title: Copy number abnormality, MYC activity and the genetic fingerprint of normal B-cells and the microRNA profile of DLBCL-77

first: Cheng

last: Li

address: 44 Binney St

department: Biostatistics and Computational Biology

institute: Dana-Farber Cancer Institute

type: Genome variation profiling by genome tiling array

summary: A better understanding of the pathogenesis of diffuse large B-cell lymphoma (DLBCL) has clarified its relationship to normal B-cells, pointed to distinct mechanisms of cell transformation, and facilitated the design of novel treatments. However, these data derive from  mRNA genes studies, whereas genome-wide integrative investigations of the role of microRNAs (miRNAs) in DLBCL are still unavailable. We used array-CGH and microarray-based expression analyses to map the abnormalities targeting miRNAs in DLBCLs (n=86). Using training and validation DLBCL cohorts, we defined a collection of miRNAs that robustly segregates these tumors in three unique subsets. This miRNA-driven substructure appears to influence disease outcome, is independent of other mRNA-based DLBCL classifications, and stems from copy number changes targeting the miRNA genome, MYC activity, and the miRNA fingerprints of mature normal B-cells. Our data have uncovered clinically relevant additional molecular complexity in DLBCL and have established a blueprint for the detailed characterizations of miRNAs that are pertinent to B-cell lymphoma biology.

overall_design: Fifty-nine primary DLBCL samples obtained from our Cancer Center tumor bank and 27 cell lines (26 DLBCL and 1 mediastinal B-cell lymphoma) were included in this study for copy number analysis. The clinical and pathological features of this collection are described in the paper. All tumors were reviewed for diagnostic accuracy by a hematopathologist (R.R.) and the categorization in germinal center B-cell-like (GCB) or non-germinal center B-cell-like (non-GCB) was performed according to the algorithm described by Hans et al. The extent of normal cells infiltration in the DLBCL was determined by semi-quantitative measurement of CD3 (T-cells) and CD68 (macrophages) staining. Four normal control hybridizations are also included. Even though we started off with 90 samples total, we inspected the sample quality visually according the log2 ratio curves and excluded 13 samples with noisy or very low signals throughout the genome. The 77 remaining samples were used in the following analyses.

first: Ricardo

last: Aguiar


title: microRNA profile of DLBCL

status: Public on Mar 16 2009

city: Boston

state: MA

country: USA

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

overall_design: Refer to individual Series


title: Gpx3 male transgenic liver expression vs B6/DBA male wildtype control

status: Public on Mar 08 2009

city: Los Angeles

first: Josh

last: Deignan

state: CA

address: 675 Charles E Young Dr South, MRL 3220

department: Cardiology

institute: UCLA

summary:
A major task in dissecting the genetics of complex traits is to identify causal genes for disease phenotypes.  We previously developed a method to infer causal relationships among genes through the integration of DNA variation, gene transcription, and phenotypic information.  Here we validated our method through the characterization of transgenic and knockout mouse models of candidate genes that were predicted to be causal for abdominal obesity.  Perturbation of eight out of the nine genes, with Gas7, Me1 and Gpx3 being novel, resulted in significant changes in obesity related traits.  Liver expression signatures revealed alterations in common metabolic pathways and networks contributing to abdominal obesity and overlapped with a macrophage-enriched metabolic network module that is highly associated with metabolic traits in mice and humans.  Integration of gene expression in the design and analysis of traditional F2 intercross studies allows high confidence prediction of causal genes, and identification of involved pathways and networks.

Keywords: Obesity study in transgenic animals

overall_design: A total of 8 mice were used for this study (4 Gpx3 transgenic and 4 B6/DBA wildtype). The gene expression data is from the livers of 23 week old male wildtype and transgenic mice. These mice were fed a 4% fat chow diet until 11 weeks of age and then were fed a 6% fat chow diet until sacrifice at 23 weeks of age.

first: Xia

last: Yang

first: Hongxiu

last: Qi

first: Jun

last: Zhu

first: Su

last: Qian

first: Gevork

last: Torosyan

first: Sana

last: Majid

first: Brie

last: Falkard

first: Robert

middle: R

last: Kleinhanz

first: Jenny

last: Karlsson

first: Lawrence

middle: W

last: Castellani

first: Sheena

last: Mumick

first: Kai

last: Wang

first: John

last: Lamb

first: Tao

last: Xia

first: Michael

last: Coon

first: Chunsheng

last: Zhang

first: Daria

last: Estrada-Smith

first: Charles

last: Farber

first: Susanna

middle: S

first: Atila

last: Van Nas

first: Anatole

last: Ghazalpour

first: Bin

first: Douglas

middle: J

last: MacNeil

first: Marc

middle: L

last: Reitman

first: Katrina

last: Dipple

first: Margarete

last: Mehrabian

first: Pek

middle: Y

last: Lum

first: Eric

middle: E

last: Shadt

first: Aldons

last: Lusis

first: Thomas

middle: A

last: Drake


title: Lactb male transgenic liver expression vs FVB male wildtype control

first: Josh

last: Deignan

overall_design: A total of 16 mice were used for this study (7 Lactb transgenic and 9 FVB wildtype). The gene expression data is from the livers of 23 week old male wildtype and transgenic mice. These mice were fed a 4% fat chow diet until 11 weeks of age and then were fed a 6% fat chow diet until sacrifice at 23 weeks of age.

first: Xia

last: Yang

first: Hongxiu

last: Qi

first: Jun

last: Zhu

first: Su

last: Qian

first: Gevork

last: Torosyan

first: Sana

last: Majid

first: Brie

last: Falkard

first: Robert

last: Kleinhanz

first: Jenny

last: Karlsson

first: Lawrence

last: Castellani

first: Sheena

last: Mumick

first: Kai

last: Wang

first: John

first: Tao


title: Gyk female heterozygous liver expression vs C57Bl/6J female wildtype control

summary:
A major task in dissecting the genetics of complex traits is to identify causal genes for disease phenotypes.  We previously developed a method to infer causal relationships among genes through the integration of DNA variation, gene transcription, and phenotypic information.  Here we validated our method through the characterization of transgenic and knockout mouse models of candidate genes that were predicted to be causal for abdominal obesity.  Perturbation of eight out of the nine genes, with Gas7, Me1 and Gpx3 being novel, resulted in significant changes in obesity related traits.  Liver expression signatures revealed alterations in common metabolic pathways and networks contributing to abdominal obesity and overlapped with a macrophage-enriched metabolic network module that is highly associated with metabolic traits in mice and humans.  Integration of gene expression in the design and analysis of traditional F2 intercross studies allows high confidence prediction of causal genes, and identification of involved pathways and networks.

Keywords: Obesity study in knockout animals

overall_design: A total of 10 mice were used for this study (5 Gyk heterozygotes and 5 B6 wildtype). The gene expression data is from the livers of 23 week old female wildtype and transgenic mice. These mice were fed a 4% fat chow diet until 11 weeks of age and then were fed a 6% fat chow diet until sacrifice at 23 weeks of age.

last: Lamb

last: Xia

first: Michael

last: Coon

first: Chunsheng

last: Zhang

first: Daria

last: Estrada-Smith

first: Charles

last: Farber

first: Susanna

first: Atila

last: Van Nas

first: Anatole

last: Ghazalpour

first: Bin

first: Douglas

last: MacNeil

first: Marc

last: Reitman

first: Katrina

last: Dipple

first: Margarete

last: Mehrabian

first: Pek

last: Lum

first: Eric

first: Aldons


title: Gas7 male transgenic liver expression vs FVB male wildtype control

first: Josh

last: Deignan

overall_design: A total of 14 mice were used for this study (7 Gas7 transgenic and 7 FVB wildtype). The gene expression data is from the livers of 23 week old male wildtype and transgenic mice. These mice were fed a 4% fat chow diet until 11 weeks of age and then were fed a 6% fat chow diet until sacrifice at 23 weeks of age.

first: Xia

last: Yang

first: Hongxiu

last: Qi

first: Jun

last: Zhu

first: Su

last: Qian

first: Gevork

last: Torosyan

first: Sana

last: Majid

first: Brie

last: Falkard

first: Robert

last: Kleinhanz

first: Jenny

last: Karlsson

first: Lawrence

first: Sheena

first: Kai

first: John

last: Shadt

last: Lusis

first: Thomas

last: Drake


title: Nuclear scaffold and matrix attachment of Aortic Adventitial Fibroblast (AoAF) chromatin

status: Public on Mar 11 2009

city: Detroit

first: Stephen

last: Krawetz

state: MI

address: 275 East Hancock Avenue

institute: Wayne State University

type: Genome binding/occupancy profiling by genome tiling array

summary: The nuclear scaffold/matrix provides an anchor for higher order genome structure that has both structural and functional implications.  Different extraction protocols, i.e., utilizing either 25 mM LIS or 2 M NaCl, isolate somewhat different protein constituents of either the nuclear scaffold or nuclear matrix respectively.  We have mapped, by array CGH, the locations of attachment to each of these residual protein bodies relative to non-attached DNA along the entire length of human chromosomes 14, 15, 16, 17 and 18 in AoAF cells.

overall_design: LIS (lithium 3,5-diiodosalicylate) or 2 M NaCl solutions followed by restriction digestion with EcoR1 facilitates the separation from scaffold/matrix bound DNA from non bound DNA.  Genomic CGH arrays were used to map the relative differences between attached (scaffold/matrix) and non-attached (loop) portions of AoAF DNA. The expression profile of the AoAF cells used for aCGH analysis was determined.

first: Amelia

middle: K

last: Linnemann


title: PPARg dependent expression changes during differentiation of 3T3L1 cell into adipocytes

status: Public on Mar 24 2009

city: Singapore

first: Vinsensius

middle: B

last: Vega

address: 60 Biopolis Street

country: Singapore

institute: Genome Institute of Singapore

summary:
We profiled PPARg dependent gene expression changes during differntiation of 3T3L1 cell using PPARg siRNA
3T3-L1 (Pre-adipocyte) cell line was induced to differentiate using standard adipocyte differentiation media (IBMX, Dex and Insulin) 48hrs post-confluency. RNA was harvested at day -2 (confluent fibroblasts), 48hrs post-induction with IBMX, DEX and Insulin (day=0) and for each subsequent day after rosiglitazone treatment. Illumina beadchip microarrays were used to determine expression profiles of genes differentially regulated in cells transfected with either siRNA targeting PPARgamma or a non-targeting control siRNA.

overall_design: 3T3L1 cell were induced to differentiate into adipocytes using IBMX, DEX and Insulin. RNA from cell treated with PPARg-specific siRNA and non-specific siRNA was isolated at different timepoints. Illumina MouseRef-8 v1.1 Bead chips were used for expression profiling

first: Mohamed

last: Hamza

first: Sebastian

last: Pott

first: Jane

last: Thomsen

first: Gopalan

last: Kandhadayar

first: Kuo

middle: P

last: Chiu

first: Chia

last: Wei

first: Yijun

last: Ruan

first: Edison

middle: T

last: Liu


title: Expression data from human neonatal melanocytes and Notch transformed human neonatal melanocytes

status: Public on Mar 03 2009

city: Philadelphia

first: Gao

last: Zhang

state: PA

address: 3601 SPRUCE ST

department: Molecular and Cellular Oncogenesis Program

institute: Wistar Institute

summary:
Notch1 Activation Confers Transforming Properties to Primary Human Melanocytes and Promotes Human Melanoma Progression
We used microarrays to detail the global programme of gene expression underlying cellularisation and identified distinct classes of up-regulated genes during this process.

