title: Role of Notch signaling on hematopoietic stem cell differentiation

status: Public on Jun 02 2009

first: Thomas

last: Mercher

address: 1 blackfan circle

department: Hematology

institute: Harvard Medical School

summary:
Although Notch signaling has been clearly implicated in lymphoid differentiation, its role in myeloid lineages differentiation is unclear.
Expression data from hematopoietic stem cells plated on OP9 stroma expressing or not the Notch ligand Delta-like1. Results provide insight into the role of Notch signalling in megakaryocyte fate specification.

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

overall_design: LSK cells were co-cultured for 3 days on OP9, OP9-DL1 and OP9-DL1+CompoundE (gamma-secretase inhibitor) and expression analysis was performed.

first: Gary

middle: D

last: Gilliland


title: PTIP-regulated genes in MEF

status: Public on May 29 2009

city: Bethesda

first: Young-Wook

last: Cho

state: MD

address: 9000 Rockville Pike

department: CEB

country: USA

institute: NIDDK

summary: PPARg and C/EBPa cooperate to control preadipocyte differentiation (adipogenesis). However, the factors that regulate PPARg and C/EBPa expression during adipogenesis remain largely unclear. Here we show PTIP, a protein that associates with histone H3K4 methyltransferases, regulates PPARg and C/EBPa expression in mouse embryonic fibroblasts (MEFs) and during preadipocyte differentiation. PTIP deletion in MEFs leads to marked decreases of PPARg expression and PPARg-stimulated C/EBPα expression. Further, PTIP is essential for induction of PPARg and C/EBPa expression during preadipocyte differentiation. Deletion of PTIP impairs the enrichment of H3K4 trimethylation and RNA polymerase II on PPARg and C/EBPa promoters. Accordingly, PTIP-/- MEFs and preadipocytes all show striking defects in adipogenesis. Furthermore, rescue of the adipogenesis defect in PTIP-/- MEFs requires co-expression of PPARg and C/EBPa. Finally, deletion of PTIP in brown adipose tissue significantly reduces tissue weight in mice. Thus, by regulating PPARg and C/EBPa expression, PTIP plays a critical role in adipogenesis.

overall_design: To identify PTIP-regulated genes, immortalized PTIP conditional knockout PTIPflox/flox MEFs were infected with retroviruses expressing either Cre recombinase or vector alone. We prepared duplicated RNAs from either vector or Cre infected cells (PTIP+/+ or PTIP-/-) for 4 affymetrix microarrays.

first: SunHwa

last: Hong

first: Qihuang

last: Jin

first: Lifeng

last: Wang

first: Ji-Eun

last: Lee

first: Oksana

last: Gavrilova

first: Kai

last: Ge


title: Consequences of GATA1 expression in Gata1- G1ME Murine Megakaryocyte/Erythrocyte Progenitor Cell Line

status: Public on Jun 10 2009

city: Philadelphia

first: Charles

last: Bailey

state: PA

address: Oncology/4 Wood, 1 Children's Center

department: Pediatrics

institute: Children's Hospital of Philadelphia

summary: G1ME cells are GATA1-deficient murine bipotential megakaryocyte/erythrocyte progenitor cells derived from Gata1-negative murine ES cells.  In order to assess the impact of GATA1 on gene regulation and cell differentiation, an expression construct was used to transiently produce high levels of GATA1.  Cells transduced with this construct or a vector control were harvested at 18 and 42 hours, and gene expression was analyzed using Affymetrix MOE430 version 2 arrays.

overall_design: Both vector control and GATA1-expressing cells were isolated by FACS for GFP and 18 and 42 hours.  Biologic triplicates were performed for each construct at each timepoint.

first: Stella

middle: T

last: Chou

first: Eugene

last: Khandros

first: Kim

middle: E

last: Nichols

first: Christopher

last: Vakoc

first: Yu

last: Yao

first: Zan

last: Huang

first: John

last: Crispino

first: Gerd

middle: A

last: Blobel

first: Mitchell

middle: J

last: Weiss


title: Identification of differentiation status of human embryonic stem cell cultures.

status: Public on Jun 01 2009

city: Piscataway

first: Ronald

middle: P.

last: Hart

state: NJ

address: 604 Allison Rd Rm B430

department: Cell Biology & Neuroscience

institute: Rutgers University

summary: Two stages of culture were tested.  ESC is the pluripotent stage expressing SSEA4 and Oct4 but not SSEA1.  NSC is a neural-committed stage expressing Nestin but not beta-tubulin.  Three cell lines were tested: H1, HSF1, and HSF6.

overall_design: One sample was assayed from each cell line at each stage of culture. Clustering reveals primary grouping is by stage.

middle: P


title: Expression data from young and adult mice after over expression of self MHC class l protein in skeletal muscle.

city: London

first: Lucy

middle: R

last: Wedderburn

address: 30 Guilford Street

department: Rheumatology

country: United Kingdom

institute: Institute of Child Health, UCL

summary:
Over expression of MHC Class l protein in skeletal muscle causes myositis. Phenotype after expression in young mice is  more severe.
We performed gene expression profiling on young and adult mice after over expression of self MHC class l protein in skeletal muscle

overall_design: Muscle from young ( early) , adult (Late) and cntrol (control) mice , n=3 each group, was used for gene expression profiling

middle: K

last: Li

first: Halima

last: Moncrieffe


title: Development and Diversification of Retinal Amacrine Interneurons at Single Cell Resolution

city: Boston

first: jeffrey

last: trimarchi

state: MA

address: 77 Avenue Louis Pasteur

department: Genetics

summary: The vertebrate retina uses diverse neuronal cell types arrayed into complex neural circuits to extract, process and relay information from the visual scene to the higher order processing centers of the brain.  Amacrine cells, a diverse class of inhibitory interneurons, are thought to mediate the majority of the processing of the visual signal that occurs within the retina.  Despite morphological characterization, the number of known molecular markers of amacrine cell types is still much smaller than the 26 morphological types that have been identified. Furthermore, it is not known how this diversity arises during development.  Here, we have combined in vivo genetic labeling and single cell genome-wide expression profiling to: 1) Identify specific molecular types of amacrine cells; 2) Demonstrate the molecular diversity of the amacrine cell class.

overall_design: It is difficult to identify new markers of amacrine cells, due to the fact that they only comprise a small percentage of the total cells in the retina.  Additionally, given that there are at least 26 distinct types of amacrine cells, population based approaches fail to achieve the precision necessary to discover markers of each type.  To facilitate the identification of new markers for different amacrine cell classes and to more fully characterize the molecular signatures of these classes, we isolated single amacrine cells.  To accomplish this goal, we introduced genetic reporters (pNdrg4::GFP or pSynapsin::GFP) into the developing retina (P0) by either in vivo or ex vivo electroporation.  These reporters were observed to label morphologically distinct sets of amacrine cells at the different timepoints harvested in this study.  Electroporated retinas were then dissociated at different time points and single retinal amacrine cells were isolated by virtue of their GFP expression and placed in tubes containing lysis buffer.  Then, their mRNAs were reverse transcribed, and the resulting cDNAs were PCR amplified for 35 cycles. Labeled cDNA samples were hybridized to Affymetrix 430 2.0 microarrays and the data was normalized using MAS5.0 software.

first: Timothy

last: Cherry

first: Jeffrey

middle: M

last: Trimarchi

first: Michael

last: Stadler

first: Constance

middle: L

last: Cepko


title: WT and Get1 +/- Bladder Time Course

status: Public on May 20 2009

city: Irvine

first: Amelia

middle: Soto

last: Hopkin

state: CA

address: 250 Sprague Hall

department: Biological Chemistry & Medicine

institute: University of California Irvine

summary: Skin and bladder epithelia form effective permeability barriers through the activation of distinct differentiation gene programs. Employing a genome-wide gene expression study, we identified transcription regulators whose expression correlates highly with that of differentiation markers both in bladder and skin, including the Grainyhead factor Get1/Grhl3, already known to be important for epidermal barrier formation. In the bladder, Get1 is most highly expressed in the differentiated umbrella cells and its mutation in mice leads to a defective bladder epithelial barrier formation due to failure of apical membrane specialization. Genes encoding components of the specialized urothelial membrane, the uroplakins, were downregulated in Get1-/- mice. At least one of these genes, Uroplakin II, is a direct target of Get1. The urothelial-specific activation of the Uroplakin II gene is due to selective binding of Get1 to the Uroplakin II promoter in urothelial cells, most likely regulated by histone modifications. These results demonstrate a key role for Get1 in urothelial differentiation and barrier formation.

overall_design: To gain insights into common and unique transcriptional regulatory programs during bladder differentiation, we profiled global gene expression in whole mouse bladder at E14.5, E16.5, and E18.5.

first: Zhengquan

last: Yu

first: Jaana

last: Mannik

last: Soto

first: Kevin

last: Lin

first: Bogi

last: Andersen


title: Reversal of oncogene transformation and suppression of tumor growth by the novel IGF1R kinase inhibitor A-928605.

status: Public on May 27 2009

city: Abbott Park

first: Paul

last: Jung

state: IL

address: 100 Abbott Park Road

department: Gene Expression Analysis

institute: Abbott Laboratories

summary:
The insulin-like growth factor (IGF) axis is an important signaling pathway in the growth and survival of many cell types and has been implicated in multiple aspects of cancer progression from tumorigenesis to metastasis. The multiple roles of IGF signaling in cancer suggest that selective inhibition of the pathway might yield clinically effective therapeutics. Here we describe A-928605, a novel small molecule inhibitor of the receptor tyrosine kinase responsible for IGF signal transduction. This small molecule is able to abrogate activation of the pathway as shown by effects on the target and downstream effectors and is shown to be effective at inhibiting the proliferation of an oncogene addicted tumor model cell line (CD8-IGF1R 3T3) both in vitro and in vivo.

Keywords: Treatment Response

overall_design: CD8-IGF1R 3T3 cells and 3T3 Vector control treated for 24 h with vehicle or IGF1R inhibitor A-928605, all with 3 replicates.

first: William

middle: N

last: Pappano

first: Jonathan

last: Meulbroek

first: Yi-Chun

last: Wang

first: Robert

last: Hubbard

first: Qian

last: Zhang

first: Meagan

last: Grudzien

first: Niru

middle: B

last: Soni

first: Eric

middle: F

last: Johnson

first: George

middle: S

last: Sheppard

first: Cherrie

last: Donawho

first: Fritz

middle: G

last: Buchanan

first: Steven

last: Davidsen

first: Randy

last: Bell

first: Jieyi


title: Get1 in urothelial differentiation and barrier formation

first: Amelia

last: Hopkin

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

overall_design: Refer to individual Series


title: Cytoprotective Nrf2 pathway is induced in chronically Txnrd1-deficient hepatocytes

status: Public on Jun 03 2009

city: Bozeman

first: Ed

last: Schmidt

state: MT

department: M&I Dept

institute: Montana State University

summary:
Metabolically active cells require robust mechanisms to combat oxidative stress.  The cytoplasmic thioredoxin reductase/thioredoxin (Txnrd1/Txn1) system maintains reduced protein dithiols and provides electrons to some cellular reductases, including peroxiredoxins.

Here we generated mice in which the txnrd1 gene, encoding Txnrd1, was specifically disrupted in all parenchymal hepatocytes.

