title: Gene expression profiling of human alveolar macrophages infected by B. anthracis spores

status: Public on Sep 07 2009

city: Richmond

first: Mikhail

last: Dozmorov

state: VA

address: 830 E Main St

department: Biostatistics

institute: Virginia Commonwealth University

summary: Bacillus anthracis is a gram-positive, aerobic, spore-forming, rod-shaped bacterium which has recently been used as an agent of bioterrorism. Because there is a significant delay between the initial contact of the spore with the host and clinical evidence of disease, there appears to be temporary containment of the pathogen by the innate immune system. Contact with the human alveolar macrophage (HAM) plays a key role in the innate immune response to B. anthracis spores. Therefore, the early macrophage response to anthrax exposure is important in understanding the pathogenesis of this disease.   The majority of genes modulated by spores were upregulated, and a lesser number were downregulated. The data was subjected to Ingenuity Pathway analysis, the Database for Annotation, Visualization and Integrated Discovery (DAVID) analysis, and the Promoter Analysis and Interaction Network Toolset (PAINT). Among the upregulated genes, we identified a group of chemokine ligands, apoptosis genes and, interestingly, keratin filament genes. Central hubs regulating the activated genes were TNF-alpha, NF-κB and their ligands/receptors. Other activated genes included IL-1alpha and IL-18. RNA for these, and several additional cytokines including IL-6, IL-1gamma, IP-10 and GM-CSF, were differentially expressed from 1.6- to 27-fold. The microarray cytokine data is consistent with our previously published findings which demonstrated that there was 4- to 43-fold induction of these cytokines at the transcriptional and translational levels as determined by RNase protection assays and ELISA. The PAINT analysis revealed that the majority of the genes affected by spores contain the binding site for c-Rel, a member of the NF-κB family of transcription factors. Other transcription regulatory elements contained in many of the upregulated genes were c-Myb, CP2, Barbie Box, E2F and CRE-BP1. This study is the first detailed microarray analysis to describe the HAM response to B. anthracis.

overall_design: In this paper, we exposed HAM to B. anthracis Sterne spores at an MOI of 1 for 6 hours. RNA was extracted from HAM and analyzed by the Affymetrix Human Genome U133 Plus 2.0 Array. The transcriptional profile of Bacillus anthracis spore-treated HAM was compared with mock infected cells, and differentially expressed genes were identified.

first: Wenxin

last: Wu

first: Kaushik

last: Chakrabarty

first: Leland

last: Booth

first: Robert

middle: E

last: Hurst

first: Mark

last: Coggeshall

first: Jordan

middle: P

last: Metcalf


title: Molecular profiling of ERBB2-amplified breast cancers

status: Public on Sep 02 2009

city: Marseille

first: Pascal

last: FINETTI

state: BdR

address: 232 Bd Ste Marguerite

department: Centre de cancérologie de Marseille

country: France

institute: Institut Paoli-Calmettes

type:
Expression profiling by array
Genome variation profiling by array

summary: 15-25% of breast cancers (BC) show ERBB2-amplification and overexpression of the encoded ERBB2 tyrosine kinase receptor. They are associated with a poor prognosis but can benefit from targeted therapy. A better knowledge of these BCs may help understand their behavior and design new therapeutic strategies. In this study, we defined the high resolution genome and gene expression profiles of 54 ERBB2-amplified BCs using 244K oligonucleotide array-comparative genomic hybridization and whole-genome DNA microarrays.   We first identified the ERBB2-C17orf37-GRB7 genomic segment as the minimal common amplicon, and CRKRS and IKZF3 as the most frequent centromeric and telomeric amplicon borders, respectively. Second, we identified 17 genome regions affected by copy number aberration (CNA). The expression of 37 genes of these regions was deregulated. Third, two types of heterogeneity were observed in ERBB2-amplified BCs. The genomic profiles of estrogen receptor-postive (ER+) and negative (ER-) ERBB2-amplified BCs were different. The WNT/ß-catenin signaling pathway was involved in ER- ERBB2-amplified BCs, and PVT1 and TRPS1 were candidate oncogenes associated with ER+ ERBB2-amplified BCs. The size of the ERBB2-amplicon was different in inflammatory (IBC) and non inflammatory BCs. ERBB2-amplified IBCs were characterized by the downregulated and upregulated mRNA expression of ten and two genes in proportion to CNA, respectively.  We have shown that ERBB2 BCs are heterogeneous and identified genomic features that may be useful in the design of therapeutical strategies

overall_design: Tumor tissues were collected from 340 patients with invasive adenocarcinoma who underwent initial surgery at the Institut Paoli-Calmettes (Marseilles, France) between 1987 and 2007 (from a cohort of 2,175 patients with frozen tumor sample) and from a series of 91 Tunisian T4d tumors (TNM, UICC) treated between 1994 and 1998 at the Salah Azaiz Institute (Tunis, Tunisia). Each patient gave informed consent and the study was approved by our institutional review board. Samples were macrodissected and frozen in liquid nitrogen within 30 minutes of removal. Genomic imbalances were determined by aCGH using 244K CGH oligonucleotide microarrays (Hu-244A, Agilent Technologies). Gene expression data of 51 of the 54 BCs and 4 normal breast (NB) samples (NB1, NB2, NB3 and NB4, representing samples from 4 women and 3 commercial pools of respectively 1, 2 and 4 normal breast RNA, Clontech, Palo Alto, CA) were quantified by using whole-genome DNA microarrays (HG-U133 plus 2.0, Affymetrix).

first: F

last: Sircoulomb

first: P

last: Finetti

first: J

last: Adelaïde

last: Bertucci

first: D

last: Birnbaum

first: M

last: Chaffanet


title: EMT Time series in ARPE19

status: Public on Aug 26 2009

city: Kisarazu

first: Rui

last: Ose

state: Chiba

address: 2-6-7 Kazusa-Kamatari

department: Department of Human Genome Research

country: Japan

institute: Kazusa DNA Research Institute

summary:
The purpose of this study is to determine the changes in gene expression by a human retinal pigment epithelium (RPE) cell line (ARPE-19) in response to combination treatment of TGF and TNF, which induces phenotypic changes in vitro that mimic the EMT (Epithelial-to-Mesenchymal Transition).
For this purpose, total RNA was extracted from TGF and TNF-treated ARPE-19 cells and differential gene expression between each time point (0, 1, 6, 16, 24, 42, and 60 hours) was determined using genechip arrays (Affymetrix, Human Genome U133).

Keywords: time course

overall_design: ARPE19 cell lines treated with TGF and TNF for 0, 1, 6, 16, 24, 42, and 60 hour. Each experiment were repeated three times. But 1 hour experiment was repeated two times.


title: Expression data in WT and ES cells overexpressing Satb1

status: Public on Sep 01 2009

city: Freiburg

first: Rudolf

last: Engelke

address: Stuebeweg 51

country: Germany

institute: Max Planck Institute of Immunobiology

summary:
Satb1 and Satb2 are regulators of higher order chromatin in T cells and osteoblasts repectively. We were interested if Satb1 and Satb2 play a role in the regulation of gene expression in ES cells.
ES cells which can inducibly express Satb1 or Satb2 were generated using the tet-ON system. In this experiment, we compared gene expression in uninduced cells to that in cells which had been induced for 24h to express Satb1 ectopcially at very high levels.

overall_design: Cells were cultured, passaged every 3 days. After trypsinisation cells were grown for 1 day in the presence of LIF only, after 1 day a set of cultures was induced by the addition of DOX to the culture medium. Induction took place for 24h, RNA was generated 48h after passaging.

first: Fabio

last: Savarese

last: Nechanitzky

last: Grosschedl


title: Trachea Epithelium as a “ Canary” for Cigarette Smoking-induced Biologic Phenotype of Small Airway Epithelium

status: Public on Aug 27 2009

city: New York

first: Yael

last: Strulovici-Barel

state: NY

address: 1300 York Avenue

department: Department of Genetic Medicine

country: USA

institute: Weill Cornell Medical College

summary: The initial site of smoking-induced lung disease is the small airway epithelium, which is difficult and time consuming to sample by fiberoptic bronchoscopy. We developed a rapid, office-based procedure to obtain trachea epithelium without conscious sedation from healthy nonsmokers (n=26) and healthy smokers (n=19, 27 ± 15 pack-yr). Gene expression differences [fold-change >1.5, p< 0.01, Benjamini-Hochberg correction] were assessed with Affymetrix microarrays. 1,057 probe sets were differentially expressed in healthy smokers vs nonsmokers, representing >500 genes. Trachea gene expression was compared to an independent group of small airway epithelial samples (n=23 healthy nonsmokers, n=19 healthy smokers, 25 ± 12 pack-yr). The trachea epithelium is more sensitive to smoking, responding with 3-fold more differentially-expressed genes than small airway epithelium. The trachea transcriptome paralleled the small airway epithelium, with 156 of 167 (93%) genes that are significantly up- and down-regulated by smoking in the small airway epithelium showing similar direction and magnitude of response to smoking in the trachea. Trachea epithelium can be obtained  without conscious sedation, representing a less invasive surrogate “canary” for smoking-induced changes in the small airway epithelium. This should prove useful in epidemiologic studies correlating gene expression with clinical outcome in assessing smoking-induced lung disease.

overall_design: Tracheal gene expression: matched group of small airway epithelial samples (n=23 healthy non-smokers, n= 19 healthy smokers)

first: Meredith

middle: L

last: Turetz

first: Timothy

last: O'Connor

first: Ann

last: Tilley

first: Jacqueline

last: Salit

first: David

last: Dang

first: Matthew

last: Teater

first: Jason

last: Masey

first: Andrew

middle: G

last: Clark

first: Ronald

last: Crystal


title: A Predictive Response Signature to Infliximab Treatment in Ulcerative Colitis

status: Public on Aug 25 2009

city: Spring House

first: K

last: Li

state: PA

address: 1400 McKean Road

institute: Janssen R&D

summary:
Infliximab, an anti-TNFa monoclonal antibody, is an effective treatment for ulcerative colitis (UC) inducing over 60% of patients to respond to treatment. Consequently, about 40% of patients do not respond. This study analyzed mucosal gene expression from patients enrolled in ACT1 to provide a predictive response signature for infliximab treatment.

