title: Expression data from unactivated vs. activated PBMCs

status: Public on May 14 2010

city: Hamburg

first: Christoph

last: Ogrodowczyk

address: Falkenried 94

department: Institute for Neural Signal Transduction

institute: ZMNH

summary: Long-lasting activation of T cells requires up-regulation of many genes, for example of transcription factors, cytoskeletal proteins and cell surface proteins encluding ion channels. An increase of ion channel density at the cell surface reflects the needs to manage increased Ca2+ influx into the activated T cell. Using oligonucleotide-based arrays we have surveyed changes in ion channel mRNA expression that occur upon T cell activation. We used Affymetrix Analysis to confirmate our data achieved by self-designed glass array analysis.

overall_design: We isolated PBMCs from blood of healthy donors and divided those into two parts. One was activated for 72 hr by adding PHA-L whereas the other part was incubated in the same way but not stimulated.

first: C

first: O

last: Pongs


title: Global Gene Expression Profiles in Cultured Skin Fibroblasts Derived from Patients with Gaucher Disease

status: Public on May 20 2010

city: Bethesda

first: Kory

middle: R

last: Johnson

state: MD

address: 10/3B01, 9000 Rockville Pike

department: DIR IT & Bioinformatics

country: USA

institute: NINDS/NIH

summary: This work presents the discovery of genes that are dysregulated in patients with Type I and Type III Gaucher Disease. It provides insight into the unique pathogenesis of these phenotypes, improved diagnostic accuracy and potential novel therapies for these patients.

overall_design: Control and patient fibroblast cultures were established from full-thickness, skin biopsies obtained under a protocol approved by the IRB of the National Institute of Neurological Disorders and Stroke.  Patient cultures were homoallelic for either the N370S mutation (non-neuronopathic, Gaucher Disease Type I, n=5), the L444P mutation (neuronopathic, Gaucher Disease Type III, n=5), or Wild-Type (Control, n=4).

first: Roscoe

middle: O

last: Brady


title: Expression data from mesothelial (LP9) and mesothelioma cells (HMESO) inhibited for extracellular-regulated kinase (ERK) 1, 2, or 5

status: Public on May 08 2010

city: Burlington

first: Arti

last: Shukla

state: VT

address: 89 Beaumont Avenue

department: Pathology

institute: University of Vermont

summary: Extracellular-regulated kinases (ERK1/2 and 5) are known to play important roles in growth and drug resistance of various cancers. Here we show roles of inhibition of ERK1, ERK2, or ERK5 on gene expression profiles of epithelioid malignant mesothelioma (MM) cells (HMESO).

overall_design: Untransfected human mesothelial cells (LP9), MM cells (HMESO) or HMESO cells stably transfected with shControl, shERK1, shERK2 , or shERK5 plasmids were used. Microarrays were performed on samples from 3 independent experiments. Each sample was analyzed on a separate array (3 independent biological replicates).


title: Reorganization of nuclear lamina – genome interactions upon differentiation of embryonic stem cells.

status: Public on May 27 2010

city: Amsterdam

first: Bas

last: van Steensel

address: Plesmanlaan 121

department: division of Molecular Biology

country: Netherlands

institute: Netherlands Cancer Institute

type: Genome binding/occupancy profiling by genome tiling array

summary:
The three-dimensional organization of chromosomes within the nucleus and its dynamics during differentiation are largely unknown. We present a genome-wide analysis of the interactions between chromatin and the nuclear lamina during differentiation of mouse embryonic stem cells (ESCs) into lineage-committed neural precursor cells (NPCs) and terminally differentiated astrocytes. Chromatin in each of these cell types shows a similar organization into large lamina associated domains (LADs), which represent a transcriptionally repressive environment. During sequential differentiation steps, lamina interactions are progressively modified at hundreds of genomic locations. This remodeling is typically confined to individual transcription units and involves many genes that determine cellular identity. From ESCs to NPCs, the majority of genes that move away from the lamina are concomitantly activated. Strikingly, a significant amount remain inactive yet become primed for activation by further differentiation. These results suggest that lamina-genome interactions are widely involved in the control of gene expression programs during lineage commitment and terminal differentiation.

UPDATE WM20121119:
We have added per-probe Hidden Markov Model state calls for the 4 cell types. These calls are not used in Peric-Hupkes, Meuleman et al. (2010), but are used in Meuleman, Peric-Hupkes et al. (2012).
The procedure to arrive at this HMM state calls is as follows:
We fitted a two-state HMM whereby emissions are distributed as Student's t variables. Mean and variance of DamID signals differ between states, but the degree of freedom (nu) is the same. Gaps in the probe coverage were filled by evenly spaced null probe-values. The parameters were estimated by an adaptation of the ECME algorithm to the HMM framework, showing faster convergence than regular EM when nu is unknown (Filion et al., Cell, 2010). State calls were derived through the Viterbi algorithm. This process was repeated separately for each cell type, yielding per-probe calls. Probes in the ‘bound’ (1) state are indicated as LAD-probes, probes in the ‘unbound’ (0) state as inter-LAD-probes.

UPDATE WM20130301:
We have also added (1-based) BED files of constitutive LAD and inter-LAD regions (mm9) across the 4 cell types. These regions are the result of simply concatenating directly adjacent cLAD/ciLAD Hidden Markov Model state calls. Again, these calls are not used in Peric-Hupkes, Meuleman et al. (2010), but are used in Meuleman, Peric-Hupkes et al. (2012).

overall_design: laminB1-chromatin interactions were assayed in 4 different mouse cell-types. For each cell-type there were 2 biological replicates, that were hybridized in a dye-swap design.

first: Daan

last: Peric-Hupkes

first: Wouter

last: Meuleman

first: Ludo

last: Pagie

first: Stefan

last: Gräf

first: Paul

last: Flicek

first: Wim

last: Brugman

first: Ron

last: Kerkhoven

first: Lodewyk

last: Wessels

first: Marcel

last: Reinders


title: Parkinson's disease-associated DJ-1 is required for the expression of GDNF receptor Ret in human neuroblastoma cells

status: Public on Apr 16 2010

city: Trieste

first: Paola

last: Roncaglia

state: TS

address: via Bonomea 265

department: Neurobiology

country: Italy

institute: SISSA/ISAS (International School for Advanced Studies)

summary: DJ-1 is an atypical peroxiredoxin-like peroxidase that may act as a redox-dependent chaperone and a regulator of transcription. To explore DJ-1-mediated transcriptional control in Parkinson’s disease (PD), we generated human neuroblastoma cells with inducible knock-down of DJ-1 expression. We then used functional genomic techniques to identify novel pathways dysregulated by loss of DJ-1 function. Using microarray gene expression profiling, we found that DJ-1 silencing alters the expression of 26 genes, with 10 down-regulated and 16 up-regulated transcripts. Among the down-regulated genes we found Ret, tyrosine kinase receptor for the neurotrophic factor GDNF. Taking advantage of Ingenuity Pathways Analysis, we identified hypoxia inducible factor 1 alpha (Hif1a) as a possible mediator of the interplay between DJ-1 and Ret. We show that Hif1a is stabilized in the absence of DJ-1, and that loss of DJ-1 generates hypoxia and accumulation of free radical species (ROS). Overexpression of wt DJ-1, but not of C106A and L166P mutants deficient in ROS scavenger activity, rescues Ret expression in neuroblastoma cells. These findings reveal novel players in PD pathogenesis and provide evidence for additional pathways involved in DJ-1-mediated neurodegeneration.

overall_design: Comparison between DJ-1-silenced human SH-SY5Y neuroblastoma cells and control cells. Two clones were selected for each condition, and each clone was analyzed in duplicate, for a total of 8 samples.


title: Expression data from Human Kidney (HK) samples

status: Public on Apr 13 2010

city: Bronx

first: Han

last: Si

state: NY

address: 1300 Morris Park

department: Medicine/Nephrology

institute: AECOM

summary:
Males are 50% more likely to develop end stage kidney failure compared to women. In this study we wanted to find out the molecular mechanism responsible for this increased risk. We collected kidney samples from patients with and without kidney disease and performed a comprehensive gene expression analysis in healthy and diseased male and female kidneys.
Interestingly, the set of gender biased genes in healthy kidneys were different from those in diseased kidneys indicating not only baseline gene expression differences but also that the male and female kidney respond differently to disease condition. Our studies indicate that men and women with kidney problems might need to be treated differently.

Keywords: Gender difference

overall_design: We collected 42 kidney samples from healthy living transplant donors, nephrectomies and from diagnostic kidney biopsies. Preliminary studies did not show significant gene expression differences in control kidneys based on the collection method (i.e. living kidney biopsy vs. unaffected portion of tumor nephrectomy). We grouped the tissue samples based on the histological readings of the kidney biopsies. Samples with evidence of glomerular and tubulointerstitial fibrosis were assigned into the diseased group. Characteristics of the research participants indicate diverse ethnic and disease groups and mild (StageIII) CKD. The tissue was microdissected into glomerular and tubulointerstitial fractions and expression arrays were performed separately.

first: Katalin

last: Susztak


title: Anthracycline resistance in four human cancer cell lines (HGU95A)

status: Public on Apr 15 2010

city: Budapest

first: Balazs

last: Györffy

address: Bókay 53

country: Hungary

institute: Hungarian Academy of Sciences

summary:
Reliable clinical tests for predicting cancer chemotherapy response are not available and individual markers failed to correctly predict resistance against anticancer agents. We hypothesized that gene expression patterns attributable to chemotherapy-resistant cells can be used as a classification tool for chemoresistance and provide novel candidate genes involved in anthracycline resistance mechanisms. We contrasted the expression profiles of 4 different human tumor cell lines of gastric, pancreatic, colon and breast origin and of their counterparts resistant to the topoisomerase inhibitors daunorubicin or doxorubicin. We also profiled the sensitive parental cells treated with doxorubicin for 24h. We interrogated Affymetrix HGU133A and U95A arrays independently. We applied two independent methods for data normalization and used Prediction Analysis of Microarrays (PAM) for feature selection. In addition, we established data sets related to drug resistance by using a “virtual array” composed of features represented on both types of oligonucleotide arrays. We identified 71 candidate genes associated with doxorubicine/daunorubicine resistance. To validate the microarray data, we also analyzed the expression of 12 selected genes by quantitative RT-PCR or immunocytochemistry, respectively. While the comparison of drug-sensitive versus drug-resistant cells yields candidates associated with drug resistance, the 24h treatment of sensitive parental cells produced a distinct transcriptional profile related to short-term drug effects.
Keywords: cell type comparison

overall_design: See Summary. Note: this series includes Affymetrix HGU95A chips (difference between resistant and parental cell lines).


title: Oncogenic Functions of the Transcription Factor LSF

status: Public on Apr 06 2010

city: Richmond

first: Devanand

last: Sarkar

state: VA

address: 1220 E Broad St, Room 7044

department: Department of Human and Molecular Genetics

institute: Virginia Commonwealth University

summary:
Overexpression of LSF in less aggressive hepatocellular carcinoma (HCC) cells resulted in highly aggressive, angiogenic and multi-organ metastatic tumors while inhibition of LSF significantly abrogated growth and metastasis of highly aggressive HCC cells in nude mice.
We used microarrays to identify the downstream genes mediating the effects of LSF in HCC cells.

overall_design: Gene expression profiles were compared between Control-8 and LSF-17 clones of HepG3 cells.

first: Catherine

middle: I

last: Dumur


title: Metabolism-based resistance to herbicides in black-grass

status: Public on May 04 2010

city: Beaucouze

first: sandrine

last: balzergue

address: Rue G. Morel

country: France

institute: INRA

summary:
Resistance to herbicides in weeds can be due to alteration(s) in the gene encoding the herbicide target site, or to herbicide degradation via a deviation in plant general metabolism. If target-site-based resistance is easy to study, the multigenic control of metabolism-based resistance renders it much more complex to study. Metabolism-based resistance to herbicides represents the major part of herbicide resistance in black-grass. Its most likely basis is an overexpression of genes encoding enzymes degrading herbicides. We thus seek to identify such overexpressed genes by comparing the transcriptomes of resistant and sensitive black-grass individuals belonging to an F2 line segregating for two resistance genes. Given there are no genomic tools developed for black-grass, this approach will use heterologous hybridisation onto a wheat Affymetrix microarray.