Keywords: Triplicate comparisons, two conditions.

overall_design: Transcriptome analyses on whole mouse livers.

first: E


title: Identification of Myelin-gene Regulatory Factor as a Critical Transcriptional Regulator Required for CNS Myelination

status: Public on May 31 2009

city: Stanford

first: Ben

last: Emery

address: 299 Campus Dr. W.

department: Neurobiology

institute: Stanford University

summary:
The transcriptional control of CNS myelin gene expression is poorly understood. Here we identify gene model 98, which we have named Myelin-gene Regulatory Factor (MRF), as a transcriptional regulator required for CNS myelination. Within the CNS, MRF is specifically expressed by postmitotic oligodendrocytes. MRF is a nuclear protein containing an evolutionarily conserved DNA binding domain homologous to a yeast transcription factor. Knockdown of MRF in oligodendrocytes by RNA interference prevents expression of most CNS myelin genes; conversely, overexpression of MRF within cultured oligodendrocyte progenitors or the chick spinal cord promotes expression of myelin genes. In mice lacking MRF within the oligodendrocyte lineage, pre-myelinating oligodendrocytes are generated but display severe deficits in myelin gene expression and fail to myelinate. These mice display severe neurological abnormalities, and die due to seizures during the third postnatal week. These findings establish MRF as a critical transcriptional regulator essential for oligodendrocyte maturation and CNS myelination.
We used microarrays to compare cultured oligodendrocytes (differentiated in vitro for 4 days) from MRF conditional knockouts and control litteramates to look at the effects of MRF deficiency on myelin gene expression. Mouse OPCs grown in vitro in the presence of PDGF serve as a baseline for gene expression prior to differentiation.

overall_design: Mouse OPCs from MRF conditional knockout (MRF fl/fl, Olig2 wt/cre) mice and control littermates (MRF wt/fl; Olig2 wt/cre) were isolated from enzymatically dissociated P7 mouse brains as previously described (Cahoy et al., 2008), positively immunopanning for PDGFR-alpha following a depletion of microglia with BSL1. Cells were grown in defined serum-free media as previously described (Dugas et al., 2006), but with the addition of 2% B-27 (Invitrogen). Cells were proliferated for several days in the presence of PDGF-AA (10 ng/ml, PeproTech) and then differentiation induced by withdrawal of PDGF-AA and addition of triiodothyronine (T3) (40 ng/ml; Sigma). RNA was isolated from cells 4 days after induction of differentiation; OPCs maintained in PDGF-AA serve as a baseline of OPC gene expression. Total RNA was isolated from cells with the RNeasy micro kit (Qiagen, Valencia, CA) using Qiagen on-column DNase treatment to remove any contaminating genomic DNA. The integrity of RNA was assessed using an Agilent 2100 Bioanalyzer (Agilent Technologies) and RNA concentration was determined using a NanoDrop ND-1000 spectrophotometer (NanoDrop, Rockland, DE). Biotinylated cRNAs for hybridization to Affymetrix 3'-arrays were prepared from 1ug total RNA using the Affymetrix two-cycle target labeling assay with spike in controls (Affymetrix Inc., Santa Clara, CA, 900494). Labeled-cRNA was fragmented and hybridized to Mouse Genome 430 2.0 Arrays (3'-arrays, Affymetrix, 900495) following the manufacturer's protocols. Raw image files were processed using Affymetrix GCOS 1.3 software to calculate individual probe cell intensity data and generate CEL data files. Using GCOS and the MAS 5.0 algorithm, intensity data was normalized per chip to a target intensity TGT value of 500 and expression data and present/absent calls for individual probe sets calculated. Gene symbols and names for data analyzed with the MAS 5.0 algorithm were from the Affymetrix Netaffx Mouse430_2 annotations file (http://www.affymetrix.com/support/technical/byproduct.affx?product=moe430-20). Quality control was performed by examining raw DAT image files for anomalies, confirming each GeneChip array had a background value less than 100, monitoring that the percencelle present calls was appropriate for the cell type, and inspecting the poly(A) spike in controls, housekeeping genes, and hybridization controls to confirm labeling and hybridization consistency.

first: Jason

middle: C

last: Dugas


title: Expression profiling of Get1 -/-  bladder

overall_design: To gain insights into molecular mechanisms of bladder defects in Get1-/- mice, we profiled bladders of three Get1-/- and three Get +/- E18.5 mice.

first: Zhengquan

last: Yu

first: Jaana

last: Mannik

last: Soto

first: Kevin

last: Lin

first: Bogi

last: Andersen


title: Exon Array expression data from 13 areas of the late second trimester human brain

status: Public on May 18 2009

city: New Haven

first: Matthew

middle: Bonser

state: CT

address: 333 Cedar St., SHM C-316

institute: Yale University School of Medicine

summary:
Tissue was microdissected from 13 regions, including 9 distinct neocortical areas, from both left and right sides of four late second trimester human brain specimens. Gene- and exon-level differential expression analyses were performed by mixed model, nested analysis of variance using the XRAY software from Biotique Systems. Further details available in Johnson, Kawasawa, et al., "Functional and Evolutionary Insights into Human Brain Development through Global Transcriptome Analysis" Neuron, Volume 62, Issue 4, 2009

Keywords: gene- and exon-level analyses

overall_design: For initial analysis, all neocortex samples were grouped together. In subsequent analyses, neocortex areas were compared with each other. In most analyses reported, left and right side samples were treated as additional biological replicates.

first: Yuka

middle: I

last: Kawasawa

first: Nenad

last: Sestan


title: Transcriptional signatures of Itk-deficient CD3+, CD4+ and CD8+ T-cells

city: Huddinge

first: Emelie

last: Blomberg

address: Novum level 5

department: Department of Laboratory Medicine, Clinical Research Center

country: Sweden

institute: Karolinska Institutet


title: Transcriptional signatures of Itk-deficiency using CD3+ T-cells

summary: The Tec-family kinase Itk plays an important role during T-cell activation and function, and controls also conventional versus innate-like T-cell development. We have characterized the transcriptome of Itk-deficient CD3+ T-cells, including CD4+ and CD8+ subsets, using Affymetrix microarrays. The largest difference between Itk-/- and Wt CD3+ T-cells was found in unstimulated cells, e.g. for killer cell lectin-like receptors. Compared to anti-CD3-stimulation, anti-CD3/CD28 significantly decreased the number of transcripts suggesting that the CD28 co-stimulatory pathway is mainly independent of Itk. The signatures of CD4+ and CD8+ T-cell subsets identified a greater differential expression than in total CD3+ cells. Cyclosporin (CsA)-treatment had a stronger effect on transcriptional regulation than Itk-deficiency, suggesting that only a fraction of TCR-mediated calcineurin/NFAT-activation is dependent on Itk. Bioinformatic analysis of NFAT-sites of the group of transcripts similarly regulated by Itk-deficiency and CsA-treatment, followed by chromatin-immunoprecipitation, revealed NFATc1-binding to the Bub1, IL7R, Ctla2a, Ctla2b, and Schlafen1 genes. Finally, to identify transcripts that are regulated by Tec-family kinases in general, we compared the expression profile of Itk-deficient T-cells with that of Btk-deficient B-cells and a common set of transcripts was found. Taken together, our study provides a general overview about the global transcriptional changes in the absence of Itk.

overall_design: CD3+ T-cells from pooled suspensions of spleen and lymph nodes of Wt and Itk knockout mice on C57BL/6 background were isolated after negative depletion. Unstimulated as well as stimulated T-cells were studied. Stimulations were done with anti-CD3 (1 mg/ml) with or without anti-CD28 (3 mg/ml) in the presence or absence of CsA (1 mg/ml) for 24 hrs. For each stimulus, at least duplicate samples were used.

first: K

middle: E

first: Nicole

last: Boucheron

first: Jessica

last: Lindvall

first: Liang

first: Julia

last: Raberger

first: Anna

last: Berglöf

first: Wilfried

last: Ellmeier

first: C

last: Smith


title: Cell-Specific Kras and Pten Mutations Document Proliferation Arrest in Granulosa Cells vs. Oncogenic Insult to OSE Cells

status: Public on May 15 2009

city: Houston

first: JoAnne

last: Richards

state: TX

address: One Baylor Plaza

department: Molecular and Cellular Biology

institute: Baylor College of Medicine

summary: The small G-protein KRAS is crucial for mediating gonadotropin-induced events associated with ovulation. However, constitutive expression of KrasG12D in granulosa cells disrupted normal follicle development leading to the persistence of abnormal follicle-like structures containing non-mitotic cells. To determine what factors mediate this potent effect of KrasG12D, gene profiling analyses were done. We also analyzed KrasG12D;Cyp19-Cre and KrasG12D;Pgr-Cre mutant mouse models that express Cre prior to or after the initiation of granulosa cell differentiation, respectively. KrasG12D induced cell cycle arrest in granulosa cells of the KrasG12D;Cyp19-Cre mice but not in the KrasG12D;Pgr-Cre mice, documenting the cell context specific effect of KrasG12D. Expression of KrasG12D silenced the Kras gene, reduced cell cycle activator genes and impaired expression of granulosa cell and oocyte specific genes. Conversely, levels of PTEN and phosphorylated p38MAPK increased markedly in the mutant granulosa cells. Because disrupting Pten in granulosa cells leads to increased proliferation and survival, Pten was disrupted in the KrasG12D mutant mice. The Pten/Kras mutant mice were infertile but lacked GCTs. By contrast, the Ptenfl/fl;KrasG12D;Amhr2-Cre mice developed aggressive ovarian surface epithelial (OSE) cell tumors that did not occur in the Ptenfl/fl;KrasG12D;Cyp19-Cre or Ptenfl/fl;KrasG12D;Pgr-Cre mouse strains. These data document unequivocally that Amhr2-Cre is expressed in and mediates allelic recombination of oncogenic genes in OSE cells. That KrasG12D/Pten mutant granulosa cells do not transform but rather undergo cell cycle arrest indicates that they resist the oncogenic insults of Kras/Pten by robust self-protecting mechanisms that silence the Kras gene and elevate PTEN and phospho-p38MAPK.

overall_design: Whole ovaries were collected from 26-day-old wild type mice, 26-day-old K-ras conditional mutant mice and 3-month-old K-ras conditional mutant mice. The gene expression profiles of these samples were compared using microarray method.

first: Heng-Yu

last: Fan

first: Zhilin

last: Liu

first: Marilene

last: Paquet

first: Jinrong

last: Wang

first: John

last: Lydon

first: Francesco

middle: J

last: DeMayo


title: A Regulatory Pathway Involving Notch1/β-Catenin/Isl1 Determines Cardiac Progenitor Cell Fate

status: Public on May 13 2009

city: San Francisco

first: Chulan

last: Kwon

address: 1650 Owens Street

institute: Gladstone Institutes

summary: The regulation of multipotent cardiac progenitor cell (CPC) expansion and subsequent differentiation into cardiomyocytes, smooth muscle, or endothelial cells is a fundamental aspect of basic cardiovascular biology and cardiac regenerative medicine. However, the mechanisms governing these decisions remain unclear. Here, we show that Wnt/β-Catenin signaling, which promotes expansion of CPCs, is negatively regulated by Notch1-mediated control of phosphorylated β-Catenin accumulation within CPCs, and that Notch1 activity in CPCs is required for their differentiation.  Notch1 positively, and β-Catenin negatively, regulated expression of the cardiac transcription factors, Isl1, Myocd and Smyd1. Surprisingly, disruption of Isl1, normally expressed transiently in CPCs prior to their differentiation, resulted in expansion of CPCs in vivo and in an embryonic stem (ES) cell system. Furthermore, Isl1 was required for CPC differentiation into cardiomyocyte and smooth muscle cells, but not endothelial cells. These findings reveal a regulatory network controlling CPC expansion and cell fate that involve unanticipated functions of β-Catenin, Notch1 and Isl1 that may be leveraged for regenerative approaches involving CPCs.

