title: Expression data from normal adrenal gland and aldosterone-producing adenoma

status: Public on Jul 19 2007

city: Augusta

first: Ping

last: Ye

state: GA

address: 1120 15th Street

department: Physiology

institute: Medical College of Georgia

summary:
The source of aldosterone in 30 to 40 % of patients with primary hyperaldosteronism (PA) is unilateral aldosterone-producing adenoma (APA). The mechanisms causing elevated aldosterone production in APA are unknown. Herein, we examined expression of G-protein coupled receptors (GPCR) in APA and demonstrate that compared to normal adrenals there is a general elevation of certain GPCR in many APA and/or ectopic expression of GPCR in others. RNA samples from normal adrenals (n = 5), APAs (n = 10), and cortisol-producing adenomas (CPAs) (n=13) were used on 15 genomic expression arrays, each of which included 223 GPCR transcripts presented in at least one out of 15 of the independent microarrays. The array results were confirmed using real-time RT-PCR (qPCR). Four GPCR transcripts exhibited a statistically significant increase that was greater than 3-fold compared to normal adrenals, suggesting a general increase in expression compared to normal adrenal glands. Four GPCR transcripts exhibited a greater than 15-fold increase of expression in one or more of the APA samples compared to normal adrenals. qPCR analysis confirmed array data and found the receptors with the highest fold increase in APA expression to be luteinizing hormone receptor (LH-R), serotonin receptor 4 (HTR4), gonadotropin-releasing hormone receptor (GnRHR), glutamate receptor metabotropic 3 (GRM3), endothelin receptor type B-like protein (GPR37), and ACTH receptor (MC2R). There are also sporadic increased expressions of these genes in the CPAs. Together, these findings suggest a potential role of altered GPCR expression in many cases of PA and provide candidate GPCR for further study.
Keywords: disease state analysis

overall_design: Normal human adult adrenals were obtained through the Cooperative Human Tissue Network (Philadelphia, PA) and Clontech (Palo Alto, CA). All the normal adrenal samples came from patients that underwent adrenalectomy along with a renalectomy due to renal carcinoma. The adrenal samples acquired from autopsies were obtained no more than six hours after death, and it was confirmed that the causes of death were unrelated to adrenal function. Real-time quantitative RT-PCR was used to check the legitimacy of the normal control adrenal samples. Examination of tissue sections from each adrenal gland also suggested normal histology.  The APA samples were obtained from the Departments of Medicine at UT Southwestern and Division of Endocrinology at the University of Padua. All APA samples were from Conn’s Syndrome patients with significantly elevated circulating aldosterone that was lowered to the normal range after surgical removal of the tumor. Thirty-two adenomas were collected from patients with primary hyperaldosteronism. Twenty-eight of these adenomas had levels of CYP11B2 that were two standard deviations (SD) greater than seen in normal adrenal glands and these samples were used for analyses. It is currently not clear if the adenomas with low CYP11B2 expression are the source of aldosterone in these patients. Other studies suggest that subcapsular micronodules have varying expression of steroidogenic enzymes-some with aldosterone synthase and some without (Shigematsu et al., 2006). Future studies are required to determine if some tumor-bearing adrenals from primary hyperaldosteronsim patients also have small CYP11B2 positive micronodules as the source of aldosterone production.  The cortisol-producing adenoma (CPA) tissues were obtained from Division of Endocrinology at the University of Padua from patients with Cushing’s syndrome. Each of the patients was cured following surgical removal of the adenoma. The use of these tissues was approved by the Institutional Review Boards of the University of Texas Southwestern Medical Center (Dallas, TX), the Medical College of Georgia (Augusta, GA), and University of Padua (Italy). In addition, this study was approved by the ethical committee at the University of Padua, and informed consent was obtained from every patient.  Microarray Analysis  Total RNA isolated from normal adult adrenal glands (n = 5) and APA (n = 10) were used on 15 genomic expression arrays. In brief, RNA was hybridized to an Affymetrix human HG-U133+2 oligonucleotide microarray set containing 54,675 probe sets representing approximately 40,500 independent human genes. The arrays were scanned at high resolution using an Affymetrix GeneChip Scanner 3000 located at Medical College of Georgia Microarray Core Facility (Augusta, GA). Results were analyzed using GeneSpring GX 7.3.1 software (Silicon Genetics, Redwood City, CA) to identify differences in expression of GPCR between normal adult adrenal and APA.  RNA Extraction   The tissue was pulverized in liquid nitrogen to a powder. Total RNA was extracted using TRIzol Reagent (Invitrogen, Carlsbad, CA). The purity and integrity of the RNA were assessed by the Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA) and its quantity was determined by NanoDrop spectrophotometer (NanoDrop Technologies, Wilmington, DE).  cDNA Synthesis  2 µg of total RNA was reverse transcribed using the High Capacity cDNA Archive Kit (Applied Biosystems, Foster City, CA) following manufacturer recommendations and incubated at 25 ºC for 10 min, and 37 ºC for 2 h. The synthesized cDNA was subjected to 1:10 dilution and stored at -20ºC.  Real-Time Quantitative PCR  Primers and probes for the amplification of the selected GPCR sequences were performed using the TaqMan Gene Expression Assays (Applied Biosystems, Foster City, CA) based on published sequences for genes encoding the respective human GPCR. The gene symbols and AB assay numbers are listed in Table 1.  The primer and probe set for human CYP11B2 was designed using Primer Express 3.0 (Applied Biosystems, Foster City, CA) and purchased from IDT (Integrated DNA Technologies, Inc, IA) as published previously (Saner-Amigh et al., 2006). In brief:  Forward: 5’-GGCAGAGGCAGAGATGCTG-3’,   Reverse: 5’- CTTGAGTTAGTGTCTCCACCAGGA-3’,   Probe:  5’-CTGCACCACGTGCTGAAGCACT-3’.  PCR reactions were performed using the ABI 7500 Fast Real-Time PCR System (Applied Biosystems, Foster City, CA) with a total volume of 20 μl per reaction following the reaction parameters recommended by the manufacturer, which includes denaturation at 95ºC for 20 s followed by amplification for 40 cycles (95ºC at 3 s, 60ºC at 30 s, fluorescence measurement). For each GPCR the 20 μl total volume consisted of 10 µl TaqMan Fast Universal PCR Master Mix (2X) (Applied Biosystems, Foster City, CA), 900 nM of each primer, 400 nM of the probe and 5 μl of each first-strand cDNA sample. CYP11B2 reaction mix consisted of 10µl TaqMan PCR Master Mix (2X) (Applied Biosystems, Foster City, CA), 100 nM of primer/probe mix and 5 μl of each first-strand cDNA sample. 18s rRNA was detected and quantified using the TaqMan Ribosomal RNA Control Reagents (Vic Probe) (Applied Biosystems, Foster City, CA). Each reaction included 10 µl of TaqMan PCR Master Mix (2X) (Applied Biosystems, Foster City, CA), 100 nM probe and 50 nM primers. Negative controls contained water instead of first-strand cDNA. Quantitative normalization of cDNA in each tissue-derived sample was performed using expression of 18s rRNA as an internal control. The generated Ct value of each gene was normalized by its respected Ct value of 18s rRNA (∆Ct). Then each gene was further normalized using the average ∆Ct value of the normal adult adrenal (∆∆Ct). The final fold-expression changes were calculated using the equation 2-∆∆Ct (Livak & Schmittgen, 2001).  Statistical Analysis  Data were analyzed and compared to control values (mean of normal adrenal samples) using the Mann-Whitney Rank Sum test with the SigmaStat 3.0 software package (SPSS, Chicago, IL). Results were considered significantly different when p value was less than 0.05.

first: William

last: Rainey


title: Genome-wide molecular profiles of HCV-induced dysplasia and hepatocellular carcinoma

status: Public on Jun 21 2007

city: New York

first: Elisa

last: Wurmbach

state: NY

address: 1 Gustave L. Levy Place

department: Hematology/Oncology

country: USA

institute: Mount Sinai School of Medicine

summary:
Gene expression profiles of 75 tissue samples were analyzed representing the stepwise carcinogenic process from pre-neoplastic lesions (cirrhosis and dysplasia) to HCC, including four neoplastic stages (very early HCC to metastatic tumors) from patients with HCV infection. Gene signatures that accurately reflect the pathological progression of disease at each stage were identified and potential molecular markers for early diagnosis uncovered. Pathway analysis revealed dysregulation of the Notch and Toll-like receptor pathways in cirrhosis, followed by deregulation of several components of the Jak/STAT pathway in early carcinogenesis, then up-regulation of genes involved in DNA replication and repair and cell cycle in late cancerous stages.
Keywords: 8 pathological stages of HCV-induced HCC

overall_design: 75 samples covering 8 stages of HCV induced HCC, with up to 13 biological repeats for each stage. Normal liver were used as control.

first: Ying-bei

last: Chen

first: Greg

last: Khitrov

first: Weijia

last: Zhang

first: Sassan

last: Roayaie

first: Myron

last: Schwartz

first: Isabel

last: Fiel

first: Swan

last: Thung

first: Vincenzo

last: Mazzaferro

first: Jordi

last: Bruix

first: Erwin

last: Bottinger

first: Scott

last: Friedman

first: Samuel

last: Waxman

first: Josep

middle: M

last: Llovet


title: E coli O157:H7 w/t LB-Glu 7 hr biofilm cells with various chemicals

status: Public on Jul 09 2007

city: College Station

first: Tarun

last: Bansal

state: TX

address: 3122 TAMU

department: Chemical Engineering

institute: Texas A & M University

summary:
E coli O157H7 (EHEC) wildtype 7 hour biofilm cells studied in LB glucose medium with and without chemicals - Epinephrine, Norepinephrine and Indole. Biofilm cells were cultured from glass wool.
Keywords: Effect of different chemicals on EHEC gene expression

overall_design:
Strain: E coli O157H7 (EHEC)
Medium: LB-Glucose
Chemicals: Epinephrine, Norepinephrine and Indole
Time: 7 hours
Cell type: Biofilm cells grown on glass wool

first: Jintae

last: Lee

first: Arul

last: Jayaraman

first: Thomas

last: Wood


title: Genetic variants regulating ORMDL3 expression are determinants of susceptibility to childhood asthma

status: Public on Jul 01 2007

city: Ann Arbor

first: Goncalo

last: Abecasis

state: MI

address: 1420 Washington Heights

institute: University of Michigan

summary:
Asthma is caused by a combination of poorly understood genetic and environmental factors. We found multiple markers on chromosome 17q21 to be strongly and reproducibly associated with childhood onset asthma in family and case-referent panels with a combined P < 10-12.  In independent replication studies the 17q21 locus showed strong association with diagnosis of childhood asthma in 2,320 subjects from a cohort of German children (P = 0.0003) and in 3,301 subjects from the British 1958 Birth Cohort (P = 0.0005). We systematically evaluated the relationships between markers of the 17q21 locus and transcript levels of genes in EBV-transformed lymphoblastoid cell lines from children in the asthma family panel used in our association study. The SNPs associated with childhood asthma were consistently and strongly associated (P <10-22) in cis with transcript levels of ORMDL3, a member of a gene family that encode transmembrane proteins anchored in the endoplasmic reticulum. The results indicate that genetic variants regulating ORMDL3 expression are determinants of susceptibility to childhood asthma.
Keywords: association study, global gene expression, asthma, ORMDL3

overall_design:
Gene expression levels were evaluated in 404 children. We then evaluated the relationship between SNPs in the 17q21 region (which show association to asthma in the same children) with gene expression levels.