Comparison using heterologous hybridisation onto a wheat whole-genome microarray of transcriptome of three pools of black-grass plants obtained 2h30 after herbicide spraying at field rate. The three pools correspond to: · Sensitive plants (killed by herbicide). · Moderately resistant plants (growth impaired by herbicide but plants still alive) · Resistant plants (growth unimpaired by herbicide)

overall_design: 6 arrays - wheat

first: Sandrine

last: Balzergue

first: Christophe

last: Délye

first: Cécile

last: Petit


title: affy_seed_kinetic_wheat-Transcriptomic wheat seed

status: Public on May 01 2010

summary:
affy_seed_kinetic_wheat - affy_seed_kinetic_wheat - Study gene expression during the grain developmental -The aim of the study is to identify the genes that are differentially expressed during the grain development in wheat.-Study gene expression during the grain developmental Sample at 100 degree days, year 2004 and 2006 Sample at 200 degree days, year 2004 and 2006 Sample at 250 degree days, year 2004 and 2006 Sample at 300 degree days, year 2004 and 2006 Sample at 400 degree days, year 2004 and 2006

Keywords: time course

overall_design: 10 arrays - wheat

first: Jerome

last: Salse

first: Marie-Laure

middle: L

last: Martin-Magniette


title: Expression profiles of MDA-MB-231, MDA-231 S1a and S1b

status: Public on Mar 25 2010

city: New York

first: Mi-Young

last: Kim

address: 1275 York Ave

institute: MSKCC

summary: Identification of genes that are involved in self-seeding by comparing gene expression profiles between parental MDA-MB-231 cells and seeder cells (MDA-231-S1a and S1b)

overall_design: 2 replicates from each sample (parental  MDA-MB-231, MDA-MB-231 S1a and  MDA-MB-231 S1b) were analyzed


title: [hexa-] Transcriptomic changes in synthetic hexaploid wheat

status: Public on Feb 18 2010

summary:
affy_hexaploid_wheat - hexa - Changes upon polyploidization in hexaploid wheat - transcriptomic changes in synthetic hexaploid derived from a cross between tetraploid and natural diploid.
Study aims to understand regulation of gene expression in synthetic and natural wheat allohexaploids (Triticum aestivum), that combines the AB genome of T. turgidum and the D genome of Aegilops tauschii; and which we have recently characterized as genetically stable.
We conducted a comprehensive genome-wide analysis of gene expression that allowed us to compare the effect of variability of the D genome progenitor, the trans-generation stability as well as comparison to natural wheat allohexaploid. We used the Affymetrix GeneChip® Wheat Genome Array, on which 55,049 transcripts are represented.
Additive expression was shown to represent majority of expression regulation in the synthetic allohexaploids, where expression for more than 93% of transcripts was equal to the one evaluated from equal mixture of parental RNA. This leaves ~2000 (~7%) transcripts, which expression was non-additive. No global gene expression bias or dominance towards any of the progenitor genomes was observed whereas high trans-generational stability and low effect of the D genome progenitor variability were revealed.
Our study suggests that gene expression regulation in wheat allohexaploids is early established upon allohexaploidization and highly conserved over generations, as demonstrated by the high similarity of expression with natural wheat allohexaploids.
Keywords: genotype and ecotype comparison

sample_organism:
Triticum aestivum
Triticum turgidum subsp. durum
Aegilops tauschii
Aegilops tauschii x Triticum turgidum

overall_design: 18 arrays - wheat

first: Veronique

last: Chague

first: Boulos

last: Chalhoub


summary:
Reliable clinical tests for predicting cancer chemotherapy response are not available and individual markers failed to correctly predict resistance against anticancer agents. We hypothesized that gene expression patterns attributable to chemotherapy-resistant cells can be used as a classification tool for chemoresistance and provide novel candidate genes involved in anthracycline resistance mechanisms. We contrasted the expression profiles of 4 different human tumor cell lines of gastric, pancreatic, colon and breast origin and of their counterparts resistant to the topoisomerase inhibitors daunorubicin or doxorubicin. We also profiled the sensitive parental cells treated with doxorubicin for 24h. We interrogated Affymetrix HGU133A and U95A arrays independently. We applied two independent methods for data normalization and used Prediction Analysis of Microarrays (PAM) for feature selection. In addition, we established data sets related to drug resistance by using a “virtual array” composed of features represented on both types of oligonucleotide arrays. We identified 71 candidate genes associated with doxorubicine/daunorubicine resistance. To validate the microarray data, we also analyzed the expression of 12 selected genes by quantitative RT-PCR or immunocytochemistry, respectively. While the comparison of drug-sensitive versus drug-resistant cells yields candidates associated with drug resistance, the 24h treatment of sensitive parental cells produced a distinct transcriptional profile related to short-term drug effects.

This SuperSeries is composed of the SubSeries listed below.

overall_design:
This reference Series links data in the following related Series:
GSE3926 Anthracycline treatment and resistance in four human cancer cell lines (HGU133A)
GSE3927 Anthracycline resistance in four human cancer cell lines (HGU95A)


title: Gene expression profiles induced by knockdown and overexpression of p63 variants in MCF-10A mammary epithelial cell line

status: Public on Mar 12 2010

city: Boston

first: Joan

middle: S.

last: Brugge

state: MA

address: 240 Longwood Avenue

department: Department of Cell Biology

institute: Harvard Medical School

summary: p63 is critical for epithelial development yet little is known about the transcriptional programmes it regulates. The p63 transactivating (TA) isoforms contain an amino-terminal exon that encodes a p53-like transactivation domain, whereas ΔN-isoforms lack this domain but contain the common DNA binding domain (DBD), suggesting that TAp63 and ΔNp63 isoforms may have opposing functions.  By characterising transcriptional changes and cellular effects following modulation of p63 expression, we have defined a vital role for p63 in cellular adhesion. Knockdown of p63 expression caused downregulation of cell adhesion-associated genes, cell detachment and anoikis in mammary epithelial cells and keratinocytes. Conversely, overexpression of the TAp63γ or ΔNp63α isoforms of p63 upregulated cell adhesion molecules, increased cellular adhesion and conferred resistance to anoikis.

overall_design: Total RNA was isolated 48 h after retroviral or adenoviral infection and was subjected to reverse transcription, labelling and hybridization.  The knockdown samples were performed in duplicate with shRNA vector control, shRNA targetting all isoforms of p63 (shDBD) and shRNA targetting p63 isoforms containing the transactivating (TA) domains.  The overexpression samples were performed in triplicate with vector control, overexpression of the ΔNp63α isoform, and overexpression of the TAp63γ isoforms.

first: Danielle

middle: K

last: Carroll

first: Jason

middle: S

first: Chee-Onn

last: Leong

first: Fang

last: Chen

first: Myles

last: Brown

first: Alea

middle: A

last: Mills

first: Leif

middle: W

last: Ellison


title: RIP-chip analysis of Tino/hMEX-3D mRNPs

status: Public on Feb 28 2010

city: Firenze

first: martino

last: donnini

state: Firenze

address: viale GB Morgagni

department: Pathology and Oncology

institute: University of Florence

summary:
Tino is an A+U-Rich Element (ARE) binding protein first identified through its ability to bind to bcl-2 mRNA and to contribute to its degradation. It has recently been recognized as a shorter form of the human Mex-3D protein (hMex-3D), one of the four members of the family of Mex-3 RNA-binding phosphoproteins. In C. elegans, ceMex-3 is a translational regulator that plays a key role in early embryonic development and in the maintenance of worm germ line totipotency. To examine the potential functional conservation between ceMex-3 and hMex3, we have used complementary microarray-based approaches to identify mRNAs directly bound to Tino/hMex-3D. Computational analysis of these target mRNAs resulted in the identification of an U-rich, 34- to 39-nucleotide long, consensus, forming loops of variable sizes. Remarkably, more than half of Tino/hMex-3D targets also contain the consensus for Quaking, which is the human ortholog of GLD-1, a regulator of nematode gametogenesis. All together, our results suggest that Tino/hMex-3D belongs to a regulatory circuit of mRNA trans-acting factors involved in cell fate and differentiation.

Keywords: RIP-chip and (recombinant)RIP-Chip analysis of Tino/hMEX-3D mRNPs

overall_design:
To identify the transcripts directly bound in vivo by Tino/hMex-3D, we have used two complementary strategies. First, using Tino-his transfected HEK293 cells, we have analysed the RNAs immunoprecipitated by Tino/hMex-3D (RNA-IP complexes). Second, we have developed an in vitro nitrocellulose RNA-protein binding assay using a truncated form of Tino, TinoΔRING-his. First for the in vivo approach, we have transiently transfected HEK293 cells with a His-tagged Tino expressing construct (see Materials) and isolated Tino target mRNAs by immunoprecipitation (IP) assays carried out under conditions preserving mRNA-protein complex integrity. Second, for the in vitro approach, cRNAs were prepared from a human placenta cDNA library and incubated with TinoΔRING-his. This deleted form of Tino-his protein conserves its mRNA binding activity. mRNAs directly bound to TinoΔRING-his protein were subsequently isolated using a nitrocellulose RNA-protein binding assay.