overall_design: YFP+ cells marking precardiac mesoderm (Isl1-cre domain) were FACS-sorted (w/wo stabilized Beta-catenin). Their total RNA was amplified with the WT-Ovation Pico RNA Amplification System, fragmented and labeled with the FL-Ovation cDNA Biotin Module V2 (Nugen). The hybridization, staining and scanning of the Affymetrix GeneChips were performed in the Gladstone Genomics Core Lab. Raw data generated from at least three independent experiments were further analyzed by the group of Dr. Ru-Fang Yeh at the Center for Informatics and Molecular Biostatistics, UCSF.

first: Deepak

last: Srivastava


title: Mesenchymal signature of post-pneumonectomy lung regeneration in adult mice

status: Public on May 08 2009

city: North Grafton

middle: A

last: Paxson

address: 200 Westboro Road

department: Clinical Sciences

institute: Tufts Cummings School of Veterinary Medicine

summary: The adult human lung has a very limited capacity to regenerate functional alveoli. In contrast, adult mice have a remarkable capacity for neoalveolarization following either lung resection or injury. The molecular basis for this unique capability to regenerate lung tissue in mice is largely unknown. We examined the transcriptomic responses to single lung pneumonectomy in adult mice in order to elucidate prospective molecular signaling used in this species during lung regeneration. Unilateral left pneumonectomy or sham thoracotomy was performed under general anesthesia (n = 8 mice per group for each of the four time points). Total RNA was isolated from the remaining lung tissue at four time points post-surgery (6 hours, 1 day, 3 days, 7 days) and analyzed using microarray technology. The observed transcriptomic patterns revealed mesenchymal cell signaling, including up-regulation of genes previously associated with activated fibroblasts (Tnfrsf12a, Tnc, Eln, Col3A1), as well as modulation of Igf1-mediated signaling. The data set also revealed early down-regulation of pro-inflammatory cytokine transcripts, up-regulation of genes involved in T cell development and function, but few similarities to transcriptomic patterns observed during embryonic or post-natal lung development. Immunohistochemical analysis suggests that early fibroblast but not myofibroblast proliferation is important during lung regeneration and may explain the preponderance of mesenchymal-associated genes that are over-expressed in this model.   This appears to differ from embryonic alveologenesis.  These data suggest that modulation of mesenchymal cell signaling and proliferation may act in concert with immunomodulation to control inflammation during post-pneumonectomy lung regeneration in adult mice.

overall_design: For each of the four time points (6 hr, 1 day, 3 day, 7 day), the mice were divided into two groups: (1) pneumonectomy (PNY) and (2) sham operated (SHAM - thoracotomy without lung resection), with eight animals in each group. One microarray was performed on pooled lung tissue from these 8 animals for each time point.  Analysis of expression in PNY vs SHAM animals was performed in two ways: using all four microarrays as replicates in a time-independent analysis, as well as analysis of each time point separately (time-dependent) without replicates.

first: Chris

last: Parkin

first: Lak

last: Iyer

first: Andrew

last: Hoffman


title: WT Dorsal Skin Time Course

first: Amelia

last: Hopkin

overall_design: To gain insights into common and unique transcriptional regulatory programs during epidermal differentiation, we profiled global gene expression in mouse dorsal skin at E14.5, E16.5, and E18.5.

first: Zhengquan

last: Yu

first: Jaana

last: Mannik

last: Soto

first: Kevin

last: Lin

first: Bogi

last: Andersen


title: Hematopoietic Stem Cells Reversibly Switch from Dormancy to Self-Renewal during Homeostasis and Repair

city: Heidelberg

first: Andreas

last: Trumpp

address: Im Neuenheimer Feld 280

department: Division of Stem Cells and Cancer

country: Germany

institute: Deutsches Krebsforschungszentrum (DKFZ)

summary: Bone marrow hematopoietic stem cells (HSCs) are crucial to maintain lifelong production of all blood cells. Although HSCs divide infrequently, it is thought that the entire HSC pool turns over every few weeks, suggesting that HSCs regularly enter and exit cell cycle. Here, we combine flow cytometry with label-retaining assays (BrdU and histone H2B-GFP) to identify a population of dormant mouse HSCs (d-HSCs) within the lin(-)Sca1+cKit+CD150+CD48(-)CD34(-) population. Computational modeling suggests that d-HSCs divide about every 145 days, or five times per lifetime. d-HSCs harbor the vast majority of multilineage long-term self-renewal activity. While they form a silent reservoir of the most potent HSCs during homeostasis, they are efficiently activated to self-renew in response to bone marrow injury or G-CSF stimulation. After re-establishment of homeostasis, activated HSCs return to dormancy, suggesting that HSCs are not stochastically entering the cell cycle but reversibly switch from dormancy to self-renewal under conditions of hematopoietic stress

overall_design: cDNA microarray analysis was performed on sorted mouse bone marrow after 170 days of doxycycline treatment for CD34 Neg, Lin Neg, Sca1+, cKit+, CD150+, CD48 Neg, HSCs. HSCs were further sorted for GFP+ (LRC) and GFP neg (non-LRC) populations.  Per condition, 3 independent biological replicates were analysed.

first: Anne

last: Wilson

first: Elisa

last: Laurenti

first: Gabriela

last: Oser

first: Richard

last: Van der Wath

first: William

last: Blanco-Bose

first: Maike

last: Jaworski

first: Sandra

last: Offner

first: Cyrille

last: Dunant

first: Leonid

last: Eshkind

first: Ernesto

last: Bockamp

first: Pietro

last: Lio

first: H

last: MacDonald


title: Gene expression profiling in the lung and liver of Perfluorooctane sulfonate (PFOS) exposed mouse fetuses

status: Public on May 12 2009

city: Research Triangle Park

first: Mitchell

last: Rosen

state: NC

department: Reproductive Toxicology

institute: U.S. EPA

summary: Most of the transcriptional changes induced by PFOS in the fetal mouse liver and lung were related to activation of PPARalpha.  When compared to the transcript profiles induced by PFOA (Pubmed ID 17681415), few remarkable differences were found other than up-regulation of Cyp3a genes.  Because PFOS and PFOA have been shown to differ in their mode of action in the murine neonate, these data suggest that changes related to PFOS-induced neonatal toxicity may not be evident in the fetal transcriptome at term.

overall_design: Thirty timed-pregnant CD-1 mice were orally dosed from gestation day 1-17 with either 0, 5, or 10 mg/kg/day PFOS in 0.5% Tween 20.  At term, fetal lung and liver were collected, total RNA prepared, and samples pooled from three fetuses per litter.  Five biological replicates consisting of individual litter samples were then evaluated for each treatment group using Affymetrix mouse 430_2 microarrays.

first: Judith

last: Schmid

first: Kaberi

middle: P

last: Das

first: Carmen

last: Wood

first: Robert

middle: D

last: Zehr

first: Christopher

last: Lau


title: Identification of genes expressed preferentially in the developing peripheral margin of the optic cup

status: Public on May 01 2009

first: jeffrey

last: trimarchi

summary: Identification of genes expressed in a preferential manner in the developing ciliary body/iris will provide a starting point for future functional analyses.  To identify candidate genes expressed in a variety of ocular tissues during development, we have profiled single cells from the developing eye. Post hoc identification of the origin of these cells showed that they included cells from the periphery of the developing optic cup.  By comparing the expression profiles of these cells to many retinal cell types, candidate genes for preferential expression in the periphery were identified.

overall_design: Single retinal cells were isolated in tubes containing lysis buffer, their mRNAs were reverse transcribed, and the resulting cDNAs were PCR amplified for 35 cycles. Labeled cDNA samples were hybridized to Affymetrix 430 2.0 microarrays and the data was normalized using MAS5.0 software.  Since the retinal cells isolated during these developmental times had not yet adopted their mature morphologies, a post hoc strategy was employed to classify each cell based upon the expression of clusters of genes (Trimarchi et al., 2008).  This classification method allowed for the identification of 21 single cells as developing retinal ganglion (RGC), amacrine (AC) or photoreceptor cells (PR) (Trimarchi et al., 2007) and an additional 42 cells as cycling RPCs (Trimarchi et al., 2008).  However, a number of the single cells did not score significantly for any of the retinal cell types.  Two cells in particular expressed genes that suggested these cells originated from the developing ciliary body/iris region.

first: Jeffrey

last: Trimarchi

first: Constance

last: Cepko


title: Gene expression changes in As4.1 cells during treatment with interleukin (IL) or hydrogen peroxide (HP)

city: Iowa City

first: Henry

last: Keen

state: IA

department: Pharmacology

institute: University of Iowa

summary: As4.1 cells are a renin-expressing cell line commonly used to study the molecular regulation of the mouse renin gene.  In the present study, the global gene expression profile was assessed in these cells under control conditions (VEHICLE) and after treatment with interleukin (IL) or hydrogen peroxide (HP), both of which negatively regulate mouse renin gene expression.

overall_design: For each experimental group, 2 separate cultures of As4.1 cells were used.  Cells were treated with interleukin (IL), hydrogen peroxide (HP) or vehicle control (VEHICLE). Cellular RNA was prepared using conventional methods and quality was assessed using the Bioanalyzer 2100 (Agilent Technologies).  All the microarray procedures were conducted at the University of Iowa DNA Core facility using standard Affymetrix protocols.  In brief, approximately 5 ug of total RNA was used as input to a one-step amplification procedure to generate biotin-labeled RNA fragments for hybridization to the Affymetrix GeneChip Mouse Genome 430 2.0 array.

first: Hana

last: Itani

first: Xuebo

last: Liu

first: Ehab

middle: H

last: Sarsour

first: Prabhat

last: Goswami

first: Ella

last: Born

first: Curt

last: Sigmund


title: Comparison of genetic and pharmacologic activation of Nrf2 signaling

city: Baltimore

first: Thomas

middle: W

last: Kensler

address: 615 N Wolfe St SPH Rm E7541

institute: Johns Hopkins School of Medicine

summary:
Genetic versus chemoprotective activation of Nrf2 signaling: overlapping yet distinct hepatic gene expression profiles between Keap1 knockout and triterpenoid treated mice

Loss of Nrf2 signaling increases susceptibility to acute toxicity, inflammation, and carcinogenesis in mice due to the inability to mount adaptive responses. By contrast, disruption of Keap1 (a cytoplasmic modifier of Nrf2 turnover) protects against these stresses in mice; although dominant negative mutations in Keap1 have been identified recently in some human cancers.  Global characterization of Nrf2 activation is important to exploit this pathway for chemoprevention in healthy, yet at-risk individuals and also to elucidate the consequences of hijacking the pathway in Keap1-mutant human cancers.  This analysis also enables a global characterization of the pharmacodynamic action of CDDO-Im at a low dose that is relevant to chemoprevention.

overall_design: Liver-targeted conditional Keap1-null (CKO) mice provide a model of genetic activation of Nrf2 signaling.  By coupling global gene expression analysis of CKO mice with analysis of pharmacologic activation using the synthetic oleanane triterpenoid CDDO-Im, we are able to gain insight into pathways affected by Nrf2 activation.  CDDO-Im is an extremely potent activator of Nrf2 signaling.  CKO mice were used to identify genes modulated by genetic activation of Nrf2 signaling.  The CKO response was compared to hepatic global gene expression changes in wild-type mice treated with CDDO-Im at a maximal Nrf2 activating dose.  n=3/group, male 9 week old mice were used.  Mice were treated with a single dose of vehicle (10% Cremophor-EL, 10% DMSO, and PBS) or 30 umol CDDO-Im/kg body weight by gavage and sacrificed 6 h later.

