title: Blood expression profiles define penetrance in DYT1 dystonia patients

status: Public on May 27 2010

city: Tuebingen

first: Michael

middle: H.

last: Walter

address: Calwer Str. 7

department: Department of Medical Genetics

institute: University of Tuebingen

summary: DYT1 dystonia is an autosomal-dominantly inherited movement disorder, which is usually caused by a GAG deletion in the TOR1A gene. Due to the reduced penetrance of ~30-40%, the determination of the mutation in a subject is of limited use with regard to actual manifestation of symptoms. In the present study, we used Affymetrix oligonucleotide microarrays to analyze global gene expression in blood samples of 15 manifesting and 15 non-manifesting mutation carriers in order to identify a susceptibility profile beyond the GAG deletion which is associated with the manifestation of symptoms in DYT1 dystonia.We identified a genetic signature which distinguished between asymptomatic mutation carriers and symptomatic DYT1 patients with 86.7% sensitivity and 100% specificity. This genetic signature could correctly predict the disease state in an independent test set with a sensitivity of 87.5% and a specificity of 85.7%.Conclusively, this genetic signature might provide a possibility to distinguish DYT1 patients from asymptomatic mutation carriers.

overall_design: Comparison of whole blood expression profiles of patients with DYT1 dystonia with non manifesting mutation carriers and non mutation carriers

last: Bonin

first: Rachel

last: Pullman

first: Enza

middle: M

last: Valente

first: Mario

last: Loi

first: Mattia

last: Gambarin

first: Deborah

last: Raymond

first: Michele

last: Tinazzi

first: Christoph

last: Kamm

first: Nicola

last: Glöckle

first: Sven

last: Poths

last: Gasser

first: Susan

last: Bressman

first: Christine

last: Klein

first: Laurie

last: Ozelius

first: Olaf

last: Riess

first: Kathrin

last: Grundmann


title: Global expression profiling of ruptured and unruptured human intracranial aneurysms

status: Public on Mar 04 2010

city: Krakow

first: Marcin

last: Piechota

address: Smetna 12

department: Molecular Neuropharmacology

country: Poland

institute: Institute of Pharmacology PAS

summary:
The primary aim of the present study was to identify differences on the transcription level between ruptured and unruptured intracranial aneurysms as well as normal intracranial arteries in human.

Keywords: Expression profiling by array

overall_design: Global gene expression profiling was performed in human intracranial aneurysms both ruptured (n=8) and unruptured (n=6) as well as in control intracranial arteries (middle meningeal artery, MMA; n=5) using oligonucleotide microarrays.

first: Joanna

last: Pera

first: Andrzej

last: Szczudlik


title: miR-29 targets in human fetal lung fibroblast IMR-90 cells

status: Public on Mar 01 2010

city: BoSTON

first: jining

last: lu

address: 715 Albany street R304

country: USA

institute: Boston University

summary: TGFβ is one of most intensively studied regulators of extracellular matrix formation, and has been implicated in the development of pulmonary fibrosis in different models. However, little is know about the role of miRNAs in TGFβ mediated fibrogenic gene regulation.  By using miRNA qRT-PCR array, we have identified miRNAs whose expression are regulated by TGFβ in IMR-90 cells. Among those down-regulated miRNAs are miR-29 family members. Knockdown miR-29 in IMR-90 cells results in up-regulation of a large number of extracellular matrix and fibrogenic genes including family members of collagen, laminin, integrin, ADAM and MMP, many of them are predicted or confirmed miR-29 targets.  Hierarchichal clustering analysis of mRNA array data revealed that many extracellular matrix and fibrogenic genes up-regulated by TGFβ in IMR-90 cells, are also up-regulated in miR-29 KD cells. Moreover, the similar set of extracellular matrix and fibrogenic genes is also significantly up-regulated in bleomycin treated mouse lungs. Together, our data strongly suggest that downstream of the TGFβ, miR-29 is a master modulator of genes involved in extracellular matrix formation and might play a significant role in pulmonary fibrosis.

overall_design: To identify targets of miR-29 in human fetal lung fibroblast cells (IMR-90), cells were transfected with 100nM scrambled, 50nM or 100nM of miR-29 LNA KD oligo. Total RNA were isolated 48hrs after transfection and subject to the mRNA array analysis using Affymetrix human Gene 1.0 ST Whole Genome Array

first: Jining

last: Lu

first: Leah

last: Cushing

first: Ping-ping

last: Kuang


title: Reprogramming of fibroblasts from Fragile-X patients to induced pluripotent stem cells (iPS) with defined factors

status: Public on May 10 2010

city: Ramat Gan

first: Achia

last: Urbach

address: Bar Ilan

department: Faculty of Life Sciences

country: Israel

institute: Bar Ilan University

summary: Human iPS cells derived from normal and Fragile-X fibroblasts in order to assess the capability of Fragile-X iPS cells to be used as a model for different aspects of Fragile-X syndrome. Microarry analysis used to compare global gene expression between human ES cells, the normal and the mutant iPS cells and the original fibroblasts, to demonstrate that the overall reprogramming process succeeded, and that the FX-iPS cells are fully reprogrammed cells.

overall_design: RNA samples from hES cells, human fibroblasts and iPS derived from the fibroblasts. Gene expression of those cells was analyzed.

first: Ori

last: Bar-Nur

first: George

middle: Q

last: Daley

first: Nissim

last: Benvenisty


title: Expression data from H1299 and H460 lung cancer cell lines

status: Public on Feb 27 2010

city: Seoul

first: Woong Yang

last: Park

address: 28 Yongondong, Chongnogu

department: Biochemistry and Molecular Biology

country: South Korea

institute: Seoul National University

summary: To identify a set of genes related to radioresistance, we analyzed the time-series gene expression profiles of radioresistant H1299 and radiosensitive H460 lung cancer cells in response to 2 Gy of ionizing radiation (IR) by performing quadratic regression (QR) analysis. Out of the 21,331 genes, we selected 6,538 genes by QR analysis from the gene expression profile of H460 cells and 6,086 genes from that of H1299 cells.  Most of the genes identified in the H460 cells were classified into continuously up- or down-regulated groups, while the major QR groups were transiently changed groups in the H1299 cell line.  From gene ontology analysis of the major QR groups, the DNA damage response was commonly enriched in both cell lines.  DNA repair-related genes such as ATM, ATR, TP53BP1, BRCA1, MRE11, NBN and RAD50 were particularly up-regulated in H1299 cells. Suppression of these DNA repair-related genes using siRNA made H1299 cells radiosensitive to ionizing radiation. The data suggest that differential responses to DNA damage confer radioresistance to cancer cells, and provide potential novel targets for sensitizing radiotherapy.

overall_design: We analyzed the time-series gene expression profiles of radioresistant H1299 and radiosensitive H460 lung cancer cells in response to 2 Gy of ionizing radiation (IR) by performing quadratic regression (QR) analysis to  identify genes associated with radioresistance

first: Woong-Yang


title: Identification and Characterization of Enhancers Controlling the Inflammatory Gene Expression Program in Macrophages

status: Public on Mar 05 2010

city: Vienna

first: Iros

last: Barozzi

address: Borschkegasse 8a

country: Austria

institute: Medical University Vienna

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Enhancers determine tissue-specific gene expression programs. Enhancers are marked by high histone H3 lysine 4 mono-methylation (H3K4me1) and by the acetyl-transferase p300, which has allowed genome-wide enhancer identification. However, the regulatory principles by which subsets of enhancers become active in specific developmental and/or environmental contexts are unknown. We exploited inducible p300 binding to chromatin to identify, and then mechanistically dissect, enhancers controlling endotoxin-stimulated gene expression in macrophages. In these enhancers, binding sites for the lineage-restricted and constitutive Ets protein PU.1 coexisted with those for ubiquitous stress-inducible transcription factors such as NF-kappaB, IRF, and AP-1. PU.1 was required for maintaining H3K4me1 at macrophage-specific enhancers. Reciprocally, ectopic expression of PU.1 reactivated these enhancers in fibroblasts. Thus, the combinatorial assembly of tissue- and signal-specific transcription factors determines the activity of a distinct group of enhancers. We suggest that this may represent a general paradigm in tissue-restricted and stimulus-responsive gene regulation.

overall_design: Chromatin immuno-precipitations of p300, PU.1 and mono-methylated H3 lysine 4 followed by multiparallel sequencing were performed in bone marrow-derived macrophages. Experiments for p300 and PU.1 were also carried out in cells treated for 2hrs with lipopolysaccharide (LPS). In case of p300, reads obtained from two different biological replicates were merged.

first: Serena

last: Ghisletti

first: Flore

last: Mietton

first: Sara

last: Polletti

first: Francesca

last: De Santa

first: Elisa

last: Venturini

first: Lorna

last: Gregory

first: Lorne

last: Lornie

first: Adeline

last: Chew

first: Chia-Lin

last: Wei

first: Jiannis

last: Ragoussis

first: Gioacchino

last: Natoli


title: MicroRNAs in Myeloma

status: Public on Mar 18 2010

city: Salamanca

first: Norma

middle: Carmen

last: Gutierrez

department: Hematology

country: Spain

institute: University Hospital

type:
Expression profiling by array
Other
Expression profiling by RT-PCR

summary: Specific microRNA (miRNA) signatures have been associated with different cytogenetic subtypes in acute leukemias. This finding prompted us to investigate potential associations between genetic abnormalities in multiple myeloma (MM) and singular miRNA expression profiles. Moreover, global gene expression profiling was also analyzed to find correlated miRNA-gene expression and select miRNA target genes that show such correlation. For this purpose, we analyzed the expression level of 365 miRNAs and the gene expression profiling in sixty newly diagnosed MM patients, selected to represent the most relevant recurrent genetic abnormalities. Supervised analysis showed significantly deregulated miRNAs in the different cytogenetic subtypes as compared to normal PC. Interestingly, miR-1 and miR-133a clustered on the same chromosomal loci, were specifically overexpressed in the cases with t(14;16). The analysis of the relationship between miRNA expression and their respective target genes showed a conserved inverse correlation between several miRNAs deregulated in MM cells and CCND2 expression level. These results illustrate, for the first time, that miRNA expression pattern in MM is associated with genetic abnormalities, and that the correlation of the expression profile of miRNA and their putative mRNA targets is useful to find statistically significant protein-coding genes in MM pathogenesis associated to changes in specific miRNAs.

overall_design:
Patients
Sixty patients with symptomatic newly diagnosed MM were included in the study. Five healthy controls of bone marrow (BM) samples were obtained from subjects undergoing BM harvest for allogeneic transplantation. In all the BM samples a CD138 positive plasma cell (PC) isolation using the AutoMACs automated separation system (Miltenyi-Biotec, Auburn, CA) was performed (purity was above 90%). All patients as well as healthy donors provided written informed consent in accordance with the Helsinki Declaration, and the research ethics committee of the University Hospital of Salamanca approved the study. The total 65 samples were analyzed from both miRNAs and mRNA gene expression profiling.