A supplementary file of ordered genes based on z-ratios, derived from experimental and negative control samples for each Affymetrix probe, is appended below.

first: Simone

last: Minardi

first: Elisa

last: Venturini

first: Matteo

last: Lulli

first: Martino

last: Donnini

first: Andrea

last: Lapucci


title: Expression data from LCM captured prostate cancer cells

status: Public on Apr 10 2010

city: kent

first: Gail

middle: C

last: Fraizer

state: OH

address: 251 Cunningham Hall

department: Biological Sciences

institute: Kent State University

summary:
The prostate represents a complex mix of cell types and there is a need to analyze distinct cell populations to better understand their potential interactions.  This study of cell-type specific gene expression patterns will contribute to understanding of how tumor epithelial cells may be affected by adjacent interstitial stromal cells within the tumor microenvirnonment.
LCM and microarray analysis were used to identify gene expression patterns in prostate cancer  epithelial and interstitial stromal cell populations. More than 500 genes whose expression levels were significantly different in epithelial cells versus adjacent stromal tissue were identified. Several chemokines and growth factors were more highly expressed in prostate cancer stromal cells  while several transcription factors and receptors were more highly expressed in epithelial cells.

overall_design: Five micron serial sections from three different intermediate grade prostate tumors were provided. One section from each had been stained by hematoxylin and eosin and the tumor areas marked for identification. To obtain RNA from 2 different tissue types (glandular epithelium and interstitial stroma) tumor areas observed to have abundant cells of interest were identified. Tumor epithelium and adjacent stromal tissue were collected separately from each slide. We sought to obtain similar populations of cells of each tissue type, so to that end, we examined each slide and and marked areas to direct the laser capture. Approximately 1,000 cells were  captured per cap and RNA was extracted and hybridized to Affymetrix microarrays as recommended by manufacturer.

first: Eric

middle: M

last: Mintz

first: Helen

last: Piontkivska

first: Jennifer

last: Gregg

first: Kathleen

middle: E


title: The trait of MS: Altered transcription regulation of nuclear receptors networks operate in the pre-disease state

status: Public on Feb 17 2010

city: Ramat Gan

first: Michael

last: Gurevich

address: Tel Hashomer

department: Multiple Sclerosis Center

country: Israel

institute: Sheba Medical Center

summary:
Molecular mechanisms that influence susceptibility to multiple sclerosis are poorly understood. We conducted a gene expression study in healthy subjects that subsequently developed the disease. Gene expression profiles (HG U133A and A2, Affymetrix,  22,215 transcripts) of peripheral blood mononuclear cells were analyzed in 9 healthy subjects (mean age 19.8+1.1 years) up to 9 years (mean 5.1±1.2 years) before onset of MS (MS to be, MS2b), 11 age-, gender-, and origin-matched subjects that remained MS-free (MSf), and 31 clinically isolated syndrome (CIS) patients. Most informative genes (p<0.05) and significant biological processes were compared. 1051 genes (611 up-regulated, 440 down-regulated) were significantly different between MS2b and MSf subjects. MS2b signature was characterized by down-regulation of the nuclear receptor (NR) family genes including NR subfamily 4 group A member1 (NR4A1, p=0.01), member 3 (NR4A3, p=0.01), NR subfamily 2 group F member 2 (NR2F1, p=0.03) and vitamin D receptor (VDR, p=0.02), all known to be involved in T-cell regulation by apoptosis. Comparison between MS2b and CIS operating networks demonstrated evolution of the altered NR dependent apoptosis regulation. Decreased NR4A1 expression was verified at the mRNA and protein level in an independent cohort of 20 relapsing-remitting MS patients. The identified MS trait is associated with suppressed transcription of NR networks that leads to altered apoptosis of activated T cells and the development of clinical disease. MS2b subjects have already an ongoing process that eventually will lead to clinical disease and our finding are of importance as they suggest the possibility of early detection and prevention of MS.

Keywords: disease state analysis

overall_design: Blood samples of healthy subjects that later developed MS (MS-to-be, MS2b, N=9) were identified and used for gene expression study. For each sample of MS2b subject, a sample of an age and gender matched subject that remained MS-free (MSf, N=11) was randomly selected. A cohort of 31 CIS patients who gave blood sample for gene expression analysis within 3 months of the onset of their first neurological event, was used for comparison with the MS2b gene expression signature. For establishment of the CIS gene expression signature, microarray data from 13 age- and sex- matched healthy subjects were used.

first: Anat

last: Achiron

first: Itamar

last: Grotto

first: Ran

last: alicer

first: David

last: Magalashvili

first: Anna

last: Feldman


title: Expression data from HMC-1 mast cells

status: Public on Mar 31 2010

city: Tel-Aviv

first: Dana

last: Baram

institute: Tel-Aviv University

summary:
We demonstrate that the G protein Gi3 is the cellular target of the adenosine A3 receptor (A3R). By using a cell permeable peptide comprising the C-terminal end of Gαi3 fused to an importation sequence (ALL1) as a selective inhibitor of Gi3 signaling, we show that by coupling to Gi3, the  A3R stimulates multiple signaling pathways in human mast cells, leading to upregulation of cytokines, chemokines and growth factors.Following contact with activated T cell membranes, endogenous adenosine binds to and activates the A3R, resulting in Gi3-mediated signaling. Specifically, the majority of ERK1/2 signaling initiated by contact with activated T cell membranes, is mediated by Gi3, giving rise to ALL1-inhibitable cellular responses. These results unveil the physiological GPCR that couples to Gi3 and establish the important role played by this G-protein in inflammatory conditions that involve adenosine-activated mast cells.
We used microarrays to detail the effect of ALL1 on  gene expression of HMC-1 cells activated directly by the A3 receptor, or by contact with activated T cell membranes.

overall_design: The experiment consisted of 6 treatments in duplicates, a total of 12 samples. The cells were activated via two different activation pathways (A3 and T membranes), with or without pretreatment with the drug ALL1.

first: Ronit

last: Sagi-Eisenberg

first: Ornit

last: Dekel


title: SAFB1 mediates repression of immune regulators and apoptotic genes in breast cancer cells

status: Public on Jan 28 2010

city: Houston

first: Chad

last: Creighton

state: TX

address: One Baylor Plaza, Mail Stop: BCM305

department: Biostatistics, Ducan Cancer Center

institute: Baylor College of Medicine

summary: The scaffold attachment factors SAFB1 and SAFB2 are paralogs, which are involved in cell cycle regulation, apoptosis, differentiation, and stress response. They have been shown to function as estrogen receptor co-repressors, and there is evidence for a role in breast tumorigenesis. To identify their endogenous target genes in MCF-7 breast cancer cells, we utilized gene expression array analysis, which was set up in a two-by-four design, with vehicle and estrogen treatment, and control, SAFB1, SAFB2, and SAFB1/SAFB2 siRNA as variables. Using custom chips containing 1.5 kb upstream regulatory region, we identified 541 SAFB1/SAFB2 binding sites in promoters of known genes, with significant enrichment on chromosome 1 and 6. Gene expression analysis revealed that the majority of target genes were induced in the absence of SAFB1 or SAFB2, and less were repressed. In contrast to SAFB2, which shared most of its target genes with SAFB1, SAFB1 had many unique target genes, most of them involved in regulation of the immune system. A subsequent analysis of the estrogen treatment group revealed that twelve percent of estrogen-regulated genes were dependent on SAFB1, with the majority being estrogen-repressed genes. These were primarily genes involved in apoptosis, such as BBC3, NEDD9, and OPG. Thus, this study confirms SAFB1/SAFB2’s primary role as co-repressors, and also uncovers a previously unknown role for SAFB1 in regulation of immune genes, and in estrogen-mediated repression of genes.

overall_design: To identify transcripts which were altered upon decrease of SAFB1 or SAFB2 expression we performed gene expression array analysis in MCF-7 cells. A secondary goal was to identify ERα-target genes for which estrogen regulation was dependent upon SAFB expression. We also added an experimental group in which we co-transfected siRNAs targeting both SAFB1 and SAFB2, which allowed us to address the question of interactions between the two paralogs. We therefore set up an experiment reflecting a 2x4 design, with a total of eight experimental groups.

last: Tsimelzon

first: Stephanie

last: Hammerich-Hille

first: Steffi

last: Oesterreich


title: MCF-7 Luciferase, PARP-1, PARG, SIRT1, and macroH2A Knockdown

status: Public on Jan 31 2010

city: Dallas

first: W. Lee

last: Kraus

address: 5323 Harry Hines Blvd.

institute: UT Southwestern Medical Center

summary: Affymetrix expression arrays were used to compare expression patterns upon knockdown of PARP-1,  PARG, SIRT1, or macroH2A in comparison to Luciferase control.

overall_design:
Total RNA isolated from three independently generated control-matched MCF-7 knockdown cell lines targeting Luciferase (control), PARP-1, PARG, SIRT1, and macroH2A was used to probe Affymetrix Human U133A 2.0 Expression Microarrays.

The raw array data was processed by Affymetrix GeneChip Operating Software (GCOS) to obtain detection calls (ABS_CALL) and signal values (VALUE_2). The signals were then normalized by scaling to a target value of 500 using GCOS. To adjust for batch effects due to day-to-day differences in RNA isolations, the empirical Bayes method was applied to the data set. After adjusting any values less than 0.01 to 0.01, the data was log2 transformed, median centered for each array, and median centered for each individual probe set (VALUE). Filters were then applied to obtain final gene lists.

first: Kristine

last: Frizzell

first: Matthew

middle: J

last: Gamble

first: Tong

last: Zhang

first: WL


title: Pregnancy changes expression in peripheral blood mononuclear cells of Multiple Sclerosis (MS) patients

status: Public on Jan 10 2010

city: Torino

first: Raffaele

last: Calogero

state: To

address: Via Nizza 52

department: Molecular Biotechnology Center

institute: University of Torino

summary:
Background: pregnancy is associated with reduced activity of multiple sclerosis (MS). However, the biological mechanisms underlying this pregnancy-related decrease in disease activity are poorly understood.
This data series contains the subset of data used to generate a MS signature comparing female MS specimens before pregnancy with respect to female MS specimens at ninth month pregnancy.

overall_design:
Subjects were followed in the outpatients clinic and blood was collected before pregnancy and at the following time points during pregnancy: first trimester (gestational age at sampling 12 weeks), second trimester (24 weeks), and third trimester (36 weeks). Before-pregnancy samples were obtained in a treatment-free period and after anticonceptional drug withdrawal.
Peripheral blood mononuclear cells (PBMCs) obtained from 17 women (8 MS patients before pregnancy and 9 MS patients at 9th month pregnancy) were analyzed by oligonucleotide microarray technology.

first: F

last: Gill

first: R

last: Lindberg

first: P

last: Valentino

last: Marnetto

first: S

last: Malucchi

first: A

last: Sala

first: M

last: Capobianco

last: di Sapio

last: Sperli

first: L

last: Kappos

last: Bertolotto


title: Changes in gene expression induced by miR29b in HTM cells

status: Public on Dec 20 2009

city: Durham

first: Coralia

last: Luna

state: NC

address: 2351 Erwin Road

department: Ophthalmology

institute: Duke University

summary: To investigate the role of miR-29b on the changes in expression of genes involved in the synthesis and deposition of extracellular matrix  in human trabecular meshwork cells (HTM).

overall_design: One HTM cell line was transfected by triplicate with microRNA 29b or scramble control. After  3 days  total RNA was extracted.

first: Guorong

last: Li

first: Jianming

last: Qiu

first: David

last: Epstein

first: Pedro

last: Gonzalez


title: Germline Mutations in TMEM127 Confer Susceptibility to Pheochromocytoma

status: Public on Feb 14 2010

city: San Antonio

first: Patricia

middle: L.