Definition of abbreviations used in Sample records:

ped: an anonymized family id, helps identify related individuals
id: an anonymized unique id within each family
fatid: an anonymized id for the father of each individual, helps
          identify siblings
motid: an anonymized id for the mother of each individual, helps
          identify siblings
Sex: 1=male, 2=female
Country: UK = United Kingdom, D = Germany
DDAST: Doctor diagnosed asthma; 0 - unknown, 1 - unaffected, 2 - affected

Note: id, fatid and motid may be recycled across families. A zero indicates the parents are not available and the individual has no siblings in the dataset

first: Miriam

middle: F

last: Moffatt

first: Michael

last: Kabesch

first: Liming

last: Liang

first: Anna

middle: L

last: Dixon

first: David

last: Strachan

first: Simon

last: Heath

first: Martin

last: Depner

first: Andrea

last: von Berg

first: Albrecht

last: Bufe

first: Ernst

last: Rietschel

last: Heinzmann

first: Burkard

last: Simma

last: Frischer

first: Saffron

middle: A

last: Willis-Owen

first: Kenny

middle: C

last: Wong

last: Illig

first: Christian

last: Vogelberg

first: Stephan

middle: K

last: Weiland

first: Erika

last: von Mutius

first: Gonçalo

middle: R

last: Farrall

first: Ivo

middle: G

last: Gut

first: G

last: Lathrop

first: William

middle: O

last: Cookson


title: Harmane Alters the Expression of a Subset of Genes with Diverse Functions in Primary Human Astrocytes

status: Public on Jun 14 2007

first: Sarah

last: Mense

address: 722 West 168th Street

department: Environmental Health Sciences

institute: Columbia University

summary:
Essential tremor (ET) is the most common movement disorder in adults, but little is known about the molecular mechanisms underlying ET pathogenesis.  Harmane is a member of a group of tremorogenic chemicals.  In humans, increased blood harmane concentration is associated with increased risk of ET.  Astrocytes are essential for brain function, and astrocyte dysfuctions are associated with many neurodegenerative diseases.  Therefore, we identified the molecular targets of harmane in primary human astrocytes by using microarray gene expression profiling and computational analysis algorithms. We found that harmane altered the expression of a limited number of genes encoding diverse functions.  Notably, the transcript levels of two GABA receptors and a GABA transporter were altered by harmane, consistent with previous evidence suggesting that the GABAergic neurotransmission system may be disrupted in ET.  Also, we found that the transcript levels of two prominent proinflammatory enzymes, the inducible nitric oxide synthase NOS2A and the cyclooxygenase COX2, and 10 other targets of the proinflammatory IFN-gamma signaling pathway were up-regulated by harmane.  These results together raise the possibility that perturbation of the expression of functions involved in neurotransmission and inflammatory activation of astrocytes might be important mechanisms underlying the neurotoxicity of harmane and ET pathogenesis.
Keywords: treatment comparison

overall_design: Primary human astrocytes (passage 4) were treated with or without 1 uM harmane for 7 days.  Total RNA was extracted by using TRIzol reagent (GIBCOBRL Life Technologies).  The quality of RNA was high as assessed by measuring absorbance at 260 and 280 nm, by gel electrophoresis, and by the quality of microarray data (see below).  We isolated RNA from three independent batches of human astrocytes, which were from different subjects.  No identifiable information from these subjects was available, in keeping with the guidelines on human subjects.  Three independent batches of astrocytes from three different subjects, not the same subject, were used to ensure that our data and conclusions would not totally rely on one unidentified subject, who may have experienced influential environmental or genetic conditions.  The synthesis of cDNAs and biotin-labeled cRNAs were carried out exactly as described in the Affymetrix GeneChip Expression Analysis Technical Manual (2000).  The human genome U133 plus 2.0 arrays were purchased from Affymetrix, Inc.  Probe hybridization and data collection were carried out by the Columbia University Affymetrix GeneChip processing center.  Initial data analysis was performed by using the Affymetrix Microarray suite.

first: Li

last: Zhang


title: A cryptic VEGF T-cell epitope: Identification and characterization by mass spectrometry and T-cell assays.

status: Public on Jun 07 2007

city: Tuebingen

first: Andreas

middle: Oliver

last: Weinzierl

address: Auf der Morgenstelle 15

department: Department of Immunology

country: Germany

institute: University of Tuebingen

summary:
The vascular endothelial growth factor A (VEGF) is involved in various physiological processes such as angiogenesis or wound healing but is also crucial in pathological events such as tumor growth. Thus, clinical anti-VEGF treatments have been developed which could already prove to have enormous beneficial effects for cancer patients. In this article we describe the first VEGF-derived CD8+ T-cell epitope. The natural HLA ligand SRFGGAVVR was identified by differential mass spectrometry in two primary renal cell carcinomas (RCC) and was significantly over-presented on both tumor tissues. SRFGGAVVR is derived from a cryptic translated region of VEGF presumably by initiation of translation at the non-classical start codon CUG499. SRFGGAVVR specific T-cells were generated in vitro using peptide loaded dendritic cells or artificial antigen presenting cells. They were identified by HLA tetramer analysis after in vitro stimulation. SRFGGAVVR specific CD8+ T-cells were fully functional T-effector cells, which were able to secrete IFN-gamma upon stimulation and killed tumor cells in vitro. Additionally, we have quantitatively analyzed VEGF mRNA and protein levels in RCC tumor and normal tissue samples by gene chip analysis, qRT-PCR, in situ hybridization, and bead based immuno assay. In the future, T-cells directed against VEGF as a tumor associated antigen may represent a possible way of combining peptide-based anti-VEGF immunotherapy with already existent anti-VEGF cancer therapies.
Keywords: disease state analysis

overall_design: Two clear cell renal cell carcinomas including autologous normal tissue were analyzed.

first: Dominik

last: Maurer

first: Florian

last: Altenberend

first: Nicole

last: Schneiderhan-Marra

first: Karin

last: Klingel

first: Oliver

last: Schoor

first: Dorothee

last: Wernet

first: Thomas

last: Joos

first: Hans-Georg

last: Rammensee

first: Stefan

last: Stevanovic


title: Analysis of clinical bladder cancer classification according to microarray expression profiles

city: Barcelona

first: Moises

middle: Burset

last: Albareda

state: Barcelona

address: Cartagena, 340-350

department: Molecular biology

country: Spain

institute: Fundacio Puigvert

summary:
Using Affymetrix microarray technology we analyzed the gene expression profiles of the most important pathological categories of bladder cancer in order to detect potential marker genes. Applying an unsupervised cluster algorithm we observed clear differences between tumor and control samples, as well as between superficial and muscle invasive tumors. According to cluster results, the T1 high grade tumor type presented a global genetic profile which could not be distinguished from invasive cases. We described a new measure to classify differentially expressed genes and we compared it against the B-rank statistic as a standard method. According to this new classification method,  the biological functions overrepresented in top differentially expressed genes when comparing tumor versus control samples were associated with growth, differentiation, immune system response, communication, cellular matrix and enzyme regulation. Comparing superficial versus invasive samples, the most important overrepresented biological category was growth and, specifically, DNA synthesis and mitotic cytoskeleton. On the other hand, some under expressed genes have been clearly related to muscular tissue contamination in control samples. Finally, we demonstrated that a pool strategy could be a good option to detect the best differentially expressed genes between two compared conditions.
Keywords: disease state analysis

overall_design: We analyzed gene expression profiles in normal bladder tissues (controls), low grade superficial tumor samples (pathologically classified as Ta low grade, named as Ta), high grade superficial tumors with an unclear clinical  behavior (T1 high grade, named as T1) and high grade muscle invasive tumors (pathologically classified as T2, T3 or T4, named as T2+). We analyzed data using a sub-pooling strategy. The number of individual samples on every pool was: controls (4, 4, 4), Ta (5, 5, 5), T1 (5, 4, 4) and T2+ (5, 5, 5).


title: Identification of human miR-34a-responsive transcripts

city: Baltimore

first: Joshua

middle: T

last: Mendell

state: MD

address: 733 N. Broadway, BRB 471

department: Institute of Genetic Medicine

institute: Johns Hopkins University School of Medicine

summary:
In order to examine the consequences of human miR-34a induction on the transcriptome, HCT116 cells (a colon cancer cell line) were infected with a retrovirus that produces miR-34a.  Gene expression profiles were then monitored using Affymetrix microarrays.
Affymetrix microarrays were used to examine the transcriptomes of HCT116 cells infected with an empty retroviral vector (pMSCV-PIG) or a retroviral vector that expresses human miR-34a.
Keywords: comparison of cells with or without enforced miR-34a expression

overall_design: RNA from retrovirally-infected cells was isolated and hybridized to Affymetrix microarrays.

first: Tsung-Cheng

last: Chang

first: Erik

last: Wentzel


title: Expression data of substantia nigra from postmortem human brain of Parkinson's disease patients (PD)

city: Gaithsburg

first: Jarlath

last: ffrench-Mullen

address: 610 Professional Drive

department: CNS Program

institute: GeneLogic, Inc.

summary:
We used microarrays to detail the global program of gene expression underlying Parkinson's disease
Keywords: normal versus diseased

overall_design: Substantia nigra tissue from postmortem brain of normal and Parkinson disease patients were used for RNA extraction and hybridization on Affymetrix microarrays: 9 replicates for the controls and 16 replicates for the Parkinson's disease patients were used. Both cohorts included males and females.

last: Ffrench-Mullen


title: Expression data from Human MPNST cancer cells infected with G207 and oncolytic HSV

status: Public on Aug 08 2007

city: Cincinnati

first: Bruce

middle: J

last: Aronow

state: OH

institute: Cincinnati Children's Hospital Medical Center

summary:
We used microarrays to identify genes regulated during oncolytic HSV infection. Oncolytic herpes simplex viruses (oHSV) are promising anticancer therapeutics. We sought to identify alterations in gene expression during oHSV infection of human cancer cells. Human malignant peripheral nerve sheath tumor (MPNST) cells were infected with G207, an ICP34.5-deleted oHSV previously evaluated in clinical trials. G207-infected cells demonstrated massive degradation of cellular mRNAs, while a subset were upregulated. A gene signature of 21 oHSV-induced genes contained 7 genes known to be HSV-induced. Go ontology classification revealed that a majority of upregulated genes are involved in Jak/STAT signaling, transcriptional regulation, nucleic acid metabolism, protein synthesis and apoptosis. Ingenuity-defined functional networks highlighted nodes for AP-1 subunits and interferon signaling via STAT1, SOCS1, SOCS3 and RANTES. Upregulation of SOCS1 correlated with sensitivity of MPNST lines to G207 and depletion of SOCS1 reduced virus replication >1-log. The transcriptome of oHSV-induced genes may predict oncolytic efficacy and provides rationale for next generation oncolytics.
Keywords: treated vs non treated

overall_design: 5 human MPNST cancer cell lines were infected with G207 or mock infected for 6 hours followed by RNA extraction and hybridization on Affymetrix microarrays.