Cytogenetic Analysis
The selection of patients was based on cytogenetic features in order to include a representative number of samples with the most relevant and recurrent genetic abnormalities. The systematic screening for genomic aberrations in our institution includes interphase FISH studies for the detection of IGH rearrangements, RB1 and P53 deletions (Abbott Molecular/Vysis, Des Plaines, IL, USA) as previously described,18 and 1q gains (ON 1q21/SRD 1p36, Kreatech Diagnostics, Amsterdam). Furthermore, only patients with more than 80% of PC exhibiting genetic abnormalities were considered for the analysis, with the exception of gains on 1q and P53 deletions, which were present in a median of 77% (range, 28-100%) and 72% (range, 46-88%) of the PC, respectively. The distribution of cytogenetic abnormalities in the 60 MM patients is summarized in Table 1.

RNA Extraction
Total RNA was extracted from normal and tumor plasma cells using miRNEasy Mini Kit (Qiagen, Valencia, USA) following manufacturer's protocol. The RNA integrity was assessed using Agilent 2100 Bioanalyzer (Agilent Tech.Inc., Palo Alto, CA, USA).

MicroRNA profiling
cDNA was synthesized from total RNA using the so-called hair-pin RT-primer according to the TaqMan MicroRNA Reverse Transcription Kit (PE Applied Biosystems, Foster City, CA). Reverse transcriptase reactions contained: 20ng of RNA, 1.5 μl 10x RT buffer, 0.15 μl dNTP mix (100 mM total), 1 μl MultiScribe Reverse Transcriptase (50 U/μl), 0.19 μl AB RNase Inhibitor (20 U/μl), 1 μl multiplex RT primers and 4.16 μl H2O (final volume 10 μl). Reactions were incubated in an Applied Biosystems GeneAmp PCR System 9700 for 30 min at 16°C, 30 min at 42°C, 5 min at 85°C and then held at 4°C. A total of eight independent RT reactions must be run per sample. Diluted RT reaction product is mixed with TaqMan® Universal PCR Mastermix (No AmpErase UNG) and loaded into the corresponding TaqMan® Low Density Arrays fill ports (Applied Biosystems, Part number: 4384792). This panel contains 368 TaqMan® MicroRNA Assays enabling accurate quantification of 365 human miRNAs and three endogenous controls (RNU48, RNU48 and RNU6B) to aid in data normalization. Real-time PCR was performed using an Applied Biosystems 7900 HT Fast Real Time PCR Sequence Detection system. The reactions were incubated at 94.5 ºC for 10 min, followed by 50 cycles of 97 ºC for 30 s and 59.7 ºC for 1 min. The threshold cycle (Ct) data was determined using 0.3 as a threshold. The Ct is defined as the fractional cycle number at which the fluorescence passes the fixed threshold.
MiRNAs with Ct values higher than 35 were excluded from the analysis, leaving a set of 192 miRNAs. Normalization was performed with the mean of RNU44 and RNU48, as they were uniformly expressed across the patient dataset. Relative quantification of miRNA expression was calculated with the 2-deltaCt and 2-deltadeltaCt methods, where deltaCtdeltaCt(miRNA)-Ct(miRNA control) and deltadeltaCt deltaCt(MM)-deltaCt(average normal PC), depending upon whether comparisons were made between MM samples and normal PC or between MM samples, respectively.19 The data was presented as log10 of the relative quantity of each miRNA.20 We used hierarchical clustering (Cluster and TreeView software) based on the average-linkage method with the centered correlation metric for unsupervised analysis.21 Differentially expressed miRNAs were identified using Significant Analysis of Microarrays (SAM) algorithm, by using the two-class (unpaired) format, not considering equal variances.22 Significant genes were selected based on false discovery ratio (FDR) and controlling the q-value for the gene list.
Target prediction was performed using miRecords, an integrated resource for animal miRNA–target interactions.23 The Predicted Targets component of miRecords integrates the predicted targets of the following miRNA target prediction tools: DIANA-microT, MicroInspector, miRanda, MirTarget2, miTarget, NBmiRTar, PicTar, PITA, RNA22, RNAhybrid, and TargetScan.

mRNA gene expression profiling
RNA labeling and microarray hybridization have been previously reported.24 Total RNA was extracted from purified cell populations using RNeasy Mini Kit (Qiagen, Valencia, CA, USA) according to the manufacturer’s instructions. Labelling and hybridizations were performed according to protocols from Affymetrix. Briefly, 100-300 ng of total RNA were amplified and labeled using the WT Sense Target labelling and control reagents kit (Affymetrix Inc., Santa Clara, CA, USA), and then hybridized to Human Gene 1.0 ST Array (Affymetrix). Washing and scanning were performed using GeneChip System of Affymetrix (GeneChip Hybridization Oven 640, GeneChip Fluidics Station 450 and GeneChip Scanner 7G). Expression value for each probe set was calculated using RMAExpress program that uses RMA (Robust Multi-Array Average) algorithm.25 A differential expression analysis was carried out on the data using SAM algorithm to identify genes with statistically significant changes in expression between different classes.
To identify the putative mRNA targets of the significant miRNAs, which at the same time were selected as significantly deregulated genes in the class comparison analysis of gene expression data, a Pearson correlation analysis was performed using a cutoff p-value <0.05 (2-tailed). Calculation of the p-values assigned to the Pearson's correlation coefficients was done assuming that the variables follow a bivariate normal distribution. The functional analysis to identify the most relevant functional categories in the datasets of miRNA target genes selected by statistical analysis, was generated through the use of Ingenuity Pathways Analysis (Ingenuity Systems, Mountain View, CA).

middle: C

first: Maria

middle: E

last: Sarasquete

first: Manuel

last: Delgado

first: Fany

middle: V

last: Ticona

first: Javier

last: De las Rivas

first: Encarna

last: Fermiñan

first: Patricia

last: Martin-Jimenez

first: Carmen

last: Chillon

first: Alberto

last: Risueño

first: Jesus

last: Hernandez

first: Ramon

last: Garcia-Sanz

first: Marcos

last: Gonzalez

middle: F

last: San Miguel


title: Genes differentially expressed in primary and metastatic liver GISTs.

status: Public on Apr 13 2010

city: Hamamatsu

first: Hirotoshi

last: Kikuchi

address: 1-20-1 Handayama, Higashi-ku

department: Second Department of Surgery

country: Japan

institute: Hamamatsu University School of Medicine

summary:
Although the main cause of gastrointestinal stromal tumor (GIST) is due to gain-of-function mutation of the c-kit gene in the interstitial cells of Cajal, concomitant genetic or epigenetic changes other than c-kit are thought to occur in the development of metastasis.
We used microarrays to identify genes that were up-regulated and down-regulated in the metastatic liver GIST.

overall_design: Three primary gastric GISTs without synchronous or metachronous metastasis and five metastatic liver tumors originated from gastric GIST were utilized for RNA extraction and hybridization on Affymetrix microarrays. No patients had received imatinib therapy before surgery.

first: Tomohiko

last: Setoguchi

first: Masayoshi

last: Yamamoto

first: Naoko

last: Gto-Inoue

first: Mitsutoshi

last: Setou


title: Gene expression analysis of lung biopsies from patients with two different forms of pulmonary sarcoidosis

status: Public on Jan 31 2010

city: Oxford

first: Ling-Pei

last: Ho

address: John Radcliffe Hospital

department: MRC Human Immunology Unit

country: United Kingdom

institute: Weatherall Institute for Molecular Meicine, University of Oxford

summary:
Approximately 60-70% of patients with pulmonary sarcoidosis have a good outcome, with disease that resolves spontaneously. It is unclear why some patients progress to fibrotic disease, and there is currently no marker that differentiates these patients from those with self-limiting lung disease.
Using the Affymetrix Human Gene 1.0 ST Array, we examined gene expression in lung tissue samples obtained using bronchoscopic transbronchial biopsy from 8 patients with active but self-limiting pulmonary sarcoidosis and 7 with active, progressive, fibrotic pulmonary disease. Analyses were performed to identify differentially expressed genes between the two groups, and to test for enrichment of sets of genes associated with defined biological processes from the Gene Ontology Consortium, as well as with gene sets derived from published data for idiopathic pulmonary fibrosis (IPF) and hypersensitivity pneumonitis (HP).

overall_design: Expression levels were examined in lung biopsy samples from 8 patients with nodular, self-limiting pulmonary sarcoidosis and 7 patients with progressive, fibrotic pulmonary sarcoidosis.

first: Helen

last: Lockstone

first: Sharon

last: Sanderson

first: Nina

last: Kulakova

first: Dilair

last: Baban

first: Andrew

last: Leonard

first: Wai

middle: L

last: Kok

first: Simon

last: McGowan

first: Chris

middle: I

last: Newbold

middle: J

last: McMichael


title: GSK-3 inhibitor treatment effect on MLL leukemia cell

status: Public on May 01 2010

city: Stanford

first: Michael

last: Cleary

state: CA

address: Room L232, Stanford Medical Center

department: Pathology

web_link: http://    http://med.stanford.edu/profiles/pathology/researcher/Michael_Cleary/  At Stanford

institute: Stanford University

summary: Human leukemia cell line RS4.11 was treated with GSK-3 inhibitor SB216763 for 20 hours.  Gene expression profiling was performed to analyze genes affected by GSK-3 inhibition.

overall_design: Human leukemia cell line RS4.11 was grown in vitro and treated with 10 uM GSK-3 inhibitor SB216763 or solvent DMSO for 20 hours.  Total RNA was extracted and microarray analysis was performed.  Two controls and three inhibitor treatment samples were analyzed.

first: Zhong

last: Wang


title: Expression data from Che-1 (AATF) depleted SKBR3 and MDA-468 cell lines

city: rome

first: maurizio

last: fanciulli

address: via e chianesi 53

department: molecular medicine

country: Italy

institute: regina elena cancer institute

summary: Che-1 is a RNA Polymerase II binding protein involved in the regulation of gene transcription. We have observed that Che-1 depletion induces apoptosis in several cancer cells expressing mutated forms of p53. We used microarrays to investigate classes of genes regulated by Che-1 in one of these cell lines.

overall_design: Total RNA was extracted from control and Che-1 depleted SKBR3 and MDA-468 cells 48 hours after transient transfection of siRNA.

first: Tiziana

last: Bruno

first: Barbara

last: Benassi


title: Expression data from TK6 exposed to low-dose metals

status: Public on Apr 01 2010

city: Chapel Hill

first: Rebecca

middle: Catherine

last: Fry

address: 1213 MHRC

department: Environmental Sciences and Engineering

institute: UNC-Chapel Hill

summary:
We are investigating the response of human lymphoblastoid cells to low-dose exposure of environmental metals
We used microarrays to detail the global programme of gene expression upon response to low-dose metals

Keywords: dose

overall_design: TK6 cells  were grown to mid log-phase and exposed to low-doses of arsenic and cadmium. RNA was collected 24 hrs after exposure.

first: Margaret

middle: A

last: Benton

first: Julia

last: Rager

first: Lisa

last: Smeester


title: Expression data from fibroblasts cultured from oesophageal biopsies, taken from metaplasia, dysplasia and EAC specimens.

status: Public on Jan 22 2010

city: Cambridge

first: Nicholas

middle: B

last: Shannon

address: Robinson Way

department: Oncology

institute: Cambridge Research Institute

summary: Since fibroblasts are a key component of the stroma with an established role in cancer, we investigated the contribution of fibroblasts to the signature observed in the stromal compartment. 13 clonally derived primary stromal fibroblasts were generated from metaplasia, dysplasia and EAC specimens. Expression of a panel of known fibroblast markers and concomitant absence of epithelial markers confirmed their fibroblastic origin. Gene expression profiling of these esophageal fibroblasts demonstrated that three ontologies related to an invasive phenotype (chemotaxis, cell adhesion, regulation of angiogenesis) differentiated cancer associated from BE fibroblasts. Furthermore, the ontologies and KEGG pathways relating to inflammation were all statistically upregulated in the fibroblast signature.