Keywords: predictive response signature

overall_design: Twenty-two patients underwent colonoscopy with biopsy before infliximab treatment. Response to infliximab was defined as endoscopic and histologic healing at week 8 (P2, 5, 9, 10, 14, 15, 16, 17, 24, 27, 36, and 45 as responders; P3, 12, 13, 19, 28, 29, 32, 33, 34, and 47 as non-responders). Messenger RNA was isolated from pre-infliximab biopsies, labeled and hybridized to Affymetrix HGU133Plus_2.0 Array. The predictive response signature was verified by an independent data set.

first: Katherine

first: Frédéric

last: Baribaud

first: Gary

last: Toedter

first: Keying

last: Ma

first: Xiao-Yu

last: Song

first: Colleen

last: Marano


title: GRE specific DNA-binding polyamide effect on dexamethasone-stimulated lung adenocarcinoma cell line

city: Pasadena

first: Peter

last: Dervan

state: CA

address: 1200 East California Blvd

institute: California Institute of Technology

summary: Analysis of dexamethasone-stimulated A549 lung adenocarcinoma epithelial cells treated with a glucocorticoid response (GR) element (GRE) specific DNA binding polyamide. Polyamide designed to target the sequence 5'-WGWWCW-3' and disrupt GR-mediated gene expression. Effects of the GR antagonist mifepristone  also examined.

overall_design: A549 cells were treated with compounds for 48 hours before RNA extraction and hybridization on Affymetrix microarrays.

first: Katy

middle: A

last: Muzikar

first: Nicholas

last: Nickols

middle: B


title: Expression data from human fibroblasts

status: Public on Aug 21 2009

city: Boston

first: Ben

middle: S.

last: Wittner

state: MA

address: 149 13th Street

department: Center for Cancer Research

institute: Massachusetts General Hospital

summary: Cancer cells interact with surrounding stromal fibroblasts during tumorigenesis, but the complex molecular rules that govern these interactions remain poorly understood, thus hindering the development of therapeutic strategies to target cancer stroma. We have taken a mathematical approach to begin defining these rules by performing large-scale quantitative analysis of fibroblast effects on cancer cell proliferation across more than four hundred heterotypic cell line pairings. Systems-level modeling of this complex dataset using singular value decomposition revealed that normal tissue fibroblasts variably express at least two functionally distinct activities, one which reflects transcriptional programs associated with activated mesenchyme, that act either coordinately or at cross-purposes to modulate cancer cell proliferation. To gain insight into the molecular identity of these fibroblast activities, we isolated RNA from 36 human skin and lung fibroblast cell line monocultures from Coriell Repositories or ATCC and performed microarray-based gene expression profiling using Affymetrix gene chips.

overall_design: Human skin and lung fibroblasts purchased from Coriell Repositories or ATCC were grown using standard cell culture conditions followed by flash freezing in liquid nitrogen, RNA isolation using Qiagen RNeasy kits, and microarray profiling

first: R

middle: C

last: Wadlow

first: B

middle: S

first: S

last: Finley

first: H

last: Bergquist

last: Upadhyay

last: Finn

first: M

last: Loda

first: U

last: Mahmood

last: Ramaswamy


title: Expression data of human neuroblastoma tissue samples

city: Hiroshima

first: Eiso

last: Hiyama

state: Hiroshima

address: 1-2-3, Kasumi, Minami-ku

department: Natural Science Center for Basic Research and Deve

institute: Hiroshima University

summary:
The prognosis of the patients with neuroblastoma largely depends on the biological characterisitics. Neuroblastoma tissues obtained before any treatments were analyzed for gene expression using Affymetrix array.
We used microarrays to detail the global programme of gene expression distinguishing tumor biology of neuroblastoma tisses from the prognosis of the patients.

overall_design: 51 neuroblastoma samples were selected those resected from the patients who died of tumor progression and those whose tumor regressed or matured spontaneously. Total RNA was extracted from these tissues samples and hybridized on Affymetrix microarrays. We sought to obtain genes to distinguish unfavorable tumors from favorable tumors.

first: Keiko

first: Megu

last: Ohtaki

last: Otani


title: HMVEC cells treated with vascular endothelial growth factor, anthrax edema toxin, and an Epac activator

city: Chicago

first: Robert

last: Doebele

state: IL

address: 5841 S. Maryland Ave. MC 2115

department: Medicine

institute: University of Chicago

summary:
Human microvascular endothelial cells (HMVEC) treated with vascular endothelial growth factor (VEGF), Antrhax Edema Toxin (ET), or the Epac activator, 8-pCPT-2'-O-Me-cAMP (8CPT)

Human microvascular endothelial cells (HMVEC) were treated with VEGF alone or VEGF in combination with either the the Epac-specific cAMP-mimetic, 8-pCPT-2'-O-Me-cAMP (8CPT), or anthrax edema toxin (ET), an adenylyl cyclase.  ET or 8CPT can inhibit VEGF-mediated chemotaxis and angiogenesis.  The goal of the study was to identify genes regulated by cAMP production (ET) or by activation of Epac/Rap (8CPT) that may mitigate the effects of VEGF treatment.

overall_design: Gene expression was measured 4 hours after treatment with VEGF, VEGF + 8CPT, VEGF + ET or mock treatment.  Each sample contained one replicate.

first: Jia

last: Hong

first: Marsha

middle: R

last: Rosner


title: Transcriptional profiling of human neural crest cells

first: Heather

last: Etchevers

address: 27 boulevard Jean Moulin

department: INSERM U1251

institute: INSERM/Aix-Marseille Univ

summary: The expression profiles of five human trunk level neural crest cell lines were determined on Affymetrix chips HG U133 Plus 2.0.

overall_design: Total RNA was extracted from a confluent 10 cm dish of hNCC (lines N1-N5) from five human embryos at 26-32 dpf (Carnegie stages 12 & 13).

first: Sophie

last: THOMAS


title: Inhibition of Chlamydia pneumoniae Replication in Human Dendritic cells by TNF-alpha-Induced Indoleamine 2,3-Dioxygenase

city: Braunschweig

last: Geffers

address: Inhoffenstr. 7

department: Dep. Molecular Bacteriology

institute: HCI - Helmholtz Centre for Infection Research

summary:
Infection with Chlamydia pneumoniae, a human respiratory pathogen, has been associated with various chronic diseases such as asthma, coronary heart disease and importantly atherosclerosis. Possibly because the pathogen can exist in a persistent form. TNF-a has been reported to induce chlamydial persitence in epithelial cell lines, however the mechanism of TNF-a-induced persistence has not been reported. Moreover, C. pneumoniae persistently infect human dendritic cells (DCs) and activate DCs to produce cytokines including TNF-a. Induction of chlamydial persistence by other cytokines such as IFN-g is known to be due to indoleamine 2,3-dioxygenase (IDO) activity. The present study therefore, investigated whether C. pneumoniae infection can induce IDO activity in dendritic cells, and whether the restriction of chlamydial growth in the DCs by TNF-a is IDO-dependent.  Our data indicate that infection of DCs with C. pneumoniae resulted in the induction of IDO expression. Reporting on our use of anti-TNF-a antibody adalimumab and varying concentrations of TNF-a, we further demonstrate that IDO induction following infection of DCs with C. pneumoniae is TNF-a-dependent. The anti-chlamydial activity induced by TNF-a and the expression of chlamydial 16S rRNA gene, euo, groEL1, ftsk and tal genes was correlated with the induction of IDO. Addition of excess amounts of tryptophan to the DC cultures resulted in abrogation of the TNF-a-mediated chlamydial growth restriction. These findings suggest that infection of DCs by C. pneumoniae induces production of functional IDO, which subsequently causes depletion of tryptophan. This may represent a potential mechanism for DCs to restrict bacterial growth in chlamydial infections.

Keywords: Chlamydia pneumoniae, Dendritic cells, TNF-a, Indoleamine 2,3-dioxygenase

overall_design: Human monocyte-derived DCs from five heathy donors were infected with C. pneumoniae at multiplicity of infection of 5 for 72 h. Mock-infected (uninfected) DCs were used as a control for the infection. Total RNA from 72 h chlamydiae-infected or mock-infected DCs was isolated with RNeasy Mini Kit (Qiagen, Hilden, Germany) and DNA contamination was removed with RQ1 DNase (Promega). Total RNA from all donors was pooled and the samples were processed and analyzed at the array facility of Helmholtz center for infection biology, Braunschweig. The integrity of the RNA was verified on the Agilent Bioanalyzer system. We used the Affymetrix Gene Chip Human Genome U133 2.0 Plus. Data were processed using Affymetrix software and signal intensity measured was normalized by the Affymetrix MAS5 algorithm.  Data were filtered to exclude genes at background levels (non-expressors, signal intensity at 100).

first: F

last: Njau

first: AD

last: Wagner


title: Gene expression profiles differentiating gastrointestinal stromal tumours according to KIT mutations and expression

city: Warszawa

first: Krzysztof

last: Goryca

address: Banacha 2c

department: CeNT

country: Poland

institute: Uniwersytet Warszawski

summary: Gastrointestinal stromal tumours (GISTs) represent a heterogeneous group of tumours of mesenchymal origin characterized by gain-of-function mutations in KIT or PDGFRA of the type III receptor tyrosine kinase family. Although mutations in either receptor are thought to drive an early oncogenic event through similar pathways, two previous studies reported the mutation-specific gene expression profiles. However, their further conclusions were rather discordant. To clarify the molecular characteristics of differentially expressed genes according to GIST receptor mutations, we combined microarray-based analysis with detailed functional annotations.

overall_design: 29 samples: 15 with KIT mutation detected, 11 with PDGFRA mutation detected, 3 with no mutation detected

first: Jerzy

last: Ostrowski

first: Marcin

last: Polkowski

first: Agnieszka

last: Paziewska

first: Magdalena

last: Skrzypczak

first: Tymon

last: Rubel

first: Katarzyna

last: Kokoszyńska

first: Piotr

last: Rutkowski

first: Zbigniew

middle: I

last: Nowecki

first: Anna

last: Jerzak

first: Dorota

last: Jarosz

first: Wlodzimierz

last: Ruka

first: Lucjan

last: Wyrwicz


status: Public on Aug 20 2009

city: Ann Arbor

first: Rork

last: Kuick

state: MI

address: University of Michigan, SPH II, 1415 Washington Heights, Room M2533

institute: University of Michigan

summary:
Time Course of TGF-beta treatment of A549 lung adenocarcinoma cell line on Affymetrix HG_U133_plus_2 arrays; triplicate experiments.

The goal of the experiment is to profile temporal gene expression changes during TGF-beta-induced epithelial-mesenchymal transition (EMT).   During EMT cancer cells loose their epithelial specifc proteins and gain mesenchymal proteins to acquire migratory and invasive phenotype essential for metastasis. Human A549 lung adenocarcinoma cell line was treated with 5 ng/mL TGF-beta for 0, 0.5, 1, 2, 4, 8, 16, 24, and 72 h to induce EMT.  The experiment was repeated 3 times.  Samples were assayed using Affymetrix HG_U133_plus_2 arrays with 54675 probe-sets, using standard techniques. We provide the raw .CEL files and a supplementary Excel spreadsheet with log-transformed data and selected results from a statistical analysis.

overall_design: Human A549 lung adenocarcinoma cell line was treated with 5 ng/mL TGF-beta for 0, 0.5, 1, 2, 4, 8, 16, 24, and 72 h.  The experiment was repeated 3 times.  Samples were assayed using Affymetrix HG_U133_plus_2 arrays with 54675 probe-sets, using standard techniques.  The 2 h sample of the third experiment was not run on an array due to poor RNA, so that only 26 arrays were run.

first: Venkateshwar

last: Keshamouni

first: Maureen

last: Sartor

first: Vasudeva

last: Mahvisno

first: James

last: Cavalcoli

first: Zachary

last: Wright

first: Alla

last: Karnovsky

first: H

middle: V

last: Jagadish

first: Barbara

last: Mirel

first: Terry

last: Weymouth

first: Brian

last: Athey

first: Gilbert

last: Omenn


title: Identifying the molecular signature of the interstitial del(7q) subgroup of uterine leiomyomata using a paired analysis

status: Public on Aug 17 2009

city: Rochester

first: Jennelle

last: Hodge

state: MN

address: 200 First Street SW

department: Department of Laboratory Medicine and Pathology

institute: Mayo Clinic

summary:
Uterine leiomyomata (UL), the most common neoplasm in reproductive-age women, have recurrent cytogenetic abnormalities including del(7)(q22q32).  To develop a molecular signature, matched del(7q) and non-del(7q) tumors identified by FISH or karyotyping from 11 women were profiled with expression arrays.  Our analysis using paired t-tests demonstrates this matched design is critical to eliminate confounding effects of genotype and environment that underlie patient variation.  A gene list ordered by genome-wide significance showed enrichment for the 7q22 target region.  Modification of the gene list by weighting each sample for percent of del(7q) cells to account for the mosaic nature of these tumors further enhanced the frequency of 7q22 genes.  Pathway analysis revealed two of the 19 significant functional networks were associated with development and the most represented pathway was protein ubiquitination, which can influence tumor development by stabilizing oncoproteins and destabilizing tumor suppressor proteins.  Array CGH (aCGH) studies determined the only consistent genomic imbalance was deletion of 9.5 megabases from 7q22-7q31.1.  Combining the aCGH data with the del(7q) UL mosacism-weighted expression analysis resulted in a list of genes that are commonly deleted and whose copy number is correlated with significantly decreased expression.  These genes include the proliferation inhibitor HPB1, the loss of expression of which has been associated with invasive breast cancer, as well as the mitosis integrity-maintenance tumor suppressor RINT1.  This study provides a molecular signature of the del(7q) UL subgroup and will serve as a platform for future studies of tumor pathogenesis.
Keywords: uterine leiomyomata, fibroids, del(7)(q22q32), gene expression, aCGH, microarray

overall_design: Matched del(7q) and non-del(7q) tumors identified by FISH or karyotyping from each of eleven woman were profiled using Affymetrix GeneChip U133 Plus 2.0 oligonucleotide gene expression arrays.