overall_design: Human neonatal melanocytes and Notch transformed human neonatal melanocytes were selected for RNA extraction and hybridization on Illumina gene expression array chip. Expression intensities were calculated and normalized for each gene probed on the array for all hybridizations using illumina Beadstudio#3 software. Microarray analyses were subsequently performed in GeneSpring to aid in the identification of genes differentially-expressed between Notch-infected and control melanocytes that may be responsible for the phenotypic changes described in the NIC-infected cells.

first: Pinnix

last: Chelsea

first: Lee

last: John Jr


title: Obesity study in transgenic and knockout animals

first: Josh

last: Deignan

summary:
A major task in dissecting the genetics of complex traits is to identify causal genes for disease phenotypes.  We previously developed a method to infer causal relationships among genes through the integration of DNA variation, gene transcription, and phenotypic information.  Here we validated our method through the characterization of transgenic and knockout mouse models of candidate genes that were predicted to be causal for abdominal obesity.  Perturbation of eight out of the nine genes, with Gas7, Me1 and Gpx3 being novel, resulted in significant changes in obesity related traits.  Liver expression signatures revealed alterations in common metabolic pathways and networks contributing to abdominal obesity and overlapped with a macrophage-enriched metabolic network module that is highly associated with metabolic traits in mice and humans.  Integration of gene expression in the design and analysis of traditional F2 intercross studies allows high confidence prediction of causal genes, and identification of involved pathways and networks.

This SuperSeries is composed of the SubSeries listed below.

first: Xia

last: Yang

first: Hongxiu

last: Qi

first: Jun

last: Zhu

first: Su

last: Qian

first: Gevork

last: Torosyan

first: Sana

last: Majid

first: Brie

last: Falkard

first: Robert

last: Kleinhanz

first: Jenny

last: Karlsson

first: Lawrence

last: Castellani

first: Sheena

last: Mumick

first: Kai

last: Wang

last: Lamb

first: Tao

last: Xia

first: Michael

last: Coon

first: Chunsheng

first: Daria

last: Estrada-Smith

first: Charles

last: Farber

first: Susanna

first: Atila

last: Van Nas

first: Anatole

last: Ghazalpour

first: Bin

first: Douglas

last: MacNeil

first: Marc

last: Reitman

first: Katrina

last: Dipple

first: Margarete

last: Mehrabian

first: Pek

last: Lum

first: Eric

last: Shadt

first: Aldons


title: Gene expression profiling of colorectal cancers to understand molecular changes and clinicopathological features

status: Public on Mar 01 2009

city: Daejon

first: Seon-Young

last: Kim

address: 52 Eoeun-dong, Yuseong-gu

department: Genome Research Center

country: South Korea

institute: Korea Research Institute of Bioscience and Biotechnology

summary:
Gene expression was analyzed in terms of canonical molecular changes and clinicopathological features to elucidate alternative or subordinate pathways during colorectal tumorigenesis and tumor growth.
Eighty-four sporadic colorectal cancer patients, standardized by tumor location, were consecutively enrolled. Representative molecular changes including APC, TP53, Wnt, RAF, and mismatch repair defect (MMR) were recorded for each sample.
Keywords: disease state analysis; sub-type analysis within colorectal cancers

overall_design: 84 samples from colorectal patients were analyzed. Paired tumor and adjacent normal tissues from the same patient were used for hybridization onto custom-made, 21k dual channel cDNA arrays. We prepared a similar number of samples from each of the three tumor locations (ascending 27, descending 29, and rectum 28) and recorded for each sample important molecular changes such as APC, TP53, RAF, WNT, and MMR mutations.

first: Seon-Youg

first: Yong

first: Jin

middle: C


title: Subclassification of lung squamous cell carcinoma

status: Public on Apr 01 2009

city: Koto-ku

first: Yuichi

last: Ishikawa

state: Tokyo

address: 3-8-31 Ariake

country: Japan

institute: Cancer Insitute

summary:
Two subclasses of lung squamous cell carcinoma with different gene expression profiles and prognosis identified by hierarchical clustering and validated by non-negative matrix factorization .

BACKGROUND: Current clinical and histopathological criteria used to define lung squamous cell carcinomas (SCCs) are insufficient to predict clinical outcome. We attempted to make a clinically-useful classification based on gene expression profiling. METHODS: We used cDNA microarrays with 40386 elements to analyze the gene expression profiles of 48 surgically resected samples of lung SCC. 9 samples of lung adenocarcinoma and 30 of normal lung were also included to give a total of 87 samples analyzed. After gene filtering, the data were subjected to hierarchical clustering and consensus clustering with the non-negative matrix factorization (NMF) approach. FINDINGS: Initial analysis by hierarchical clustering allowed division of SCCs into two distinct subclasses. An additional independent round of hierarchical clustering and consensus clustering with the NMF approach provided a validation for the classification. Kaplan-Meier analysis with the log rank test pointed to a non-significant difference in survival (p=0.071) but the likelihood of survival to 6 years was significantly different between the two groups (40.5% vs 81.8%, p=0.014, Z-test). Biological process categories characteristic for each subclass were identified statistically and up-regulation of cell-proliferation related genes was evident in the subclass with a poor prognosis. In the subclass with the better survival, genes involved in differentiated intracellular functions, such as the MAPKKK cascade, ceramide metabolism, or regulation of transcription, were up-regulated.
Keywords: repeat sample

first: Michael

middle: H

last: Jones

first: Carl

last: Virtanen


title: Hela Mitotic Selection - gene expression time series in early G1 phase of the human cell cycle

status: Public on Feb 26 2009

city: Hanover

last: Whitfield

state: NH

address: 7400 Remsen

department: Genetics

institute: Dartmouth Medical School

summary:
HeLa mitotic cells were collected using a mitotic shakeoff apparatus to study gene expression in early G1 phase of the human cell cycle. This set includes microarray data from two shake-off experiments : shake 1 (0h-14h) which was conducted over  14 hours and RNA samples collected every 2 hrs, and shake 2 (0min-120min) which was conducted over 2 hours and samples collected every 15 minutes. Total RNA was prepared using ULTRASPEC RNA isolation system, and Reference RNA was extracted from asynchronously growing HeLa cells using TRIzol. For cDNA synthesis and microarray hybridization, refer to Whitfield ML. et al., Mol Biol Cell, 2002. 13(6): p. 1977-2000.
Groups of assays that are related as part of a time series.
Age: g1 progression after mitotic shake-off
Replicate: two biological replicates (shake1 and shake2)

Keywords: time_series_design

overall_design: Computed


title: The impact of copy number variation on local gene expression in mouse hematopoietic stem and progenitor cells

status: Public on Mar 09 2009

city: Baltimore

first: Patrick

last: Cahan

state: MD

address: 733 North Broadway, MRB 653

department: ICE and Biomedical Engineering

institute: Johns Hopkins University School of Medicine

summary: The extent to which differences in germ line DNA copy number contribute to natural phenotypic variation is unknown. We analyzed the copy number content of the mouse genome to a sub-10 kb resolution. We identified over 1,300 copy number variant regions (CNVRs), most of which are < 10 kb in length, are found in more than one strain, and, in total, span 3.2% (85 Mb) of the genome. To assess the potential functional impact of copy number variation, we mapped expression profiles of purified hematopoietic stem and progenitor cells, adipose tissue and hypothalamus to CNVRs in cis. Of the more than 600 significant associations between CNVRs and expression profiles, most map to CNVRs outside of the transcribed regions of genes. In hematopoietic stem/progenitor cells, up to 28% of strain-dependent expression variation is associated with copy number variation, supporting the role of germ line CNVs as major contributors to natural phenotypic variation in the laboratory mouse.

overall_design: To map the CNV content of the mouse genome, we selected 17 Tier 1-3 Mouse Phenome Project strains and three additional strains of biomedical interest, representing all major inbred lineages.  We performed comparative genomic hybridization using a long-oligonucleotide array containing 2,149,887 probes evenly spaced across the reference genome with a median inter-probe spacing of 1,015 bases.  Labeling, hybridization, washing and array imaging were performed as previously described (PMID:16075461).  We performed segmentation using wuHMM, a Hidden Markov Model algorithm that utilizes sequence-level information and can detect CNVs less than 5 kb in length (fewer than five probes) at a low false positive rate (PMID:18334530).  To estimate the overall impact of CNV on gene expression in vivo, we performed expression profiling of hematopoietic stem/progenitors cells using the Illumina Mouse Beadchip-6v1 platform. See manuscript for further details.

first: Yedda

last: Li

first: Masayo

last: Izumi

first: Timothy

last: Graubert


title: Circulating Monocytes in HIV-1 infected Viremic Subjects Exhibit an Anti-Apoptotic Gene Signature

city: West Point

last: Nebozhyn

address: Mail Stop WP53B-120, 770 Sumneytown Pike, Building 53, P.O. Box 4

department: GpGx/Computational System biology

institute: Merck, Inc

summary:
Mechanisms that may allow circulating monocytes to persist as CD4 T cells diminish in HIV-1 infection have not been investigated. We have characterized steady-state gene expression signatures in circulating monocytes from HIV-infected subjects and have identified a stable anti-apoptosis gene signature comprised of 38 genes associated with p53, CD40L, TNF and MAPKinase signaling networks. The significance of this gene signature is indicated by our demonstration of cadmium chloride- or Fas ligand-induced apoptosis resistance in circulating monocytes in contrast to increasing apoptosis in CD4 T cells from the same infected subjects. As potential mechanisms in vivo, we show that monocyte CCR5 binding by HIV-1 virus or agonist chemokines serve as independent viral and host modulators resulting in increased monocyte apoptosis resistance in vitro. We also show evidence for concordance between circulating monocyte apoptosis-related gene expression in HIV-1 infection in vivo and available datasets following viral infection or envelope exposure in monocyte derived macrophages in vitro. The identification of in vivo gene expression associated with monocyte resistance to apoptosis is of relevance to AIDS pathogenesis since it would contribute to: (1) maintaining viability of infection targets and long-term reservoirs of HIV-1 infection in the monocyte/macrophage populations, and (2) protecting a cell subset critical to host survival in spite of sustained high viral replication.

