome 21 restricted expression alteration was found in trisomy 21.
Keywords: Trisomy, Down syndrome, Patau syndrome, Edward syndrome, microarray, gene expression, amniocytes, chorion villus cells, gene dosage effect, DSCR1

overall_design: The study included the following samples: Three samples with normal chromosomes in Amniocytes (AC) and chorion villus cells (CV) each, three samples with trisomy 13 in AC, three samples with trisomy 18 in CV, and three samples with trisomy 21 in AC and CV each.

first: Michael

last: Bonin

first: Ulrike

middle: A

last: Mau-Holzmann

first: Andreas

last: Dufke

first: Heike

last: Stappert

first: Ismail

last: Tekesin

first: Helmut

last: Heilbronner

first: Kay

last: Nieselt

first: Olaf

last: Riess

last: Walter


title: Identification of genes specifically over-expressed in lupus CD4 T and B cells

city: Brussels

first: Bernard

middle: Robert

last: Lauwerys

address: Avenue Hippocrate 10

department: Institut de Recherches Expérimentales et Cliniques

country: Belgium

institute: Université catholique de Louvain

summary:
Objective: Gene expression studies performed on PBMC from systemic lupus erythematosus (SLE) patients provided strong evidence of a type I interferon signature, underscoring the potential role of these cytokines in the physiopathology of SLE. In this work, we performed microarray analyses of differential gene expression using purified CD4 T and B cells sorted from SLE PBMC. In order to discriminate genes specific to SLE from those induced by inflammatory responses in general, control samples were obtained not only from healthy individuals but also from rheumatoid arthritis (RA) patients.
Results: A strong interferon signature was found both in the CD4 T and the B lymphocytes from SLE patients, thereby confirming the results obtained on total PBMC. Interestingly, many interferon-induced genes were also over-expressed in CD4 and B cells from RA patients. Some genes were more specifically over-expressed in SLE lymphocytes, and 3 of them, SLAMF1, BRDG1 and RASGRP1, were exclusively up-regulated in SLE B cells. SLAMF1 and BRDG1 are localized in disease-associated loci, thereby suggesting that they might play a role in the physiopathology of the disease.
Keywords: Comparative analysis of global gene expression in PBMC subsets

overall_design: CD4 T and B cells were sorted by flow cytometry from PBMC of patients with SLE, RA and healthy controls. GeneChip® Human genome U133 Plus 2.0 arrays were hybridized in monoplicates and the genes differentially expressed among the three groups of patients were identified using ANOVA tests with corrections for multiple comparisons.

middle: R

first: Jamila

last: Louahed

first: Laurent

last: Knoops

first: Anne-Lise

last: Maudoux

first: Edward

middle: K

last: Wakeland

first: Benoit

last: Van den Eynde

first: Frederic

last: Houssiau


title: Prostate Adenocarcinoma Progression

status: Public on Nov 27 2006

city: Kaohsiung

first: Kuang-Den

last: Chen

address: 123, Ta-Pei Rd., Niao-Sung Dist.

department: Translational Research Center in Biomedical Sicences

country: Taiwan

institute: Kaohsiung Chang Gung Memorial Hospital

summary:
Expression profiling of prostate carcinoma tissue samples of benign and metastatic in disease progression.
Keywords: disease-state analysis

overall_design: In our studies, microarray results compared to those obtained from real-time polymerase-chain reactions to identify concordant changes in mRNA levels of target genes.


title: Cultured A549 lung cancer cells treated with actinomycin D and sapphyrin PCI-2050

city: Los Angeles

first: Joseph

last: Hacia

address: 2250 Alcazar Street, IGM 261

department: Biochemistry and Molecular Biology

institute: University of Southern California

summary:
The goal of this project was to characterize changes in gene expression in response to the anti-cancer agent sapphyrin PCI-2050.  Cultured A549 human lung cancer cells were treated with sapphyrin PCI-2050 or actinomycin D, a known transcripitonal inhibitor.  The gene expression profiles of drug-treated and control A549 cultures were determined using Human Genome U133 Plus 2.0 Arrays (Affymetrix, Santa Clara, CA).  Further details are provided in our published manuscript: <http://www.molecular-cancer.com/content/6/1/9>.
Keywords: responses to treatments with anti-cancer agents

overall_design: A549 human lung cancer cells (1 x 10^5 cells per T-25 flask in 7 mL complete RPMI 1640 medium) were seeded eight days prior to treatment of non-cycling plateau phase cultures with drug.  At four hours prior to RNA isolation, sapphyrin PCI-2050 (1.25 or 2.5 μM final concentration), actinomycin D (5 μg/mL final concentration), or control (5% mannitol) solution was added to the cultures.  Each experiment was performed in triplicate.  After incubation, all cultures were washed twice with HBSS supplemented with 0.5% BSA and total RNA was isolated and subjected to analysis on Human Genome U133 Plus 2.0 Arrays (Affymetrix, Santa Clara, CA).

middle: G

first: Darren

last: Magda


title: Spheroid Formation and Recovery of Human Foreskin Fibroblasts and T98G Glioma Cells at Ambient Temperature

city: Blacksburg

first: Richard

middle: F.

