title: HDAC7 is a repressor of myeloid genes whose downregulation in pre-B cells is required for transdifferentiation into macrophages

status: Public on Mar 25 2014

city: Barcelona

first: Maribel

last: Parra

state: Barcelona

address: Avda. Gran Via 199-203, L'Hospitalet de Llobregat

department: Cancer Epigenetics and Biology Programme (PEBC)

institute: Bellvitge Biomedical Research Institute (IDIBELL)

summary: In the immune system HDAC7 is expressed in T cells where it regulates the expression of key genes for T cell development and function. Here we report that HDAC7 is also highly expressed in B cell precursors, where it is recruited by MEF2C to repress the activity of key genes for myeloid cell function. While HDAC7 is down-regulated during the conversion of pre-B cells into macrophages, re-expression of HDAC7 interferes with both the acquisition of the myeloid gene transcriptional program and macrophage specific cell functions. Thus, HDAC7 is a novel transcriptional repressor of lineage inappropriate genes in B cells.

overall_design: Biological duplicates of C10-MSCV and C10-HDAC7 cells were un-iduced or induced to transdifferentiate for 48 and 72 hors. Total RNA from cultured cells was extracted by Trizol and then purified. PCR amplified RNAs were hybridized against Affymetrix mouse arrays chip (Mouse Genome 430 PM strip).

first: Abul

last: Islam


title: Selective enhancement and repression of glucocorticoid receptor signaling by coregulator Hic-5

status: Public on Mar 21 2014

city: Los Angeles

first: Dai-Ying

last: Wu

state: CA

address: 1441 Eastlake Ave, NOR 6314

department: Biochemistry

country: USA

institute: University of Southern California

summary:
The glucocorticoid receptor (GR) recruits many coregulators via the well characterized AF2 interaction surface in the GR ligand binding domain, but LIM domain coregulator Hic-5 binds to the relatively uncharacterized tau2 activation domain in the hinge region of GR. Requirement of Hic-5 for glucocorticoid-regulated gene expression in U2OS osteosarcoma cells was defined by Hic-5 depletion and global gene expression analysis. Hic-5 depletion had selective and dramatic effects, positive and negative, on both activation and repression of GR target genes. For some hormone-induced genes, Hic-5 facilitated recruitment of the Mediator complex and RNA polymerase II. In contrast, many genes were not regulated by hormone until Hic-5 was depleted. On these genes Hic-5 acted at a very early step of the regulatory process, preventing efficient GR binding on enhancers, chromatin remodeling, and thus preventing glucocorticoid-driven transcriptional regulation. Overall, Hic-5 has selective and diverse roles on GR target genes, functioning as coactivator on some genes and corepressor on others, and either facilitating or opposing the glucocorticoid-driven actions of GR. Hic-5 exhibits multiple mechanisms of action, either regulating GR binding to DNA and chromatin remodeling, or facilitating later steps in transcription complex assembly.
We investigate the relationship between GR and Hic5 and identify classes of genes that respond differently when cells are induced with hormone and when Hic5 is knocked down

overall_design: We knock down Hic-5 (TGFB1I1) in U2OS cells using siRNA (siHic5_2) along with nonspecific siRNA (shNS) and assay gene expression changes at 4 different time points of hormone treatment. We also include non-infected control (NI) as a second control at each time point.

first: Rajas

last: Chodankar

first: Kimbery

middle: D

last: Siegmund

first: Michael

middle: R

last: Stallcup


title: A molecular profile of chronic cocaine abuse includes differential expression of genes regulating transcription, chromatin and dopamine cell phenotype

status: Public on Apr 01 2014

city: Detroit

middle: J

last: Bannon

state: MI

address: 540 E Canfield

department: Pharmacology

institute: Wayne State University School of Medicine

summary: Midbrain dopamine (DA)-synthesizing neurons play a key role in the addiction process, providing a compelling rationale for determining drug-induced molecular changes arising in these cells. This microarray-based study determined the profiles of midbrain gene expression in chronic cocaine abusers (n = 10) and well-matched drug-free control subjects (n = 10). Array-related procedures were performed in triplicate for each subject. Data analysis revealed that 98 array probes (corresponding to 91 genes) exhibited robust, statistically significant expression differences between chronic cocaine abusers and matched control subjects (p ≤ 0.05, FDR = 5%; with ≥ 1.4 fold-change cut-off applied). Changes in transcript abundance identified by microarray were validated by qPCR analysis in every instance examined (n = 11), regardless of the direction or magnitude of change, supporting the validity of the larger dataset of genes differentially expressed in cocaine abusers. Many of the genes exhibiting robust differential expression were associated with the regulation of transcription, chromatin function, or dopamine cell phenotype. For approximately one-half of these genes, transcript abundance was significantly predictive for subject assignment to the cocaine-abusing versus control cohort. The findings suggest that there is a molecular signature associated with core pathophysiological changes in the DA neurons of chronic cocaine abusers that can be exploited for the development of potential biomarkers and novel therapeutic targets for addiction.

overall_design: Human post-mortem midbrain gene expression derived from cocaine-related deaths are compared to midbrain gene expression of non-cocaine related deaths. Each subject was was arrayed in triplicates.

first: Sharon

middle: K

last: Michelhaugh

first: Zachary

last: Hartley

first: Steven

last: Halter

first: James

middle: A

last: David

first: Carl

last: Schmidt

first: Magen

middle: M

last: Johnson


title: Human mesenchymal cells treated with various supplements (dataset to support blima package for bead level Illumina microarray analysis)

status: Public on Mar 24 2014

city: Prague 2

first: Vojtěch

last: Kulvait

address: U Nemocnice 5

department: Institute of Pathological Physiology

country: Czech Republic

institute: Charles University in Prague - 1st Faculty of Medicine

summary: This dataset contains 9 Illumina array spots representing 3 different treatment conditions of human mesenchymal cells. First two conditions in quadruplicate and third condition on single array. The data are intended to support blima package as testing data included in blimaTestingData package available at https://bitbucket.org/kulvait/blima page. The package was developed to make analysis of Illumina bead level data possible without summarizing them to the level of probes prior to differential expression testing.

overall_design: We label the arrays used for downstream analysis A1, A2, A3, A4 for condition A (cells grown in alfa-MEM medium with 10% fetal bovine serum) and E1, E2, E3, E4 for condition E (cells grown in CellGro medium, with human serum and suplements FGF-2, EGF, M-CSF and insulin) in the set there is also included array labeled D4 for condition D (cells grown in CellGro medium, with human serum and supplements PDGF-BB, EGF, M-CSF and insulin).

first: Robert

last: Pytlík


title: Genetic differences underlying chromatin discordance between monozygotic twins [expression profile]

city: Choongchung-Buk-do

first: Hyo-Jeong

last: Ban

address: OHTAC, Yeonje-ree 643, Osong-eup, Chungwon-Koon

department: bioinformatics

country: South Korea

institute: KCDC

summary: Open chromatin is implicated in regulatory processes, and thus variation in chromatin structure may contribute to variation in gene expression and other molecular phenotypes. In this work, we performed a targeted deep sequencing to identify somatic mutations and genetic polymorphisms underlying accessible chromatin in the genomes of 72 monozygotic twins.

overall_design: Expression profiling by illumina beads array  for 36 pairs of monozygotic twins

first: Kwoneel

last: Kim


title: Expression data from Mesenchymal progenitor cells knocked down for HIF-1a or HIF-2a expression

city: Milan

first: Rosa

last: Bernardi

address: via Olgettina 60

country: Italy

institute: San Raffaele Scientific Institute

summary:
HIF-1a and HIF-2a are expressed at high levels in mesenchymal progenitors compared to more committed mesenchymal cells and hematopoietic cells. HIF-factors could therefore have a role in the regulation the biology of mesenchymal progenitors and their functions, like the non cell-autonomous maintenance of hematopoietic progenitors.
We used microarrays to detail the global program of gene expression regulated by HIF-1a or HIF-2a in mesenchymal progenitors

overall_design: Mesenchymal progenitors were FACS-sorted and cultured in low oxygen concentration for few days. Once cells started to form CFU-F colonies, we transduced them with shRNAs targeting specifically HIF-1a or HIF-2a. Four days after transduction, cells were collected and RNA extracted for microarray analysis.

first: Ylenia

last: Guarnerio


title: RUNX2 selectively attenuates or augments androgen-driven transcription in prostate cancer

status: Public on Mar 20 2014

first: Gillian

last: Little

address: 2250 Alcazar Street

summary: Prostate carcinogenesis is associated with changes in androgen signaling from driving cellular differentiation to promoting oncogenic behaviors.  RUNX2 binds the androgen receptor (AR), and ectopic expression of RUNX2 is linked to prostate cancer (PCa) progression.  We therefore investigated genome-wide the influence of RUNX2 on androgen-induced gene expression and AR DNA binding in PCa cells.  The predominant function of RUNX2 is to inhibit the androgen response, attributable in part to dissociation of AR from target genes such as the tumor suppressor NKX3-1.  At a minority of AR target genes, however, AR activity persists in the presence of RUNX2.  Some of these genes are co-operatively stimulated by androgen and RUNX2 signaling and are characterized by the presence of putative enhancers co-occupied by AR and RUNX2.  Genes synergistically stimulated by AR and RUNX2 include the invasion-promoting transcription factor SNAI2.  Indeed, co-activation of AR and RUNX2, but neither alone, stimulated PCa cell invasiveness, which was abolished by SNAI2 silencing.  Accordingly, PCa biopsies most strongly stained for SNAI2 exhibit high nuclear expression of both RUNX2 and AR.  The RUNX2-mediated locus-dependent modulation of AR activity in PCa opens a research avenue that may guide the development of novel diagnostic and therapeutic approaches to patient management.

overall_design: total RNA from C4-2B/Rx2dox cells was extracted in biological triplicates from four different conditions.  Ethanol vehicle control, dox to induce RUNX2 expression, DHT to activate androgen receptor and DHT+dox combined.

middle: H

first: Baruch

last: Frenkel


title: A Transcriptomic Reporter Assay Employing Neutrophils to Measure Immunogenic Activity of Septic Patients' Plasma (Neutrophil)

status: Public on Mar 17 2014

city: Dallas

first: Damien

last: Chaussabel

state: TX

address: 3434 Live Oak

institute: Baylor Institute for Immunology Research

summary: The wide array of molecules carried by plasma regulates critical immune functions and constitutes valuable biomarkers and therapeutic targets. In recent years the introduction of “systems approaches” has provided investigators with powerful means for assessing immune responses in patient samples on a global scale. However, while the use of genome-wide profiling technologies has become widespread, measuring the plasma proteome still presents considerable challenges. An alternative approach that consists in measuring transcriptome responses in reporter cells exposed in vitro to patient plasma has been successfully employed in a limited number of studies. Here we devised such a “Transcriptomic Reporter Assay” system to assess the immunogenicity of plasma from septic patients and evaluate its potential for biomarker discovery. Sepsis is a common, severe systemic infectious process for which physicians still lack efficient diagnostic or prognostic tools. Of the three different cell reporter systems tested, neutrophils were identified as the most capable “plasma sensor”. Compared to peripheral blood mononuclear cells and dendritic cell preparations neutrophils were best able to discriminate between plasma from septic and control subjects and responded by upregulating a robust immune transcriptional program. Additionally, the amplitude of the neutrophil transcriptomic response was shown to be associated with disease severity in two additional sets of patients. Overall, our results demonstrate both the suitability and potential clinical relevance of a neutrophil reporter assay for assessing immunopathogenic processes in a complex and severe condition such as sepsis.

