title: Dynamic patterning at the pylorus: formation of an epithelial intestine-stomach boundary in late fetal life

city: Ann Arbor

last: Li

state: MI

address: 109 Zina Pitcher Place,045 BSRB

department: Bioinformatics Program & CDB

institute: Universtiy of Michigan

summary: In the adult mouse, distinct morphological and transcriptional differences separate stomach from intestinal epithelium. Remarkably, the epithelial boundary between these two organs is literally one cell thick. This discrete junction is established suddenly and precisely at embryonic day (E) 16.5, by sharpening a previously diffuse intermediate zone. In the present study, we define the dynamic transcriptome of stomach, pylorus and intestinal tissues between E14.5 and E16.5. We show that establishment of this boundary is concomitant with the induction of over a thousand genes in intestinal epithelium, and these gene products provide intestinal character. Hence, we call this process intestinalization. We identify specific transcription factors (Hnf4g, Creb3l3 and Tcfec) and examine signaling pathways (Hedgehog and Wnt) that may play a role in this process. Finally, we define a unique expression domain at the pylorus itself and detect novel pylorus-specific patterns for the transcription factor Gata3 and the secreted protein nephrocan.

overall_design: Stomach, pylorus and duodenum tissue from E14.5 and E16.5 mouse embryos were collected for RNA extraction and hybridization on Affymetrix microarrays. We sought to study the gene expression profiles and identify genes and pathways enriched in these three tissues at two important developmental times.

first: Aaron

last: Udager

first: Chunbo

last: Hu

first: Tracy

last: Qiao

first: Neil

last: Richards

first: William

last: Zacharias

first: Deborah

middle: L

last: Gumucio


title: Expression data from NIH-3T3 cells used for half-life determination

status: Public on Sep 30 2009

city: Cambridge

first: Lars

last: Dölken

state: Cambridgeshire

address: Addenbrookes Hospital, Hills Road

department: Medicine

country: United Kingdom

institute: University of Cambridge

summary:
Data from tc-, nt- and p-RNA as well as 1 and 2h of actinomycin-D treatment (5µg/ml) of NIH-3T3 cells used to determine half-lives. RNA was labeled for 15, 30 or 60 minutes with 4-thiouridine. After preparation of tc-RNA, thiol-labeled RNA was biotinylated using biot-HPDP and subsequently tc-RNA was separated into nt- and p-RNA using streptavidin coated magnetic beads. All three fractions were used for microarray analysis. For actinomycin-D experiments only tc-RNA was used prepared from cell before and 1 an 2h after addition of act-D.
We used microarrays to analyze the effects of 1 and 3h of IFNalpha and gamma treatment in total cellular RNA
Keywords: determination of RNA half-lives in NIH-3T3

overall_design: NIH-3T3 cells ( 5th to 15th passage after thawing) were split from confluent plates 24h before start of the experiment. At the begin of the experiment about 80% confluency was reached. The experiment was started by applying fresh, prewarmed, CO2-equilibrated medium containing either 200µM 4sU (for 1h labeling), 500µM 4sU (for <=30min labeling) or 5µg/ml actinomycin-D.

first: Zsolt

last: Ruzsics

first: U

middle: H

last: Koszinowski


title: Expression data from NIH-3T3 cells treated with mock, 100 U/ml IFN alpha or 100 U/ml gamma for 1or 3h

summary:
Differential gene expression caused by 1h and 3h of IFN alpha or gamma treatment was analyzed in total cellular RNA of NIH-3T3 cells compared to mock
We used microarrays to analyze the effects of 1 and 3h of IFNalpha and gamma treatment in total cellular RNA
Keywords: time course in total cellular RNA (tc-RNA)

overall_design: NIH-3T3 cells ( 5th to 15th passage after thawing) were split from confluent plates 24h before start of the experiment. At the begin of the experiment about 80% confluency was reached. The experiment was started by applying fresh, prewarmed, CO2-equilibrated medium containing either mock, 100U/ml IFN alpha or 100 U/ml IFN gamma. Labeling was either started together with addition of IFN for 60 minutes or for 30 minutes starting 150 minutes after begin of treatment.


title: newly transcribed RNA (nt-RNA) for IFN alpha and gamma time course

summary:
Expression data from NIH-3T3 cells treated with mock, 100 U/ml IFN alpha or 100 U/ml gamma for 1 or 3h on nt-RNA labeled for 30-60 min at different times of interferon treatment
Differential gene expression caused by IFN alpha or gamma was analyzed in newly transcribed RNA (nt-RNA) of NIH-3T3 cells treated for 1 or 3 h. RNA was labeled for 30 to 60 min and separated from total cellular RNA (tc-RNA) following Trizol RNA preparation and thiol-specific biotinylation
We used microarrays to analyze the effects of IFNalpha and gamma treatment in newly transcribed RNA (nt-RNA)
Keywords: time course

overall_design: NIH-3T3 cells (5th to 15th passage after thawing) were split 24 h before start of the experiment. When starting the experiment 80% confluency was reached. The experiment was started by applying fresh, prewarmed, CO2-equilibrated medium containing either mock, 100U/ml IFN alpha or 100 U/ml IFN gamma. Labeling was either started simultaneously, 30 or 150 minutes later.


title: Interleukin-7 promotes monocyte/macrophage arrest on endothelial cells

status: Public on Oct 01 2009

city: Houston

first: Kazuhiro

last: Oka

state: TX

address: One Baylor Plaza

department: Molecular & Cellular Biology

country: USA

institute: Baylor College of Medicine

summary:
Background: It is recognized that atherosclerosis can regresses at least in animal models. However, little is known about the mechanisms. We induced regression of advanced atherosclerosis in apolipoprotein E deficient (APOE/) mice and studied underlying mechanisms. Unexpectedly, our study led to the role of interleukin-7 (IL-7) in atherogenesis.
Methods and Results: We treated APOE/ mice fed a high cholesterol diet for 30 weeks to induce advanced lesions with a helper-dependent adenoviral vector expressing human apoE3 (HDAd-gE3), and analyzed the regression of atherosclerosis after 41 weeks. Using microarray analysis, we identified IL-7 as one of most significantly affected genes by lowering cholesterol. To answer why IL-7 is downregulated by reduced cholesterol, we studied effects of IL-7 on endothelial cells (ECs). Our major findings were (1) long-term lowering cholesterol induced regression of advanced atherosclerosis. (2) Microarray analysis identified multiple signaling pathways affected by lowering cholesterol. (3) Correction for multiple testing revealed that IL-7 expression was downregulated, whereas gamma-sarcoglycan and Î±-actin were upregulated. (4) Oxidized LDL upregulated IL-7 expression in macrophages but not in aorta ECs or smooth muscle cells. (5) IL-7 increased the expression of cell adhesion molecules and chemokine in ECs and promoted monocyte adhesion to ECs. (6) Systemic elevation of IL-7 induced inflammatory response and recruited monocyte/macrophage to the lesions without increasing plasma cholesterol.
Conclusion: Our finding suggest that IL-7 inflames endothelium and triggers the adhesion/recruitment of monocyte/macrophages to the atherosclerotic lesions and thus plays a direct role in development of atherosclerosis.

Key Words: arteriosclerosis, gene therapy, hypercholesterolemia, interleukins, cell adhesion molecules

overall_design:
Female APOE-/- mice (8 weeks of age) on a C57BL/6 background were purchased from Jackson Laboratory and were fed a diet containing 0.2% (w/w) cholesterol and 10% (v/w) coconut oil to induce atherosclerosis for 30 weeks. Mice were then divided into 3 groups,  (n=15/group), treated with a tail vein injection of helper-dependent adenovirus expressing human apolipoprotein E3 (HDAd-gE3, 5 x 1012 viral particles/kg, n=11, vector), empty vector (HDAd-0, 5 x 1012 VP/kg, n=11, vector control) or phosphate buffered saline (PBS, vehicle) and sacrificed 10 days after. Three aortas extending sinus to arch were pooled for RNA extraction with an RNeasy kit (Qiagen).

Sample ID
12031: E4: vector control (HDAd-0)
12043: E5: vector control
12046: V2: vector
12047: V3: vector
12048: V4: vector
12051: B1: vehicle (PBS)
12053: B3: vehicle
12054: B4: vehicle
12039: E3: vector control

first: Rongying

first: Antonio

last: Paul

first: Kerry

last: Ko

first: Michael

last: Sheldon

first: Carrie

last: Decker

first: Shelly

last: Cormier

first: Lan

first: Elie

middle: A

last: Nour

first: Lawrence

last: Chan


title: Comparative analysis of gene regulation by the transcription factor PPARα_mouse

status: Public on Sep 21 2009

city: Wageningen

first: Guido

last: Hooiveld

address: HELIX, Stippeneng 4

department: Div. Human Nutrition & Health

country: Netherlands

institute: Wageningen University

summary:
Studies in mice have shown that PPARα is an important regulator of hepatic lipid metabolism and the acute phase response. However, little information is available on the role of PPARα in human liver. Here we set out to compare the function of PPARα in mouse and human hepatocytes via analysis of target gene regulation. Primary hepatocytes from 6 human and 6 mouse donors were treated with PPARα agonist Wy14643 and gene expression profiling was performed using Affymetrix GeneChips followed by a systems biology analysis. Baseline PPARα expression was similar in human and mouse hepatocytes. Depending on  species and time of exposure, Wy14643 significantly induced the expression of 362-672 genes. Surprisingly minor overlap was observed between the Wy14643-regulated genes from mouse and human, although more substantial overlap was observed at the pathway level. Xenobiotics metabolism and apolipoprotein synthesis were specifically regulated by PPARα in human hepatocytes, whereas glycolysis-gluconeogenesis was regulated specifically in mouse hepatocytes. Most of the genes commonly regulated in mouse and human were involved in lipid metabolism and many represented known PPARα targets, including CPT1A, HMGCS2, FABP, ACSL, and ADFP. Several genes were identified that were specifically induced by PPARα in human (MBL2, ALAS1, CYP1A1, TSKU) or mouse (Fbp2, lgals4, Cd36, Ucp2, Pxmp4). Furthermore, several putative novel PPARα targets were identified that were commonly regulated in both species, including CREB3L3, KLF10, KLF11 and MAP3K8. Our results suggest that PPARα activation has a major impact on gene regulation in human hepatocytes. Importantly, the role of PPARα as master regulator of hepatic lipid metabolism is generally well-conserved between mouse and human. Overall, however, PPARα regulates a mostly divergent set of genes in mouse and human hepatocytes.