first: Yonatan

middle: Y

last: Mahller

first: Bhuvaneswari

last: Sakthivel

first: William

middle: H

last: Baird

first: Ya-Hsuan

last: Hsu

first: Timothy

middle: P

last: Cripe

first: Ruty

last: Mehrian-Shai


title: HDAC-42 modulates acid-induced gene expression in human SEG-1 esophageal adenocarcinoma cells

status: Public on Jul 30 2007

city: Columbus

first: Laura

last: Kresty

address: CCC Building, Rm 302B

department: Internal Medicine

institute: The Ohio State University

summary:
The role of acid and bile salts in the pathogenesis of esophageal carcinoma arising from Barrett's metaplasia has been well established.  Cell proliferation of Barrett's epithelium in response to pulsatile acid exposure has since been confirmed in vivo using endoscopy specimens.
Histone deactylases (HDACs) modulate nucleosomal packaging of DNA, thereby influencing gene transcription and multiple cancer-associated processes.  Thus, we conducted microarray analysis to assess the ability of HDAC-42 to modulate key acid-induced changes as well as to impact other genes altered as the normal esophageal epithelium progresses along the metaplasia-dysplasia-esophageal adenocarcinoma continuum.
Keywords: acid-pulsed cells pretreated with HDAC inhibitor or vehicle

overall_design: SEG-1 cells were pretreated for 24 hours with vehicle or HDAC-42, pulsed with media or acidified media (pH 3.5; 20 mins at 37C), replenished with media or HDAC-42, and harvested 6 hours later.  Global gene expression analysis was conducted using the human genome chip U133 2.0 Plus.


title: Comparative gene expression profile analysis between native human odontoblasts and pulp tissue

city: Oulu

first: Virve

last: Pääkkönen

address: Aapistie 3

department: Institute of Dentistry

country: Finland

institute: University of Oulu

summary:
Knowledge of differential gene expression between pulp and odontoblasts might give insight to the regulation of these spatially related but functionally diverse cells. Our aim was large-scale analysis of expression profiles of native human pulp tissue and odontoblasts, and search for genes expressed only in odontoblasts.
Keywords: comparative expression analysis

overall_design: Microarray using Affymetrix HGU133A (for pulp) and  HGU133plus 2.0 array (for odontoblasts) was performed to pooled pulp and odontoblasts of native human third molars. Microarray analysis’ repeatability was estimated by comparing our pulp sample with expression profiles of two pulp samples downloaded from the GEO-database. The genes expressed only in our pulp sample or in odontoblasts were divided into categories, and the expression of selected odontoblast-specific genes of extracellular matrix (ECM) organization and biogenesis category was confirmed with RT-PCR and Western blot.

first: Jussi

last: Vuoristo

first: Tuula

last: Salo

first: Leo

last: Tjäderhane


title: Profiling of Angiotensin II Rapid Response Genes in Human, Bovine, and Rat Adrenocortical Cells.

status: Public on Jul 18 2007

first: Edson

middle: da Fonseca

last: Nogueira

summary:
Angiotensin II (Ang-II) regulates adrenal steroid production and gene transcription through several signaling pathways. Changes in gene transcription occur within minutes after Ang-II stimulation, causing an acute increase in aldosterone production and subsequent increase in the overall capacity to produce aldosterone. Our goal was to compare the Ang-II regulation of early gene expression and confirm the upregulation of selected genes using quantitative real-time RT-PCR (qPCR) across three species: human, bovine, and rat.
Keywords: species comparison, adrenal glomerulosa, microarray, angiotensin II

sample_organism:
Homo sapiens
Bos taurus
Rattus norvegicus

overall_design: Microarray analysis was performed using samples from control and Ang-II-(10 nM) treated (1 hour) cells from human adrenocortical tumor cell line H295-R, and primary adrenal glomerulosa cells from bovine and rat, applied respectively to human HG-133 + 2 , bovine, and rat 230-2 Affymetrix chips. qPCR was performed to confirm upregulation of selected genes using mRNA. Dye Swap was not used. Human samples (H295R) include 3 replicates of Basal (controls) and 3 repeats of Angiotensin II samples. Human H295R cells include also a group of cycloheximide (protein synthesis blocker) and angiontensin II + cycloheximide in order to check if the genes were direct targets of angiotensin II.

first: Wendy

middle: B

last: Bollag

first: Nicole

last: Gallo-Payet

first: Claudia

last: Vargas

middle: E

last: Rainey


title: Identification of the JNK Signaling Pathway as a Functional Target of the Tumor Suppressor PTEN

status: Public on Jun 01 2007

city: Los Angeles

first: Igor

last: Vivanco

state: CA

address: 700 Westwood plaza

department: Mol. & Md. Pharmacology

institute: UCLA

summary:
Although most of the oncogenic phenotypes of PTEN loss have been attributed to AKT activation, AKT alone is not sufficient to induce all of the biological activities associated with PTEN inactivation.  We searched for additional PTEN-regulated pathways through gene set enrichment analysis (GSEA) and found that PTEN inactivation causes an enrichment of genes associated with JNK activation.  Biochemically, PTEN-null cells exhibit higher JNK activity, and genetic studies demonstrate that JNK functions parallel to and independently of AKT.  Furthermore, PTEN deficiency sensitizes cells to JNK inhibition.  We also found that negative feedback regulation of PI3K was impaired in PTEN-null cells.  These findings implicate JNK in PI3K-driven cancers and demonstrate the utility of GSEA to identify functional pathways using genetically defined systems.
Keywords: Genetic modification:  PTEN RNAi

overall_design: A431, HCC827 and SKBR-3 cells were retrovirally and stably transduced with a small hairpin RNA targeting human PTEN.  PTEN knockdown was confirmed at the protein level by western blot to be approximately 90%.  PTEN-deficient sublines were named by adding the suffix "PR"(for PTEN RNAi) at the end of each of the parental cell line names.  For example, PTEN deficient A431 cells are called A431-PR.  RNA was extracted from each of the parental and PTEN-deificient lines.  Each sample was done in duplicate (ie two chips per sample) for a total of twelve samples.

first: Nicolaos

last: Palaskas

first: Chris

last: Tran

first: Stephen

last: Finn

first: Gad

last: Getz

first: Norman

last: Kennedy

first: Jing

last: Jiao

first: Joshua

last: Rose

first: Wanling

last: Xie

first: Massimo

last: Loda

first: Todd

last: Golub

first: Ingo

last: Mellinghoff

first: Roger

last: Davis

first: Hong

last: Wu

first: Charles

middle: L

last: Sawyers


title: Robustness of gene expression signatures in leukemia: comparison of three distinct total RNA preparation procedures.

status: Public on May 31 2007

city: Padova

first: Andrea

last: Zangrando

address: Via Giustiniani, 3

department: Woman and Child's Health

country: Italy

institute: University of Padova

summary:
Microarray gene expression (MAGE) signatures allow insights into the transcriptional processes of leukemias and may evolve as a molecular diagnostic test. Introduction of MAGE into clinical practice of leukemia diagnosis will require comprehensive assessment of variation due to the methodologies.
For the first time, we present a study focused on analyzing the impact of different RNA preparation procedures on gene expression data for different subtypes of pediatric acute leukemias.
Keywords: comparison of various total RNA preparation procedures

overall_design: We systematically assessed the impact of three different total RNA isolation procedures on variation in expression data: method A: lysis of mononuclear cells, followed by lysate homogenization and RNA extraction; method B: organic solvent based RNA isolation, and method C: organic solvent based RNA isolation followed by purification.

first: Marta

last: Campo Dell'Orto

first: Alexander

last: Kohlmann


title: Predicting Features of Breast Cancer with Gene Expression Patterns

status: Public on May 26 2007

city: Boston

first: James

middle: Dirk

last: Iglehart

state: MA

address: 44 Binney St.

department: Cancer Biology

institute: Dana Farber Cancer Institute

summary:
Predictors built from gene expression data accurately predict ER, PR, and HER2 status, and divide tumor grade into high-grade and low-grade clusters; intermediate-grade tumors are not a unique group.  In contrast, gene expression data cannot be used to predict tumor size or lymphatic-vascular invasion.
Keywords: disease state analysis

overall_design:
Microarray data from the tumors of 129 patients were analyzed for the ability to predict biomarkers (ER, PR, HER2), histologic features (grade and lymphatic-vascular invasion), and stage-related information (tumor size and lymph node metastasis). Multiple statistical predictors were used and the prediction accuracy determined by error rates of prediction and by dimensional scaling and visualization of the states under study. Models to predict lymph node metastasis were built by combinations of molecular, histologic and anatomic features.

***GSM125119.CEL and GSM125120.CEL are corrupt***

first: Xuesong

last: Lu

first: Xin

first: Zhigang

last: Wang

middle: D

first: Xuegong

last: Zhang

last: Richardson


title: A toxicogenomic profile of Mn  in Human Astrocytes

first: Sarah

last: Mense

summary:
Exposure of adult humans to manganese (Mn) has long been known to cause neurotoxicity.  Recent evidence also suggests that exposure of children to Mn is associated with developmental neurotoxicity.  Astrocytes are critical for the proper functioning of the nervous system, and they play active roles in neurogenesis, synaptogenesis and synaptic neurotransmission.  In this report, to help elucidate the molecular events underlying Mn neurotoxicity, we systematically identified the molecular targets of Mn in primary human astrocytes by using microarray gene expression profiling and computational data analysis algorithms.  We found that Mn altered the expression of diverse genes ranging from those encoding cytokines and transporters to signal transducers and transcriptional regulators.  Particularly, 28 genes encoding proinflammatory chemokines, cytokines and related functions were up-regulated whereas 15 genes encoding functions involved in DNA replication and repair and cell cycle checkpoint control were down-regulated.  These results are further supported by data from real-time RT-PCR, Western blotting and flow cytometric analyses.  In addition, analysis of common regulators revealed that 16 targets known to be positively affected by the IFN-gamma signaling pathway were up-regulated by Mn, suggesting that the proinflammatory IFN-gamma signaling pathway was likely activated.  These results raise the possibility that inflammatory activation of astrocytes and the increased expression of proinflammatory cytokines and chemokines and/or the activation of related signaling pathways might be an important mechanism leading to Mn neurotoxicity.
Keywords: treatment comparison

overall_design: Primary human astrocytes (passage 4) were treated with or without 200 uM MnCl2 for 7 days.  Total RNA was extracted by using TRIzol reagent (GIBCOBRL Life Technologies).  The quality of RNA was high as assessed by measuring absorbance at 260 and 280 nm, by gel electrophoresis, and by the quality of microarray data (see below).  We isolated RNA from three independent batches of human astrocytes, which were from different subjects.  No identifiable information from these subjects was available, in keeping with the guidelines on human subjects.  Three independent batches of astrocytes from three different subjects, not the same subject, were used to ensure that our data and conclusions would not totally rely on one unidentified subject, who may have experienced influential environmental or genetic conditions.  The synthesis of cDNAs and biotin-labeled cRNAs were carried out exactly as described in the Affymetrix GeneChip Expression Analysis Technical Manual (2000).  The human genome U133 plus 2.0 arrays were purchased from Affymetrix, Inc.  Probe hybridization and data collection were carried out by the Columbia University Affymetrix GeneChip processing center.  Initial data analysis was performed by using the Affymetrix Microarray suite.