last: Dahia

address: 7703 Floyd Curl Dr MC7880

department: Medicine/Cel&Struct Biology

institute: UTHSCSA

summary: Pheochromocytomas, catecholamine-secreting tumors of neural crest origin, are frequently hereditary. However, the molecular basis of the majority of these tumors is unknown. We identified the transmembrane-encoding gene TMEM127 on chromosome 2q11 as a new pheochromocytoma susceptibility gene. In a cohort of 103 samples, we detected truncating germline TMEM127 mutations in approximately 30% of familial tumors and about 3% of sporadic-appearing pheochromocytomas without a known genetic cause. The wild-type allele was consistently deleted in tumor DNA, suggesting a classic mechanism of tumor suppressor gene inactivation. Pheochromocytomas with mutations in TMEM127 are transcriptionally related to tumors bearing NF1 mutations and, similarly, show hyperphosphorylation of mammalian target of rapamycin (mTOR) effector proteins. Accordingly, in vitro gain-of-function and loss-of-function analyses indicate that TMEM127 is a negative regulator of mTOR. TMEM127 dynamically associates with the endomembrane system and colocalizes with perinuclear (activated) mTOR, suggesting a subcompartmental-specific effect. Our studies identify TMEM127 as a tumor suppressor gene and validate the power of hereditary tumors to elucidate cancer pathogenesis.

overall_design: We performed comparative analysis of expression profiling of 125 Pheochromocytomas and paragangliomas with a variety of mutations representatives of distinct pheochromocytoma susceptibility syndromes, sporadic tumors and familial tumors without an identifiable mutation. This dataset combines data from 75 initial tumors and 50 new tumor samples, three samples were processed in duplicate.

first: Yuejuan

last: Qin

first: Li

last: Yao


title: Transcription signature of Multiple Sclerosis

last: Calogero

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

overall_design: Refer to individual Series


title: Imatinib and Nilotinib induced molecular changes in Philadelphia chromosome-positive Chronic Myelogenous Leukemia

status: Public on Dec 19 2009

city: Duesseldorf

first: Daniela

last: Bruennert

address: Universitaetsstr. 1

department: Clinic for Hematology, Oncology and clin. Immunology

country: Germany

institute: University Clinic Duesseldorf

summary:
Chronic myelogenous leukemia (CML) is a malignant stem cell disease characterized by a reciprocal translocation between chromosome 9 and 22. The selective bcr-abl tyrosine-kinase inhibitor Imatinib has become the therapy of choice for patients with newly diagnosed CML including those previously considered candidates for allogeneic haematopoietic stem cell transplantation. The tyrosine-kinase inhibitor Nilotinib is a derivate of Imatinib with higher potency.
To examine the molecular and functional effects of Nilotinib and Imatinib in chronic myelogenous leukemia, we performed gene expression and functional analyses in K562 cells following treatment with the two tyrosine kinase inhibitors.

overall_design: Affymetrix U133A 2.0 microarrays were used to examine the gene expression profile of K562 cells after in vitro treatment with Imatinib (0.5 µM) or Nilotinib (0.05 µM) for 24 hours. Gene expression data of the treated cells were compared with data of untreated cells.

first: Frank

last: Neumann


title: Personalized medicine in psoriasis: developing a genomic classifier to predict histological response to Alefacept

status: Public on Dec 01 2009

first: Mayte

last: Suarez-Farinas

address: 1425 Madison Ave, L2-70C, Box 1077,

institute: Mount SinaiSchool of Medicine

summary:
Alefacept treatment is highly effective in a select group patients with moderate-to-severe psoriasis, and is an ideal candidate to develop systems to predict who will respond to therapy. A clinical trial of 22 patients with moderate to severe psoriasis treated with alefacept (7.5mg weekly i.v. x12 weeks) was conducted in 2002-2003, as a mechanism of action study. Patients were classified as responders or non-responders to alefacept based on histological criteria. Microarray data from PBMCs of 16 of these patients was analyzed to generate a treatment response classifier. We used a discriminant analysis method that performs sample classification from gene expression data, via nearest shrunken centroid method''.  A disease response classifier using 23 genes was created to accurately predict response to alefacept (12.3% error rate in favour of responders). This preliminary study may provide a useful tool to predict response of psoriatic patients with alefacept.

Keywords: class prediction

overall_design: Microarray data from 16  patients was analyzed to generate a treatment response classifier. We used a discriminant analysis method that performs sample classification from gene expression data, via nearest shrunken centroid method.

first: James

middle: G

last: Krueger


title: Chronic myelogenous leukemia hematopoietic stem cells

status: Public on Jan 05 2010

city: Modena

first: Rossella

last: Manfredini

address: Via Gottardi 100

department: Life Sciences

institute: Centre for Regenerative Medicine

summary:
We show the molecular and functional characterization of a novel population of lineage-negative CD34-negative (Lin- CD34-) hematopoietic stem cells (HSCs) from chronic myelogenous leukemia (CML) patients at diagnosis. Molecular caryotyping and quantitative analysis of BCR/ABL transcript demonstrated that about one third of CD34- was leukemic. CML CD34- cells showed kinetic quiescence and limited clonogenic capacity. However, stroma-dependent cultures and cytokines induced CD34 expression on some HSCs, cell cycling, acquisition of clonogenic activity and increased expression of BCR/ABL transcript. CML CD34- cells showed an engraftment rate in immunodeficient mice similar to that of CD34+ cells. Gene expression profiling revealed the down-regulation of cell cycle arrest genes together with genes involved in antigen presentation and processing, while the expression of angiogenic factors was strongly up-regulated when compared to normal counterparts. Flow cytometry analysis confirmed the significant down-regulation of HLA class I and II molecules in CML CD34-cells. Increasing doses of imatinib mesilate (IM) did not affect fusion transcript levels, BCR-ABL kinase activity and the clonogenic efficiency of CML CD34- cells as compared to leukemic CD34+cells.
Thus, we identified in CML a novel CD34- leukemic stem cell subset with peculiar molecular and functional characteristics which may be a potential target for CML therapeutics.

overall_design:
Leukemic cells were obtained from 12 chronic phase Ph+ CML  patients at diagnosis and before treatment. Normal samples were leukapheresis products from 12 healthy stem cell donors receiving recombinant human granulocyte colony-stimulating factor (G-CSF; Lenograstim, Sanofi-Aventis, Milan, Italy). The protocol was approved by the ethical committee of the University Hospital and each patient/donor gave written informed consent. Hemopoietic stem/progenitor cell purification and phenotypic analyses were performed as previously described (Lemoli et al, Br J Haematol, 2003; Lemoli RM et al., Blood, 1997). Aliquots of sorted Lin-CD34-, Lin-CD34+ and Lin+CD34+  were reanalyzed by FacScan (Becton Dickinson, Franklin Lakes, NJ) to assess their purities.  Total cellular RNA was extracted from 0.5x105 cells of each sample using RNeasy Micro kit (Qiagen, Valencia, CA) following the protocol supplied by the manufacturer. Disposable RNA chips (Agilent RNA 6000 Nano LabChip kit, Agilent Technologies, Waldbrunn, Germany) were used to determine the concentration and purity/integrity of RNA samples using Agilent 2100 Bioanalyzer. RNAs originating from 12 normal donors or from 12 CML patients were pooled in order to obtain at least 2 mg per sample.
One-cycle target labeling assays, as well as the Affymetrix Human HG-U95Av2 GeneChip arrays hybridization, staining, and scanning, were performed, using Affymetrix standard protocols (Affymetrix, Santa Clara,CA).


title: Expression data from human melanoma cell lines treated or not with inflammatory cytokines

status: Public on Dec 11 2009

city: Brussels

first: Nicolas

last: van Baren

address: 74 avenue Hippocrate, UCL7459

country: Belgium

institute: Ludwig Institute for Cancer Research

summary:
Melanomas are often infiltrated by activated inflammatory cells. Thus, melanoma cells are very likely stimulated by inflammatory cytokines.
In order to assess the impact of common inflammatory cytokines, we investigated the gene expression profile of melanoma cell lines before and after cytokine treatment in vitro.

overall_design: 5 human melanoma cell lines were treated with either IFN-α 1,000 U/ml, IFN-γ 100 U/ml or TNF-α 10 ng/ml for 72 hours, or were left untreated. We analyzed their expression profile with Affymetrix expression arrays.

first: Olga

last: Kholmanskikh

first: Francis

last: Brasseur

first: Etienne

last: De Plaen


title: Pregnancy changes expression in peripheral blood mononuclear cells of healthy donors

first: Raffaele

summary:
Background: pregnancy is associated with reduced activity of multiple sclerosis (MS). However, the biological mechanisms underlying this pregnancy-related decrease in disease activity are poorly understood.
This data series contains the subset of data used to generate a healthy donors signature comparing female healthy specimens before pregnancy with respect to female healthy specimens at ninth month pregnancy.

overall_design:
Subjects were followed in the outpatients clinic and blood was collected before pregnancy and at the following time points during pregnancy: first trimester (gestational age at sampling 12 weeks), second trimester (24 weeks), and third trimester (36 weeks). Before-pregnancy samples were obtained in a treatment-free period and after anticonceptional drug withdrawal.
Peripheral blood mononuclear cells (PBMCs) obtained from 11 women (7 healthy donors before pregnancy and 4 healthy donors at 9th month pregnancy) were analyzed by oligonucleotide microarray technology.

first: F

last: Gilli

first: R

last: Lindberg

first: P

last: Valentino

last: Marnetto

first: S

last: Malucchi

first: A

last: Sala

first: M

last: Capobianco

last: di Sapio

last: Sperli

first: L

last: Kappos

last: Bertolotto


title: Loss of Syk in normal breast cells

status: Public on Nov 20 2009

city: Washington

first: Junfeng

last: Sun

state: DC

address: Suite 180, 4000 Reservior Rd NW

institute: Georgetown Univ

summary:
Loss of Syk in normal breast cells in vivo and in vitro:  gene expression and phenotypic switch to stem-cell like with induction of invadopodia

Morphological and gene microarray analysis following Syk knockdown in MCF10A reveals a loss of luminal and differentiated epithelial features with epithelial to mesenchymal transition and a gain in invadopodia and stem/ progenitor cell marker expression.

These results support the role of Syk in limiting normal stem/progenitor proliferation and invasion during morphogenesis, and support the role of Syk as a critical tumor suppressor for breast cancer.

overall_design: Global gene expression profiles were measured using Affymetrix U133A 2.0 microarrays in MCF10A cell lines (control and Syk siRNA knockdown cells on collagen gels or plastic).

first: You

last: Sung

first: Xuehua

last: Xu

first: Duane

last: Mueller

first: Kinza

last: Sentissi

first: Peter

last: Johnson

first: Elana

last: Urbach

first: Francoise

last: Seillier-Moiseiwitsch

first: Michael

first: Susette


title: Expression Data From HCMV-Infected Human Monocytes 48 Hours Post-Infection: Role of PI(3)K

status: Public on Jan 07 2010

city: Shreveport

first: Andrew

middle: D.

last: Yurochko

state: LA

address: 1501 Kings Highway

department: Microbiology & Immunology

institute: LSUHSC-S

summary:
Human cytomegalovirus (HCMV) induces pro-inflammatory monocytes following infection and we have evidence that phosphatidylinositol 3-kinase [PI(3)K] is a key mediator in this activation. To begin to address how this signalling pathway is responsible for the functional changes in infected monocytes, we examined the role this pathway played in the transcriptome of infected monocytes. Global transcriptional profiling using cDNA microarrays revealed a significant number of genes were regulated in a PI(3)K-dependent manner, identifying this pathway as a key cellular control point in the conversion of monocytes to an activated pro-inflammatory state following HCMV infection.