Keywords: two group study design

overall_design: 33 samples hybridized, including 13 HIV-1 Patients, 12 Healthy Controls and 4 HIV-1 Patients and 4 Controls followed 6 months later

first: Malavika

last: Giri

first: Andrea

last: Raymond

first: Bethsebah

last: Gekonge

first: Aidan

last: Hancock

first: Shenoa

last: Creer

first: Calen

last: Nicols

first: Malik

last: Yousef

last: Foulkes

first: Karam

last: Mounzer

first: Jane

last: Shull

first: Guido

last: Silvestri

first: Jay

last: Kostman

first: Ronald

middle: G

last: Collman

first: Louise

last: Showe

first: Luis

last: Montaner


title: Nuclear scaffold and matrix attachment of HeLa chromatin

status: Public on Feb 27 2009

first: Stephen

last: Krawetz

address: 275 East Hancock, CS Mott Center

summary: The nuclear scaffold/matrix provides an anchor for higher order genome structure that has both structural and functional implications.  Different extraction protocols, i.e., utilizing either 25 mM LIS or 2 M NaCl, isolate somewhat different protein constituents of either the nuclear scaffold or nuclear matrix respectively.  We have mapped, by array CGH, the locations of attachment to each of these residual protein bodies relative to non-attached DNA along the entire length of human chromosomes 14, 15, 16, 17 and 18 in HeLa cells.

overall_design: LIS or 2 M NaCl solutions followed by restriction digestion with EcoR1 facilitates the separation from scaffold/matrix bound DNA from non bound DNA.  Genomic CGH arrays were used to map the relative differences between attached (scaffold/matrix) and non-attached (loop) portions of HeLa DNA. The expression profile of the HeLa cells used for aCGH analysis was also determined.

first: Amelia

last: Linnemann

first: Adrian

last: Platts


title: Characterisation of LAP/LIP transcriptional signature in human hepatoma cells

city: ROUEN

first: Jean

middle: Philippe

last: SALIER

address: Bvd Gambetta

department: Inserm Unit 519

country: France

institute: Faculty of Medicine and Pharmacy

summary: BACKGROUND & AIMS: C/EBPbeta is involved in numerous process as carcinogenesis but its role is still not clear due to the existence of an active form (LAP) and an inhibitory form (LIP) of this transcription factor. The main goals of the present research were (i) the identification of genes inversely regulated by LAP and LIP i-e the genuine C/EBPbeta molecular signature in the Hep3B human hepatoma cell line (ii) a better understanding of LAP and LIP respective role in hepatic cells survival and proliferation (iii) the search of the C/EBPbeta signature among hepatocellular carcinomas.  METHODS: Using Tet-off expression system we engineered Hep3BLAP and Hep3BLIP cells, in which LAP and LIP were over-expressed respectively. Then, using both expression profiling (DNA arrays) and ChIP-on-chip analysis, we identified genes inversely and/or directly regulated by each of the C/EBPbeta isoforms. The expression levels of these genes regulated by LAP/LIP were compared in controls and HCCs patients.   RESULTS: We identified 676 genes inversely regulated by LAP and LIP and among these, 45 are direct targets. Using functional studies, we displayed the opposite role of LAP and LIP in staurosporine-induced cell death and the implication of LAP in the repression of Hep3B cells proliferation. Finally we identified a subgroup of HCCs with a deregulation of 165 genes belonging to C/EBPbeta signature and coding for proteins involved in chemoresistance and metastasis formation.  CONCLUSIONS: Our study increases knowledge on LAP and LIP functions and provides first evidence that their molecular signature in the HCCs could predict tumor evolution.

overall_design: Total genomic DNA were extracted from 3 Hep3BLAP expressing LAP and were labelled Cy3 fluorochrome. Genomic DNA were extracted from 3 Hep3BLAP expressing LAP, were immunoprecipited with anti-CEBPbeta antibody  and were labelled with Cy5 fluorochrome. Each sample was hybridized on an Agilent two-color microarray G4489A (Human Promoter ChIP-on-Chip Set 244K).

first: Gaëlle

last: Saint-Auret

first: Jean-Louis

last: Danan

first: Xavier

last: Gidrol

first: Martine

last: Hiron

first: Céline

last: Blache

first: Simon

last: Tendil

first: Maryvonne

last: Daveau

first: Jean-philippe

last: Salier


title: Expression profiling of p53-regulated genes by cDNA microarray analysis

status: Public on Feb 24 2009

city: Tokyo

first: Hidewaki

last: Nakaagwa

address: 4-6-1 Shirokanedai, Minato-ku,

department: Institute of Medical Science

institute: The University of Tokyo

summary:
To characterize the molecular features of p53 signaling pathway, we generated the  gene-expression profiles of  p53-introduced U373MG cells by genome-wide cDNA microarrays

Keywords: time course

overall_design: U373MG cells were infected with 20 MOI of Ad-p53 or Ad-LacZ. mRNA from infected cells were collected at 0h, 6h, 12h, 24h and 48h.  we used the RNA mixtures from Ad-LacZ infected cells as a reference of cDNA microarray analysis.

first: Koichi

last: Matsuda

first: Yusuke

last: Nakamura


title: The protein kinase Tor1 regulates adhesin expression in Candida albicans

status: Public on Feb 21 2009

city: Durham

first: Robert

last: Bastidas

address: 322 CARL, Research Drive Box 3546

department: Molecular Genetics and Microbiology

institute: Duke University

summary: Eukaryotic cell growth is coordinated in response to nutrient availability, growth factors, and environmental stimuli, enabling cell–cell interactions that promote survival. The rapamycin-sensitive Tor1 protein kinase, which is conserved from yeasts to humans, participates in a signaling pathway central to cellular nutrient responses. To gain insight into Tor-mediated processes in human fungal pathogens, we have characterized Tor signaling in Candida albicans. Global transcriptional profiling revealed evolutionarily conserved roles for Tor1 in regulating the expression of genes involved in nitrogen starvation responses and ribosome biogenesis. Interestingly, we found that in C. albicans Tor1 plays a novel role in regulating the expression of several cell wall and hyphal specific genes, including adhesins and their transcriptional repressors Nrg1 and Tup1. In accord with this transcriptional profile, rapamycin induced extensive cellular aggregation in an adhesin-dependent fashion. Moreover, adhesin gene induction and cellular aggregation of rapamycin-treated cells were strongly dependent on the transactivators Bcr1 and Efg1. These findings support models in which Tor1 negatively controls cellular adhesion by governing the activities of Bcr1 and Efg1. Taken together, these results provide evidence that Tor1-mediated cellular adhesion might be broadly conserved among eukaryotic organisms.

overall_design: Study consists of wild type Candida albicans cells grown in Spider liquid medium at 35C and treated either with vehicle or 20 nM rapamycin.  Analysis is based off 4 independent biological replicates

first: Joseph

last: Heitman

first: Maria

last: Cardenas


title: ETS2, ERG promote megakaryopoiesis, synergize with alterations in GATA1 to immortalize hematopoietic progenitor cells

status: Public on Feb 16 2009

city: Chicago

first: monika

last: stankiewicz

state: IL

address: 303 E. Superior St. Lurie Bldg 5-250

department: Hematology/Oncology

institute: Northwestern University

summary: Serial replated GFP + lineage negative cells from Gata1deltaneodeltaHS fetal liver hematopoietic progenitors for 4 generations in methocult M3234 under megakaryocyte promoting conditions, IL3, TPO, IL6, and IL11. Cells were subject to retro-viral infection with MIGR1GFP, MIGR1ETS2, MIGR1ERG, and MIGR1FLI1.

sample_organism:
Homo sapiens
Mus musculus

overall_design: Comparison of gene expression profile for GFP + immortalized MIGR1-GFP Gata1deltaneodeltaHS fetal liver progenitors to that of GFP+ Gata1deltaneodeltaHS MIGR1-ETS2, MIGR1-ERG, and MIGR1FLI1 fetal liver progenitors


title: Hepatocellular carcinoma

status: Public on Feb 13 2009

city: Daejeon

first: Bu-Yeo

last: Kim

address: 1672 Yuseongdae-ro, Yuseong-gu

department: Herbal Medicine Research Division

institute: Korea Institute of Oriental Medicine

summary:
Transcription profiling of pair-matched tumor and adjacent liver tissues from 56 HCC patients were performed using cDNA microarray.

Keywords: Disease state analysis

overall_design: Common reference. One replicate per array.

first: Kyung-Suk

last: Suh

first: Je-Geun

last: Lee

first: Sun-Hoo

last: Park

first: Chul-Ju

last: Han

first: Sang-Bum

first: Sook-Hyang

last: Jeong

first: Young-Il

last: Yeom

first: Chang-Min

first: Su-Jin

last: Cho

first: Ja-June

last: Jang

first: Dong-Wook

last: Choi

first: Kee-Ho


title: Salmonella typhimurium infection of zebrafish embryo

status: Public on Mar 02 2009

city: Leiden

first: Anna

middle: Magdalena

last: Zakrzewska

address: Wassenaarseweg 64

department: IBL

country: Netherlands

institute: University of Leiden

summary: Characterization of the zebrafish embryonic host response to systemic bacterial infection with Salmonella typhimurium wild type strain (SL1027) and its isogenic LPS O-antigen mutant Ra (SF1592) by means of a time-resolved global expression analysis.

overall_design: All infection experiments were performed using mixed egg clutches from three tanks of AB strain zebrafish. Embryos were staged at 27 hours post fertilization (hpf) by morphological criteria (Kimmel et al., 1995) and approximately 250 cfu of DsRed expressing S. typhimurium wt and Ra mutant bacteria were injected into the caudal vein close to the urogenital opening. As a control an equal volume of PBS was likewise injected. Injections were controlled using a Leica MZ Fluo 3 stereomicroscope with epifluorescence attachment together with a Femtojet microinjector (Eppendorf) and a micromanipulator with pulled microcapillary pipettes. Pools of 20-40 embryos were collected at 2, 5, 8 and 24 hours post infection (hpi). For the microarray analysis, the whole infection procedure was preformed in triplicate on separate days. The triplicates are marked A,B and C. The order of injecting wt bacteria, Ra bacteria and PBS control was randomized in the different experiments. The general reference sample is a mixture of all RNA samples from this infection study and it is named Common Reference (CR).

first: Oliver

last: Stockhammer

first: Zoltan

last: Hegedus

first: Herman

last: Spaink

first: Annemarie

last: Meijer


title: Metastatic breast cancer to the brain: set 2

status: Public on Feb 11 2009

city: Brisbane

first: Leonard

last: Da Silva

address: Herston Road

country: Australia

institute: University of Queensland

summary:
Background:
Metastases to the brain from breast cancer have a high mortality. Basal-like and HER2 positive breast cancers appear to have a high propensity to spread to the brain. The mechanisms that allow cells to colonise the brain are unclear.
Methods:
We have analysed matched and unpaired samples of breast cancer and brain metastases using morphology, immunophenotype and expression profiling and validated the data using in vitro cell culturing models and in vivo mice model.
Results:
Most of the brain metastases were triple negative and had a basal-like phenotype. Metastatic cells to the brain over-expressed HER3 compared to primaries and showed evidence of higher activation of the MAPK pathways. Inhibition using anti-neuregulin antibody, Herceptin and Lapatinib reduced tumour growth in vitro and in vivo.
Conclusions:
The data demonstrate activation of neuregulin-HER3 pathway in brain metastases from breast cancer and suggest that even in absence of HER2 amplification (as with triple negative and basal cancers), anti-epidermal growth factor receptor family inhibitors may have a role in treating these patients.

overall_design:
An initial set of 29 matched pairs of primary breast cancer and their brain metastases and 22 unmatched brain metastases of from breast cancer was were available as formalin fixed-paraffin embedded (FFPE) tumor blocks. A second set of validation samples comprisinged of 10 matched pairs of primary breast cancer and their corresponding brain metastases and a further 11 non-breast metastatic brain cancers tumours fromn non-breast sites (one melanoma, six lung adenocarcinomas, one prostate carcinoma, one colon carcinoma  and two renal cell carcinomas was also studied. After filtering DASL data on the first set of samples, 15/29 matched pairs and 22/22 unmatched metastases remained available for analysis for the initial set. In the second set all samples were analysed.