first: Li

middle: M


title: Proinflammatory Responses of Human Airway Cells to Ricin Involve Stress-Activated Protein Kinases and NF-κB

status: Public on May 22 2007

city: Portland

first: John

last: Wong

state: OR

address: 3181 SW Sam Jackson Park Rd. (L215)

department: Cell & Developmental Biology

institute: Oregon Health & Science University

summary:
Ricin is a potential bioweapon because of its toxicity, availability, and ease of production.  When delivered to the lungs, ricin causes severe pulmonary damage with symptoms that are similar to those observed in acute lung injury and adult respiratory distress syndrome.  The airway epithelium plays an important role in the pathogenesis of many lung diseases, but its role in ricin intoxication has not been elucidated.  Exposure of cultured primary human airway epithelial cells to ricin resulted in the activation of stress-activated protein kinases (SAPKs) and NF-κB and in the increased expression of multiple proinflammatory molecules. Among the genes upregulated by ricin and identified by microarray analysis were those associated with transcription, nucleosome assembly, inflammation, and response to stress.  Sequence analysis of the promoters of these genes identified NF-κB as one of the transcription factors whose binding sites were over-represented.  Although airway cells secrete TNF-α in response to ricin, blocking TNF-α did not prevent ricin-induced activation of NF-κB.   Inhibition of p38 MAPK by a chemical inhibitor and NF-κB by short interfering RNA resulted in a marked reduction in the expression of proinflammatory genes, demonstrating the importance of these two pathways in ricin intoxication.  Therefore, the p38 MAPK and NF-κB pathways are potential therapeutic targets for reducing the inflammatory consequences of ricin poisoning.
Keywords: Comparative genomic hybridization

overall_design: Control RNA from untreated primary human airway cells was compared to RNA from ricin-treated airway cells

first: Veselina

last: Korcheva

first: David

last: Jacoby

first: Bruce

last: Magun


title: Identification of Cellular Genes Targeted by KSHV-Encoded MicroRNAs

status: Public on May 11 2007

city: Gainesville

first: M Cecilia

last: Lopez

state: FL

address: 1376 Mowry Road

department: Molecular Genetics and Microbiology

institute: University of Florida

summary:
MicroRNAs are 19 to 23 nt RNAs that post-transcriptionally regulate gene expression. Human cells express several hundred miRNAs which regulate important biological pathways such as development, proliferation, and apoptosis. Recently, 12 microRNA genes have been identified within the genome of Kaposi’s sarcoma-associated herpesvirus; however, their functions are still unknown. To identify host cellular genes that may be targeted by these novel viral regulators, we performed gene expression profiling in cells stably expressing KSHV-encoded miRNAs. Data analysis revealed a set of 81 genes whose expression was significantly changed in the presence of miRNAs. While the majority of changes were below 2-fold, eight genes were down-regulated between 4- and 20-fold. We confirmed miRNA-dependent regulation for three of these genes and found that protein levels of thrombospondin 1 (THBS1) were decreased >10-fold. THBS1 has previously been reported to be down-regulated in KS lesions and has known activity as a strong tumor suppressor and anti-angiogenic factor, exerting its anti-angiogenic effect in part by activating the latent form of TGF-b. We show that reduced THBS1 expression in the presence of viral miRNAs translates into decreased TGF-b activity. These data suggest that KSHV-encoded miRNAs may contribute directly to pathogenesis by down-regulation of THBS1, a major regulator of cell adhesion, migration, and angiogenesis.
Keywords: comparison, experiemental versus control

overall_design: 293 cells were stably transfected with either a vector expressing 10 KHSV encoded microRNAs or empty vector control.  Cells were selected through G418.  4 samples were collected for each cell line and RNA collected for Affymetrix microarray analysis.  Results show which transcripts are changed in the presence of KSHV microRNA expression.

first: Mark

middle: A

last: Samols

first: Rebecca

last: Skalsky

first: Ann

last: Maldonado

first: Alberto

last: Riva

first: MCecilia

first: Henry

middle: V

last: Baker

first: Rolf

last: Renne


title: Modulating hypoxia-inducible transcription by disrupting the HIF-1-DNA interface

status: Public on May 17 2007

city: Pasadena

first: Peter

last: Dervan

address: 1200 East California Blvd

institute: California Institute of Technology

summary:
Transcription mediated by hypoxia inducible factor (HIF-1) contributes to tumor angiogenesis and metastasis but is also involved in the activation of cell-death pathways and normal physiological processes.  Given the complexity of HIF-1 signaling it could be advantageous to target a subset of HIF-1 effectors rather than the entire pathway.  We compared the genome-wide effects of three molecules that each interfere with the HIF-1-DNA interaction: a polyamide targeted to the hypoxia response element (HRE), siRNA targeted to HIF-1α, and echinomycin, a DNA binding natural product with a similar but less specific sequence preference to the polyamide.  The polyamide affects a subset of hypoxia-induced genes that are consistent with the binding site preferences of the polyamide.  For comparison, siRNA targeted to HIF-1α and echinomycin each affect the expression of nearly every gene induced by hypoxia.  Remarkably, the total number of genes affected by either polyamide or HIF-1α siRNA over a range of thresholds is comparable.  The data shows how polyamides can be used to affect a subset of a pathway regulated by a transcription factor.  In addition, this study offers a unique comparison of three complementary approaches towards exogenous control of endogenous gene expression.
Keywords: Gene expression changes in cultured U251 cells after DFO-stimulation and various treatment conditions

overall_design: Hypoxia-mimetic DFO (deferoxamine)-stimulated U251 cells that were treated with polyamide 1, HIF-1a siRNA, and echinomycin were compared to control cells that were also DFO-stimulated.  Cells not stimulated with DFO were also compared to the DFO-stimulated controls.  Three biological replicates were included for each treatment/condition.

first: Nicholas

middle: G

last: Nickols

first: Claire

middle: S

last: Jacobs

first: Michelle

last: Farkas


title: Suppression of androgen receptor mediated gene expression by a sequence-specific DNA binding polyamide

status: Public on May 05 2007

summary:
Androgen Receptor (AR) is essential for the growth and progression of prostate cancer in both hormone-sensitive and hormone-refractory disease.  We have designed a sequence-specific DNA binding polyamide (1) that targets the consensus androgen response element (ARE).  This polyamide binds the PSA promoter ARE, inhibits androgen-induced expression of PSA and several other AR-regulated genes in cultured prostate cancer cells, and reduces AR occupancy at the PSA promoter and enhancer.  Down-regulation of PSA by this polyamide was comparable to that produced by the synthetic anti-androgen bicalutamide (Casodex) at the same concentration.  Genome-wide expression analysis reveals that a similar number of transcripts are affected by treatment with the polyamide and with bicalutamide.  Direct inhibition of AR-DNA binding by sequence-specific DNA binding small molecules could offer an alternative approach to antagonizing AR activity.  A polyamide (2) that targets a different DNA sequence is included as a control.
Keywords: Gene expression changes in cultured LNCaP cells after DHT-stimulation and various treatment conditions

overall_design: DHT (dihydrotestosterone)-stimulated LNCaP cells that were treatment with polyamide 1, polyamide 2, bicalutamide were compared to control cells that were also DHT-stimulated.  Cells not stimulated with DHT were also compared to the DHT-stimulated controls.  Three biological replicates were included for each treatment/condition except the no-DHT induced controls, which were in biological duplicate.


title: Microarray analysis of NL and PCOS oocytes

status: Public on Jun 05 2007

city: Lincoln

first: Jennifer

middle: R

last: Wood

state: NE

address: 3800 Fair St.

department: Animal Science

institute: University of Nebraska

summary:
Polycystic ovary syndrome (PCOS), the most common cause of anovulatory infertility, is characterized by increased ovarian androgen production, arrested follicle development, and is frequently associated with insulin resistance.  These PCOS phenotypes are associated with exaggerated ovarian responsiveness to FSH and increased pregnancy loss.  To examine whether the perturbations in follicle growth and the intrafollicular environment affects development of the mature PCOS oocyte, genes that are differentially expressed in PCOS compared to normal oocytes were defined using microarray analysis.  This analysis detected approximately 8000 transcripts.  Hierarchical clustering and principal component analysis revealed differences in global gene expression profiles between normal and PCOS oocytes.  374 genes had a statistically-significant increase or decrease in mRNA abundance in PCOS oocytes.  A subset of these genes was associated with chromosome alignment and segregation during mitosis and/or meiosis, suggesting that increased mRNAs for these proteins may negatively affect oocyte maturation and/or early embryonic development.  Of the 374 differentially expressed genes, 68 contained putative androgen receptor, retinoic acid receptor, and/or peroxisome proliferating receptor gamma binding sites, including 9 of the genes involved in chromosome alignment and segregation.  These analyses demonstrated that normal and PCOS oocytes that are morphologically indistinguishable and of high quality exhibit different gene expression profiles.  Furthermore, altered mRNA levels in the PCOS oocyte may contribute to defects in meiosis and/or mitosis which might impair oocyte competence for early development and therefore contribute to poor pregnancy outcome in PCOS.
Keywords: disease state analysis

overall_design: A single MII oocyte, defined by one polar body in the perivitelline space and no visible nuclear structure in the cytoplasm, was collected from 6 individual NL and 6 individual PCOS ovaries, placed immediately in TRIzol (Sigma, St. Louis MO), and stored at -80 C until further study.  Total RNA was isolated from each oocyte and subjected to three rounds of linear amplification with the Ovation Biotin RNA Amplification and Labeling System (NuGen Technologies, San Carlos CA) per the manufacturer’s instructions.  RNA from the GeneChip Eukaryotic Poly-A RNA Control Kit (Affymetrix, Santa Clara CA) was amplified and labeled under the same conditions for a positive control.  Affymetrix GeneChip Human Genome U133 Plus 2.0 microarray chips (Affymetrix, Santa Clara, CA) were hybridized at the University of Pennsylvania Microarray Core Facility.  Briefly, the linear-amplified, biotin-labled cDNA from 6 NL (N1-N6) and 6 PCOS (P1-P6) oocytes was hybridized to individual Affymetrix U133 chips.  The fluorescence intensity of each chip was normalized to a trimmed mean signal of 150.  Each transcript on the U133 chip was defined as present or absent in each oocyte sample using the Affymetrix Microarray Suite 5.0.