overall_design: 13 clonally derived primary stromal fibroblasts were generated from metaplasia, dysplasia and esophageal adenocarcinoma (EAC) specimens.

first: Amel

last: Saadi

last: Fitzgerald


title: Down-regulation of ERBB3 by shRNA in the T47D cell line

status: Public on Jan 16 2010

city: Rehovot

first: Amit

last: Zeisel

address: Weizmann Institute

department: Physics of complex systems

institute: Weizmann Institute of Science

summary: ERBB3 expression levels were down-regulated by shRNA. Two such stable lines were made and gene expression was measured by Affymetrix arrays.

overall_design: Four types of T47D cell lines were compared: WT and shEGFP as controls, versus sh2608 and sh3293 as the ERBB3 down-regulated cells. Two replicates per cell line.

first: Erez

last: Bublil

first: Yosef

last: Yarden


title: Expression data from serum-starved control and CDK8 depleted cells following serum stimulation

status: Public on Jan 19 2010

city: Boulder

first: Aaron

last: Donner

state: CO

department: Molecular Cellular and Developmental Biology

institute: University of Colorado Boulder

summary:
The Mediator complex allows communication between transcription factors and RNA polymerase II (RNAPII). CDK8, the kinase found in some variants of Mediator, has been characterized mostly as a transcriptional repressor. Recently, CDK8 was demonstrated to be a potent oncoprotein. Here we show that CDK8 is predominantly a positive regulator of gene expression within the serum response network, as it is required for expression of several members of the AP-1 and EGR family of oncogenic transcription factors (e.g. FOS, JUN, EGR1-3). Mechanistic studies demonstrate that CDK8 is not required for recruitment of RNAPII and promoter escape at these loci. Instead, CDK8 depletion leads to the appearance of slower elongation complexes carrying hypophosphorylated RNAPII. We show that CDK8-Mediator regulates precise steps in the assembly of a functional elongation complex, including the recruitment of P-TEFb and BRD4, but is dispensable for recruitment of SPT5 and FACT. Furthermore, CDK8-Mediator specifically interacts with P-TEFb. Thus, we uncovered a novel role for CDK8 in transcriptional regulation that may contribute to its oncogenic effects.
We used microarrays to analyze the expression of immediate early genes in response to serum in cells with and without the Mediator associated kinase, CDK8.

overall_design: HCT116 cells expressing control and CDK8-directed short-hairpin RNAs were starved of serum for 40 hours, then RNA was isolated at time 0 and 30 minutes following serum stimulation.

first: Joaquin

last: Espinosa


title: Bone marrow-derived MSC as endometrial stromal progenitor cells

status: Public on Mar 31 2010

city: Valencia

first: Jose

middle: Antonio

last: Martinez-Conejero

address: C/ Guadassuar 1, bajo

institute: iGenomix

summary:
In the current study, we hypothesized that if bone-marrow derived MSC contribute to endometrial regeneration and are progenitors to hESF, their treatment with agents known to regulate hESF differentiation could promote their differentiation down the stromal fibroblast lineage. To this end, we treated bone marrow-derived MSC with estradiol (E2) and progesterone (P4), BMP2, and activators of the PKA pathway and investigated specific markers of hESF differentiation (decidualization). Furthermore, we investigated the transcriptome of these cells in
response to cAMP and compared this to the transcriptome of hESF decidualized in response to activation of the PKA pathway. The data support that MSC can be differentiated down the hESF pathway, as evidenced by changes in cell shape and common expression of decidual markers and other genes important in hESF differentiation and function.

overall_design:
Bone marrow-derived MSC were expanded in DMEM (high glucose) medium containing 10% fetal bovine serum, 1% penicillin and streptomycin. At 80% confluency MSC were trypsinized and plated at 5x103 cells/cm2 in 6cm plates. Nearly confluent cells were serumstarved overnight and thereafter treated for 0, 3, 7, 14 and 21 days in low-serum medium (2%
FBS) with the following treatments: (a) 10nM E2/1mM P4; (b) 0.5mM or 1mM 8-Br-cAMP (cAMP); (c) 50nM or 100nM BMP-2; (d) 0.5mM and 1mM 8-Br-cAMP + BMP-2; (d) 0.5mM and 1mM 8-BrcAMP + BMP-2 + E2/P4; (e) 50nM or 100nM human FSH, with vehicle controls at each time point. Each experiment was conducted in duplicate and repeated at least 3
times. Media were changed every third day. Endpoints were prolactin (PRL) and IGF binding protein-1 (IGFBP1) mRNA, assessed by quantitative real-time RT-PCR, and protein in conditioned media, assessed by ELISA (see below), after 3, 7, 14 and 21 days of incubation.
Immunostaining for vimentin, cytokeratin 7, and Oct-4 was performed before and after MSC treatments to assess the stromal lineage of MSC and their differentiation status. MSC expressed vimentin and did not express cytokeratin or Oct-4, independent of culture conditions
or duration of culture or treatment. The morphological appearance of MSC under different treatment conditions was photographed using a Leica CTR 6500 microscope.
Microarray analysis was performed with MSC, n=3, at t=0 (untreated, uncultured control), 14day vehicle control, and 14day treated with 8-Br-cAMP. Only high quality samples (RIN=9.1-10) were used for microarray analysis. Hybridization was performed with Affymetrix Human Gene 1.0 ST arrays.

first: Lusine

last: Aghajanova

first: José

last: Horcajadas

first: Francisco

last: Esteban

first: Linda

last: Giudice


title: Expression, MeDIP and ChIP-on-chip data from normal Prostate epithelial cells (PrEC) and the LNCaP cancer cell line

status: Public on Jan 04 2010

city: Darlinghurst

last: Statham

state: NSW

address: 384 Victoria St

department: Cancer Department

country: Australia

institute: Garvan Institute of Medical Research

type: Methylation profiling by genome tiling array

type: Genome binding/occupancy profiling by genome tiling array

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

overall_design: Refer to individual Series


title: Expression data from normal Prostate epithelial cells (PrEC) and the LNCaP cancer cell line

summary:
To identify genomic regions which display concordant gene expression in prostate cancer, we performed expression profiling of normal prostate epithelial cells (PrEC) and the prostate cancer cell line LNCaP.
These expression arrays were integrated ChIP-on-chip studies of active and repressive epigenetic marks in same cells to discover and characterise regions of Long Range Epigenetic Silencing (LRES) in prostate cancer.

overall_design: Duplicate expression arrays of PrEC and LNCaP

first: Marcel

middle: W

last: Coolen

first: Clare

last: Stirzaker

first: Jenny

middle: Z

last: Song

first: Zena

last: Kassir

first: Carlos

middle: S

last: Moreno

first: Andrew

middle: N

last: Young

first: Vijay

last: Varma

first: Terence

middle: P

last: Speed

first: Mark

last: Cowley

first: Paul

last: Lacaze

first: Warren

last: Kaplan

middle: D

last: Robinson

first: Susan

last: Clark


title: Gene expression analysis of bone tumors

status: Public on Dec 15 2009

city: Lund

first: Fredrik

last: Mertens

address: Lund University Hospital

department: Dept of Clinical Genetics

country: Sweden

institute: Lund University

summary:
Subungual exostosis is a benign bone- and cartilage-forming tumor known to harbour a pathognomonic t(X;6)(q22;q13-14). Using global gene expression analysis and quantitative real-time PCR we could show that this translocation results in increased expression of the IRS4 gene, presumably due to disruption and/or exchange of regulatory sequences with the translocation partner, the COL12A1 gene. A corresponding deregulation at the protein level could be demonstrated in primary cell cultures using a combination of fluorescence in situ hybridization and immunostaining. As the t(X;6) usually is the sole cytogenetic aberration in subungual exostosis, the deregulated expression of IRS4 is likely to be pathogenetically essential.
Gene expression analysis of bone tumors.

overall_design: Total RNA was extracted from five frozen bone tumor biopsies (two osteochondromas, two cases of BPOP, one subungual exostosis), cDNA was generated, fragmented and end-labelled. The fragmented and biotinylated cDNA was hybridized to the GeneChipR Human Gene 1.0 ST Arrays (Affymetrix).


title: Expression data from normal peripheral blood mononuclear cells, cell lines K562 and HL60

status: Public on Jan 08 2010

city: New Delhi

first: Alok

last: Bhattacharya

state: New Delhi

address: New Mehrauli Road

department: School of Life Sciences

country: India

institute: Jawaharlal Nehru University

summary: We did microarray to compare the gene expression profile of peripheral blood mononuclear cells (PBMC from normal volunteer) and two leukemic cell lines that is K562 (Chronic myelogenous leukemia cell line), HL60 (Promyelocytic leukemia cell line) in order to find differentially expressed genes in these samples.

overall_design: We used three RNA samples 1. Peripheral blood mononuclear cells (PBMC) from normal volunteer 2. HL60 cell line (This cell line is used to study promyelocytic leukemia) 3. K562 cell line (This cell line is used as a model to study Chronic Myelogenous Leukemia). Total RNA was extracted from normal PBMC, K562 and HL60 and quality was checked by running it on 1.2% formaldehyde agarose. The RNA was used as a starting template to synthesize double-stranded cDNA with random hexamers tagged with T7 promoter sequence. The fragmented DNA was labeled and used for overnight hybridization with Gene ST 1.0 arrays, followed by washing, staining and scanning. The normalized data for the normal PBMC as control was compared with the two cancer cell lines K562 and HL60.

first: Hafeez

last: Ahmad

first: Ritu

last: Kulshreshtha

first: Candida

last: Vaz


title: Expression data of influenza A infected human macrophages

status: Public on Oct 31 2009

city: Hong Kong

first: SMY

last: Lee

address: 21 Sassoon Road

country: Hong Kong

institute: The University of Hong Kong

summary:
Human disease caused by highly pathogenic avian influenza (HPAI) H5N1 can lead to a rapidly progressive viral pneumonia leading to acute respiratory distress syndrome. There is increasing evidence suggests a role for virus-induced cytokine dysregulation in contributing to the pathogenesis of human H5N1 disease. The key target cells for the virus in the lung are the alveolar epithelium and alveolar macrophages, and previous data has shown that compared to seasonal human influenza viruses, equivalent infecting doses of H5N1 viruses markedly up-regulate pro-inflammatory cytokines in both primary cell types in vitro. The dysregulation of H5N1-induced host responses is therefore important for understanding the viral pathogenesis.
We used microarrays to analyze and compare the gene expression profiles in primary human macrophages after influenza A virus infection.

overall_design: Peripheral-blood leucocytes were separated from buffy coats of three healthy blood donors and cells were differentiated for 14 days before use. Differentiated macrophages were infected with H1N1 and H5N1 at a multiplicity of infection (MOI) of two. Total RNA was extracted from cells after 1, 3, and 6h post-infection, and gene expression profiling was performed using an Affymetrix Human Gene 1.0 ST microarray platform.