Keywords: Analysis of target gene regulation by using microarrays

overall_design: Primary hepatocytes from 6 mice from 6 different strains were treated with the PPARα agonist Wy14643 for 6 and 24 hours, and gene expression profiling was performed using Affymetrix GeneChips.  Mouse primary hepatocytes were isolated by two-step collagenase perfusion from 6 different strains of mouse; NMRI, SV129, FVB, DBA, BALB/C and C57BL/6J.   Cells were plated on collagen-coated six-well plates. Viability was determined by Trypan Blue exclusion, and was at least 75%. Hepatocytes were suspended in William's E medium (Lonza Bioscience, Verviers, Belgium) supplemented with 10% (v/v) fetal calf serum, 20 m-units/mL insulin, 10 nM dexamethasone, 100 U/mL penicillin, 100 μg/mL of streptomycin, 0.25 μg/mL fungizone and 50 μg/mL gentamycin. After four hours the medium was discarded and replaced with fresh medium. The next day, cells were incubated in fresh medium in the presence or absence of Wy14643 (10 microM) dissolved in DMSO for 6 and 24 hours, followed by RNA isolation. A 5-fold lower concentration of Wy14643 was used in mouse primary hepatocytes to take into account the higher affinity of Wy14643 for mouse PPARα compared to human PPARα.

first: Maryam

last: Rakhshandehroo

last: Müller

first: Sander

last: Kersten


title: Genome wide mRNA expression correlates of viral control in CD4+T cells from HIV-1 infected individuals

city: Lausanne

first: Margalida

last: Rotger

address: Bugnon 48

department: University of Lausanne

country: Switzerland

institute: Institute of Microbiology

summary: There is great interindividual variability in HIV-1 viral set point after seroconversion, some of which is known to be due to genetic differences among infected individuals. Here, our focus is on determining, genome-wide, the contribution of variable gene expression to viral control, and to relate it to genomic DNA polymorphism. RNA was extracted from purified CD4+ T-cells from 137 HIV-1 seroconverters, 16 elite controllers, and 3 healthy blood donors. Expression levels of more than 48,000 mRNA transcripts were assessed by the Human-6 v3 Expression BeadChips (Illumina). Genome-wide SNP data was generated from genomic DNA using the HumanHap550 Genotyping BeadChip (Illumina). We observed two distinct profiles with 260 genes differentially expressed depending on HIV-1 viral load. There was significant upregulation of expression of interferon stimulated genes with increasing viral load, including genes of the intrinsic antiretroviral defense. Upon successful antiretroviral treatment, the transcriptome profile of previously viremic individuals reverted to a pattern comparable to that of elite controllers and of uninfected individuals. Genome-wide evaluation of cis-acting SNPs identified genetic variants modulating expression of 190 genes. Those were compared to the genes whose expression was found associated with viral load: expression of one interferon stimulated gene, OAS1, was found to be regulated by a SNP (rs3177979, p=4.9E-12); however, we could not detect an independent association of the SNP with viral set point. Thus, this study represents the first attempt to integrate genome-wide SNP signals with genome-wide expression profiles in the search for biological correlates of HIV-1 control. It underscores the paradox of the association between increasing levels of viral load and greater expression of antiviral defense pathways. It also shows that elite controllers do not have a distinctive mRNA expression pattern in CD4+ T cells. Changes in RNA expression reflect responses to viral replication rather than a mechanism that might explain viral control

overall_design: Total RNA from 202 samples obtained from CD4 T cells from HIV infected individuals to identify associations between variation in gene expression viral set point. Samples belong to different groups as follows (Paired Untreated/treated samples: 38/38, only untreated samples: 89, successfully treated samples: 29. Eight samples from 3 healthy blood donors in replicates.

first: Kristen

middle: K

last: Dang

first: Jacques

last: Fellay

first: E

last: Heinzen

first: Sheng

last: Feng

first: Patrick

last: Descombes

first: Kevin

middle: V

last: Shianna

first: G

last: Dongliang

first: Huldrich

last: Günthard

first: David

middle: B

last: Goldstein

first: Amalio

last: Telenti


title: Microarray analysis of Cbfb-deficient regulatory T cells

status: Public on Sep 19 2009

city: Kyoto

first: Akihiko

last: Kitoh

address: 53 Shogoin Kawahara-cho

department: Institute for Frontier Medical Sciences

country: Japan

institute: Kyoto University

summary:
Gene expression profiles of Cbfb-deficient and control Treg cells were compared.

Abstract: Naturally arising regulatory T (Treg) cells express the transcription factor FoxP3, which critically controls the development and function of Treg cells. FoxP3 interacts with another transcription factor Runx1 (also known as AML1). Here we showed that Treg cell-specific deficiency of Cbfβ, a cofactor for all Runx proteins, or that of Runx1, but not Runx3, induced lymphoproliferation, autoimmune disease, and hyper-production of IgE. Cbfb-deleted Treg cells exhibited impaired suppressive function in vitro and in vivo, with altered gene expression profiles including attenuated expression of FoxP3 and high expression of interleukin-4. The Runx complex bound to more than 3000 gene loci in Treg cells, including the Foxp3 regulatory regions and the Il4 silencer. In addition, knockdown of RUNX1 showed that RUNX1 is required for the optimal regulation of FoxP3 expression in human T cells. Taken together, our results indicate that the Runx1-Cbfβ heterodimer is indispensable for in vivo Treg cell function, in particular, suppressive activity and optimal expression of FoxP3.

overall_design: CD4+CD25hi cells, most of which were Foxp3+ Treg cells, were isolated from Cbfb-flox/flox: Foxp3-ires-Cre (n = 3) and control Cbfb-flox/wt: Foxp3-ires-Cre (n = 3) mice. Total RNA was extracted from those purified Cbfb-deficient or control Treg cells.

first: Masahiro

last: Ono

first: Yoshinori

last: Naoe

first: Naganari

last: Ohkura

first: Tomoyuki

last: Yamaguchi

first: Hiroko

last: Yaguchi

first: Issay

last: Kitabayashi

first: Toshihiko

last: Tsukada

first: Takashi

last: Nomura

first: Yoshiki

last: Miyachi

first: Ichiro

last: Taniuchi

first: Shimon

last: Sakaguchi


title: Analysis of gene expression levels in BBF2H7-/- chondrocytes

status: Public on Sep 11 2009

city: Kiyotake, Miyazaki

first: Atsushi

last: Saito

address: Kihara, 5200

department: Anatomy

institute: Miyazaki university

summary: BBF2H7 (BBF2 human homolog on chromosome 7), an ER-resident basic leucine zipper transcription factor, is activated in response to ER stress and abundantly expresses in chondrocytes. While BBF2H7 is widely expressed in many tissues and organs, the most intense signals were detected in the proliferating zone of the cartilage. We compared gene expressions in primary cultured chondrocytes prepared from rib cartilage between WT and BBF2H7-/- mice at E18.5. Primary cultured chondrocytes were prepared from E18.5 rib cartilage of WT and BBF2H7-/- mice. Chondrocytes were isolated using 0.2% collagenase D (Roche) after adherent connective tissue was removed by 0.2% trypsin (Sigma) and collagenase pretreatment. Isolated chondrocytes were maintained in α-MEM (Gibco) supplemented with 10% FCS and 50 µg/mL ascorbic acid. Adenovirus vectors expressing the mouse p60 BBF2H7 (1-377 aa, BBF-N) were constructed with the AdenoX Expression system (Clontech), according to the manufacturer’s protocol. The cells were infected with adenoviruses 30 h before analysis.

overall_design: We compared gene expressions in primary cultured chondrocytes prepared from rib cartilage between WT and BBF2H7-/- mice at E18.5 using a microarray and various genes associated with protein secretory pathway and ER biogenesis were significantly down-regulated in BBF2H7-/- chondrocytes. We infected primary cultured chondrocytes prepared from BBF2H7-/- mice with adenovirus expressing p60 BBF2H7. Several genes were up-regulated and we picked up them as the direct target of BBF2H7.

first: Kazunori

last: Imaizumi


title: Gender-specific gene repression of PPAR-alpha KO mice in liver and heart

first: Sylvain

last: Pradervand

address: Genopode

department: CIG

institute: UNI Lausanne

summary:
Most metabolic studies are conducted in male animals; thus, the molecular mechanism controlling gender-specific pathways has been neglected, including sex-dependent responses to peroxisome proliferator-activated receptors (PPARs). Here we show that PPARalpha has broad female-dependent repressive actions on hepatic genes involved in steroid metabolism and inflammation. In males, this effect is reproduced by the administration of synthetic PPARalpha ligand. Using the steroid hydroxylase gene Cyp7b1 as a model, we elucidated the molecular mechanism of this PPARalpha-dependent repression. Initial sumoylation of the ligand-binding domain of PPARalpha triggers the interaction of PPARalpha with the GA-binding protein alpha bound to the target promoter. Histone deacetylase is then recruited, and histones and adjacent Sp1-binding site are methylated. These events result in the loss of Sp1-stimulated expression, and thus the down-regulation of Cyp7b1. Physiologically, this repression confers protection against estrogen-induced intrahepatic cholestasis, paving the way for a novel therapy against the most common hepatic disease during pregnancy.

Keywords: Genetic modification

overall_design: Expression profile difference between male and female PPARalpha wild-type and knock-out mice in liver and heart (3 pools of 4 animals in each group). Wild-type (12 males and 12 females) and knock-out PPARalpha SV129 mice (12 males and 12 females) approximately 10 to 12 weeks of age were killed at ZT14 and their livers and hearts quickly removed and frozen.

first: Nicolas

last: Leuenberger

first: Walter

last: Wahli


title: Gene expression and alternative splicing in human gastric cancer

status: Public on Sep 24 2009

city: shanghai

first: Zhang

last: Qinghua

address: 151# libing road

country: China

institute: Shanghai shbiochip co

summary:
27 sample pairs were conducted exon microarrays and 25 sample pairs were retain for our further alternative splicing analysis. 260 genes showed differential expression in more than 50% of our samples and 21 genes among them were validated by real-time PCR. Six genes with splicing events, which were firstly found in gastric cancer, were confirmed by RT-PCR.