first: Daniel

last: Dumesic

last: Abbott

first: Jerome

middle: F

last: Strauss III


title: Expression data from human breast tissue

status: Public on May 30 2007

first: Andrea

last: Richardson

address: 75 Francis Street

department: Pathology

institute: Brigham and Women's Hospital

summary:
bulk breast tumor RNA from patient
Abstract: Sporadic basal-like cancers (BLC) are a distinct class of human breast cancers that are phenotypically similar to BRCA1-associated cancers. Like BRCA1-deficient tumors, most BLC lack markers of a normal inactive X chromosome (Xi). Duplication of the active X chromosome and loss of Xi characterized almost half of BLC cases tested. Others contained biparental but nonheterochromatinized X chromosomes or gains of X chromosomal DNA. These abnormalities did not lead to a global increase in X chromosome transcription but were associated with overexpression of a small subset of X chromosomal genes. Other, equally aneuploid, but non-BLC rarely displayed these X chromosome abnormalities. These results suggest that X chromosome abnormalities contribute to the pathogenesis of BLC, both inherited and sporadic.
total 62 sample incudes 43 tumor, 7 normal breast and  12 normal organelle
Keywords: disease state analysis

overall_design: compare expression of tumor to normal breast tissue


title: Hyperexpression and Downregulation of Melanotransferrin on Various Cell Lines

city: Sydney

first: Des

middle: Raymond

state: New South Wales

address: 92-94 Parramatta Rd, Sydney

department: Bosch Institute

country: Australia

institute: University of Sydney

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series

first: Yohan

last: Suryo Rahmanto

first: Louise

last: Dunn


title: Expression data from overnight minimal medium culture

status: Public on May 01 2007

first: Yanqin

last: Yang

address: 600 N. Wolfe Street

institute: The Johns Hopkins University

summary:
Comparison of gene experission profiles of Ecoli WT3110 and phoU mutant
Keywords: one time point

overall_design: DNA microarray analysis of the PhoU mutant with the wild type strain W3110 as a control. The PhoU mutant and the wild type strain were grown in MOPS minimal medium overnight when the RNA was isolated using MasterPure RNA Purification Kit and reverse transcribed for making probes for array hybridization on Affymetrix microarrays.


title: shRNA-mediated knock down of Bmi-1 and Mel-18 in medulloblastoma cells

city: Cambdridge

first: Dmitri

last: Wiederschain

address: 250 Massachusetts Avenue

department: Oncolology

institute: Novartis

summary:
Bmi-1 and Mel-18 are close structural homologues that belong to the polycomb group (PcG) of transcriptional regulators of homeotic gene expression in development. They are believed to stably maintain repression of gene expression by altering the state of chromatin at specific promoters.  A number of clinical and experimental observations have also implicated Bmi-1 in tumorigenesis and stem cell maintenance.  Bmi-1 overexpression or amplification has been observed in a number of human malignancies, particularly in B-cell lymphomas, medulloblastomas and breast cancer. We report here that shRNA-mediated knock-down of either Bmi-1 or Mel-18 in human medulloblastoma DAOY cells results in the inhibition of proliferation, loss of clonogenic survival and anchorage-independent growth, and suppression of xenograft tumor formation in nude mice. Furthermore, overexpression of both Bmi-1 and Mel-18 significantly increased clonogenic survival of Rat1 fibroblasts.  In contrast, stable downregulation of Bmi-1 or Mel-18 alone did not affect the growth of SK-OV-3 or U2OS cancer cell lines or normal human WI38 fibroblasts.  Gene expression analysis of shRNA-expressing DAOY cells has demonstrated a significant overlap in the Bmi-1- and Mel-18-regulated genes and revealed novel gene targets under their control.  Taken together, these results suggest that Bmi-1 and Mel-18 might have overlapping functions in human tumorigenesis.
Keywords: shRNA knock-down

overall_design: DAOY cells were transduced with the indicated lentiviral shRNA and stable cell lines were generated by selection in puromycin (1 µg/ml). 2 x 105 cells of each stable cell line were plated onto 60 mm dishes in triplicates and harvested 4 days later. Triplicate total RNA samples from shRNA-expressing and mock-transduced DAOY cells were isolated using affinity resin (Qiagen RNeasy Mini Kit, Qiagen AG). RNA Integrity and purity were assessed with the RNA 6000 Nano LabChip system on Bioanalyzer 2100 (Agilent Technologies).   Gene array analysis was conducted at the Genomics Factory at Novartis PHARMA AG, Basel, Switzerland using Gene Chip Human Genome 133 2.0 Plus Expression Array (Affymetrix Inc.).

first: Michael

last: Jones

last: Morrissey

first: Paul

last: Fordjour

first: John

last: Benson


title: AML mRNA hybridization to HGU133 Plus 2.0

city: Shimotsukeshi

first: Hiroyuki

last: Mano

state: Tochigi

address: 3311-1 Yakushiji

department: Division of Functional Genomics

country: Japan

institute: Jichi Medical University

summary:
CD34-positive fractions were used for mRNA preparation, which was hybridized to HGU 133 Plus 2.0 arrays.
Keywords: Disease state analysis

overall_design: CD34-positive fractions of 22 AML patients were analyzed.


title: Hyperexpression of Human Melanotransferrin on SK-N-MC Cell Line

summary:
Melanoma tumor antigen p97 or melanotransferrin (MTf) is an iron (Fe)-binding protein with high homology to serum transferrin. MTf is expressed at very low levels in normal tissues and in high amounts in melanoma cells. The over-expression of MTf in tumor cells was hypothesized to assist rapidly proliferating neoplastic cells with their increased Fe requirements. However, our recent characterization of the MTf knockout (MTf -/-) mouse demonstrated that MTf did not have an essential role in Fe metabolism. To understand the function of MTf, we utilized whole-genome microarray analysis to examine the gene expression profile of five models after modulating MTf expression. These models included two new stably transfected MTf hyper-expression models (SK-N-MC neuroepithelioma and LMTK- fibroblasts) and one cell type (SK-Mel-28 melanoma) where MTf was down-regulated by post-transcriptional gene silencing. These findings were compared to alterations in gene expression identified using the MTf -/- mouse. In addition, the changes identified from the gene array data were also assessed in a new model of MTf down-regulation in SK-Mel-2 melanoma cells. In the cell line models, MTf hyper-expression led to increased cellular proliferation, while MTf down-regulation resulted in decreased proliferation. Across all five models of MTf down- and up-regulation, we identified three genes modulated by MTf expression. These included ATP-binding cassette sub-family B member 5 (Abcb5), whose change in expression mirrored MTf down- or up-regulation. In addition, thiamine triphosphatase (Thtpa) and transcription factor 4 (Tcf4) were inversely expressed relative to MTf levels across all five models. The products of these three genes are involved in membrane transport, thiamine phosphorylation and cell proliferation/survival, respectively. This study identifies novel molecular targets directly or indirectly regulated by MTf and potential pathways involved in its function. These molecular targets could be involved, at least in part, to the role of MTf in modulating proliferation.
Keywords: Melanotransferrin, hyperexpression cell lines, comparative genomic hybridization

overall_design: Hyper-expression constructs of the human MTf cDNA was generated using the pCMV-Script® (Stratagene, La Jolla, CA) expression vector. The inserts were sequenced and confirmed against human MTf (Genbank Accession: NM_005929) using the public NCBI database. Transfections of pCMV-Script® transgenes/vectors into human SK-N-MC neuroepithelioma cells was performed with Lipofectin® reagent (Invitrogen, Melbourne, Australia) according to the manufacturer’s protocol. Total RNA was isolated from the cells using TRIzol Reagent® (Sigma-Aldrich) according to the manufacturer’s protocol. Total RNA from the stably-transfected SK-N-MC neuroepithelioma cell lines were prepared and hybridized onto Human Genome U133 Plus 2.0 Array. The human GeneChip® U133 Plus 2.0 consists of greater than 47,000 transcripts and variants from over 38,500 well characterized human genes (Affymetrix, Santa Clara, CA).


title: EHEC hydroxyindole project

status: Public on Apr 26 2007

city: College Station

first: Jintae

last: Lee

department: Chemical Engineering

institute: Texas A&M University

summary:
For the microarray experiments, 10 g glass wool (Corning Glass Works, Corning, N.Y.) were used to form biofilms (30) in 250 mL in 1 L Erlenmeyer shake flasks which were inoculated with overnight cultures diluted that were 1:100.  For EHEC with 7-hydroxyindole and isatin, 1000 mM 7-hydroxyindole in 250 mL DMF, 250 mM isatin in 250 mL DMF, or 250 mL DMF alone were added to cells grown in LB.  The cells were shaken at 250 rpm and 30°C for 7 hours to form biofilms on the glass wool, and RNA was isolated from the suspension cells and the biofilm.
Keywords: Biofilm study

overall_design:
Strain: EHEC EDL933 (ATCC 43895) wild type
Medium: LB
Cells: Suspension cells and biofilm cells on glass wool
Time: 7h

first: Thomas

middle: K

first: Tarun

last: Bansal

first: Arul

last: Jayaraman

first: William

last: Bentley


title: Spermatozoal RNA Profiles from Normally Fertile and Heterogeneously Teratozoospermic Subjects

status: Public on Apr 27 2007

city: Detroit

first: Stephen

last: Krawetz

address: 275 East Hancock, CS Mott Center

institute: Wayne State University

first: Adrian

last: Platts

first: David

middle: J

last: Dix

first: Hector

last: Chemes

first: Kary

last: Thompson

first: Robert

last: Goodrich

middle: C

last: Rockett

first: Vanesa

middle: Y

last: Rawe

first: Silvina

last: Quintana

last: Diamond

first: Lillian

last: Strader


title: Mapping of HNF4α binding sites, acetylation of histone H3 and expression in Caco2 cells

status: Public on May 10 2007

city: Copenhagen

first: Jesper

middle: T

last: Troelsen

address: Blegdamsvej 3b

country: Denmark

institute: Univ. of Copenhagen

type: Genome binding/occupancy profiling by genome tiling array

summary:
Background & Aims: The role of HNF4α has been extensively studied in hepatocytes and pancreatic β cells, but emerging evidence indicates that HNF4α is a key regulator of intestinal epithelial cell differentiation as well. The aim of the present work is to identify HNF4α target genes in the intestine in order to elucidate the role of HNF4α in differentiation of the intestinal epithelial cells.    Results: One thousand one hundred and seventy-six genes were identified as HNF4α targets, many of which have not previously been described as being regulated by HNF4α. The 1,176 genes contributed significantly to gene ontology (GO) pathways categorized by lipid and amino acid transport and metabolism. A thorough analysis of Cdx-2, trehalase, and cingulin promoters verified that these genes are regulated by HNF4α. In each case we were able to identify a functional HNF4α binding site in their promoters.  Conclusions: HNF4α regulation of the Cdx-2 promoter unravels a transcription factor network also including HNF1α and β, all of which are transcription factors involved in intestinal development and gene expression.
Keywords: ChIP-CHIP and expression data

overall_design: Methods: We have performed a ChIP-chip analysis of the human intestinal cell line Caco-2 in order to make a genome-wide identification of HNF4α binding to promoter regions. The HNF4α ChIP-chip data was matched with gene expression and histone H3 acetylation status of the promoters in order to identify HNF4α binding to actively transcribed genes with an open chromatin structure.