first: Suki

middle: M

first: Jennifer

last: Gardy

first: C

last: Cheung

first: Timothy

middle: K

first: Kenrie

last: Hui

first: Nancy

middle: Y

last: Ip

first: Y

last: Guan

first: Robert

last: Hancock

first: J

last: Peiris


title: Deletion of the WWOX gene and frequent loss of its protein expression 3 in human osteosarcoma

status: Public on Nov 26 2009

city: HOUSTON

first: wei

last: zhang

state: TX

address: 1515 Holcombe Blvd, Unit 85

institute: M. D. Anderson Cancer Center

summary: Array comparative genomic hybridization on 10 frozen osteosarcoma specimens

overall_design: 6 treated Sarcoma vs. 2 untreated Sarcoma vs. 2 before-after treatment replicates

first: Jilong

last: Yang

first: David

last: Cogdell

first: Da


title: The PKA Pathway of Endometrial Stromal Fibroblasts Reveals Differentiation and Proliferative Potential in Endometriosis

status: Public on Dec 19 2009

first: Jose

last: Martinez-Conejero

summary: Intrinsic abnormalities in transplanted eutopic endometrium are believed to contribute to the pathogenesis of pelvic endometriosis. Herein, we investigated transcriptomic differences in human endometrial stromal fibroblasts (hESF) from women with (hESFendo) versus without (hESFnon-endo) endometriosis, in response to activation of the PKA pathway with 8-Br-cAMP. hESFnon-endo (n=4) and hESFendo (mild endometriosis, n=4) were isolated from eutopic endometrium and treated +/- 0.5mM 8-Br-cAMP for 96 hours. Purified total RNA was subjected to microarray analysis using the whole genome Gene 1.0 ST Affymetrix platform. 733 genes were regulated in cAMP-treated hESFnon-endo versus 172 genes in hESFendo, suggesting a blunted response to cAMP/PKA pathway activation in women with disease. Real-time PCR and ELISA validated the decreased expression of decidualization markers in hESFendo compared to hESFnon-endo. In the absence of disease, 8-Br-cAMP down-regulated progression through the cell-cycle due to a decrease in Cyclin D1, cyclin-dependent kinase 6 and cell division cycle 2, and an increase in cyclin-dependent kinase inhibitor 1A. However, cell cycle components in hESFendo were not responsive to 8-Br-cAMP, resulting in persistence of a proliferative phenotype. hESFendo treated with 8-Br-cAMP exhibited altered expression of immune response, extracellular matrix, cytoskeleton, and apoptosis genes. Changes in phosphodiesterase expression and activity were not different among experimental groups. Thus, eutopic hESF with increased proliferative potential may seed the pelvic cavity via retrograde menstruation and promote establishment, survival, and proliferation of endometriosis lesions, independent of hydrolysis of cAMP and likely due to an inherent abnormality in the PKA pathway in the presence of disease.

overall_design: Endometrial tissue biopsies were obtained from 14 reproductive age women. Six subjects had endometriosis (n=1 minimal, n=2 mild, n=1 moderate-severe, n=2 severe), based on visualization of lesions during laparoscopy and their histologic evaluation. Staging of endometriosis was according to the revised American Fertility Society classification system. Participating subjects with endometriosis were 22-46 years old (mean 33.5+/-3.0), not pregnant, and did not use hormonal medications within 3 months before surgery. Controls were eight cycling premenopausal subjects (37-49 years old, mean 44.4+/-1.7) undergoing endometrial biopsy or hysterectomy for benign reasons, such as fibroids, pelvic organ prolapse, pelvic pain or endometrial polyps. Controls had regular menstrual cycles (25-35 days), were documented not to be pregnant, had no history of endometriosis and no evidence of endometriosis at laparoscopy and had not been on hormonal therapies for at least 3 months before tissue sampling.

first: Lusine

last: Aghajanova

last: Horcajadas

first: James

last: Weeks

first: Francisco

last: Esteban

first: Camran

last: Nezhat

first: Marco

last: Conti

first: Linda

middle: C

last: Giudice


title: Gene Expression Profiling of an Immortalized Human Neural Stem Cell Line HB1.F3

status: Public on Oct 22 2009

city: Tokyo

first: Jun-ichi

last: Satoh

address: 2-522-1 Noshio, Kiyose

department: Bioinformatics

institute: Meiji Pharmaceutical University

summary: Neural stem cells (NSC) with self-renewal and multipotent properties serve as an ideal cell source for transplantation to treat spinal cord injury, stroke, and neurodegenerative diseases. To efficiently induce neuronal lineage cells from NSC for neuron replacement therapy, we should clarify the intrinsic genetic programs involved in a time and place-specific regulation of human NSC differentiation. Recently, we established an immortalized human NSC clone HB1.F3 to provide an unlimited NSC source applicable to genetic manipulation for cell-based therapy. To investigate a role of neurogenin 1 (Ngn1), a proneural basic helix-loop-helix (bHLH) transcription factor, in human NSC differentiation, we established a clone derived from F3 stably overexpressing Ngn1. Genome-wide gene expression profiling identified 250 upregulated genes and 338 downregulated genes in Ngn1-overexpressing F3 cells (F3-Ngn1) versus wild-type F3 cells (F3-WT). Notably, leucine-rich repeat-containing G protein-coupled receptor 5 (LGR5), a novel stem cell marker, showed a robust increase in F3-Ngn1.

overall_design: Primary cultures of fetal human telencephalon cells were transformed with a retroviral vector pLSNmyc carrying the v-myc oncogene and the neomycin resistance gene. Following selection with G418, a single continuously dividing clone with a capacity to self-renew and differentiate into neurons and glial cells both in vitro and in vivo was isolated and designated HB1.F3. It carried normal human karyotype of 46 XX. After transducing a retroviral vector pBabePNgn1 carrying the open-reading frame (ORF) of the human Ngn1 gene and the puromycin resistance gene into HB1.F3 cells, a single puromycin-resistant clone was selected, expanded, and designated HB1.F3-Ngn1. In the present study, the wild-type HB1.F3 cells and the HB1.F3-Ngn1 cells are abbreviated as F3-WT and F3-Ngn1. Total RNA of F3-WT and F3-Ngn1 isolated from subconfluent cells was processed for two times different hybridization experiments, named as 1st and 2nd experiments.

first: Seung

middle: U

last: Kim


title: Gene expression profiles of dental follicle cells before and after osteogenic differentiation in vitro

status: Public on Sep 12 2009

city: Regensburg

first: Christian

last: Morsczeck

address: Franz-Josef Strauss Allee 11

country: Germany

institute: University of Regensburg

summary: We analysed gene expression profiles in dental follicle cells before and after osteogenic differentiation with dexamethasone.

overall_design: Total RNAs were isolated from dental follicle cells before and after 28 days of differentiation in a standard osteogenic differentiation medium supplemented with dexamethasone


title: Diabetes biomarker disease progression study in rat liver

status: Public on Oct 21 2009

city: Washington

first: Svetlana

last: Ghimbovschi

state: DC

address: 111 Michigan Ave., NW

department: Research Center for Genetic Medicine

institute: Children's National Medical Center

summary:
The goals and objectives: To study Type 2 diabetes progression and the development of insulin resistance in two animal models with and without a high fat diet  superimposed on these models.  Background: Diabetes is a systemic metabolic imbalance involving multiple tissues/organs, and an early hallmark feature of this disease state is insulin resistance. Multifactorial interactions of genetics, prenatal environmental factors (fetal programming) and postnatal environmental factors (nutrition and activity) likely contribute to the diabetic phenotype.Animal models can serve as a valuable tool for studying diabetes disease progression and for identifying useful biomarkers of type 2 diabetes. Several inbred rodent models are available for diabetes related studies.  The GK rat is an obvious choice among available inbred models as the genetic basis for this inheritable form of diabetes is polygenic (5), unlike most other inbred rodent models that exhibit single gene defects.  Many of the characteristics of the GK rat mirror human diabetes (hyperglycemia, glucose intolerance, insulin resistance), although hyperlipidemia does not appear to be prominent in the GK rat.  Due to its polygenic mode of inheritance and 100% penetrance, the GK rat may be a useful model for human diabetes. Induced animal models can also be useful in diabetes studies. One such model is metabolic syndrome resulting from experimentally induced fetal programming (produced by maternal malnutrition or by exposure to corticosteroids in the third trimester).  Both in humans and animals,  accumulating evidence suggests that alterations in the human fetal environment can result in permanent physiologic changes that manifest as increased incidence of adult onset pathology.  Numerous epidemiological studies have forged a strong link between low birth weight and the development of metabolic syndrome in adulthood.  From such observations has arisen the concept of “fetal programming” whereby exposure to some factor(s) during crucial stages in development can permanently alter or “reset” physiologic/metabolic functions.  In the rat, exposure to corticosteroids during a “window” in third trimester gestation (CS programming) results in fetal growth retardation and insulin resistance in adult offspring.  Genetic factors play a primarily role in the etiology of diabetes in the GK rat, whereas fetal environmental factors are causative in CS programming.  (It should be noted that although altered fetal environmental effects, most likely stemming from maternal hyperglycemia, have been implicated to play some role in the decreased pancreatic B cell mass in GK rats, these effects occur earlier in gestation and therefore differ from programming by CS in late gestation.)  A comparison of the development of insulin resistance in the GK rat with development in the CS programmed rat will provide insight into genetic and fetal environmental factors in disease development. Superimposing dietary alterations (i.e., high fat feeding) (11) on both animal models may aid in the dissection of multiple interacting factors (genetic, fetal environmental factors, postnatal environmental factors) on the development and progression of insulin resistance and type 2 diabetes. Such studies may also aid in the identification of useful biomarkers for insulin resistance and type 2 diabetes in humans.
Proposed research:   Experiments are designed to study disease progression and the development of insulin resistance in two animal models:  the GK rat and the CS programmed rat, with and without a high fat diet superimposed on these models.  Animals will be maintained in our facility from weaning (GK rats) or birth (CS programmed - WKY), and body weights taken weekly.  Appropriate diets (normal or high fat) will be introduced at weaning.  Groups of animals (N=6) will be sacrificed at 5 different ages:  4, 8, 12, 16, and 20 weeks.  Plasma samples will be analyzed for markers of hyperglycemia, hyperinsulinemia, dyslipidimia, and for selected other hormonal factors which may contribute to disease etiology (adiponectin, leptin, corticosterone). At sacrifice, muscle is harvested, flash-frozen in liquid nitrogen, and warehoused in our tissue collection maintained at - 80 degrees C for this and possible future work.  Study will initially focus on examination of selected molecular markers of insulin resistance at the mRNA level in rat liver (PEPCK, PDK4).