RNA extraction and Real-Time RT-PCR
Three 5μm thick FFPE sections of clinical samples, parental cell line MB-MDA-231 cell line (fixed in paraformoldehyde and embedded in paraffin) and intracranial mouse tumours were cut and de-waxed in two lots of xylene and two lots of 100% ethanol, 10 minutes each. RNA was extracted using the High Pure RNA Paraffin Kit® (Roche Applied Science, Mannheim, Germany) according to the manufacturer’s protocol. RNA was quantified using the NanoDrop® ND-1000 (NanoDrop Technologies, DE, USA). RT-PCR for HER3 and housekeeping genes GAPDH and RPL13a was carried out using the Rotor-gene system (Corbett Life Science, Australia). Complementary DNA (cDNA) was synthesized from 240ng of total RNA using oligodT16 primers and the SuperScriptTM III Reverse Transcriptase kit (Invitrogen, Carsbald, CA, USA). A standard curve for gene expression of HER3 and GAPDH and RPL13a was constructed using total RNA from a human breast cancer cell line (SKBR-3). Replicates of 10-fold dilutions of RNA in water were made to give 3 different calibrators, each with a known amount of SKBR-3 RNA for which arbitrary copy number were given. The PCR conditions were: 50°C for 2 minutes, 95°C for 2 minutes, followed by 40 cycles with the following profile: denaturation at 95°C for 20 seconds, annealing at 60°C for 15 seconds, extension at 72°C for 20 seconds. Flourescent incorporation was acquired in the green channel during the 72°C incubations. Intron spanning primer sequences were: GAPDH (5’CTGCACCACCAACTGCTTAG3’, 5’GTCTTCTGGGTGGCA GTGAT3’), RPL13a (5’GTTGGTGTTCATCCGTTG3’, 5’GTACGCTGTGAAGCATCAA3’), HER3 (5’TGGTTCATGGGCATGTATCC3’, 5’CTAGACCTAAGACTTGGAGC3’).

DASL gene expression profilingGene expression profiling was performed using the DASL assay (cDNA-mediated annealing, selection extension and ligation, Illumina Inc., California, USA) interrogating the DASL Cancer Panel that contained 512 cancer related genes. The first and second sets of samples were analyzed in two independent experiments. Tumours arising from the animal experiment were analyzed in conjunction with the second set of clinical samples. All protocols were as specified by Illumina Inc. Briefly, RNA (250ng) was converted to cDNA through a reverse transcription reaction with biotinylated oligo-d(T)18 and random nonamers. Oligonucleotides (three unique pairs for each of the 512 genes) were annealed to the biotinylated cDNA, the duplexes were bound to streptavidin conjugated paramagnetic particles to remove non-hybridized oligos. The annealed oligos were then extended and ligated (to incorporate an address sequence and primer site) to generate amplifiable products. These products were subjected to PCR amplification using fluorescently labeled (Cy3 and Cy5) primers. The labeled PCR products were hybridized to a Sentrix array or Beadchip. Following hybridization, the arrays were scanned with a BeadArray Reader (Illumina) and data was extracted using BeadStudio software (Illumina). Samples exhibiting a median hybridization intensity across all probes of <600 (background corrected) were excluded from analysis. Probes with a BeadStudio detection score greater than 0.99 in more than 15 conditions were included. Array data transformation and normalization (per chip normalized to 50th percentile and per gene normalized to median) was done in Genespring® version 7.0 software (Agilent Technologies, Santa Clara, USA). Hierarchical clustering was performed using: Similarity measure; Pearson Correlation and clustering algorithm; Average Linkage. Genes differentially expressed between primary tumors and brain metastases were identified using analysis of variance (ANOVA, p<0.01) or parametric test, variances not assumed equal (Welch t-test, p < 0.005). Up and down regulation was considered for normalized data values that were greater or less than a factor of 2 fold for the conditions compared.

first: Peter

last: Simpson


title: Renal Cell Carcinoma: Hypoxia and Endocytosis

status: Public on Feb 10 2009

city: Grand Rapids

first: Karl

last: Dykema

address: 333 Bostwick Ave. NE

institute: Van Andel Institute

summary:
Hypoxia signature in Clear cell RCC

A set of clear cell renal cell carcinomas were examined for the deregulation of hypoxia inducible and endocytosis regulated genes

overall_design:
Tumor samples and Normal renal tissue

GSE14762.rda contains an R data object

first: Kyle

last: Furge


title: Metastatic breast cancer to the brain: implications of the local microenvironment for colonization and treatment

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

overall_design: Refer to individual Series


title: Murine Spleen: Wild type vs Brx haploinsufficiency

status: Public on Feb 09 2009

city: Bethesda

first: Tomoshige

last: Kino

address: 10 Center Drive MSC 1109

department: Program in Reproductive and Adult Endocrinology

institute: National Institute of Child Health & Human Development

summary:
Comaprison of the gene expression profiles of the spleen between wild type and Brx haploinsuffieint mice

Nuclear factor of activated T-cells 5 (NFAT5) is a transcription factor that regulates hyperosmolarity-responsive genes and helps activated T lymphocytes adapt to and discharge their functions in hyperosmolar environments. We report here that the Rho-type guanine nucleotide exchange factor (GEF) Brx is essential for increased NFAT5 expression in response to osmotic stress in lymphoid tissues. Indeed, brx haploinsufficient mice expressed significantly less NFAT5 in their spleens than wild type controls and their splenocytes had a defective response to osmotic stress in vitro. Haploinsufficient mice also had smaller-sized spleens containing fewer splenocytes, as well as a defective immunoglobulin response to ovalbumin compared to wild type mice. The Brx GEF domain and the p38 mitogen-activated kinase (MAPK) cascade were both required for osmotic stress-mediated induction of NFAT5 in Jurkat cells. Brx physically interacted with the cJun kinase (JNK)-interacting protein (JIP) 4, a scaffold protein for activation of the p38 MAPK cascade that was required for osmotic stress-induced NFAT5 expression. Thus, Brx is a signal integrator for the adaptive response to osmotic stress in the immune system, activating small G proteins, attracting JIP4 and stimulating p38 MAPK, ultimately increasing the expression of NFAT5 and activating hyperosmolarity protective genes, a phenomenon crucial for proper immune function in hyperosmolar environments, such as inflammatory sites and immune organs.
Keywords: Genetic modification

overall_design: Two-condition experiment: Wild type and Brx haploinsuffient, 3 replicates, labeling swap for each

first: Yan

last: Su

first: Manoli

last: Irini

first: Yonghong

last: Wang


title: The use of zebrafish reward mutants to search for novel transcripts mediating the behavioral effects of amphetamine

status: Public on Jan 30 2009

city: Neuherberg

first: Katharine

last: Webb

address: Ingolstaedterlandstr. 1

department: Institute for developmental genetics

country: Germany

institute: Helmholtz Zentrum Muenchen

summary:
Addiction is a pathological dysregulation of brain reward systems, determined by several complex genetic pathways.  The conditioned place preference (CPP) test provides an evaluation of the effects of drugs in animal models, allowing the investigation of substances at a biologically relevant level with respect to reward.  Our lab has previously reported the development of a reliable CPP paradigm for zebrafish.  Here, this CPP test was used to isolate a dominant ENU-induced mutant, no addiction (naddne3256), which fails to respond to amphetamine, and which we used as an entry point towards identifying the behaviorally- relevant transcriptional response to amphetamine. Using microarray technology, we compare the wildtype with the mutant response to the drug, we compare the transcriptomes of the wildtype and mutants without drug and we also compared the transcriptomes of wildtype fish with the drug and wildtype fish without the drug.

Through the combination of microarray experiments comparing the adult brain transcriptome of mutant and wild-type siblings under normal conditions, as well as their response to amphetamine, we identified genes that correlate with the mutants’ altered CPP behavior. In addition to pathways classically involved in reward, this gene set shows a striking enrichment in transcription factor-encoding genes classically involved in brain development, which later appear to be re-used within the adult brain.  We selected a subset of them for validation by quantitative PCR and in situ hybridization, revealing that specific brain areas responding to the drug through these transcription factors include domains of ongoing adult neurogenesis.  Finally, network construction revealed functional connections between several of these genes. Together, our results identify a new network of coordinated gene regulation that influences or accompanies amphetamine-triggered CPP behavior and that may underlie the susceptibility to addiction.

overall_design: Overall design includes three separate comparisons.  Firstly, we took newly identified nad/+ mutants and their wildtype siblings.  Both groups have had amphetamine administered.  Secondly, we took nad/+ mutants and their wildtype siblings and compared them without drug.  In a third experiment, we compared wildtype with drug vs. wildtype fish without drug.  We pooled 4-5 brains per biological replicate and had in total 3 biological replicates per treatment.  We used one-color arrays and compared the control and reference samples.

first: William

last: Norton

first: Laure

last: Bally-Cuif


title: Gene Expression in Castration-Resistant C4-2 Cells after AR Knockdown

status: Public on Feb 20 2009

city: Vancouver

first: Helen

middle: SW

last: Cheng

state: BC

address: 2660 Oak Street

country: Canada

institute: The Prostate Centre at VGH

summary: Prostate cancer is the most common, lethal malignancy in men. Although androgen withdrawal therapies are used to treat advanced disease, progression to a castration-resistant, end-stage is the usual outcome. In this study, the tested hypothesis was that the androgen receptor remains essential for the growth and viability of castration-resistant disease. Knocking down the androgen receptor in well-established tumors grown in castrated mice caused growth arrest, decreased serum PSA, and frequently regression and total eradication of tumors. Growth control of castration-resistant tumors appeared to be linked to the extent of androgen receptor knockdown, which triggers upregulation of many genes involved in apoptosis, cell cycle arrest, and inhibition of tumorigenesis and protein synthesis. Our findings provide proof of principle that in vivo knockdown of the androgen receptor is a viable therapeutic strategy to control and possibly eradicate prostate cancers that have progressed to the lethal castration-resistant state.

overall_design: C4-2 human prostate cancer cells stably expressing a tetracycline-inducible AR-targeted short hairpin RNA (shRNA) or scrambled shRNA were generated. These two cell lines were incubated in the absence of androgens with or without doxycycline hyclase (DOX). Comparison analysis of the gene expression profiles of C4-2 cells stably expressing AR shRNA + DOX and control cells (AR shRNA - DOX and scrambled shRNA ± DOX) was conducted to identify differentially regulated genes due to AR knockdown after normalization and data filtering. Genes were considered to be significantly different if the expression in the induced AR shRNA + DOX cells was at least 1.7-fold greater or 1.7-fold less than that seen in the control cells, with P< 0.05.