first: Melinda

last: Yates

first: Quynh

last: Tran

first: Patrick

last: Dolan

first: William

last: Osburn

first: Soona

last: Shin

first: Colin

last: McCulloch

first: Jay

last: Silkworth

first: Keiko

last: Taguchi

first: Masayuki

last: Yamamoto

first: Charlotte

last: Williams

first: Karen

last: Liby

first: Michael

last: Sporn

last: Sutter


title: Gene expression of Mus exposed to chronic hypoxia

status: Public on Apr 30 2009

city: Reno

first: Monica

middle: M

last: Baze

state: NV

address: Dept. of Biology/ 314

department: Biology

institute: University of Nevada, Reno

summary:
High altitude environments are characterized by the unique and unavoidable stress of chronic hypoxia. While much is known about gene expression responses to acute or in vitro hypoxia, less is known about the gene expression profiles of animals exposed to systemic chronic hypoxia, such as that experienced at high elevations. Here we simulated the hypoxic environment of two high altitude elevations,and a third chamber recieved ambient Reno air. Mice were housed in the hypoxic chambers for 32 days.
We used microarrays to characterize the differential gene expression in the livers of mice housed in hypoxic environment of 4500 m versus 3000 and 1400 m. We used this data to draw hypotheses related to novel physiological responses to chronic systemic hypoxia

overall_design: Mice were housed one of three chambers; the first received ambient Reno air (1400 m) and the other two received air mixed with nitrogen such that one chamber simulated the hypoxic environment of 3000 m and the third chamber simulated hypoxic environment of 4500 m. Twelve mice were housed in each chamber for 32 days. Liver were extracted, and RNA from livers of 4 mice were pooled such that each treatment was represented by 3 pooled samples.  	Eight of the nine arrays were used for data analysis; one array was excluded as several Affymetrix quality control metrics indicated data quality that was not reliable.

first: Jack

last: Hayes

last: Schlauch


title: Expression data from colon epithelium of STAT3IEC-KO in acute DSS colitis

city: Erlangen

first: Clemens

last: Neufert

address: Ulmenweg 18

department: First Department of Medicine

summary: STAT3 is a pleiotropic transcription factor with important functions in cytokine signalling in a variety of tissues. However, the role of STAT3 in the intestinal epithelium is not well understood. Here we demonstrate that development of colonic inflammation is associated  with the induction of STAT3 activity in intestinal epithelial cells (IEC). Studies in genetically engineered mice showed that epithelial STAT3 activation in DSS colitis is dependent on IL-22 rather than IL-6. IL-22 was secreted by colonic CD11c+ cells in response to Toll-like receptor stimulation. Conditional knockout mice with an IEC specific deletion of STAT3 activity were highly susceptible to experimental colitis, indicating that epithelial STAT3 regulates gut homeostasis. STAT3IEC-KO mice, upon induction of colitis, showed a striking defect of epithelial restitution. Gene chip analysis indicated that STAT3 regulates the cellular stress response, apoptosis and pathways associated with wound healing in IEC. Consistently, both IL-22 and epithelial STAT3 were found to be important in wound-healing experiments in vivo. In summary, our data suggest that intestinal epithelial STAT3 activation regulates immune homeostasis in the gut by promoting IL-22-dependent mucosal wound healing.

overall_design: 4 samples of colon epithelium were analyzed from 4 mice (2 per group Stat3flfl VillinCre- and Stat3flfl VillinCre+, respectively) after they had been treated with DSS (2.5%) for 5 days

first: Christoph

last: Becker

first: Geethanjali

last: Pickert

first: Alexei

last: Nikolaev

first: Markus

last: Neurath


title: Ovarian Development in Mice Requires GATA4/FOG2 Transcriptional Complex

city: Hanover

first: Sergei

last: Tevosian

state: NH

address: 7400 Hinman Box

institute: Dartmouth Medical School

summary:
We have demonstrated previously that mammalian sexual differentiation requires both GATA4 and FOG2 transcription regulators to assemble the functioning testis. We have now determined that the sexual development of female mice is profoundly affected by the loss of GATA4-FOG2 interaction. We have also identified the Dkk1 gene, encoding a secreted inhibitor of canonical β-catenin signaling as a target of GATA4/FOG2 repression in the developing ovary. The tissue-specific ablation of the β-catenin gene in the gonads disrupts female development while in the Gata4ki/ki/Dkk1-/- or Fog2-/-/Dkk1-/- embryos the normal ovarian gene expression pattern is partially restored. Control of ovarian development by the GATA4/FOG2 complex presents a novel insight into the crosstalk of transcriptional regulation and extracellular signaling in ovarian development.
Keywords: genetic modification

overall_design: 6 control and 6 Gata4ki/ki mutant samples were analyzed

first: Nikolay

middle: L

last: Manuylov

first: Danielle

last: Maatouk

first: Fatima

middle: O

last: Smagulova

first: Lyndsay

last: Leach

first: Blanche

last: Capel


title: SPARC-null vs. wild-type lens epithelium

city: Seattle

first: Teri

last: Greiling

state: WA

department: Biological Structure

institute: University of Washington

summary: SPARC is a matricellular glycoprotein involved in regulation of the extracellular matrix, growth factors, adhesion, and migration.  SPARC-null mice have altered basement membranes and develop posterior sub-capsular cataracts with cell swelling and equatorial vacuoles.  Exchange of fluid, nutrients, and waste products in the avascular lens is driven by a unique circulating ion current.  Here we demonstrate that SPARC-null mouse lenses exhibit abnormal circulation of fluid, ion, and small molecules which leads to altered fluorescein distribution in vivo, loss of resting membrane polarization, and altered distribution of small molecules.  Microarray analysis of SPARC-null lenses showed changes in gene expression of ion channels and receptors, matrix and adhesion genes, cytoskeleton, immune response genes, and cell signaling molecules.  Our results demonstrate that the regulation of SPARC on cell-capsular matrix interactions can influence the circulation of fluid and ions in the lens, and the phenotype in the SPARC-null mouse lens is the result of multiple intersecting pathways.

overall_design: Lens epithelial cells from 7 lenses of littermate mice were isolated by laser capture microdissection.  3 wild-type lenses from 3 different mice and 4 knock-out lenses from 3 different mice were used as biological replicates.

first: Brad

last: Stone

first: John

last: Clark


title: Transcriptional signatures of Itk-deficiency using CD3+, CD4+ and CD8+ T-cells

first: Emelie

last: Blomberg

overall_design: CD3+ CD4+ and CD8+ T-cells from pooled suspensions of spleen and lymph nodes of Wt and Itk knockout mice on C57BL/6 background were isolated after negative depletion. Unstimulated as well as stimulated T-cells were studied. Stimulations were done with anti-CD3 (1 mg/ml) for 24 hrs. For the CD4+ T-cells we collected triplicates from the Itk knockout mice and duplicates from the Wt group. For the CD8+ T-cells, we got duplicates from Itk knockout , while we obtained a single sample from Wt owing to the low cell yield for resting Wt CD8+ T-cells. After CD3-stimulation we got a single sample from the CD8+ subset of both Wt and Itk knockout, while for the CD4+ subsets we collected duplicates.

first: K

first: Nicole

last: Boucheron

first: Jessica

last: Lindvall

first: Liang

last: Yu

first: Julia

last: Raberger

first: Anna

last: Berglöf

first: Wilfried

last: Ellmeier

first: C

last: Smith


title: Gene expression changes in Bmi1 knock-out MEFs as compared to wild-type.

city: Barcelona

first: Eric

middle: M.

last: Kallin

state: Barcelona

address: C/ Dr. Aiguader, 88

department: Differentiation and Cancer

country: Spain

institute: Center for Genomic Regulation

summary: Polycomb group (PcG) proteins control organism development by regulating the expression of developmental genes. Transcriptional regulation by PcG proteins is achieved at least partly through the PRC2-mediated methylation on lysine 27 of histone H3 (H3K27) and PRC1-mediated ubiquitylation on lysine 119 of histone H2A (uH2A). As an integral component of PRC1, Bmi1 has been demonstrated to be critical for H2A ubiquitylation. Although recent studies have revealed the genome wide binding patterns of some of the PRC1 and PRC2 components, as well as the H3K27me3 mark, there have been no reports describing genome wide localization of uH2A. Using the recently developed ChIP-Seq technology, here we report genome wide localization of the Bmi1-dependent uH2A mark in MEF cells. Gene promoter averaging analysis indicates a peak of uH2A just inside the transcription start site (TSS) of well annotated genes.  This peak is enriched at promoters containing the H3K27me3 mark and represents the least expressed genes in WT MEF cells.  In addition, peak finding reveals regions of local uH2A enrichment throughout the mouse genome, including almost 700 gene promoters.  Genes with promoter peaks of uH2A exhibit lower level expression when compared to genes that do not contain promoter peaks of uH2A.  Moreover, we demonstrate that genes with uH2A peaks have increased expression upon Bmi1 knockout.  Importantly, local enrichment of uH2A is not limited to regions containing the H3K27me3 mark.  We provide evidence to suggest that DNA methylation is tightly linked to H2A ubiquitylation in high density CpG promoters.  Thus, our work not only reveals Bmi1-dependent H2A ubiquitylation but also suggests that uH2A targeting in differentiated cells may employ a different mechanism from that in ES cells.

overall_design: RNA was extracted from wild-type MEF cells which were immortalized through TBX2 overexpression, amplified, labeled, and hybridized to an Affymetrix 430 2 microarray. In parallel, RNA was prepared from TBX2-immortalized Bmi1 null MEF cells and hybridized to an Affymetrix 430 2 microarray.

first: Ru

last: Cao

first: Raja

last: Jothi

first: Kai

last: Xia

first: Kairong

last: Cui

first: Keji

last: Zhao

first: Yi

last: Zhang


title: Gene expression analysis of SH-SY5Y neuroblastoma cells overexpressing ZNF423

status: Public on Apr 29 2009

city: Amsterdam

first: Sidong

last: Huang

address: Plesmanlaan 121

country: Netherlands

institute: The Netherlands Cancer Institute

summary: We have identified ZNF423 (also known as Ebfaz, OAZ or Zfp423) as a component critically required for retinoic acid (RA)-induced differentiation. ZNF423 associates with the RARα/RXRα nuclear receptor complex and is essential for transactivation in response to retinoids. Down-regulation of ZNF423 expression by RNA interference in neuroblastoma cells results in a growth advantage and resistance to RA-induced differentiation, whereas overexpression of ZNF423 leads to growth inhibition and enhanced differentiation. Futhermore, we show that low ZNF423 expression is associated with poor disease outcome of neuroblastoma patients. To identify the other key pathways regulated by ZNF423 in human neuroblastoma, we expressed elevated levels of ZNF423 in SH-SY5Y cells and performed full genome gene expression analysis in these cells.

overall_design: Human neuroblastoma SH-SY5Y cells (expressing the murine ecotropic receptor)  were retrovirally infected with MSCV-ZNF423 or MSCV vector control.  The gene expression analysis of these cells was then performed using the Illumina platform. The representative experiments are shown.