Keywords: Transcriptome analysis

overall_design: Total 49 gastric caners and corresponding normal mucosas were choose to reexamine the pathological information and procure pure cells using laser capture microdissection. 25 sample pairs passed microarray quality control and were retained for further alternative splicing analysis. One sample per array.

first: Yanqing

last: Yang

first: Wen

last: Zhang

first: Hengjun

last: Gao

first: Qinghua


title: Microarray of RNA from calvaria in WT and OASIS-/- (KO) mice

status: Public on Sep 10 2009

city: Miyazaki

address: 5200 Kihara, Kiyotake

institute: Division of Molecular and Cellular Biology, Department of Anatomy, Faculty of Medicine, University of Miyazaki

summary:
Investigation of whole genome gene expression level changes in OASIS KO calvaria compared to wild-type calvaria.
To gain further insight into the potential mechanisms underlying the defective bone formation in OASIS KO mice, we compared the gene expression in calvaria between WT and OASIS KO mice using a microarray.

overall_design: Each sample of total RNA was collected from a number of  mice.

first: Imaizumi

last: Kazunori


title: The transcription factor AP2γ regulates the number of basal progenitors

status: Public on Sep 22 2009

city: Neuherberg

first: Johannes

last: Beckers

address: Ingolstaedter Landstr. 1

department: Institute of Experimental Genetics

country: Germany

institute: Helmholtz Zentrum Muenchen

summary:
Understanding the mechanisms that specify neuronal subtypes is important to unravel the complex mechanisms of neuronal circuit assembly. Here we have identified a novel role for the transcription factor AP2γ in progenitor and neuronal subtype specification in the cerebral cortex. Conditional deletion of AP2γ causes misspecification of basal progenitors starting at
mid-neurogenesis and gain-of-function experiments show that AP2γ directly regulates the expression of several genes characteristic for basal progenitors, such as Math3 and Tbr2. The misspecification of basal progenitors upon loss of AP2γ resulted in their increased death and
the ultimate reduction of callosal layer II/III projection neurons. This had pronounced effects on visual performance with a strong reduction of visual acuity. Thus, we have identified a novel regulator of basal progenitor fate regulating the number of layer II/III neurons in an area-specific manner and revealing their importance for accurate function of the visual cortex.

overall_design: We analysed 12 samples from the cortex (rostral and caudal) of AP2gamma k.o. mice and control mice. For each of the 4 groups (caudal ko, caudal wt, rostral ko, rostral wt) three biological replicates were analysed.

first: Luisa

last: Pinto

first: Daniela

last: Drechsel

first: Marie-Theres

last: Schmid

first: Jovica

last: Ninkovic

first: Martin

last: Irmler

first: Monika

last: Brill

first: Laura

last: Restani

last: Gianfranceschi

first: Victor

last: Tarabykin

first: Kristin

last: Baer

first: Francois

last: Guillemot

first: Colette

last: Dehay

first: Nada

last: Zecevic

first: Matteo

last: Caleo

first: Hubert

last: Schorle

first: Magdalena

last: Götz


title: Effects of irradiation on mouse lung

city: Montreal

first: Christina

middle: Kathleen

last: Haston

state: QC

address: 3626 St. Urbain

country: Canada

institute: McGill University

summary:
Total lung RNA from 3 mouse strains after 18Gy thoracic irradiation.
Thoracic cavity radiotherapy is limited by the development of alveolitis and fibrosis in susceptible patients.  To define the response to 18 Gy pulmonary irradiation in mice, at the expression level, and to identify pathways which may influence the alveolitis and fibrosis phenotypes expression profiling was completed.  Male mice of three strains, A/J (late alveolitis response), C3H/HeJ (C3H, early alveolitis response) and C57BL/6J (B6, fibrosis response) were exposed to thoracic radiation, euthanised when moribund and lung tissue gene expression was assessed with microarrays.

overall_design: treated vs. control in 3 strains


title: Expression of Wnt Receptors in Adult Spiral Ganglion Neurons: Fzd 9 Located at Growth Cones of Regenerating Neurites

status: Public on Sep 16 2009

city: Urbana

first: Richard

last: Kollmar

state: IL

address: 407 S. Goodwin Ave., MC-114

institute: University of Illinois at Urbana-Champaign

summary: The fidelity of sound transmission by cochlear implants in patients with sensorineural hearing loss could be greatly improved by increasing the number of frequency channels. This could be achieved by stimulating and guiding neurite outgrowth to reduce the distance between the implant's electrodes and the remnants of the spiral ganglion neurons. However, little is known about signaling pathways, besides those of neurotrophic factors, that are operational in the adult spiral ganglion. To systematically identify neuronal receptors for guidance cues in the adult cochlea, we conducted a genome-wide cDNA microarray screen with two-month-old CBA/CaJ mice. A meta-analysis of our data and those from older mice in two other studies revealed the presence of neuronal transmembrane receptors that represent all four established guidance pathways—ephrin, netrin, semaphorin, and slit—in the mature cochlea as late as 15 months. In addition, we observed the expression of all known receptors for the Wnt morphogens, whose neuronal guidance function has only recently been recognized. In situ hybridizations located the mRNAs of the Wnt receptors frizzled 1, 4, 6, 9, and 10 specifically in adult spiral ganglion neurons. Finally, frizzled 9 protein was found in the growth cones of adult spiral ganglion neurons that were regenerating neurites in culture. We conclude from our results that adult spiral ganglion neurons are poised to respond to neurite damage, owing to the constitutive expression of a large and diverse collection of guidance receptors. Wnt signaling, in particular, emerges as a candidate pathway for guiding neurite outgrowth towards a cochlear implant after sensorineural hearing loss.

overall_design:
To determine which potential guidance receptors were present in the adult cochlea and whether any of them were chronically up- or downregulated after damage, we conducted a microarray hybridization screen with modioli from three unexposed and noise-exposed mice each. The noise treatment was administered at four weeks of age, and the modioli were harvested at eight weeks of age. The pre-processed hybridization signals (GSM321743-8) were further analyzed with Bioconductor tools (release 2.1; www.bioconductor.org) in R software (version 2.6.1; http://www.r-project.org) according to the procedures outlined in Gentleman et al. (2005). See linked supplementary files for Bioconductor details and results of analysis:

GSE12810_BioconductorCode.txt: Step-by-step description of probe-set selection based on (i) hybridization signal and (ii) Gene Ontology annotation; (iii) of significance analysis of microarrays; and (iv) of meta-analysis of our data and those of Someya and colleagues

GSE12810_GO_Eset.xls: Significance analysis of microarrays  for neuronal transmembrane receptors to detect differential expression between noise-exposed and unexposed modioli

GSE12810_Meta_GO_Eset_tTests.xls: Meta-analysis of neuronal transmembrane receptors in our study (GSE12810) and two previous studies by Someya and colleagues (GSE4786 & GSE4866) to estimate absolute expression levels in adult cochlea

first: Samit

middle: M

last: Shah

first: Young-Jin

last: Kang

first: Barbara

last: Christensen

first: Albert

middle: S

last: Feng


title: Study of Differential Gene Expression in Metastatic Breast Tumors

status: Public on Sep 09 2009

city: Savannah

first: Nagendra

last: Ningaraj

state: GA

address: 4700 Waters Avenue

department: Anderson Cancer Institute

institute: Memorial University Medical Center

summary: Introduction: The prognosis for patients with breast tumor metastases to brain is extremely poor. Identification of prognostic molecular markers of the metastatic process is critical for designing therapeutic modalities for reducing the occurrence of metastasis. Although ubiquitously present in most human organs, calcium-activated potassium (BK) channel is significantly upregulated in breast cancer cells. In this study we investigated the role of KCNMA1 gene, which encodes α subunit of KCa channels (BK channels) in breast cancer metastasis and invasion.   Methods: We performed Global exon array to study the expression of KCNMA1 in metastatic breast cancer in brain, compared its expression in primary breast cancer and breast cancers metastatic to other organs, and validated the findings by RT-PCR.   Immunohistochemistry was performed to study the expression and localization of α subunit of KCa channel protein in primary and metastatic breast cancer tissues and breast cancer cell lines. We performed matrigel invasion, transendothelial migration and membrane potential assays in established lines of  normal breast cells (MCF-10A), non-metastatic breast cancer (MCF-7), non-brain metastatic breast cancer cells (MDA-MB-231), and brain-specific metastatic breast cancer cells (MDA-MB-361) to study whether KCa channel inhibition attenuates breast tumor invasion and metastasis using KCNMA1 knockdown with siRNA and biochemical inhibition with IBTX (Iberiotoxin).  Results: The Global exon array and RT-PCR showed higher KCNMA1 expression in metastatic breast cancer in brain compared to metastatic breast cancers in other organs. Our results clearly show that metastatic breast cancer cells exhibit increased BK channel activity, leading to greater invasiveness and transendothelial migration, both of which could be attenuated by blocking KCNMA1.  Conclusion: Determining the relative abundance of BK channel over expression in breast cancer metastatic to brain and the mechanism of its action in brain metastasis will provide a unique opportunity to identify and differentiate between low grade breast tumors that are at high risk for metastasis from those at low risk for metastasis. This distinction would in turn allow for the appropriate and efficient application of  effective  treatments while sparing patients  with low risk for metastasis from the toxic side effects of chemotherapy.

overall_design: Three cell lines representing primary breast tumor (MCF-7), metastatic breast tumors to other organs excluding brain (MDA-MB-231), and metastatic breast tumors to brain (MDA-MB 361) were run in triplicates to identify differentially expressed genes. Metastatic breast tumor cell lines were compared with primary breast tumor cell line with respect to their gene expression.

first: Umesh

middle: T

last: Sankpal

first: Divya

last: Khaitan


title: Cdcs1 a major colitis susceptibility locus in mice; subcongenic analysis reveals genetic complexity

city: Hannover

first: Andre

last: Bleich

institute: Institute for Laboratory Animal Science and Central Animal Facility, Hannover Medical School

summary: Background and Aims: In the interleukin-10-deficient (Il10-/-) mouse model of IBD, 10 quantitative trait loci (QTL) have been shown to be associated with colitis susceptibility by linkage analyses on experimental crosses of highly susceptible C3H/HeJBir (C3Bir)-Il10-/- and partially resistant C57BL/6J (B6)-Il10-/- mice. The strongest locus (C3Bir-derived cytokine deficiency-induced colitis susceptibility [Cdcs]1 on Chromosome [Chr] 3) controlled multiple colitogenic subphenotypes and contributed the vast majority to the phenotypic variance in cecum and colon. This was demonstrated by interval-specific Chr 3 congenic mice wherein defined regions of Cdcs1 from C3Bir or B6 were bred into the IL-10-deficient reciprocal background and altered the susceptible or resistant phenotype. Furthermore, this locus likely acts by inducing innate hypo- and adaptive hyperresponsiveness, associated with impaired NFΚB responses of macrophages. The aim of the present study was to dissect the complexity of Cdcs1 by further development and characterization of reciprocal Cdcs1 congenic strains and to identify potential candidate genes in the congenic interval. Material and Methods: In total, 15 reciprocal congenic strains were generated from Il10-/- mice of either C3H/HeJBir or C57BL/6J backgrounds by 10 cycles of backcrossing. Colitis activity was monitored by histological grading. Candidate genes were identified by fine mapping of congenic intervals, sequencing, microarray analysis and a high-throughput real-time RT-PCR approach using bone marrow-derived macrophages. Results: Within the originally identified Cdcs1-interval, three independent regions were detected that likely contain susceptibility-determining genetic factors (Cdcs1.1, Cdcs1.2, and Cdcs1.3). Combining results of candidate gene approaches revealed Fcgr1, Cnn3, Larp7, and Alpk1 as highly attractive candidate genes with polymorphisms in coding or regulatory regions and expression differences between susceptible and resistant mouse strains. Conclusions: Subcongenic analysis of the major susceptibility locus Cdcs1 on mouse chromosome 3 revealed a complex genetic structure. Candidate gene approaches revealed attractive genes within the identified regions with homologs that are located in human susceptibility regions for IBD.