first: Peter

middle: J

last: Park

first: Jonathan

middle: M

last: Dreyfuss

first: Iman

last: Assil-Kishawi

first: Priya

last: Somasundaram

first: Luwam

last: Semere

first: Bradley

last: Quade

first: Allison

last: Lynch

first: Elizabeth

last: Stewart

first: Cynthia

last: Morton


title: Expression data from CD4-positive HTLV-positive cell lines and from CD4-positive HTLV-negative primary cells.

city: Erlangen

first: Ralph

last: Grassmann

address: Schlossgarten 4

department: Institute of Clinical and Molecular Virology

institute: Universität Erlangen Nürnberg

summary: We used microarrays to compare the global programme of gene expression in HTLV-positive, ATL-derived and HTLV-positive in vitro-transformed cell lines with that of uninfected primary CD4 T cells.

overall_design: Cells were harvested one day after splitting, in case of primary T cells, cells were harvested after reaching a postmitotic state.

first: Andrea

middle: K

last: Kress


title: Gene expression profiling from MM1.S cells with control or beta-catenin knockdown.

status: Public on Aug 24 2009

first: Daniel

middle: Ruben

last: Carrasco

address: 44 Binney St

department: Medical Oncology

institute: Dana-Farber Cancer Institute

summary: MM1.S cells stably transduced with control or b-catenin shRNA were established. Total RNA was isolated from 5x 10^6 cells of each in triplicate.

overall_design: Comparative gene expression profiling in multiple myeloma.

first: Jui

last: Dutta-Simmons

first: Mala

last: Mani

first: Kenneth

last: Anderson


title: Comparison of Human and Rat Uterine Leiomyoma: Identification of a Dysregulated mTOR Pathway

city: Cambridge

first: Scott

last: Jelinsky

address: 610 Main Street

department: Inflammation and Immunology

institute: Pfizer

summary:
Uterine leiomyomata, or fibroids, are benign tumors of the uterine myometrium that significantly affect up to 30% of reproductive-age women. Despite being the primary cause of hysterectomy in the United States, accounting for up to 200,000 procedures annually, the etiology of leiomyoma remains largely unknown.  Due to the lack of an effective medicinal therapy for these tumors, this disease continues to have a tremendous negative impact on women’s health. As a basis for understanding leiomyoma pathogenesis and identifying targets for pharmacotherapy, we conducted transcriptional profiling of leiomyoma and unaffected myometrium from humans and Eker rats, the best characterized preclinical model of leiomyoma. A global comparison of mRNA from leiomyoma versus myometrium in human and rat identified a highly significant overlap of dysregulated gene expression in leiomyoma.  An unbiased pathway analysis using a method of gene set enrichment based on the Sigpathway algorithm detected the mammalian target of rapamycin (mTOR) pathway as one of the most highly upregulated pathways in both human and rat tumors. Activation of this pathway was confirmed in both human and rat leiomyomata at the protein level via Western.  Inhibition of mTOR in female Eker rats with the rapamycin analog WAY-129327 for 2 weeks decreased mTOR signaling and cell proliferation in tumors, and treatment for 4 months significantly decreased tumor incidence, multiplicity and size. These results identify dysregulated mTOR signaling as a component of leiomyoma etiology across species and directly demonstrate the dependence of these tumors on mTOR signaling for growth in the Eker rat. Modulation of this pathway warrants additional investigation as a potential therapy for uterine leiomyoma.

Keywords: Disease State Analysis: Animal Model Validation

sample_organism:
Homo sapiens
Rattus norvegicus

platform_organism:
Homo sapiens
Rattus norvegicus

overall_design: We analyzed 1-3 leiomyoma or normal myometrium biopsies from each 23 woman undergoing hysterectomy for the treatment of uterine fibroids.  tment and compared it leiomyoma and normal myometrium from the Eker rat model of uterine fibroids (N=14-15)


city: Berlin

first: Lars

last: Bullinger

address: Augustenburger Platz 1

department: Hematology, Oncology and Tumorimmunology

institute: Charité

summary:
Thalidomide Exerts Distinct Molecular Antileukemic Effects and Combined Thalidomide/Fludarabine Therapy is Clinically Effective in High-Risk Chronic Lymphocytic Leukemia

Background: Thalidomide represents a promising immunomodulatory drug that targets both leukemia cells and the tumor microenvironment.
Methods: We treated chronic lymphocytic leukemia (CLL) patients with a combined thalidomide/fludarabine regimen and monitored cellular and molecular changes induced by thalidomide in-vivo prior to fludarabine treatment. Thalidomide was given daily (100mg p.o./day) and fludarabine was administered on days 7-11 (25 mg/m² i.v./day) within each 4-week cycle (maximum of 6 cycles). Twenty patients received thalidomide/fludarabine as first line therapy and 20 patients were previously treated. Unmutated IgVH mutation status was found in 36 cases and 13 had high-risk cytogenetic aberrations (deletion of 17p13 or 11q22-q23).
Results: The overall response rate was 80% and 25% for untreated and previously treated patients, respectively. While thalidomide effectively reduced the number of CLL cells, the number of CD3 lymphocytes showed no significant change, but the number of CD4+CD25hiFOXP3+ T-regulatory cells was significantly decreased. Gene expression profiling revealed a thalidomide induced signature containing both targets known to play a role in immunomodulatory drug action as well as novel candidate genes.
Conclusions: Combined thalidomide/fludarabine therapy demonstrated efficacy in high-risk CLL patients. Furthermore, our study provides novel biological insights into thalidomide effect, which might act by enhancing apoptosis of CLL cells and reducing Tregs, thereby enabling T-cell dependent anti-tumor effect.

overall_design: Patients ("preTHAL") received thalidomide (100mg p.o./day) starting at day 0 and at day 7 Fludarabine therapy was added to the therapy. To evaluate the in-vivo influence of thalidomide on the transcriptome in CLL, paired GEP was performed in 20 cases at treatment start at day 0 (preTHAL) and prior to fludarabine treatment at day 7 (postTHAL) using HumanGenome U133 Plus2.0 Arrays according to the manufacturer's recommendations (Affymetrix; RNA was isolated from peripheral blood samples enriched for leukemic cells by Ficoll-density gradient centrifugation, percentage of leukemic cells > 90%). Fluorescence ratios were evaluated by both the GCOS software and normalized by applying the RMA algorithm using the BRB Array Tools software.


title: gene expression in human subcutaneous adipose tissue after CLA intervention

city: Kiel

first: Julia

last: Herrmann

address: hermann-Weigmann Str

department: PBE

institute: MIR/BFEL

summary: Iisomer-specific effects of conjugated linoleic (CLA) supplementation on gene expression with particular consideration of the PPAR 2 Pro12Ala SNP in human adipose tissue.

overall_design: Effect of CLA supplementation on genome wide gene expression in adipose tissue biopsies from 5 PPARg2 Ala12Ala and 5 PPARg2 Pro12Pro men were investigated. Subjects underwent four intervention periods (4wk) in a randomized double blind cross-over design receiving f either cis-9, trans-11 CLA, trans-10,cis-12 CLA, 1:1 mixture of both isomers or a reference oil preparation. After each intervention biopsies were taken and whole genome expression microarrays were applied.


title: Role of FAK, dominant-negative FAK, FAK-CD and FAKsiRNA in MCF-7 human breast cancer cell tumorigenesis

city: Gainesville

first: Vita

last: Golubovskaya

state: FL

address: 1376 Mowry Road

department: Surgery

institute: university of florida, Shands Cancer Center

summary: Focal adhesion kinase (FAK) is a non-receptor tyrosine kinase that plays an important role in proliferation, motility, adhesion, invasion, angiogenesis, and survival signaling. Focal adhesion kinase has been shown to be overexpressed in many types of tumors, including breast cancer at early stages of tumorigenesis.  To study the biological role of FAK in breast tumorigenesis, we used FAKsiRNA to down-regulate FAK in MCF-7 cell lines.

overall_design: Eight samples were analyzed in MCF-7, MCF-7-Vector, MCF-7 control (luciferase) siRNA and FAKsiRNA#1, FAKsiRNA#2

first: Jian-Liang

last: Li

first: William

last: Cance


title: Letrozole (Femara) early response to treatment

status: Public on Aug 15 2009

city: Houston

first: Chad

last: Creighton

state: TX

address: One Baylor Plaza, Mail Stop: BCM305

department: Biostatistics, Ducan Cancer Center

institute: Baylor College of Medicine

summary:
Tumorigenic breast cancer cells characterized by high CD44 and low or undetectable CD24 levels (CD44+/CD24-/low) may be resistant to conventional therapies and responsible for cancer relapse. We defined a “signature” expression pattern of hundreds of genes associated with CD44+/CD24-/low, mammosphere-forming cells. In a panel of patient breast tumors, this tumorigenic gene signature was found exclusively manifested in tumors of the recently identified “claudin-low” molecular profile subtype characterized by overexpression of many mesenchymal-associated genes, suggesting that these tumors have pre-existing higher levels of tumorigenic cells. Furthermore, when comparing the expression profiles of paired breast cancer core biopsies before versus after hormone therapy or chemotherapy, both the tumorigenic and “claudin-low” signatures were more active in about half of tumors after treatment, indicative of a greater enrichment of tumorigenic cells as a result of treatments targeting the bulk tumor cells.
Keywords: two group comparison

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

first: Michael

last: Lewis

first: Jenny

last: Chang


title: Gene expression data from cancer mammospheres and bulk tumors

summary:
Tumorigenic breast cancer cells characterized by CD44 expression and low or undetectable CD24 levels (CD44+/CD24-/low) may be resistant to chemotherapy and therefore responsible for cancer relapse. Paired breast cancer core biopsies before and after neoadjuvant chemotherapy or lapatinib were obtained and as single cell suspensions stained using antibodies against CD24, CD44, and lineage markers, and then analyzed by flow cytometry. Mammosphere (MS) formation in culture was compared before and after treatment. Global gene expression differences between cancer cells bearing CD44+/CD24-/low cells and all other sorted cells, and between cancer MS and the primary bulk invasive cancers were analyzed. We report that CD44+/CD24-/low tumorigenic breast cancer cells were intrinsically chemoresistant ─ chemotherapy led to increased CD44+/CD24-/low cells, increased self-renewal capacity on MS assays, and enhanced tumorigeneicity in immunocompromised SCID/Beige mice. Conversely, in patients with HER2 overexpressing tumors, the EGFR/HER2 tyrosine kinase inhibitor, lapatinib decreased CD44+/CD24-/low cells, with the majority of these patients after conventional therapy achieving pathologic complete response, a validated surrogate marker for long-term survival. Gene transcription pathways that underlie chemoresistant, MS-forming CD44+/CD24-/low cells involve genes belonging to stem cell self-renewal, Wnt signaling, and early development pathways.
Keywords: two group comparison

overall_design:
Cells from human breast tumors were grown as mammospheres (MS).
Isolated single cell suspensions from primary breast cancers were plated onto non-adherent (polyhema-coated) plastic, counted with a hematocytometer, and 20,000 cells were then seeded into a 6-well ultra-low attachment plate supplemented with 2mL MEGM, with the addition of 2 mL of freshly unfrozen MEGM every 3-4 days.  Gene expression profiles were taken of both MS and primary bulk tumors and compared with each other.


title: Gene expression data from CD44+/CD24- cells sorted by flow cytometry

overall_design:
Human breast tumor samples were sorted using flow cytometry to select for cells that were CD44+ and CD24-.  Gene expression profiles of these cells were compared with profiles of the other sorted cells (CD24+ and CD44-/CD24-).