Keywords: Type 2 diabetes, biomarker, rat, liver, time series

overall_design: The study contains: 50 liver samples from GK (GotoKakizake) rats and 51 liver sample from WKY (WistarKyoto) rats  feeded with normal diet (ND) and high fat diet (HFD) and sacrificed at 5 different ages:  4, 8, 12, 16, and 20 weeks. So, each time point contains 5 GK samples with ND, 5 GK samples with HFD, 5 WKY samples with ND and 5 WKY samples with HFD.

first: Richard

last: Almon

first: Debra

last: DuBois


title: Diabetes biomarker disease progression study in rat adipose tissue

summary:
The goals and objectives: To study Type 2 diabetes progression and the development of insulin resistance in two animal models with and without a high fat diet  superimposed on these models.  Background: Diabetes is a systemic metabolic imbalance involving multiple tissues/organs, and an early hallmark feature of this disease state is insulin resistance. Multifactorial interactions of genetics, prenatal environmental factors (fetal programming) and postnatal environmental factors (nutrition and activity) likely contribute to the diabetic phenotype.Animal models can serve as a valuable tool for studying diabetes disease progression and for identifying useful biomarkers of type 2 diabetes. Several inbred rodent models are available for diabetes related studies.  The GK rat is an obvious choice among available inbred models as the genetic basis for this inheritable form of diabetes is polygenic (5), unlike most other inbred rodent models that exhibit single gene defects.  Many of the characteristics of the GK rat mirror human diabetes (hyperglycemia, glucose intolerance, insulin resistance), although hyperlipidemia does not appear to be prominent in the GK rat.  Due to its polygenic mode of inheritance and 100% penetrance, the GK rat may be a useful model for human diabetes. Induced animal models can also be useful in diabetes studies. One such model is metabolic syndrome resulting from experimentally induced fetal programming (produced by maternal malnutrition or by exposure to corticosteroids in the third trimester).  Both in humans and animals,  accumulating evidence suggests that alterations in the human fetal environment can result in permanent physiologic changes that manifest as increased incidence of adult onset pathology.  Numerous epidemiological studies have forged a strong link between low birth weight and the development of metabolic syndrome in adulthood.  From such observations has arisen the concept of “fetal programming” whereby exposure to some factor(s) during crucial stages in development can permanently alter or “reset” physiologic/metabolic functions.  In the rat, exposure to corticosteroids during a “window” in third trimester gestation (CS programming) results in fetal growth retardation and insulin resistance in adult offspring.  Genetic factors play a primarily role in the etiology of diabetes in the GK rat, whereas fetal environmental factors are causative in CS programming.  (It should be noted that although altered fetal environmental effects, most likely stemming from maternal hyperglycemia, have been implicated to play some role in the decreased pancreatic B cell mass in GK rats, these effects occur earlier in gestation and therefore differ from programming by CS in late gestation.)  A comparison of the development of insulin resistance in the GK rat with development in the CS programmed rat will provide insight into genetic and fetal environmental factors in disease development. Superimposing dietary alterations (i.e., high fat feeding) (11) on both animal models may aid in the dissection of multiple interacting factors (genetic, fetal environmental factors, postnatal environmental factors) on the development and progression of insulin resistance and type 2 diabetes. Such studies may also aid in the identification of useful biomarkers for insulin resistance and type 2 diabetes in humans.
Proposed research:   Experiments are designed to study disease progression and the development of insulin resistance in two animal models:  the GK rat and the CS programmed rat, with and without a high fat diet superimposed on these models.  Animals will be maintained in our facility from weaning (GK rats) or birth (CS programmed - WKY), and body weights taken weekly.  Appropriate diets (normal or high fat) will be introduced at weaning.  Groups of animals (N=6) will be sacrificed at 5 different ages:  4, 8, 12, 16, and 20 weeks.  Plasma samples will be analyzed for markers of hyperglycemia, hyperinsulinemia, dyslipidimia, and for selected other hormonal factors which may contribute to disease etiology (adiponectin, leptin, corticosterone). At sacrifice, muscle is harvested, flash-frozen in liquid nitrogen, and warehoused in our tissue collection maintained at - 80 degrees C for this and possible future work.  Study will initially focus on examination of selected molecular markers of insulin resistance at the mRNA level in rat abdominal fat (PEPCK, IRS-1).

Keywords: Type 2 diabetes, biomarker, adipose tissue, rat, time series

overall_design: The study contains: 50 adipose tissue samples from GK (GotoKakizake) rats and 51 adipose tissue samples from WKY (WistarKyoto) rats  feeded with normal diet (ND) and high fat diet (HFD) and sacrificed at 5 different ages:  4, 8, 12, 16, and 20 weeks. So, each time point contains 5 GK samples with ND, 5 GK samples with HFD, 5 WKY samples with ND and 5 WKY samples with HFD.


title: Expression data for EglN2 conditional knockdown in T47D breast carcinoma cells

status: Public on Sep 19 2009

city: Boston

first: Qing

last: Zhang

state: MA

address: 44 Binney St

department: Medical Oncology

institute: DFCI

summary: mRNA expression was assayed from T47D SCR and T47D EGLN2 conditional knock down cell lines in order to profile the gene expression pattern regulated by EGLN2.

overall_design: T47D cells that were infected with lentivirus encoding either conditonal EglN2 shRNA or Scr control shRNA were treated with Doxycycline for either 0 hr or 48hr followed by RNA extraction.

first: Jiayun

last: Liu

first: William

last: Kaelin


title: Diabetes biomarker disease progression study in rat liver, gastrocnemius muscle, and adipose tissue


title: The differential transcriptome and ontology profiles of mural and cumulus granulosa cells in stimulated antral follicles

status: Public on Oct 15 2009

city: Perth

first: Sulev

last: Koks

state: Western Australia

address: 90 South Street

department: Centre for Molecular Medicine and Innovative Therapeutics

institute: Murdoch University

summary: Communication between various ovarian cell types is a prerequisite for folliculogenesis and ovulation. In antral follicles granulosa cells divide into two distinct populations of mural (MGC) and cumulus granulosa cells (CGC), enveloping the antrum and surrounding the oocyte, respectively. IVF offers a good opportunity for analysing their functional properties since granulosa cells can be retrieved during the puncturing procedure of stimulated follicles. The aim of this study was to compare the transcriptomes of MGC and CGC in stimulated antral follicles obtained from 19 women undergoing IVF-ICSI procedure. MGC were obtained from follicular fluid and CGC were acquired after oocyte denudation for micromanipulation. Gene expression analysis was conducted using the genome-wide Affymetrix transcriptome array. The expression profile of the two granulosa cell populations varied significantly. Out of 28 869 analysed transcripts 4 480 were differentially expressed (q-value < 10-4) and 489 showed ≥ 2-fold difference in the expression level with 222 genes up-regulated in MGC and 267 in CGC. The transcriptome of MGC showed higher expression of genes involved in immune response, hematological system function and organismal injury, while CGC had genes involved in protein degradation and nervous system function up-regulated. Cell-to-cell signalling and interaction pathways were noted in both cell populations. Furthermore, numerous novel transcripts that have not been previously described in follicular physiology were identified. In conclusion, our results provide a solid basis for future studies in follicular biology that will help to identify molecular markers for oocyte and embryo viability in IVF.

overall_design: 38 samples, 19 cumuls and 19 mural cells, paired design.

first: Agne

last: Velthut

first: Aili

last: Sarapik

first: Signe

last: Altmäe

first: Eva

last: Reinmaa

first: Leo

last: Schalkwyk

first: Cathy

last: Fernandes

first: Heena

middle: V

last: Lad

first: Ursel

last: Soomets

first: Ulle

last: Jaakma

first: Andres

last: Salumets


title: The gamma secretase inhibitor LLNle induces apoptosis of human GBM tumor initiating cells

status: Public on Dec 21 2009

city: Genova (GE)

first: Massimiliano

last: Monticone

address: Largo R. Benzi, 10

institute: IRCCS AOU San Martino – IST, Genova

summary:
Glioblastomas (GBM) are poorly differentiated astrocytic tumors arising in the Central Nervous System (CNS), which despite aggressive treatments are still characterized by a fatal outcome. Several studies have shown the existence of a subpopulation of cells within glioma tumors displaying cancer stem cells properties. As the term “tumor initiating cells” (TICs) is frequentely used to describe cells as these with cancer stem cells capacity. Because TICs promotes the tumor chemo- and radio-resistance and angiogenesis it is conceivable that finding a mean to kill these cells would lead to a better therapy for GBM.
The NOTCH gene has an important role during the CNS development, in the maintenance of dividing cells in promoting neural lineage entry of Embryonic Stem Cells and the differentiation of astroglia from the rat adult hippocampus-derived multipotent progenitors. The activation of the NOTCH signaling requires the proteolytic processing of this type I integral membrane protein by a two step process catalyzed first by a metalloprotease and then by the gamma-secretase. An increased activation of the NOTCH signaling has been implicated in several tumors types. Recently some studies showed that this pathway induces the survival/proliferation in GBM and glioma cells, and the expression of stem cell properties in glioma cells. Accordingly to these findings, the inhibition of this pathway leads to depletion of stem-like cells and blocks the engraftment in embryonal brain tumors. Furthermore, enhanced NOTCH signaling may lead to one of the tumor resistance mechanisms deployed by GBM. Targeting the NOTCH pathway specifically in GBM TICs appears therefore as a rational approach for exploring novel and hopefully more effective therapeutic strategies for the management of this malignancy.
Several molecular tools are available for targeting the Notch pathway such as specific siRNAs, shRNAs or drugs such as gamma-secretase inhibitors. Among these tools, the latter are small peptides/molecules able to inhibit the gamma-secretase by distinct mechanisms. In this study we used two drugs known as gamma-secretase inhibitors, to investigate by gene expression profiling, their ability to interfere specifically with the proliferative properties of GBM TICs previously obtained in our laboratory. Our data show that one of these two drugs, LLNle, is effective in killing these cells in vitro by activating protein catabolic process mediated by the proteasome, suggesting that preclinical studies should definitely be carried on to evaluate whether LLNle is able to significantly improve the survival in hybrid human GBM-animal models.

overall_design:
Gamma-secretase inhibitors have been proposed as drugs able to kill cancer cells by targeting the NOTCH pathway. In this study we employed two of such inhibitors, namely the z-Leu-leu-Nle-CHO (LLNle) and N-[N-(3,5-difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester (DAPT), to verify whether they were effective in vitro in the killing of human GBM tumor initiating cells (TICs).
We first established that of the two drugs only LLNle reduces the viability of GBM TICs obtained from three different patients in the low micromolar range. Cells were treated with 7.5 μM LLNle or DAPT or vehicle alone (DMSO 0.1%) and kept in a humidified 5% CO2 atmosphere at 37°C for the indicated time period (24 or 48 hours).
To establish which cellular processes are activated in GBM TICs by LLNle we generated and analyzed the gene expression profile after treatment with this compound and with DAPT and DMSO (vehicle). Our data show that LLNle induces upregulation of genes coding for proteasome subunits and subsequently mitotic arrest in these cells by repressing genes required for DNA synthesis and mitotic progression and by activation of genes acting as mitotic inhibitors. Our data are consistent with proteasome inhibition by LLNle, subsequent upregulation of proteasome activity and subsequent unleash of the apoptotic process in GBM TICs.

first: Emanuela

last: Biollo

first: Andrea

last: Fabiano

first: Francesco

last: Romeo

first: Massimo

last: Maffei

first: Marina

last: Fabbi

first: Alice

last: Melotti

first: Antonio

last: Daga

first: Walter

last: Giaretti

first: Giorgio

last: Corte

first: Patrizio

last: Castagnola


title: Gene expression of human paediatric osteosarcoma tumour samples relative to normal human osteoblasts

status: Public on Sep 08 2009

city: Toronto

first: Bekim

last: Sadikovic

state: Ontario

address: 555 University Avenue

country: Canada

institute: The SickKids Hospital and Princess Margaret Hospital

summary:
Gain or loss of genes and deregulation of gene expression can result in cumulative and progressive disruptions of normal cellular functions.
Cancer-specific changes in gene expression play an essential role in cancer occurrence, and ultimately lead to cancer-related self-sufficiency in growth signals, insensitivity to growth inhibitory signals, evasion of apoptosis, limitless replicative potential, angiogenesis, and metastasis. We aimed to analyse such changes in gene expression related to osteosarcoma.