first: Robert

last: Snoek

first: Katia

last: Margiotti

first: Colleen

last: Nelson

first: Paul

last: Rennie


title: Metastatic breast cancer to the brain: set 1

summary:
Background:
Metastases to the brain from breast cancer have a high mortality. Basal-like and HER2 positive breast cancers appear to have a high propensity to spread to the brain. The mechanisms that allow cells to colonise the brain are unclear.
Methods:
We have analysed matched and unpaired samples of breast cancer and brain metastases using morphology, immunophenotype and expression profiling and validated the data using in vitro cell culturing models and in vivo mice model.
Results:
Most of the brain metastases were triple negative and had a basal-like phenotype. Metastatic cells to the brain over-expressed HER3 compared to primaries and showed evidence of higher activation of the MAPK pathways. Inhibition using anti-neuregulin antibody, Herceptin and Lapatinib reduced tumour growth in vitro and in vivo.
Conclusions:
The data demonstrate activation of neuregulin-HER3 pathway in brain metastases from breast cancer and suggest that even in absence of HER2 amplification (as with triple negative and basal cancers), anti-epidermal growth factor receptor family inhibitors may have a role in treating these patients

overall_design:
An initial set of 29 matched pairs of primary breast cancer and their brain metastases and 22 unmatched brain metastases of from breast cancer was were available as formalin fixed-paraffin embedded (FFPE) tumor blocks. A second set of validation samples comprisinged of 10 matched pairs of primary breast cancer and their corresponding brain metastases and a further 11 non-breast metastatic brain cancers tumours fromn non-breast sites (one melanoma, six lung adenocarcinomas, one prostate carcinoma, one colon carcinoma  and two renal cell carcinomas was also studied. After filtering DASL data on the first set of samples, 15/29 matched pairs and 22/22 unmatched metastases remained available for analysis for the initial set. In the second set all samples were analysed.
RNA extraction and Real-Time RT-PCR
Three 5μm thick FFPE sections of clinical samples, parental cell line MB-MDA-231 cell line (fixed in paraformoldehyde and embedded in paraffin) and intracranial mouse tumours were cut and de-waxed in two lots of xylene and two lots of 100% ethanol, 10 minutes each. RNA was extracted using the High Pure RNA Paraffin Kit® (Roche Applied Science, Mannheim, Germany) according to the manufacturer’s protocol. RNA was quantified using the NanoDrop® ND-1000 (NanoDrop Technologies, DE, USA). RT-PCR for HER3 and housekeeping genes GAPDH and RPL13a was carried out using the Rotor-gene system (Corbett Life Science, Australia). Complementary DNA (cDNA) was synthesized from 240ng of total RNA using oligodT16 primers and the SuperScriptTM III Reverse Transcriptase kit (Invitrogen, Carsbald, CA, USA). A standard curve for gene expression of HER3 and GAPDH and RPL13a was constructed using total RNA from a human breast cancer cell line (SKBR-3). Replicates of 10-fold dilutions of RNA in water were made to give 3 different calibrators, each with a known amount of SKBR-3 RNA for which arbitrary copy number were given. The PCR conditions were: 50°C for 2 minutes, 95°C for 2 minutes, followed by 40 cycles with the following profile: denaturation at 95°C for 20 seconds, annealing at 60°C for 15 seconds, extension at 72°C for 20 seconds. Flourescent incorporation was acquired in the green channel during the 72°C incubations. Intron spanning primer sequences were: GAPDH (5’CTGCACCACCAACTGCTTAG3’, 5’GTCTTCTGGGTGGCA GTGAT3’), RPL13a (5’GTTGGTGTTCATCCGTTG3’, 5’GTACGCTGTGAAGCATCAA3’), HER3 (5’TGGTTCATGGGCATGTATCC3’, 5’CTAGACCTAAGACTTGGAGC3’).

DASL gene expression profilingGene expression profiling was performed using the DASL assay (cDNA-mediated annealing, selection extension and ligation, Illumina Inc., California, USA) interrogating the DASL Cancer Panel that contained 512 cancer related genes. The first and second sets of samples were analyzed in two independent experiments. Tumours arising from the animal experiment were analyzed in conjunction with the second set of clinical samples. All protocols were as specified by Illumina Inc. Briefly, RNA (250ng) was converted to cDNA through a reverse transcription reaction with biotinylated oligo-d(T)18 and random nonamers. Oligonucleotides (three unique pairs for each of the 512 genes) were annealed to the biotinylated cDNA, the duplexes were bound to streptavidin conjugated paramagnetic particles to remove non-hybridized oligos. The annealed oligos were then extended and ligated (to incorporate an address sequence and primer site) to generate amplifiable products. These products were subjected to PCR amplification using fluorescently labeled (Cy3 and Cy5) primers. The labeled PCR products were hybridized to a Sentrix array or Beadchip. Following hybridization, the arrays were scanned with a BeadArray Reader (Illumina) and data was extracted using BeadStudio software (Illumina). Samples exhibiting a median hybridization intensity across all probes of <600 (background corrected) were excluded from analysis. Probes with a BeadStudio detection score greater than 0.99 in more than 15 conditions were included. Array data transformation and normalization (per chip normalized to 50th percentile and per gene normalized to median) was done in Genespring® version 7.0 software (Agilent Technologies, Santa Clara, USA). Hierarchical clustering was performed using: Similarity measure; Pearson Correlation and clustering algorithm; Average Linkage. Genes differentially expressed between primary tumors and brain metastases were identified using analysis of variance (ANOVA, p<0.01) or parametric test, variances not assumed equal (Welch t-test, p < 0.005). Up and down regulation was considered for normalized data values that were greater or less than a factor of 2 fold for the conditions compared.

first: Nic

last: Waddell


title: HNSCC samples vs. normal tonsil samples: self-self hybridization design

status: Public on Jan 22 2009

city: Houston

first: Yuan

last: Qi

state: TX

address: 1400 Pressler St

department: Bioinformatics & Computational Biology

institute: University of Texas M.D. Anderson Cancer Center

summary:
transcription profiling of human head and neck squamous cell carcinoma (HNSCC) samples vs. normal tonsil samples using a self-self hybridization experimental design on two-color arrays. Used to identify differentially expressed genes in tumor/normal samples, and compare the result to that of the same samples using the reference design experimental setting.

Keywords: tumor/normal comparison

overall_design: 8 HNSCC tumor samples and 8 normal tonsil samples. Each was profiled using a self-self hybridization design, where two aliquots of the same sample were labeled with two different dyes (cy3 and cy5) and hybridize to the same array.

first: D

middle: N

last: Hayes


title: HNSCC samples vs. normal tonsil samples: reference design

summary:
transcription profiling of human head and neck squamous cell carcinoma (HNSCC) samples vs. normal tonsil samples using a two-color reference design experimental setting. Used to identify differentially expressed genes in tumor/normal samples, and compare the result to that of the same samples using the self-self hybridization experimental setting.

Keywords: tumor/normal comparison

overall_design: 8 HNSCC tumor samples and 8 normal tonsil samples. One sample per array. Two-color reference design with a common reference sample (a modified version of the Stratagene Human Universal Reference) on each array.


title: LNCaP cells treated with chemoprevetive agents

status: Public on Jan 21 2009

city: Stanford

last: Stanford Microarray Database

address: 300 Pasteur Drive

department: Stanford University, School of Medicine

institute: Stanford Microarray Database (SMD)

summary:
BACKGROUND. Prostate cancer is thought to arise as a result of oxidative stresses and induction of antioxidant electrophile defense (phase 2) enzymes has been proposed as a prostate cancer prevention strategy. The isothiocyanate sulforaphane, derived from cruciferous vegetables like broccoli, potently induces surrogate markers of phase 2 enzyme activity in prostate cells in vitro and in vivo. To better understand the temporal effects of sulforaphane and broccoli sprouts on gene expression in prostate cells, we carried out comprehensive transcriptome analysis using cDNA microarrays. METHODS. Transcripts significantly modulated by sulforaphane over time were identified using StepMiner analysis. Ingenuity Pathway Analysis (IPA) was used to identify biological pathways, networks, and functions significantly altered by sulforaphane treatment. . StepMiner and IPA revealed significant changes in many transcripts associated with cell growth and cell cycle, as well as a significant number associated with cellular response to oxidative damage and stress. Comparison to an existing dataset suggested that sulforaphane blocked cell growth by inducing G2/M arrest. Cell growth assays and flow cytometry analysis confirmed that sulforaphane inhibited cell growth and induced cell cycle arrest. CONCLUSIONS. Our data suggest that in prostate cells sulforaphane primarily induces cellular defenses and inhibits cell growth by causingG2/Mphase arrest. Furthermore, based on the striking similarities in the gene expression patterns induced across experiments in these cells, sulforaphane appears to be the primary bioactive compound present in broccoli sprouts, suggesting that broccoli sprouts can serve as a suitable source for sulforaphane in intervention trials.
A compound treatment design type is where the response to administration of a compound or chemical (including biological compounds such as hormones) is assayed.
Time: length of treatment
Compound Based Treatment: LNCaP cells were treated with 10 or 25 uM L-sulforaphane, broccoli extract or DMSO

Keywords: compound_treatment_design

overall_design: Computed

first: Suvarna

last: Bhamre

first: Sunita

last: Jones

first: Hongjuan

last: Zhao


title: Pilot-study for a novel universal tool for detection of oncogenic fusion transcripts

status: Public on Jan 19 2009

city: Oslo

first: Gard

middle: Olav

last: Thomassen

address: Radiumhospitalet, Rikshospitalet HF

department: Institute for Cancer Research

country: Norway

institute: Rikshospitalet

summary:
A novel oligonucleotide microarray design is described whereby one can screen for all known oncogenic fusion transcripts by one microarray hybridization. Measurements of chimeric transcript junctions are combined with exon-wise measurements of individual fusion partners.

Keywords: Nimblegen custom-design

overall_design: The pilot data included a design with 68,861 oligonucleotide probes covering all combinations of chimeric exon-exon junctions from 275 pairs of fusion genes, as well as sets of oligos internal to all the exons of the fusion partners. Proof of principle was demonstrated by detection of known fusion genes (such as TCF3:PBX1, ETV6:RUNX1, and TMPRSS2:ERG) from six positive controls consisting of leukemia cell lines and prostate cancer biopsies.

first: Rolf

middle: I

last: Skotheim

middle: O

first: Marthe

last: Eken

first: Guro

last: Lind

first: Francesca

last: Micci

first: Franclim

middle: R

last: Ribeiro

first: Nuno

last: Cerveira

first: Manuel

last: Teixeira

first: Sverre

last: Heim

first: Torbjørn

last: Rognes

first: Ragnhild

last: Lothe


title: Estimating networks from RNA interference and microarray data

status: Public on Jan 29 2009

city: Heidelberg

first: Mark

last: Fellmann

address: INF 580

department: Molecular Genome Analysis

institute: DKFZ

summary:
We here used the technology of RNA interference and cDNA microarrays to reconstruct a network between genes known to be disregulated in ER-positive breast cancer diseases. Therefor single genes of high importance were silenced and the effect of this pertubation was measured with cDNA microarrays
Keywords: network_reconstruction_RNAi

overall_design: human ER-positive breast cancer cell line was cultivated as described and transfection with chemically synthesized siRNAs was performed according to standard HiPerFect transfection protocol by Qiagen (Hilden, Germany) At least 4 cDNA-microarrays were hybridized in dye-swap method for the silencing of every gene of interest. To minimize off-target effects we used at least two different siRNAs per gene.