overall_design: Polymorphonuclear neutrophils (PMNs) were isolated from two healthy donors. Plasma samples were obtained from patients with culture-confirmed sepsis (n=12) and from uninfected controls (n=12). PMNs were cultured for 6 hours in medium alone, plasma from patients with sepsis, plasma from uninfected controls, and LPS using a final concentration of 20%. Transcriptional profiles were acquired using Illumina HumanHT12 V4 BeadChips.

first: Prasong

last: Khaenam

first: Darawan

last: Rinchai

first: Laurent

last: Chiche

first: Matthew

middle: C

last: Altman

first: Surachat

last: Buddhisa

first: Chidchamai

last: Kewcharoenwong

first: Duangchan

last: Suwannasaen

last: Mason

first: Elizabeth

last: Whalen

first: Wattanachai

last: Susaengrat

first: Kimberly

last: O’Brien

first: Quynh-Ahn

last: Nguyen

first: Vivian

last: Gersuk

first: Peter

last: Linsley

first: Ganjana

last: Lertmemongkolchai


title: A Transcriptomic Reporter Assay Employing Neutrophils to Measure Immunogenic Activity of Septic Patients' Plasma (Expt. 2)

overall_design: Polymorphonuclear neutrophils (PMNs) were isolated a healthy donor. Plasma samples were obtained from a first set of patients with culture-confirmed sepsis (n=29) and from uninfected controls (n=17). PMNs were cultured for 6 h in medium alone, plasma from patients with sepsis, plasma from uninfected controls, and LPS using a final concentration of 20%. Transcriptional profiles were acquired using Illumina HumanHT12 V4 BeadChips.

first: Michael


title: A Transcriptomic Reporter Assay Employing Neutrophils to Measure Immunogenic Activity of Septic Patients' Plasma (DC)

overall_design: Monocytes were isolated from two healthy donors. Monocytes were cultured with interleukin 4 (IL4) and granulocyte-macrophage colony-stimulating factor (GM-CSF) to generate monocyte derived dendritic cells (MoDCs). Plasma samples were obtained from patients with culture-confirmed sepsis (n=12) and from uninfected controls (n=12). MoDCs were cultured for 6 h in medium alone, plasma from patients with sepsis, plasma from uninfected controls, and LPS using a final concentration of 20%. Transcriptional profiles were acquired using Illumina HumanHT12 V4 BeadChips.


title: A Transcriptomic Reporter Assay Employing Neutrophils to Measure Immunogenic Activity of Septic Patients' Plasma (PBMC)

overall_design: Peripheral blood mononuclear cells (PBMCs) were isolated from two healthy donors. Plasma samples were obtained from patients with culture-confirmed sepsis (n=12) and from uninfected controls (n=12). PBMCs were cultured for 6 h in medium alone, plasma from patients with sepsis, plasma from uninfected controls, and LPS using a final concentration of 20%. Transcriptional profiles were acquired using Illumina HumanHT12 V4 BeadChips.


title: Molecular pathways reflecting poor intrauterine growth are imprinted in Wharton’s jelly derived Mesenchymal Stem Cells [set4]

status: Public on Mar 31 2014

city: Singapore

first: Dennis

last: Wang

address: 30 Medical Drive

country: Singapore

institute: Singapore Institute for Clinical Sciences

summary:
In order to identify gene-expression patterns in mesenchymal stem cells associated with different birth weights and intrauterine growth parameters,
we conducted comparative microarray gene expression experiments prior and post acute insulin stimulation

overall_design: Mesenchymal stem cells (MSC) were extracted from the Wharton's jelly of the umbilical cord of normal and SGA babies. The MSCs were expended in culture. The MSCs were subjected to acute insulin treatment for various time points. The RNA of these cells were collected for gene-expression study.

first: Walter

last: Stunkel


title: A Human Pluripotent Stem Cell Surface N-Glycoprotein resource including New Markers, Extracellular Epitopes, and Drug Targets

status: Public on Mar 12 2014

city: Hong Kong

first: Kenneth

last: Boheler

address: 5 Sassoon Road

country: China

institute: The University of Hong Kong

summary: Detailed knowledge of cell surface proteins for isolating well-defined populations of human pluripotent stem cells (hPSCs) would significantly enhance their characterization and translational potential. Normal H9 human embryonic stem cells and the KB3 human induced pluripotent stem cell lines were analyzed by Cell Surface Capture Technology, and in parallel transcript profiles from five independent samples (i.e., Replicas 1-5 for each) were performed to facilitate protein and transcriptomic comparisons.

overall_design: The study compared gene expression profiles of pluripotent stem cells with Cell Surface Capture technology generated N-glycoprotein surfaceome analyses of the same cell types.


title: A Transcriptomic Reporter Assay Employing Neutrophils to Measure Immunogenic Activity of Septic Patients' Plasma

first: Damien

last: Chaussabel

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

overall_design: Refer to individual Series


title: Evoking potent RNAi response in cancer using novel 2’-OMe-phosphorodithioated modified siRNAs

status: Public on Mar 10 2014

city: Houston

first: Sangbae

last: Kim

address: 1 Baylor plaza

department: HGSC

institute: Baylor Colleage of Medicine

summary: Use of MePS2-modified siRNAs to target GRAM Domain Containing 1B (GRAMD1B), a novel protein in taxane resistance.

overall_design: Two groups of samples are included: 1. siControl treated HeyA8-MDR and 2. siGRAMD1B treated HeyA8-MDR. Gene expression profiles of siGRAMD1B-HeyA8-MDR cells were compared to that of siControl-HeyA8MDR cells.

first: Sherry

last: Wu

first: Anil

last: Sood

first: Ju-Seog

last: Lee


title: Genome-wide analysis of vascular smooth muscle cells treated by etonogestrel (ETO), medroxyprogesterone acetate (M) or progesterone (P)

status: Public on Mar 11 2014

city: Columbus

first: S.

middle: Joseph

last: Huang

state: Ohio

department: Ob/Gyn

institute: The Ohio State University

summary: Analysis of long-acting progestin-only contraceptive (LAPC)-mediated differential gene expression in vascular smooth muscle cells (VSMCs). We tested the hypothesis that LAPCs influence expression of several common genes associated with differentiation, survival and migration of VMSCs. Results provide important information of several common and unique gene expression profiles in cultured VSMC treated with ETO, M or P. Among ETO and M responsive genes, up- or down-regulated common genes were determined and used further confirmation and in vitro functional analyses.

overall_design: Total RNA (n=3) obtained from cultured aortic VSMCs treated with vehicle only (CON) or ETO or M or P for 6h.

first: Murat

last: Basar

first: Umit

last: Kayisli

first: Ozlem

last: Guzeloglu-Kayisli

first: S


title: Excision of viral reprogramming cassettes by Cre protein transduction enables rapid, robust and efficient derivation of transgene-free human iPS cells

city: Bonn

first: Asifiqbal

last: Kadari

state: NRW

address: sigmund-freud strasse 25

department: stem cell engineering group

country: Germany

institute: Institute of reconstructive neurobiology, University of Bonn

summary: Emergence of induced pluripotent stem cells (iPSC) technology has paved novel routes for regenerative medicine. iPSCs offer the possibilities of disease modeling, drug toxicity studies as well as cell replacement therapies by autologous transplantation. Classical protocols of iPSC generation harness infection by retro- or lenti-viruses. Although such integrating viruses represent very robust tools for reprogramming, the presence of viral transgenes in iPSCs is deleterious as it holds the risk of insertional mutagenesis leading to malignant transformation. Moreover, remaining reprogramming transgenes have been shown to affect the differentiation potential of iPSCs. More recently, alternative protocols have been explored to derive transgene-free iPSC, including use of transposons, mRNA transfection, episomal plasmid transfection, and infection with non-integrating viruses such as Sendai virus. However, the utility of such protocols remains limited due to low efficiency and narrow range of cell specificity. In this study we aim at combining the robustness of lentiviral reprogramming with the high efficacy of Cre recombinase protein transduction to readily delete reprogramming transgenes from iPSCs. We demonstrate rapid generation of transgene-free human iPSCs by excising the loxP-flanked reprogramming cassette employing direct delivery of biologically active Cre protein. By genome-wide analysis and targeted differentiation towards the cardiomyocyte lineage, we show that transgene-free iPSCs do resemble more to human ESCs and has better differentiation potential than iPSCs before Cre transduction. Our study provides a simple, rapid and robust protocol for the generation of superior transgene-free iPSCs suitable for disease modeling, tissue engineering and cell replacement therapies.

overall_design: mRNA extracted from human Fibroblasts (AR1034ZIMA), human Embryonic Stem Cell line I3 (hES I3), three human induced Pluripotent Stem Cell clones 1, 1.2 and 1.4 (fl-ARiPS cl1, del-ARiPS cl 1.2, del-ARiPS cl1.4) has been hybridized on Illumina Human HT-12 (version 4 revision 2) arrays for genome wide expression analysis. Samples were run at least as duplicate technical replicates. Differential gene expression analysis has been performed on the grouped expression data with the human embryonic stem cells (hES I3) group as the reference.

first: Min

last: Lu

first: Ming

last: Li

first: Thileepan

last: Sekaran

first: Rajkumar

middle: P

last: Thummer

first: Naomi

last: Guyette

first: Vi

last: Chu

first: Frank

last: Edenhofer


title: Genome-wide effect of conditioned media supernatant (CMS) derived from human endometrial stromal cells treated by long-acting progestin-only contraceptives (LAPCs) on Human endometrial endothelial cells

status: Public on Mar 08 2014

summary: Analysis of differential gene expression in Human endometrial endothelial cells (HEECs) incubated with CMS derived from human endoemtrial stromal cells (HESCs) treated by LAPCs. We tested the hypothesis that paracrine factors sectreted from HESCs treated LAPCs, etonogestrol (ETO) or medorxyprogesterone acetate (M) influence several common genes associated with survival in HEECs. Thefore, whole genome analyses were performed in HEECs treated with HESC derived CMS obtained from vehicle (estrodial (E) as control) or progesterone (P) or ETO or M incubations under hypoxia (HX) and normoxia (NX). Results provide important information of several common and unique gene expression profiles in cultured HEECs treated with  HESC CMS from P or ETO or M incubations. Among M and ETO responsive genes, up- or down-regulated survival related common genes were determined and used further confirmation and in vitro functional analyses.

overall_design: Total RNA (n=3) obtained from cultured HEECs incubated 6h with vehicle- or P4- or ETO- or MPA- treated NX or HX conditioned HESC-derived CMS.