title: Gene expression profiles in dendritic cells from Peyer's patches and epidermal of skin

status: Public on Apr 28 2009

city: Wuhan

first: quan

last: xing

address: Luoyu Road 237

country: China

institute: wuhan university

summary: Many studies have shown that the mucous membranes and skin are at the interface with different external environments and face the disparity of pathogenic effects, such as biological agents, chemical or physical environment. This difference may demand distinct immune responses. However, the mechanism to induce the distinct immune responses in mucous and skin is largely unknown. Dendritic cells of mucosa and skin are crucial in the initiation of immune responses, maintenance of self-tolerance and antigens presentation T cells. The different functions between mucosal and epidermal dendritic cells may play an important role in different immune responses. To compare the different gene expression of the mucosal DC and Langerhans cells (LC), we utilized microarrays to investigate different gene expression profiles in mucosal DC isolated from PPs (PDC) and epidermal LC from skin (ELC). 3548 genes were shown to be differentially expressed between PDC and ELC. According to genes annotations, 105 genes may be involved in immunity process. The genes involved in immune process were categorized to five groups related to DC function, including antigen presentation, antigen uptake, cytokines chemokines, and receptors, cell surface molecules and signal transduction. 11 of the highest expressed genes were selected as the candidate genes and reformed by real-time PCR. These 11 selected genes might be suitable candidates to further study the difference of gene expression between mucosal DC and epidermal LC and would be used for design for new vaccine.

overall_design: Beads and FACS sort were used to isolate dendritic cells from Peyer's patches and epidermal of skin from 80 four-week-old female BALB/c mice. Dendritic cells from Peyer's patches were pooled as one sample, and dendritic cells from epidermal of skin were pooled as another sample. RNA isolation, amplification, cDNA labelling, microarray hybridization and analyses were performed according to the Affymetrix manual book.

first: Quan

last: Xing

first: Yuhong

last: Li

first: Mingwen

last: Fan


title: Gene expression profiles of E13.5 developing podocyte in the developing kidney isolated from MafB-GFP transgenic mice using FACS on 430 2.0 Array chip. (GUDMAP Series ID: 27)

status: Public on Apr 25 2009

city: Edinburgh

first: GUDMAP

last: Developers

address: Crewe Road

institute: IGMM MRC Human Genetics Unit

summary: The long term objective is to create an encyclopedia of the expression levels of all genes in multiple components of the developing kidney.  The central thesis is straightforward. The combination of fluorescent activated cell sorting (FACS) plus microarray analysis offers a powerful, efficient and effective method for the creation of a global gene expression atlas of the developing kidney. Microarrays with essentially complete genome coverage can be used to quantitate expression levels of every gene in FACS isolated components of the developing kidney. The ensuing rapid read-out provides an expression atlas that is more sensitive, more economical and more complete than would be possible by in situ hybridizations alone.

overall_design: At E13.5 we isolated developing cells of the podocyte using Mafb transgenic mice and fluorescent activated cell sorting (FACS) and then defined the gene expression profiles with microarrays.

first: Steve

last: Potter

last: Brunskill


title: Gene expression profiles of E15 ureter. (GUDMAP Series ID: 26)

summary: The long term objective is to create an encyclopedia of the expression levels of all genes in multiple components of the developing genitourinary tract.  The central thesis is straightforward. The combination of microdissected tissues and FACS sorted cells plus microarray analysis offers a powerful, efficient and effective method for the creation of a global gene expression atlas of the developing urogenital system. Microarrays with essentially complete genome coverage can be used to quantitate expression levels of every gene. The ensuing rapid read-out provides an expression atlas that is more sensitive, more economical and more complete than would be possible by in situ hybridizations alone. The data submitted here delineates the gene expression profiles of the e15 mouse ureter.  Comparisons with the same compartments in the neonate will be made to determine changes in gene expression from relatively undifferentiated tissue (e15) to a highly differentiated structure (neonate).

overall_design: FVB/N mice were time mated. At embryonic day 15 mice were euthanized by decapitation and the ureters were microdissected, cut from pelvis to the bladder insertion point and snap frozen in liquid nitrogen. Total RNA was isolated for gene expression analysis using the Affymetrix GeneChip Mouse Genome 430 2.0 Array.

first: James

last: Lessard

first: Alfor

last: Lewis


title: Irradiated stroma selects for invasive and metastatic tumoc cells

city: Lausanne

first: Mauro

last: Delorenzi

address: 155 Boveresses

country: Switzerland

institute: ISREC

summary:
Radiotherapy is widely used to treat human cancer. Patients locally recurring after radiotherapy, however, have increased risk of metastatic progression and poor prognosis. The clinical management of post-radiation recurrences remains an unresolved issue. Tumors growing in pre-irradiated tissues have an increased fraction of hypoxic cells and are more metastatic, a condition known as tumor bed effect. Here we demonstrate that tumor cells growing in a pre-irradiated bed, or selected in vitro though repeated cycles of severe hypoxia, retain an invasive and metastatic capacities when returned to normoxia. HIF activity, while it facilitates metastatic spreading of tumors growing in a pre-irradiated bed, is not essential. Through gene expression profiling and gain and loss of function experiments, we identified the matricellular protein CYR61 and aVb5 integrin, as proteins cooperating to mediate these effects. Inhibition of aVb5 integrin suppressed invasion and metastasis induced by CYR61 and attenuated metastasis of tumors growing within a pre-irradiated field. These results represent a conceptual advance to the understanding of the tumor bed effect and identify CYR61 and aVb5 integrin as proteins that co-operate to mediate metastasis. They also indicate aV integrin inhibition a potential therapeutic approach for preventing metastasis in patients at risk for post-radiation recurrences, which can be promptly tested in the clinic.
Keywords: Tumour profiling

overall_design: The overall experimental design consist of a single channel experiment where a total of 9 independent samples of 3 different conditions were analyzed.  The SCCVII murine cell line were grown either in an iradiadiated (IR) or non-irradiated (NIR) environment.  The parental cell line (PA) was also hybridized as control.

first: Yan

last: Monnier

first: Pierre

last: Farmer

first: Gregory

last: Bieler

first: Natsuko

last: Imaizumi

first: Thierry

last: Sengstag

first: Gian-Carlo

last: Alghisi

first: Curzio

last: Ruegg


title: Professional memory CD4+ T lymphocytes preferentially reside and rest in the bone marrow

status: Public on Apr 21 2009

city: Berlin

first: Pawel

last: Durek

address: Charitéplatz 1

institute: Deutsches Rheuma-Forschungszentrum

summary: CD4+ T lymphocytes are key to immunological memory, but little is known about the lifestyle of memory CD4+ T lymphocytes. We showed that in the memory phase of specific immune responses to antigens, most of the memory CD4+ T lymphocytes relocated into the bone marrow (BM) within 3-8 weeks after their generation, a process involving integrin a2. Antigen-specific memory CD4+ T lymphocytes expressed Ly-6C to a high degree, unlike most splenic CD44hiCD62L- CD4+ T lymphocytes. In adult mice, more than 80% of Ly-6Chi CD44hiCD62L- memory CD4+ T lymphocytes were in the BM. In the BM, they are located next to IL-7-expressing VCAM-1+ stroma cells, and were in a resting state. Upon challenge with antigen, they rapidly expressed cytokines and CD154 and induced the production of high-affinity antibodies, indicating their functional activity in vivo and marking them as professional memory T helper cells

overall_design:
FACSAria sorted CD44highCD62L-CD25- CD4+ T cells of murine (C57BL/6 mice) bone marrow were compared to those of the spleen using Affymetrix GeneChip Mouse Genome 430A 2.0 Array. After total RNA extraction, reverse transcription, cDNA extraction, the biotinylated cRNA was transcribed, fragmented, and 15 µg cRNA hybridized in triplicates for each of the two groups to the GeneChip arrays. Group of bone marrow chips: GSM393497, GSM393940, GSM393941, group of spleen chips: GSM393942, GSM393943, GSM393944.
Lists of differentially regulated genes were created using High Performance Chip Data Analysis (HPCDA) with Bioretis database (http://www.bioretis-analysis.de). To see the list of significant genes, go to Bioretis database, register and with your username and password log into the database. Then check under Account, User Details the two boxes Public Chip Rights and Public Analyses. Then select under Comparison, View single results in the window Select an existing Analysis the analysis SPL_294. To find the list of significant genes, click an next, then on Choose an existing parameterset and select first a default increase parameterset (be shure that on row "t-Test EG vs. BG *" in the first 4 left columns <0.05 is included and check the box "* use Bonferroni correction"). Then click on Search only and you will get the list of differentially íncreased genes. Perform the same procedure with a default parameterset for decrease genes. You can get instruction/help about Bioretis database before Login on sheet introduction.

first: Koji

last: Tokoyoda

first: Sandra

last: Zehentmeier

first: Ahmed

middle: N

last: Hegazy

first: Inka

last: Albrecht

first: Joachim

middle: R

last: Grün

first: Max

last: Löhning

first: Andreas

last: Radbruch


title: Inhibitor trials in chondrocytes - MAS 5.0 normalization

city: London

first: veronica

last: ulici

state: Ontario

address: 1151 Richmond Street

department: Physiology and Pharmacology

country: Canada

institute: University of Western Ontario

summary:
Objectives: To identify similarities and differences in gene expression data in the MEK/ERK and PI3K pathways and to determine how histone modification affects these same pathways. Goal: To identify and functionally characterize novel targets of these signaling pathways in the context of chondrocyte differentiation.

Keywords: Treatment, signaling, one-colour array, signaling pathway comparison

overall_design:
Cartilage derived from the hind limbs of CD1 mice staged at 15.5 days of embryonic development were dissected and plated in primary chondrocyte monolayer cultures. Cells were treated with MEK/ERK pathway inhibitors (SB202190 and UO126), a PI3K inhibitor (LY294002), a histone deacetylase inhibitor (Trichostatin A) and the vehicle control (DMSO) for 24 hours. Total RNA was isolated from these samples and hybridized to Affymetrix MOE 430 2.0 chips at the London Regional Genomics facility.
A total of 15 genechips were analyzed between 4 treatments and the vehicle (DMSO) control. Three biological replicates per treatment were executed.

first: Claudine

middle: G

last: James

first: Veronica

last: Ulici

first: Frank

last: Beier

first: Lee-Anne

last: Stanton


title: Regulation of T cell gene expression by Halofuginone (HF)

status: Public on Apr 10 2009

city: Boston

first: CBDM

last: Lab

address: 77 Avenue Louis Pasteur

department: Microbiology and Immunobiology

institute: Harvard Medical School

summary:
T cell differentiation to the Th17 effector subset requires stimulation through the T cell and co-stimulatory receptors, together with cytokine stimulation by TGFb and IL-6. The small molecule halofuginone (HF) inhibits Th17 cell development and induces a pattern of stress-regulated gene expression that mimics amino acid starvation.
We used global transcript profiling to ask how halofuginone modulates gene expression induced during T cell activaiton and Th17 differentiation

Keywords: T cell activation/ differentiation timecourse

overall_design: Purified mouse CD4+ CD25- T cells were activated under Th17 polarizing cytokine conditions and treated with either halofuginone or its inactive derivative, MAZ1310, for 3- or 6-hours.