overall_design: Bone marrow derived macrophages (BMDM) of colitis susceptible C3Bir-Il10-/- and colitis resistant B6-Il10-/- as well as the Il10-/- reciprocal congenic strains CB-R1 (C3Bir genetic background carrying a long congenic chromosome 3 element containing Cdcs1 from B6, rendering this formerly susceptible background resistant) and BC-R3 (B6 genetic background that carries the Cdcs1-region of C3Bir) were cultured and stimulated with flagellin or left unstimulated.   BMDM were obtained from 3 male mice per genotype and cultured in polystyrene 6-well culture plates. Before RNA-isolation, 3 wells per plate were stimulated with Cbir1 flagellin (and subsequently pooled for RNA isolation), the other 3 wells left unstimulated (and also pooled). In total, these experiments were replicated three times in order to perform microarray analyses in triplicates.

first: Gwen

last: Büchler

first: Jason

last: Beckwith

first: Lydia

last: Petell

middle: P

last: Affourtit

first: Benjamin

last: King

first: Daniel

middle: J

last: Shaffer

first: Derry

middle: C

last: Roopenian

first: Hans

last: Hedrich

first: John

last: Sundberg

first: Edward

last: Leiter


title: Transcriptional Biomarkers to Predict Female Mouse Lung Tumors in Rodent Cancer Bioassays - A 26 Chemical Set

city: Research Triangle Park

first: Russell

middle: Scott

last: Thomas

state: NC

address: 6 Davis Drive

institute: The Hamner Institutes for Health Sciences

summary: The process for evaluating chemical safety is inefficient, costly, and animal intensive.  There is growing consensus that the current process of safety testing needs to be significantly altered to improve efficiency and reduce the number of untested chemicals. In this study, the use of short-term gene expression profiles was evaluated for predicting the increased incidence of mouse lung tumors.  Animals were exposed to a total of 26 diverse chemicals with matched vehicle controls over a period of three years.  Upon completion, significant batch-related effects were observed.  Adjustment for batch effects significantly improved the ability to predict increased lung tumor incidence.  For the best statistical model, the estimated predictive accuracy under honest five-fold cross-validation was 79.3% with a sensitivity and specificity of 71.4 and 86.3%, respectively.  A learning curve analysis demonstrated that gains in model performance reached a plateau at 25 chemicals, indicating that the size of the current data set was sufficient to provide a robust classifier.  The classification results showed a small subset of chemicals contributed disproportionately to the misclassification rate.  For these chemicals, the misclassification was more closely associated with genotoxicity status than efficacy in the original bioassay. Statistical models were also used to predict dose-response increases in tumor incidence for methylene chloride and naphthalene.  The average posterior probabilities for the top models matched the results from the bioassay for methylene chloride.  For naphthalene, the average posterior probabilities for the top models over-predicted the tumor response, but the variability in predictions were significantly higher. The study provides both a set of gene expression biomarkers for predicting chemically-induced mouse lung tumors as well as a broad assessment of important experimental and analysis criteria for developing microarray-based predictors of safety-related endpoints.

overall_design: Five-week-old female B6C3F1 mice were exposed for 13 weeks to 26 chemicals. The chemical and dose information are provided with the individual sample annotations.  With each chemical treatment, a matched vehicle control group was run concurrently with the exposure.  A subset of two chemicals, methylene chloride and naphthalene, were performed in a five-point dose response with a matched purified air control group.  The concentrations for these exposures overlapped those in the original cancer bioassay. Gavage exposures were administered 5 days per week and feed exposures were provided 7 days per week.  For inhalation exposures, mice were exposed 6 hr per day, 5 days per week.  After 13 weeks, animals were euthanized and lungs were collected. The right lobe was used for microarray analysis. Microarray analysis was performed on the lungs of three to four mice per treatment group.


title: GM-CSF-mediated granulopoiesis is regulated by the transcription factor STAT5A/B

city: Bethesda

first: WeiPing

last: Chen

state: MD

address: Bldg 8, Room 1A11, NIDDK/NIH

department: GCL

institute: NIDDK/NIH

summary: GM-CSF controls the development of granulocytes but little is known about the contribution of the downstream mediating transcription factor STAT5A/B.  To elucidate this pathway, we generated mice lacking the Stat5a and 5b genes in blood cells.  Peripheral neutrophils were decreased and administration of 5-FU and GM-CSF failed to induce granulopoiesis in Stat5a/b-mutant mice. GMPs were isolated and cultured with GM-CSF.  Both the number and size of STAT5A/B-null colonies were reduced and GM-CSF-induced survival of mature STAT5A/B-null neutrophils was impaired.  Time-lapse cinematography and single cell tracking of GMPs revealed that STAT5A/B-null cells were characterized by a longer generation time and an increased cell death.  Gene expression profiling experiments suggested that STAT5A/B directs GM-CSF signaling through the regulation of cell survival genes.

overall_design: Mice lacking or with  the Stat5a and 5b genes in blood cells, which were treated w/o GMP

first: Akiko

last: Kimura

first: Michael

last: Rieger

first: James

last: Simone

first: Mark

last: Wickre

last: O’Shea

first: Timm

last: Schroeder

first: Lothar

last: Hennighausen


title: Erythroid differentiation: G1E model

city: University Park

first: Ross

last: Hardison

state: PA

address: 303 Wartik Lab

institute: Pennsylvania State University

summary: Analysis of erythroid differentiation using Gata1 gene-disrupted G1E ER4 clone cells. Estradiol addition activates an ectopically expressed Gata-1-estrogen receptor fusion protein, triggering synchronous differentiation. 30 hour time course corresponds roughly to late burst-forming unit-erythroid stage (t=0 hrs)  through orthochromatic erythroblast stage (t=30 hrs).

overall_design: G1E ER4 cells cultured in G1E medium were treated at 6 time points with estradiol to initiate erythroid differentiation by activating Gata1 transcription factor and total RNAs from treated cells were extracted for microarray experiment. The erythroid differentiation status was confirmed by cell pellet color and expression of microRNA miR451. The design was similar to an earlier studies (Welch, J. J., Watts, J. A., Vakoc, C. R., Yao, Y., Wang, H., Hardison, R. C., Blobel, G. A., Chodosh, L. A., and Weiss, M. J. (2004)). Global regulation of erythroid gene expression by transcription factor GATA-1. Blood 104, 3136-3147), except that a more recent version of Affymetric chip was used to acheive greater transcriptome coverage.

first: Duonan

last: Yu

first: Mitchell

last: Weiss

first: Yong

last: Cheng


title: Expression data of murine GPI-deficient bone marrow cells in a mouse model of targeted Pig-a deletion

first: Valeria

last: Visconte

address: 10 Center Dr

department: 10-CRC Clinical Research Center

institute: NIH

summary:
Somatic mutation in the X-linked phosphatidylinositol glycan class A (PIG-A) gene causes glycosylphosphatidylinositol (GPI) anchor deficiency in humans with Paroxysmal Nocturnal Hemoglobinuria (PNH). Clinically, patients with PNH have intravascular hemolysis, venous thrombosis and bone marrow failure. We produced a conditional Pig-a knock-out mouse model specifically inactivating the Pig-a gene in hematopoietic cells to study the role of PIG-A deficiency in PNH pathophysiology. We used Affymetrix Mouse Genome 430 2.0 chips to investigate the gene expression pattern in the mouse model of targeted Pig-a deletion.

Keywords: Gene expression comparison between untransplanted and transplanted GPI-deficient bone marrow cells

overall_design: We performed microarray analysis on 3 pools of sorted GPI-deficient (GPI-) and GPI normal (GPI+) bone marrow cells derived from the same Pig-a knock-out animals, and identified 1275/669 genes of the 45,101 transcripts potentially available for screening using 2-fold change, <10% false discovery rate (FDR) and percentage of present calls as selection criteria. The major representative genes belong to the category of immune response. We tested whether the molecular differences between GPI- and GPI+  bone marrow cells could be preserved when GPI- cells were transplanted in lethally-irradiated wild type mice (C57BL/6). Microarray analysis was performed using sorted bone marrow cells from 4 animals transplanted with GPI-deficient bone marrow cells (T-GPI-) from Pig-a knock-out donors, 3 animals transplanted with GPI-normal bone marrow cells from C57BL/6 donors (T-GPI+), and  GPI-normal bone marrow cells from 4  wild-type C57BL/6 mice (WT-GPI+). We found 296 probesets with  2-fold change cutoff, of which T-GPI- cells had 145 up-regulated genes in comparison to WT-GPI+ cells, and had 123 genes differentially expressed when compared to T-GPI+ cells. The gene expression of the GPI-deficient cells was very similar between the two sets of microarray experiments affirming the maintenance of the phenotype before and after bone marrow cell transplantation.


title: Regulated SMAD signalling in development

city: Pullman

first: Lizhong

last: Yang

state: WA

address: 531 Fulmer Hall/POB 644660

department: School of Molecular Biosciences

institute: Washington State University

summary: Phosphorylation and subsequent nuclear translocation of SMAD proteins determine the cellular response to activin.  Here we identify a novel means by which activin signalling is regulated to enable developmental stage-specific SMAD gene transcription.  In response to activin A, immature proliferating mouse Sertoli cells exhibit nuclear accumulation of SMAD3, but not SMAD2, although both proteins are phosphorylated.  In post-mitotic differentiating cells, both SMAD2 and SMAD3 accumulate in the nucleus.  Furthermore, immature Sertoli cells are sensitive to activin dosage; at higher concentrations maximal SMAD3 nuclear accumulation is observed, accompanied by a small, but significant, increase in nuclear SMAD2.  Microarray analysis confirmed that differential SMAD utilization correlated with altered transcriptional outcomes and identified new activin target genes, Gja1 and Serpina5, which are known to be required for Sertoli cell development and male fertility.  In immature Sertoli cells, genetic or transient knockdown of SMAD3 enhanced SMAD2 nuclear accumulation in response to activin, with increased Serpina5 mRNA levels associated with nuclear localized SMAD2.  In transgenic mice with altered activin bioactivity that display male fertility phenotypes, testicular Gja1 and Serpina5 mRNA levels reflected altered in vivo activin levels.  We conclude that regulated nuclear accumulation of phosphorylated SMAD2 is a novel determinant of developmentally regulated activin signalling.