Core biopsies of primary breast tumors were taken and placed immediately in cold RPMI-1640 supplemented with 10% heat-inactivated newborn calf serum (HINCS, Invitrogen, Carlsbad, CA). Within an hour, the samples were minced and then digested in 10-15 mL of MEGM with 250-300 units/mL collagenase at 370C. The samples were filtered, washed, and then subjected to hypotonic shock  to lyse red blood cells. About 106 single cells were re-suspended, incubated for 15 min at 40C with anti-CD44 (APC), anti-CD24 (FITC), and anti-lineage cocktail antibodies (PE-conjugated anti-CD2, CD3, CD10, CD16, CD18, CD31 and CD 140B) (Pharmingen, San Diego, CA) using the manufacturer’s suggested concentrations. The cells were then washed twice, re-suspended with the viability dye propidium iodide, and analyzed using Dako MoFlo flow cytometry.  Side- and forward- scatter were used to eliminate debris and cell doublets, and the Lin- cells were further analyzed by CD44 and CD24 markers.


title: Antileukemic activity of valproic acid in chronic lymphocytic leukemia cells defined by microarray analysis

city: Bruxelles

first: Basile

last: Stamatopoulos

address: Bd de Waterloo n°121

department: Experimental Hematology

country: Belgium

institute: Jules Bordet Institute

summary:
Epigenetic code modifications by histone deacetylase inhibitors (HDACi) have recently been proposed as potential new therapies for hematological malignancies. Chronic Lymphocytic Leukemia (CLL) remains incurable despite the introduction of new treatments. CLL cells are characterized by an apoptosis defect rather than excessive proliferation, but proliferation centers have been found in organs such as bone marrow and lymph nodes.
Here, we investigated gene expression modifications in CLL cells following treatment with valproic acid (VPA), a well-tolerated anti-epileptic drug with HDAC inhibitory activity.

overall_design: CLL cells obtained from 14 patients were treated in vitro with a concentration of 1mM VPA for 4 hours. VPA effects on gene expression were thereafter studied using Affymetrix technology

last: STAMATOPOULOS

first: Nathalie

last: MEULEMAN

first: Cécile

last: DE BRUYN

first: Philippe

last: MINEUR

last: MARTIAT

first: Dominique

last: BRON

first: Laurence

last: LAGNEAUX


title: Expression data from human Ishikawa cells treated with Bisphenol A

status: Public on Aug 13 2009

city: MASON

first: Jay

middle: Patrick

last: Tiesman

state: OH

address: PO BOX 8006

department: Global Biotechnology

institute: Procter & Gamble

summary:
This study provides a comprehensive evaluation of changes in gene expression during treatment with Bisphenol A in vitro.
Ishikawa cells were maintained and grown (to confluency or at a desired cell density between 500,000 - 1 million cells/mL) in DMEM/F-12 Medium (supplemented with 10% Fetal Bovine Serum + 1X penicillin/streptomycin) in xenoestrogen-free Corning plasticware.  Cells were gently washed in warm PBS and transferred to phenol red-free DMEM/F-12 Medium (supplemented with 10% Charcoal-Stripped Fetal Bovine Serum + 1X penicillin/streptomycin) in Corning Cell Culture Cluster wells overnight and then challenged with 1pM (very low, vL), 100pM (low, L), 10nM (high, H), and 1uM (very high, vH) levels of BPA.  Prior to collection, cells were washed in warm PBS, resuspended & briefly incubated in TRI-Reagent, & finally collected (in quintuplicate replicates) at each specific time point: 8 hours, 24 hours, and 48 hours.  Following RNA isolation, the best RNA yields for each quadruplicate set was selected for target preparation and GeneChip processing.

Keywords: Dose Response/Time Course

overall_design: 60 total samples:  5 Doses [Vehicle Control, Very Low (1 nM), Low (100 nM), High (10 uM), Very High (100 uM)]; 3 Timepoints [8 hour, 24 hour, 48 hour]; 4 Replicates each.


title: Expression profile of grafted human engineered skin substitutes compared with intact human

status: Public on Aug 07 2009

city: Cincinnati

first: Bruce

last: Aronow

institute: Cincinnati Children's Hospital Medical Center

summary:
The goal of the experiment: To characterize the dynamic gene expression profile of engineered human skin in vitro and after grafting, and compare with expression profile of uninjured human skin.

Bioengineered skin substitutes can facilitate wound closure in massively burned patients, but deficiencies limit their outcomes compared to native skin autografts.  To identify gene programs associated with their in vivo capabilities and limitations, we extended previous gene expression profile analyses to now compare engineered skin following in vivo grafting to both in vitro maturation and to normal human skin.  Cultured skin substitutes were grafted to full-thickness wounds in athymic mice; biopsies for microarray analyses were collected at multiple in vitro and in vivo time points.  Over 10,000 transcripts exhibited large-scale expression pattern differences during in vitro and in vivo maturation.  Using hierarchical clustering, eleven different expression profile clusters were partitioned based on differential sample type and temporal stage-specific activation or repression.  Analyses show that the wound environment exerts a massive influence on gene expression in skin substitutes. For example, in vivo healed skin substitutes gained expression of many native skin-expressed genes, including those associated with epidermal barrier and multiple categories of cell-cell and cell-basement membrane adhesion. In contrast, immunologic, trichogenic, and endothelial gene programs were largely lacking.  These analyses suggest important areas to guide further improvement of engineered skin for both increased homology with native skin and enhanced wound healing.

overall_design: In the study, we hybridized RNA isolated from skin substitutes from days 3, 7, or 14 of in vitro incubation, and 3, 7, 14, 28, 42, or 56 days after transplantation to athymic mice, to Affymetrix Human U133 Plus 2.0 gene chips.

first: Aaron

last: Friedman

first: Steven

middle: T

last: Boyce

first: Dorothy

last: Supp


title: Expression data from Chd1-deficient mouse ES cells (E14 cell lines) and genome-wide binding of Chd1 in parental ES cells

city: San Francisco

first: Miguel

last: Ramalho-Santos

address: 35 Medical Center Way

department: Eli and Edythe Broad Center of Regeneration Medicine

institute: UCSF

type: Genome binding/occupancy profiling by genome tiling array

summary:
We used microarrays to study the effect of Chd1 loss of function in mouse ES cells.
We used ChIP-chip to analyze genome-wide binding of Chd1 in normal ES cells.

overall_design:
Mouse Embryonic Stem (ES) cells were infected with a lentiviral vector (pSicoR-mCherry) for expression of a shRNA against Chd1 and GFP (as a control). mCherry-sorted ES cells were plated and individual clones were selected and grown under normal ES cell conditions. 6 ES cell clonal lines were isolated and analyzed along with the parental E14 cell line. The final analysis consisted in 3 control cell lines (E14, E1 and G8) and 4 Chd1-deficient cells (3 different clones using a shRNA sequence #1- C1i5, C1i6 and C1i9; 1 clone using a different shRNA (#4) sequence targeting Chd1- C4i2). RNA samples were isolated from these Chd1-deficient ES cells and control ES cells, and hybridized on Affymetrix chip.
For ChIP-chip, parental E14 ES cell line was used.

first: Alexandre

last: Gaspar-Maia

first: Adi

last: Alajem

first: Fanny

last: Polesso

first: Rupa

last: Sridharan

last: Mason

first: Amy

last: Heidersbach

first: João

last: McManus

first: Kathrin

last: Plath

first: Eran

last: Meshorer


title: Comparison of post-mortem tissue from brain BA10 region between schizophrenic and control patients.

city: Stevenage

middle: MH

last: Molesworth

state: Hertfordshire

address: Gunnels Wood Road

department: R&D Computational Biology

country: United Kingdom

institute: GlaxoSmithKline

summary:
Analysis of gene expression in two large schizophrenia cohorts identifies multiple changes associated with nerve terminal function.
Schizophrenia is a severe psychiatric disorder with a world-wide prevalence of 1%. The pathophysiology of the illness is not understood, but is thought to have a strong genetic component with some environmental influences on aetiology. To gain further insight into disease mechanism, we used microarray technology to determine the expression of over 30 000 mRNA transcripts in post-mortem tissue from a brain region associated with the pathophysiology of the disease (Brodmann area 10: anterior prefrontal cortex) in 28 schizophrenic and 23 control patients.

overall_design: Post-mortem derived BA10 tissue from 28 schizophrenic and 23 control patients were compared. Age, gender, post-mortem delay and pH of brain lysates data were also captured.

first: P

last: Maycox

first: F

last: Kelly

first: A

last: Taylor

first: S

last: Bates

first: J

last: Reid

first: R

last: Logendra

first: M

last: Barnes

first: C

last: Larminie

first: N

last: Jones

last: Lennon

last: Davies

last: Hagan

last: Scorer

last: Angelinetta

first: MT

last: Akbar

last: Hirsch

last: Mortimer

first: T

first: Jackie

last: de Belleroche


title: Transcriptional effects of CRP* expression in Escherichia coli

status: Public on Aug 12 2009

city: University Park

first: Patrick

last: Cirino

email: cirino@engr.psu.edu, rxk249@psu.edu

address: 226A Fenske lab

department: Chemical Engineering

institute: Penn State

summary:
Escherichia coli exhibits diauxic growth in sugar mixtures due to CRP-mediated catabolite repression and inducer exclusion related to phosphotransferase system enzyme activity. Replacement of the native crp gene with a catabolite repression mutant (referred to as crp*) alleviates diauxic effects in E. coli and enables co-utilization of glucose and other sugars. While previous studies have examined the effects of expressing CRP* mutants on the expression of specific catabolic genes, little is known about the global transcriptional effects of CRP* expression.
In this study, we compare the transcriptome of E. coli W3110 (expressing wild-type CRP) to that of mutant strain PC05 (expressing CRP*) in the presence and absence of glucose.

overall_design: Four different conditions were tested in this study: W3110 in LB medium (WT), W3110 in LB+glucose medium (WT G), PC05 in LB medium (CRP*), and PC05 in LB+glucose medium (CRP* G).

first: Reza

last: Khankal

first: Jonathan

middle: W

last: Chin

first: Debashis

last: Ghosh


title: Expression data from FoxM1 siRNA knock down Human aortic vascular smooth muscle cells

first: I-Ching

last: Wang

address: 3333 Burnet Ave.

department: Pulmonary Biology

institute: CCHMC

summary:
Transcription factor FoxM1 is expressed in proliferating cells, and its expression is critical for cell proliferation in embryos and tumors. FoxM1 regulates a multi-gene transcriptional network for cell cycle regulation.