title: Tamoxifen induces expression of immune-response related genes in cultured NHMEC

first: Ofelia

middle: Ana

last: Olivero

address: 37 Convent Dr-MSC 4255

department: CDI

institute: NCI-CCR-

summary: Administration of tamoxifen (TAM) as breast cancer adjuvant therapy is associated with a 50% reduction in breast cancer risk and an increase in endometrial cancer risk. To investigate underlying mechanisms, we documented TAM-induced gene expression changes in cultured normal human mammary epithelial cell (NHMEC) strains (numbered 5, 16 and 40) established from tissue taken at reduction mammoplasty from three different individuals. Cells exposed to 0 and 10 uM TAM for 48 hours were evaluated for survival, TAM-DNA adduct formation by TAM-DNA chemiluminescence immunoassay (CIA), and gene expression changes using DNA-oligonucleotide microarray and real time reverse transcriptase-PCR (RT-PCR). At 48 hr, cells exposed to 10 uM TAM were 78% viable, respectively, and there were no measurable TAM-DNA adducts (limit of detection [LOD] = 0.3 adducts/10^8 nucleotides). For microarray analysis, RNA was extracted from cells exposed on three occasions to 10 uM TAM and the corresponding oligonucleotide probes were labeled with fluorescent dyes and hybridized to NCI microarrays (>20,000 human genes). Expression changes of > 3-fold were considered significant, and by these criteria, thirteen genes were up-regulated and one was down-related in NHMEC strain 16, seventeen genes were up-regulated in strain 05, and eleven genes were up-regulated in strain and 40. Elements that were up-regulated in all three cell strains included several interferon-induced genes (IFITM1, IFIT1, IFNA1, MXI and GIP3), and a potassium ion channel (KCNJ1). No significant expression changes were found for genes related to estrogen or xenobiotic metabolism. RT-PCR revealed TAM-induced up-regulation of the five genes of interest, and interferon á (IFNA1), in all three NHMEC strains, with the exception of GIP3 and MX1, which were unchanged in strain 40. The magnitude of TAM-induced up-regulation ranked: strain 16 > strain 05> strain 40. The consistent induction of interferon-related genes in all three NHMEC strains suggests that, in addition to hormonal effects, TAM exposure may enhance immune response, through interferon induction, in normal breast tissue.

overall_design: RNA was extracted from three cell strains (NHMEC 98016, 98040 and 99005) and exposed on three separate occasions to 10 uM TAM. Comparisons were made using 16 arrays with 8 dye swaps for cell strain NHMEC 99005, 9 arrays with 4 dye swaps for NHMEC 98016, and 12 arrays with 6 dye swaps for NHMEC 98040.


title: Hypoxia and biofilms in Candida parapsilosis

status: Public on Jan 15 2009

city: Dublin

first: Geraldine

last: Butler

address: Belfield

department: School of Biomolecular and Biomedical Science

country: Ireland

institute: University Colege Dublin Conway Institute


title: Transcriptional profile of Candida parapsilosis in SD media

summary: Partial genome microarray  and plankton cells

overall_design: RNA was isolated from biofilms and from planktonic cells grown in SD media. Samples were labeled with either Cy5 or Cy3.  Four independent biological replicates were compared, including dye swaps.

first: Tristan

last: Rossignol


title: Global gene-expression analyses of the Esrrb reprogrammed cells

city: Singapore

first: Vinsensius

last: Vega

address: 60 Biopolis Street

institute: Genome Institute of Singapore

summary:
In the murine system, Oct4, Sox2, c-Myc and Klf4 are sufficient to convert fibroblasts to induced pluripotent stem (iPS) cells that exhibit many characteristics of embryonic stem (ES) cells. Herein, we show that the orphan nuclear receptor Esrrb works in conjunction with Oct4 and Sox2 to mediate reprogramming of mouse embryonic fibroblasts (MEFs) to iPS cells. Esrrb reprogrammed cells share similar expression and epigenetic signatures as ES cells. These cells are also pluripotent and can differentiate in vitro and in vivo into the three major embryonic cell lineages. Furthermore, these cells contribute to mouse chimeras and are germline transmissible. In ES cells, Esrrb targets many genes involved in selfrenewal and pluripotency. This suggests that Esrrb may mediate reprogramming through the up-regulation of ES cell-specific genes. Our findings also indicate that it is possible to reprogram MEFs without exogenous Klf transcription factors and link a nuclear receptor to somatic cell reprogramming.
We used microarrays to detail the global programme of gene expression of  ES cells, Esrrb reprogrammed iPS cell lines and MEFs.

Keywords: comparative

overall_design: Two biological replicates for ES cells, each iPS cell line and each type of MEF. The global gene expression profiles of the Esrrb reprogrammed iPS cell lines were compared to ES cells and MEFs.

first: B

last: Feng

first: J

last: Jiang

first: P

last: Kraus

first: JH

last: Ng

first: JCD

last: Heng

first: YS

last: Chan

first: LP

last: Yaw

first: W

last: Zhang

first: YH

last: Loh

last: Han

first: V

last: Cacheux Rataboul

last: Lim

first: T

last: Lufkin

first: HH


title: Global gene-expression analyses of the Esrrb knockdown cells

summary:
In the murine system, Oct4, Sox2, c-Myc and Klf4 are sufficient to convert fibroblasts to induced pluripotent stem (iPS) cells that exhibit many characteristics of embryonic stem (ES) cells. Herein, we show that the orphan nuclear receptor Esrrb works in conjunction with Oct4 and Sox2 to mediate reprogramming of mouse embryonic fibroblasts (MEFs) to iPS cells. Esrrb reprogrammed cells share similar expression and epigenetic signatures as ES cells. These cells are also pluripotent and can differentiate in vitro and in vivo into the three major embryonic cell lineages. Furthermore, these cells contribute to mouse chimeras and are germline transmissible. In ES cells, Esrrb targets many genes involved in selfrenewal and pluripotency. This suggests that Esrrb may mediate reprogramming through the up-regulation of ES cell-specific genes. Our findings also indicate that it is possible to reprogram MEFs without exogenous Klf transcription factors and link a nuclear receptor to somatic cell reprogramming.
Global gene expression effects of silencing the Esrrb gene. We used microarrays to detail the global programme of gene expression after silencing the Esrrb gene.

Keywords: time-course

overall_design: Three biological replicates each for control GFP and Esrrb RNAi. The global gene expression profiles of the Esrrb knockdown cells were compared to control GFP knockdown cells for days 2, 4 and 6.


title: Global gene-expression analyses of the Esrrb reprogrammed cells and Esrrb knockdown cells

summary:
In the murine system, Oct4, Sox2, c-Myc and Klf4 are sufficient to convert fibroblasts to induced pluripotent stem (iPS) cells that exhibit many characteristics of embryonic stem (ES) cells. Herein, we show that the orphan nuclear receptor Esrrb works in conjunction with Oct4 and Sox2 to mediate reprogramming of mouse embryonic fibroblasts (MEFs) to iPS cells. Esrrb reprogrammed cells share similar expression and epigenetic signatures as ES cells. These cells are also pluripotent and can differentiate in vitro and in vivo into the three major embryonic cell lineages. Furthermore, these cells contribute to mouse chimeras and are germline transmissible. In ES cells, Esrrb targets many genes involved in selfrenewal and pluripotency. This suggests that Esrrb may mediate reprogramming through the up-regulation of ES cell-specific genes. Our findings also indicate that it is possible to reprogram MEFs without exogenous Klf transcription factors and link a nuclear receptor to somatic cell reprogramming.

This SuperSeries is composed of the SubSeries listed below.


title: Age-Dependent Differences in Gene Expression in Response to Calvarial Injury

last: Stanford Microarray Database

summary:
Children less than 2 years of age are capable of healing large calvarial defects, whereas adults have been found to lack this endogenous ability. In this study, we used microarray analysis to compare genomewide expression patterns during active regeneration after injury with calvaria in skeletally immature and mature mice. Parietal bone defects were created in 6-day-old (juvenile) and 60-day-old (adult) mice using a 4-mm trephine bit (n = 20 mice per age group). The calvarial disc was removed, leaving the underlying dura mater intact. Two weeks after injury, the region of regeneration with the underlying dura mater was harvested, and RNA was extracted for microarray analysis. The 25 most differentially upregulated genes in juvenile regenerates compared with adults were listed, as well as selected bone-related genes. In addition, QRT-PCR confirmation of specific genes was performed for validation. Juvenile regenerates expressed significantly greater amounts of BMP-2, -4, -7, as well as FGF-2 and its receptor FGFR-1. Various other growth factors were also noted to be upregulated, including IGF-2 and Ptn. This corresponded with the increased expression of markers for osteogenic differentiation of Sparc and Oc. Markers of osteoclast activity, Acp5, Ctsk, and Mmp2, were noted to be greater in juvenile regenerates compared with adults. The observation of Mmp14 upregulation, however, highlights the importance of balanced osteoclast-mediated bone resorption for ultimate healing. The 2 most differentially regulated genes, transthyretin (Ttr) and prostaglandin D2 synthase (Ptgds), highlight the potential role of retinoic acid signaling and the prostaglandin axis on skeletal regeneration. These findings underscore the multitude of biomolecular mechanisms at play, allowing juvenile calvaria to heal after injury. The identification of various growth factors and cytokines involved also suggests novel therapeutic strategies for tissue-engineering purposes.
Set of arrays that are part of repeated experiments
Elapsed Time: Calvarial regeneration in 6 day old  vs 60 day old mice

overall_design: Biological Replicate

first: Zhen

last: Wang


title: Temporal Expression of Chemokines Dictates the Hepatic Inflammatory Infiltrate in a Murine Model of Schistosomiasis.

status: Public on Jan 10 2009

city: Herston

first: Melissa

middle: Louise

last: Burke

state: QLD

address: 300 Herston Road

department: Division of Infectious Diseases and Immunology

institute: Queensland Institute of Medical Research

summary:
Schistosomiasis continues to be an important cause of parasitic morbidity and mortality world-wide. Determining the molecular mechanisms regulating the development of granulomas and fibrosis will be essential for understanding how schistosome antigens interact with the host environment. We report here the first whole genome microarray analysis of the murine liver during the progression of Schistosoma japonicum egg-induced granuloma formation and hepatic fibrosis. Our results reveal a distinct temporal relationship between the expression of chemokine subsets and the recruitment of cells to the infected liver. Genes up-regulated earlier in the response included T- and B-cell chemoattractants, reflecting the early recruitment of these cells illustrated by flow cytometry. The later phases of the response corresponded with peak recruitment of eosinophils, neutrophils, macrophages and myofibroblasts/hepatic stellate cells (HSCs) and the expression of chemokines with activity for these cells including CCL11 (eotaxin 1), members of the Monocyte-chemoattractant protein family (CCL7, CCL8, CCL12) and the Hepatic Stellate Cell/Fibrocyte chemoattractant CXCL1. Peak expression of macrophage chemoattractants (CCL6, CXCL14) and markers of alternatively-activated macrophages (e.g. Retnla) during this later phase provides further evidence of a role for these cells in schistosome-induced pathology. Additionally, we demonstrate that CCL7 immunolocalises to the fibrotic zone of granulomas. Furthermore, striking up-regulation of neutrophil markers and the localisation of neutrophils and the neutrophil chemokine S100A8 to fibrotic areas suggests the involvement of neutrophils in S. japonicum¬-induced hepatic fibrosis. These results further our understanding of the immunopathogenic and, especially, chemokine signalling pathways that regulate the development of S. japonicum-induced granulomas and fibrosis and may provide correlative insight into the pathogenesis of other chronic inflammatory diseases of the liver where fibrosis is a common feature.