last: Helm

state: VA

address: West Campus Drive

department: Biochemistry, Fralin Biotechnology Center

institute: Virginia Polytechnic Institute and State University

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

overall_design: Refer to individual Series


title: Spheroid Formation and Recovery of Human T98G Glioma Cells at Ambient Temperature

summary:
Mammalian cells were grown as multicellular aggregates (spheroids) in an effort to determine the signaling events required for two cellular transformations states; primary foreskin fibroblasts (HFF-2) and glioblastoma cancer (T98G) cells, to survive at room temperature under oxygen and nutrient-deprived conditions for extended periods of time (2 weeks) and subsequently grown out from the arrested state as adherent monolayers. HFF-2 cells were cultured in DMEM supplemented with 15% fetal bovine serum and 5% carbon dioxide humidified air at 37 degrees C. T98G cells were cultured in EMEM with 10% FBS, 5% non-essential amino acids and 5% carbon dioxide humidified air at 37 degreesC.  Monolayers were grown in T-185 flasks to 60% confluency then split into T-185 flasks coated with a 1% agarose mix in a 2:1 media/water ratio. Cells were suspended in 30 ml of supplemented media and grown for 4 days in order to form multicellular spheroids as described previously by our group (J. Cell. Physiol., 206 [2006] 526-536; see GSE1364 and GSE1455 for similar experiments with HEK293 cells). The suspension was removed from the flasks and centrifuged (1500 x g, 2 min) and the media removed. The pellet was returned to the flasks and then placed in vacuum bags (Dri-shield 2000 moisture barrier bag from Surmount Inc., USA; Cat. number 70068), which were sealed immediately under vacuum (Deni Magic Vac, Champion model; Keystone Manufacturing, USA). Vacuum-sealed flasks were stored for 2 weeks (in the dark) at room temperature. Recovery was initiated by removing the flask from the bag and resuspending the spheroids in supplemented media and placing the flasks in a 5% CO2/humidified air incubator maintained at 37 degreesC. Timepoints for transcriptional analysis were monolayer (control), 4 day growth spheroids, 2 week stored spheroids and 7 day growth back to monolayers.

There were 67 HFF-2 and 50 T98G genes that exhibited at least a 10-fold change in expression during the course of arrest and recovery.  Eleven of these genes were common to both cell lines.  While the trends were the same between the two cell lines for genes that were in common, the timing of the responses were clearly different. The primary fibroblasts showed significant gene expression changes during spheroid formation and returned to their initial monolayer state after the two week arrest, whereas the glioblastoma cells exhibited major changes in gene expression during the recovery process.  Furthermore, the T98G cells never returned to the monolayer expression levels during the time course of the recovery phase (7 days growth under monolayer conditions).  Major differences in the transcriptional data were also found between the cell lines with regards to the putative cellular location of the encoded proteins.  Of the genes with significant fold changes (+/- 10-fold) for which location information is available, the primary fibroblasts were significantly enriched in genes encoding extracellular (32%) and membrane-associated (26%) proteins.  The glioblastoma cells, on the other hand, had only 17% extracellular and 13% membrane-associated.  The expression level trends for all genes above the 10-fold cutoff were as observed for the "in common" genes; HFF-2 genes had returned to monolayer expression levels after 7 days of recovery whereas the T98G line was still exhibiting expression values much different from that of the monolayer control.

Keywords = human foreskin fibroblasts, desiccation, rehydration, monolayer, spheroid, stabilization, ambient temperature
Keywords: other

first: Graham

middle: D

last: Jack

first: Stephen

middle: M

last: Slaughter

first: Malcolm

last: Potts


title: Expression data of small airway epithelium from phenotypically normal smokers and non-smokers

status: Public on Nov 21 2006

first: Yael

last: Strulovici-Barel

summary:
Modification of Gene Expression of the Small Airway Epithelium in Response to Cigarette Smoking
The earliest morphologic evidence of changes in the airways associated with chronic cigarette smoking is in the small airways. To help understand how smoking modifies small airway structure and function, we developed a strategy using fiberoptic bronchoscopy and brushing to sample the human small airway (10th-12th order) bronchial epithelium to assess gene expression (HG-133 Plus 2.0 array) in phenotypically normal smokers (n=10, 33 ± 7 pack-yr) compared to matched non-smokers (n=12). Even though the smokers were phenotypically normal, analysis of the small airway epithelium of the smokers compared to the non-smokers demonstrated up- and -down-regulation of genes in multiple categories relevant to the pathogenesis of chronic obstructive lung disease (COPD), including genes coding for cytokines/innate immunity, apoptosis, mucin, response to oxidants and xenobiotics, and general cellular processes. In the context that COPD starts in the small airways, these gene expression changes in the small airway epithelium in phenotypically normal smokers are candidates for the development of therapeutic strategies to prevent the onset of COPD.
Keywords: smokers vs non-smokers

overall_design: comparison of gene expression in airway epithelial cells of the small airways of healthy smokers vs healthy non-smokers

first: Adriana

last: Heguy

first: Ben-Gary

last: Harvey

first: Phil

last: Leopold

first: Brendan

last: Carolan

first: Barbara

last: Ferris

first: Ronald

last: Crystal


title: Spheroid Formation and Recovery of Human Foreskin Fibroblasts at Ambient Temperature