overall_design: We performed genome-wide comparison of gene expression and identified genes that are differentialy expressed in osteosarcoma tumour samples relative to normal human osteoblasts (HOB)


title: HIF-2alpha Knockdown in A498 (VHL-/-) ccRCC cells

status: Public on Aug 27 2009

city: Jenkintown

first: Amar

last: Majmundar

state: PA

address: 434 Hillside Avenue

institute: University of Pennsylvania

summary:
HIF-2alpha is essential for (VHL-/-) ccRCC subcutaneous tumor growth in mice, and in tumor cell lines, its inhibition results in increased ROS accumulation, tumor cell death and responsiveness to radiation treatment. We have utilized transcriptional profiling to screen for putative HIF-2alpha targets genes that serve an anti-oxidant and, thus, cell survival function.
A498 ccRCC cell line was treated with control siRNA or mixture of two HIF-2alpha specific siRNA for 48 hours, and RNA was harvested.  4 independent experiments were performed, and expression was compared between control and HIF-2alpha knockdown groups.

overall_design: 8 total samples were applied to Affymetrix Human Gene 1.0 ST Arrays.  We performed two-class paired analysis using Significance Analysis of Microarrays (SAM) software to compare expression in the CT (control siRNA) and H2 (Hif2-alpha siRNA) groups.

first: Jessica

last: Bertout

middle: J

first: John

last: Gordan

first: Jennifer

last: Lam

first: Dara

last: Ditsworth

first: Brian

last: Keith

first: Eric

last: Brown

first: Katherine

middle: L

last: Nathanson

first: M

last: Simon


title: Immunomodulatory effect of 5-azacytidine (5-azaC): potential role in the transplantation setting.

status: Public on Sep 02 2009

city: Salamanca

first: José

last: Pérez Simón

state: Salamanca

address: Paseo de San Vicente, s/n

department: Hematology Service

institute: University Hospital

summary: Cytokine genes are targets of multiple epigenetic mechanisms in T lymphocytes. 5-azacytidine (5-azaC) is a nucleoside-based DNA methyltransferases (DNMT) inhibitor which induces demethylation and gene reactivation. In the current study, we analyzed the effect of 5-azaC in T-cell function and observed that 5-azaC inhibits T-cell proliferation and activation, blocking cell cycle in G0-G1 phase and decreasing the production of proinflammatory cytokines such as TNFα and IFNγ. This effect was not due to a pro-apoptotic effect of the drug but to the down-regulation of genes involved in T-cell cycle progression and activation such as CCNG2, MTCP1, CD58, and ADK and up-regulation of genes which induce cell growth arrest, such as DCUN1D2, U2AF2, GADD45B or p53. In spite of being also up-regulated, we did not find any effect of 5-azaC on the methylation pattern of FOXP3. Finally, the administration of 5-azaC at 60 and 84 hours post-transplant prevented the development of GVHD leading to a significant increase in survival in a fully mismatched BMT mouse model. In conclusion, the current study shows the effect of 5-azaC in T-lymphocytes and illustrates its role in the allogeneic transplantation setting as an immunomodulatory drug, describing new pathways which must be explored in order to prevent graft-versus-host disease.

overall_design: In order to elucidate the mechanisms involved in the effect of 5-azaC on T-cells we performed Affymetrix gene expression assays in T-cells untreated versus treated with 5-azaC at 1 μM after 2 and 4 days of culture, making a total of 12 samples (coming from time-series studies of 3 healthy individuals)

first: Luis

middle: I

last: Sánchez-Abarca

first: Silvia

last: Gutierrez-Cosio

first: Carlos

last: Santamaría

first: Teresa

last: Caballero-Velazquez

first: Belen

last: Blanco

first: Carmen

last: Herrero-Sánchez

first: Juan

last: Garcia

first: Francisco

last: González

first: Soraya

last: Carrancio

first: Pilar

last: Hernández-Campo

last: Flores

first: Consuelo

last: Del-Cañizo

first: Jesús

middle: F

last: San-Miguel

middle: A

last: Pérez-Simón


title: Genome-wide changes in DNA methylation and copy number play a role in deregulation of gene expression in osteosarcoma

first: Bekim

last: Sadikovic

type: Genome variation profiling by genome tiling array

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

overall_design: Refer to individual Series


title: Isoform-specific transcriptional activity of overlapping targets that respond to thyroid hormone receptors a1 and b1

status: Public on Aug 08 2009

city: Davis

first: Michael

last: Goodson

address: One Shields Ave

institute: University of California Davis

summary: Thyroid hormone receptors (TRs) are hormone-regulated transcription factors that control multiple aspects of physiology and development.  TRs are expressed in vertebrates as a series of distinct isoforms that exert distinct biological roles.  We wished to determine if the two most widely expressed isoforms, TRa1 and TRb1, exert their different biological effects by regulating different sets of target genes.  Using stably transformed HepG2 cells and a microarray analysis, we were able to demonstrate that TRa1 and TRb1 regulate a largely overlapping repertoire of target genes in response to T3 hormone.  However, these two isoforms display very different transcriptional properties on each individual target gene, ranging from a much greater T3-mediated regulation by TRa1 than by TRb1, to near equal regulation by both isoforms. We also identified TRa1 and TRb1 target genes that were regulated by these receptors in a hormone-independent fashion.  We suggest that it is this gene-specific, isoform-specific amplitude of transcriptional regulation that is the likely basis for the appearance and maintenance of TRa1 and TRb1 over evolutionary time.  In essence, TRa1 and TRb1 adjust the magnitude of the transcriptional response at different target genes to different levels; by altering the ratio of these isoforms in different tissues or at different developmental times, the intensity of T3 response can be individually tailored to different physiological and developmental requirements.

overall_design: TRa1, TRb1, or empty plasmid control stably transfected HepG2 cells were treated with 100 nM T3 or with ethanol carrier alone for 6h.  Three independent biological repeats were analyzed for each of the three transformant pools (empty plasmid control, TRa1, and TRb1).

first: Ivan

middle: H

last: Chan

first: Martin

last: Privalsky


title: Global expression profiling of CD4 T-cell responses to house dust mite allergens in human atopics and nonatopics.

status: Public on May 05 2009

city: Subiaco

first: Anthony

last: Bosco

state: WA

address: 100 Roberts Road

institute: Telethon Kids Institute

summary:
The aim of this study was to employ a systems-level analysis to elucidate gene expression networks operating in the CD4 T-cell responses which underpin human atopic disease.

Keywords: Disease state analysis; T-cell response

overall_design: The study design consisted of 45 subjects, including 30 HDM-sensitized atopics, and 15 nonatopic controls. For each subject, gene expression was profiled in paired samples of HDM-stimulated CD4 T-cells and in unstimulated CD4 T-cells (baseline control), thus the microarray data set consists of 90 microarrays.

first: Kathy

last: McKenna

last: Firth

first: Peter

last: Sly

first: Patrick

middle: G

last: Holt


title: Expression data from MDA-MB-231 reference and in vitro-derived subpopulations with distinct invasive potentials

status: Public on Jun 27 2009

city: Paris

first: Nadejda

last: Vintonenko

address: 1 Avenue Claude Vellefaux

country: France

institute: Inserm

summary:
To understand the link between invasion behavior and the steps of metastasis formation, we isolated invasive subpopulations from MDA-MB-231 cells in vitro using matrigel coated boyden chambers. Whole genome transcriptional profiling was used to characterize the expression changes uniquely related to invasive abilities of these cells.

In this dataset, we include expression data obtained from MDA-MB-231 invasive cells (INV cells) and those that failed to invade the coated membrane during selection (REF cells). These subpopulations have been characterized in vitro by the differences in motility and invasion, cell morphology, adhesion properties to endothelial cells or fibronectin, proliferation, and resistance to doxorubicin as well as to growth factor starvation. INV tumors xenografted in nude mice presented higher volume with more angiogenesis. In addition, when injected into blood circulation, INV cells induced more sites of metastasis in nude mice as compared to REF cells, and dramatically diminished survival by about 80%. Transcriptomic analysis showed differences in genes involved in proliferation, chemotaxis and cytoskeleton, but particularly up-regulated genes involved in negative regulation of apoptosis and down-regulated genes involved in cell adhesion or cell-cell junction. In conclusion, invasive behavior was sufficient to reveal 134 differentially expressed genes correlated with invasive cell aggressivity.

overall_design: 8 total samples were analyzed (4 for each condition). To determine differentially expressed genes, differences between both conditions were calculated on each day of measurement and analyzed using a two-sided paired t-test implemented in the anapuce R package (http://www.agroparistech.fr/mia/outil.html). Pairing was made to remove variation due to the day of measurement in the comparison between the two conditions. The variance was split between subgroups of genes with homogeneous variance (Delmar P, Robin S, Daudin J.-J. VarMixt: efficient variance modelling for the differential analysis of replicated gene expression data. Bioinformatics, 2005, 21: 4,502-508). Statistical raw p-values were adjusted for multiple comparisons using the Benjamini-Hochberg procedure (Benjamini Y, Hochberg Y. Controlling the false discovery rate: a practical and powerful approach to multiple testing. J. R. Statist. Soc. B, 1995, 57:1, 289-300) which controls the False Discovery Rate (FDR) after the estimation of the proportion of genes not differentially expressed using the smoother method (Storey JD, Tibshirani R, Statistical significance for genome-wide experiments. Proc. Nat. Acad. Sci. USA, 2003, 100, 9440-9445). The level of statistical significance was set at 0.05 for all the comparisons.

first: N

first: M

last: DiBenedetto


title: Profiling gene expression in HeLa cells following TFEB transient transfection

status: Public on Jul 01 2009

city: Houston

first: Marco

last: Sardiello

address: One Baylor Plaza

department: MHG

institute: Baylor College of Medicine

summary: Total RNA samples from three biological replicates in which TFEB was transiently overexpressed in HeLa cells by transfection using a pcDNA3 vector. As negative control, we used total RNA samples from HeLa cells transfected with an empty pcDNA3 vector.

overall_design: TFEB transfection

first: Andrea

last: Ballabio


title: TNF activates a distinct cellular program in human regulatory T cells

status: Public on Dec 31 2009

city: Tel Hashomer

first: Itamar

last: Goldstein

address: Tel Hashomer

institute: Sheba Medical Center and Tel Aviv University

summary:
Here we show that tumor necrosis factor (TNF) induced in human T-regulatory cells (Treg), as compared to conventional T cells (Tcon), a transcription program highly enriched for typical NF-κB target genes, such as: the cytokines LTA and TNF; the TNF-receptor super family members FAS, 4-1BB and OX-40; various anti-apoptotic genes; and other important immune-response genes.
As an initial approach to examine the cellular program induced by TNF in Tregs versus Tcon cells, we employed microarray gene expression analysis at 2 and 24 hrs following TNF treatment.