Keywords: Time course, disease state analysis

sample_organism:
Schistosoma japonicum
Mus musculus

overall_design: The gene expression profile of the murine liver were examined at 4, 6 and 7 weeks post infection with S. japonicum in comparison to that of uninfected controls. Microarray analysis was performed on pooled RNA from 4 mice per group. Two technical replicates were performed for each cRNA sample.  Microarray analysis was also performed on RNA derived from S. japonicum eggs to examine the contribution of increasing levels of egg derived RNA to the gene expression patterns observed in the liver.

middle: L

first: Donald

last: McManus

first: Grant

middle: A

last: Ramm

first: Mary

last: Duke

first: Yuesheng

last: Li

first: Malcolm

last: Jones

first: Geoffrey

last: Gobert


title: GSTM3 single nucleotide polymorphism (rs7483) associated with peripheral BMC expression in Alzheimer disease

status: Public on Jan 12 2009

city: Montreal

first: Olivier

middle: Charles

last: Maes

state: Quebec

address: 3999 Cote Ste Catherine

department: Bloomfield Centre for Research in Aging

institute: Lady Davis Institute for Medical Research

summary:
Comparisons between the sample groups (normal elderly control (NEC) and Alzheimer disease (AD)) allowed the identification of genes with disease expression patterns associated with the glutathione S-transferase M3 single nucleotide polymorphism rs7483.
Keywords: biological repeat

overall_design: Peripheral blood mononuclear cells (BMC) were obtained from normal elderly control (NEC) and Alzheimer disease (AD) subjects.  Targets from biological replicates of NEC (n=18) and AD (n=16) were generated and the expression profiles were determined using the NIA Human MGC cDNA microarray.

first: George

last: Chong

first: Howard

last: Chertkow

first: Hyman

middle: M

last: Schipper

first: Eugenia


title: Transcription profiling of human head and neck squamous cell carcinoma (HNSCC) samples vs. normal tonsil samples

first: Yuan

last: Qi

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

overall_design: Refer to individual Series


title: Mouse Sirt6-/- RelA+/- tissues

status: Public on Jan 09 2009

city: South San Francisco

first: Adam

middle: S

last: Adler

address: 1 DNA Way

institute: Genentech

summary:
We harvested spleen tissue from Sirt6-/- or Sirt6-/- RelA+/- animals and then completed genome-wide microarray analysis. Total RNA was extracted with TRIzol (Invitrogen) and amplified with Amino Allyl MessageAmp II Amplification kit. Amplified age-matched wild-type tissues were used as reference RNA for analysis. Array hybridization of mouse MEEBO arrays is as described (Hendrickson et al., PLoS ONE, 2008).
Set of arrays that are part of repeated experiments

Keywords: Biological Replicate

overall_design:
Biological Replicate
Computed

first: Tatiparthi

last: Reddy


title: Mouse Sirt6-/- tissues

summary:
We harvested liver, spleen, thymus, and skin tissues from 21 or 29 day old Sirt6-/- animals and then completed genome-wide microarray analysis. Total RNA was extracted with TRIzol (Invitrogen) and amplified with Amino Allyl MessageAmp II Amplification kit. Amplified age-matched wild-type tissues were used as reference RNA for analysis. Array hybridization of mouse MEEBO arrays is as described (Hendrickson et al., PLoS ONE, 2008).
Set of arrays that are part of repeated experiments
Tissue type: Age & tissue harvested

Keywords: Biological Replicate

first: Tiara

last: Kawahara


title: Sirt6 study

platform_organism:
Homo sapiens
Mus musculus


title: Mouse Sirt6-/- TNF-alpha timecourse

summary:
We completed genome-wide microarray analysis in Sirt6 knockout (ko) mouse embryonic fibroblasts (MEFs) and control wildtype (wt) MEFs. Prior to RNA extraction, MEFs were treated with TNF-alpha (10 ng/ml) for the indicated times. Total RNA was extracted with TRIzol (Invitrogen) and amplified with Ambion Amino Allyl MessageAmp II Amplification kit. Amplified mouse Universal Reference RNA (Stratagene) was used as reference RNA for analysis. Array hybridization of mouse MEEBO arrays is as described (Hendrickson et al., PLoS ONE, 2008).
Groups of assays that are related as part of a time series.
Compound Based Treatment: 10 ng/ml TNF-alpha

Keywords: time_series_design


title: Comparison of consecutive microarray platforms

status: Public on Dec 31 2008

city: Bonn

first: Joachim

last: Schultze

state: NRW

address: Carl-Troll-Strasse 31

department: Genomics and Immunoregulation

institute: LIMES (Life and Medical Sciences Center Genomics and Immunoregulation)

summary:
The MAQC project demonstrated that microarrays with comparable content show inter- and intra-platform reproducibility. However, since the content of gene databases still increases, the development of new generations of microarrays covering new content is mandatory. To better understand the potential challenges updated microarray content might pose on clinical and biological projects we developed a methodology consisting of in silico analyses combined with performance analysis using real biological samples. Here we clearly demonstrate that not only oligonucleotide design but also database content and annotation strongly influence comparability and performance of subsequent generations of microarrays. Additionally, using human blood samples and purified T lymphocyte subsets as two independent examples, we show that a performance analysis using biological samples is crucial for the assessment of consistency and differences. This study provides an important resource assisting investigators in comparing microarrays of updated content especially when working in a clinical or regulatory setting.
Keywords: microarray comparison

overall_design: Samples performed on different microarray platforms were compared. We therefore used Skleroderma and Bacteremia samples to compare technical replicates on consecutive array platforms.

first: Daniela

last: Eggle

first: Svenja

last: Debey-Pascher

first: Marc

last: Beyer


title: Alzheimer disease diagnostic prediction using peripheral BMC gene expression

summary: Using prediction analysis for microarrays (PAM), peripheral blood mononuclear cells (BMC) gene expression proves useful as predictive markers of sporadic AD. We obtained 90% accuracy for correct AD classification by PAM (36 of 40; four NEC classified as AD) using BMC gene expression. Investigating the contribution of these genes to the pathology of AD and their usefulness as prognostic biomarkers holds promise in the prevention of this devastating disease.

overall_design: Peripheral blood mononuclear cells (BMC) were obtained from normal elderly control (NEC) and Alzheimer disease (AD) subjects. Targets from biological replicates of NEC (n=22) and AD (n=18) were generated and the expression profiles were determined using the NIA Human MGC cDNA microarray. Disease classification was preformed by the PAM software provided by Stanford University, using a subset of differentially expressed genes in AD vs. NEC.


title: Gene expression in early phases of Autosomal Dominant Polycystic Kidney Disease

status: Public on Dec 28 2008

city: Taipei

first: wencheng

last: chen

address: 128 Sec. 2, Academia Rd

department: Insititute of Molecular Biology

country: Taiwan

institute: Academia Sinica

summary: Little is known about the genes involved in the initial cyst formation and disease progression in autosomal dominant polycystic kidney disease (ADPKD). To uncover the genetic determinants and molecular mechanisms of ADPKD, we analyzed 4-point time-series DNA microarrays from Pkd1L3/L3 mice to generate high-resolution gene expression profiles at different stages of disease progression. We found different characteristic gene expression signatures in the kidneys of Pkd1L3/L3 mice compared to age-matched controls during the initial phase of the disease. By postnatal week 1, the Pkd1L3/L3 kidney already had a distinctive gene expression pattern considerably different from the corresponding normal controls.  The genes differentially expressed, either induced or repressed, in ADPKD are important in immune defense, cell structure and motility, cellular proliferation, apoptosis and metabolic processes, and include members of three pathways (Wnt, Notch, and BMP) involved in morphogenetic signaling. Further analysis of the gene expression profiles from the early stage of cystogenesis to end stage disease identified a possible gene network involved in the pathogenesis of  ADPKD.

overall_design:
A total of 13 microarray data sets were generated with RNA samples from age-matched control and disease littermates at the four time points, with at least three biological replicates for each time point (PNW1, n = 3; PNW2, n = 4; PNW3, n = 3; PNW3.5, n = 3).
13 2-color experiments including:
3 replicates of: Wt mice vs. Pkd1L3/L3 mice at PNW 1
4 replicates of: Wt mice vs. Pkd1L3/L3 mice at PNW 2
3 replicates of: Wt mice vs. Pkd1L3/L3 mice at PNW 3
3 replicates of: Wt mice vs. Pkd1L3/L3 mice at PNW 3.5

first: Wen-Cheng

last: Chen


title: T7-based linear amplification of low concentration mRNA samples using beads and microfluidics

status: Public on Jan 01 2009

city: gaithersburg

first: audrey

middle: n

last: player

address: 8717 grovemont circle

department: microarray facility

institute: national cancer institute

summary:
We have demonstrated in vitro transcription (IVT) of cDNA sequences from purified Jurkat T-cell mRNA immobilization on microfluidic packed beads down to single-cell quantities.  The microfluidic amplified aRNA was nearly identical in length and quantity compared with benchtop reactions using the same starting sample quantities.  Microarrays were used to characterize the number and population of genes in each sample, allowing comparison of the microfluidic and benchtop processes.

Keywords: gene expression transcripts detected

overall_design: In this study we examined the IVT reaction in a microfluidic system apart from the other steps in the linear amplification process. Stock concentrations of cDNA were generated and known concentrations (down to single-cell equivalents) were packed onto the microfluidic or utilized as a benchtop process. IVT reactions were performed and efficiencies compared using microarray platforms.

first: Audrey

last: Player

first: Jason

last: Kralj


title: Cell cycle dependent variation of a CD133 epitope in human embryonic stem cell, colon cancer and melanoma cell lines.

city: La Jolla

first: Roy

last: Williams

address: 3030 Science Park Road

department: Regenerative Medicine

institute: The Scripps Research Institute

summary:
CD133 (Prominin1) is pentaspan transmembrane glycoprotein expressed in several stem cell populations and cancers. Reactivity with an antibody (AC133) to a glycoslyated form of CD133 has been widely used for the enrichment of cells with tumor initiating activity in xenograph transplantation assays. We have found by fluorescence-activated cell sorting that increased AC133 reactivity in human embryonic stem cells, colon cancer and melanoma cells is correlated with increased DNA content and reciprocally, that the least reactive cells are in the G1/G0 portion of the cell cycle. Continued cultivation of cells sorted on the basis of high and low AC133 reactivity results in a normalization of the cell reactivity profiles indicating that cells with low AC133 reactivity can generate highly reactive cells as they resume proliferation. The association of AC133 with actively cycling cells may contribute to the basis for enrichment for tumor initiating activity.