overall_design:
Murine 6dpp Sertoli Cell treated with 0ng, 5ng, 50ng activin (duplicates).
Murine 15dpp Sertoli Cell treated with 0ng, 5ng activin (duplicates). Total 10 samples.

first: Catherine

last: Itman

first: Chris

last: Small

middle: D

last: Griswold

first: Ankur

last: Nagaraja

first: Martin

last: Matzuk

first: Chester

last: Brown

first: Matthias

last: Ernst

first: David

last: Jans

first: Kate

last: Loveland


title: Quantification of synthetic miRNAs

status: Public on Sep 08 2009

city: Kgs. Lyngby

first: Hanni

last: Willenbrock

address: Kemikalievej, bygning 208

department: Center for Biological Sequence Analysis

country: Denmark

institute: DTU

type: Non-coding RNA profiling by high throughput sequencing

summary: Recently, next-generation sequencing has been introduced as a promising, new platform for assessing the copy number of transcripts, while the existing microarray technology is considered less reliable for absolute, quantitative expression measurements. Nonetheless, so far, results from the two technologies have only been compared based on biological data, leading to the conclusion that, although they are somewhat correlated, expression values differ significantly. Here, we use synthetic RNA samples, resembling human microRNA samples, to find that microarray expression measures actually correlate better with sample RNA content than expression measures obtained from sequencing data. In addition, microarrays appear highly sensitive and perform equivalently to next-generation sequencing in terms of reproducibility and relative ratio quantification.

platform_organism:
Homo sapiens
Mus musculus
Rattus norvegicus
Human alphaherpesvirus 1
Human betaherpesvirus 5
Murid betaherpesvirus 1
human gammaherpesvirus 4
JC polyomavirus
Human immunodeficiency virus 1
Murid gammaherpesvirus 4
Human gammaherpesvirus 8
Betapolyomavirus hominis
Betapolyomavirus macacae

overall_design: Two artificial samples (sample A and sample B) were constructed by mixing different amounts of 744 synthetic RNAs.

first: Jesper

last: Salomon

first: Rolf

last: Søkilde

first: Thomas

middle: N

last: Hansen

first: Kim

last: Barken

first: Finn

last: Nielsen

first: Søren

last: Møller

last: Litman


title: Transcriptional maturation of neocortical fast-spiking GABAergic interneurons

status: Public on Sep 05 2009

city: Ashburn

first: Ken

last: Sugino

state: VA

address: 19700 Helix Dr

institute: Janelia Research Campus

summary: Fast-spiking (FS) interneurons are important elements of neocortical circuitry that constitute the primary source of synaptic inhibition  in adult cortex and impart temporal organization on ongoing cortical activity. The highly specialized intrinsic membrane and firing  properties that allow cortical FS interneurons to perform these functions are attributable to equally specialized gene expression, which is  ultimately coordinated by cell-type-specific transcriptional regulation. Although embryonic transcriptional events govern the initial  steps of cell-type specification in most cortical interneurons, including FS cells, the electrophysiological properties that distinguish adult  cortical cell types emerge relatively late in postnatal development, and the transcriptional events that drive this maturational process are  not known. To address this, we used mouse whole-genome microarrays and whole-cell patch clamp to characterize the transcriptional  and electrophysiological maturation of cortical FS interneurons between postnatal day 7 (P7) and P40. We found that the intrinsic and  synaptic physiology of FS cells undergoes profound regulation over the first 4 postnatal weeks and that these changes are correlated with  primarily monotonic but bidirectional transcriptional regulation of thousands of genes belonging to multiple functional classes. Using  our microarray screen as a guide, we discovered that upregulation of two-pore K leak channels between P10 and P25 contributes to one  of the major differences between the intrinsic membrane properties of immature and adult FS cells and found a number of other  candidate genes that likely confer cell-type specificity on mature FS cells.

overall_design: We characterized the transcriptional maturation of genetically labeled FS interneurons in mouse S1  cortex using whole-genome microarrays.mRNA was harvested from manually sorted GFPexpressing  neurons dissociated from acutely prepared brain slices  at a range of developmental time points (P7, P10, P15, P25, and  P40) and subsequently reverse transcribed, amplified, labeled,  and hybridized to Affymetrix 430 2.0 whole-genome microarrays  (three microarrays from three different animals per condition). We used the G42 transgenic mice described by Chattopadhyaya  et al. (2004) for all experiments.

first: Benjamin

middle: W

last: Okaty

last: Hempel


title: Gene expression analysis in intestinal epithelial cells of germ-free versus wildtype mice

status: Public on Sep 04 2009

city: Lund

first: Knut

last: Kotarsky

state: Skane

address: Tornavägen 10

department: Experimental Medical Siences

country: Sweden

institute: Lund University

summary: Gene expression was analyzed in intestinal epithelial cells of germ-free and wildtype mice.

overall_design: Ileums and proximal colons of 4 C57/Bl6 control and 4 C57/BL6 germ-free mice were collected and embedded in OCT. Tissue sectioned (10 mikroM) and laser capture microscopy was performed using a Zeiss/Palm laser capture system. Total RNA was extracted from about 16000 cells of the catapulted samples using RNA microprep kit (Stratagene). These 16 000 cells derive from all 4 animals (pooled approx. 4 000 cells from each mouse). We retrieved between 50 and 150 ng total RNA. Fourty ng total RNA was expanded by two rounds of in vitro cRNA amplification and affymetrix genechip Moe403 were probed at the Swegene Resource Center Lund, Sweden. Array data analysis was performed using dCHIP2004 software.

first: William

last: Agace

first: Hanna

last: Stenstad


title: CAFs are activated in incipient neoplasia to orchestrate tumor promoting inflammation in an NF-κB-dependent manner.

status: Public on Sep 02 2009

city: San Francisco

first: Neta

last: Erez

state: CA

address: 513 Parnassus Ave.

institute: UCSF

summary:
Full title: Cancer Associated Fibroblasts are activated in incipient neoplasia to orchestrate tumor promoting inflammation in an NF-κB-dependent manner.
Cancer Associated Fibroblasts (CAFs) support tumorigenesis by stimulating angiogenesis, cancer cell proliferation, and invasion. We demonstrate that CAFs also mediate tumor-enhancing inflammation. Using a mouse model of squamous skin cancer, we found a pro-inflammatory gene signature in CAFs isolated from dysplastic skin. This signature was also evident in CAFs from skin as well as mammary and pancreatic tumors in mice, and in human cancer. Surprisingly, the inflammatory signature was already activated in CAFs isolated from the incipient hyperplastic stage in multistep tumorigenesis. CAFs from this pathway functioned to promote macrophage recruitment, neovascularization and tumor growth in vivo, activities abolished when NF-κB signaling was inhibited. Additionally, we show that normal dermal fibroblasts can be “educated” by carcinoma cells to express pro-inflammatory genes.

overall_design: We performed expression profiling analysis of dermal fibroblasts sorted from dysplastic skin tissue of K14-HPV16 mice, and from age matched non transgenic controls. This array analysis was repeated in triplicate. Amplified RNA was hybridized to the 430 2.0 Affymetrix mouse genome arrays.

first: Douglas

last: Hanahan


title: Expression data from regulatory T cells with Stat3 ablation

city: New York

first: Ashutosh

last: Chaudhry

state: NY

department: Immunology

institute: Memorial Sloan-Kettering Cancer Center

summary: We used microarrays to detail the global programme of gene expression dependent upon Stat3 in regulatory T cells

overall_design: Regulatory T cells isolated from Foxp3<Cre/wtS> Stat3<fl/wt> or Foxp3 <Cre/wt> Stat3 <fl/fl> were isolated using FACS based sorting followed by RNA extraction, amplification, labeling and hybridization on Affymatrix Microarray

first: A

middle: Y

last: Rudensky


title: Gene Expression in White Adipose Tissue of KSR2-/- mice compared to wild type mice

status: Public on Sep 03 2009

city: Omaha

first: Robert

last: Lewis

state: NE

address: 12706 Lied Transplant Center

institute: Univ. of Nebraska Medical Center

summary: The goal of this study was to define the gene expression changes that take place in the white adipose tissue of KSR2-/- mice at 6 months of age.

overall_design: Three wild type mice and three KSR2-/- mice were used for total RNA extraction from white adipose tissue.  The RNA from each mouse was analyzed on one Affymetrix gene chip and the differences were compared using Gene Set Enrichment Analysis.

first: Kurt

last: Fisher


title: Central corneal thickness is a genetic dependent trait among inbred strains of mice

status: Public on Sep 01 2009

city: Iowa City

first: Markus

middle: H

last: Kuehn

state: IA

address: 375 Newton Road

department: Ophthalmology and Visual Sciences

institute: University of Iowa

summary:
Central corneal thickness (CCT) exhibits broad variability.  We determined the corneal gene expression profile three mouse strains with distinct corneal thickness: C57BLKS/J (88.6 um), SJL/J (123.5 um), and C57BL/6J (100.1 um).

Keywords: Strain comparison

overall_design: Enucleated eyes from 4 month old mice were dissected in phosphate-buffered saline and a punch of central cornea was collected utilizing a 2-mm biopsy punch. Two central corneal punches (left and right eyes) were pooled from each mouse to form one sample; three samples were analyzed per strain.

first: M

middle: G

last: Anderson

first: G

last: Lively

first: B

last: Jiang

last: Hedberg-Buenz

last: Chang

middle: E

last: Petersen


title: Global analysis of gene expression by SV40 T antigen in the mouse small intestine epithelium

city: Pittsburgh

first: Ulka

last: Sachdev-Ost

address: 300 Halket Street Suite 5414

institute: University of Pittsburgh School of Medicine

summary: SV40 large T antigen (TAg) contributes to cell transformation, in part, by targeting two well characterized tumor suppressors, pRb and p53. TAg expression affects the transcriptional circuits controlled by Rb and by p53. We have performed a microarray analysis to examine the global change in gene expression induced by wild-type TAg and TAg-mutants, in an effort to link changes in gene expression to specific transforming functions. For this analysis we have used enterocytes from the mouse small intestine expressing TAg. Expression of TAg in the mouse intestine results in hyperplasia and dysplasia. Our analysis indicates that practically all gene expression regulated by TAg in enterocytes is dependent upon its binding and inactivation of the Rb-family proteins.

overall_design: Laser capture microdissection (LCM) was used to isolate villus enterocytes from three independent non-transgenic mice, four wild-type TAg transgenic mice, five N136 transgenic mice, three D44N transgenic mice and three 3213 transgenic mice. Total RNA was extracted from the enterocytes with the Pico Pure kit and amplified twice with the Ribo Amp kit (Arcturus). Amplified RNA was processed by the Genomics and Proteomics Core Laboratories at the University of Pittsburgh and hybridized to the Affymetrix GeneChip Mouse Genome 430 2.0 array. MAS 5.0 was used to obtain the present/absent calls and CEL files were normalized by RMA to obtain log2 expression values.