We used microarrays to detail the global program of gene expression either directly or indirectly regulated by FoxM1, and distinct classes of up- and down-regulated genes.

overall_design: Human aortic vascular smooth muscle cells (HAVSMC) were transfected with FoxM1 siRNA and control siRNA for 48 hours. Total RNA was extracted, purified, and prepared for hybridization on Affymetrix microarrays.

first: I

first: Vladimir

last: Kalinichenko


title: Molecular signature of cell cycle exit induced in human T lymphoblasts by IL-2 withdrawal

first: Magdalena

last: Chechlinska

address: Roentgena 5

department: Department of Immunology

institute: Maria Sklodowska-Curie Memorial Cancer Centre and Institute of Oncology

summary:
Molecular mechanisms of cell cycle exit are poorly understood. A group of genes required for cell cycle exit and maintenance of cell quiescence in human fibroblasts following serum deprivation has been recently identified. Studies on lymphocytes following growth factor deprivation-induced cell cycle exit have predominantly focused on the initiation of apoptosis. A set of genes involved in lymphocyte quiescence have also been identified among genes highly expressed in resting lymphocytes and down-regulated after cell activation. In our study, proliferating IL-2-dependent human T cells were forced to exit cell cycle by growth factor withdrawal, and their gene expression profiles were examined.
The differential gene expression analysis was performed in primary and immortalized IL-2-dependent T lymphocytes. Cell samples were collected directly from the IL-2-containing cultures and 8-hrs following IL-2 withdrawal, before apoptosis could be evidenced by the Annexin-V staining. The three primary T lymphoblast cell populations were obtained from the peripheral blood mononuclear cells (PBMC) stimulated for 24h by wheat germ agglutinin and cultured in the presence of IL-2 up to 4-8 population doublings. As shown by the cell surface analysis, these populations were composed of T cells exclusively. Samples of these cell populations were subsequently analyzed as biological replicates. Two spontaneously immortalized IL-2-dependent T cell lines were derived from normal spleen and from PBMC derived from Nijmegen Breakage Syndrome patient. Gene expression was assessed by the Affymetrix microarray HG-U133 2.0 Plus that detects 38,500 genes. The expression of a selected number of genes was verified by the qRT-PCR method.
We have identified a set of 53 genes that we called a “T lymphocyte cell cycle exit signature”, comprised of 13 up-regulated and 40 down-regulated genes. Genes linked to transcription, cell cycle, cell growth, proliferation and differentiation, cell adhesion and immune functions were found to be overrepresented among the differentially expressed, before and after IL-2 deprivation. Among those, PIM1, BCL2, IL-8, HBEGF, DUSP6, OSM, CISH, SOCS2, SOCS3, LIF and IL13 were down-regulated and RPS24, SQSTM1, TMEM1, LRRC8D, ECOP, YY1AP1, C1orf63, ASAH1, SLC25A46 and MIA3 were up-regulated. Identification of genes involved in cell cycle exit and quiescence, may provide new insights into the mechanisms of tissue repair and regeneration as well as of cancer development.

overall_design:
Cell sources and cell sample preparation.
Samples of three primary, IL-2-dependent T lymphoblast cell lines, derived from three healthy donors (j, 43, 6) were collected from IL-2-containing culture and 8-hrs following IL-2 withdrawal (three pairs, each sample was analyzed once = 6 samples).
The two spontaneously immortalized IL-2-dependent T cell lines were derived from normal spleen (line5) and from PBMC derived from a Nijmegen Breakage Syndrome patient (S9). Samples of the two immortalized cell lines were collected in three biological replicates each, from the cultures with and without IL-2 (2 x 2 x 3 = 12 samples).

first: Jan

last: Siwicki

first: Monika

last: Gos

first: Malgorzata

last: Oczko-Wojciechowska

first: Michal

last: Jarzab

first: Aleksandra

last: Pfeifer

first: Barbara

last: Steffen

first: Anna

last: Leonowicz


title: Effect of ectotopic expression of K13 on global gene expression in HUVEC

status: Public on Aug 06 2009

city: Los Angeles

first: Vasu

last: Punj

address: Biggy Street

institute: USC

summary: Integrated microarray and multiplex cytokine analyses of Kaposi's Sarcoma Asssociated Herpesvirus viral FLICE Inhibitory Protein K13 affected genes and cytokines in human blood vascular endothelial cells. The KSHV-encoded K13 protein is one of the few proteins to be expressed in latently-infected spindle cells and the ectopic expression of K13 in human vascular endothelial cells is sufficient to transform them into spindle cells.

overall_design: We have examined the effect of ectopic K13 expression on global gene expression in human umbilical vein endothelial cells (HUVEC).

first: Hittu

last: Matta

first: Sandra

last: Schamus

first: Preet

last: Chaudhary


title: Response of bronchial epithelial cells to low doses of cigarette smoke condensate and subsequent demethylation agent

status: Public on Aug 01 2009

city: Zurich

first: Hubert

last: Rehrauer

address: Winterthurerstr. 190

department: FGCZ

country: Switzerland

institute: ETH Zurich / University of Zurich

summary:
The study seeks to identify the epigenetic changes caused by exposure of to cigarette smoke condensate. To this goal human bronchial epithelial cells, BEAS-2B, were treated with 5-aza-2’deoxycitidine and  trychostatin A (5AzaC/TSA) subsequent to a chronic exposure (1 month) to cigarette smoke condensate (CSC). As negative control served BEAS-2B cells that were untreated or treated with CSC/DMSO for one month without the subsequent application of 5Aza/TSA.

Keywords: stress response

overall_design: BEAS-2B Cells were treated for one month with CSC, DMSO, and left untreated. Subsequently half of the samples were treated with the demethylation agent. So that there were six different conditions with three biological replicates each. One sample had to be excluded because of low quality.

first: Emilija

last: Veljkovic

first: Anne

last: Anstett

first: Mirco

last: Menigatti

first: Alex

last: Soltermann

first: Josef

last: Jiricny

first: Wanjiang

last: Han


title: Detection of Notch1-IC, Notch2-IC and EBNA2 target genes in human B cells

city: Munich

first: Lothar

middle: J.

last: Strobl

address: Marchioninistr. 25

department: Department of Gene Vectors

institute: Helmholtz Zentrum München, German Research Center for Environmental Health

summary:
Notch1-IC, Notch2-IC or EBNA2 have been induced in a conditionally immortalized human B cell line (EREB2-5) in order to identify similar and unique target genes in B cells. CAT was used as a control.

Keywords: time course

overall_design: RNA was isolated at different time points after induction of Notch1-IC, Notch2-IC or EBNA2 in EREB2-5 cells. Three independent experiments were performed (except Notch1-IC at 3day).

first: Hella

last: Kohlhof


title: Highly Efficient generation of Human Hepatic Cells from Induced Pluripotent Stem Cells.

city: Charleston

first: Stephen

middle: A

last: Duncan

state: SC

address: 173 Ashley Avenue, BSB 6th Floor, Room 657A

department: Regenerative Medicine and Cell Biology

institute: Medical University South Carolina

summary: Reprogrammed somatic cells offer a valuable source of pluripotent cells that have the potential to differentiate into many cells types and provide a new tool for regenerative medicine. In the present study we differentiated induced pluripotent stem cells (iPS cells) into hepatic cells. We first showed that mouse iPS cells could from a complete liver in mouse embryo (E14.5) including hepatocytes, endothelial cells, sinusoidal cells and resident macrophages. We then designed a highly efficient hepatocyte differentiation protocol using defined factors on human embryonic stem cells (ES cells). This protocol was found to generate more than 80% albumin expressing cells that show hepatic functions and express most of liver genes as shown by microarray analyses. Similar results were obtained when human iPS cells were induced to differentiate following the same procedure.

overall_design:
Total RNA was harvested from the following sources and used for Affymetrix array analysis following manufacturer defined protocols:
1) human foreskin fibroblasts, ATCC cell line CRL2097, 3 independent cultures
2) induced pluripotent stem (iPS) cells derived from CRL2097, 3 independent undifferentiated cultures
3) induced pluripotent stem (iPS) cells derived from CRL2097, 3 independent  cultures harvested at day 20 (d20) of a hepatic differentiation protocol
4) WAO9 human embryonic stem cells, 3 independent undifferentiated cultures
5) WAO9 human embryonic stem cells, 3 independent cultures harvested at day 20 (d20) of a hepatic differentiation protocol


title: Response of bronchial epithelial cells to low doses of cigarette smoke condensate

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

overall_design: Refer to individual Series


title: The presence of RNA polymerase II, active or stalled, predicts epigenetic fate of promoter CpG islands

status: Public on Jul 31 2009

city: Shinagawa-ku

first: Hideyuki

last: Takeshima

state: Tokyo

department: Institute for Advanced Life Sciences

institute: Hoshi University

type: Methylation profiling by genome tiling array

summary: Instructive mechanisms are present for induction of DNA methylation, as shown by methylation of specific CpG islands (CGIs) by specific inducers and in specific cancers. However, instructive factors involved are poorly understood, except for involvement of low transcription and trimethylation of histone H3 lysine 27 (H3K27me3). Here, we used methylated DNA immunoprecipitation (MeDIP) combined with a CGI oligonucleotide microarray analysis, and identified 5510 and 521 genes with promoter CGIs resistant and susceptible, respectively, to DNA methylation in prostate cancer cell lines. Expression analysis revealed that the susceptible genes had low transcription in a normal prostatic epithelial cell line. Chromatin immunoprecipitation with microarray hybridization (CHiP-chip) analysis of RNA polymerase II (Pol II) and histone modifications showed that, even among the genes with low transcription, the presence of Pol II was associated with marked resistance to DNA methylation (OR = 0.22; 95% CI = 0.12-0.38), and H3K27me3 was associated with increased susceptibility (OR = 11.20; 95% CI = 7.14-17.55). The same was true in normal human mammary epithelial cells for 5430 and 733 genes resistant and susceptible, respectively, to DNA methylation in breast cancer cell lines. These results showed that the presence of Pol II, active or stalled, and H3K27me3 can predict the epigenetic fate of promoter CGIs independently of transcription levels.

overall_design: To analyze DNA methylation status in normal and cancer cells, MeDIP-CGI oligonucleotide microarray analysis was performed.  To analyze expression and histone modification status in normal cells, GeneChip analysis and ChIP-oligonucleotide microarray analysis were performed.


title: Gene Expression Profiles of CD21low B cells in Common Variable Immunodeficiency

status: Public on Jul 30 2009

city: Freiburg i. Breisgau

first: Mirzokhid

last: Rakhmanov

state: BW

address: Breisacher Str. 66

department: Centre of Chronic Immunodeficiency

institute: University Medical Center Freiburg

summary:
The homeostasis of circulating B cell subsets in the peripheral blood of healthy adults is well regulated, but in disease it can be severely disturbed. Thus, a subgroup of patients with common variable immunodeficiency (CVID) presents with an extraordinary expansion of an unusual B cell population characterized by the low expression of CD21. Since these circulating atypical B cells in the blood of CVID patients could not be assigned to any certain B cell differentiation stage in the periphery, they were designated as CD21low B cells. Although, CD21low B cells are polyclonal and unmutated IgM+IgD+ B cells like naive B cells in the peripheral blood, they reveal several distinct phenotypic and functional features.
In order to uncover the the global programme of gene expression underlying these differences and changes in CD21low B cells we have performed microarray hybridization experiments and identified differentially expressed genes which define a distinct gene expression profile in CD21low B cells compared to naive B cells.