title: Airway epithelium, small airways, normal non-smokers, phenotypic normal smokers, smokers with COPD and early COPD

summary:
Upregulation of Expression of the Ubiquitin Carboxyl Terminal Hydrolase L1 Gene in Human Airway Epithelium of Cigarette Smokers
The microarray data deposited here is from 39 HG-U133 Plus 2.0 GeneChips, from 12 normal non-smokers, 12 phenotypic normal smokers, 9 Early COPD and 6 COPD individuals, all small airways, all small airway. A subset of these samples have been already submitted under GEO Accession Number GSE 4498. These are: 12 non-smokers samples (GSM101095-GSM101106) and 10 smoker samples (GSM101107-GSM101116). These 22 samples that are also in GSE4498 were described in Harvey, B-G; Heguy, A.; Leopold, P.L.; Carolan, B.; Ferris, B. and Crystal R.G. Modification of Gene Expression of the Small Airway Epithelium in Response to Cigarette Smoking. J. Mol. Med (in press). These data are part of a study aimed at  understanding how cigarette smoking modifies neuroendocrine cells, in which microarray analysis with TaqMan confirmation was used to assess airway epithelial samples obtained by fiberoptic bronchoscopy from 81 individuals (normal nonsmokers, normal smokers, smokers with early COPD and smokers with established COPD). Of 11 genes considered to be neuroendocrine cell-specific, only ubiquitin C-terminal hydrolase L1(UCHL1), a member of the ubiquitin proteasome pathway, was consistently upregulated in smokers compared to nonsmokers. Up-regulation of UCHL1 at the protein level was observed with immunohistochemistry of bronchial biopsies of smokers compared to nonsmokers. Interestingly, however, while UCHL1 expression was present only in neuroendocrine cells of the airway epithelium in nonsmokers, UCHL1 expression was also expressed in ciliated epithelial cells in smokers, an intriguing observation in light of recent observations that ciliated cells can are capable of transdifferentiating to other airway epithelium. In the context that UCHL1 is involved in the degradation of unwanted, misfolded or damaged proteins within the cell and is overexpressed in >50% of lung cancers, its overexpression in chronic smokers may represent an early event in the complex transformation from normal epithelium to overt malignancy.
Keywords: non-smokers vs phenotypic normal smokers, smokers with early COPD, and smokers with COPD

overall_design: comparison of gene expression in airway epithelial cells of the small airways of normal non-smokers, phenotypic normal smokers, smokers with early COPD and smokers with COPD


title: rogae-affy-human-323460

status: Public on Nov 14 2006

city: Phoenix

first: Winnie

last: Liang

state: AZ

address: 445 N. Fifth Street

institute: Translational Genomics

summary:
Regulation of presenilin genes. Presenilins are intramembrane aspartic proteases. These proteases are critical proteins in pathogenesis of Alzheimer's disease. The function of recently identified presenilin-homologous proteases (IMPAS or SPP)s is unknown. Our preliminary data in C.elegans model suggested the role of these proteins in early-development and , perhaps, in pathway related- to cholesterol -regulated signalling (Grigorenko et al, 2004, PNAS).
The overall goal is to determine pattern of gene expression alterations in cells with knock-out or knock-down presenilin related proteins (IMPAS). The cultured cells,    C.elegans and mouse models are planned to be used in this study. Initially, we will determine gene expression patterns in human  cells  embryonic kidney cells (HEK 293 cell line) overexpressing different isoforms of hIMP1 protein.
We hypothesize that novel family of presenilin-related proteases is important for cholesterol-regulated intracellular signalling and early  development (including CNS/neuronal development and function).
We will examine in HEK 293 cells effect of over expression of 1) hIMP1 wt protein, 2) over expression of dominant negative isoform of hIMP1 protein  and compare their gene expression patterns to 3) HEK 293 cells treated with mock vector. HEK 293 cells were seeded on 10 cm plates and transfected next day using Lipofectamine Plus reagent. 24 hours after cells were washed twice with ice cold PBS buffer and lysed with TRizol reagent directly on plates. Total RNA was extracted, purified according the manufacturer protocol and stored at -800C. We will provide 2 RNA samples for each hIMP1 isoform used in two different transfection experiment to reduce any spurious expression differences resulting from culture condition or transfection efficiency variations.
Keywords: dose response

first: Evgeny

middle: I

last: Rogaev


title: Airway epithelium, large and small airways, phenotypically normal smokers, non-smokers, early COPD and COPD


title: Expression data in ezrin-knockdown ec109 cells(PSE1) and control(PS)

status: Public on Nov 09 2006

city: shantou

first: Enmin

last: Li

state: guangdong

address: 22 Xinling Road

country: China

institute: Shantou university, medical college

summary:
Ezrin belongs to the ezrin¨Cradixin¨Cmoesin (ERM) family proteins, which cross-link actin cytoskeleton and plasma membrane, and is actively involved in regulating the growth and metastatic capacity of cancer cells. Esophageal squamous cell carcinoma (ESCC) is one of the most fatal malignancies in the world and the expression of ezrin in ESCC has been described only recently, but its roles and mechanism still remained unclear.
we have used the pSUPER RNAi system to stably suppress the expression of the ezrin gene in EC109, an esophageal squamous carcinoma  cell line, and then cDNA microarray was performed to explore some changed genes with ezrin silence.
Keywords: Gene expression after RNAi