first: Abhilasha

middle: V

last: Rathi

first: Maria

last: Sáenz-Robles

first: Paul

last: Cantalupo

last: Whitehead

first: James

last: Pipas


title: Weighted Gene Coexpression Network Analysis Identifies Biomarkers in Glycerol Kinase Deficient Mice

city: Los Angeles

first: Nicole

middle: Kim

last: MacLennan

address: 10833 LeConte Ave

department: Pediatrics

institute: University of California, Los Angeles

summary: Symptomatic glycerol kinase deficiency (GKD) is associated with episodic metabolic and central nervous system deterioration.  We report here the first application of Weighted Gene Co-Expression Network Analysis (WGCNA) to investigate a knockout (KO) murine model of a human genetic disease.  WGCNA identified networks and key hub transcripts from liver mRNA of glycerol kinase (Gyk) KO and wild type (WT) mice.      Day of life 1 (dol1) samples from KO mice contained a network module enriched for organic acid metabolism before Gyk KO mice develop organic acidemia and die on dol3-4 and the module containing Gyk was enriched with apoptotic genes.  Roles for the highly connected Acot, Psat and Plk3 transcripts were confirmed in cell cultures and subsequently validated by causality testing.  We provide evidence that GK may have an apoptotic moonlighting role that is lost in GKD.    This systems biology strategy has improved our understanding of GKD pathogenesis and suggests possible treatments.

overall_design: Male WT and KO mouse pups were sacrificed on day of life (dol) 1 and each liver was harvested.  Total RNA from 4 KO and 3 WT livers was isolated individually.  Affymetrix mus 430 2.0 GeneChips were used to analyze differences in liver gene expression between KO and WT mice.  Dol1 and 3 Gyk KO mice represent different disease states.  Dol 1 was chosen because mice are phenotypically asymptomatic with respect to Glycerol Kinase Deficiency (GKD) and allowed us to look at alterations that occur before the overt disease state.  Dol 3 mice are phenotypically symptomatic with respect to GKD.

middle: K

first: Jun

last: Dong

first: Jason

last: Aten

first: Steve

last: Horvath

first: Lola

last: Rahib

first: Loren

last: Ornelas

first: Katrina

last: Dipple

first: Edward

middle: R

last: McCabe


title: A murine pneumonic plague model after infection with wild-type Yersinia pestis CO92 and its Braun lipoprotein mutant

status: Public on Sep 29 2009

city: Galveston

first: Ashok

last: Chopra

address: 301 University Blvd.

department: Microbiology & Immunology

institute: University of Texas Medical Branch

summary: Swiss-Webster female mice (Charles River Laboratories, Wilmington, MA) 5-6 weeks of age were infected intranasally with 5 LD50 of either WT or Δlpp mutant of Y. pestis CO92. Uninfected mice were used as controls. At either 12 or 48 h post infection (p.i.), 3 mice per group were euthanized and the lungs, livers, and spleens were harvested and homogenized in 1 ml of RNALater (Ambion/Applied Biosystems, Austin, TX) using 50-ml tissue homogenizers (Kendell, Mansfield, MA). RNA was isolated from the tissue homogenates and purified using RNAqueous (Ambion). After an overnight precipitation, the RNA was resuspended in 20 ul of diethylpyrocarbonate (DEPC)-treated water and hybridized to Affymetrix GeneChip Mouse Genome 430 2.0 arrays, performed by the Molecular Genomics Core at UTMB Galveston, Texas, per manufacture protocols. The arrays had 45,000 probe sets representing more than 39,000 transcripts derived from ~34,000 well-substantiated mouse genes. The experiments were performed in triplicate (biological replicates), generating a total of 45 arrays.


title: Expression data from murine embryonic stem cell derived cardiac progenitors

city: Boston

first: Ibrahim

last: Domian

state: MA

address: 185 Cambridge Street

department: Division of Cardiology

institute: Massachusetts General Hospital

summary:
Mamamlian cardiogenesis occurs through the development of discreate populations of first and second heart field progenitors.  We have used a dual transgenic color reproter system to isolate purified populations of these progenitors.

We used microarrays to detail the global programme of gene expression underlying cardiac development in mouse

All four populations of cells are derived from embryonic stem cells differentiating in vitro (day 6 of in vitro differentaition).  The stem cell   line has two transgenic reporters as follows:

1.  The second heart field (SHF) specific reporter of the Mef2C gene (E. Dodou, S. M. Xu, B. L. Black, Mech Dev 120, 1021 (Sep, 2003)) driving the   expression of dsRed

2.  The cardiac specific enhancer ( C. L. Lien et al., Development 126, 75 (Jan, 1999)) driving the expression of eGFP.   Thus, the red cells are SHF specific, the green cells are cardiac specific, and the red/green are SHF and cardiac  specific.  These cells are compared   to the double negative cells which serve as a control.

overall_design: Embryonic stem cell derived progenitors were isolated into four distinct populations by FACS purifying these progenitors based on a two color reporter system.  Four populations were then compared to each other by transcriptional profiling.

first: Murali

last: Chiravuri


title: Activin/Nodal signaling in mouse embryonic stem cells

status: Public on Aug 31 2009

city: Kansas City

first: Jay

last: Vivian

state: KS

address: 3901 Rainbow Blvd.

department: Pathology

institute: University of Kansas Medical Center

summary: Members of the transforming growth factor (TGF)-β superfamily play essential roles in the pluripotency, self-renewal, and differentiation of embryonic stem cells. While bone morphogenic proteins maintain pluripotency of undifferentiated mouse ES cells, the role of Activin/Nodal signaling is less clear.  To determine the target genes of Activin/Nodal-Smad2 signaling in undifferentiated embryonic stem cells, changes in gene expression were examined following stimulation with recombinant Activin (2 hours) or after inhibition of Activin/Nodal with SB431542 (24 hours) using defined media culture conditions with LIF and 20 ng/mL BMP4.  SB431542 is a specific inhibitor of ALK4/5/7 receptors and antagonizes both Activin and Nodal signaling. Via western analysis, Activin stimulation increased pSmad2 in ES cells after 2 hours, and treatment with SB431542 for 24 hours virtually eliminated pSmad2.  Total Smad2 expression remained unchanged through these manipulations.  RNA from cells treated with Activin or SB431542 was extracted by standard methods with Qiagen RNeasy columns.  The RNA was analyzed with the Mouse Genome 430A Array from Affymetrix.  Samples were performed in duplicate, and RNA from cells treated with Activin or SB431542 was compared to untreated embryonic stem cells.

middle: L

first: Katherine

last: Galvin


title: Cell cycle exit and terminal differentiation independent of the Rb gene family during embryonic development

city: Stanford

first: Julien

last: Sage

address: 265 Campus Drive

department: Pediatrics and Genetics

institute: Stanford

summary: The retinoblastoma cell cycle regulator pRb and the two related proteins p107 and p130 are thought to suppress cancer development both by inhibiting the G1/S transition of the cell cycle in response to growth-arrest signals and by promoting cellular differentiation. Here, we investigated the phenotype of Rb family triple knock-out (TKO) embryonic stem cells as they differentiate in vivo and in culture. Confirming the central role of the Rb gene family in cell cycle progression, TKO mouse embryos did not survive past mid-gestation and differentiating TKO cells displayed increased proliferation and cell death. However, patterning and cell fate determination were largely unaffected in these TKO embryos. Furthermore, a number of TKO cells, including in the neural lineage, were able to exit the cell cycle in G1 and terminally differentiate. This ability of Rb family TKO cells to undergo cell cycle arrest was associated with the repression of target genes for the E2F6 transcription factor, uncovering a pRb-independent control of the G1/S transition of the cell cycle. These results show that the Rb gene family is required for proper embryonic development but is not absolutely essential to induce G1 arrest and differentiation in certain lineages.

overall_design: Genome-wide gene expression was analyzed for wild-type and TKO ESC and EB cells. Three biological replicates were isolated for wild-type ESCs, TKO ESCs, and wild-type EBs, while five biological replicates were isolated for TKO EBs.

first: Stacey

last: Wirt

first: Adam

middle: S

last: Adler


title: Expression data of Naive Treg, allogeneic tumor-activated Treg, and GVHD-activated Treg cells

city: St Louis

first: Rekha

last: Meyer

state: MO

address: 4320 Forest Park Av

institute: Washington University School of Medicine

summary:
Regulatory T (Treg) cells play an important role in the induction and maintenance of peripheral tolerance.  Treg cells also suppress a variety of other immune responses, including anti-tumor and alloimmune responses.  We have previously reported that tumor-activated Treg cells express granzyme B and that granzyme B is important for Treg cell-mediated suppression of anti-tumor immune responses (GSE13409).  Here, we report that allogeneic mismatch also induces the expression of granzyme B.  Granzyme B-deficient mice challenged with fully mismatched allogeneic P815 mastocytoma cells have markedly improved survival compared to WT and other granzyme- or perforin-deficient mice, suggesting an immunoregulatory role for granzyme B in this setting.  Treg cells harvested from the tumor environment of P815-challenged mice express granzyme B.  Treg cells also express granzyme B in vitro during mixed lymphocyte reactions and in vivo in a mouse model of graft-versus-host disease (GVHD).  However, in contrast to findings from our previously published tumor model, granzyme B is not required for the suppression of effector T cell (Teff) proliferation in in vitro Treg suppression assays stimulated by either Concanavalin A or allogeneic antigen presenting cells.  Additionally, in an ex vivo assay, sort-purified in vivo-activated CD4+Foxp3+ Treg cells from mice with active GVHD -- under conditions known to induce granzyme B expression in Treg cells -- suppressed Teff cell proliferation in a granzyme B-independent manner.  Adoptive transfer of naive granzyme B-deficient CD4+CD25+ Treg cells into a mouse model of GVHD rescued hosts from lethatlity equivalently to naive wild-type Treg cells.  Serum analysis of GVHD-associated cytokine production in these recipients also demonstrated that Treg cells suppressed production of IL-2, IL-4, IL-5, GM-CSF, and IFN-gamma in a granzyme B-independent manner.  In order to determine whether the context in which Treg cells are activated alters the intrinsic properties of Treg cells, we used Foxp3 reporter mice to obtain gene expression profiles of CD4+Foxp3+ Treg cells purifed from naive resting spleens, spleens from mice with acute GVHD, and from ascites fluid of mice challenged intraperitoneally with allogeneic P815 tumor cells.  Unsupervised analyses revealed distinct activation signatures of Treg cells among the 3 experimental groups.  Taken together, these findings demonstrate that granzyme B is not required for Treg cell-mediated suppression of GVHD, which is in contrast to what we have previously reported for Treg cell function in the setting of tumor challenge.  Cell intrinsic differences could partially account for these differential phenotypes.  These data also suggest the therapeutic potential of targeting specific Treg cell suppressive functions in order to segregate GVHD and graft-versus-tumor effector functions.