Keywords: Disease state

overall_design: To begin to globally define how PI(3)K is involved in the HCMV-induced changes in monocyte function, we performed a transcriptome analysis in the presence of an inhibitor to the PI(3)K signalling pathway. Specifically, a cDNA microarray containing 12,626 unique probe sets was utilized to assess the modulation of the monocyte transcriptome at 48 hours post-infection in the presence of the pharmacological agent LY294002 (LY), a PI(3)K inhibitor. A total of 2 replicates from HCMV-infected monocytes, 2 replicates from LY-pretreated infected monocytes, and 2 replicates from mock-infected monocytes were analyzed in this study.

first: Elizabeth

last: Bivins-Smith

last: Smith

middle: D


title: Transcriptome analysis identifies molecular effectors of unconjugated bilirubin in human neuroblastoma SH-SY5Y cells: 4h

status: Public on Nov 19 2009

city: Trieste

first: Paola

last: Roncaglia

address: via Bonomea 265

institute: SISSA/ISAS (International School for Advanced Studies)

summary: The deposition of unconjugated bilirubin (UCB) in selected regions of the brain results in irreversible neuronal damage, or Bilirubin Encephalopathy (BE). Although UCB impairs a large number of cellular functions, the basic mechanisms of neurotoxicity have not yet been fully clarified. While cells can accumulate UCB by passive diffusion, cell protection may involve multiple mechanisms including the extrusion of the pigment as well as pro-survival homeostatic responses that are still unknown. The effects of UCB treatment to SH-SY5Y neuroblastoma cell line were examined by high-density oligonucleotide microarrays. 230 genes were induced after 24 hours. A Gene Ontology (GO) analysis showed that a large group of UCB-induced genes were components of the ER stress response. Independent experimental validation of molecular events crucial for the ER stress response is presented. The results show that UCB exposure induces the ER stress response as a major intracellular homeostatic response in neuroblastoma cells in vitro. Our finding may provide valuable information for new therapeutic strategies in the treatment of BE.

overall_design: We used high-density oligonucleotide microarrays to analyze the gene expression profile of human neuroblastoma SH-SY5Y cells upon UCB treatment. Three replicates of UCB-treated cells were analyzed. As controls, we used three replicates of the same cells treated with DMSO only (the solvent used for UCB treatment).


title: Transcription signature of Multiple Sclerosis in peripheral blood mononuclear cells.

first: Raffaele

last: Calogero

summary:
Background: pregnancy is associated with reduced activity of multiple sclerosis (MS). However, the biological mechanisms underlying this pregnancy-related decrease in disease activity are poorly understood.
This data series contains the subset of data used to generate a MS signature comparing female healthy specimens with respect to MS patients

overall_design:
Subjects were followed in the outpatients clinic and blood was collected before pregnancy and at the following time points during pregnancy: first trimester (gestational age at sampling 12 weeks), second trimester (24 weeks), and third trimester (36 weeks). Before-pregnancy samples were obtained in a treatment-free period and after anticonceptional drug withdrawal.
Peripheral blood mononuclear cells (PBMCs) obtained from 15 women (8 MS patients and 7 healthy controls) were analyzed by oligonucleotide microarray technology.

first: F

last: Gilli

first: RLP

last: Lindberg

first: P

last: Valentino

last: Marnetto

first: S

last: Malucchi

last: Sala

last: Capobianco

first: RA


title: Transcriptome analysis identifies molecular effectors of unconjugated bilirubin in human neuroblastoma SH-SY5Y cells


title: Expression profile after β-TrCP inhibition and androgen ablation in prostate cancer cells

status: Public on Nov 24 2009

city: Jerusalem

first: Gady

last: Cojocaru

address: Kiryat Hadassah

institute: hadassah medical center

summary: We examined gene expression of LAPC4 cells after knocking down β-TrCP, androgen ablation, or the combined treatments compared to non treated cells.

overall_design: Using lentiviral vectors, we stably infected LAPC4 cells with specific shRNA targeted for both β-TrCP isoforms (i.e. 1 and 2) or GFP vector as control. To establish androgen ablation we treated the cells with charcoal stripped serum (CSS).

first: Udi

last: Gluschnaider

last: Cojocaro


title: Effective treatment of psoriasis with etanercept is linked to suppression of IL17 signaling, not immediate response TNF

status: Public on Oct 30 2009

city: New York

first: Mayte

last: Suarez-Farinas

summary:
The success of TNF inhibitors for treatment of psoriasis and other inflammatory diseases was previously attributed to blockade of innate immunity.  In a clinical trial using etanercept TNF blocking agent to treat psoriasis vulgaris, we used affymetrix gene arrays to analyze broad gene profiles in lesional skin at multiple timepoints during drug treatment (baseline, and weeks 1, 2, 4 and 12) compared to non-lesional skin.  This analysis created a temporal model of TNF-dependent gene regulation that informs molecular mechanisms of TNF-mediated inflammation.  We identified four gene clusters that were differentially down-modulated during etanercept treatment: the cluster down-regulated most rapidly contained mostly dendritic cell activation genes.  Culturing human keratinocytes with TNF, IFNg and IL-17 generated a list of keratinocyte genes regulated by each cytokine.  The IL-17 pathway genes were strongly down-modulated early, whereas IFNg pathway genes were not down-modulated until final disease resolution at week 12.  Finally, we show that TNF blockade rapidly inhibits IL-12/IL-23 p40 subunit expression, and that p40 neutralization inhibits psoriatic dermal émigré-mediated Th17 polarization.  We hypothesize that etanercept inhibits myeloid dendritic cell production of IL-23, a Th17 survival cytokine, resulting in rapid downregulation of IL-17 pathway genes.  This data links effects of TNF blockade on the innate immune system with the adaptive immune system.

Keywords: time-course experiment

overall_design: In this study 15 patients with moderate-to-severe psoriasis were given 50mg of etanercept (Amgen) biweekly for 12 weeks. And analyzed using gene array on mRNA extracted from tissue collected at each biopsy time point (non-lesional Time: 0; lesional Time: 0, weeks 1, 2, 4, and 12).  Patients were stratified as “responders” or “non-responders” based on whether or not they achieved histologic disease resolution by week 12 of etanercept treatment (decreased epidermal thickening, normalization of proliferation marker Ki67, and loss of differentiation marker K16).

last: Suárez-Fariñas

first: James

last: Krueger


title: Transcriptome analysis identifies molecular effectors of unconjugated bilirubin in human neuroblatoma SH-SY5Y cells: 24h

summary: The deposition of unconjugated bilirubin (UCB) in selected regions of the brain results in irreversible neuronal damage, or Bilirubin Encephalopathy (BE). Although UCB impairs a large number of cellular functions, the basic mechanisms of neurotoxicity have not yet been fully clarified. While cells can accumulate UCB by passive diffusion, cell protection may involve multiple mechanisms including the extrusion of the pigment as well as pro-survival homeostatic responses that are still unknown. The effects of UCB treatment to SH-SY5Y neuroblastoma cell line were examined by high density oligonucleotide microarrays. 230 genes were induced after 24 hours. A Gene Ontology (GO) analysis showed that a large group of UCB-induced genes were components of the ER stress response. Independent experimental validation of molecular events crucial for the ER stress response is presented. The results show that UCB exposure induces ER stress response as major intracellular homeostatic response in neuroblastoma cells in vitro. Our finding may provide valuable information for new therapeutic strategies in the treatment of BE.

overall_design: We used high density oligonucleotide microarrays to analyze the gene expression profile of human neuroblastoma SH-SY5Y cells upon UCB treatment. Three replicates of UCB-treated cells were analyzed. As controls, we used three replicates of the same cells treated with DMSO only (the solvent used for UCB treatment).


title: Profiling of immortalized human lung epithelial cells following oncogenic KRAS expression and TBK1 suppression

status: Public on Nov 04 2009

city: La Jolla

first: Pablo

last: Tamayo

state: CA

address: 3855 Health Sciences Drive MC 0658

department: Moores Cancer Center

institute: UCSD

summary:
The purpose of the dataset is to analyze expression of genes induced by KRAS and regulated by TBK1
The proto-oncogene KRAS is mutated in a wide array of human cancers, most of which are aggressive and respond poorly to standard therapies. Although the identification of specific oncogenes has led to the development of clinically effective, molecularly targeted therapies in some cases, KRAS has remained refractory to this approach. An alternative strategy for targeting KRAS is to identify gene products that, when suppressed or inhibited, result in cell death only in the presence of an oncogenic allele. Here we have used systematic RNA interference (RNAi) to detect synthetic lethal partners of oncogenic KRAS and found that the non-canonical IkB kinase, TBK1, was selectively essential in cells that harbor mutant KRAS. Suppression of TBK1 induced apoptosis specifically in human cancer cell lines that depend on oncogenic KRAS expression. In these cells, TBK1 activated NF- B anti-apoptotic signals involving cREL and BCL-XL that were essential for survival, providing mechanistic insights into this synthetic lethal interaction. These observations identify TBK1 as a potential therapeutic target in KRAS mutant tumors and establish a general approach for the rational identification of co-dependent pathways in cancer.

overall_design: Knock out of TBK1 in the contect of KRAS activation (mutant) and control (WT)

first: David

last: Barbie


title: Influence of Sex on Gene Expression in Human Corneal Epithelial Cells

status: Public on Nov 13 2009

city: Boston

last: Sullivan

address: 20 Staniford Street

institute: Schepens Eye Research Institute

summary:
Objectives. Sex-associated differences have been identified in the anatomy, physiology and pathophysiology of the human cornea. We hypothesize that many of these differences are due to fundamental variations in gene expression. Our objective in this study was to determine whether such differences do exist in human corneal epithelial cells both in vivo and in vitro. Methods. Human corneal epithelial cells were isolated from the corneoscleral rims of male and female donors. Cells were processed either directly for RNA extraction, or first cultured in phenol red-free keratinocyte serum free media. The RNA samples were examined for differentially expressed mRNAs by using of CodeLink Bioarrays and Affymetrix GeneChips. Data were analyzed with GeneSifter.Net software. Results. Our results demonstrate that sex significantly influences the expression of over 400 genes in human corneal epithelial cells in vivo. These genes are involved a broad spectrum of biological processes, molecular functions and cellular components, such as metabolic processes, DNA replication, cell migration, RNA binding, oxidoreductase activity and nucleoli. We also identified significant, sex-related effects on gene expression in human corneal epithelial cells in vitro. However, with few exceptions (e.g. X- and Y-linked genes), these sex-related differences in gene expression in vitro were typically different than those in vivo. Conclusions. Our findings support our hypothesis that sex-related differences exist in the gene expression of human corneal epithelial cells.