overall_design: he mammalian expression vector, pSUPER.neo.circular (OligoEngine), was used for expression of siRNA in EC109 cells. Briefly, two primer pairs were synthesized, one pair encoding nucleotides 274¨C292 (TCCACTATGTGGATAATAA) followed by a 9 base ¡®¡®loop¡¯¡¯ (TTCAAGAGA) and the inverted repeat (PSE1), and the second encoding nucleotides 265¨C283 (ACTTTGGCCTCCACTATGT) again followed by the loop and inverted repeat (PSE2). And pSUPER.neo vector of nonspecific siRNA was taken as negative control (PSC). The primer pairs were annealed and inserted into the BglII and HindIII sites of pSUPER.neo and transformed into JM109 competent cells (Promega). Positive clones were verified and transfected into EC109 cells using FuGENE 6 transfection reagent (Roche) according to the manufacturer¡¯s instructions. G418 (400 ¦Ìg/ml, Calbiochem) was added into the culture medium after 24 h. Stable G418-resistant clones were obtained in 7¨C9 days. The expanded cells were then used for subsequent studies.

first: Jianjun

last: Xie

first: Liyan

last: Xu


title: An inflammation-induced mechanism for leukocyte transmigration of lymphatic vessel endothelium.

status: Public on Dec 05 2006

city: Oxford

first: Dilair

last: Baban

address: Roosevelt Drive

department: Wellcome Trust Centre Human Genetics

institute: University of Oxford

summary:
The exit of antigen-presenting cells (APC) and lymphocytes from inflamed skin to afferent lymph is vital for the initiation and maintenance of dermal immune responses. How such exit is achieved and how cells transmigrate the distinct endothelium of lymphatic vessels is however unknown. Here we show that inflammatory cytokines trigger activation of dermal lymphatic endothelial cells (LEC) leading to expression of the key leukocyte adhesion receptors ICAM-1, VCAM-1 and E-selectin, as well as a discrete panel of chemokines and other potential regulators of leukocyte transmigration. Furthermore, we show that both ICAM-1 and VCAM-1 are induced in the dermal lymphatic vessels of mice exposed to skin contact hypersensitivity where they mediate lymph node trafficking of DC via afferent lymphatics. Lastly, we show that TNF_-stimulates both DC adhesion and transmigration of dermal LEC monolayers in vitro and that the process is efficiently inhibited by ICAM-1 and VCAM-1 adhesion-blocking mAbs. These results reveal a CAM-mediated mechanism for recruiting leukocytes to the lymph nodes in inflammation and highlight the process of lymphatic transmigration as a potential new target for anti-inflammatory therapy.
Keywords: TNFalpha, Lymphatic endothelium, induction, Inflammation

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Global gene expression profile of normal dermal lymphatic endothelial cells cultured in media alone (no TNF) compared to that of normal dermal lymphatic endothelial cells stimulated with TNFalpha, 1 ng/ml for 48h.Triplicate biological samples were analyzed from human lymphatic endothelial cells (3 x controls; 3 x TNF treated) and a single sample analyzed from mouse lymphatic endothelial cells (1 x controls; 1 x TNF treated).

first: Louise

last: Johnson

first: Steven

last: Clasper

first: Andrew

last: Holt

first: Patricia

last: Lalor

first: David

last: Jackson


title: Identification of ERRalpha dependent PGC-1alpha tagret genes

status: Public on Nov 10 2006

first: Dmitri

last: Kazmin

address: 240 Pasteur Dr

department: Institute for Immunity, Transplantation and Infection

summary:
PGC-1 transcription factor was customized to limit its interations to ERRalpha. This mutant (2x9) was used to dissect the transcription activation patterns that are attributable to the PGC1-ERR interaction and PGC-1 actions that are independent of ERR. Inactive mutant with the deleted LLXXL motifs (L2L3) and wt PGC-1 were used as negative and positive controls respectively. BGAL-expressing construct was used to control for non-specific effects of adenoviral infection.
Keywords: ERRalpha, PGC-1alpha, nuclear receptor, orphan nuclear receptor, coactivator, transcription factors

overall_design: Expression constructs for BGAL control and three variants of PGC-1 were introduced into HepG2 cells via adenoviral infection. RNA was isolated 24 hrs post infection and the changes in gene expression were detected using GeneChip technology (Affymetrix). Hybridization was performed on the HG-133 plus 2 platform. The experiment was performed in three biological replicates. Raw expression data were normalized using MAS5.0 algorithm.