first: Mark

middle: S

last: Sundrud

first: Christophe

last: Benoist

first: Markus

last: Feuerer


title: Expression data from  synovial sarcoma-like tumors induced in a genetically engineered mouse model

status: Public on Apr 14 2009

city: St. Louis

first: MALAY

last: HALDAR

state: MO

address: 7751 charing square lane

department: Pathology and Immunology

institute: Washington University in St. Louis

summary: Synovial sarcoma-like tumors were generated in mice by conditionally expressing the human t(X;18) translocation-derived SYT-SSX2 fusion protein. Using a Tamoxifen-inducible CreER system, we show here that sporadic expression of SYT-SSX2 across multiple tissue types leads to exclusive formation of synovial sarcoma-like tumors while its widespread expression is lethal.  CreER-based sporadic expression both avoids the severe early developmental phenotypes     associated with widespread SYT-SSX2 expression and better models natural pathogenesis of cancers where transformed cells usually arise within an environment of largely normal cells.

overall_design: Genetically engineered mice capable of conditionally expressing the human synovial sarcoma-associated SYT-SSX2 fusion oncogene were mated with genetically engineered mice expressing the CreER fusion protein from ROSA locus. The progenies harboring both CreER and SYT-SSX2 were followed up with or without tamoxifen injection. Tumors were generated in these mice that were dissected out, RNA extracted, and subjected to expression profiling by microarray analysis.

first: Malay

last: Haldar

first: Mario

last: Capecchi


title: Zfx controls BCR-induced proliferation and survival of B lymphocytes

status: Public on Apr 17 2009

city: New York

first: Teresita

last: Arenzana

state: NY

department: Microbiology Department

institute: Columbia University

summary:
The development, homeostasis and function of B lymphocytes involve multiple rounds of B cell receptor (BCR)-controlled proliferation and prolonged maintenance. We analyzed the role of transcription factor Zfx, a recently identified regulator of stem cell maintenance, in B cell development and homeostasis. Conditional Zfx deletion in the bone marrow blocked B cell development at the pre-BCR selection checkpoint. Zfx deficiency in peripheral B cells caused impaired generation of the B-1 cell lineage, accelerated B cell turnover, depletion of mature recirculating cells, and delayed T-dependent antibody responses. Zfx-deficient B cells showed normal proximal BCR signaling, but impaired BCR-induced proliferation and survival. This was accompanied by aberrantly enhanced and prolonged integrated stress response, and delayed induction of Cyclin D2 and Bcl-xL proteins. Thus, Zfx restrains the stress response and couples antigen receptor signaling to B cell expansion and maintenance during development and peripheral homeostasis.

Keywords: Expression profiling by array

overall_design: For global gene expression analysis, 2.5 x 105 flow-sorted follicular B cells (B220+ AA4.1- CD23hi CD21int) were cultured in 200 μL B cell media containing 10 μg/mL α-IgM for 0, 2, and 12 hrs.  Cells were harvested in Trizol (Invitrogen). Total RNA was isolated using Trizol (Invitrogen), and 100 ng of each RNA sample were used for reverse transcription and two rounds of linear antisense RNA amplification/labeling (Ambion). The aRNA was amplified without labeling in the first round, and then labeled with Biotin-16-UTP(Roche Applied Sciences) and Biotin-11-CTP (PerkinElmer Life Sciences) in the second round. Labeled aRNA was fragmented in RNA Fragmentation Buffer (QIAGEN) prior to hybridization. Labeled RNA samples (10 ug/array) were hybridized in duplicate to Mouse Genome 430 2.0 arrays (Affymetrix) at the Columbia University Microarray Project core facility according to the manufacturer's instructions. Quality control and normalization were performed using positive and negative hybridization controls (Affymetrix) spiked into the RNA prior to labeling. Linear amplification of RNA was confirmed for every sample. Array scanning and raw data processing were done using GCOS 1.4 software (Affymetrix).

first: Matthew

last: Smith-Raska

first: Boris

last: Reizis


title: Linkage of Meis1 leukemogenic activity to multiple downstream effectors including Trib2 and Ccl3

status: Public on Apr 09 2009

city: Gothenburg

first: Lars

last: Palmqvist

address: Sahlgrenska University Hospital

department: Institute of Biomedicine

institute: University of Gothenburg

summary: OBJECTIVE: MEIS1, a HOX cofactor, collaborates with multiple HOX and NUP98-HOX fusion proteins to accelerate the onset of acute myeloid leukemia (AML) through largely unknown molecular mechanisms. MATERIALS AND METHODS: To further resolve these mechanisms, we conducted a structure-function analysis of MEIS1 and gene-expression profiling, in the context of NUP98-HOXD13 (ND13) leukemogenesis. RESULTS: We show, in a murine bone marrow transplantation model, that the PBX-interaction domain, the homeodomain, and the C-terminal domain of MEIS1, are all required for leukemogenic collaboration with ND13. In contrast, the N-terminal domain of MEIS1 is dispensable for collaboration with ND13, but is required for Flt3 upregulation, indicating additional roles for MEIS1 in induction of leukemia independent of alterations in Flt3 expression. Gene-expression profiling of a cloned ND13 preleukemic cell line transduced with wild-type or Meis1 mutant forms revealed deregulation of multiple genes, including a set not previously implicated as MEIS1 targets. Chromatin immunoprecipitation revealed the in vivo occupancy of MEIS1 on regulatory sequences of Trib2, Flt3, Dlk1, Ccl3, Ccl4, Pf4, and Rgs1. Furthermore, engineered overexpression of Trib2 complements ND13 to induce AML while Ccl3 potentiates the repopulating ability of ND13. CONCLUSION: This study shows that Meis1-induced leukemogenesis with ND13 can occur in the absence of Flt3 upregulation and reveals the existence of other pathways activated by MEIS1 to promote leukemia.

overall_design: Establishment of pre-leukemic bone marrow cell lines following transduction with ND13 have been previously described (Pineault, Abramovich et al. 2005). In brief, lines were established from BM cells previously treated with 5-fluorouracil (5-FU) from (C57Bl/6Ly-Pep3b x C3H/HeJ) F1 (PepC3) (Ly5.1+/Ly5.2+) mice freshly transduced with the ND13-GFP or ND13pac virus and maintained in liquid culture. To generate ND13+Meis1 or ND13+Meis1 mutant BM cell lines, the ND13 BM cells were transduced by co-cultivation on irradiated (4,000 cGy) E86 producers for Meis1-YFP (or Meis1 mutant forms), respectively, for a period of 2 days in the presence of 5 µg/ml of protamine sulfate. Three independent experiments were performed for each of the four different conditions included in the study.

first: Bob

last: Argiropoulos

first: Eric

last: Yung

first: Florian

last: Kuchenbauer

first: Michael

last: Heuser

first: Laura

last: Sly

first: Adrian

last: Wan

first: Gerald

last: Krystal

first: Keith

last: Humphries


title: Expression data from mouse normal thymus, thymus tumor, and XIST resistant thymus tumor

city: Vienna

first: christian

last: haslinger

state: Vienna

address: Dr. Boehringer-Gasse 5-11

country: Austria

institute: Boehringer Ingelheim Austria

summary: The non-coding Xist RNA triggers silencing of one of the two female X chromosomes during X inactivation in mammals. Gene silencing by Xist is restricted to special developmental contexts found in cells of the early embryo and specific hematopoietic precursors. The absence of critical silencing factors might explain why Xist cannot silence outside these contexts. Here, we show that Xist can also initiate silencing in a lymphoma model. Using the tumor context we identify the special AT rich binding protein SATB1 as an essential silencing factor. We show that loss of SATB1 in tumor cells abrogates the silencing function of Xist. In normal female lymphocytes Xist localizes along SATB1 filaments and, importantly, forced Xist expression can relocalize SATB1 into the Xist cluster. This reciprocal influence on localization suggests a molecular interaction between Xist and SATB1. SATB1 and its close homologue SATB2 are expressed during the initiation window for X inactivation in embryonic stem cells and are recruited to surround the Xist cluster. Furthermore, ectopic expression SATB1 or SATB2 enables gene silencing by Xist in embryonic fibroblasts, which normally do not provide an initiation context. Thus, SATB1 functions as a crucial initiation factor and may act to organize genes for silencing by Xist during the initiation of X inactivation.

overall_design: Normal mouse thymus, tumor, and XIST resistant tumor tissue was hybridized on the Affymetrix Mouse430_2 chip. All groups consist of triplicates.

first: Ruben

last: Agrelo

first: Abdallah

last: Souabni

first: Maria

last: Novatchkova

first: Christian

last: Haslinger

first: Martin

last: Leeb

first: Vukoslav

last: Komnenovic

first: Hiroyuki

last: Kishimoto

first: Lionel

last: Gresh

first: Terumi

last: Kohwi-Shigematsu

first: Lukas

last: Kenner

first: Anton

last: Wutz


title: Smad2 and 3 transcription factors  control muscle mass in adulthood

city: Padova

first: Reimar

last: Abraham

address: Via Orus 2

country: Italy

institute: Venetian Institute of Molecular Medicine

summary: Loss of muscle mass occurs in a variety of diseases including cancer, chronic heart failure, AIDS, diabetes and renal failure, often aggravating pathological progression. Preventing muscle wasting by promoting muscle growth has been proposed as a possible therapeutic approach. Myostatin is an important negative modulator of muscle growth during myogenesis and myostatin inhibitors are attractive drug targets. However, the role of the myostatin pathway in adulthood and the transcription factors involved in the signaling are unclear. Moreover recent results confirm that other TGFβ members control muscle mass. Using genetic tools we perturbed this pathway in adult myofibers, in vivo, to characterize the downstream targets and their ability to control muscle mass. Smad2 and Smad3 are the transcription factors downstream of myostatin/TGFβ and induce an atrophy program which is MuRF1 independent  and requires FoxO activity. Furthermore Smad2/3 inhibition promotes muscle hypertrophy independent of satellite cells but partially dependent of mTOR signalling. Thus myostatin and Akt pathways cross-talk at different levels. These findings point to myostatin inhibitors as good drugs to promote muscle growth during rehabilitation especially when they are combined with IGF1-Akt activators.

overall_design: Tansgenic mice were generated that express a fusion protein of Akt with the estrogen binding domain in skeletal muscle. Addition of tamoxifen stabilizes the protein leading to increased Akt signaling and muscle hypertrophy. RNA was extracted from gastrocnemius muscle after 24h oil or tamoxifen treatment or 48h tamoxifen treatment. RNA from the muscles of 3-4 mice were pooled for each chip (Affymetrix Mouse 430 2.0 whole genome expression).

first: Roberta

last: Sartori

first: Giulia

last: Milan

last: Patron

first: Christina

last: Mammucari

first: Bert

last: Blaauw

first: Marco

last: Sandri


title: Dppa4 is dispensable for embryonic stem cell identity and germ cell development, but essential for embryogenesis

city: Tuebingen

first: Michael

middle: H.

last: Walter

address: Calwer Str. 7

department: Department of Medical Genetics

institute: University of Tuebingen

summary: Dppa4 (Developmental pluripotency-associated 4) has been identified in several highprofile screens as a gene that is expressed exclusively in pluripotent cells. It encodes a nuclear protein with a SAP-like domain and appears to be associated preferentially with transcriptionally active chromatin. Its exquisite expression pattern and results of RNA interference experiments have led to speculation that Dppa4, as well as its nearby homolog Dppa2, might play essential roles in embryonic stem cell function and/or germ cell development. To rigorously assess suggested roles, we have generated Dppa4-deficient and Dppa4/Dppa2 double-deficient ES cells, as well as mice lacking Dppa4. Contrary to predictions, we find that Dppa4 is completely dispensable for ES cell identity and germ cell development. Instead, loss of Dppa4 in mice results in late embryonic/peri-natal death and striking skeletal defects with partial penetrance. Thus, surprisingly, Dppa4-deficiency affects tissues, which never transcribed the gene, and at least some loss-of-function defects manifest phenotypically at an embryonic stage long after physiologic Dppa4 expression has ceased. Concomitant with targeted gene inactivation, we have introduced into the Dppa4 locus a red fluorescent marker (tandem-dimer RFP), which is compatible with GFP-based proteins and allows non-invasive visualization of pluripotent cells and reprogramming events.