overall_design: Leucocytes were separated by Ficoll gradient from peripheral blood of healthy donors (HD) and CVID patients. CD19+CD27-CD38+CD21+ naive B cells of HD as well as naive and CD19hiCD27-CD38lowCD21low (CD21low) B cells of CVID patients were sorted using a MoFlow cell sorter. RNA was extracted from sorted ex vivo B cell subpopulations and hybridized on Affymetrix microarrays. Gene expression profiles were compared between CD21low B cells and naive B cells of CVID patients as well as HD naive B cells.

first: Klaus

last: Warnatz


title: Analysis of adrenocortical tumors identify IGF2 and Ki-67 as useful in differentiating carcinomas from adenomas

status: Public on Jul 29 2009

city: St Leonards, Sydney

first: Patsy

last: Soon

state: NSW

address: Royal North Shore Hospital

department: Cancer Genetics

country: Australia

institute: Kolling Institute of Medical Research

summary:
Purpose: The management of adrenocortical tumors (ACTs) is complex, compounded by the difficulty in discriminating benign from malignant tumors using conventional histology.  The Weiss score is the current most widely used system for ACT diagnosis but it has limitations, particularly with ACTs with a score of 3.  The am of this study was to identify molecular markers whose expression can discriminate adrenocortical carcinomas (ACCs) from adrenocortical adenomas (ACAs) by microarray gene expression profiling and to determine their clinical applicability by using immunohistochemistry (IHC).  Experimental design: Microarray gene expression profiling was used to identify 7 molecular markers which were significantly differentially expressed between ACCs and ACAs.  These results were confirmed with quantitative PCR for all 7 genes and IHC for 3 protein.  Results: Microarray gene expression profiling was able to accurately categorize ACTs into ACCs and ACAs.  All 7 genes were strong discriminators of ACCs from ACAs on qPCR.  IHC with IGF2, MAD2L1, CCNB1 and Ki-67, but not ACADVL or ALOX15B, had high diagnostic accuracy in differentiating ACCs from ACAs.  The best results however were obtained with a combination of IGF2 and Ki-67 with 96% sensitivity and 100% specificity in diagnosing ACCs.  Conclusion: Microarray gene expression profiling accurately differentiates ACCs from ACAs.  The combination of IGF2 and Ki-67 IHC is also highly accurate in distinguishing between the 2 groups and is particularly helpful in ACTs with Weiss score of 3.

Keywords: Adrenocortical carcinoma, adrenocortical adenoma, differential gene expression, immunohistochemistry, qPCR

overall_design: 12 ACCs, 16 ACAs and 6 normal adrenal cortices are included in this study.  The Affymetrix HGU133plus2.0 genechip was used.

first: Anthony

last: Gill

first: Diana

middle: E

last: Benn

first: Adele

last: Clarkson

first: Bruce

middle: G

last: Robinson

first: Kerrie

middle: L

last: McDonal

first: Stan

middle: B

last: Sidhu


title: BCOR-regulated genes in human oculo-facio-cardio-dental syndrome

status: Public on Jul 23 2009

first: Cun-Yu

last: Wang

address: 10833 Le Conte

institute: UCLA

summary:
Oculo-facio-cardio-dental syndrome (OFCD) is a rare genetic disorder characterized by teeth with extremely long roots (radiculomegaly), and craniofacial, eye and cardiac abnormalities. The mutation of the transcriptional co-repressor BCOR has been identified as being responsible for oculo-facio-cardio-dental (OFCD) syndrome.
Mesenchymal stem cells (MSCs) is isolated from the root apical papilla of an OFCD patient. Gene expression profiling is performed and compared between mutant MSCs and wild type MSCs.

overall_design: Total RNA were extracted from normal MSCs (MSCWT) and mutant MSCs (MSCO).

first: Zhipeng

last: Fan


title: H1299 EGF and Iressa stimulation

city: Yokohama

first: Mariko

last: Okada

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

institute: RIKEN RCAI

summary: Controlled activation of epidermal growth factor receptor (EGFR) is systematically guaranteed at the molecular level, however aberrant activation of EGFR is frequently found in cancer. Transcription induced by EGFR activation often involves coordinated expression of genes that positively and negatively regulate the original signaling pathway, therefore alterations in EGFR kinase activity may reflect changes in gene expression associated with the pathway. In this study, we investigated transcriptional changes following EGF stimulation with or without the EGFR kinase inhibitor Iressa in H1299 human non-small-cell lung cancer cells (parental H1299, H1299 cells which overexpress wild-type: EGFR-WT and mutant EGFR: L858R). Our results clearly showed differences in transcriptional activity in the absence or presence of EGFR kinase activity, and genes sharing the same molecular functions showed distinct expression dynamics. The results showed particular enrichment of EGFR/ErbB signaling-related genes in a differentially expressed gene set, and significant protein expression of MIG6/RALT(ERRFI1), an EGFR negative regulator, was confirmed in L858R. High MIG6 protein expression was correlated with basal EGFR phosphorylation and inversely correlated with EGF-induced ERK phosphorylation levels. Investigation of NCI-60 cell lines showed that ERRFI1 expression was correlated with EGFR expression regardless of tissue type. These results suggest that cells accumulate MIG6 as an inherent negative regulator to suppress excess EGFR activity when basal EGFR kinase activity is considerably high. Taken together, an EGFR mutation can cause transcriptional changes to accommodate the activation potency of the original signaling pathway at the cellular level.

overall_design: H1299 human non-small cell lung cancer cells were stimulated by the growth hormone (epidermal growth factor (EGF)) or EGFR kinase inhibitor (Iressa). Control was set as non-treated cells.

first: Takeshi

last: Nagashima

last: Hatakeyama


title: Global gene expression profiling of human pleural mesotheliomas

city: Torino

first: Raffaele

last: Calogero

state: To

address: Via Nizza 52

department: Molecular Biotechnology Center

country: Italy

institute: University of Torino

summary:
The goal of our study was to molecularly dissect mesothelioma tumor pathways by mean of microarray technologies in order to identify new tumor biomarkers, that could be used as early diagnostic markers and possibly as specific molecular therapeutic targets. We performed Affymetrix U133A plus 2.0 microarray analysis comparing 9 human pleural mesotheliomas with 4 normal pleural specimen. Stringent statistical feature selection detected a set of differentially expressed genes that were further evaluated to identify potential biomarkers to be used in early diagnostics. Selected genes were confirmed by RT-PCR. As reported by other mesothelioma profiling studies, most of genes are involved in G2/M transition. Our list contains several genes previously described as prognostic classifier. Furthermore, we found novel genes never associated before to mesothelioma and could be involved in tumor progression. Notable, the identification of MMP-14, a member of matrix metalloproteinase family. This molecule has been described as a new disease marker and could be used as biomarker also for mesothelioma early diagnosis and prognosis and that can be viewed as new and effective therapeutic target to test.
Accepted for pubblication in PLoS ONE

overall_design: 9 mesothelioma mixed type tumors versus the pleura of 4 normal donors were compared

first: Stefania

last: Crispi

first: Pasquale

last: Mellone

first: Bruno

last: Vincenzi

first: Gennaro

last: Citro

first: Mario

last: Santini

first: Simona

last: Menegozzo

first: Alfonso

last: Baldi

first: Massimo


title: Expression data from LNCaP and abl cells

status: Public on Jul 25 2009

city: Boston

first: Arjun

middle: Kumar

last: Manrai

address: 44 Binney Street, Mail Stop LW225

department: Department of Biostatistics and Computational Biology

institute: Dana Farber Cancer Institute

summary:
Androgen receptor (AR) is a ligand-dependent transcription factor that plays a key role in the onset and progression of prostate cancer. We investigated  AR-induced gene expression in prostate cancer cells LNCaP and abl by transfecting siAR / siControl or treating cells  with androgen (DHT) over a time course.
Keywords: siRNA transfection and androgen stimulation time course

overall_design: We hybridized RNA to Affymetrix human genome U133 plus 2.0 arrays.

first: Qianben

first: Myles

last: Brown


title: MLL rearrangements in pediatric ALL and AML: MLL specific and lineage specific signatures

status: Public on Jul 22 2009

city: Padova

first: Andrea

last: Zangrando

address: Via Giustiniani, 3

department: Woman and Child's Health

institute: University of Padova

summary:
Gene expression analysis identified a MLL translocation-specific signature of differentially expressed genes discriminating ALL and AML with and without MLL rearrangements.
Gene expression signatures of acute lymphoblastic and myeloblastic leukemia samples with and without MLL rearrangements were analyzed using paired supervised analyses.

overall_design: ALL and AML patients with and without MLL rearrangements have been studied using paired class comparison (SAM) and class prediction (PAM) analyses.

first: Geertruy

last: te Kronnie


title: Gene profiling of heart atria in PI3K and Mst1 mouse models

city: Sydney

first: Ruby

middle: CY

last: Lin

address: 176 Hawkesbury Road, Westmead

institute: The Westmead Institute for Medical Research

summary:
We used microarrays to detail genome-wide gene expression underlying cardiac myocyte pathologies and identified candidate genes and specific pathways affecting cardiac myopathies

Keywords: transgenics, cardiac disease analysis

overall_design: Mouse heart atria were dissected for RNA extraction and hybridisation on Affymetrix microarrays. We sought to profile gene expression from the transgenic mouse models for non-transgenics (Ntg), dnPI3K (dominant negative for phosphoinositide 3-kinase), Mst1 (mammalian sterile 20-like kinase 1) and dnPI3K-Mst1 (double mutant) mouse models with a combined background of C57BL/6/FVB/N inbred strains. Pathological insults on cardiac myocytes are investigated

first: Julie

middle: R

last: McMullen

first: RubyCY


title: The IKK2/NF-kB-pathway suppresses MYC-induced lymphomagenesis

city: Ulm

first: Kay

last: Klapproth

address: Albert-Einstein-Allee 11

department: Institute of Physiological Chemistry

institute: Ulm University

summary:
Abstract.
Deregulated c-MYC is found in a variety of cancers where it promotes proliferation as well as apoptosis. In many hematological malignancies enhanced NF-kB exerts prosurvival functions. Here we investigated the role of NF-kB in mouse and human c-MYC-transformed lymphomas. The NF-kB-pathway is extinguished in murine lymphoma cells and extrinsic stimuli typically inducing NF-kB activity fail to activate this pathway. Genetic activation of the NF-kB pathway induces apoptosis in these cells, while inhibition of NF-kB by an IkBa superrepressor provides a selective advantage in vivo. Furthermore, in human Burkitt´s lymphoma cells we find that NF-kB activation induces apoptosis. NF-kB upregulates Fas and predisposes to Fas-induced cell death, which is caspase 8 mediated and can be prevented by CFLAR overexpression. We conclude that c-MYC overexpression sensitizes cells to NF-kB-induced apoptosis and persistent inacvtivity of NF-kB signaling is a prerequisite for MYC-mediated tumorigenesis. We could also show that low immunogenicity and Fas insensitivity of MYC driven lymphoma cells is reversed by activation of NF-kB. Our observations provide a molecular explanation for the described absence of the NF-kB signaling in Burkitt´s lymphoma and question the applicability of NF-kB inhibitors as candidates for treatment of this cancer.