Keywords: Female vesus Male.

overall_design: Gene expression in corneal epithelial cells was compared between females and males. RNA was isolated from female (n=3) and male (n= 3) donors. The RNA was then used to generate labeled cRNA, which was then hybridized to CodeLink Uniset Human Bioarrays and Affymetrix GeneChip Human Genome U133A 2.0 Arrays.  CEL files for Affymetrix GeneChip Human Genome U133A 2.0 Arrays are lost.

first: T

last: Suzuki

last: Richards

last: Liu

first: R

middle: V

last: Jensen

first: D


title: TpoR Agonist signaling in AML primary cells

status: Public on Oct 22 2009

city: Philadelphia

first: Anna

last: Kalota

state: PA

address: 421 Curie Blvd

department: Medicine, Hem/Onc

institute: University of Pennsylvania

summary: Biologic characterization of SB-559457 (SB), a non-peptidyl hydrazone class of thrombopoietin receptor (Mpl) agonist, revealed toxicity towards human leukemia cells. Anti-proliferative effects followed by significant, non-apoptotic, cell death within 72 hours occurred in 24/26 AML, 0/6 ALL, and 3/6 CML patient samples exposed to SB, but not recombinant human thrombopoietin (rhTpo), in liquid suspension culture. Further investigation revealed increased phosphorylation of p70S6/S6 kinases in SB, but not in rhTpo, treated cells. Expression profiling of cells exposed to SB vs rhTpo revealed statistically significant, ~2-fold changes in GAPDH and REDD1 gene expression, confirmed by QRT-PCR. These genes, induced in energy or hypoxia stressed cells, have been implicated in cell death pathways, and may provide important clues to the mechanism of SB induced, leukemic cell death. These results suggest that nonpeptidyl, hydrazone class Mpl agonists may be clinically useful anti-leukemic agents by virtue of their combined thrombopoietic and anti-leukemic effects.

overall_design: Primary cells collected from 5 patients with acute myeloid leukemia (AML) were stimulated stimulated with MPL receptor agonist (SB559457) or recombinant human Tpo (rhTpo) for 6 hours. Next, RNA was isolated from these cells. RNA was used for gene profile analyis to compare genes regulated in AML cells by SB559457 verus rhTpo. For each sample half of the cells were stimulated with Mpl agonist and half with rhTpo.


title: Transcriptome analysis identifies molecular effectors of unconjugated bilirubin in human neuroblastoma SH-SY5Y cells: 1h

first: Paola

last: Roncaglia


title: A caspase-independent necrotic death is activated by isopeptidase inhibitor G5 in apoptosis-resistant glioblastoma cells

status: Public on Oct 13 2009

summary:
The regulation of necrotic death and its relevance in anti-cancer therapy are largely unknown. Here we have investigated the pro-apoptotic and pro-necrotic activities of two ubiquitin-proteasome system inhibitors (UPSIs): bortezomib and G5. The present study points out that the glioblastoma cell lines U87MG and T98G are useful models to study the susceptibility to apoptosis and necrosis in response to UPSIs. U87MG cells are resistant to apoptosis induced by bortezomib and G5 but susceptible to necrosis induced by G5. On the opposite T98G cells are susceptible to apoptosis induced by both inhibitors but show some resistance to G5-induced necrosis. By comparing the transcriptional profiles of the two cell lines, we have found that the resistance to G5-induced necrosis could arise from differences in glutathione synthesis/utilization and in the microenvironment. In particular collagen IV, which is highly expressed in T98G cells, and fibronectin, whose adhesive function is counteracted by tenascin-C in U87MG cells, can restrain the necrotic response to G5. Collectively, our results provide an initial characterization of the molecular signals governing cell death by necrosis in glioblastoma cell lines.

Keywords: cell line comparison

overall_design: Gene expression profiling was evaluated from 3 replicates each of T98G and U87MG cells.


title: Expression Profiling of a Mouse Xenograft Model of “Triple-Negative” Breast Cancer Brain Metastases With Vorinostat

status: Public on Oct 14 2009

city: Bethesda

first: Sean

last: Davis

address: 37 Convent Drive, Room 6138

institute: National Cancer Institute

summary:
Gene Expression Profiling of a Mouse Xenograft Model of “Triple-Negative” Breast Cancer Brain Metastases With and Without Vorinostat Treatment.
Purpose: As chemotherapy and molecular therapy improve the systemic survival of breast cancer patients, the incidence of brain metastases increases. Few therapeutic strategies exist for the treatment of brain metastases because the blood-brain barrier severely limits drug access. We report the pharmacokinetic, efficacy, and mechanism of action studies for the histone deactylase inhibitor vorinostat (suberoylanilide hydroxamic acid) in a preclinical model of brain metastasis of triple-negative breast cancer.
Experimental Design: The 231-BR brain trophic subline of the MDA-MB-231 human breast cancer cell line was injected into immunocompromised mice for pharmacokinetic and metastasis studies. Pharmacodynamic studies compared histone acetylation, apoptosis, proliferation, and DNA damage in vitro and in vivo.
Results: Following systemic administration, uptake of [14C]vorinostat was significant into normal rodent brain and accumulation was up to 3-fold higher in a proportion of metastases formed by 231-BR cells. Vorinostat prevented the development of 231-BR micrometastases by 28% (P = 0.017) and large metastases by 62% (P < 0.0001) compared with vehicle-treated mice when treatment was initiated on day 3 post-injection. The inhibitory activity of vorinostat as a single agent was linked to a novel function in vivo: induction of DNA double-strand breaks associated with the down-regulation of the DNA repair gene Rad52.
Conclusions: We report the first preclinical data for the prevention of brain metastasis of triple-negative breast cancer. Vorinostat is brain permeable and can prevent the formation of brain metastases by 62%. Its mechanism of action involves the induction of DNA double-strand breaks, suggesting rational combinations with DNA active drugs or radiation.

overall_design: We performed gene expression profiling on metastases from vehicle- or vorinostat-treated mice to determine if alterations in gene expression were observable that were consistent with the phenotypes observed. Brain metastases from five vehicle-treated mice and six 150 mg/kg vorinostat-treated mice were procured by laser capture microdissection. RNA was extracted from the captured tumor cells from each brain and two rounds of linear amplification was done. The amplified RNA from each mouse was processed separately through microarray hybridization and analysis.

first: Patricia

last: Steeg

first: Paul

last: Meltzer

first: Diane

last: Palmieri


title: Altered gene expression in Werner & Bloom syndromes is associated with sequences having G-quadruplex forming potential

status: Public on Dec 10 2009

first: Jay

last: Johnson

address: 422 Curie Blvd

department: Pathology and Laboratory Medicine

summary: The human Werner and Bloom syndromes (WS and BS) are caused by deficiencies in the WRN and BLM RecQ helicases, respectively.  WRN, BLM and their S. cerevisiae homologue Sgs1, are particularly active in vitro in unwinding G-quadruplex DNA (G4-DNA), a family of non-canonical nucleic acid structures formed by certain G-rich sequences.  Recently, mRNA levels from loci containing potential G-quadruplex-forming sequences (PQS) were found to be preferentially altered in sgs1 mutants, suggesting that G4-DNA targeting by Sgs1 directly affects gene expression.  Here, we extend these findings to human cells.  Using microarrays to measure mRNAs obtained from human fibroblasts deficient for various RecQ family helicases, we observe significant associations between loci that are upregulated in WS or BS cells and loci that have PQS.  No such PQS associations were observed for control expression datasets, however.  Furthermore, upregulated genes in WS and BS showed no or dramatically reduced associations with sequences similar to PQS but that have considerably reduced potential to form intramolecular G4-DNA.  These findings indicate that, like Sgs1, WRN and BLM can regulate transcription globally by targeting G4-DNA.

overall_design:
Cell culture conditions and media
Human fibroblast cell strains (WS: AG05229, AG12795, AG12797; BS: GM02932, GM03402, GM16891; RTS: AG18371, AG18375, AG05013; Normal/Wild-type: AG04054, AG06310, AG09975) were obtained from the Coriell Repository (Camden, NJ), from donors matched for gender and of similar ages, and were at similar passage levels.  Cells were cultured in MEM supplemented with Earle’s salts, 20% fetal bovine serum, 1x penicillin/streptomycin, and 1x fungizone in 3% O2 at 37oC and harvested for RNA extraction during active growth and at ~ 80% confluence.

GeneChip microarray expression
Total RNA from the 12 fibroblast cell strains was isolated by extraction with TRIzol (Invitrogen) and purified using the RNeasy system (Qiagen).  Total RNA was amplified by in vitro transcription using the Ovation RNA Amplification System V2 (NuGen).  The resultant cDNA was fragmented and labeled using the FL-Ovation cDNA Biotin Module V2 (NuGen), and then purified using QIAquick columns (Qiagen), as specified by the Ovation System manual.  Labeled probe was hybridized to Affymetrix U133A 2.0 GeneChips, and ultimately scanned using an Axon GenePix array scanner.

Statistical analysis of microarray expression experiment
The output files were normalized by Robust Multiarray Average (RMA), using the R package GCRMA and gene expression levels were log2-transformed.

first: Kajia

last: Cao

last: Ryvkin

first: Li-San

last: Wang

first: Frederick

middle: B


title: Expression data from hESCs differentiated in the presence or absence of nicotinamide

status: Public on Oct 02 2009

first: Reba

last: Condiotti

department: Goldyne Savad Institute of Gene Therapy

institute: Hadassah Medical Center

summary: Human ESCs are pluripotent cells that have the capacity of self renewal for a prolonged period in vitro, and can differentiate into derivatives of all three primary germ layers: endoderm, mesoderm and ectoderm. Human ESCs are responsive to a wide range of factors in vitro that can direct their differentiation into specific cell types. We analyzed the effect of nicotinamide (NIC) on differentiation of hESCs in vitro. CEL file for GSM424319 is unavailable.

overall_design: hESCs were cultured as free-floating clusters in the presence of 10% FCS for 4 weeks in the presence or absence of nicotinamide.

first: Etti

last: Ben-Shushan

first: Gideon

last: Rechavi


title: FGF-2 ENHANCES PROLIFERATION AND DELAYS LOSS OF CHONDROGENIC POTENTIAL IN HUMAN ADULT BONE MARROW-DERIVED MSCs

status: Public on Oct 29 2009

city: Cleveland

first: Luis

last: Solchaga

address: 2103 Cornell Road

institute: Case Western Reserve University

summary:
Fibroblast growth factor-2 delays the loss of chondrogenic potential in adult bone marrow-derived mesenchymal stem cells
We compared human mesenchymal stem cells (hMSCs), expanded long-term with and without fibroblast growth factor (FGF) supplementation, with respect to their proliferation rate, and ability to differentiate along the chondrogenic pathway in vitro. hMSCs expanded in FGF-supplemented medium proliferated more rapidly than those expanded under control conditions. Aggregates of FGF-treated cells exhibited chondrogenic differentiation at passages 1 through 7, although, in some of the preparations, chondrogenic differentiation was somewhat diminished after seventh passage. Aggregates made with control cells differentiated along the chondrogenic lineage at first passage but exhibited only marginal chondrogenic differentiation after four passages and failed to form cartilage after seven passages. Microarray analysis of gene expression identified 334 transcripts that were differentially expressed in fourth passage control cells which had reduced chondrogenic potential compared to fourth passage FGF-treated cells which retained this differentiation capacity and 243 transcripts that were differentially expressed when comparing them to first passage control cells which were also capable of differentiating into chondrocytes. The intersection of these analyses yielded 49 transcripts that were differentially expressed in cells that exhibited chondrogenic differentiation in vitro compared to cells that did not. These preliminary data must now be validated to verify whether the different gene expression profiles translate into functional differences. These findings suggest that care should be exercised when extensively expanding these cells for cartilage tissue engineering applications.