overall_design: Highly pure CD4+ CD25+/hi Treg and CD25- Tcon cells, obtained by FACSAria-based cell-sorting, were either treated with 50ng/mL of TNF or left untreated, for 2 and 24 hours to detect both early and late changes in gene transcription. Total RNA extracted from the various samples were analyzed using the Affymetrix HU GENE 1.0 ST Array platform for changes in gene-transcripts induced by TNF as compared to untreated media control samples.

first: Meital

last: Nagar

first: Jasmine

last: Jacob-Hirsch


title: Comparison of osteosarcoma cell lines and normal human osteoblasts

status: Public on May 01 2009


title: Knockdown of STAT1 in SCC61 tumor xenografts leads to alterations in the expression of energy metabolic pathways

status: Public on Apr 28 2009

city: Chicago

first: Sean

middle: Pravin

last: Pitroda

state: IL

address: 5841 South Maryland Avenue, MC 1105

department: Radiation and Cellular Oncology

institute: The University of Chicago

summary: Background: Signal Transducer and Activator of Transcription 1 (STAT1) has traditionally been regarded as a transmitter of interferon signaling and a pro-apoptotic tumor suppressor. Recent data have identified new functions of STAT1 associated with tumorigenesis and resistance to genotoxic stress, including ionizing radiation (IR) and chemotherapy. To investigate the mechanisms contributing to the tumorigenic functions of STAT1, we performed a combined transcriptomic-proteomic expressional analysis and found that STAT1 is associated with regulation of energy metabolism.   Methods: We generated a stable knockdown of STAT1 in the SCC61 human squamous cell carcinoma cell line, established tumor xenografts in athymic mice, and compared transcriptomic and proteomic profiles of STAT1 wild-type (WT) and knockdown (KD) untreated or irradiated (IR) tumors. Transcriptional profiling was based on Affymetrix Human GeneChip® Gene 1.0 ST microarrays. Proteomes were determined from the MS/MS data by searching against the human subset of the UniProt database. Data were analyzed using Significance Analysis of Microarrays (SAM) for RNA and Visualize software for proteins. Functional analysis was performed with Ingenuity Pathway Analysis (IPA) with statistical significance measured by Fisher’s exact test.   Results: Knockdown of STAT1 led to significant growth suppression in untreated tumors and radiosensitization of irradiated tumors. These changes were accompanied by alterations in the expression of genes and proteins of glycolysis (GG), the citrate cycle (CC) and oxidative phosphorylation (OP). Of these pathways, GG had the most concordant changes in gene and protein expression and demonstrated a STAT1-dependent expression of genes and proteins consistent with tumor-specific glycolysis. IR drastically suppressed the GG pathway in STAT1 KD tumors without significant change in STAT1 WT tumors. The STAT1 and glycolytic pathways were co-expressed in human breast tumors, and expression of STAT1-linked glycolytic genes was highly predictive of poor prognosis.   Conclusions: Our results identify a previously uncharacterized function of STAT1 in tumors: expressional regulation of genes and enzymes involved in glycolysis, the citrate cycle, and mitochondrial oxidative phosphorylation, with predominant regulation of glycolysis. STAT1-dependent transcriptional and translational regulation of glycolysis suggests a potential role for STAT1 as a transcriptional modulator of genes responsible for the Warburg effect.

overall_design: This dataset includes expression data from SCC61 cells stably transfected with a control vector or one expressing a short hairpin RNA (shRNA) to STAT1 and grown as tumor xenografts in athymic mice. A subset of tumors were treated with ionizing radiation in 5 Gy fractions over six consecutive days (total 30 Gy). Each array was hybridized with a pooled sample normalized to total RNA and consisting of RNA obtained from three independent tumor xenografts. Analysis of differentially expressed genes was based on pair-wise comparisons (STAT1 wild-type versus STAT1 knockdown for untreated or irradiated conditions) of duplicated arrays using Significance Analysis of Microarrays (SAM) version 3.02.

first: Bassam

middle: T

last: Wakim

first: Ravi

last: Sood

first: Mara

last: Beveridge

last: Beckett

first: Dhara

last: MacDermed

first: Ralph

middle: R

last: Weichselbaum

first: Nikolai

last: Khodarev


title: Inherent gene signature of stage II and III colorectal cancer leads in individual recurrence

status: Public on Jul 02 2009

city: Suita

first: masakazu

last: miyake

state: Osaka

address: Yamadaoka 2-2

department: Gastroenterological Surgery

institute: Graduate School of Medicine, Osaka University

summary: Background & Aims. The current staging system for colorectal cancer (CRC) based on TNM classification allows prediction of potential recurrence. However, it does not necessarily make reliable personalized prediction of prognosis.  In this paper we describe combination of clinicopathological data and gene signature of dissected tumor specimen with stage II and III CRC patients would improve the situation..     Methods. A total of 1978 CRC were collected over 5 years, and then 371 stage II and 322 stage III of them with more than 45.9 months records were subjected to clinicopathological feature analyses. Out of this collection, 129 stage II and III CRC cases were selected for analyses of gene expression profiles with resected specimen. The gene signatures were analyzed by repeated random divisions of the samples into training and test sets to extract discriminator genes. After testing the applicability of this discriminator set, it was subjected to validation using a newly obtained set of 69 samples.   Results. The pathological factors in solo or in combinations could not make personalized recurrence prediction, except for partial success with stage II patients. The gene signature, on the other hand, was capable of producing a set of discriminator genes, though the accuracy was yet to be improved. We observed that the best result was obtained when discriminators were selected from stage II CRC samples and used for prognosis of stage II CRC. When stage III cases were included in the process of discriminator extraction or in the process to validate samples, the results were poorer. Finally, we examined 31 independent stage II samples with a set of 30 such discriminators and were able to obtain results with 78 % accuracy, 90 % negative predictive value (NPV), and 55% positive predictive value (PPV).  Conclusions. Independent clinicopathological variables were not able to predict prognosis of individual patient, unless the factors are combined. On the other hand, gene signatures allowed accurate prediction of prognosis for individuals, especially with stage II CRC, suggesting its potential use for selection of best treatment option for individual patients. The accuracy of discriminator prediction will be further improved when we take the evolution of CRC into consideration.

overall_design: Of 198 samples, 129 represented the discovery phase and 69 represented the validation phase.


title: Expression data of human ES cells-derived CD34+CD43+CD45low cells (hemogenic endothelium-like cells) expanded upon overexpression of Sox17

status: Public on Dec 14 2012

city: Yokohama

first: Takaho

middle: A.

last: Endo

state: Kanagawa

address: 1-7-22 Suehiro, Tsurumi

department: IMS

institute: RIKEN

summary: Overexpression of transcription factor Sox17 in human ES cells-derived endothelial cells and hematopoietic cells enhances expansion of hemogenic endothelium-like cells.

overall_design: Human ES cells were differentiated for 6 days, 8 days or 12 days in EBs, then CD34+CD43-CD45- endothelial cells, CD34+CD43+CD45- pre-hematopoietic progenitor cells (HPCs) or CD34+CD43+CD45+ HPCs were isolated by fluorescence activated cell sorting (FACS) and subjected to a microarray analysis.　Some samples were plated onto OP9 cells after the isolation by FACS, and transduced with the 4OH-tamoxifen-inducible 1×FLAG-tagged Sox17-ERT retrovirus. The cells were cultured with 4OH-tamoxifen. CD34+CD43+CD45low hemogenic endothelium-like cells expanded by Sox17-ERT were collected by magnetic-activated cell sorting (MACS) and subjected to a ChIP-chip analysis.

first: Yaeko

last: Nakajima-Takagi

first: Atsushi

last: Iwama

first: Mitsujiro

last: Osawa


title: Gene expression of osteosarcoma cell (U2OS, MG63) lines relative to normal human osteoblasts (HOB)

first: Bekim

last: Sadikovic

summary:
Gain or loss of genes and deregulation of gene expression can result in cumulative and progressive disruptions of normal cellular functions.
Cancer-specific changes in gene expression play an essential role in cancer occurrence, and ultimately lead to cancer-related self-sufficiency in growth signals, insensitivity to growth inhibitory signals, evasion of apoptosis, limitless replicative potential, angiogenesis, and metastasis. We aimed to analyse such changes in gene expression related to osteosarcoma.
Keywords: Comparative

overall_design: We performed genome-wide comparison of gene expression and identified genes that are differentialy expressed in osteosarcoma (U2OS, MG63) cell lines relative to normal human osteoblasts (HOB)


title: Homogeneous monocytes and macrophages from hES cells following coculture-free differentiation in M-CSF and IL-3.

status: Public on Apr 24 2009

city: Guildford

first: Fernando

middle: O

last: Martinez Estrada

state: Surrey

address: Stag Hill

department: Faculty of Health & Medical Sciences

institute: University of Surrey

summary: In the present study we aimed at producing homogeneous populations of monocytes and macrophages from human embryonic stem (hES) cells. Human embryonic stem cell lines KCL001, KCL002, and HUES-2 were differentiated into monocytes by coculture-free differentiation with two growth factors using a three-step method. The method involved embryoid body (EB) formation in hES media, directed differentiation with macrophage colony-stimulating factor and interleukin (IL)-3, and harvest of nonadherent monocytes from the culture supernatants.  Transcriptome analysis of the esMC and the esMDM revealed a distinct myeloid signature that correlated with primary adult blood-derived monocytes and spleen tissue samples but not with other tissue samples tested.  In conclusion we have developed a simple and efficient method for producing homogeneous populations of monocytes and macrophages from hES cells. esMCs have a myeloid signature and can differentiate into functional macrophages. The method should prove useful in answering experimental questions regarding monocyte and macrophage development and biology.

overall_design: Triplicates of monocytes and macrophages derived from human embryonic stem cells were compared to a triplicate of freshly isolated human blood monocytes.

first: Karl

last: Karlsson

first: Sally

last: Cowley

first: William

last: James


title: Gene expression data from primate postnatal brain development - superior frontal gyrus

status: Public on Mar 10 2009

city: Leipzig

first: Mehmet

last: Somel

address: Deutscher Platz 6

department: Evolutionary Genetics

institute: Max Planck Institute for Evolutionary Anthropology

summary:
In development, timing is of the utmost importance, and the timing of various developmental processes are often changed during evolution. During human evolution sexual maturation has been delayed relative to other primates and this may have played a critical role for both the increase of human brain size and the rise of human-specific cognitive traits .
We measured the timing of gene expression changes in the superior frontal gyrus region of the brains of humans, chimpanzees, and rhesus macaques throughout postnatal development.