first: Mette

last: Nielsen

first: Simon

last: Bressendorff

first: Jette

last: Møller

first: Jørgen

last: Olsen


title: Gene Profiling of Endometrium Reveals Progesterone Resistance and Candidate Genetic Loci in Women with Endometriosis

city: San Francisco

first: Linda

last: Giudice

address: 513 Parnassus Ave. HSE 1619

department: OBGYN and RS

institute: University of California, San Francisco

summary:
The transition of regularly cycling endometrium from the proliferative or Estrogen-dominant phase of the menstrual cycle to the Progesterone-dominant Early and Mid Secretory phases requires wide-spread changes in gene expression that shift the endometrium from a proliferative capacity to a differentiated 'decidual' phenotype in preparation for implantation.  This process appears delayed in women with severe endometriosis, suggestive of a progesterone resistant endometrium in this disease.
Keywords: disease state analysis

overall_design: Endometrial biopsies were obtained from women both with normal endometrial pathologies and no history of endometriosis and from women with laporoscopy proven moderate-severe stage endometriosis.  Samples were collected from the Proliferative(PE), Early Secretory (ESE) and Midsecretory (MSE) phases. Samples were then processed for Total RNA isolation and Affymetrix chip hybridization.


title: Spermatozoal RNA Profiles (U133 Plus 2.0 Array)

last: Krawetz

summary:
Normal human spermatogenesis concludes with the formation of large numbers of  morphologically well developed spermatozoa.  While transcriptionally quiescent these cells carry  an RNA payload that reflects the final spermiogenic phase of transcription. We report here the spermatozoal  transcript profiles characteristic of normally fertile individuals and infertile males suffering from a  consistent and severe teratozoospermia in which under 4% of spermatozoa are morphologically normal.  RNA was  extracted from the purified sperm cells of ejaculate and hybridized to Affymetrix U133 (v2) Microarrays.
Keywords: disease state analysis

overall_design: Spermatozoal RNAs were prepared from the semen samples of 21 individuals.  An  asymmetric dual block design was adopted with biological replicates in both blocks.  13 semen samples were  assessed from normally fertile males who had fathered at least one child.  8 semen samples were assessed from  infertile individuals with a severe and consistent heterogeneous teratozoospermia who showed no other abnormal  semen parameters.

middle: E

first: John

first: Michael

middle: P


title: Gene expression changes in anti-FcgRIIb treated DCs and monocytes

status: Public on Apr 30 2007

city: New Haven

first: Kavita

last: Dhodapkar

address: 333 Cedar Street

institute: Yale

summary:
The ability of dendritic cells (DCs) to activate immunity is linked to their maturation status. In prior studies we have shown that selective antibody-mediated blockade of inhibitory FcgRIIB receptor on human DCs in the presence of activating immunoglobulin (Ig) ligands leads to DC maturation and enhanced immunity to antibody-coated tumor cells. Here we show that Fcg receptor (FcgR) mediated activation of human monocytes and monocyte-derived DCs is associated with a distinct gene expression pattern, including several inflammation associated chemokines as well as type 1 interferon (IFN) response genes including the activation of signal transducer and activator of transcription 1 (STAT1).
Keywords: response to antibody

overall_design: To further characterize FcgR mediated enhancement of DC function, we analyzed the gene expression profiles (GEP) of pure populations of monocyte-derived DCs from healthy donors (n=5) using Affymetrix Human Genome U133 Plus2.0 microarrays. Immature DCs cultured in 1% plasma were treated for 24 hours with either anti-FcgRIIB or isotype control antibody. To test whether FcgR mediated DC maturation was distinct from other maturation stimuli, we also compared DCs matured using the inflammatory cytokine cocktail (TNF-a, IL-1b, IL-6 and PGE2) commonly utilized in DC immunotherapy trials. In addition, we also treated Cd14+ monocytes (n=3) with anti-FcgRIIB antibody or isotype control. In order to better characterize the interferon responsive genes in DCs, we treated immature DCs (n=3) with 1000 U/ml of IFN-a2b.


title: Downregulation of Human Melanotransferrin on SK-Mel-28 Cell Line

first: Des

last: Richardson

summary:
Melanoma tumor antigen p97 or melanotransferrin (MTf) is an iron (Fe)-binding protein with high homology to serum transferrin. MTf is expressed at very low levels in normal tissues and in high amounts in melanoma cells. The over-expression of MTf in tumor cells was hypothesized to assist rapidly proliferating neoplastic cells with their increased Fe requirements. However, our recent characterization of the MTf knockout (MTf -/-) mouse demonstrated that MTf did not have an essential role in Fe metabolism. To understand the function of MTf, we utilized whole-genome microarray analysis to examine the gene expression profile of five models after modulating MTf expression. These models included two new stably transfected MTf hyper-expression models (SK-N-MC neuroepithelioma and LMTK- fibroblasts) and one cell type (SK-Mel-28 melanoma) where MTf was down-regulated by post-transcriptional gene silencing. These findings were compared to alterations in gene expression identified using the MTf -/- mouse. In addition, the changes identified from the gene array data were also assessed in a new model of MTf down-regulation in SK-Mel-2 melanoma cells. In the cell line models, MTf hyper-expression led to increased cellular proliferation, while MTf down-regulation resulted in decreased proliferation. Across all five models of MTf down- and up-regulation, we identified three genes modulated by MTf expression. These included ATP-binding cassette sub-family B member 5 (Abcb5), whose change in expression mirrored MTf down- or up-regulation. In addition, thiamine triphosphatase (Thtpa) and transcription factor 4 (Tcf4) were inversely expressed relative to MTf levels across all five models. The products of these three genes are involved in membrane transport, thiamine phosphorylation and cell proliferation/survival, respectively. This study identifies novel molecular targets directly or indirectly regulated by MTf and potential pathways involved in its function. These molecular targets could be involved, at least in part, to the role of MTf in modulating proliferation.
Keywords: Melanotransferrin, siRNA, comparative genomic hybridization

overall_design: To prepare a construct capable of generating hairpin siRNA specific for human MTf mRNA, the expression vector, pSilencer™ 3.1-H1 neo (Ambion, Texas, USA) was used. The vector was used to clone transgene of 66-bp that transcribe 19-mer double-stranded hairpin RNAs of the target gene. The transgene were specifically targeted to positions 2031-2049-bp in the MTf gene (Genbank Accession: NM_005929). Transfections of pS-MTf transgenes or pS-scrambled vectors into human SK-Mel-28 melanoma cells were performed with LipofectamineTM 2000 reagent (Invitrogen, Melbourne, Australia). Total RNA was isolated from the cells using TRIzol Reagent® (Sigma-Aldrich) according to the manufacturer’s protocol. Total RNA from the stably-transfected SK-Mel-28 melanoma cell lines were prepared and hybridized onto Human Genome U133 Plus 2.0 Array. The human GeneChip® U133 Plus 2.0 consists of greater than 47,000 transcripts and variants from over 38,500 well characterized human genes (Affymetrix, Santa Clara, CA).

first: Yohan

last: Suryo Rahmanto

first: Louise

middle: L

last: Dunn


title: Expression data from pre natal and post natal CCAM samples and relative controls

first: Agnes

last: Paquet

address: 1550 Fourth Street, RM 545

summary:
This study aims at giving an insight on gene expression in CCAM.
Keywords: disease state analysis

overall_design: A total of 14 microarrays were completed.  Samples were divided into four groups: Fetal CCAM samples (n=4), fetal control samples (n=2), post-natal CCAM samples (n=5), post-natal control samples (n=3)

first: Sam

last: Hawgood

first: Amy

last: Wagner


title: Dendritic Cells Compare the Similarity of Endogenous and Exogenous Antigens

status: Public on Apr 12 2007

city: Houston

first: William

last: Decker

address: 1515 Holcombe Blvd.

department: Stem Cell Transplantation and Cellular Therapy

institute: University of Texas MD Anderson Cancer Center

summary:
Here we demonstrate by the use of extensive controls and stringent statistical analysis that dendritic cells differentially regulate hundreds of different genes based upon sequence similarity of endogenously- and exogenously-loaded antigens and in a T-cell independent fashion.  When endogenously and exogenously-derived antigens are identical, dendritic cells upregulate many different components of the Th-1 response, favoring the priming of CD8+ effectors and promulgating cellular immunity.
Keywords: Loading methodology comparison

overall_design: Dendritic cells were loaded by 1 of 5 different methods, then matured for 48 hours.  Total cellular RNA was then extracted from 9 unloaded DC populations, 10 mRNA-loaded populations, 11 lysate-loaded populations, 7 doubly-loaded (matched) populations, and 7 doubly-loaded (mismatched) populations, a total of 44 different experiments.  Each population was derived from a different normal human donor.  Matched/mismatched refers to the source of the mRNA and lysate used to load the DCs.  Matched signifies that the mRNA and lysate came from the same source.  Mismatched signifies that the mRNA and lysate came from disparate sources.  Gene expression profiles were then determined according to the method by which the DCs had been loaded.  Genes were identified as differentially expressed by DCs doubly-loaded with matched antigens, only if they were significantly different from all of the other four controls.  Significance was defined as Cohen’s |d| > 1.0 (large effect) and q-value (Benjamini-Hochberg false discovery) < 0.01.

first: Will

first: Dongxia

last: Xing

first: Sufang

last: Li

middle: N

last: Robinson

first: Hong

last: Yang

first: David

last: Steiner

first: Krishna

last: Komanduri

first: Jose

last: Engelmayer

first: Elizabeth

last: Shpall


title: Human endometriosis vs normal endometrium study - transcriptional profiling

status: Public on Apr 09 2007

city: San Diego

first: Richard

middle: B

last: Roth

address: 12790 El Camino Real

department: Molecular Medicine

institute: Neurocrine Biosciences, Inc.

summary:
Normal and diseased human tissues were profiled for gene expression using the Affymetrix U133 plus 2.0 array
In total 20 samples were processed
Keywords: Human tissue transcriptional profiling

overall_design: Affymetrix human U133 plus 2.0 array was used to transcriptionally profile both normal and diseased endometrial human tissues


title: S1P mediates key targets associated with survival, proliferation and pluripotency in human embryonic stem cells

status: Public on May 25 2007

city: Sheffield

first: Paul

middle: Roy

last: Heath

state: S. Yorks

address: SITraN, 385a Glossop Road

department: Academic Neurology Unit

country: United Kingdom

institute: University of Sheffield

summary:
Human embryonic stem cells (hESCs) replicate by the process of self-renewal, whilst maintaining their pluripotency.   Understanding  the pathways involved in the regulation of this self-renewal process will assist in developing fully-defined conditions for the proliferation of hESCS required for therapeutic applications. We previously demonstrated a role for Sphingosine-1-phosphate (S1P) in the survival and proliferation of hESCs.  The present study investigates further key signalling pathways and the downstream targets of S1P.
Microarrays were used to examine changes in gene expression invoked by treatment of human embryonic stem cells grown upon different matrices
Keywords: Karyotypically normal  human Esc's

overall_design: Human embryonic stem cells (Sheff 4) were grown upon MEFs. One group were treated with S1P. Each group consisted of three replicates which were hybridised to Human U133 plus 2 Affymetrix GeneChips

first: Katie

last: Inniss

first: Stuart

last: Avery

first: Jill

last: Shepherd

first: Harry

last: Moore


title: Identification of LEDGF-responsive genes in Jurkat cells

status: Public on Apr 13 2007

city: Philadelphia

first: Gary

last: Wang

state: PA

address: 405 Johnson Pavilion, 3610 Hamilton Walk

department: Microbiology

institute: University of Pennsylvania School of Medicine

summary:
The LEDGF transcript from the PSIP1 gene was knocked down in Jurkat cells using RNAi technology. The resulting Jurkat-derived cell line (Jurkat-siJK2) was compared to a control cell line (wild type Jurkat) using microarray analysis. Genes identified as being modulated by LEDGF were preferential targets of HIV integration.
Keywords: effects of gene knockdown

overall_design: 2 cell lines were used, Jurkat-siJK2 (experimental) and wild-type Jurkat (control). 3 independent cultures of each cell line were used for independent RNA extractions, labeling reactions and array hybridizations.