first: Stephanie

last: Gaillard

first: Linda

middle: L

last: Grasfeder

first: Ching-Yi

last: Chang

first: Donald

middle: P

last: McDonnell


title: Timecourse of in vitro erythroid differentiation of human adult-derived peripheral blood CD34+ cells

city: Philadelphia

first: Sankar

last: Addya

state: PA

address: 233 South 10th Street

department: Kimmel Cancer Center

institute: Thomas Jefferson University Hospital

summary:
computational analysis of transcriptomic information during erythroid development
Keywords: erythroid differentiation

overall_design: mRNA expression profiling was used to characterize developing erythroid cells at 6 time points during differentiation

first: Margaret

last: Keller

first: Kathy

last: Delgrosso

first: Saul

last: Surrey


title: NOTCH signaling in T-cell acute lymphoblastic leukemia cell lines

city: Stockholm

last: Brodin

address: Halsovagen 7-9

department: Department of biosciences and Nutrition

institute: Karolinska Institute

summary:
In T-cell acute lymphoblastic leukemia (T-ALL) NOTCH 1 receptors are frequently mutated. This leads to aberrantly high Notch signaling, but how this translates into deregulated cell cycle control and the transformed cell type is poorly understood. In this report, we analyze downstream responses resulting from the high level of NOTCH 1 signaling in T-ALL. Notch activity, measured immediately downstream of the NOTCH 1 receptor, is high, but expression of the canonical downstream Notch response genes HES 1 and HEY 2 is low both in primary cells from T-ALL patients and in T-ALL cell lines. This suggests that other immediate Notch downstream genes are activated, and we found that Notch signaling controls the levels of expression of the E3 ubiquitin ligase SKP2 and its target protein p27Kip1. We show that in T-ALL cell lines, recruitment of NOTCH 1 ICD to the SKP2 promoter was accompanied by high SKP2 and low p27Kip1 protein levels were low. In contrast, pharmacologically blocking Notch signaling reversed this picture and led to loss of NOTCH 1 ICD occupancy of the SKP2 promoter, decreased SKP2 and increased p27Kip1 expression. T-ALL cells show a rapid G1-S cell cycle transition, while blocked Notch signaling resulted in G0/G1 cell cycle arrest, also observed by transfection of p27Kip1 or, to a smaller extent, a dominant negative SKP2 allele. Collectively, our data suggest that the aberrantly high Notch signaling in T-ALL maintains SKP2 at a high level and reduces p27Kip1, which leads to more rapid cell cycle progression.
Keywords: comparative genomic hybridization

overall_design: Three independent cultures of the MOLT4 cell line before and 48 hours after addition of the gamma-secretase inhibitor DAPT (5 uM).

first: Takeaki

last: Dohda

first: Aljona

last: Maljukova

first: Lining

last: Liu

first: Mats

last: Heyman

first: Dan

last: Grandér

first: Olle

last: Sangfelt

first: Urban

last: Lendahl


title: The effects of Serum Amyloid A on gene expression profile in HUVECs

city: Singapore

first: yulan

last: zhao

address: Level 1,  Service Block,  #B1-15

department: Paediatrics

country: Singapore

institute: National University of Singapore

summary:
This study attemps to study the global effects of Serum Amyloid A (SAA) on human endothelial gene expression profile in different time points.
Keywords: time course

overall_design: HUVECs were cultured to confluence in 6-well plates and stimulated with 20 μg/ml of SAA (PeproTech) for 0, 4, 24 and 48 h. To determine the cellular transcript profile in Control (0h) and 3 different SAA groups (S4h, S24h and S48h), the relative abundance of mRNAs in SAA groups were compared with Control by use of  Human Genome U133 Plus 2.0 Array (HG-U133 Plus 2.0, Affymetrix)). Each group had 2 individual samples. . In order to obtain the 3 group comparison datasets (S4h/C, S24h/C and S48h/C), expression patterns for each individual SAA sample (4h, 24h and 48h) were compared to each individual control sample. Hence, each group comparison has 4 individual sample comparisons. The robust increased or decreased cut-off was use the Affymetrix recommending "1" or "-1" of the signal log ratio. . If the gene is robust “increased” or “decreased” in 4 individual sample comparisons of the total 4 comparisons, it was selected as the final robust changed in group comparison. Each gene we were interested, real time PCR was carried out to validate the microarray results.

first: Yulan

last: Zhao

first: HUi-Jen

last: Lye

first: Hendrian

last: Sukardi

first: Chew-Kiat

last: Heng


title: mRNA expression profiling of p63-depleted versus wildtype ME180 cells

city: Boston

first: Kevin

last: Struhl

address: 240 Longwood Ave.

department: Dept. Of Biological Chemistry & Molecular Pharmacology

institute: Harvard Medical School

summary:
We investigated the transcriptional effects of p63 binding by analyzing ME180 cells depleted for all p63 isoforms via expression of a small hairpin RNA (shRNA) targeting the p63 oligomerization domain.
Keywords: differential expression profiling from transcription factor depletion

overall_design: Data were collected from three biological replicates for p63-depleted and control cells respectively.

first: Annie

last: Yang

first: Zhou

last: Zhu

first: Philipp

last: Kapranov

first: Frank

last: McKeon

first: George

last: Church

first: Thomas

last: Gingeras


title: HCaRG vs NEO

status: Public on Oct 27 2006

city: Montreal

first: Johanne

last: Tremblay

state: QUE

institute: CHUM Research Center, Hotel-Dieu

summary:
Summary:
HEK293 cells were transfected with control plasmid (pcDNA1/Neo; Invitrogen) or with the plasmid encoding HCaRG by a standard calcium phosphate co-precipitation method. The clones expressing the highest levels of HCaRG, HCaRG clone 8 and 9 were used in this experiment, while clone transfected with vector alone, Neo clone, served as control.
Stable transfectants were synchronized and grown in the presence of 10% FBS for 48 h. Total RNAs were purified with the mini RNeasy kit (Qiagen).