Keywords: Age series

sample_organism:
Macaca mulatta
Pan troglodytes
Homo sapiens

overall_design: Human, chimpanzee and rhesus macaque post-mortem brain samples from the superior frontal gyrus region of the prefrontal cortex were collected. The age ranges of the individuals in all three species covered the respective species' postnatal maturation period from infancy to adulthood. RNA extracted from the dissected tissue was hybridized to Affymetrix® Human Gene 1.0 ST arrays.

first: Yan

last: Zhen

first: Song

last: Guo

first: Sabine

last: Bahn

first: Maree

last: Webster

first: Cynthia

middle: S

last: Weickert

first: Michael

last: Lachmann

first: Svante

last: Pääbo

first: Philipp

last: Khaitovich


title: Understanding adult human progenitor cells

status: Public on Feb 07 2009

city: BRISTOL

first: DEBRA

middle: ANN

last: CARTER

address: BRISTOL EYE HOSPITAL, LOWER MAUDLIN STREET

department: CLINICAL SCIENCE @ SOUTH BRISTOL

institute: BRISTOL UNIVERSITY

summary:
Overarching aim is to achieve a greater understanding of the control of progenitor cells within the adult human retina within the normal and diseased retinal microenvironment. Specifically we will assess via our experimental designs: (i) the control of CD133+ retinal cell populations that display mitotic potential and differentiation and

Keywords: comparative genomic hybridization using adult human CD133+ & CD133- retinal cells

overall_design: Adult human retina was processed to produce a cell suspension, CD133+ cells were magnetically isolated to produce cell populations of CD133+ and CD133- cells. 3 retinal samples were analysed for each cell population.

first: Debra

last: Carter

first: Andrew

middle: D

last: Dick

first: Eric

last: Mayer


title: Ago2 or Dicer knockdown effects on mRNA levels in HeLa

status: Public on Feb 05 2009

city: Seoul

first: Jinju

last: Han

address: 599 Gwanangno, Gwanak-gu

institute: Seoul National University

summary:
Analysis of mRNA changes in HeLa cells following Ago2 or Dicer depletion. Dicer, a cytoplasmic RNase III, generates the mature form of small RNAs including microRNA. Ago2 is a component of an effector complex of microRNA.

Keywords: gene expression array-based (RNA / in situ oligonucleotide)

overall_design: siRNA against Ago2 or Dicer were trasnfected into HeLa cells. siRNA against GFP was used as a control. Biologically duplicated total RNAs were prepared from HeLa cells, 24 hrs and 48 hrs after siRNA transfection.

first: Young-Kook

last: Kim

first: V

middle: N


title: Diabetes biomarker disease progression study in rat gastrocnemius muscle

status: Public on Oct 21 2009

city: Washington

first: Svetlana

last: Ghimbovschi

address: 111 Michigan Ave., NW

institute: Children's National Medical Center

summary:
The goals and objectives: To study Type 2 diabetes progression and the development of insulin resistance in two animal models with and without a high fat diet  superimposed on these models.  Background: Diabetes is a systemic metabolic imbalance involving multiple tissues/organs, and an early hallmark feature of this disease state is insulin resistance. Multifactorial interactions of genetics, prenatal environmental factors (fetal programming) and postnatal environmental factors (nutrition and activity) likely contribute to the diabetic phenotype.Animal models can serve as a valuable tool for studying diabetes disease progression and for identifying useful biomarkers of type 2 diabetes. Several inbred rodent models are available for diabetes related studies.  The GK rat is an obvious choice among available inbred models as the genetic basis for this inheritable form of diabetes is polygenic (5), unlike most other inbred rodent models that exhibit single gene defects.  Many of the characteristics of the GK rat mirror human diabetes (hyperglycemia, glucose intolerance, insulin resistance), although hyperlipidemia does not appear to be prominent in the GK rat.  Due to its polygenic mode of inheritance and 100% penetrance, the GK rat may be a useful model for human diabetes. Induced animal models can also be useful in diabetes studies. One such model is metabolic syndrome resulting from experimentally induced fetal programming (produced by maternal malnutrition or by exposure to corticosteroids in the third trimester).  Both in humans and animals,  accumulating evidence suggests that alterations in the human fetal environment can result in permanent physiologic changes that manifest as increased incidence of adult onset pathology.  Numerous epidemiological studies have forged a strong link between low birth weight and the development of metabolic syndrome in adulthood.  From such observations has arisen the concept of “fetal programming” whereby exposure to some factor(s) during crucial stages in development can permanently alter or “reset” physiologic/metabolic functions.  In the rat, exposure to corticosteroids during a “window” in third trimester gestation (CS programming) results in fetal growth retardation and insulin resistance in adult offspring.  Genetic factors play a primarily role in the etiology of diabetes in the GK rat, whereas fetal environmental factors are causative in CS programming.  (It should be noted that although altered fetal environmental effects, most likely stemming from maternal hyperglycemia, have been implicated to play some role in the decreased pancreatic B cell mass in GK rats, these effects occur earlier in gestation and therefore differ from programming by CS in late gestation.)  A comparison of the development of insulin resistance in the GK rat with development in the CS programmed rat will provide insight into genetic and fetal environmental factors in disease development. Superimposing dietary alterations (i.e., high fat feeding) (11) on both animal models may aid in the dissection of multiple interacting factors (genetic, fetal environmental factors, postnatal environmental factors) on the development and progression of insulin resistance and type 2 diabetes. Such studies may also aid in the identification of useful biomarkers for insulin resistance and type 2 diabetes in humans.
Proposed research:   Experiments are designed to study disease progression and the development of insulin resistance in two animal models:  the GK rat and the CS programmed rat, with and without a high fat diet superimposed on these models.  Animals will be maintained in our facility from weaning (GK rats) or birth (CS programmed - WKY), and body weights taken weekly.  Appropriate diets (normal or high fat) will be introduced at weaning.  Groups of animals (N=6) will be sacrificed at 5 different ages:  4, 8, 12, 16, and 20 weeks.  Plasma samples will be analyzed for markers of hyperglycemia, hyperinsulinemia, dyslipidimia, and for selected other hormonal factors which may contribute to disease etiology (adiponectin, leptin, corticosterone). At sacrifice, muscle is harvested, flash-frozen in liquid nitrogen, and warehoused in our tissue collection maintained at - 80 degrees C for this and possible future work.  Study will initially focus on examination of selected molecular markers of insulin resistance at the mRNA level in rat gastrocnemius muscles (IRS-1, PDK4).

Keywords: Type 2 diabetes, biomarker, rat, muscle, time series

overall_design: The study contains: 50 gastrocnemius muscle samples from GK (GotoKakizake) rats and 51 gastrocnemius muscle samples from WKY (WistarKyoto) rats  feeded with normal diet (ND) and high fat diet (HFD) and sacrificed at 5 different ages:  4, 8, 12, 16, and 20 weeks. So, each time point contains 5 GK muscle samples with ND, 5 GK muscle samples with HFD, 5 WKY muscle samples with ND and 5 WKY muscle samples with HFD.

first: Richard

last: Almon

last: DuBois


title: Global Gene Expression in Deceased Donor Liver Transplantation with Ischemic Preconditioning

status: Public on Apr 01 2009

city: Newark

first: Ali

last: Raza

state: NJ

address: 185 South Orange Ave

department: Surgery

institute: UMDNJ

summary:
The clear benefits of ischemic preconditioning (IPC) in reducing ischemia reperfusion injury (IRI) remain indistinct in human liver transplantation.
To further understand the mechanistic aspects of IPC in human deceased donor liver transplantation (DDLT), we performed microarray analyses to determine global gene expression profiles associated with IPC administration. Donor and recipient characteristics in both groups were comparable. Clinical data from our study subset and larger trial were similar.  IPC increased expression of 10 transcripts at either time point with roles in: antioxidant defenses, immunological response, lipid biosynthesis, and xenobiotic metabolism.  IPC decreased the expression of 1 cell development related transcript.  Conclusions: 1) IPC in DDLT increased the expression of antioxidant transcripts similar to studies in animal IPC, anesthetic, and remote IPC.  2) IPC increased expression of lipogenic transcripts, which may be relevant to the clinically observed increased IRI in our IPC group. 3) Our microarray findings support our clinical observations and are compatible with the varied outcomes of hepatic IPC studies in human liver transplantation.

overall_design: We conducted a nested sub-study in 12/101 subjects enrolled in a prospective randomized trial of 10 min IPC in DDLT during 2003-2006.  Liver biopsies were performed at the end of cold storage and at 90 minutes after allograft reperfusion. Six biopsy pairs from both IPC and No IPC (STD) groups within a narrow donor risk index range were selected. Total RNA was extracted and hybridized with Affymetrix GeneChip Human Gene 1.0 ST Array. IPC effects were examined by comparing IPC vs. STD at both time points. Transcripts whose expression changed 2-fold with p<0.05 were considered significant.

first: George

last: Dikdan

first: Patricia

last: Soteropoulus

first: Helen

last: Fernandes

first: Baburao

last: Koneru


title: Two genetic pathways in myxoinflammatory fibroblastic sarcoma and morphologically similar lesions

status: Public on Jan 26 2009

city: Lund

first: Karolin

last: Nord

address: BMC C13

institute: Lund University

first: KH

last: Hallor

first: R

last: Sciot

first: J

last: Staaf

first: M

last: Heidenblad

first: A

last: Rydholm

first: HCF

last: Bauer

first: K

last: Åström

first: HA

last: Domanski

first: JM

last: Meis

first: LG

last: Kindblom

first: I

last: Panagopoulos

first: N

last: Mandahl

first: F

last: Mertens


title: Gene expression analysis of myxoinflammatory fibroblastic sarcoma, and morphologically similar lesions

summary: Global gene expression analysis was performed in cases 1, 2, 5, 6 and 8 as well as in five pooled myxofibrosarcomas using the Affymetrix GeneChip Human Gene 1.0 ST Arrays (Affymetrix Inc., Santa Clara, CA, USA). Unsupervised hierarchical clustering analysis, using Pearson correlation and average linkage clustering, separated the control pool and case 6 from the rest of the cases. Cases 1 and 5, both harboring t(1;10)(p22;q24), were in turn segregated from cases 2 and 8. Tumors with t(1;10)(p22;q24) thus seemed to have a more distinct expression pattern than tumors affected by amplification of regions on chromosome 3. Tumors with t(1;10) showed increased expression levels of NPM3 and FGF8 compared with tumors without such a translocation, including the control pool. No other gene in the 5 Mb region proximal to the breakpoint on chromosome 10 displayed a more pronounced difference between the two groups. Of the genes located in the commonly amplified region on chromosome 3, VGLL3 and CHMP2B showed a higher expression in tumors affected by the amplification.

overall_design: Cases 1, 2, 5, 6 and 8 were analyzed using the GeneChip Human Gene 1.0 ST Arrays (Affymetrix Inc, Santa Clara, CA, USA). As a control, a pool of five myxofibrosarcomas was used. Total RNA was extracted from frozen tumor biopsies using TRIzol reagent (Invitrogen, Carlsbad, CA, USA) and RNeasy (Qiagen, Valencia, CA, USA) according to the manufacturers’ instructions. RNA quality and concentration were measured using an Agilent 2100 bioanalyzer and Nanodrop ND-1000, respectively. cDNA was generated with the GeneChip® Whole Transcript (WT) cDNA Synthesis and Amplification Kit (Affymetrix) using 300 ng total RNA. Amplified cDNA was fragmented and end labeled using the GeneChip® WT Terminal Labelling Kit (Affymetrix). Subsequently, the fragmented and biotinylated cDNA was hybridized to the GeneChip® Human Gene 1.0 ST Arrays (Affymetrix). The arrays were washed and stained on a GeneChip® Fluidics Station 450 (Affymetrix) according to the manufacturer’s recommendations. Scanning was carried out with the GeneChip® Scanner 3000 and image analysis was performed using GeneChip® Operating Software (GCOS, Affymetrix). Expression data was normalized, background corrected and summarized using the RMA algorithm implemented in the Affymetrix Expression Console™ version 1.0 software. Clustering was performed with MeV 4.1.01 software (http://www.tm4.org/mev.html), using unsupervised hierarchical clustering analysis on the basis of Pearson correlation and average linkage clustering.