Keywords: Normal vs Activated

overall_design:
Six replicates of Naive CD4+Foxp3+ Treg cells were purified from resting spleens, five replicates of allogeneic tumor-activated Treg cells and three samples of GVHD-activated Treg cells.
Naive reps 1-3 are controls for the GVHD-activated samples.
Naive reps 4-6 are controls for the Allogeneic tumor-activated samples.

first: Tim

last: Ley

first: Sheng

last: Cai


title: MEF FAN TNF

city: Toulouse

first: bruno

last: Ségui

address: Av Jean Poulhes

country: France

institute: INSERM U858

summary:
FAN (Factor associated with neutral sphingomyelinase activation) is an adaptor protein that constitutively binds to TNF-R1. Microarray analysis was performed in fibroblasts derived from wild-type or FAN knockout mouse embryos to evaluate the role of FAN in TNF-induced gene expression.
We used microarrays to detail the global programme of gene expression underlying cellularisation and identified distinct classes of up-regulated genes during this process.

Keywords: genetic modification

overall_design: MEFs were derived from C57BL/6 embryos that were either wild type or FAN-/-. Cells were either grown in DMEM 0%FCS for 24h or DMEM 0% FCS for 8h followed by incubation in DMEM 0% FCS containing 50ng/ml TNF for 16h. These four conditions were each used to generate total RNA (RNeasy MidiKit, Qiagen) which was sent to AROS applied biotechnology (Sweden) for Affymetrix GeneChip Mouse Genome 430 2.0 Array analysis.

first: Bruno

first: Jason

last: Iacovoni


title: Evaluation of Estrogen receptor related gene expression in C57BL/6 vs ERKO-alpha and ERKO-beta mice

status: Public on Aug 29 2009

city: Rensselaer

first: Sridar

last: Chittur

address: One Discovery Drive, CRC 342G

department: Center for Functional Genomics

institute: SUNY-University at Albany

summary:
These samples are part of the MMRRC study conducted at UAlbany

Keywords: strain comparison

overall_design: Whole brains from C57BL/6 mice were compared to those from estrogen receptor knockout mice on the same background. Six samples (3 male and 3 female) from each strain (wild type B6, ERalphaKO, ERbetaKO) were compared in this study. These were processed on Affymetrix Mouse 430 plus 2.0 arrays to obtain expression data for each cell ine.

first: Marcy

last: Kuentzel

first: Paulette

last: McCormick

first: Gerald

last: Bothe


title: Integrative approach for prioritizing cancer genes in sporadic colon cancer

status: Public on Aug 27 2009

city: Milano

first: James

middle: F

last: Reid

address: Via Adamello, 16

department: Molecular Genetics of Cancer

country: Italy

institute: Fondazione Istituto FIRC di Oncologia Molecolare

type:
Expression profiling by array
Genome variation profiling by SNP array

summary: The current multistep carcinogenesis models of colon cancer do not fully capture the genetic heterogeneity of the disease, which is additionally complicated by the presence of passenger and driver genetic alterations. The aim of the present study was to select in the context of this significant heterogeneity additional genes functionally related to colon cancer development. Methods: High-throughput copy number and gene expression data of 36 microsatellite stable sporadic colon cancers resected from patients of a single Institution characterized for mutations in APC, KRAS, TP53 and loss of 18q were analyzed. Genes whose expression correlated with the underlying copy number pattern were selected and their association with the above listed mutations and overall survival was evaluated. Results: Gain of 20q was strongly associated with TP53 mutation, and overall survival with alterations on 7p, 8p, 13q, 18q and 20q. An association with 18q loss and gain of 8q24 was also observed. New candidate genes with a potential role in colon cancer are PLCG1 on 20q, DBC1 on 8q21 and NDGR1 on 8p24. In addition an unexpected pattern of loss and mutability was found in the region upstream of KRAS gene. Conclusions: By integrating copy number alterations with gene expression and mutations in colon cancer associated genes we have developed a strategy that identifies previously known molecular features and additional players in the molecular landscape of colon cancer.

overall_design:
A total of 48 sporadic colon cancer samples were analyzed by Affymetrix Mapping 250K Nsp SNP Arrays and 36 of them were also analyzed by Affymetrix Human Exon 1.0 ST Array [transcript (gene) version].   Short summary: Expression data was correlated to copy number data to identify genes whose expression was induced by copy number changes. Gene dosage candidates were then evaluated for their association with gene mutation status of APC, KRAS and TP53, loss of heterozigosity of 18q and overall survival.   Long summary: Raw intensity .CEL files of the SNP arrays were processed with Chromosome Copy Number Analysis Tool (v.1.5.6 Affymetrix, Santa Clara, CA) to identify chromosomal gains and losses. Forty eight normal samples from the HapMap project supplied by Affymetrix were used as an un-paired reference set [http://www.affymetrix.com/support/technical/sample_data/500k_data.affx]. All genomic coordinates of the SNP array probes were mapped to the Human Mar. 2006 assembly the UCSC genome browser. Raw intensity .CEL files of the exon arrays were processed with the Robust Multi-Array implementation of Affymetrix Power Tools (v.1.8.6) using the core set of features (22011 probesets). All plots and analysis steps following this processing were done using the R programming language version 2.9.0Bioconductor packages. The identification of statistically significant segments of aberration in the copy number data was performed using the default parameters of the KC-SMART algorithm and 1000 permutations (KCsmart v.2.2.0). The identification and annotation of genes within each aberrant segment was performed using biomaRt v.2.0.0. The list of genes was further annotated by using the cancerGenes resource, Cancer Gene Census and the list of breast and colon CAN genes listed in Wood et al. The genomic landscapes of human breast and colorectal cancers. Science 2007;318:1108-1113. Gene dosage effects across 36 samples for which gene expression and copy number data was available was assessed by evaluating the Spearman correlation of the raw continuous copy number (log-ratios) expression (log-intensity) for each genomic region surrounding each gene falling within the segments  identified by KC-SMART. Prior to performing this task, to reduce the total number of correlation tests to perform, we filtered the gene expression dataset by removing all entries having no Gene Symbol annotation 17291 probesets and half the remaining dataset exhibiting the lowest variance resulting in 8645 probesets. Category package version 2.10.0 was used to apply a linear model-based test to detect enrichment of systematic high correlation in specific chromosomal bands taking into consideration the hierarchical structure of the bands. Gene expression class comparisons and survival analysis of the selected gene dosage candidates were performed using two sample t-tests and Cox proportional hazards regression. P-value adjustment of the correlation tests and the gene expression associations was performed using the step-up false discovery rate (FDR) controlling procedure of Benjamini and Hochberg. Raw data, supplementary methods and figures supplied as reproducible documentation (Sweave file) available from the Web Link.

Clinical information of samples contains: gender (F=female, M=male), age (years), dimension (cm), stage (Duke's I-IV), status (0=Alive, 1=Dead), survival (months), apc (0=wt, 1=mut), kras (0=wt, 1=mut), tp53 (0=wt, 1=mut), chr18qloh: (0=no LOH, 1=LOH).

first: Manuela

last: Gariboldi

first: Viktorija

last: Sokolova

first: Patrizia

last: Capobianco

first: Andrea

last: Lampis

first: Federica

last: Perrone

first: Stefano

last: Signoroni

first: Aurora

last: Costa

first: Ermanno

last: Leo

first: Silvana

last: Pilotti

first: Marco

middle: A

last: Pierotti


title: Hepatic gene expression during the development of experimental biliary atresia in different mouse strains

status: Public on Aug 30 2009

city: Braunschweig

first: Robert

last: Geffers

address: Inhoffenstr. 7

department: Dep. Molecular Bacteriology

institute: HCI - Helmholtz Centre for Infection Research

summary: Biliary atresia (BA) is a rare cholestatic disease of unknown etiology that affects infants and shows an incidence of 1 out of 18,000 live births in Europe (1). The first therapeutic option is a timely performed portoenterostomy. However, the majority of patients suffer from a progressive inflammatory process, which leads to complete destruction of the extra- and intrahepatic biliary system followed by end-stage liver cirrhosis. Hence, BA is the leading indication for pediatric liver transplantation worldwide (2, 3). To understand the pathogenesis of the disease and improve theoutcome of BA patients, research has focused on the inflammatory process in liver and bile ducts, in which several factors are remarkably elevated, such as activated CD4 and CD8 T-cells, TNF alpha,IFN alpha and other proinflammatory TH1 cytokines (3-8). By the time of diagnosis, however, the disease has already reached an advanced state, characterized by the complete obstruction of the extrahepatic bile ducts with impaired bile flow and fibrosis or cirrhosis of the liver. Therefore, studies in humans focusing on the trigger mechanism of BA are limited due to the paucity of liver and availability of bile duct tissue for research. One infectious animal model has been developed, in which newborn Balb/c mice exclusively show the experimental BA phenotype after infection with rhesus rotavirus (RRV) (9, 10). This model allows the analysis of the inflammatory reactions in liver and bile ducts at early steps in the development of bile duct atresia (11-20). Furthermore, inbred mouse strains have been shown to have a different susceptibility for the development of experimental BA, suggesting that Balb/c mice have an immunological gap responsible for disease progression (10, 12). The aim of this study was to identify key genes responsible for the BA phenotype by comparing the transcriptomes at an early time point after virus infection, i.e. before bile duct atresia, between two mouse strains with different susceptibilities to BA. Differences in the virus titration and the clinical course of infected mice were analyzed, and variations in the hepatic gene response assessed by comparative microarray assays were correlated to variances in the hepatic inflammatory reaction.

overall_design: Balb/c mice and C57Black/6 (Black/6) mice were infected with RRV postpartum and signs of BA and survival were noted. Liver sections of diseased, healthy and control animals were assessed for T-cell expression, and the virus loads were determined. Second, mice were sacrificed after three days, and isolated hepatic RNA was subjected to gene expression analysis using Affymetrix Gene Chip MOE 430 2.0.We compared three individual expression profiles from RRV-infected Balb/c mice against 2 individual expression profiles from RRV-infected C57/BL6 control mice using the Affymetrix  GeneChip MOE 430 2.0.