first: Angela

last: Ciuffi

first: Jeremy

last: Leipzig

first: Charles

last: Berry

first: Frederic

middle: D

last: Bushman


title: Transcriptome maps of six different human cell lines

status: Public on Apr 23 2007

city: Amsterdam

first: Roel

last: van Driel

address: Kruislaan 318

department: BioCentrum

country: Netherlands

institute: University of Amsterdam

summary:
The three-dimensional (3D) folding of the chromosomal fibre in the human interphase nucleus is an important, but poorly understood aspect of gene regulation. Especially basic principles of 3D chromatin and chromosome organisation are still elusive. In this paper, we quantitatively analyse the 3D structure of large parts of chromosomes 1 and 11 in the G1 nucleus of human cells and relate it to the human transcriptome map (HTM). Despite a considerable cell-to-cell variation, our results show that subchromosomal domains, which are highly expressed, are more decondensed, have a more irregular shape and are located in the nuclear interior compared to clusters of low expressed genes. These aspects of chromosome structure are shared by six different cell lines and therefore are independent of cell type specific differences in gene expression within the investigated domains. Systematic measurements show that there is little to no intermingling of chromatin from different parts of the same chromosome, indicating that the chromosomal fibre itself is a compact structure. Together, our results reveal several basic aspects of 3D chromosome architecture, which are related to genome function.
Keywords: cell line comparison

overall_design: The data sets presented here together with sample GSM153780, accompany the paper: "The 3D structure of human interphase chromosomes is related to the transcriptome map“ by Goetze et al., (submitted). All cells were grown under standard conditions (see table below) and harvested at 70% confluency. Total RNA was isolated and analyzed on Affymetrix U133 Plus 2.0 to generate transcriptome maps.

first: Sandra

last: Goetze

first: Julio

last: Mateos-Langerak

first: Hinco

last: Gierman

first: Wim

last: de Leeuw

first: Osdilly

last: Giromus

first: Mireille

middle: H

last: Indemans

first: Jan

last: Koster

first: Vladan

last: Ondrej

first: Rogier

last: Versteeg


title: DC-SIGN initiates an immature dendritic cell phenotype triggering Rho activation that is utilised by HIV-1

status: Public on Apr 24 2007

city: Oxford

first: Dilair

last: Baban

address: Roosevelt Drive

department: Wellcome Trust Centre Human Genetics

institute: University of Oxford

summary:
DC-SIGN is a C-type lectin expressed by dendritic cells (DCs) that binds HIV-1, sequestering it within multivesicular bodies to facilitate transmission to CD4+ T cells. Here we characterize the molecular basis of signalling through DC-SIGN by large-scale gene expression profiling and phosphoproteome analysis. Solitary DC-SIGN activation leads to a phenotypically disparate transcriptional program from Toll-like receptor (TLR) triggering with downregulation of MHC II, CD86, and interferon response genes and with induction of the TLR negative regulator ATF3. Phosphoproteome analysis reveals DC-SIGN signals through the leukemia-associated Rho guanine nucleotide exchange factor (LARG) to induce Rho activity. This LARG activation also occurs on DC HIV exposure and is required for effective HIV viral synapse formation. Taken together HIV mediated DC-SIGN signalling provides a mechanism by which HIV evades the immune response yet induces viral spread.
Keywords: Activation state, signalling, Toll-like Receptor (TLR)

overall_design: Circulating monocyte derived DCs were isolated from buffy coats by adherence and culture in IL-4 and granulocyte-macrophage colony-stimulating factor (GM-CSF). DC preparations analyzed were more than 98% pure. At day four 10 million immature DCs were either left unstimulated or stimulated using plate bound anti-DC-SIGN antibody for 2 hr. Three replicates of non-stimulated or stimulated cells were taken and used to extract total RNA.

first: Alison

last: Simmons

first: Ashleigh

last: Hodges

first: Katherine

last: Sharrocks

first: Mariola

last: Batyka

first: Arnaud

last: Moris

first: Olivier

last: Schwartz

first: Kay

last: Davies

first: Benedikt

last: Kessler

first: Andrew

last: McMichael


title: Modulation of gene expression by decitabine in U-2OS cells in vitro and in vivo

status: Public on Apr 05 2007

city: Toronot

first: Khaldoun

last: Al-Romaih

address: 610 University Ave

department: AMO

country: Canada

institute: Ontario Cancer Institute

type:
Expression profiling by array
Methylation profiling by array

summary:
Background:  Epigenetic modifications such as methylation silencing of genes with CpG-island-associated promoters is frequently observed in cancer.  Studies regarding the implications of epigenetic modifications in osteosarcoma (OS) have been limited.  The epigenetic drug decitabine is a potential re-activator of silenced genes through de-methylation, and is currently undergoing clinical trials for cancer treatment.  No study to date has utilized decitabine to modify gene expression in OS-derived cells to identify gene-specific methylation targets that may have therapeutic importance.  The objective of this study was to measure the response of the OS cell line, U-2OS, to decitabine treatment both in vitro and in vivo.
Results: Genome-wide screening was used to distinguish decitabine-dependent changes in gene expression in U-2OS and to identify responsive loci with de-methylated CpG regions.  U-2OS xenografts were established in the sub-renal capsule of immune-deficient mice to study the effect of decitabine in vivo on tumor growth and differentiation.  Genome-wide screening of U-2OS cells in vitro revealed that decitabine treatment at low-dosage (1 µM) caused a significant induction (p<0.0025) in the expression of 88 genes, 13 had a ≥2-fold change, 11 of which had CpG-island-associated promoters.  Interestingly, 6 of these 11 were pro-apoptotic genes.  1 µM decitabine resulted in a significant induction of cell death in U-2OS cells in vitro (p<0.05).  Decitabine treatment also reduced tumor volume size significantly (p<0.05) in U-2OS in vivo at a 2.5 mg/kg dose.  Histological analysis of U-2OS xenograft sections revealed a lower mitotic activity (p<0.0001), a higher bone matrix production (p<0.0001), and higher apoptosis (p = 0.0329) after treatment.    The 6 pro-apoptotic genes were also induced to ≥2-fold in vivo.  Quantitative Methylation Pyrosequencing (Pyro Q-CpG) showed that 4 pro-apoptotic genes had CpG-island DNA de-methylation as a result of treatment in vitro and in vivo.
Conclusion: Our data provide new insights regarding the use of epigenetic modifiers in OS, and have important implications for therapeutic trials involving demethylation drugs.  The findings of this study suggest an epigenetic mechanism, whereby OS tumor cells silence expression of pro-apoptotic genes through promoter-CpG methylation, to maintain high proliferation capacity and continuous growth.
Keywords: Osteosarcoma, epigenetics, methylation, decitabine

overall_design: Total RNA was extracted using the RNeasy kit (Qiagen, Germany) from duplicate experiments of U-2OS cells at Day3 after treatment with 1 µM decitabine or medium alone (control).  In each experiment RNA yields were pooled from two independent cultures per treatment arm to minimize experimental noise.  For each case, 10 µg of RNA was labeled and hybridized to the Affymetrix HG-U133A GeneChips using the manufacturer’s protocol (Affymetrix, Santa Clara, CA) by the Center of Applied Genomics at the Hospital for Sick Children (Toronto, Canada).  Data were extracted using the Microarray Suite (MAS) version 5.0 (Affymetrix) and linearly scaled to achieve an average intensity of 150 across each chip.  The candidate gene list obtained from the MAS 5.0-extracted data was selected by eliminating genes that were not present in at least one experiment.  The arrays were subjected to a pair wise comparison using MAS 5.0, with signal intensities from the no-treatment cells as the baseline.  The statistical significance for the change of expression for each probe set between the decitabine treated and control was calculated by the MAS5.0 software. The criteria for gene selection for real-time expression validation analysis was based on the statistically significant up-regulation (p<0.0025) and fold change of ≥ 2 for expression after decitabine treatment.  The gene list was annotated based on the NetAffx data-base (http://www.affymetrix.com/analysis /index.affx) and further verified using the Human Genome Browser data base (http://genome.ucsc.edu/)

middle: I

last: Al Romaih

first: Mona

last: Prasad

first: Maria

last: Zielenska

first: Jeremy

middle: A

last: Squire


title: Human body index - transcriptional profiling

first: Richard

last: Roth

summary:
Normal and diseased human tissues were profiled for gene expression using the Affymetrix U133 plus 2.0 array

In total 677 samples were processed , representing over 90 distinct tissue types

Some tissue samples were purchased from Stratagene (SG), Ambion (AB), and Becton-Dickinson (BD)
Keywords: Human body index of gene expression

overall_design: Affymetrix human U133 plus 2.0 array was used to transcriptionally profile both normal and diseased human tissues representing over 90 distinct tissue types.