overall_design: Ramos cells were transfected in triplicates with pRTS-GFP (A+,C+,D+) or pRTS-CA-IKK2 (E+,F+,H+), selected with hygromycin and transgene expression was induced with doxycycline (0,5mg/ml) for 48hrs. RNA was isolated with RNeasy mini kit (Qiagen, Venlo, Netherlands) and Gene expression profiling (GEP) was performed using Affymetrix Human Genome U133 Plus 2.0 Array (Affymetrix, Santa Clara, CA, USA). 2 µg of total RNA were labeled using the GeneChip® One-Cycle Target Labeling assay kit (Affymetrix). After hybridization arrays were stained and washed in a FS 450 Fluidics station (Affymetrix) before imaging on an Affymetrix GeneChip (3000) scanner. Raw data were generated using the GCOS 1.4 software (Affymetrix). Probe level data were obtained using the Robust Multichip Average (RMA) normalization algorithm and CEL files were loaded into Genesifter (GeneSifter.Net, VizX Laboratories, Seattle, WA, USA). Genes were identified as differentially expressed among the two classes if a two-sample T-test revealed a nominal significance level of 0.05 and the ratio between the two classes was at least 2 fold. Calculation of false discovery rate was done according to the method of Benjamini and Hochberg. Biological significance was determined using Gene Ontolgy reports.

first: Sandrine

last: Sander

first: Dragan

last: Marinkovic

first: Bernd

last: Baumann

first: Thomas

last: Wirth


title: Global effect of RpoS on gene expression in pathogenic Escherichia coli O157:H7 strain EDL933

status: Public on Aug 03 2009

city: Hamilton

first: Herb

last: Schellhorn

state: ON

address: 1280 Main St. W.,

department: Biology

country: Canada

institute: McMaster University

summary:
RpoS is a conserved stress regulator that plays a critical role in survival under stress conditions in Escherichia coli and other γ-proteobacteria. RpoS is also involved in virulence of many pathogens including Salmonella and Vibrio species. Though well characterized in non-pathogenic E. coli K12 strains, the effect of RpoS on transcriptome expression has not been examined in pathogenic isolates. E. coli O157:H7 is a serious human enteropathogen, possessing a genome 20% larger than that of E. coli K12, and many of the additional genes are required for virulence. The genomic difference may result in substantial changes in RpoS-regulated gene expression. To test this, we compared the transcriptional profile of wild type and rpoS mutants of the E. coli O157:H7 EDL933 type strain. The rpoS mutation had a pronounced effect on gene expression in stationary phase, and more than 1,000 genes were differentially expressed (two-fold, p<0.05). By contrast, we found 11 genes expressed differently in exponential phase. Western blot analysis revealed that, as expected, RpoS level was low in exponential phase and substantially increased in stationary phase. The defect in rpoS resulted in impaired expression of genes responsible for stress response (e.g., gadA, katE and osmY), arginine degradation (astCADBE), putrescine degradation (puuABCD), fatty acid oxidation (fadBA and fadE), and virulence (ler, espI and cesF). For EDL933-specific genes on O-islands, we found 50 genes expressed higher in wild type EDL933 and 49 genes expressed higher in the rpoS mutants. The protein levels of Tir and EspA, two LEE-encoded virulence factors, were elevated in the rpoS mutants under LEE induction conditions. Our results show that RpoS has a profound effect on global gene expression in the pathogenic strain O157:H7 EDL933, and the identified RpoS regulon, including many EDL933-specific genes, differs substantially from that of laboratory K12 strains.
In this study, we characterized the RpoS regulon of E. coli O157:H7 strain EDL933 using microarray analysis.

overall_design: A precise rpoS deletion mutant of EDL933 was constructed and employed in this study. EDL933 wild type and rpoS mutants were inoculated in triplicate into LB media at a starting OD of 0.0001 and grown aerobically at 37C. Cultures were harvested at OD600 = 0.3 in exponential phase and OD600=1.5 in stationary phase. For RNA extraction, cultures were mixed directly with a boiling lysis buffer containing SDS and EDTA followed by acidic hot phenol to minimize RNA degradation. RNA samples were hybridized to Affymetrix E. coli Genome 2.0 Array according to Affymetrix's standard protocols.

first: Tao

last: Dong


title: Smoking-dependent Reprogramming of Alveolar Macrophage Polarization: Implication for Pathogenesis of COPD

status: Public on Aug 10 2009

city: New York

first: Yael

last: Strulovici-Barel

state: NY

address: 1300 York Avenue

department: Department of Genetic Medicine

institute: Weill Cornell Medical College

summary: Background: When exposed to specific stimuli, macrophages exhibit distinct activation programs, M1 and M2 polarization, that define macrophage function as inflammatory/immune effectors or anti-inflammatory/tissue remodeling cells, respectively. Due to their position on the lung epithelial surface, alveolar macrophages (AM) directly interact with environmental stimuli such as cigarette smoke, the major risk factor for the development of chronic obstructive pulmonary disease (COPD). Based on the current paradigm that, in response to smoking, AM contribute to both inflammatory and tissue remodeling processes in the lung relevant to the pathogenesis of COPD, we hypothesized that chronic exposure to cigarette smoking activates both the M1 and M2 polarization programs in AM. Methods and Findings: To assess this hypothesis, global transcriptional profiling with TaqMan confirmation and flow cytometry analysis was carried out on AM obtained by bronchoalveolar lavage of 24 healthy nonsmokers, 34 healthy smokers and 12 smokers with COPD to assess the expression of 41 M1 genes and 32 M2 genes in each group. Contrary to our expectations, while there was up-regulation of some genes typical for M2-related phenotypes, AM of healthy smokers exhibited substantial suppression of M1-related inflammatory/immune genes. These M1- and M2-related changes progressed with the development of smoking-induced lung disease, with AM of smokers with COPD exhibiting further down-regulation of M1-related genes accompanied with further up-regulation of some M2-related genes. Conclusion: The data demonstrates that the modifications of the AM transcriptome associated with smoking result in a unique phenotype characterized by reprogramming of AM towards M1-deactivated partially M2-polarized macrophages and suggests that, while AM likely contribute to smoking-induced tissue remodeling, the role of AM in the early pathogenesis of smoking-induced COPD in humans is not inflammatory. This concept is a departure from the conventional concept that AM-mediated inflammation participates in the early derangements of the lung induced by smoking, and suggests a novel paradigm for conceptualizing COPD and developing new approaches to prevent the development of smoking-induced lung disease.

overall_design: Comparison of gene expression in alveolar macrophages of normal non-smokers and normal smokers and smokers with COPD

first: Renat

last: Shaykhiev

first: Anja

last: Krause

first: Jacqueline

last: Salit

first: Ben-Gary

last: Harvey

first: Timothy

middle: P

last: O’Connor

first: Ronald

last: Crystal


title: Transcriptome of BAHD1 knock-down HEK293 cells with siRNA

status: Public on Jul 16 2009

city: Jouy-en-Josas

first: Helene

last: Bierne

address: Domaine de Vilvert, bat 442, INRA Micalis

department: Micalis Institute

institute: INRAE - University PARIS SACLAY

summary:
Gene silencing via heterochromatin formation plays a major role in cell differentiation and maintenance of homeostasis. Here, we report the identification and characterization of a novel heterochromatinization factor in vertebrates, Bromo Adjacent Homology Domain-containing protein 1 (BAHD1). BAHD1 interacts with HP1, MBD1, HDAC5 and with several transcription factors. Through electron and immunofluorescence microscopy studies, we show that BAHD1 overexpression directs HP1 to specific nuclear sites and promotes formation of large heterochromatic domains, which lack acetyl histone H3 and are enriched in H3 trimethylated at lysine 27. Furthermore, ectopically expressed BAHD1 colocalizes with the heterochromatic X inactive chromosome. As highlighted by whole genome microarray analysis of BAHD1 knock down cells, BAHD1 represses several proliferation and survival genes and in particular, the insulin-like growth factor II gene (IGF2). BAHD1 specifically binds the CpG-rich P3 promoter of IGF2. This region contains DNA binding sequences for the transcription factor SP1, with which BAHD1 co-immunoprecipitates. Collectively, these findings provide evidence that BAHD1 acts as a silencer by recruiting proteins that coordinate heterochromatin assembly at specific sites in the genome.
We used microarrays to identify BAHD1 gene targets. We compared the transcriptome profile of BAHD1 depleted cells with siRNA to that of cells treated with control siRNA.

overall_design: Six samples were analysed, including three biological replicates of control cells and three biological replicates of BAHD1 KD cells.

first: Hélène

first: Béatrice

last: Regnault

first: Pascale

last: Cossart


title: Meta analysis of gene expression in human islets after in vitro expansion.

city: Seattle

first: Burak

last: Kutlu

state: WA

address: 1441 North 34th Street

institute: Institute for Systems Biology

summary: Pancreatic islet transplantation as a cure for type 1 diabetes (T1D) cannot be scaled up due to a scarcity of human pancreas donors. In vitro expansion of beta cells from mature human pancreatic islets provides an alternative source of insulin-producing cells. The exact nature of the expanded cells produced by diverse expansion protocols, and their potential for differentiation into functional beta cells, remain elusive. We performed a large-scale meta-analysis of gene expression in human pancreatic islet cells, which were processed using three different previously described protocols for expansion and attempted re-differentiation. All three expansion protocols induced dramatic changes in the expression profiles of pancreatic islets; many of these changes are shared among the three protocols. Attempts at re-differentiation of expanded cells induce a limited number of gene expression changes. Nevertheless, these fail to restore a pancreatic islet-like gene expression pattern. Comparison with a collection of public microarray datasets confirmed that expanded cells are highly comparable to mesenchymal stem cells. Genes induced in expanded cells are also enriched for targets of transcription factors important for pluripotency induction. The present data increases our understanding of the active pathways in expanded and re-differentiated islets. Knowledge of the mesenchymal stem cell potential may help development of drug therapeutics to restore beta cell mass in T1D patients.

overall_design: In this study, we have tested three different protocols to expand human pancreatic islets in monolayer and after attempted maneuvers to re-differentiate the expanded cells back to islets. We have characterized the resulting cells in detail by performing microarray analyses with fresh pancreatic islets, expanded islet cells and re-differentiated cells. Genes modified by either of three protocols have 70 to 80% overlap with the genes changed by the other two protocols. Although there are promising changes in the right direction, none of the three protocols could achieve a return to a functional islet state. The expanded cells highly resemble Mesenchymal Stem Cells (MSC), and similar gene regulatory networks seem to be active in both cell types. On the other hand, the expanded islet cells are different from MSC in that they seem to retain activity of some islet gene modules. The current results highlight the importance of designing new strategies that take into account the MSC potential of expanded cells.


title: Networking of differentially expressed genes in human Saos-2 osteosarcoma cells resistant to methotrexate

status: Public on Sep 08 2009

city: Barcelona

first: Carlos

middle: J

last: Ciudad

state: Barcelona

address: Av. Juan XXIII-27

department: Biochemistry and Molecular Biology

country: Spain

institute: University of Barcelona

summary:
A summary of the work associated to these microarrays is the following:

The need for an integrated view of all data obtained from high-throughput technologies gave rise to network analyses. These are especially useful to rationalize phenomena in terms of how external perturbations propagate through the expression of genes. To address this issue in the case of drug resistance, we constructed Biological Association Networks of genes differentially expressed in cell lines resistant to methotrexate (MTX). Seven cell lines representative of different types of cancer including colon cancer (HT29 and Caco2), breast cancer (MCF7 and MDA-MB-468), pancreatic cancer (MIA PaCa-2), erythroblastic leukemia (K562) and osteosarcoma (Saos-2), were used. The differential expression pattern between sensitive and MTX-resistant cells was determined by microarrays covering the whole human genome and analyzed with the GeneSpring GX software package, v.7.3.1. Genes deregulated in common in the two colon cancer cell lines studied, were subject of Biological Association Networks construction. Dikkopf homolog-1 (DKK1) was a clear node of this network, and functional validations of this target using a siRNA showed a chemosensitization toward MTX. Members of the UDP-glucuronosyltransferase 1A (UGT1A) family formed a network of differentially expressed genes in the two breast cancer cell lines studied. siRNA treatment against UGT1A showed also an increase in MTX sensitivity. Eukaryotic translation elongation factor 1 alpha 1 (EEF1A1) was a gene overexpressed in common among the pancreatic cancer, leukemia and osteosarcoma cell lines, and siRNA treatment against EEF1A1 produced a chemosensitization toward MTX. Biological Association Networks identified DKK1, UGT1As and EEF1A1 as important gene nodes in MTX-resistance. Treatments using iRNA technology against these three genes show chemosensitization toward MTX.