overall_design: Comparison of Dppa4 knockout ES cells (CI#243.12) against Dppa4 hemizygous parental ES cell line (CI#84.2). Both cell lines were grown in presence of mouse embrionic fibroblasts as feeder cells

first: Babita

last: Madan

first: Vikas

first: Odile

last: Weber

first: Philippe

last: Tropel

first: Carmen

last: Blum

first: Emmanuelle

last: Kieffer

first: Stephane

last: Viville

first: HansJoerg

last: Fehling


title: Expression data from retroviral vector-infected immortalized mouse embryonic fibroblasts (MEFs)

city: Singapore

first: Seiichi

last: Mori

state: Singapore

address: Centre for Life Sciences #02-07, 28 Medical Drive

department: Cancer Science Institute Singapore

country: Singapore

institute: National University of Singapore

summary:
Cultured cancer cells exhibit substantial phenotypic heterogeneity when measured in a variety of ways such as sensitivity to drugs or the capacity to grow under various conditions.  Among these, the ability to exhibit anchorage-independent cell growth (colony forming capacity in semisolid media) has been considered to be fundamental in cancer biology because it has been connected with tumor cell aggressiveness in vivo such as tumorigenic and metastatic potentials, and also utilized as a marker for in vitro transformation.  Although multiple genetic factors for anchorage-independence have been identified, the molecular basis for this capacity is still largely unknown.  To investigate the molecular mechanisms underlying anchorage-independent cell growth, we have used genome-wide DNA microarray studies to develop an expression signature associated with this phenotype.  Using this signature, we identify a program of activated mitochondrial biogenesis associated with the phenotype of anchorage-independent growth and importantly, we demonstrate that this phenotype predicts potential for metastasis in primary breast and lung tumors.

Keywords: c-Myc or v-Src retroviral vector-infected immortalized mouse embryonic fibroblasts.

overall_design: Expression data of c-Myc and v-Src transformed MEFs was used to validate an expression signature generated from human cultured breast cancer cell lines with anchorage-independent growth ability.

first: Jeffrey

last: Chang

first: Joseph

last: Nevins


title: ligand-activated and -unactivated AHR in wiltype and mutant hepatoma cells

city: Ann Arbor

first: Maureen

last: Sartor

state: MI

address: 2017 Palmer Commons-100 Washtenaw Ave.

institute: University of Michigan

summary: Starting with our early global expression analyses of TCDD-treated human hepatoma cells {Puga, 2000 4679 /id}, the AHR transcriptional induction profile has been extensively studied, whether activated by TCDD, B[a]P or in the absence of exogenous ligands (reviewed in {Frericks, 2007 5618 /id}).  In addition to using prior knowledge to integrate expression profiles into the AHR gene target network, we performed a new set of expression profile analyses of wild type Hepa-1c1c7 and c35 cell lines and compared the responses in naïve cells with responses in TCDD or B[a]P exposed cells for 8 hours.  Results of our expression array studies are in close agreement with current knowledge.

overall_design: Three biological replicates were each performed for wildype naïve cells  (DMSO), TCDD activated cells, and B[a]P activated cells, and for c35 (AHR mutant) naïve cells (DMSO), TCDD activated cells, and B[a]P activated cells.

last: Schnekenburger

first: Jennifer

last: Marlowe

first: John

middle: F

last: Reichard

first: Ying

last: Wang

first: Yunxia

last: Fan

first: Ci

last: Ma

first: Saikumar

last: Karyala

first: Danielle

last: Hableib

first: Xiangdong

last: Liu

first: Mario

last: Medvedovic

first: Alvaro

last: Puga


title: Hepatic gene expression profile of mice exposed to social stress

city: Aomori

first: Yuji

last: Nakai

state: Aomori

address: 2-1-1, Yanagawa

department: Institute of Regional Innovation

country: Japan

institute: Hirosaki University

summary: Social stress is well known to be involved in the occurrence and exacerbation of mental illness, and also various life-style related diseases such as hyperinsulinemia, hyperglycemia, cardiovascular diseases and cancer. However, there is little information on tissue-specific gene expression in response to social stress, which reflects our daily life. Liver is one of the most important organs, owing to its biological functions such as energy metabolic homeostasis, metabolization and detoxification of endo- and exogenous substances. In order to elucidate the mechanism underlying response to social stress in the liver, we investigated hepatic gene expression in mice exposed to isolation stress using DNA microarray. Male BALB/c mice (4 weeks old) were housed 5 per cage for 10 days acclimatization. Then mice were exposed to isolation stress for 30 days. After stress treatment, the mouse liver RNA was subjected to DNA microarray analysis. Taking the false discovery rate into account, isolation stress altered expression of 420 genes. Moreover, Gene Ontology analysis of these differentially expressed genes indicated that isolation stress remarkably down-regulated lipid metabolism-related pathway through peroxisome proliferator-activated receptor-alpha (PPARalpha), while lipid biosynthesis pathway regulated by sterol regulatory element binding factor-1 (SREBF-1), Golgi vesicle transport and secretory pathway-related genes were significantly up-regulated. These results suggested that isolation for 30 days, mild and consecutive social stress, not only regulate the systems for lipid metabolism but also cause the endoplasmic reticulum stress in mouse liver.

overall_design: Male BALB/c mice (4 weeks old, Japan SLC, Shizuoka, Japan) weighing 14-18 g were housed 5 per cage. After acclimatization for 10 days, the mice were exposed to isolation (1 mouse per cage). All cages were placed in a foam plastic box in order to avoid social contact. To enhance the feeling of isolation, the bed volume in each cage for the isolated mice was reduced to one-tenth of that in the control group. The weight of bedding chips was about 2 g. All mice were housed in an air-conditioned room ( room temperature: 23 ± 1°C, humidity: 55 ± 5 %) under 12 h dark/12 h light cycles, with free access to tap water and MF diet (Oriental Yeast Co., Tokyo, Japan).

first: Keiko

last: Motoyama

first: Tomoya

last: Miyashita

first: Yuichiro

last: Fukui

first: Maki

last: Morita

first: Kazutsuka

last: Sanmiya

first: Hiroyuki

last: Sakakibara

first: Ichiro

last: Matsumoto

last: Abe

first: Takashi

last: Yagabe

first: Nobuhiro

last: Yajima

first: Kayoko

last: Shimoi


title: Expression profile of Sall4-null ES cells and Sall4 heterozygous ES cells

status: Public on Apr 01 2009

city: Kumamoto

first: Ryuichi

last: Nishinakamura

address: 2-2-1 Honjo

department: Institute of Molecular Embryology and Genetics

institute: Kumamoto University

summary:
Sall4 is a mouse homolog of a causative gene of the autosomal dominant disorder known as Okihiro syndrome. We previously showed that Sall4 absence leads to lethality during peri-implantation and that Sall4-null embryonic stem (ES) cells proliferate poorly with intact pluripotency when cultured on feeder cells. However, a subsequent report indicated that shRNA-mediated Sall4 inhibition in ES cells led to a severe reduction in Oct3/4 and a secondary increase in Cdx2, which resulted in complete differentiation into the trophectoderm when cultured in the feeder-free condition. So we profiled gene expression changes when Sall4 is deleted in ES cells in the presence or absence of feeder cells.

key word: embryonic stem (ES) cell, Sall4, feeder

overall_design:
ES cells were cultured with or without mouse embryonic fibroblast (MEF) feeder cells in LIF-supplemented medium as described. To maintain the expression of Oct3/4, all ES cells were cultured in the presence of Blasticidin.

Four samples were analyzed.

GSM356329, GSM356330 : cultured in the absence of feeders
GSM356331, GSM356332 : cultured on the feeders


title: Molecular Signatures of Fetal and Adult Prostate Stem Cells

first: Roy

last: Blum

address: 552 First Ave

department: Cancer Institute

institute: New York University

summary: The global gene expression profiles of adult and fetal murine prostate stem cells were determined to define common and unique regulators whose misexpression might play a role in the development of prostate cancer. A distinctive core of transcriptional regulators common to both fetal and adult primitive prostate cells was identified as well as molecules that are exclusive to each population. Elements common to fetal and adult prostate stem cells include expression profiles of Wnt, Shh and other pathways identified in stem cells of other organs, signatures of the aryl-hydrocarbon receptor, and up-regulation of components of the aldehyde dehydrogenase/retinoic acid receptor axis. There is also a significant lipid metabolism signature, marked by overexpression of lipid metabolizing enzymes and the presence of the binding motif for Srebp1. The fetal stem cell population, characterized by more rapid proliferation and self-renewal, expresses regulators of the cell cycle, such as E2f, Nfy, Tead2 and Ap2, at elevated levels, while adult stem cells show a signature in which TGF-β has a prominent role. Finally, comparison of the signatures of primitive prostate cells with previously described profiles of human prostate tumors identified stem cell molecules and pathways with deregulated expression in prostate tumors including chromatin modifiers and the oncogene, Erg.  Our data indicate that adult prostate stem or progenitor cells may acquire characteristics of self-renewing primitive fetal prostate cells during oncogenesis and suggest that aberrant activation of components of prostate stem cell pathways may contribute to the development of prostate tumors.

overall_design: In order to identify molecules and pathways that are active in primitive prostate populations we determined the transcriptional profiles of four populations of cells: (i) fetal urogenital sinus epithelial cells, enriched in fetal prostate stem cells, (ii) Sca-1Hi, cells that express high levels of Sca-1, enriched in adult prostate stem cells, (iii) Sca-1Lo, cells that express medium to low levels of Sca-1 and are enriched in transit-amplifying cells, and (iv) Sca-1Neg, cells with no Sca expression, that represent the most mature population and have almost no in vivo regenerative potential.

first: Rashmi

last: Gupta

first: Patricia

middle: E

last: Burger

first: Christopher

last: Ontiveros

first: Sarah

last: Slam

first: Xiaozhong

last: Xiong

first: Alexander

last: Kamb

first: Holger

last: Wesche

first: Lisa

last: Marshall

first: Gene

last: Cutler

first: Sean

last: Wang

first: Jiri

last: Zavadil

first: David

last: Moscatelli

first: E

last: Wilson


title: Gene Expression Profiling of Cellular Dose-Response to Hypochlorous Acid (HOCl)

city: Research Triangle Park

first: Courtney

last: Woods

address: PO Box 12137, 6 Davis Drive

department: Computational Biology

institute: The Hamner Institutes

summary:
  Hypochlorous acid (HOCl) is a potent oxidant that is produced endogenously in mammalian tissue by phagocytes. Exogenous exposure to HOCl also can occur following inhalation of chlorine gas. HOCl has been implicated as a source of oxidative stress associated atherosclerosis and other diseases.
  The purpose of this study was to identify dose-dependent transitions in cellular response to hypochlorous acid (HOCl), with a  focus on understanding how various cellular defense and stress dose-responses overlap.