Keywords: time course and treatment

overall_design: hMSCs from three different donors were expanded for up to seven passages with or without FGF supplementation. At the end of first, fourth and seventh passages the chondrogenic potential and gene expression progfiles were analyzed.

first: Kitsie

last: Penick

first: Victor

last: Goldberg

first: Arnold

middle: I

last: Caplan

first: Jean

middle: F

last: Welter


title: Profiling genes induced by oncogenic KRAS (G12V) and genes regulated by TBK1, a synthetic lethal partner of KRAS

status: Public on Oct 21 2009

first: Pablo

last: Tamayo

summary:
The proto-oncogene KRAS is mutated in a wide array of human cancers, most of which are aggressive and respond poorly to standard therapies. Although the identification of specific oncogenes has led to the development of clinically effective, molecularly targeted therapies in some cases, KRAS has remained refractory to this approach. An alternative strategy for targeting KRAS is to identify gene products that, when suppressed or inhibited, result in cell death only in the presence of an oncogenic allele. Here we have used systematic RNA interference (RNAi) to detect synthetic lethal partners of oncogenic KRAS and found that the non-canonical IkB kinase, TBK1, was selectively essential in cells that harbor mutant KRAS. Suppression of TBK1 induced apoptosis specifically in human cancer cell lines that depend on oncogenic KRAS expression. In these cells, TBK1 activated NF- B anti-apoptotic signals involving cREL and BCL-XL that were essential for survival, providing mechanistic insights into this synthetic lethal interaction. These observations identify TBK1 as a potential therapeutic target in KRAS mutant tumors and establish a general approach for the rational identification of co-dependent pathways in cancer.

This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Expression data from Trk-overexpressing NB cell line SY5Y

status: Public on Oct 05 2009

city: Essen

first: Alexander

last: Schramm

address: Hufelandstr. 55

department: Childrens Hospital

institute: University Hospital Essen

summary: Neurotrophin receptors play a pivotal role in the biology and prognosis of neuroblastoma (NB). While expression of TrkA/NTRK1 confers a good prognosis to NB patients, expression of TrkB/NTRK2 is associated with unfavorable outcome. We have transfected the neurotrophin-receptor null cell line SY5Y with either full-length TrkA or TrkB and performed transcriptional profiling to analyse the effects of Trk-expression without activation and in a time course after activation.

overall_design: SY5Y transfected with either full length TrkA- or TrkB cDNA or vector control were cultivated in the absence or presence of their ligands NGF and BDNF, respectively. At the indicated time points cells were harvested and analysed using Affymetrix U95Av2 chips.

first: Johannes

middle: H

last: Schulte


title: PIF modulated gene expression in human Peripheral Blood Mononuclear Cell (PBMC)

status: Public on Sep 26 2009

city: Cherry Hill,

first: Eytan

middle: R

last: Barnea

state: NJ

address: 1697 Lark Lane

institute: BioIncept LLC

summary:
Pregnancy is an immune paradox: where the mother accepts a semi-allograft/allograft (donor embryo), while avoiding maternal immune suppression. Indeed, impaired development of maternal immune tolerance towards the conceptus may be an important cause of implantation failure and pregnancy loss. Successful implantation rates for allogenic donor embryos are unexpectedly high, while abnormal embryos may be unable to initiate maternal tolerance, suggesting that embryo-specific compounds can play a vital role in this process. Both a tolerant T-suppressor (TH2), and inflammatory, T-helper 1 (TH1) cytokine profiles are an integral part of pregnancy and the viable embryo/fetus plays a critical role in creating this delicate balance. Maternal recognition of pregnancy occurs prior to implantation. There were several embryo derived factors reported. However, we previously found that preimplantation factor (PIF) is distinct from the other non-pregnancy-specific embryo-derived factors. Herein we aimed to characterize PIF and examine its role in early pregnancy events by observing the isolated novel peptide's effect on human Peripheral Blood Mononuclear Cell (PBMC).
We did affymetrix, GeneChip Human Genome U133 plus 2.0 array on RNA isolated from normal human PBMC cultured for 24hrs in the presence or absence of PIF to study the genes modulated by PIF.

overall_design: Blood was collected from a male volunteer and PBMC were separated using the Ficoll Hypaque method. Cells were cultured in RPMI culture media for 24 hours in four different groups in duplicate viz., Media alone, Media + 50 nM PIF, Media + 50nM PIF +CD3 (10µg/ml)+ CD28 (10µg/ml) and Media + CD3 (10µg/ml) +CD28 (10µg/ml). At the end of incubation cells were removed from the media, washed and collected in RNAlater (Qiagen) and transferred to Biocodon Sciences (Tx, USA) for Gene array analysis on dry ice. Samples were analyzed using the Agilent Bioanalyzer Process found to be adequate. Each 100 µl contained 2.7-5.4 µg RNA (Nanodrop analyzer). Each sample was tested integrity. RNA from duplicate samples was combined and array was performed on an Affymetrix chip (Human Genome U 133 Plus 2.0 Array). Subsequently, arrays were processed, scanned, data was extracted and statistical evaluation was carried out comparing the various groups. Data generated were categorized as robust changes or mild changes based on differences in expression.

first: David

last: Kirk

first: Sivakumar

last: Ramu

first: Graciela

last: Krikun

first: Benjamin

last: Rivnay

first: Thillainayagam

last: Sathiyaseelan

first: Chris

last: Stamatkin

first: S

last: Albini

first: Roumen

last: Roussev

first: Chandrakant

last: Tayade

first: Carolyn

last: Coulam

first: Michael

last: Paidas


title: Differential gene expression patterns of EBV infected EBNA-3 positive and negative human B lymphocytes

status: Public on Sep 05 2009

city: Erlangen

first: Roland

last: Lang

address: Wasserturmstr. 3-5

institute: University Hospital Erlangen

summary: The genome of Epstein-Barr virus (EBV) encodes 86 proteins but only a limited set is expressed in EBV-growth transformed B cells, termed lymphoblastoid cell lines (LCLs). These cells proliferate via the concerted action of EBV nuclear antigens (EBNAs) and latent membrane proteins (LMPs), some of which are rate limiting to establish a stable homeostasis of growth promoting and anti-apoptotic activities. We show here that EBV mutants, which lack the EBNA-3A gene, are impaired but can still initiate cell-cycle entry and proliferation of primary human B cells in contrast to an EBNA-2-deficient mutant virus. Surprisingly and in contrast to previous reports, these viral mutants are attenuated in growth transformation assays but give rise to permanently growing EBNA-3A negative B cell lines which exhibit reduced proliferation rates and elevated levels of apoptosis. Expression profiles of EBNA-3A deficient LCLs are characterized by 129 upregulated and 167 downregulated genes, which are significantly enriched for genes involved in apoptotic processes or cell cycle progression like the tumor suppressor gene p16/INK4A or might contribute to essential steps in the viral life cycle. In addition EBNA-3A cellular target genes remarkably overlap with previously identified targets of EBNA-2.

overall_design: Comparison of gene expression profiles from lymphoblastoid cell lines (LCLs) infected with EBNA-3A positive or negative EBV. LCLs were generated from three donors. A total of five EBNA-3A positive and nine negative LCLs were analysed.

first: Bettina

last: Kempkes

first: Marie

middle: L

last: Hertle

first: Jörg

last: Mages


title: Effect of Plasmodium falciparum infected erythrocytes on primary human brain microvascular endothelial cell

status: Public on Nov 10 2009

city: Baltimore

first: Abhai

middle: K

last: Tripathi

address: 615 N Wolfe St

department: Mol Microbiol and Immunology, SPH and Neurology, SOM

institute: Johns Hopkins University

summary: Cerebral malaria is a severe multifactorial condition associated with the interaction of high numbers of infected erythrocytes to human brain endothelium without invasion into the brain. The result is coma and seizures with death in more than 20% of cases. Because the brain endothelium is at the interface of these processes, we investigated the global gene responses of human brain endothelium after the interaction with Plasmodium falciparum–infected erythrocytes with either high- or low-binding phenotypes. The most significantly up-regulated transcripts were found in gene ontology groups comprising the immune response, apoptosis and antiapoptosis, inflammatory response, cell-cell signaling, and signal transduction and nuclear factor B (NF-B) activation cascade. The proinflammatory NF-B pathway was central to the regulation of the P falciparum–modulated endothelium transcriptome. The proinflammatory molecules, for example, CCL20, CXCL1, CXCL2, IL-6, and IL-8, were increased more than 100-fold, suggesting an important role of blood-brain barrier (BBB) endothelium in the innate defense during P falciparum–infected erythrocyte (Pf-IRBC) sequestration. However, some of these diffusible molecules could have reversible effects on brain tissue and thus on neurologic function. The inflammatory pathways were validated by direct measurement of proteins in brain endothelial supernatants. This study delineates the strong inflammatory component of human brain endothelium contributing to cerebral malaria.

overall_design: Total of 8 samples (4 control and 4 treated) were analyzed. 4 control samples included two normal RBC control and two medium controls. 4 treated samples includes 2 exposed to low binding Pf-IRBC and 2 exposed to high binding Pf-IRBC (Pf-IRBC-P). Medium and RBC controls were finally used as four replicates of control and all four Pf-IRBC or Pf-IRBC-P exposed endothelial cells were used as 4 separate treated controls.

first: Monique

last: Stins

first: Wei

last: Sha

first: Vladimir

last: Shulaev

last: Sullivan


title: Expression Analysis Upon PARP-1 or PARG Knockdown in MCF-7 Cells

status: Public on Oct 23 2009

city: Dallas

first: W. Lee

last: Kraus

address: 5323 Harry Hines Blvd.

institute: UT Southwestern Medical Center

summary: Poly(ADP-ribose) polymerase-1 (PARP-1) and poly(ADP-ribose) glycohydrolase (PARG) are enzymes that modify target proteins in the nucleus by the addition and removal, respectively, of ADP-ribose polymers.  Although a role for PARP-1 in gene regulation has been well established, the role of PARG is less clear.  To investigate how PARP-1 and PARG coordinately regulate global patterns of gene expression, we used short hairpin RNAs (shRNAs) to stably knockdown PARP-1 or PARG in MCF-7 cells, followed by expression microarray analyses.

overall_design:
Two shRNAs targeting Luciferase (Luc, control), PARP-1, or PARG were cloned into the pSUPER.retro vector (puro or neo resistant) and sequentially introduced into parental MCF-7 cells using a standard retroviral infection technique.  Luc, PARP-1, and PARG control-matched cell lines were generated three independent times.