overall_design: Two cell lines are compared, which are Saos-2 osteosarcoma cells sensitive to methotrexate and Saos-2 cells resistant to 10e-6M methotrexate. Six samples are provided which correspond to triplicates of each cell line. The samples provided were  analyzed using the specific software GeneSpring GX.

first: Elisabet

last: Selga

first: Carlota

last: Oleaga

last: de Almagro

first: Véronique

last: Noé


title: Expression profiling of Bmal mutant dorsal skin at telogen of hair follicle cycling

status: Public on Jul 17 2009

city: Irvine

first: Kevin

middle: K

last: Lin

state: CA

address: 250 Sprague Hall

department: Biological Chemistry & Medicine

institute: University of California, Irvine

summary: Hair follicles undergo recurrent cycling of controlled growth (anagen), regression (catagen), and relative quiescence (telogen) with a defined periodicity. Taking a genomics approach to study gene expression during synchronized mouse hair follicle cycling, we discovered that, in addition to circadian fluctuation, CLOCK-regulated genes are also modulated in phase with the hair growth cycle. During telogen and early anagen, circadian clock genes are prominently expressed in the secondary hair germ, which contains precursor cells for the growing follicle. Analysis of Clock and Bmal1 mutant mice reveals a delay in anagen progression, and the secondary hair germ cells show decreased levels of phosphorylated Rb and lack mitotic cells, suggesting that circadian clock genes regulate anagen progression via their effect on the cell cycle. Consistent with a block at the G1 phase of the cell cycle, we show a significant upregulation of p21 in Bmal1 mutant skin. While circadian clock mechanisms have been implicated in a variety of diurnal biological processes, our findings indicate that circadian clock genes may be utilized to modulate the progression of non-diurnal cyclic processes.

overall_design: To gain molecular understanding of the the hair cycle delay in Bmal mutant mice, we profiled the dorsal skin of Bmal knockout (-/-) and their heterozygous (+/-) littermates at P22. At P22, the skin samples are comparable because all the samples are in telogen just prior to the hair cycle delay was observed. Histological sections were used to classify each sample into specific stage of the hair growth cycle based on established morphological guidelines. RNA from each mouse dorsal skin were separately hybridized to an Affymetrix Mouse Gene 1.0 ST array.

first: Vivek

last: Kumar

first: Mikhail

last: Geyfman

first: Darya

last: Chudova

first: Alexander

last: Ihler

first: Padhraic

last: Smyth

first: Ralf

last: Paus

first: Joseph

middle: S

last: Takahashi

first: Bogi

last: Andersen


title: Effects of chronic exposure of human bronchial epithelial cells to low doses of cigarette smoke condensate

city: Zurich

first: Hubert

last: Rehrauer

address: Winterthurerstr. 190

institute: ETH Zurich / University of Zurich

summary:
BEAS-2B cells have been treated with low doses (20µg/ml) of CSC for 4 months. As negative control BEAS-2B cells were treated with DMSO (the CSC solvent). Non-treated cells were cultivated in parallel.

Keywords: time course, stress response

overall_design: After each month total RNA was extracted from three replicates of CSC, DMSO and non-treated BEAS-2B cells and hybridized to Affymetrix GeneChips.

first: Emilija

last: Veljkovic

first: Anne

last: Anstett

first: Mirco

last: Menigatti

first: Alex

last: Soltermann

first: Josef

last: Jiricny

first: Wanjiang

last: Han


title: Comparison of the migration profile of MSCs

status: Public on Jul 14 2009

city: Singapore

first: Ivy

last: Ho

address: 11 Hospital Drive

country: Singapore

institute: National Cancer Center

summary: To compare the gene expression profile of MSCs harvested from bone marrow in the context of cell migration.

overall_design: A total of 4 MSCs isolated from different donors were used.  Duplicate samples were used for each hybridization.  The experiment was repeated twice.  CEL files obtained were imported into Partek Genomics suite for analysis.  The final processed data were obtained from selecting for genes that have at least 2-fold difference in expression.  The gene list was further narrow down to the keywords cell adhesion, chemokine, cytokine, and metalloproteinases.

first: Paula

last: Lam


title: Biomarkers for Early and Late Stage Chronic Allograft Nephropathy by Genomic Profiling of Peripheral Blood

status: Public on Jul 13 2009

city: La Jolla

first: Sunil

last: Kurian

address: 10550 N Torrey Pines Road

department: Molecular and Experimental Medicine

institute: The Scripps Research Institute

summary:
Despite significant improvements in life expectancy of kidney transplant patients due to advances in surgery and immunosuppression, Chronic Allograft Nephropathy (CAN) remains a daunting problem. A complex network of cellular mechanisms in both graft and peripheral immune compartments complicates the non-invasive diagnosis of CAN, which still requires biopsy histology. This is compounded by non-immunological factors contributing to graft injury.  There is a pressing need to identify and validate minimally invasive biomarkers for CAN to serve as early predictors of graft loss and as metrics for managing long-term immunosuppression.
This study attempts to identify sets of unique transcript biomarkers with high predictive accuracy for both mild and moderate/severe CAN. These biomarkers are the necessary first step to a genomic classification of CAN based on peripheral blood and the targets for a prospective, serial-monitoring clinical study.

overall_design: We used DNA microarrays and bioinformatics to identify candidate genomic markers of mild and moderate/severe CAN in peripheral blood of two distinct cohorts (n=42 and n=35, respectively) of kidney transplant patients with biopsy-documented histology.

middle: M


title: Effects of demethylation on lung cancer cell lines

summary:
The goal of this study is to identify those genes that are commonly silenced by DNA-methylation in lung cancer cell lines.

Keywords: cell line comparison, stress response

overall_design: Ten lung cancer cell lines (Calu 1, Hotz, U1752, Calu 6, ZL25, Calu3, SW2, A549, H125, and H 460) were analyzed before and after the treatment with 5-aza-2’deoxycitidine and trychostatin A (5AzaC/TSA).


title: Antitumor Activity and Molecular Effects of the Novel Hsp 90 Inhibitor, IPI-504, in Pancreatic Cancer

status: Public on Jul 15 2009

city: Baltimore

first: Aik Choon

last: Tan

state: MD

address: 1650 Orleans Street, RM 1M86

department: The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins

institute: Johns Hopkins University

summary:
Tumors from pancreatic cancer specimens obtained at surgery were used for efficacy testing and biologic analysis.  These tumors were s.c. explanted in xenograft models for subsequent treatment experiments.
This study aimed to assess the antitumor activity of the Hsp90 inhibitor, IPI-504, in pancreatic cancer and to determine the biological effects of the agent.

Keywords: Pharmacogenetics

overall_design: Five tumor samples were profiled in treated and untreated with IPI504.  Untreated samples were profiled in duplicate.  The overall design is to identify differentially expressed genes in IPI504 treated versus untreated.

first: Aik

middle: C

first: Manuel

last: Hidalgo


title: Coordinate Control of Nrf2 Mediated Genes in the Human Small Airway Epithelium Highly Responsive to Smoking

first: Yael

last: Strulovici-Barel

summary: Nuclear factor erythroid 2-related factor 2 (NFE2L2, Nrf2) is an oxidant responsive transcription factor known to induce phase 2 detoxifying and antioxidant genes to protect cells from oxidative stress. Cigarette smoke, with its large oxidant content, is a major stressor to the small airway epithelium, the cells of which are vulnerable to oxidant damage and consequent malignant transformation. In this study, we assessed the role of cigarette smoke in activation of Nrf2 in the human small airway epithelium in vivo. Fiberoptic bronchoscopy was used to sample a pure population of small airway epithelium in 38 healthy nonsmokers and 45 healthy smokers, and gene expression was assessed using Affymetrix HG-U133 Plus 2.0 microarrays. Compared to that of  healthy nonsmokers, Nrf2 protein was significantly activated in the small airway epithelium of healthy normal smokers and localized in the nucleus (p<0.05). Of the human homologs of 201 known murine Nrf2-mediated genes, 13 highly smoking-responsive genes were identified (p<10-4, all comparisons smokers to nonsmokers). Using a “Nrf2-index” to quantify the extent of expression in the small airway epithelium of these 13 known Nrf2 genes, variability in the level of expression was observed among the 45 healthy smokers, but the variability was coordinately modulated among the 13 genes, an observation confirmed by TaqMan quantitative PCR. This variability in the coordinate level of expression of the 13 Nrf2-mediated genes was independent of the smoking history. Based on these observations, the “Nrf2 index” was used to evaluate whether other genes modulated by smoking in the small airway epithelium were also coordinately up- or down- modulated among the 45 healthy smokers. Two genes, pirin (PIR) and UDP glucuronosyltransferase 1 family polypeptide A4 (UGT1A4), not previously known to be modulated by Nrf2 were identified as being coordinately modulated among the 45 smokers. Both genes contain several functional antioxidant response elements in the promoter region. Using an electrophoretic mobility shift assay, these antioxidant response elements in the promoters of PIR and UGT1A4 responded in vitro to activated Nrf2. These observations are consistent with the concept that Nrf2 plays an important role in regulating cellular defenses against smoking in the highly vulnerable small airway epithelium cell population, and that there is variability among the population in the relative Nrf2 responsiveness to a similar oxidant burden.

overall_design: Affymetrix arrays were used to assess gene expression data in small airway epithelium obtained by fiberoptic bronchoscopy of 38 healthy non-smokers and 45 healthy smokers

first: Ralf-Harto

last: Hübner

first: Jamie

middle: D

last: Schwartz

first: Bishnu

last: De

first: Barbara

last: Ferris

first: Neil

last: Hackett

first: Ronald

last: Crystal


title: Onconase Responsive Genes in Human Mesothelioma Cells: Implications for an RNA Damaging Therapeutic Agent

status: Public on Jul 09 2009

city: Philadelphia

first: JIANMING

last: PEI

state: PA

address: 333 Cottman Ave.

department: Population Science

institute: Fox Chase Cancer Center

summary:
Onconase represents a new class of RNA-damaging drugs. Mechanistically, Onconase is thought to internalize, where it degrades intracellular RNAs such as tRNA and double-stranded RNA, and thereby suppresses protein synthesis.  However, there may be additional or alternative mechanism(s) of action.
Microarray analysis was used to compare gene expression profiles in untreated human malignant mesothelioma (MM) cell lines and cells exposed to 5 ug/ml Onconase for 24 h.

overall_design: In this study, microarray analysis was used to compare gene expression profiles in untreated human malignant mesothelioma (MM) cell lines and cells exposed to 5 ug/ml Onconase for 24 h.

first: Deborah

last: Altomare

first: Susanna

last: Rybak

first: Jianming

last: Pei

first: Jacob

middle: V

last: Maizel

first: Mitchell

last: Cheung

first: Joseph

last: Testa

first: Kuslima

last: Shogen