title: Identification of rheumatoid arthritis and osteoarthritis patients by transcriptome-based rule set generation [Jena]

status: Public on Mar 05 2014

city: Eisenberg

first: Raimund

middle: W

last: Kinne

address: Klosterlausnitzer Str. 81

department: Department of Orthopedics

institute: Jena University Hospital

summary:
Discrimination of rheumatoid arthritis (RA) patients from patients with other inflammatory/degenerative joint diseases or healthy individuals purely on the basis of genes differentially expressed in high-throughput data has proven very difficult. Thus, the present study sought to achieve such discrimination by employing a novel unbiased approach using rule-based classifiers.  Three multi-center genome-wide transcriptomic data sets (Affymetrix HG- U133 A/B) from a total of 79 individuals, including 20 healthy controls (control group - CG), as well as 26 osteoarthritis (OA) and 33 RA patients, were used to infer rule- based classifiers to discriminate the disease groups. The rules were ranked with respect to Kiendl’s statistical relevance index, and the resulting rule set was optimized by pruning. The rule sets were inferred separately from data of one of three centers and applied to the two remaining centers for validation. All rules from the optimized rule sets of all centers were used to analyze their biological relevance applying the software Pathway Studio.
The optimized rule sets for the three centers contained a total of 29, 20, and 8 rules (including 10, 8, and 4 rules for ‘RA’), respectively. The mean sensitivity for the prediction of RA based on six center-to-center tests was 96% (range 90% to 100%), that for OA 86% (range 40% to 100%). The mean specificity for RA prediction was 94% (range 80% to 100%), that for OA 96% (range 83.3% to 100%). The average overall accuracy of the three different rule-based classifiers was 91% (range 80% to 100%). Unbiased analyses by Pathway Studio of the gene sets obtained by discrimination of RA from OA and CG with rule-based classifiers resulted in the identification of the pathogenetically and/or therapeutically relevant interferon-gamma and GM-CSF pathways.  First-time application of rule-based classifiers for the discrimination of RA resulted in high performance, with means for all assessment parameters close to or higher than 90%. In addition, this unbiased, new approach resulted in the identification not only of pathways known to be critical to RA, but also of novel molecules such as serine/threonine kinase 10.

first: Dirk

last: Woetzel

first: Rene

last: Huber

first: Peter

last: Kupfer

last: Pohlers

first: Michael

last: Pfaff

first: Dominik

last: Driesch

first: Thomas

last: Haeupl

last: Koczan

last: Stiehl

first: Reinhard

last: Guthke


title: Identification of rheumatoid arthritis and osteoarthritis patients by transcriptome-based rule set generation [Leipzig]

city: Leipzig

address: Liebigstr. 24

department: Institute of Pathology

institute: University of Leipzig

summary: Discrimination of rheumatoid arthritis (RA) patients from patients with other inflammatory/degenerative joint diseases or healthy individuals purely on the basis of genes differentially expressed in high-throughput data has proven very difficult. Thus, the present study sought to achieve such discrimination by employing a novel unbiased approach using rule-based classifiers.  Three multi-center genome-wide transcriptomic data sets (Affymetrix HG- U133 A/B) from a total of 79 individuals, including 20 healthy controls (control group - CG), as well as 26 osteoarthritis (OA) and 33 RA patients, were used to infer rule- based classifiers to discriminate the disease groups. The rules were ranked with respect to Kiendl’s statistical relevance index, and the resulting rule set was optimized by pruning. The rule sets were inferred separately from data of one of three centers and applied to the two remaining centers for validation. All rules from the optimized rule sets of all centers were used to analyze their biological relevance applying the software Pathway Studio.  The optimized rule sets for the three centers contained a total of 29, 20, and 8 rules (including 10, 8, and 4 rules for ‘RA’), respectively. The mean sensitivity for the prediction of RA based on six center-to-center tests was 96% (range 90% to 100%), that for OA 86% (range 40% to 100%). The mean specificity for RA prediction was 94% (range 80% to 100%), that for OA 96% (range 83.3% to 100%). The average overall accuracy of the three different rule-based classifiers was 91% (range 80% to 100%). Unbiased analyses by Pathway Studio of the gene sets obtained by discrimination of RA from OA and CG with rule-based classifiers resulted in the identification of the pathogenetically and/or therapeutically relevant interferon-gamma and GM-CSF pathways.  First-time application of rule-based classifiers for the discrimination of RA resulted in high performance, with means for all assessment parameters close to or higher than 90%. In addition, this unbiased, new approach resulted in the identification not only of pathways known to be critical to RA, but also of novel molecules such as serine/threonine kinase 10.


title: Kin28 Regulates the Transient Association of Mediator with Core Promoters

city: Montreal

first: Francois

last: Robert

state: QC

address: 110 av des Pins Ouest

country: Canada

institute: IRCM

type: Genome binding/occupancy profiling by genome tiling array

summary: Mediator is an essential, broadly utilized eukaryotic transcriptional co-activator. How and what it communicates from activators to RNA polymerase II remains an open question. Here we performed genome-wide location profiling of yeast Mediator subunits. Mediator is not found at core promoters but rather occupies the upstream activating sequence (UAS), upstream of the pre-initiation complex. In the absence of Kin28/CDK7 kinase activity, or in cells where the CTD is mutated to replace Ser5 with alanines, however, Mediator accumulates at core promoters together with RNAPII. We propose that Mediator is quickly released from promoters upon Ser5 phosphorylation by Kin28/CDK7, which also allows for RNAPII to escape from the promoter.

overall_design: We took a systematic approach to examine the genome-wide distribution (using ChIP-chip) of the various Mediator subunits. Mediator occupancy was also assayed in mutants for most of the CTD kinases and in strains where some CTD serines had been replaced by alanines. Mediator ChIPs were performed with Myc-tagged subunits, except in some preliminary experiments where polyclonal antibodies were used. Most ChIPs (in Cy5) were hybridyzed against a control ChIP sample from an isogenic non-tagged strain (in Cy3). In some of the preliminary experiments, non immunoprecipitated DNA (input) was used as the control. In addition to Mediator ChIPs, the project includes TFIIB and RNAPII (Rpb3) ChIP-chip datasets. All ChIP-chip experiments were done in duplicates except for the preliminary experiments that were done in monoplicat for the most part. Each microarray was normalized using the Lima Loess and replicates were combined using a weighted average method as previously described (Pokholok et al., 2005).

first: Célia

last: Jeronimo

first: François


title: Gene expression analysis of PBMC from HIV and HIV/TB co-infected patients

city: Philadelphia

first: Louise

last: Showe

state: PA

address: 3601 Spruce St

institute: The Wistar Institute

summary: Diagnosis of TB, especially in the presence of an HIV co-infection, can be challenging when using conventional diagnostic methods.  In this study, we analyzed global gene expression data from PBMC of patients that were either mono-infected with HIV or co-infected with HIV and TB in order to identify a TB-specific gene signature.

overall_design: Total RNA obtained from PBMC from a South African cohort.  Microarry analysis was performed to compare gene expression in patients either infected with HIV or co-infected with HIV/TB.

first: Noor

last: Dawany

first: Andrew

middle: V

last: Kossenkov

first: Celia

last: Chang

first: Prudence

last: Ive

first: Francesca

last: Conradie

first: Wendy

last: Stevens

first: Ian

last: Sanne

first: Livio

last: Azzoni

first: Luis

last: Montaner


title: Systematic design and functional analysis of artificial microRNAs as an approach for multi-gene targeting

status: Public on Feb 28 2014

city: Seattle

first: Jason

last: Arroyo

state: WA

address: 1100 Fairview Ave N

department: Human Biology

institute: Fred Hutchinson Cancer Research Center

summary: Unlike short interfering RNAs (siRNAs), which are commonly designed to repress a single messenger RNA (mRNA) target through perfect base pairing, microRNAs (miRNAs) are endogenous small RNAs that have evolved to concurrently repress multiple mRNA targets through imperfect complementarity. MicroRNA target recognition is primarily determined by pairing of the miRNA seed sequence (nucleotides 2–8) to complementary match sites in each mRNA target. Whereas siRNA technology is well established for single target knockdown, the design of artificial miRNAs for multi-target repression is largely unexplored. We designed and functionally analysed over 200 artificial miRNAs for simultaneous repression of pyruvate carboxylase and glutaminase by selecting all seed matches shared by their 3′ untranslated regions. Although we identified multiple miRNAs that repressed endogenous protein expression of both genes, seed-based artificial miRNA design was highly inefficient, as the majority of miRNAs with even perfect seed matches did not repress either target. Moreover, commonly used target prediction programs did not substantially discriminate effective artificial miRNAs from ineffective ones, indicating that current algorithms do not fully capture the features important for artificial miRNA targeting and are not yet sufficient for designing artificial miRNAs. Our analysis suggests that additional factors are strong determinants of the efficacy of miRNA-mediated target repression and remain to be discovered.

overall_design: 293T cells were transiently transfected with artificial miRNAs or non-targeting control (Allstars siRNA, Qiagen). Three replicate transfections were performed for each miRNA or control. Total RNA was extracted 48 hours after transfection.

first: Emily

middle: N

last: Gallichotte

first: Muneesh

last: Tewari


title: Histone demethylase RBP2 is critical for breast cancer progression and metastasis

status: Public on Feb 27 2014

city: New Haven

first: Qin

last: Yan

state: CT

address: 310 Cedar St

department: Pathology

institute: Yale University

summary: To determine the roles of RBP2 in breast cancer metastasis, MDA-MD-231 cells were transfected with siRNAs against RBP2 or luciferase control, followed with gene expression microarray analysis and gene set enrichment analysis. These analyses revealed that RBP2 knockdown significantly decreased expression of genes linked to breast cancer metastasis to lung.

overall_design: Total RNA obtained from the breast cancer cell line MDA-MB231 72 hours after transfection with siRNA targeting KDM5A/RBP2/JARID1A, and targeting Luceferase gene as a control.

first: Jian

last: Cao

first: Zongzhi

last: Liu

first: Don

last: Nguyen


title: Notch signaling regulates neural crest differentiation from human pluripotent stem cells

status: Public on Mar 03 2014

city: Turku

first: Kartiek

last: Kanduri

address: Tykistokatu 6

country: Finland

institute: Turku Centre for Biotechnology

summary: The aim of the dataset was to study on genome-wide level the effect of Notch inhibition in gene expression on neural crest differentiation of human embryonic stem cells under chemically defined conditions.

overall_design: Total RNA from hESCs, hESC-derived neural crest, hESC-derived neural crest+DAPT, and hESC-derived neural stem cells was collected and compared at their global gene expression level. Samples from 3 biological replicates were analysed.

first: Parinya

last: Noisa

first: Carina

last: Lund

first: Riikka

first: Harri

last: Lähdesmäki

first: Riitta

last: Lahesmaa

first: Karolina

last: Lundin

first: Hataiwan

last: Chokechuwattanalert

first: Timo

last: Otonkoski

last: Tuuri

first: Taneli

last: Raivio


title: Analysis of global gene expression in pluripotent stem cell (PSC) derived neural stem cells (NSCs) and neurons and astrocytes differentiated from these NSCs

city: Bethesda

first: Nasir

last: Malik

state: MD

address: 50 South Drive/Room 1146

department: NIAMS

institute: NIH

summary: Gene expression was compared between two control PSC lines and neurons and astrocytes differentiated from each line.

overall_design: Total RNA obtained from fetal astrocytes from two sources compared to total RNA obtained from two neuronal stem cell lines.

first: Malik

last: Nasir

first: Rao

last: Mahendra


title: An extracellular matrix-driven positive feedback loop regulates translation in Idiopathic Pulmonary Fibrosis

status: Public on Mar 04 2014

city: Minneaplois

first: Matthew

last: Parker

state: MINNESOTA

address: 420 Delaware St. SE

department: Department of Medicine

institute: University of Minnesota

summary: Analysis of gene expression of lung fibroblasts seeded onto decellularized extracellular matrix (ECM). Experiment had 2x2 design where fibroblasts from idiopathic pulmonary fibrosis (IPF) or control patients were seeded onto decelluarized lung tissue from IPF or control patients allowing for determination of gene expression differences that were driven by IPF ECM and which differences were driven by the IPF fibroblast.