Keywords: parallel sample

overall_design:
Chips HG-U133 Plus 2.0:
- Labeling protocol: GeneChip IVT  Labeling Kit.
- Hybridization: According to the manufacturer's protocol (Affymetrix).
- Scanner: GeneChip Scanner 3000.
- Normalization: Employing GCOS software, chips were normalized using all probe sets scaling option and target signal at 500.

first: Carlos

last: El Hader

first: Pavel

last: Hamet


title: Transcriptome analysis of the arginine regulon in E.coli

status: Public on Nov 01 2006

city: Leuven

first: Rekin's

last: Janky

address: Herestraat 49 Box 816

department: Nucleomics Core

institute: VIB

summary:
Analysis of the response to arginine of the Escherichia coli K-12 transcriptome by microarray hybridization and real-time quantitative PCR provides a first coherent quantitative picture of the ArgR-mediated repression of arginine biosynthesis and uptake genes. Transcriptional repression was shown to be the major control mechanism of the biosynthetic genes, leaving only limited room for additional transcriptional or post-transcriptional regulations. The art genes coding for the specific arginine uptake system are subject to ArgR-mediated repression,
with a strong repression of artJ, coding for the periplasmic binding protein of the system. The hisJQMP genes of the histidine transporter (part of the LAO uptake system) were discovered to be a part of the arginine regulon. Analysis of their control region with reporter gene fusions and electrophoretic mobility shift in the presence of pure ArgR repressor showed the involvement in repression of the ArgR protein and of an ARG box 120 bp upstream of hisJ.
No repression of the genes of the AO uptake system was observed. Finally, comparing the time course of arginine repression of gene transcription with the evolution of the specific
activities of the cognate enzymes showed that while full genetic repression was achieved two minutes after arginine addition, enzyme concentrations were diluted at the rate of cell
division. This emphasizes the importance of the feedback-inhibition of the first enzymatic
step of the pathway in controlling the metabolic flow through the biosynthesis in the period following the onset of repression.
Keywords: dose response, replicate design, genetic modification design

overall_design:
3 groups of 3 replicate samples were analyzed:
(1) Wild type on minimal medium (strain P4X, slides 1753, 1765, 1989), this created a reference condition

(2) Wild type grown in the presence of arginine (strain P4X plus 100µg/ml of arginine, slides 1754, 1766, 1990), this gave a list of all the genes repressed by the system: ArgR+arginine

(3) Mutant where no active repressor is present; grown in the presence of arginine (strain P4XB2 (argR-) plus 100µg/ml arginine, slides 1992, 1993, 1994). This condition told us which genes are indeed regulated by the system: ArgR+arginine. Those genes are in this case derepressed.

first: Marina

last: Caldara

first: Daniel

last: Charlier

first: Raymond

last: Cunin


title: Gene expression signature of colorectal cancer with microsatellite instability.

status: Public on Oct 17 2006

city: Bunkyo-ku

first: Toshiaki

last: Watanabe

department: Department of Surgical Oncology

institute: University of Tokyo

summary:
Samples were taken from colorectal cancers in surgically resected specimens in 84 colorectal cancer patients. The expression profiles were determined using Affymetrix Human Genome U133 Plus 2.0 arrays. Comparison between the sample groups allow to identify a set of discriminating genes between MSI and MSS cancers and, furthermore, to determine the distinct characteristics of proximal and distal MSI cancers.
Keywords: repeat

overall_design: Eighty-four colorectal cancer patients who had undergone surgical resection of colorectal cancer were studied. To identify molecular signatures of MSI cancers, gene expression profiles were compared between MSI and MSS cancers. Next, we examined the difference in gene expression profiles between proximal and distal MSI cancers.

first: Takashi

last: Kobunai

first: Etsuko

last: Toda

first: Yoshihiro

last: Okayama

first: Yoshikazu

last: Sugimoto

first: Toshinori

last: Oka


title: Cross platform microarray analysis for robust identification of differentially expressed genes

status: Public on Oct 28 2006

city: Nerviano

first: Roberta

last: Bosotti

state: MI

address: viale Pasteur,10

department: Biotechnology

institute: Nerviano Medical Sciences srl

summary:
Microarrays have been widely used for the analysis of gene expression and several commercial platforms are available. The combined use of multiple platforms can overcome the inherent biases of each approach, and may represent an alternative that is complementary to RT-PCR for identification of the more robust changes in gene expression profiles.
In this paper, we combined statistical and functional analysis for the cross platform validation of two oligonucleotide-based technologies, Affymetrix (AFFX) and Applied Biosystems (ABI), and for the identification of differentially expressed genes.