Keywords: Gene expression profiles of cells that express the AC133 epitiope of CD133 vs. AC133 negative cells

overall_design: Total RNA from AC133 high and negative sorted Caco2 and WM115 cells was extracted using the TRI Reagent (Molecular Research Center, Inc., Cincinnati, OH) according to the manufacturer’s protocol. Two samples each of two cell lines were analyzed as biological duplicates (8 in total). Labeled cRNA was prepared from 500 ng RNA using the Illumina® RNA Amplification Kit from Ambion (Austin, TX, USA). The Biotin labeled cRNA (750 ng) was hybridized 18 hr at 58ºC to the HumanRef-8 v2 Expression BeadChip (>22,000 gene transcripts; Illumina, San Diego, CA, USA) according to the manufacturer’s instructions. BeadChips were scanned with an Illumina BeadArray Reader and hybridization efficiency was monitored using BeadStudio software (Illumina) to measure internal controls built into the Illumina system. Data analysis was done in three stages. First, expression intensities were calculated for each gene probed on the array for all hybridizations (8 in total) using illumina’s Beadstudio#3 software. Second, intensity values were quality controlled and normalized: quality control was carried out by using the illumina Beadstudio detection P-value set to <0.05 as a cutoff. This removed genes which were effectively absent from the array (that is, were not detected). All the arrays were then normalized using the normalize quantiles routine from the Affy package in Bioconductor. This procedure accounted for any variation in hybridization intensity between the individual arrays.  Finally, these normalized data were imported into GeneSpring and analyzed for differentially expressed genes. The groups of biological replicates were described to the software and significantly differentially expressed genes determined on the basis of t-tests and fold difference changes in expression level. The list of significant differentially expressed genes was compared to published gene expression results with the use of Nextbio search engine.

first: Marie

last: Jaksch

first: Jorge

last: Múnera

first: Ruchi

last: Bajpai

first: Alexey

last: Terskikh

first: Robert

middle: G

last: Oshima


title: Saccharomyces cerevisiae cells: W303-1A vs W303-1A delta MSN2, MSN4

status: Public on Dec 23 2008

city: Randwick

first: Anthony

middle: G.

last: Beckhouse

state: NSW

address: High St

department: BABS

institute: Ramaciotti Centre for Gene Function Analysis

summary:
Transcriptional profiling of Saccharomyces cerevisiae cells comparing the W303-1A wildtype with the W303-1A double mutant for MSN2 and MSN4 during zinc deficient conditions

Keywords: Genetic modification with zinc limitation

overall_design: Two condition experiment, W303-1A vs W303-1A delta MSN2, MSN4. Biological replicates: 2 wildtype, 2 knock-out, independently grown and harvested.

first: Victoria

last: Lyons

middle: J

last: Gauci

first: Vincent

last: Higgins


title: Human shSIRT6 TNF-alpha timecourse

first: Adam

last: Adler

summary:
We established HeLa cell lines stably expressing a short hairpin RNA against SIRT6 (S6 sh2), and then completed genome-wide microarray analysis in shSIRT6 knock-down and control pSR HeLa cells. Prior to RNA extraction, HeLa cells were treated with TNF-alpha (5 ng/ml) for the indicated times. Total RNA was extracted with TRIzol (Invitrogen) and amplified with Ambion MessageAmp RNA Amplification kit. Amplified human Universal Reference RNA (Stratagene) was used as reference RNA for analysis. Construction and array hybridization of human cDNA microarrays were performed as previously described (Perou et al., 2000).
Groups of assays that are related as part of a time series.
Compound Based Treatment: 5 ng/ml TNF-alpha

Keywords: time_series_design

overall_design: Computed

first: Tatiparthi

last: Reddy


title: Gene expression profile of H187-63AR, an ABT-737 resistant derivative of H187

status: Public on Dec 19 2008

city: Baltimore

first: Christine

last: Hann

address: 1550 Orleans St., CRB2, Rm 562

department: Oncology

institute: Johns Hopkins University School of Medicine

summary:
ABT-737 is a highly potent small molecule inhibitor of Bcl-2 which has demonstrated efficacy in preclinical studies.  To identify factors which mediate ABT-737 sensitivity we created an ABT-737-resistant cell line derivative. This cell lines maintained its ABT-737 resistance in vitro and in vivo.  We isolated RNA from H187-63AR xenografts and compared their global gene expression to H187 xenografts.
Keywords: Tumor comparison

overall_design:
Gene expression profiles of H187 and H187-63AR derivative tumors were compared. Two tumors from each group were compared.

The raw data file (21208_Hann_nonorm_nobkgd_GEOarchive_Matrix.txt) contains the following:
1688628038_B - H187 Control 1 (57)
1688628038_C - H187 Control 2 (58)
1688628038_E - H187-63AR 1 (513)
1688628038_H - H187-63AR 1 (565)

first: Charles

last: Rudin


title: Subcutaneous adipose tissue from lean and obese subjects

status: Public on Dec 22 2008

city: Paris

first: David

last: Mutch

address: Centre de Recherche des Cordeliers (CRC), 15 rue de l'école de médecine

department: Nutrition and Metabolism

institute: INSERM UMRS U872 (Eq 7) Nutriomique

summary:
Obtaining adipose tissue samples are paramount to the understanding of human obesity. We have examined the impact of needle-aspirated and surgical biopsy techniques on the study of subcutaneous adipose tissue (scAT) gene expression in both obese and lean subjects. Biopsy sampling methods have a significant impact on data interpretation and revealed that gene expression profiles derived from surgical tissue biopsies better capture the significant changes in molecular pathways associated with obesity. We hypothesize that this is because needle biopsies do not aspirate the fibrotic fraction of scAT; which subsequently results in an under-representation of the inflammatory and metabolic changes that coincide with obesity. This analysis revealed that the biopsy technique influences the gene expression underlying the biological themes commonly discussed in obesity (e.g. inflammation, extracellular matrix, metabolism, etc), and is therefore a caveat to consider when designing microarray experiments. These results have crucial implications for the clinical and physiopathological understanding of human obesity and therapeutic approaches.

Keywords: subject and tissue biopsy technique comparison

overall_design:
Tissue samples from lean and obese subjects were analyzed: total of 36 hybridizations. The goal was to compare the effect of biopsy sampling methods on global subcutaneous adipose tissue gene expression analyses. The following subject groups were used for the analysis:
9 lean subjects: needle biopsy
9 lean subjects: surgical biopsy
9 obese subjects: needle biopsy
9 obese subjects: surgical biopsy

first: Karine

last: Clément

first: Joan

last: Tordjman

first: Veronique

last: Pelloux


title: Microarray analysis of the regulation of TNFalpha-stimulated PGE2 production in gingival fibroblasts

status: Public on Dec 11 2008

city: Stockholm

first: Johan

last: Lindberg

address: Roslagstullsbacken 21, Floor 3

country: Sweden

institute: Royal Institute of Technology

summary:
Prostaglandin E2 (PGE2) is involved in several inflammatory conditions including periodontitis. The aim of this study was to investigate the global gene expression profile of tumor necrosis factor alpha (TNFalpha) stimulated human gingival fibroblasts, focusing on signal pathways related to PGE2 production and the new PGE2-synthesizing enzymes, prostaglandin E synthases (PGES). The expression of microsomal prostaglandin E synthase-1 (mPGES-1) as well as the upstream cyclooxygenase-2 (COX-2) was up-regulated by TNFalpha, accompanied by increased PGE2 production. In contrast, the expression of microsomal prostaglandin E synthase-2 (mPGES-2) and cytosolic prostaglandin E synthase (cPGES) was unaffected by TNFalpha. Using microarray analysis in a time-course factorial design including time points 1, 3 and 6 h, we identified differentially expressed genes in response to TNFalpha treatment. Enrichment analysis of microarray data identified two positively regulated signal transduction pathways: c-Jun N-terminal kinase (JNK) and Nuclear Factor-kappaB (NF-kappaB). We used specific inhibitors and phosphorylation analysis to confirm their role in PGE2 regulation. Both JNK and NF-kappaB inhibitors reduced the TNFalpha-stimulated up-regulation of mPGES-1 and COX-2 as well as subsequent PGE2 production. The novel finding that TNFalpha-stimulated mPGES-1 is regulated by JNK suggests this kinase as a potential future target for treatment strategies in inflammatory disorders, including periodontitis.

Keywords: Time course, gene expression, factorial design.

overall_design: Three human gingival fibroblast cell lines were established from gingival biopsies obtained from 3 healthy patients, 3 to 12 years of age, with no clinical signs of periodontal disease. Cells were incubated with or without TNF-alpha (20 ng/ml) for 1, 3 or 6h. After incubation for 1, 3 or 6 h in the two conditions, the cells were immediately frozen in liquid nitrogen and then stored at -70°C for subsequent isolation of total RNA. The experimental design of the microarray study was set up as a time-course factorial design, to best observe the TNF-alpha induced gene expression changes over time. C++ program (G.F. Glonek, P.J. Solomon, Factorial and time course designs for cDNA microarray experiments, Biostatistics 5 (2004) 89-111) was used to determine the exact layout of the design in order to estimate the interaction effect between treatment and time, i.e. genes that are differentially expressed over time, with optimal statistical efficiency. Cyanine 5 and cyanine 3 were used for labeling. For each cell line 12 hybridizations were performed in a time-course factorial design. In total, 36 hybridizations were performed.

first: Tove

last: Båge

first: Joakim

last: Lundeberg

first: Thomas

last: Modéer

first: Tülay

last: Yucel-Lindberg


title: Atopic Eczema expression profiling, Saaf_2008

status: Public on Dec 17 2008

first: Annika

last: Saaf

department: Department of Molecular Medicine & Surgery

institute: Karolinska Institute, Karolinska University Hospital, Solna

summary:
Background: Atopic eczema (AE) is a common chronic inflammatory skin disorder. In order to dissect the genetic background several linkage and genetic association studies have been performed. Yet very little is known about specific genes involved in this complex skin disease, and the underlying molecular mechanisms are not fully understood.

Results: We used human DNA microarrays to identify a molecular picture of the programmed responses of the human genome to AE. The transcriptional program was analyzed in skin biopsy samples from lesional and patch-tested skin from AE patients sensitized to Malassezia sympodialis (M. sympodialis), and corresponding biopsies from healthy individuals. The most notable feature of the global gene-expression pattern observed in AE skin was a reciprocal expression of induced inflammatory genes and repressed lipid metabolism genes. The overall transcriptional response in M. sympodialis patch-tested AE skin was similar to the gene-expression signature identified in lesional AE skin. In the constellation of genes differentially expressed in AE skin compared to healthy control skin, we have identified several potential susceptibility genes that may play a critical role in the pathological condition of AE. Many of these genes, including genes with a role in immune responses, lipid homeostasis, and epidermal differentiation, are localized on chromosomal regions previously linked to AE.

Conclusion: Through genome-wide expression profiling, we were able to discover a distinct reciprocal expression pattern of induced inflammatory genes and repressed lipid metabolism genes in skin from AE patients. We found a significant enrichment of differentially expressed genes in AE with cytobands associated to the disease, and furthermore new chromosomal regions were found that could potentially guide future region-specific linkage mapping in AE.

A disease state experiment design type is where the state of some disease such as infection, pathology, syndrome, etc is studied.
Disease State: Skin biopsies from normal ('normal') and lesional and patch-tested ('leisonal') skin from Atopic eczema patients sensitized to Malassezia sympodialis
Organism Part: location of skin biopsy

Keywords: disease state analysis