overall_design:
Lung fibroblasts from 5 patients with idiopathic pulmonary fibrosis and 5 control patients were cultured on decellularized ECM from IPF or control lung. Total RNA and polyribosome RNA were isolated after the cells were cultured on the decellularized ECM for 18 hours.
When possible, a control cell line and a diseased cell line were cultured (and processed) simultaneously to minimize the effect of experimental variance induced by running the experiment at different times.Samples with the same batch number (provied in the sample 'characteristics' field) were cultured and processed at the same time.

first: Daniel

last: Rossi

first: Mark

middle: S

last: Peterson

first: Kristina

last: Sikstrom

first: Eric

last: White

first: Craig

last: Henke

first: Ola

last: Larsson

first: Peter

middle: B

last: Bitterman


title: miR-146a promotes the initiation and progression of melanoma  by activating Notch signaling

status: Public on Feb 18 2014

city: Birmingham

first: Narendra

last: Wajapeyee

state: AL

address: 720 20th Street South, Kaul 540A

department: Department of Biochemistry and Molecular Genetics

institute: University of Alabama at Birmingham

summary: Oncogenic mutations in BRAF and NRAS occur in 70% of melanomas. Here we identify a microRNA, miR-146a, that is highly upregulated by oncogenic BRAF and NRAS. Expression of miR-146a increases the ability of human melanoma cells to proliferate in culture and form tumors in mice, whereas knockdown of miR-146a has the opposite effects. We show these oncogenic activities are due to miR-146a targeting the NUMB mRNA, a repressor of Notch signaling. Previous studies have shown that pre-miR-146a contains a single nucleotide polymorphism (C>G rs2910164). We find that the ability of pre-miR-146a/G to activate Notch signaling and promote oncogenesis is substantially higher than that of pre-miR-146a/C. Analysis of melanoma cell lines and matched patient samples indicates that during melanoma progression pre-miR-146a/G is enriched relative to pre-miR-146a/C, resulting from a C-to-G somatic mutation in pre-miR-146a/C. Collectively, our results reveal a central role for miR-146a in the initiation and progression of melanoma.

overall_design: SKMEL28 melanoma cell line stably expressing either an empty vector or  pre-miR146a with either C or G SNP (SKMEL28-FG12, SKMEL28-miR-146a/C and SKMEL28-miR-146a/G) were used to prepare total RNA.  Microarray analysis was performed by using biological replicates for each stable cell lines for a total of 6 samples.

first: Matteo

last: Forloni

first: Shaillay

last: Dogra

first: Yuying

last: Dong

first: Darryl

last: Conte

first: Julie

middle: L

last: Zhu

first: Jianhong

last: Ou

first: April

last: Deng

first: Meera

last: Mahalingam

first: Michael

last: Green


title: Identification of rheumatoid arthritis and osteoarthritis patients by transcriptome-based rule set generation

status: Public on Feb 21 2014

city: Berlin

first: Thomas

last: Häupl

address: Charitéplatz 1

department: Rheumatology

institute: Charité

summary:
Discrimination of rheumatoid arthritis (RA) patients from patients with other inflammatory/degenerative joint diseases or healthy individuals purely on the basis of genes differentially expressed in high-throughput data has proven very difficult. Thus, the present study sought to achieve such discrimination by employing a novel unbiased approach using rule-based classifiers.  Three multi-center genome-wide transcriptomic data sets (Affymetrix HG- U133 A/B) from a total of 79 individuals, including 20 healthy controls (control group - CG), as well as 26 osteoarthritis (OA) and 33 RA patients, were used to infer rule- based classifiers to discriminate the disease groups. The rules were ranked with respect to Kiendl’s statistical relevance index, and the resulting rule set was optimized by pruning. The rule sets were inferred separately from data of one of three centers and applied to the two remaining centers for validation. All rules from the optimized rule sets of all centers were used to analyze their biological relevance applying the software Pathway Studio.
The optimized rule sets for the three centers contained a total of 29, 20, and 8 rules (including 10, 8, and 4 rules for ‘RA’), respectively. The mean sensitivity for the prediction of RA based on six center-to-center tests was 96% (range 90% to 100%), that for OA 86% (range 40% to 100%). The mean specificity for RA prediction was 94% (range 80% to 100%), that for OA 96% (range 83.3% to 100%). The average overall accuracy of the three different rule-based classifiers was 91% (range 80% to 100%). Unbiased analyses by Pathway Studio of the gene sets obtained by discrimination of RA from OA and CG with rule-based classifiers resulted in the identification of the pathogenetically and/or therapeutically relevant interferon-gamma and GM-CSF pathways.
First-time application of rule-based classifiers for the discrimination of RA resulted in high performance, with means for all assessment parameters close to or higher than 90%. In addition, this unbiased, new approach resulted in the identification not only of pathways known to be critical to RA, but also of novel molecules such as serine/threonine kinase 10.

overall_design: Synovial tisssue from healthy joints, OA joints, and RA joints.

first: Dirk

last: Woetzel

first: Rene

last: Huber

last: Kupfer

last: Pohlers

last: Pfaff

first: Dominik

last: Driesch

last: Häupl

last: Koczan

last: Stiehl

first: Reinhard

last: Guthke

first: Raimund

last: Kinne


title: Expression profiling of VCaP prostate cancer cells with ERG knockdown or WP1130 treatment

city: Dallas

first: Ralf

last: Kittler

state: Texas

address: 5323 Harry Hines Blvd

institute: University of Texas Southwestern Medical Center

summary: Microarray studies was performed to analyze gene expression profiling in VCaP cells after ERG was silenced by two different siRNAs or VCaP cells were treated by WP1130.

overall_design: RNA samples, containing 3 replicates for each sample, were extracted from VCaP cells at 72 hours siERG post-transfection or 24 hours WP1130 treatment. After biotin labeling, samples were hybrided on the Illumina Human HT12v4.0 Expression Beadchip


title: Contribution of the scavenger receptor MARCO to TLR-induced activation of dendritic cells

city: Boston

first: M. Simo

last: Arredouani

state: MA

address: 330 Brookline Ave.

department: Surgery

institute: Beth Israel Deaconess Medical Center

summary: To gain additional insight into the role of MARCO in dendritic cell activation and function, we profiled transcriptomes of mouse splenic dendritic cells obtained from MARCO deficient mice and their wild type counterparts under resting and activating conditions.

overall_design: Pure dendritic cells from the spleens of WT and MARCO KO mice were cultured overnight in PBS or Lipopolysaccharide (LPS). Gene expression profiling was performed using Affymetrix platform.

first: M


title: Genome wide expression analysis of FABP4 modulation in urothelial cancer cell UM-UC14 and UM-UC9

status: Public on Feb 17 2014

first: Tiewei

last: Cheng

state: United States

address: 7777 Knight Rd.

institute: UT MD Anderson Cancer Center

summary: Analysis of gene expression profiling in FABP4 modulation UM-UC14 and Um-UC9 cells. The overall objective was to identify genes regulated by PPARγ signaling pathway in particular FABP4

overall_design: Total RNA was obtained from FABP4 modulated (either by FABP4 knockdown or rosiglitazone treatment) UM-UC14 or UM-UC9 cells compared to untreated control

first: Beat

last: Roth

first: Woonyoung

last: Choi


title: Genome-wide espression kinetics of children progressing to clinical Type 1 diabetes (T1D).

status: Public on Feb 20 2014

first: Henna

last: Kallionpää

address: P.O. Box 123

department: Turku Centre for Biotechnology

institute: University of Turku

summary: To unravel genes and molecular pathways involved in the pathogenesis of type 1 diabetes (T1D), we performed genome-wide gene expression profiling of prospective venous blood samples from children developing T1D-associated autoantibodies or progressing towards clinical diagnosis.

overall_design: 247 peripheral blood RNA samples from 18 prediabetic children and their matched controls were analyzed with Illumina Human HT-12 Expression BeadChips version 3 arrays, in order to study the gene expression changes occuring during the pathogenesis of Type 1 diabetes (T1D). Each case child (with T1D-specific autoantibodies) was matched with a persistently autoantibody-negative control child, with the same HLA-DQB1 risk category, gender, and place and date of birth. Two control children were selected for T1D cases 3, 5, 13 and 17. Seroconversion is determined as the first detection of T1D-specific autoantibody/autoantibodies (ICA titre >4 JDFU, IAA >3.47 RU, GADA >5.4 RU, IA-2A >0.43 RU, ZnT8A >0.61 RU).

first: Laura

last: Elo

first: Juha

last: Mykkänen

first: Essi

last: Laajala

first: Matti

last: Vaarma

first: Teemu

first: Heikki

last: Hyöty

first: Jorma

last: Ilonen

first: Riitta

last: Veijola

first: Tuula

last: Simell

first: Mikael

last: Knip

first: Harri

last: Lähdesmäki

first: Olli

last: Lahesmaa

first: Isis

last: Ricaño-Ponce

first: Cisca

last: Wijmenga


title: Genome-wide gene expression analysis of the whole blood leukocyte transcriptional response to endotoxin treatment

status: Public on Feb 24 2014

city: Msida

first: Brendon

last: Scicluna

address: Msida campus

country: Malta

institute: University of Malta

summary: Several inflammatory diseases respond to TNFα inhibitors, implicating TNFα signaling in the pathogenesis of these conditions. Here, we set out to map the systemic genome-wide transcriptome influenced by TNFα release. We performed an intravenous endotoxin challenge in 16 healthy subjects, half of whom were pretreated with the soluble TNF receptor fusion protein, etanercept. Whole-blood leukocyte total RNA was isolated using the PAXgeneTM tube and PAXgeneTM RNA isolation system (PreAnalytiXTM, Qiagen) as described by the manufacturer. Total RNA yield and purity (260nm:280nm) were determined spectrophotometrically. The integrity of the re-suspended total RNA was determined using the RNA Nano Chip Kit on the Bioanalyzer 2100 and the 2100 Expert software (Agilent). To increase the sensitivity of our gene expression assay, the overload of globin mRNA of whole blood samples was reduced by applying the human GLOBINclear kit (Ambion/Appied Biosystems). Synthesis, amplification and purification of anti-sense RNA was performed using 300 ng of enriched mRNA per sample and the Illumina TotalPrep RNA Amplification Kit (Ambion/Applied Biosystems) following the Illumina Sentrix Array Matrix expression protocol. A total of 750 ng biotinylated cRNA was hybridized onto the HumanHT-12v3 expression BeadChips (Illumina). The BeadChips were scanned according to the protocol described in the Illumina Whole Genome Gene Expression for BeadStation Manual v3.2, Revision A using scanning software BeadScan 3.5.31. The GenomeStudio® Data Analysis Software (Illumina) was used for data collection. Missing values were imputed by using the k-nearest-neighbor algorithm (k=15). The raw scan and imputed data were read using the lumi available through Bioconductor. This was carried out using the R statistical package (version 2.10.1; R Foundation for Statistical Computing, Vienna, Austria). Quality control checks of the raw non-normalized data included visualization of the similarity matrix and hierarchical clustering analysis. Samples for one placebo-treated individual did not pass our quality control checks, and thus were removed from subsequent normalization and analysis. Total RNA isolated from a collection of human tissue-derived cell lines was used as an internal control; this sample also was omitted from subsequent data normalization. Variance stabilizing normalization (Biconductor library vsn) was applied to all other samples. All subsequent analyses were performed on vsn-transformed intensity values.