In this study, we analysed differentially expressed genes after treatment of an ovarian carcinoma cell line with a cell cycle inhibitor. Treated versus control RNA was analysed for expression of 16425 genes represented on both platforms.
We assessed reproducibility between replicates for each platform using CAT plots, and we found it high for both, with better scores for AFFX. We then applied integrative correlation analysis to assess reproducibility of gene expression patterns across studies, bypassing the need for normalizing expression measurements across platforms. We identified 930 genes as differentially expressed on AFFX and 908 on ABI, with ~80% common to both platforms. Despite the different absolute values, the range of intensities of the differentially expressed genes detected by each platform was similar. ABI showed a slightly higher dynamic range in FC values, which might be associated with its detection system. 62/66 genes identified as differentially expressed by Microarray were confirmed by RT-PCR.

In this study we present a cross-platform validation of two oligonucleotide-based technologies, AFFX and ABI. We found good reproducibility between replicates, and showed that both platforms can be used to select differentially expressed genes with substantial agreement. Pathway analysis of the affected functions identified themes well in agreement with those expected for a cell cycle inhibitor, suggesting that this procedure is appropriate to facilitate the identification of biologically relevant signatures associated with compound treatment. The high rate of confirmation found for both common and platform-specific genes suggests that the combination of platforms may overcome biases related to probe design and technical features, thereby accelerating the identification of trustworthy differentially expressed genes.

accepted for publication on BMC Bioinformatics
Keywords: platform comparison

overall_design: Three biological replicates were performed for treatment and control. Biological replicates were pooled to obtain a unique sample for treatment or control, which was then divided to generate three aliquots for each condition (technical replicates) per platform.

first: Giuseppe

last: Locatelli

first: Healy

last: Sandra

first: Emanuela

last: Scacheri

first: Luca

last: Sartori

first: Ciro

last: Mercurio

first: Raffaele

last: Calogero

first: Antonella

last: Isacchi


title: Airway epithelium, large airways, phenotypically normal smokers vs non-smokers, MAS5 (HG-U133 2.0)

city: New York

first: Yael

last: Strulovici-Barel

state: NY

address: 1300 York Avenue

institute: Weill Cornell Medical College

summary:
Upregulation of Expression of the Ubiquitin Carboxyl Terminal Hydrolase L1 Gene in Human Airway Epithelium of Cigarette Smokers
The microarray data deposited here is from 9 HG-U133 Plus 2.0 GeneChips, from 4 normal non-smokers, and 5 phenotypic normal smokers, all large airways. These data are part of a study aimed at  understanding how cigarette smoking modifies neuroendocrine cells, in which microarray analysis with TaqMan confirmation was used to assess airway epithelial samples obtained by fiberoptic bronchoscopy from 81 individuals (normal nonsmokers, normal smokers, smokers with early COPD and smokers with established COPD). Of 11 genes considered to be neuroendocrine cell-specific, only ubiquitin C-terminal hydrolase L1(UCHL1), a member of the ubiquitin proteasome pathway, was consistently upregulated in smokers compared to nonsmokers. Up-regulation of UCHL1 at the protein level was observed with immunohistochemistry of bronchial biopsies of smokers compared to nonsmokers. Interestingly, however, while UCHL1 expression was present only in neuroendocrine cells of the airway epithelium in nonsmokers, UCHL1 expression was also expressed in ciliated epithelial cells in smokers, an intriguing observation in light of recent observations that ciliated cells can are capable of transdifferentiating to other airway epithelium. In the context that UCHL1 is involved in the degradation of unwanted, misfolded or damaged proteins within the cell and is overexpressed in >50% of lung cancers, its overexpression in chronic smokers may represent an early event in the complex transformation from normal epithelium to overt malignancy.
Keywords: smokers vs non-smokers

overall_design: comparison of gene expression in airway epithelial cells of the large airways of healthy smokers vs healthy non-smokers

first: Brendan

last: Carolan

first: Adriana

last: Heguy

first: Ben-Gary

last: Harvey

first: Phil

last: Leopold

first: Barbara

last: Ferris

first: Ronald

last: Crystal


title: Immunoprecipitation of U2AF65 associated mRNAs

status: Public on Oct 14 2006

city: Lisboa

first: Margarida

last: Gama-Carvalho

address: Av. Prof. Egas Moniz

country: Portugal

institute: Institute of Molecular Medicine

summary:
U2AF65 is an essential splicing factor involved in the 3'splice site recognition dureing the first steps of spliceosome assembly. In addition, this protein has nucleocytoplasmic shuttling activity and the Drosophila homologue has been implicated in mRNA export.
We have used microarray hybridization coupled to RNA immunoprecipitation from HeLa cell cytoplasmic extracts using anti-U2AF65 antibodies to identify mRNA molecules associated with the U2AF65 splicing factor