title: Expression profiling of MCF-7 cells with inducible LMO4 and DN-Clim expression

status: Public on Apr 01 2007

city: Irvine

first: Kevin

last: Lin

address: 250 Sprague Hall

department: Biological Chemistry & Medicine

institute: University of California, Irvine

summary:
The nuclear LIM-only protein LMO4 is upregulated in breast cancer, especially estrogen receptor negative tumors, and its overexpression in mice leads to hyperplasia and tumor formation. Here, we show that deletion of LMO4 in the mammary glands of mice leads to impaired lobuloalveolar development due to decreased epithelial cell proliferation. With the goal of discovering potential LMO4-target genes, we also developed a conditional expression system in MCF-7 cells for both LMO4 and a dominant negative (DN) form of its co-regulator, Co-factor of LIM domains (Clim/Ldb/Nli). We then used DNA microarrays to identify genes responsive to LMO4 and DN-Clim upregulation. One of the genes common to both datasets was BMP7, whose expression is also significantly correlated with LMO4 transcript levels in a large dataset of human breast cancers, suggesting that BMP7 is a bona fide target gene of LMO4 in breast cancer. Inhibition of BMP7 partially blocks the effects of LMO4 on apoptosis, indicating that BMP7 mediates at least some functions of LMO4. Gene transfer studies show that LMO4 regulates the BMP7 promoter, and chromatin immunoprecipitation studies show that LMO4 and its co-factor Clim2 are recruited to the BMP7 promoter.  Furthermore, we demonstrate that HDAC2 recruitment to the BMP7 promoter is inhibited by upregulation of LMO4 and that HDAC2 knockdown upregulates the promoter. These studies suggest a novel mechanism of action for LMO4: LMO4, Clim2 and HDAC2 are part of a transcriptional complex, and increased LMO4 levels can disrupt the complex, leading to decreased HDAC2 recruitment and increased promoter activity.
Keywords: Inducible cell lines

overall_design: We used the Tet-off system to establish several distinct MCF-7 cell clones, referred to as MCF7-LMO4-TetOff and MCF7-DN-Clim-TetOff cells, in which the expression of Myc-tagged LMO4 or DN-Clim was repressed by the presence of doxycycline in the medium. Total RNA was isolated from three distinct MCF7-LMO4-TetOff cell lines and three distinct MCF7-DN-Clim-TetOff cell lines in presence and absence of doxycycline after 6 days. Treatment with doxycycline To decrease variability, we pooled RNAs from three experiments for each of the three cell lines.

first: Ning

last: Wang

first: Zhongxian

last: Lu

first: Kaye-Starr

last: Lam

first: Ryan

last: Newton

first: Xiaoman

last: Xu

first: Zhengquan

last: Yu

first: Gordon

last: Gill

first: Bogi

last: Andersen


title: 63 Melanoma cell lines

status: Public on Mar 31 2007

city: Herston

first: Leisl

last: Packer

state: QLD

address: 300 Herston Rd

department: Cancer and Cell Biology

institute: Queensland Institute of Medical Research

summary:
63 melanoma cell lines hybridized to Affymetrix Hu133_Plus 2 oligo arrays. The aim of this study was to identify potential downstream targets of key oncogenes and TSGs in melanoma (including p14ARF, p16INK4A, BRAF etc).
Publications relevant to this series include:
Johansson et al. Pigment Cell Res 2007.
Keywords: melanoma

overall_design: 63 melanoma cell lines hybridized to Affymetrix Hu133_Plus 2 oligo arrays. These cell lines were sequenced for key tumour suppressor genes (TSGs) and oncogenes known to be involved in melanoma development. The microarray data was then analysed together with a particular genotype status (see relevant publications) to identify genes that potentially act downstream of these oncogenes and TSGs to contribute to melanoma development. See publications for analysis methods.

first: Nicholas

last: Hayward

first: Sandra

last: Pavey

middle: M

first: Glen

last: Boyle

first: Peter

middle: G

last: Parsons

first: Mitchell

middle: S

last: Stark


title: AML hybridizations

status: Public on Apr 04 2007

first: Hiroyuki

last: Mano

type: Genome variation profiling by genome tiling array

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

overall_design: Refer to individual Series


title: Expression profiling in monozygotic twins discordant for bipolar disorder

city: Brisbane

last: Matigian

state: Queensland

address: 170 Kessels Rd, Nathan

institute: National Adult Stem Cell Centre

summary:
To identify genes dysregulated in bipolar disorder (BD1) we carried out global gene expression profiling using whole-genome microarrays. To minimize genetic variation in gene expression levels between cases and controls we compared expression profiles in lymphoblastoid cell lines from monozygotic twin pairs discordant for the disease. We identified 82 genes that were differentially expressed by ≥ 1.3-fold in 3 BD1 cases compared to their co-twins, and which were statistically (p ≤ 0.05) differentially expressed between the groups of BD1 cases and controls. Using qRT-PCR we confirmed the differential expression of some of these genes, including: KCNK1, MAL, PFN2, TCF7, PGK1, and PI4KCB, in at least 2 of the twin pairs. In contrast to the findings of a previous study by Kakiuchi and colleagues with similar discordant BD1 twin design1 our data do not support the dysregulation of XBP1 and HSPA5. From pathway and gene ontology analysis we identified up-regulation of the WNT signalling pathway and the biological process of apoptosis. The differentially regulated genes and pathways identified in this study may provide insights into the biology of BD1.
Keywords: Bipolar Disorder, Monozygotic Twins, Discordant, Trasnsformed Lymphocytes

overall_design: Three pairs of Monozygotic Twins which were discordant for Bipolar Disorder with psychotic features were examined. (n=6)

first: Louisa

last: Windus

first: Heather

last: Smith

first: Cheryl

last: Filippich

first: Christos

last: Pantelis

first: John

last: McGrath

first: Bryan

last: Mowry


title: Melanoma cell lines involved in the p14ARF genotype analysis

summary:
35 Melanoma cell lines hybridized to Affymetrix Hu133_Plus 2 microarrays were analysed for genes differentially expressed between cell lines carrying wild-type p14ARF and those with mutant 14ARF. All of these cell lines contained wild-type p53 (so that the effects of p14ARF mutations could be analysed without contamination from p53).
Keywords: Affymetrix Hu133_Plus microarrays

overall_design: The aim of this study was to identify downstream effectors of p14ARF in melanoma. Relevant paper: Packer et al. (2007) Int J Cancer.

middle: K

middle: J


title: Gene-resolution analysis of DNA copy number variation using oligonucleotide expression microarrays

status: Public on Mar 29 2007

city: Columbus

first: David

middle: Lawrence

last: Newsom

address: 700 Children's Drive

department: Microbial Pathogenesis

institute: Nationwide Children's Hospital

summary:
We present a novel method of using commercial oligonucleotide expression microarrays for aCGH, enabling DNA copy number measurements and expression profiles to be combined using the same platform. This method yields aCGH data from genomic DNA without complexity reduction at a median resolution of approximately 17,500 base pairs. Due to the well-defined nature of oligonucleotide probes, DNA amplification and deletion can be defined at the level of individual genes and can easily be combined with gene expression data.
Keywords: genomic DNA, CGH, Copy Number Variation

overall_design: Genomic DNA isolated from peripheral blood samples and from Neuroblastoma cell lines was fragmented by DNAseI and biotin labeled by terminal transferase. Labeled DNAs were hybridized to U133plus2.0 GeneChips (Affymetrix). Hybridization and other conditions were slightly modified from those suggested for 10K Mapping Arrays (Affymetrix) and washing conditions were carried out as suggested for 100K Mapping Arrays. Probe set signals were either generated using the RMA algorithm  or using the in-house developed WPP algorithm. Copy-number variation between Neuroblastoma and normal DNA was calculated.

first: Herbert

last: Auer

first: Norma

last: Nowak

first: M

last: McHugh

first: S

last: Singh

first: C

middle: Y

last: Yu

first: Y

last: Yang

first: G

last: Wenger

first: J

last: Gastier-Foster

first: K

last: Kornacker


title: Expression profiling in early onset colorectal cancer

status: Public on Mar 20 2007

city: Singapore

first: Yi

last: Hong

email: hong.yi@sgh.com.sg, fbap8570@yahoo.com

address: Outram Road

department: Colorectal Surgery

country: Singapore

institute: Singapore General Hospital

summary:
Background: Causative genes for autosomal dominantly inherited familial adenomatous polyposis (FAP) and hereditary non-polyposis colorectal cancer (HNPCC) have been well characterized. There is, however, another 10-15 % early onset colorectal cancer (CRC) whose genetic components are currently unknown.  In this study, we used DNA chip technology to systematically search for genes differentially expressed in early onset CRC.
Keywords: disease state analysis

overall_design: Methods: RNA extracted from colonic mucosa of healthy controls(10samples) and patients(12samples) were analyzed using GeneChip U133-Plus 2.0 Array. Patients and controls were age- (50 or less), ethnicity- (Chinese) and tissue-matched.  T-test, hierarchical clustering, mean fold-change and principal component analysis were used to identify genes that differentiate between patients and controls. These were subsequently verified by real-time polymerase chain reaction (PCR) technology.


title: Expression data from different karyotypic human embryonic stem cells

status: Public on Mar 10 2007

city: changsha

first: sheng

last: yang

address: xiangya Road

country: China

institute: Institute of Human Reproduction & Stem Cell Engineering, Central South University

summary:
Recent reports indicated that during long term culture HESCs acquired cytogenetic abnormality which are similar to those occurred in tumorigenesis. This raise concerns about the safety use of cultured HESCs. The cause of genetical instability in cultured HESCs is poorly understood and whether these changes will progressively developed and ultimately transform HESCs into malignancy has not been systematically investigated. In our lab, chHES3 was found to have progressive chromosomal changes during long term suboptimal culture. In order to understand the molecular mechanism underlying the karyotypic changes, we compared the expression profile of differnt karyotypic chHES-3 cells and signa pathways involved in the process. We found that the expression profiles of oncogenes, suppress tumor genes and genes related with apoptosis had tendency to EC with complexity and compared with the normal karyotypic HES cells, the activating Wnt pathway gene up-regulated and the suppressing gene down-regulated in the aberrant karyotypic HES cells. The species and abundance of the wnt gene family had obvious difference in difference batch mouse embryonic fibroblast feeder cells.
We used microarrays to detail the global programme of gene expression underlying cellular mailignant transformation, identified distinct genes or gene clusters and analysed some signal pathways involved in this process.
Keywords: different karyotype

overall_design: Different karotypic human embryonic stem cells and human embryoinc carcinoma cells were collected for RNA extraction and hybridization on Affymetrix microarrays. By the karyotype analysis we selected three different karyotypic chHES-3 cells and human embryonic carcinoma cells to compared.

first: Yang

last: Sheng

first: Ge

last: Lin

first: Shu-Wei

last: Luo

first: Di

last: Zhou

first: LeiYu

last: Deng

first: TianChen

last: Liu

first: YangQi

last: Ou

first: Jing

first: Ding

last: Yuan

first: PingYuan

last: Xie


title: Cytokine treated normal human epidermal keratinocytes

status: Public on Mar 09 2007

city: South San Francisco

first: Kenneth

last: Jung

address: 1 DNA Way MS93

institute: Genentech

summary:
Normal human epidermal keratinocytes (NHEK) from neonatal foreskin were cultured in serum-free EpiLife medium with human KC growth supplement (0.2% bovine pituitary extract (v/v), 5ug bovine insulin, 5ug/ml bovine transferrin, 0.5ng/ml human EGF, and 0.18 ug/ml hydrocortisone) from Cascade Biologics.  Cultures were treated with recombinant cytokines from R&D Systems.  J Immunol. 2007 Feb 15;178(4):2229-40.
Keywords: cytokine response

overall_design: NHEK were treated with IL-19, IL20, IL-22, and IL24, with controls untreated, along with IL1b, IFN gamma, and KGF.

first: Susan

last: Sa

first: Patricia

last: Valdez

first: Jianfeng

last: Wu

first: Fiona

last: Zhong

first: Linda

last: Hall

first: Ian

last: Kasman

first: Jane

last: Winer

first: Zora

last: Modrusan

first: Dimitry

last: Danilenko

first: Wenjun

last: Ouyang