overall_design: This study investigated the genome-wide transcriptional response to endotoxin-induced TNF-alpha-mediated signaling in whole-blood leukocytes. Total RNA was isolated from PAXgene-treated whole blood of 16 healthy volunteers before and 4 hours after infusion of 1ng/kg E. coli lipopolysaccharide. Eight of these subjects were treated with the TNF-alpha inhibitor, etanercept.

first: Tom

last: van der Poll


title: Mus musculus gene expression profiling for Wt, SKM-Tg, C and SKM-KO in Affymetrix 430PM strip arrays

status: Public on Feb 13 2014

city: Barcelona

first: Oscar

last: Reina Garcia

address: C/Baldiri Reixac 10

department: Biostatistics and Bioinformatics

country: Spain

institute: IRB Barcelona

summary:
We analyzed the functional role of DOR (Diabetes and Obesity Regulated gene) (also named Tp53inp2) in skeletal muscle. We show that DOR has a direct impact on skeletal muscle mass in vivo. Thus, using different transgenic mouse models, we demonstrate that while muscle-specific DOR gain-of-function results in reduced muscle mass, loss-of-function causes muscle hypertrophy. DOR has been described as a protein with two different functions, i.e., a nuclear coactivator and an autophagy regulator (Baumgartner et. al., PLoS One, 2007; Francis et. al., Curr Biol, 2010; Mauvezin et. al., EMBO Rep, 2010; Nowak et. al., Mol Biol Cell, 2009). This is why we decided to analyze which of these two functions could explain the phenotype observed in our mice models. In this regard, we performed a transcriptomic analysis using microarrays looking for genes differentially expressed in the quadriceps muscle of WT and SKM-Tg mice as well as in C and SKM-KO animals. Surprisingly, only a reduced number of genes were dysregulated upon DOR manipulation and most of the genes underwent mild changes in expression. These data strongly suggest that DOR does not operate as a nuclear co-factor in mouse skeletal muscle under the conditions subjected to study. In contrast, DOR enhances basal autophagy in skeletal muscle and promotes muscle wasting when autophagy is a contributor to muscle loss.
To determine the functional role of DOR in skeletal muscle, we generated transgenic mice (SKM-Tg) overexpressing DOR specifically in skeletal muscle under the Myosin-Light Chain 1 promoter/enhancer. The open reading frame of DOR was introduced in an EcoRI site in the MDAF2 vector, which contains a 1.5 kb fragment of the MLC1 promoter and 0.9 kb fragment of the MLC1/3 gene containing a 3' muscle enhancer element (Rosenthal et. al., PNAS, 1989; Otaegui et. al., FASEB J, 2003). The fragment obtained after the digestion of this construct with BssHII was the one used to generate both transgenic mouse lines. Nontransgenic littermates were used as controls for the transgenic animals (Wt). In addition, a muscle-specific DOR knock-out mouse line (SKM-KO) was also generated by crossing homozygous DOR loxP/loxP mice with a mouse strain expressing Cre recombinase under the control of the Myosin-Light Chain 1 promoter (Bothe et. al., Genesis, 2000). Deletion of exons 3 and 4 driven by Cre recombinase caused the ablation of DOR expression. Non-expressing Cre DOR loxP/loxP littermates were used as controls for knockout animals (C). Four-month-old male mice were used in all experiments. Mice were in a C57BL/6J pure genetic background.

overall_design:
We used microarrays to analyze the effect of DOR gain-of-function and DOR ablation on skeletal muscle gene expression
Total RNA from quadriceps muscles from 4-month-old male mice was extracted and used for hibridization on Affimetrix microarrays

first: David

last: Sala

first: Antonio

last: Zorzano


title: LRH-1 governs vital transcriptional programs in endocrine sensitive and resistant breast cancer cells: Expression profiling

city: Sherbrooke

first: Nicolas

last: Gévry

state: Québec

address: 2500 boulevard de l'université

institute: Université de Sherbrooke

summary: Tumor characteristics are decisive in the determination of treatment strategy for breast cancer patients. Patients with estrogen receptor-α (ERα) positive breast cancer can benefit from long-term hormonal treatment. Nonetheless, the majority of patients will develop resistance to these therapies. Here, we investigated the role of the liver receptor homolog-1 (LRH-1, NR5A2) in anti-estrogen (AE) sensitive and resistant breast cancer cells. We identified genome-wide LRH-1 binding sites using ChIP-seq, uncovering preferential binding to regions distal to transcriptional start sites (TSS). We further characterized these LRH-1 binding sites by integrating overlapping layers of specific chromatin marks, revealing that many LRH-1 binding sites are active and could be involved in long-range enhancer-promoter looping. Combined with transcriptome analysis of LRH-1 depleted cells, these results show that LRH-1 regulates specific subsets of genes involved in cell proliferation in AE-sensitive and AE-resistant breast cancer cells. Furthermore, the LRH-1 transcriptional program is highly associated with signature of poor outcome breast cancer tumors in vivo. Herein report the genome-wide location and molecular function of LRH-1 in breast cancer cells and reveal its therapeutic potential for the treatment of breast cancers, notably for tumors resistant to treatments currently used in therapies.

overall_design: Total RNA was obtained from two biological replicates of breast cancer cells MCF7, LCC2 and LCC9 transduced with LRH-1 shRNA (shLRH-1) or control shRNA (shCTL) lentiviruses.

first: Stephanie

last: Bianco

last: Gevry


title: Gene expression profiles among hiPSCs, hiPSC-EpSCs and hEpSCs

status: Public on Feb 11 2014

first: Xiaowei

last: Xu

address: 3400 Spruce st

institute: University of Pennsylvania

summary: Phenotypically comparing the hiPSC-EpSCs with hiPSCs and hEpSCs.

overall_design: There are two or three biologic replicates.

first: Ruifeng

last: Yang

first: Zhi

last: Wei


title: Transcriptional response to TNF-alpha in a panel of 17 human melanoma cell lines

status: Public on Feb 10 2014

city: Bonn

first: Michael

last: Holzel

address: Sigmund-Freud Str. 25

department: Institute of Clinical Chemistry and Clinical Pharmacology

institute: University Hospital Bonn

summary:
A panel of 17 human melanoma cell lines with known BRAF and NRAS mutation status was stimulated with TNF-alpha for 72 hours.
The goal of the study was to correlate the transcriptional response in BRAF versus NRAS mutated melanoma cell lines.

overall_design:
Total RNA was obtained from a panel of 17 human melanoma cell lines treated for 72 hours with TNF-alpha or left untreated.
Gene expression profiling was done using the Illumina Human HT12 v4 platform.

first: Tobias

last: Bald

first: Judith

last: Kohlmeyer

first: Jennifer

last: Landsberg

first: Meri

last: Rogava

first: Stefanie

last: Riesenberg

first: Dirk

last: Schadendorf

last: Hölzel

first: Thomas

last: Tüting


title: Chronic exposure to fluticasone propionate shapes gene expression and modulates innate and adaptive activation pathways in human macrophages (LPS)

city: Guildford

first: Fernando

middle: O

last: Martinez Estrada

state: Surrey

address: Stag Hill

department: Faculty of Health & Medical Sciences

country: United Kingdom

institute: University of Surrey

summary: Here we investigate how glucocorticoids affect the response to LPS in human macrophages.

overall_design: Total RNA obtained from monocyte-derived macrophages exposed to lipopolysacharide presceding exposure to fluticasone propionate or left untreated.

first: Martijn

last: van de Garde

last: Martinez

first: Rene

middle: E

last: Jonkers

first: Jörg

last: Hamann


title: Chronic exposure to fluticasone propionate shapes gene expression and modulates innate and adaptive activation pathways in human macrophages (INF)

summary: Here we investigate how glucocorticoids affect the response to Interferon gamma in human macrophages.

overall_design: Total RNA obtained from monocyte-derived macrophages exposed to Interferon gamma presceding exposure to fluticasone propionate or left untreated.

middle: D


title: Chronic exposure to fluticasone propionate shapes gene expression and modulates innate and adaptive activation pathways in human macrophages

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

overall_design: Refer to individual Series


title: Strigolactone analogs induce apoptosis through activation of p38 and the stress response pathway in cancer cell lines and in conditionally reprogrammed primary prostate cancer cells

city: Washington

first: Ronit

middle: Iris

last: Yarden

state: DC

address: 3900 Reservoir Rd., NW

department: Human Science

institute: Georgetown University

summary: Strigolactones are a novel class of plant hormones produced in roots and regulate shoot and root development. We have previously shown that synthetic strigolactone analogues potently inhibit growth of breast cancer cells and breast cancer stem cells. Here we show that strigolactone analogues inhibit the growth and survival of an array of cancer-derived cell lines representing solid and non-solid cancer cells including: prostate, colon, lung, melanoma, osteosarcoma and leukemic cell lines, while normal cells were minimally affected. Furthermore, we tested the response of patient-matched conditionally reprogrammed normal and prostate cancer cells. The tumor cells exhibited significantly higher sensitivity to the two most potent SL analogues with increased apoptosis compared to their normal counterpart cells.  Treatment of cancer cells with strigolactone analogues was hallmarked by increased expression and activity of genes involved in stress signaling, cell cycle arrest and apoptosis. All five strigolactone analogues induced G2/M cell cycle arrest, accompanied with a decrease in the expression level of cyclin B1. Apoptosis was marked by   increased percentages of cells in the sub-G1 fraction and was confirmed by Annexin V staining. In conditionally reprogramed matched tumor and normal prostate cells, the cleavage of PARP1 confirmed the specific increase in apoptosis of tumor cells. In summary, Strigolactone analogues are promising candidates for anticancer therapy by their ability to specifically induce cell cycle arrest, cellular stress and apoptosis in tumor cells with minimal effects on growth and survival of normal cells.

overall_design: There are duplicate samples for each condition. U2OS cells were treated with 2 different strigolactone analogues: ST362 or MEB55 at the concentration of 5 ppm for either 6 hr or 24 hr.Control samples were those treated with vehicle only .

first: Claire

last: Pollock

first: Sara

last: McDoungh

first: Victor

last: Wang

first: Hyojung

last: Lee

first: Lymor

last: Ringer

first: R

middle: J

first: Adam

last: Feldman

first: Cristina

last: Prandi

first: Hinanit

last: Koltai

first: Yoram

last: Kapulnik

first: Olga

last: Rudrigaz

first: Richard

last: Schlegel

first: Christopher

last: Albanese

middle: I


title: The identification of downstream gene of p63, STAT3 and PPAR gamma

city: Houston

first: Woonyoung

last: Choi

address: 1515 Holcombe

institute: Md Anderson Cancer center

summary: To identify the downstream gene of p63, STAT3 and PPARg signling

overall_design: Total RNA from p63 knock down UC14 and STAT3 knockdown Scaber cells were compared to control cells. Also Total RNA from Rosiglitazone treated UC7 cells were compared to untreated control.

first: I-ling

first: Tiewei

last: Cheng

first: David

last: McConkey


title: Effect on peripheral blood gene expression profile of lenalidomide plus rituximab in patients with follicular lymphoma

status: Public on Feb 06 2014

first: Min

last: Zhang

address: 7455 Fannin Street

institute: MD Anderson Cancer Center

summary: Peripheral blood was collected into PAXgene tubes from 7 patients with follicular lymphoma, in a phase 2 clinical trial of lenalidomide plus rituximab as initial therapy for advanced-stage indolent non-Hodgkin's lymphoma. Blood was collected at baseline and on Day 15 of the second 28-day cycle, for determination of the gene expression profile after globin mRNA reduction.

overall_design: The effect of treatment on gene expression profiles was largely based on the paired t test, using the Significance Analysis of Microarrays method.