overall_design:
RAW 264.7 cells were grown to 80-90% confluency in DMEM supplemented with supplemented with 10% fetal bovine serum (FBS), 10 mM HEPES, 100 U penicillin/ml, and 100 µg streptomycin/ml. Cells were treated with 0, 0.14, 0.35, 0.7, 1.4, 2.1, 2.8 or 3.5 mM HOCl in medium for 6 hr. Total RNA was isolated from cells with TRIzol  according to manufacturer's instructions and then subjected to cleanup using RNase-Free DNase Set and RNeasy Mini kit. The resultant DNA-free RNA was diluted in RNase-free H2O and quantified by Nanodrop  at 260 nm. The quality of RNA samples was confirmed using RNA Nano Chips with Agilent 2100 Bioanalyzer. RNA samples were stored at –70 oC until use.
 From 5 micrograms of total RNA, cDNA was synthesized using a one-cycle cDNA synthesis kit (Affymetrix Corp., Santa Clara, CA). cDNA was transcribed to cRNA which was then biotin-labeled using GeneChip IVT labeling kit. Fifteen micrograms of labeled cRNA were then hybridized to an Affymetrix Mouse Genome 430 2.0 Array at 45°C for 16 hr.  Biological cRNA replicates (n = 3) were each hybridized to an individual array. After being washed using the GeneChip Fluidics Station 450, arrays were scanned using a GeneChip 3000 scanner and intensity values were extracted from the CEL file using Array Assist software (Stratagene, La Jolla, CA).

first: Jingqi

last: Fu

first: Peng

last: Xue

first: Yongyong

last: Hou

first: Linda

middle: J

last: Pluta

first: Longlong

last: Yang

first: Qiang

last: Zhang

first: Russell

last: Thomas

first: Melvin

last: Andersen

first: Jingbo

last: Pi


title: Gene expression profiles of E13 bladder neck/urethral compartments. (GUDMAP Series ID: 25)

first: GUDMAP

last: Developers

institute: IGMM MRC Human Genetics Unit

summary: The long term objective is to create an encyclopedia of the expression levels of all genes in multiple components of the developing bladder.  The central thesis is straightforward. The combination of micro dissected tissues and FACS sorted cells plus microarray analysis offers a powerful, efficient and effective method for the creation of a global gene expression atlas of the developing urogenital system. Microarrays with essentially complete genome coverage can be used to quantitate expression levels of every gene. The ensuing rapid read-out provides an expression atlas that is more sensitive, more economical and more complete than would be possible by in situ hybridizations alone. The data submitted here delineates the gene expression profiles of the mesenchymal and epithelial compartments of the E13 mouse bladder neck/urethral compartment.

overall_design: FVB/N mice were time mated. At embryonic day 13 mice were euthanized by decapitation and the bladders were microdissected and cut just below the ureters. Since the EDTA dissociation was ineffective, the bladder neck and urethra were collected and treated with 1 mg/ml trypsin in Tyrode's solution for 20 minutes at 37°C. The layers were separated using a fine needle and rimming. The samples were placed in RLT and stored at -80°C. RNA was prepared and the Epicentre 2-round amplification scheme described under the Lessard Group Protocols on the GUDMAP pages were performed. The amplified RNA was examined with the Affymetrix GeneChip Mouse Genome 430 2.0 Array.

first: James

last: Lessard

first: Alfor

last: Lewis


title: Induction of lipid oxidation gene expression by polyunsaturated fatty acids of marine origin in small intestine of mice

city: Wageningen

first: Guido

last: Hooiveld

address: HELIX, Stippeneng 4

department: Div. Human Nutrition & Health

institute: Wageningen University

summary: Dietary polyunsaturated fatty acids (PUFA) act as potent natural hypolipidemics and are linked to many health benefits in humans and in animal models. Mice fed long-term a high fat diet, in which medium-chain alpha linoleic acid (ALA) was partially replaced by long-chain docosahexaenoic (DHA) and eicosapentaenoic (EPA) fatty acids, showed reduced accumulation of body fat and prevention of insulin resistance, besides increased mitochondrial beta-oxidation in white adipose tissue and decreased plasma lipids. ALA, EPA and DHA all belong to PUFA of n-3 series. The intestine is a gatekeeper organ for ingested lipids. To examine the potential contribution of the intestine in the beneficial effects of EPA and DHA, this study assessed gene expression changes using whole genome microarray analysis on small intestinal scrapings. The main biological process affected was lipid metabolism. Fatty acid uptake, peroxisomal and mitochondrial beta-oxidation, and omega-oxidation of fatty acids were all increased. Quantitative real time PCR and intestinal fatty acid oxidation measurements ([14C(U)]-palmitate) confirmed significant gene expression differences in a dose-dependent manner. Furthermore, no major changes in the expression of lipid metabolism genes were observed in colonic scrapings. In conclusion, we show that marine n-3 fatty acids regulate small intestinal gene expression patterns. Since this organ contributes significantly to whole organism energy use, this adaptation of the small intestine may contribute to the complex and observed beneficial physiological effects of these natural compounds under conditions that will normally lead to development of obesity and diabetes.

overall_design: Male 4-month-old C57BL/6J mice were maintained for 4 weeks on semisynthetic high-fat (20% wt/wt) diets differing in the composition of n-3 PUFA. Two isocaloric diets were used (n=12): control sHFf diet which contained flax-seed oil (rich in ALA) as the only lipid source, or the sHFf-F2 diet, which had the same composition except that 44 % of the lipids were replaced by a n-3 PUFA concentrate (EPA&DHA) containing 6 % EPA and 51 % DHA (EPAX 1050TG; EPAX AS, Lysaker, Norway). Quality control of RNA samples showed that four samples did not pass quality thresholds, and these samples were excluded from array analysis (two control small intestine samples and one EPA&DHA small intestine sample). The remaining RNA samples were pooled per tissue per diet group. Thus; the flax-seed (control) array was hybridized with RNA pooled from of 10 mice, the PUFA array was hybridized with RNA pooled from 11 mice.

first: Evert

last: van Schothorst

first: Nicole

last: Franssen-van Hal

first: Jaap

last: Keijer


title: Comparisons of epithelial and mesenchymal murine breast tumor cell lines

city: Rochester

first: Keith

last: Knutson

state: MN

address: 200 First Street SW

department: Immunology

institute: Mayo Clinic

summary:
Epithelial  tumor cells (E) underwent EMT in vivo in FVB/N mice generating mesenchymal tumors. Mesenchymal cell lines (M1-M4) were each derived from a different mouse. This study compares gene expression between these two different tumor types.

Keywords: cell type comparison

overall_design: Two replicates were prepared for each of five cell lines, one epithelial cell line (E) and 4 mesenchymal cell lines M1-M4.

first: Michael

middle: K

last: Asiedu

first: Jennifer

last: Reiman


title: Gene expression profiles of heterozygous periphilin knockout mice compared to wt littermates

last: Bonin

state: BW

address: CAlwerstr. 7

department: Medical Genetics

institute: University Tuebingen

summary: Periphilin is a protein which is involved in multiple processes in vivo, including terminal differentiation of keratinozytes as well as cell-cycle and cancer related functions. Here we generated mice with a targeted disruption of the periphilin-1 gene to explore its physiological role from an organismic perspective. In accordance with a ubiquitous expression of periphilin in the murine embryo, the homozygous deficiency of periphilin is lethal in early embryogenesis. We therefore characterized mice with a disruption of only one periphilin allele. As heterozygous periphilin knockout mice show no obvious histological alterations and have no apparent behavioural phenotypes, we compared whole transcriptome RNA expression profiles of total brain tissue of periphilin+/- knockout mice and wild type littermates for an in-depth analysis of heterozygous animals. In periphilin+/- knockout mice, 3 probe sets were indicative of significant differential expression (p ≤ 0.05, SLR ≥ 1 which is equivalent to 2 fold). Two of these probe sets (1438864_at and 1439170_at) refer to unannotated or unclassifiable genes. According to the NetAffx™ Analysis Center, the third probe set (1447831_s_at) is assigned to myotubularin related protein 7 (Mtmr7). The probe set, however, lies upstream of the Mtmr7 gene (Ensembl gene ENSMUSG00000039431), and its signal is therefore not representative for the Mtmr7 gene. Quantitative RT-PCR (qRT-PCR) analysis with a primer set specific for Mtmr7 confirmed that its expression is not altered in periphilin+/- knockout mice (relative expression compared to wild type: 0.924; p = 0.419). In summary, none of more than 45.000 covered transcripts is differentially expressed in periphilin+/- knockout mice in comparison to wild type littermates.

overall_design: Whole brains of three male 2-months-old periphilin+/-knockout mice and three C57BL/6-wild type littermates

first: Anne

last: Söhn

first: Thu-Trang

last: Pham

first: Karin

last: Schaeferhoff

first: Thomas

last: Floss

first: Daniela

last: Vogt Weisenhorn

first: Wofgang

last: Wurst

first: Olaf

last: Riess


title: Gene Expression Profiling of the MLL-AF4 and Flt3 tyrosine kinase domain (TKD) genes in 32Dc cell

status: Public on Mar 31 2009

city: Tokyo

first: Hiroki

last: Yamaguchi

address: 1-1-5 Sendagi, Bunkyo-Ku

department: Division of Hematology, Department of Internal Medicine

institute: Nippon Medical School

summary: The tumorigenesis capacity of MLL-AF4 alone is insufficient for causing leukemia. Based on the finding that an Flt3 gene mutation in the tyrosine kinase domain (TKD) was observed in approximately 15% of MLL leukemia, we investigated synergistic leukemogenesis effects of the two genes in vitro. In a mouse IL3-dependent cell line, 32Dc, the expression of MLL-AF4 and Flt3 TKD was induced using a lentiviral vector. We performed gene expression profiling in the MLL-AF4 and the Flt3 TKD+MLL-AF4 expressing 32Dc cells. The enhancement of Hox genes expression was not identified. However, instead, the expression of S100A6, which was involved in the control of cell proliferation, was synergistically enhanced in the presence of both MLL-AF4 and Flt3 TKD genes.

overall_design: We performed gene expression profiling: 32Dc vs. MLL-AF4 expressing 32Dc, 32Dc vs. Flt3 TKD+MLL-AF4 expressing 32Dc, and MLL-AF4 expressing 32Dc vs. Flt3 TKD+MLL-AF4 expressing 32Dc. A single sample for each expressing cells was analyzed.

first: K

last: Dan

last: Inokuchi


title: US28-expressing and mock-transfected stable NIH-3T3 cell lines

status: Public on Mar 30 2009

city: Amsterdam

first: David

last: Maussang

address: De Boelelaan 1083

department: Division of Medicinal Chemistry, Faculty of Sciences

institute: Vrije Universiteit Amsterdam, Leiden/Amsterdam Center for Drug Research

summary:
The human cytomegalovirus (HCMV) encodes the chemokine receptor US28 that exhibits constitutive activity. NIH-3T3 cells stably transfected with US28 present a pro-angiogenic and transformed phenotype both in vitro and in vivo.
We used gene expression profiling to determine how US28 constitutive activity modulates expression of genes compared to mock-transfected cells.

Keywords: Comparison of stably transfected cell lines

overall_design: We isolated RNA from two different clonal stable cell lines of NIH-3T3 cells transfected either either mock or US28-WT and analysed these 4 RNA samples using Affymetrix mouse genome arrays.

first: Ellen

last: Langemeijer

first: Carlos

middle: P

last: Fitzsimons

first: Marijke

last: Stigter-van Walsum

first: Remco

last: Dijkman

first: Martin

last: Borg

first: Erik

last: Slinger

first: Andreas

last: Schreiber

first: Detlef

last: Michel

first: Cornelis

last: Tensen

first: Guus

middle: A

last: van Dongen

first: Rob

last: Leurs

first: Martine

last: Smit