Total RNA was isolated from stable Luc, PARP-1, and PARG knockdown cells was analyzed for global patterns of gene expression.  Briefly, 7 μg of total RNA was labeled using Affymetrix's standard one-cycle amplification and labeling protocol.  The labeled cRNA was then hybridized to Affymetrix Human U133A 2.0 GeneChips, which were scanned using a GeneChip Scanner 3000.  Each biological replicate (1, 2 or 3) represents a single RNA isolation from each set of cell lines.

These 9 samples were samples were part of a 15-sample microarray analysis (GSE12971) examining expression regulation by SIRT1, PARP-1, PARG and macroH2A. All 15 samples were included for the data normalization steps.

The raw array data was processed by Affymetrix GeneChip Operating Software (GCOS) to obtain detection calls (ABS_CALL) and signal values (VALUE_2).  The signals were then normalized by scaling to a target value of 500 using GCOS.  To adjust for batch effects due to day-to-day differences in RNA isolations, the empirical Bayes method was applied to the data set.  After adjusting any values less than 0.01 to 0.01, the data was log2 transformed, median centered for each array, and median centered for each individual probe set (VALUE).  Filters were then applied to obtain final gene lists.

Specifically, the criteria for a gene to be considered regulated by PARP-1 or PARG was: (1) detection call flagged as present or marginal in 2 out of 3 array replicates for both control and factor knockdown cell lines and (2) significance of values between control and knockdown cell lines for any given gene had a two-tailed Student’s t-test with a p-value < 0.05.  To determine the genes most robustly regulated by PARP-1 or PARG, a fold change criteria was applied, namely log2 fold change > 0.5 or < -0.5 when compared to the Luc control knockdown cells.

first: Kristine

last: Frizzell

first: Matthew

last: Gamble

first: Tong

last: Zhang

first: WL


title: Gene expression analysis of chronically inflamed human peri-implant and periodontal ligament cells in vivo

status: Public on Aug 30 2009

city: Braunschweig

first: Robert

last: Geffers

address: Inhoffenstr. 7

department: Dep. Molecular Bacteriology

institute: HCI - Helmholtz Centre for Infection Research

summary:
The goal of this study was to compare the gene expression profiles of chronically inflamed human peri-implant and chronically inflamed human periodontal tissues in order to elucidate potential changes at the molecular level. Cells out of the pocket depth of the inflamed peri-implant and periodontal ligament as well as from the middle third of healthy periodontal ligament were applied. Genome-wide gene expression was compared with the help of microarray analysis, and the data were validated by real-time RT-PCR. The expression rates of 14,239 genes were investigated and 2,079 of them were found differentially expressed by at least two-fold; the expression rates of 1,093 genes were significantly up-regulated and the expression rates of 986 genes were significantly down-regulated in inflamed peri-implant cells compared to inflamed periodontal cells. We focused on genes coding for extracellular matrix components and those that degrade them. Only genes of non-fibril-forming collagens (types IV, VI, VII, and XVII) were increased in inflamed peri-implant tissue, whereas only the genes of two fibril-forming collagens (types III and XVII) were decreased, suggesting that peri-implant tissue re-models faster than periodontal tissue. Furthermore, cathepsin D and cathepsin S might participate to a greater extent in connective tissue destruction of peri-implant tissue. The present investigation demonstrated that, despite their clinical similarities, periimplantitis and periodontitis are two different diseases at the genetic level.
Keywords: inflammation, peri-implant, periodontal ligament cells

overall_design:
The peri-implant tissue was scraped of the implant and the periodontal tissue was scraped off the roots of the teeth. Both tissue probes were treated with the RNeasy mini Kit (Qiagen, Hilden, Germany) to isolate total RNA from chronically inflamed peri-implant and chronically inflamed PDL cells, according to the manufacturer’s instructions. Healthy PDL cells were taken from the middle third of the root clearly avoiding sampling the gingiva. The chronically inflamed peri-implant and periodontal cells were taken directly out of the depth of the pocket again avoiding the ingrown gingiva. Quality control and quantitation of total RNA samples were determined prior to the microarray experiments (Agilent 2100 Bioanalyzer, Agilent Technologies, Palo Alto, CA, USA). Equal amounts of total RNA were pooled into an inflamed peri-implant and an inflamed and healthy periodontal cell pool.  To monitor the relative abundance of mRNA for full-length human genes, the Affymetrix GeneChip HG_U133A was used, representing 22,283 probe sets (14,239 unique human genes). Labelled cRNA, fragmented to an average size of 100–150 bases, was hybridized to the GeneChips. For each pool of cRNA, two GeneChips were hybridized.
Chronically inflamed peri-implant as well as healthy and chronically inflamed periodontal tissue was collected from 48 patients at the Dental Medical School of the University of Goettingen (21 men, 27 women, aged between 18 and 72 years) from March 2005 to August 2006, as already described by Gersdorff et al. (2007). The peri-implant tissue probes were taken from endosteal implants (Astra Tech®, ITI Bonefit®, or Brånemark®) with chronic peri-implant lesions (Schwarz et al. 2007). The periodontal tissue probes were taken from teeth, either extracted for orthodontic reasons or due to carious lesions (healthy periodontium) or from teeth with generalized chronic periodontitis (Armitage 2004). Teeth with healthy PDL had no clinical signs of inflammation (no bleeding on probing), exhibited a probing depth of ?3,5 mm and had no radiographic detectable bone loss. In contrast, implants and teeth with generalized chronic inflammation exhibited obvious clinical signals of inflammation (bleeding on probing), exhibited a probing depth of >4 mm and had radiographic evidence of bone loss (1 to 7 mm). A detailed anamnesis of each patient was explored. All patients were without a medical history, i.e. heart diseases or diabetes, and were not on medication. In addition, the tissue samples used in our study were only taken from non-smokers. The explanted implants and extracted teeth were immediately frozen and stored at -80 °C. All patients who participated in the study were informed about the nature and aim of this project and gave their written informed consent. The study was approved according to the regulations of the Ethics Committee of the Medical Faculty of the University of Goettingen.

first: R

first: N

last: Gersdorff

last: Miosge


title: Expression Data From HCMV-Infected Human Monocytes: Role of EGFR

status: Public on Sep 03 2009

first: Andrew

last: Yurochko

summary:
Human cytomegalovirus (HCMV) induces pro-inflammatory monocytes following infection and we have evidence that EGFR is a key mediator in this early activation.  To begin to address how this signalling pathway is responsible for the rapid activation of infected monocytes, we examined the role this pathway played in the transcriptome of infected monocytes.  Global transcriptional profiling using cDNA microarrays revealed a significant number of genes, including inflammatory genes, were regulated in a EGFR-dependent manner, identifying this pathway as a key cellular control point in the conversion of monocytes to an activated pro-inflammatory state following HCMV infection.

Keywords: Disease state

overall_design: To begin to globally define how EGFR is involved in the HCMV-induced changes in monocyte function, we performed a transcriptome analysis in the presence of inhibitors to the EGFR signalling pathway.  Specifically, a cDNA microarray containing 12,625 unique probe sets was utilized to assess the modulation of the monocyte transcriptome at 24 hours post infection in the presence of blocking anti-EGFR antibody and pharmacological agent AG1478 (AG; an EGFR inhibitor).  A total of 4 replicates from mock-infected, HCMV-infected, anti-EGFR antibody-pretreated infected and AG-pretreated infected monocytes were analyzed in this study.

first: Gary

last: Chan

first: Maciej

middle: T

last: Nogalski


title: Ependymoma gene expression analysis

status: Public on Oct 18 2009

city: Richmond

first: Timothy

last: Van Meter

address: 1101 E. Marshall St.

department: Neurosurgery

institute: Virginia Commonwealth University

summary: This study was designed to assess ependymoma global gene expression analyses.

overall_design: Frozen tumor ependymoma biopsy samples from pediatric patients.

first: William

middle: C

last: Broaddus

first: Catherine

last: Dumur


title: Expression profiling of Burkitt's lymphoma cells 24h after FOXM1 shRNA or MYB shRNA lentivirus-mediated transduction

status: Public on Jul 27 2009

first: Mariano

middle: Javier

last: Alvarez

address: 1130 St. Nicholas ave.

department: Systems Biology

institute: Columbia University

summary: Human Burkitt's lymphoma ST486 cells were transduced with non-target control shRNA lentiviral vectors, FOXM1 shRNA, and MYB shRNA lentiviral vectors.  Total RNA was isolated 24h later. cRNA was produced with the standard one-step IVT protocol (Affymetix) and hybridized in U95Av2 gene chips (Affymetrix).

overall_design:
Experiment consists in 3 independent samples:
Expression profiling of Burkitt's lymphoma cells 24h after non-target control shRNA lentiviral mediated transduction.
Expression profiling of Burkitt's lymphoma cells 24h after FOXM1 shRNA lentiviral mediated transduction.
Expression profiling of Burkitt's lymphoma cells 24h after MYB shRNA lentiviral mediated transduction.
Data processing performed using MAS5 or GCRMA.

first: Pavel

last: Sumazin

first: Presha

last: Rajbhandari

first: Andrea

last: Califano


title: Lung adenocarcinoma cell line Wnt responsive signature

status: Public on Jul 10 2009

first: Don

middle: X

last: Nguyen

address: 1275 York Box 116

department: Cancer Biology and Genetics

institute: MSKCC

summary: Identify Wnt3A responsive signature in lung adenocarcinoma cells

overall_design: 4 different cell lines +/- Wnt3A; replicate samples

first: Xiang

first: Joan

last: Massague


title: Whole genome profiling of fibroblasts from Diamond-Blackfan Anemia patients

status: Public on Sep 14 2009

first: Paola

last: Roncaglia

address: via Bonomea 265

department: Neurobiology

institute: SISSA/ISAS (International School for Advanced Studies)

summary:
Diamond-Blackfan Anemia (DBA) is a rare inherited red cell hypoplasia characterized by a defect in the maturation of erythroid progenitors and is in some cases associated to malformations. Patients have an increased risk of solid tumors. Mutations have been found  in several ribosomal protein (RP) genes. Studies in hematopoietic progenitors from patients show that the haploinsufficiency of an RP impairs rRNA processing and ribosome biogenesis. DBA lymphocytes and fibroblasts show reduced protein synthesis, and the latter display abnormal rRNA processing and impaired proliferation.
To evaluate the involvement of non-hematopoietic tissues in DBA, we have analyzed global gene expression in fibroblasts from DBA patients compared to healthy controls. Microarray expression profiling using Affymetrix GeneChip Human Genome U133A 2.0 Arrays revealed that 421 genes are statistically significantly differentially expressed in DBA patients' fibroblasts relative to controls. These genes include a large cluster of ribosomal proteins and factors involved in protein synthesis and amino acid metabolism, as well as genes with roles in cell death, cancer and tissue development. Our results report for the first time into abnormal expression in a non-hematopoietic cell type in DBA, and support the hypothesis that DBA may be due to a defect in general or specific protein synthesis.

Keywords: Disease state analysis.

overall_design: Analyzed samples include fibroblasts from 4 DBA patients and 6 healthy controls.