first: Nathan

last: Fowler

first: Zhiqiang

first: Sattva

last: Neelapu

last: Davis


title: The Functional Consequences of Variation in Transcription Factor Binding

status: Public on Feb 05 2014

city: Tucson

first: Darren

middle: A.

last: Cusanovich

state: AZ

address: 1230 N Cherry Avenue, BSRL 421

department: Cellular and Molecular Medicine

institute: University of Arizona

summary: One goal of human genetics is to understand how the information for precise and dynamic gene expression programs is encoded in the genome. The interactions of transcription factors (TFs) with DNA regulatory elements clearly play an important role in determining gene expression outputs, yet the regulatory logic underlying functional transcription factor binding is poorly understood. Many studies have focused on characterizing the genomic locations of TF binding, yet it is unclear to what extent TF binding at any specific locus has functional consequences with respect to gene expression output. To evaluate the context of functional TF binding we knocked down 59 TFs and chromatin modifiers in one HapMap lymphoblastoid cell line. We then identified genes whose expression was affected by the knockdowns. We intersected the gene expression data with transcription factor binding data (based on ChIP-seq and DNase-seq) within 10 kb of the transcription start sites of expressed genes. This combination of data allowed us to infer functional TF binding. Using this approach, we found that only a small subset of genes bound by a factor were differentially expressed following the knockdown of that factor, suggesting that most interactions between TF and chromatin do not result in measurable changes in gene expression levels of putative target genes. We found that functional TF binding is enriched in regulatory elements that harbor a large number of TF binding sites, at sites with predicted higher binding affinity, and at sites that are enriched in genomic regions annotated as “active enhancers”.

overall_design: 201 samples were analyzed - 57 transcription factors were knocked down in triplicate in the same cell line, 2 factors were knocked down in 6 replicates each and an additional 18 control knockdowns (siRNA not targeting any known gene) were analyzed

middle: A

first: Bryan

last: Pavlovic

first: Jonathan

middle: K

last: Pritchard

first: Yoav

last: Gilad


title: Interferon Lambda Alleles Predict Innate Antiviral Immune Responses and Hepatitis C Virus Permissiveness.

status: Public on Feb 12 2014

city: Chapel Hill

first: Timothy

last: Sheahan

state: North Carolina

address: 3113 MHRC 135 Dauer Drive, Campus Box 7435

department: Epidemiology

institute: University of North Carolina at Chapel Hill

summary: Hepatitis C virus (HCV) infection can result in viral chronicity or clearance. Although host genetics and particularly genetic variation in the interferon lambda (IFNL) locus are associated with spontaneous HCV clearance and treatment success, the mechanisms guiding these clinical outcomes remain unknown. Using a laser capture microdissection-driven unbiased systems virology approach, we isolated and transcriptionally profiled HCV-infected and adjacent primary human hepatocytes (PHH) approaching single cell resolution. An innate antiviral immune signature dominated the transcriptional response, but differed in magnitude and diversity between HCV-infected and adjacent cells. Molecular signatures associated with more effective antiviral control were determined by comparing donors with high and low infection frequencies. Cells from donors with clinically unfavorable IFNL genotypes were infected at a greater frequency and exhibited dampened antiviral and cell death responses. These data suggest that early virus-host interactions, particularly host genetics and induction of innate immunity, critically determine the outcome of HCV infection.

overall_design: Cell populations of primary human hepatocytes were collected via laser capture microdissection, their transcriptomes were amplified and analyzed via whole Illumina genome microarray. Three populations were collected: virus infected cells, cells adjacent to infected cells (“adjacent”) and mock infected cells. Cells from multiple donors were employed to address host genetic variability on our observed phenotypes. Nine different donors were assessed on 1-day post infection and three donors were assessed 3 days post infection and and four donors were assessed on 7 days post infection. For each cell population (virus infected, adjacent or mock), four biological replicate arrays were performed. In total, we analyzed 186 microarrays

first: Naoko

last: Imanaka

first: Svetlana

last: Marukian

first: Marcus

last: Dorner

first: Peng

last: Liu

first: Alexander

last: Ploss

first: Charles

last: Rice


title: Analysis of global gene expression in fetal astrocytes and pluripotent stem cell derived neural stem cells

status: Public on Feb 02 2014

first: Nasir

last: Malik

address: 50 South Drive/Room 1146

institute: NIH

summary: Gene expression was compared between two sets of commercially obtained fetal astrocytes and two sets of neuronal stem cell lines derived from iPSC and ES cells.  Pathways and genes critical for astrocyte development and maintenance were identified in the analysis.

overall_design: Total RNA obtained from fetal astrocytes from two sources compared to total RNA obtained from two neuronal stem cell lines.

first: Malik

last: Nasir

first: Rao

last: Mahendra


title: Whole blood transcriptome of survivors and nonsurvivors of sepsis

status: Public on Feb 04 2014

city: Kingswood

first: Grant

last: Parnell

state: NSW

address: Nepean Hospital

department: Nepean Clinical School

country: Australia

institute: University of Sydney

summary: There is currently no reliable tool available to measure immune dysfunction in septic patients in the clinical setting. This proof-of-concept study assesses the potential of gene expression profiling of whole blood as a tool to monitor immune dysfunction in critically ill septic patients. Whole blood samples were collected daily for up to 5 days from patients admitted to the intensive care unit with sepsis. RNA isolated from whole blood samples was assayed on Illumina HT-12 gene expression microarrays consisting of 48,804 probes. Microarray analysis identified 3677 genes as differentially expressed across 5 days between septic patients and healthy controls. Of the 3677 genes, biological pathway analysis identified 86 genes significantly down-regulated in the sepsis patients were present in pathways relating to immune response. These 86 genes correspond to known immune pathways implicated in sepsis including lymphocyte depletion, reduced T lymphocyte activation and deficient antigen presentation. Furthermore, expression levels of these genes correlated with clinical severity, with a significantly greater degree of down-regulation found in non-survivors compared to survivors. The results show that whole blood gene-expression analysis can capture systemic immune dysfunctions in septic patients. Our study provides an experimental basis to support further study on the use of a gene expression based assay, to assess immunosuppression and guide immunotherapy in future clinical trials.

overall_design: Daily PAXgene samples for up to 5 days for sepsis survivors (n=26), sepsis nonsurvivors (n=9), and healthy controls (n=18).

first: Benjamin

middle: M

last: Tang


title: PSN1 Polysome and INK128

status: Public on Feb 03 2014

city: New Haven

first: Thomas

last: Hayman

address: 15 York Street

department: Department of Therapeutic Radiology

institute: Yale University

summary: Polysome-bound mRNA was analyzed to determine the effects of INK128 and radiation on mRNA translation

overall_design: 4 samples were analyzed with 3 biological replicates

first: Heekyong

last: Bae

first: Tamalee

last: Kramp

first: Muhammad

last: Jamal

first: Uma

middle: T

last: Shankavaram

first: Kevin

last: Camphausen

first: Philip

last: Tofilon


title: Gene expression profiles of wild type and Actinin 4 point mutant knock-in adult podocyte cells at 2, 5 and 44 weeks following TRAP purification. (GUDMAP Series ID: 49)

status: Public on Feb 01 2014

city: Edinburgh

first: GUDMAP

last: Developers

address: Crewe Road

institute: IGMM MRC Human Genetics Unit

summary:
The long term goal is to create an encyclopaedia of podocyte gene expression both in homeostasis, development and disease.  Podocytes are identified by their expression of an eGFPL10a fusion protein whose expression is driven by a Col1a1 promoter fragment.  Podocyte-specific RNA is generated by eGFP-affinity purification of polysomes and then subject to microarray analysis.
Expression profiling of wild type and  Actinin 4 point mutant knock-in kidney podocytes by TRAP.

overall_design: Col1a1-eGFPL10a mice were crossed against Actn4+/tm1.1Plk (KI) mice.  Wild-type or KI mice were subject to TRAP at the following time points: 2, 5 and 44 weeks.

first: Ben

last: Humphreys


title: Genome-wide analysis of gene expression changes in human alveolar epithelial cells grown on collagen- or matrigel-coated dishes

city: Raleigh

first: Helena

last: Morales Johansson

address: 1060 William Moore Drive

department: Molecular Biomedical Sciences

institute: NCSU

summary: Analysis of human alveolar epithelial cell signatures at gene expression level. The aim of the study was to identify candidate genes that are induced in human alveolar epithelial type II cells on collagen-coated dishes. Results provide important information about changes hAT2 cells undergo in the transformation to AT1-like cells.

overall_design: Total RNA obtained from human alveolar epithelial cells grown on collagen- or matrigel-coated dishes for 12, 24, 48 and 72 hr.

last: Sannes

first: Donna

middle: R

last: Newman


title: Lambda light chain knockdown in ALMC1 human myeloma cells

status: Public on Jan 30 2014

first: Lakshmanan

last: Iyer

address: 136 Harrison Ave

department: Neuroscience

institute: Tufts University

summary: We used siRNA to knockdown lambda light chain expression in ALMC1 cells that express an intact IgG lambda monoclonal protein.

overall_design: We compared knockdown cells to cells treated with control siRNA.

first: Ping

last: Zhou

first: Xun

last: Ma

first: Raymond

last: Comenzo


title: D. simulans with variable Y chromosomes

first: Alan

last: Branco

address: 665 Huntington Ave.

department: Environmental Health

institute: Harvard School of Public Health

summary: Y chromosome from different Drosophila simulans were introgressed into the same genetic background. Strains showing distinct sex-ratio distortion were clustered according to the ratio of males and females observed in the progeny. Strains showing disproportionally high number of female flies in the progeny were contrasted with strains displaying the proportion of male and females flies close to 1:1. In addition, interspecific Y chromosome from Drosophila sechellia (Ysec) was compared with D. simulans Y chromosomes. Total RNA was extracted from whole flies using TRIzol (Life Technologies). The synthesis of cDNA and its labeling with fluorescent dyes (Cy3 and Cy5) as well as hybridization reactions were carried out using 3DNA protocols and reagents (Genisphere). A minimum of 2 biological replicas was used, however this number varied according to the hybridization design (See 'Branco et. al, Natural variation of the Y chromosome suppresses sex ratio distortion and modulates testis-specific gene expression in Drosophila simulans' for details of the hybridization design). Microarrays were ~18,000-feature cDNA arrays spotted with D. melanogaster cDNA PCR products. We have recently annotated these probes using the D. simulans genome. Possible biases introduced by using a D. melanogaster platform for a D. simulans genome are minimized because the probed genomes are essentially identical except for the Y chromosomes. Slides were scanned using Axon 400B scanner (Axon Instruments) and GenePix Pro 6.0 software. Foreground fluorescence of dye intensities was normalized by the Loess method in Bioconductor / Limma.

overall_design: Dye "swaps," loop design.

first: Bernardo

last: Lemos


title: Involvement of endocytosis and alternative splicing in the formation of the pathological process in Parkinson's Disease

status: Public on Jan 31 2014

city: Moscow

first: Anelya

last: Alieva

address: 2 Kurchatova sq.

country: Russia

institute: IMG RAS

summary: We performed a whole-transcriptome analysis of the peripheral blood of untreated patients with stage 1 PD (Hoehn–Yahr scale).