Sample preparation methods (Gama-Carvalho et al, 2006, submitted)
Suspension HeLa cells were grown in DMEM, 10% FCS, Pen/Strep and split 1:2 the day before harvesting. Post-nuclear cytoplasmic extracts were obtained as described (Herbert and Hecht 1999), using 5x107 cells/ml of lysis buffer. mRNAs were isolated by immunoprecipitation from 200ul of precleared HeLa cell extract, using the anti-U2AF65 MC3 mAb (Gama-Carvalho, Krauss et al. 1997) or the Bb7 anti-PTB mAb (ATCC) for 2h at 4ºC. Immune complexes were precipitated with a slurry with 50% of protein A/protein G agarose beads (Amersham), blocked with 100ug/ul of tRNA and RNAse free BSA (Ambion), by rotating for 1h at 4ºC and washed  with lysis buffer. Complexes bound to the beads were eluted with TES buffer (10mM Tris, 0.5M EDTA, 0.5% SDS, pH 8.0) by heating at 65ºC for 10 minutes and Trizol extracted. Polyadenilated RNA in the cell extract (input) and in the immunoprecipitation sample was reverse transcribed, end-tagged and PCR amplified using the Super SMART PCR cDNA synthesis kit from Clontech, following manufacturer instructions. 1ug of input or precipitated RNA was used per RT-PCR reaction to produce samples for microarray hybridization. For each set of samples (input and immunoprecipitated) a 100ul test PCR reaction was performed according to manufacturer instructions and analyzed by agarose gel electrophoresis to select ideal conditions for amplification in the linear range. PCR reactions were pooled to obtain enough material for microarray hybridization. 15ug of PCR amplified cDNA from input and immunoprecipitated samples were fragmented, end labeled with biotin and hybridized to Affymetrix GeneChip Human Genome U133 Plus 2.0 Arrays as described (Brodsky, Meyer et al. 2005), following standard Affymetrix hybridization protocols.

Brodsky, A. S., C. A. Meyer, et al. (2005). "Genomic mapping of RNA polymerase II reveals sites of co-transcriptional regulation in human cells." Genome Biol 6(8): R64.
Gama-Carvalho, M., R. D. Krauss, et al. (1997). "Targeting of U2AF65 to sites of active splicing in the nucleus." J Cell Biol 137(5): 975-87.
Herbert, T. P. and N. B. Hecht (1999). "The mouse Y-box protein, MSY2, is associated with a kinase on non-polysomal mouse testicular mRNAs." Nucleic Acids Res 27(7): 1747-53.

Keywords: RNA immunoprecipitation

overall_design: RNA immunoprecipitation was performed from HeLa cell extracts with anti-U2AF65 antibodies. For each immunoprecipitation experiment, two Affimetrix microarrays were hybridized with equal amount of total extract cDNA or cDNA from the immunoprecipitated sample. This series contains microarray data from three independant RNA immunoprecipitation experiments

first: Nuno

last: Barbosa-Morais

first: Alexander

last: Brodsky

first: Pamela

last: Silver

first: Maria

last: Carmo-Fonseca


title: Immunoprecipitation of PTB65 associated mRNAs

summary:
PTB is multifunctional RNA binding protein reported to be involved in splicing, 3' -end processing, stability and translational regulation.

Sample preparation methods (Gama-Carvalho et al, 2006, submitted)
Suspension HeLa cells were grown in DMEM, 10% FCS, Pen/Strep and split 1:2 the day before harvesting. Post-nuclear cytoplasmic extracts were obtained as described (Herbert and Hecht 1999), using 5x107 cells/ml of lysis buffer. mRNAs were isolated by immunoprecipitation from 200ul of precleared HeLa cell extract, using the anti-U2AF65 MC3 mAb (Gama-Carvalho, Krauss et al. 1997) or the Bb7 anti-PTB mAb (ATCC) for 2h at 4ºC. Immune complexes were precipitated with a slurry with 50% of protein A/protein G agarose beads (Amersham), blocked with 100ug/ul of tRNA and RNAse free BSA (Ambion), by rotating for 1h at 4ºC and washed  with lysis buffer. Complexes bound to the beads were eluted with TES buffer (10mM Tris, 0.5M EDTA, 0.5% SDS, pH 8.0) by heating at 65ºC for 10 minutes and Trizol extracted. Polyadenilated RNA in the cell extract (input) and in the immunoprecipitation sample was reverse transcribed, end-tagged and PCR amplified using the Super SMART PCR cDNA synthesis kit from Clontech, following manufacturer instructions. 1ug of input or precipitated RNA was used per RT-PCR reaction to produce samples for microarray hybridization. For each set of samples (input and immunoprecipitated) a 100ul test PCR reaction was performed according to manufacturer instructions and analyzed by agarose gel electrophoresis to select ideal conditions for amplification in the linear range. PCR reactions were pooled to obtain enough material for microarray hybridization. 15ug of PCR amplified cDNA from input and immunoprecipitated samples were fragmented, end labeled with biotin and hybridized to Affymetrix GeneChip Human Genome U133 Plus 2.0 Arrays as described (Brodsky, Meyer et al. 2005), following standard Affymetrix hybridization protocols.

Brodsky, A. S., C. A. Meyer, et al. (2005). "Genomic mapping of RNA polymerase II reveals sites of co-transcriptional regulation in human cells." Genome Biol 6(8): R64.
Gama-Carvalho, M., R. D. Krauss, et al. (1997). "Targeting of U2AF65 to sites of active splicing in the nucleus." J Cell Biol 137(5): 975-87.
Herbert, T. P. and N. B. Hecht (1999). "The mouse Y-box protein, MSY2, is associated with a kinase on non-polysomal mouse testicular mRNAs." Nucleic Acids Res 27(7): 1747-53.

Keywords: RNA immunoprecipitation

overall_design: We have used microarray hybridization coupled to RNA immunoprecipitation from HeLa cell cytoplasmic extracts using anti-PTB antibodies to identify mRNA molecules associated with the PTB splicing factor