overall_design: 10 samples were investigated

first: Maria

last: Shadrina

first: Elena

middle: V

last: Filatova

first: Alexey

last: Karabanov

first: Sergey

last: Illarioshkin

first: Svetlana

last: Limborska

first: Petr

last: Slominsky


title: Expression profiling of migrated and invaded breast cancer cells predicts early metastatic relapse and reveals Krüppel-like factor 9 as a potential suppressor of invasive growth in breast cancer

status: Public on Jan 29 2014

city: Edegem

first: Ken

last: Op de Beeck

address: Universiteitsplein 1

country: Belgium

institute: University Of Antwerp

summary: Cell motility and invasion initiate metastasis. However, only a subpopulation of cancer cells within a tumor will ultimately become invasive. Due to this stochastic and transient nature, in an experimental setting, migrating and invading cells need to be isolated from the general population in order to study the gene expression profiles linked to these processes. This report describes microarray analysis on RNA derived from migrated or invaded subpopulations of triple negative breast cancer cells in a Transwell set-up, at two different time points during motility and invasion, pre-determined as “early” and “late” in real-time kinetic assessments. Invasion- and migration-related gene expression signatures were generated through comparison with non-invasive cells, remaining at the upper side of the Transwell membranes. Late-phase signatures of both invasion and migration indicated poor prognosis in a series of breast cancer data sets. Furthermore, evaluation of the genes constituting the prognostic invasion-related gene signature revealed Krüppel-like factor 9 (KLF9) as a putative suppressor of invasive growth in breast cancer. Next to loss in invasive vs non-invasive cell lines, KLF9 also showed significantly lower expression levels in the “early” invasive cell population, in several public expression data sets and in clinical breast cancer samples when compared to normal tissue. Overexpression of EGFP-KLF9 fusion protein significantly altered morphology and blocked invasion and growth of MDA-MB-231 cells in vitro. In addition, KLF9 expression correlated inversely with mitotic activity in clinical samples, indicating anti-proliferative effects.

overall_design: After selection of two time points (“early” and “late”), RNA from invasive and migratory MDA-MB-231 cells was isolated from Transwell membranes and hybridized onto an Illumina HumanHT-12 v4 Expression beadchip. Gene expressions of migrated/invaded subpopulations vs non-motile cells were compared.

first: Ridha

last: Limame

first: Steven

last: Van Laere

first: Lieselot

last: Croes

first: Annemieke

last: De Wilde

first: Luc

last: Dirix

first: Guy

last: Van Camp

first: Marc

last: Peeters

first: Olivier

last: De Wever

first: Filip

last: Lardon

first: Patrick

last: Pauwels


title: Raptor/mTORC1-dependent metabolic programming of exit from quiescence underlies T cell activation and fate decisions

status: Public on Jan 27 2014

city: Memphis

first: Geoffrey

last: Neale

state: TN

address: 262 Danny Thomas Place

department: Hartwell Center

institute: St Jude Childrens Research Hospital

summary: Naïve T cells respond to antigen stimulation by exiting from quiescence into clonal expansion and functional differentiation, but the control mechanism is elusive. Here we describe that Raptor/mTORC1-dependent metabolic reprogramming is a central determinant of this transitional process. Loss of Raptor abrogates T cell priming and Th2 cell differentiation, although Raptor function is less important for continuous proliferation of actively cycling cells. mTORC1 coordinates multiple metabolic programs in T cells including glycolysis, lipid synthesis and oxidative phosphorylation to mediate antigen-triggered exit from quiescence. mTORC1 further links glucose metabolism to the initiation of Th2 differentiation by orchestrating cytokine receptor expression and cytokine responsiveness. Activation of Raptor/mTORC1 integrates T cell receptor (TCR) and CD28 co-stimulatory signals in antigen-stimulated T cells. Our studies identify a Raptor/mTORC1-dependent pathway linking signal-dependent metabolic reprogramming to quiescence exit, and this in turn coordinates lymphocyte activation and fate decisions in adaptive immunity.

overall_design: We used microarrays to explore the gene expression profiles differentially expressed in CD4+ T-cells from wild-type (WT) and CD4(cre) x Raptor(fl/fl) mice before and after stimulation with anti CD3/CD28 antibodies.

first: Kai

last: Yang

first: Hongbo

last: Chi


title: miR-638 promotes melanoma metastasis and protects melanoma cells from apoptosis and autophagy (mRNA)

city: Leipzig

first: Animesh

last: Bhattacharya

state: Saxony

address: Johannisallee 30, MBFZ, Room no B-214

department: Dermatology

institute: University of Leipzig, Medical Faculty

summary: MicroRNA (miRNA) expression profiling identified miR-638 as one of the most significantly overexpressed miRNAs in metastatic lesions compared with primary melanomas. miR-638 enhanced the tumourigenic properties of melanoma cells in vitro and lung colonization in vivo. mRNA expression profiling of miR-638 and antagomir-transduced cells identified new candidate genes as miR-638 targets, the majority of which is involved in p53-mediated apoptosis regulation. miR-638 depletion stimulated expression of p53 and its downstream target genes and induced apoptosis and autophagy in melanoma cells. miR-638 promoter analysis revealed transcription factor associated protein 2-α (TFAP2A) as a direct negative regulator of miR-638. Further analyses provided strong evidence for a double negative regulatory feedback loop between miR-638 and TFAP2A. Taken together, miR-638 may support melanoma progression by suppressing p53-mediated apoptosis pathways and by targeting the transcriptional repressor TFAP2A.

overall_design: Whole genome cDNA microarray (Illumina Human HT-12 v4 Expression BeadChip Kit, San Diego, CA 92122 USA) analyses were performed in duplicates using RNA extracted from SK-Mel-147 cells transfected with a non-targeting control, miR-638 or antagomiR-638.

first: Ulf

last: Schmitz

first: Yvonne

last: Raatz

first: Madeleine

last: Schönherr

first: Tina

last: Kottek

first: Marianne

last: Schauer

first: Sandra

last: Franz

first: Anja

last: Saalbach

last: Anderegg

first: Olaf

last: Wolkenhauer

first: Dirk

last: Schadendorf

first: Jan

middle: C

last: Simon

first: Thomas

last: Magin

first: Julio

last: Vera

first: Manfred

last: Kunz


title: miR-638 promotes melanoma metastasis and protects melanoma cells from apoptosis and autophagy

type:
Expression profiling by array
Other

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

overall_design: Refer to individual Series


title: Genome wide analysis of Visceral adipose tissue CD14+ cells from Obese and obese diabetic subjects

status: Public on Jan 24 2014

city: Paris

first: Michele

last: Guerre-Millo

address: 91 Boulevard de l'Hopital

country: France

institute: INSERM

summary:
Identification of the inflammatory signature in visceral adipose tissue CD14+ cells (adipose tissue macrophage)
Total RNA obtained from CD14+ cells (Immunoselcted cells from stromal adipose tissue cells)

overall_design: Adipocytes and cells of the stroma vascular fraction (SVF) were obtained by collagenase digestion of adipose tissue. SVF cells were resuspended in endotoxin-free PBS supplemented with 2% FCS and 1 mM EDTA. Isolation of CD14+ using positive selection magnetic beads (Stemcell technologies) and CD4+ cells was performed using positive selection magnetic beads (Stemcell Technologies, Vancouver, Canada). An Illumina (San Diego, CA) RNA amplification kit (NuGEN, BiotinIL Module) was used according to the manufacturer's instructions to obtain biotinlabeled cDNA from 50 ng of total RNA extracted from adipose tissue CD14+ cells.

first: Michèle

first: Nicolas

last: Venteclef

first: Patricia

last: Ancel


title: A Pilot Study on the Potential of RNA-Associated to Urinary Vesicles as a Suitable Non-Invasive Source for Diagnostic Purposes in Bladder Cancer

city: Derio

first: Ana

middle: Maria

last: Aransay

state: BIZKAIA

address: Parque tecnologico de Bizkaia, Building 801-A

department: Genome Analysis Platform

institute: CIC bioGUNE

summary: Abstract: Bladder cancer is one of the most common cancers and, together with prostate carcinoma, accounts for the majority of the malignancies of the genitourinary tract. Since prognosis ameliorates with early detection, it will be beneficial to have a repertoire of diagnostic markers that could complement the current diagnosis protocols. Recently, cell-secreted extracellular vesicles have received great interest as a source of low invasive disease biomarkers because they are found in many body fluids, including urine. The current work describes a pilot study to generate an array-based catalogue of mRNA associated to urinary vesicles, and also a comparison with samples obtained from bladder cancer patients. After an analysis of presence/absence of transcripts in bladder cancer EVs, a list of genes was selected for further validation using PCR technique. We found four genes differentially expressed in cancer samples. LASS2 and GALNT1 were present in cancer patients, while ARHGEF39 and FOXO3 were found only in non-cancer urinary vesicles. Previous studies have pointed to the involvement of those genes in tumour progression and metastasis.

overall_design: Total RNA isolated from Extracellular Vesicles obtained from urine of bladder cancer patients was compared with RNA isolated from urinary vesicles of non-cancer patients.

first: A

last: Perez

last: Loizaga

first: R

last: Arceo

first: I

last: Lacasa

last: Rabade

first: K

last: Zorroza

first: D

last: Mosen-Ansorena

first: E

last: Gonzalez

first: J

last: Falcon-Perez

first: M

last: Unda-Urzaiz

first: F

last: Royo


title: Expression data of CHK1 shRNA knockdown and control shRNA in U2OS cells

first: Chun-Jen

middle: Curtis

last: Lin

address: 7345 Fannin Street

department: Systems Biology

institute: University of Texas MD Anderson Cancer Center

summary: Homologous recombination-mediated DNA repair deficiency (HRD) predisposes to cancer development, but also provides therapeutic opportunities. Here, we identified an HRD gene signature that robustly predicted HRD status. Unexpectedly, concurrent loss of PTEN in BRCA1-deficient cells might extensively rewire the HR repair network and confer resistance to PARP inhibitor, partially through over-expression of TTK. We used the HRD gene signature as a drug discovery tool and found several PARP-inhibitor-synergizing agents through the connectivity map. Thus gene expression profiling can be used to define the functional status of the HR repair network providing prognostic and therapeutic information.

overall_design:
Various shRNAs that target genes involved in homologous recombination (HR) were transfected in MCF-10A non-transformed breast cells lines. Stable HR gene knockdown MCF-10A cells were seeded 200000 at 10 cm plate. Cells were harvested after 48 hours culturing and used for gene expression profiling.

The shRNA that target CHK1 gene was transfected in human osteosarcoma U2OS cell line by lentiviral particles and selected stable CHK1 knockdown U2OS cells. Scrambled control shRNA-transfected U2OS cells were applying as control.  Both stable CHK1 knockdown and control U2OS cells were seeded with 2 x 10^5 cells at 10 cm culture plate. Cells were cultured in McCOY 5A medium with 10% FBS and harvested after 48 hours culturing.  mRNA was extracted from collected cells and performing gene expression profiling. Three biological replicates were applied.

first: Guang

last: Peng

first: Curtis

first: Wei

last: Mo

first: Hui

last: Dai

first: Yun-Yong

last: Park

first: Soo-Mi

last: Kim

first: Qianxing

first: Yang

first: Stefan

last: Siwko

first: Ruozhen

last: Hu

first: Ju-Seog

last: Lee

first: Bryan

last: Hennessy

first: Samir

last: Hanash

first: Gordon

middle: B

last: Mills

first: Shiaw-Yih


title: Akt inhibitor MK2206 prevents influenza A(H1N1)pdm09 virus infection in vitro

status: Public on Jan 23 2014

city: Helsinki

first: Denis

last: Kainov

address: Tukholmankatu 8

department: Institute for Molecular Medicine Finland

institute: Helsinki University

summary: The influenza A(H1N1)pdm09 virus caused a global flu pandemic in 2009 and contributes to seasonal epidemics. Different treatment and prevention options for influenza have been developed and applied with limited success. Here we report that an Akt inhibitor MK2206 possesses potent antiviral activity against influenza A(H1N1)pdm09 virus in vitro. We showed that MK2206 blocks the entry of different A(H1N1)pdm09 strains into cells. Moreover, MK2206 prevented A(H1N1)pdm09-mediated activation of cellular signaling pathways and the development of cellular immune responses. Importantly, A(H1N1)pdm09 virus was unable to develop resistance to MK2206. Thus, MK2206 is a potent anti-influenza A(H1N1)pdm09 agent.

overall_design: Total RNA obtained from NCI-H1666 cells, which are non-small cell lung cancer cell line. NCI-H1666 cells  were non- or MK2206-treated (10 μM) and mock- or virus-infected (A/Helsinki/p14/2009) at moi of 3.

first: Oxana

last: Denisova

first: Salla

last: Virtanen

first: Carina

last: VonSchantz

first: Dmitrii

last: Bychkov

first: Jens

last: Desloovere

first: Sandra

last: Soderholm

first: Linda

middle: L

last: Theisen

first: Janne

last: Tynell

first: Niina

last: Ikonen

first: Elena

last: Vashchinkina

first: Tuula

last: Nyman

first: Sampsa

last: Matikainen

first: Olli

last: Kallioniemi

first: Ilkka

last: Julkunen

first: Claude

middle: P

last: Muller

first: Xavier

last: Saelens

first: Vladislav

last: Verkhusha


title: Aberrant choline metabolism in epithelial ovarian cancer: relevance of choline kinase activity and expression

city: Milan

first: Loris

last: De Cecco

address: via Venezian 1

institute: IRCSS Istituto Nazionale Tumori

summary: Detection of the abnormal phosphatidylcholine (PtdCho) metabolism in EOC by analysis of magnetic resonance profile (MRS), showed a significant increase of PCho content in EOC cells as compared with non tumoral counterparts associated with activation of choline kinase (ChoK), the enzyme responsible for PCho production following choline phosphorylation in the PtdCho biosynthetic pathway (Kennedy’s pathway). The α-isoform of ChoK has an essential role in growth control and signal transduction and it has been implicated in the carcinogenic process. Aim of the study is the evaluation of the biological relevance of ChoK expression and activity to define its possible role as a non-invasively detectable EOC biomarker and druggable target.   We specifically silenced ChoKα expression by transient RNA interference in two different EOC cell lines and evaluated the biological effects related to metabolic profiles, cell proliferation, cell cycle regulation and alteration of global gene expression to possibly identify new pathways implicated in altered choline metabolism.  Following ChoKα silencing we observed a 70% reduction of mRNA and protein expression with a similar reduction in PCho accumulation as assessed by HMRS analysis. The ChoKα silencing was accompanied by 20% inhibition of cell growth and similar percentage of cells blocked in the G1-phase of cell cycle. No alteration in cell morphology and modulation of the main survival signaling pathways (PI3K and MAPK-related signaling) were observed. By gene expression analysis we found 476 differentially expressed genes (p< 0.01; FDR 25%) in silenced cells. By transcriptome analysis of CHKA silenced cells as compared to controls, among the most relevant co-repressed genes we found CyclinA, related to regulation of cell cycle progression, and cytokines genes (IL-6 and IL-8) related to inflammation and EOC aggressiveness. Acyl-CoA synthetase medium-chain family member 3 (ACSM3), phosphatidic acid phosphatase type 2 (PPAP2A) and Osteoprotegerin were among the most up-regulated genes. All co-modulated genes have been validated by RTqPCR also in independent biological replicates. Ingenuity System Pathways Analysis (IPA) identified a network of molecules most significantly affected by CHKA silencing whose main functions is related to cell morphology cellular assembly and organization, cellular function and maintenance. Also, the cellular functions predicted to be mostly affected by silencing were cellular movement and cell death that are expected to be respectively decreased and increased in silenced cells.

overall_design: a total of 12 samples were analyzed: Two EOC cell line SKOV3 and INTOV11 were transiently silenced with unrelated or CHKA-specific siRNA pool. Three independent experiment were run for each cell line.

first: L

first: M

last: Bagnoli


title: Expression data of ATM, ATR, CHK1, CHK2, and 53BP1 shRNA knockdown and control shRNA in MCF-10A cells

first: Chun-Jen

last: Lin

overall_design:
Various shRNAs that target genes involved in homologous recombination (HR) were transfected in MCF-10A non-transformed breast cells lines. Stable HR gene knockdown MCF-10A cells were seeded 200000 at 10 cm plate. Cells were harvested after 48 hours culturing and used for gene expression profiling.

The shRNAs that target ATM, ATR, CHK1, CHK2, and 53BP1 genes were transfected in MCF-10A non-transformed breast cell line by lentiviral particles and selected stable ATM, ATR, CHK1, CHK2, and 53BP1 knockdown MCF-10A cells. Scrambled control shRNA-transfected and wild type MCF-10A cells were applying as control.  All knockdown and control MCF-10A cells were seeded with 2 x 10^5 cells at 10 cm culture plate. Cells were cultured in MCF-10A medium and harvested after 48 hours culturing.  mRNA was extracted from collected cells and performing gene expression profiling. Three biological replicates were applied.

first: Guang

last: Peng

first: Curtis

first: Wei

last: Mo

first: Hui

last: Dai

first: Yun-Yong

last: Park

first: Soo-Mi

last: Kim

first: Qianxing

first: Yang

first: Stefan

last: Siwko

first: Ruozhen

last: Hu

first: Ju-Seog

last: Lee

first: Bryan

last: Hennessy

first: Samir

last: Hanash

first: Gordon

last: Mills

first: Shiaw-Yih


title: sVEGFR-1 signaling through α5β1 integrin

status: Public on Jan 20 2014

city: Genoa

first: Paolo

last: Uva

address: Via Gerolamo Gaslini, 5

institute: IRCCS Istituto Giannina Gaslini

summary: Soluble VEGFR-1 (sVEGFR-1) acts both as a decoy receptor for VEGFs and as an extracellular matrix protein for α5β1 integrin. A sVEGFR-1-derived peptide that interacts with α5β1 integrin promotes angiogenesis. However, canonical signal downstream integrin activation is not induced, resulting into lack of focal adhesion maturation. We performed a gene expression profile of endothelial cells adhering on sVEGFR-1 compared to that of cells adhering on fibronectin, the principal α5β1 integrin ligand. Three protein kinase-C substrates, adducin, MARCKS, and radixin were differently modulated. Adducin and MARCKS were less phosphorylated whereas radixin was higher phosphorylated in sVEGFR-1 adhering cells, the latter leading to prolonged small GTPase Rac1 activation and induction of a pathway involving the heterotrimeric G protein α13. Altogether, our data indicated endothelial cell acquisition of an highly motile phenotype when adherent on sVEGFR-1. Finally, we indicated radixin as accountable for the angiogenic effect of α5β1 integrin interaction with sVEGFR-1 that in turn depends on an active VEGF-A/VEGFR-2 signaling.

overall_design: Endothelial cells were let adhere in Petri dishes coated with fibronectin or sVEGFR-1 before RNA extraction and hybridization on Affymetrix microarrays. Endothelial cells plated on BSA-treated Petri dishes were used as non-adhesion control. Each hybridization was performed in triplicate.

first: Angela

last: Orecchia

first: Amel

last: Mettouchi

first: Glenn

last: Simon

first: Diego

last: Arcelli

first: Simona

last: Avitabile

first: Gianluca

last: Ragone

first: Guerrino

last: Meneguzzi

first: Karl

last: Pfenninger

first: Giovanna

last: Zambruno

first: Cristina

last: Failla


title: CD40-activation of human B cells

status: Public on Jan 14 2014

city: Cologne

first: Alexander

last: Shimabukuro-Vornhagen

address: Kerpener Str. 62

department: Department I of Internal Medicine

institute: University Hospital of Cologne

summary: Antibody-independent effector functions of B cells, such as antigen presentation and cytokine production, have been shown to play an important role in a variety of immune-mediated conditions such as autoimmune diseases, transplant rejection and graft-versus-host disease.  Therapeutic strategies, which interfere with B cell activation could therefore be a useful addition to the current immunosuppressive armamentarium. CD40 is one of the strongest activation stimuli for B cells. The aim of this study was to characterise the gene expression changes that occurr after B cell activation via CD40.

overall_design: Purified B cells isolated from PBMCs from four donors were compared to B cells which

first: Andreas

last: Draube

first: Alexey

last: Popov


title: Overcoming chemoresistance in B lymphoma cells: elucidating the role of NF-kB in caspase-independent apoptosis by transcriptomic assessment.

status: Public on Jan 19 2014

city: Lund

first: Ida

last: Johansson

address: Department of Oncology Clinical Sciences Lund University BMC C13

department: Oncology

country: Sweden

institute: Lund University

summary: Although therapy responsiveness to therapy in Burkitt lymphoma (BL) is high, relapsed disease and and chemoresistance remain a clinical challenge, and complete mechanisms underlying BL chemoresistance and how it can be circumvented is yet to be fully elucidated. In this study we present data showing that chymotrypsin-like serine proteases inhibitor Nα-tosyl-L-phenylalanine chloromethylketone (TPCK) and specific NF-κB inhibitor Bay-11 7082 can induce caspase-independent apoptosis in chemoresistant BL cells. We also demonstrate that both TPCK and Bay-11 7082-treatment leads to decreased NF-κB nuclear activity and that this is associated with sensitization of chemoresistant Burkitt lymphoma cells. Furthermore we investigated global transcriptional changes induced by Bay-11 7082 and TPCK in the DG-75 and Raji cell lines, respectively, by microarray analysis using Illumina BeadChips. TPCK-treatment of Raji and DG-75 cells resulted in 59 and 21 differently expressed genes, respectively, while Bay-11-treated Raji and DG-75 cells displayed 1403 and 8 differently expressed genes, respectively. Gene Ontology (GO) categorization confirmed enrichment of multiple GOs in Bay 11-treated Raji and DG-75 cells. Fifty percent of the 61 categories in Raji cells were categories sorting under Biological Processes and represented mostly increased gene expression. In DG-75 cells Bay-11 7082 induced significant gene ontology enrichment in only two categories, where the increased/decreased ratio was 1:1. Further unsupervised and supervised bioinformatics processing by Ingenuity Pathway Analysis indicated significant networks in response to TPCK and Bay 11 respectively, including association to NF-κB.  Bay-11 7082 demonstrated deregulated NF-κB related members of receptor mediated cell death signaling, i.e TRAF2 and TRADD, as well as deregulated members of the NF-κB signaling pathway from the cytoplasmic compartment, i.e RELB, in Raji cells. Comparably NF-κB network analysis of Raji- and DG-75 cells treated with Bay-11 7082 and Raji cells treated with TPCK demonstrated deregulation of NF-κB target genes CD69 and IL8. These data indicates that NF-kB may play a role in overcoming chemoresistance in BL cells with defective classical apoptosis signaling.

first: Astrid

last: Ottosson-Wadlund

first: Siriporn

last: Jitkaew

first: Suthat

last: Fucharoen

first: Ingrid

last: Hedenfalk

first: Bengt

last: Fadeel