first: J

last: Leonhardt

first: R

first: C

last: Pertersen


title: Gene expression profiling from memory P14 T cells with control or mutated ThPOK

city: seattle

first: Ruka

last: Setoguchi

address: 1959 NE Pacific Street

institute: University of Washington

summary: We noticed that ThPOK repression is readily abrogated upon in vitro TCR stimulation of peripheral CD8 T cells. This observation prompted us to investigate a role of ThPOK in the CD8 T cell response to an acute viral infection. We observed that clonal expansion is significantly less in both primary and secondary CD8 T cell responses in the absence of functional ThPOK. To approach this mechanism, we carried out a microarray analysis for comparison of gene expression between ThPOKhd/hd and ThPOKwt/wt P14 memory T cells.

overall_design: To identify genes either directly or indirectly regulated by ThPOK in CD8 T cells, we performed a microarray analysis of ThPOKhd/hd and ThPOKwt/wt memory P14 T cells. We analyzed gene expression profiles of memory cells since they showed the highest and most uniform ThPOK expression compared to not only naïve but also effector CD8 T cells. The two memory subsets were FACS purified during the memory phase, more than 100 days after LCMV infection.

first: Michael

middle: J

last: Bevan


title: Expression Profiling in Subcellular Compartments of Human Cell Line K562

status: Public on Aug 25 2009

first: Sridar

last: Chittur

summary:
These samples are part of the ENCODE consortium’s proposed time-limited Pilot Study for confirmation of the utility of RNA abundance measurements as a standard reference phenotyping tool.
Affy expression profiling of Subcellular RNA from ENCODE cell line K562

Keywords: cell fraction comparison

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: RNA samples from a single biological replicate

first: Carrie

last: Davis

first: Scott

last: Tenenbaum


title: ENCODE Tier2 cell phenotyping study

status: Public on Aug 24 2009

summary:
These samples are part of the ENCODE consortium’s proposed time-limited Pilot Study for confirmation of the utility of RNA abundance measurements as a standard reference phenotyping tool.

Keywords: cell type comparison

For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and  http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf

overall_design: Each of the 7 ENCODE laboratories submitted biological triplicates for at least one of the four Tier2 cell lines. These were processed on Affymetrix Exon 1.0 ST arrays to obtain phenotyping data for each cell ine.


title: Knee joint synovium from different grades of inflammation in mouse collagen induced arthritis (CIA)

status: Public on Aug 26 2009

city: Basel

first: Anton

last: Glück

institute: Novartis Institutes for BioMedical Research

summary:
Knee joint synovium was used for gene expression analysis of mouse collagen induced arthritis (CIA).  Synovium was prepared at day 30 after initial sensitization from: healthy controls, CIA animals with no, with mild, with moderate, or with severe joint inflammation.  Each sample group is represented by three replicates, each consisting of tissue collected from three to four animals.

Keywords: disease severity analysis

overall_design: The data set consists of 15 samples: five groups with three replicates each.  One sample group is from healthy controls, the other groups are from CIA animals with different degress of joint inflammation.

first: Leo

last: Joosten

first: Nicole

last: Hartmann

first: Martin

last: Letzkus

first: Nozomi

last: Takahashi


title: Gene expression from TCDD treated C57BL6/J and human Aryl hydrocarbon Receptor expressing primary mouse hepatocytes

city: Univeristy Park

first: Gary

last: Perdew

address: 309A Life Sciences Building

institute: The Pennsylvania State University

summary: The human and mouse aryl hydrocarbon receptor (hAHR and mAHRb) share limited (58%) transactivation domain sequence identity. Compared to the mAHRb allele, the hAHR displays 10-fold lower relative affinity for prototypical ligands such as 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD). However, in previous studies we have demonstrated that the hAHR can display a higher relative ligand binding affinity than the mAHRb for specific AHR ligands such as indirubin. Each receptor has also been shown to differentially recruit LXXLL co-activator-motif proteins and to utilize different TAD subdomains in gene transactivation. Using hepatocytes isolated from C57BL6/J mice (Ahrb/b) and AHRTtr transgenic mice which express hAHR protein specifically in hepatocytes, we investigated whether the hAHR and mAHRb differentially regulate genes. Microarray and quantitative-PCR analysis of Ahrb/b and AHRTtr primary-mouse hepatocytes treated with 10 nM TCDD revealed that a number of established AHR target genes such as Cyp1a1 and Cyp1b1 are significantly induced by both receptors. Remarkably, of the 1752 genes induced by mAHRb and 1186 genes induced by hAHR, only 265 genes (<10%) were significantly activated by both receptors in response to TCDD. Conversely of the 1100 and 779 genes significantly repressed in mAHRb and hAHR hepatocytes respectively, only 462 (<25%) genes were significantly repressed by both receptors in response to TCDD treatment. Genes identified as differentially expressed are known to be involved in a number of biological pathways, including cell proliferation and inflammatory response which suggests that compared to the mAHRb, the hAHR may play contrasting roles in TCDD-induced toxicity and endogenous AHR-mediated gene regulation.

overall_design: Isolated mouse hepatocytes from wild type and treated with 10nM TCDD for 6h were analyzed. 1 array per mouse (3 mice per treatment group) was used.GeneChip® Operating Software (Affymetrix) was utilized to preprocess CAB/CEL files generated from the 12 scanned microarrays which represented hepatocytes isolated from one mouse each. Data quality was initially assessed by checking the array image, B2 oligo performance, average background to noise ratios, poly-A controls, hybridization controls and the 3' to 5'  probe-set ratios for control genes (e.g. ß-actin or GAPDH).  Microarray data was normalized using Probe Logarithmic Intensity Error Approximation PLIER-MM algorithm (Affymetrix Expression Console™ Software 1.1). Normalized microarray data outputs from TCDD and control treated Ahrb/band AHRTtr hepatocytes were compared for differential expression using Significance Analysis of Microarrays (SAM, version 2.23A (Pan 2002; Tusher et al. 2001)) with 100 permutations, KNN-10.

first: Colin

last: Flaveny


title: Gene Expression Profiles Associated with Ethanol-Induced Teratogenic Effects in a C57BL/6 Mouse Embryo Culture Model

city: Indianapolis

first: Feng

last: Zhou

state: IN

address: 635 Barnhill Dr.

department: Anatomy & Cell Biology

institute: Indiana University School of Medicine

summary:
Mouse model for Fetal Alcohol Syndrome. Embryos exposed to alcohol in controlled environment to assess teratogenic effects.
Fetal Alcohol Syndrome (FAS) is a leading developmental disorder. To date,  a holistic view of molecular gene changes is largely unexplored. Using microarray analysis of whole embryo mouse culture with strict-control over alcohol-level, we found, directly related alcohol-metabolism, a reduction of retinol binding protein 1(Rbp1), and a, de novo expression of aldehyde dehydrogenase 1B1 (ALDH1B1).  Remarkably, four key hemopoiesis genes (glycophorin A, adducin 2, beta-2 microglobulin, and ceruloplasmin) became absent, and many histone variants genes were reduced.  Hypothesis-driven informatics analysis and intersection analysis of two independent experiments indicated that the altered genes are involved in cell growth, hemopoiesis, histone modification, eye and heart development, and a collective reduction in expression of growth factor genes (Igf1, Efemp1, Tieg, and Edil3) and neural specification genes (neurogenin, Sox 5, bHLHb5).  Down-regulated neural specification phenotypes further supported the above findings.  Further more, the gene expression profile indicated distinct subgroups which overlapped with the teratogenesis of the open- and the closed-neural tubes known in FAS.  In summary, our data reveal genes alteration with causal potential for dysmorpology (e.g. retinoic acid, neuronal specification, and neurotrophic factors, and epigenetics related histone genes) and those downstream responsive genes related to alcohol metabolism, and developmental teratogenesis.
Keywords: comparison of gene expression profiles for treated vs. control

overall_design: Comparison of whole embryo gene expression after exposure to ethanol for 46 hours. Note 2 independent experiments completed.

middle: C

first: Qianqian

last: Zhao

first: Makiko

last: Kuwagata

first: Yunlong

last: Liu

first: Charles

last: Goodlett

first: Jeanette

last: McClintick

first: Tiebing

last: Liang

first: Howard

last: Edenberg

first: Lang

last: Li


title: Transcriptional profiling of mouse glomerulus in lipopolysaccharide-induced proteinuria model

city: Stockholm

first: ying

last: sun

address: Scheeles Vag 2

institute: Karolinska Institutet

summary:
The pathogenic mechanisms of common kidney glomerular diseases, including the vast majority of cases of proteinuria, remain unknown.
To gain insight into the pathogenesis of proteinuria development, we characterized the glomerular gene expression changes that accompany early stages of proteinuria induced by lipopolysaccharide (LPS) in mice.

overall_design: Nine-week-old female C57Bl6 mice were  allocated to LPS-treated group (n=6) or PBS control group (n=4), and subsequently treated either by a single intraperitoneal injection LPS or control buffer.

first: Ying

last: Sun

first: Liqun

last: He

first: Minoru

last: Takemoto

first: Jaakko

last: Patrakka

first: Timo

last: Pikkarainen

first: Guillem

last: Genové

first: Jenny

last: Norlin

first: Katarina

last: Truvé

first: Karl

last: Tryggvason

first: Christer

last: Betsholtz


title: Skeletal muscle growth and fiber composition in mice are regulated through the transcription factors STAT5a/b

status: Public on Aug 18 2009

city: Bethesda

first: WeiPing

last: Chen

address: Building 8 room 1a11

department: Genomic Core Lab

institute: NIDDK/NIH

summary:
In skeletal muscle, STAT5a/b transcription factors are critical for normal postnatal growth, whole-animal glucose homeostasis, and local IGF-1 production.  These observations have led us to hypothesize that STAT5a/b are critical for maintenance of normal muscle mass and function.  To investigate this, mice with a skeletal muscle-specific deletion of the Stat5a/b genes (Stat5MKO) were used.  Stat5MKO mice displayed reduced muscle mass, altered fiber-type distribution and reduced activity.  On a molecular level, gene expression in skeletal muscle of Stat5MKO and control mice was analyzed by microarrays and real-time PCR, both in the presence and absence of growth hormone (GH) stimulation.  Several genes involved in muscle growth, fiber-type and metabolism were significantly changed.  Specifically in the quadriceps, a muscle almost exclusively composed of type II fibers, the absence of STAT5a/b led to increased expression of several genes associated with type I fibers and the de novo appearance of type I fibers.  Additionally, it is shown here that expression of the androgen receptor gene (Ar) is controlled by GH through STAT5a/b.  The link between STAT5a/b and Ar gene is likely through direct transcriptional regulation, as chromatin immunoprecipitaion of the Ar promoter region in C2C12 myoblasts was accomplished by antibodies against STAT5a. These experiments demonstrate an important role for STAT5a/b in skeletal muscle physiology and they provide a direct link to androgen signaling.

Keywords: stat5 KO or WT treated or w/o GH

overall_design: Total 6 WT controls and 6 stat5a/b KO mice, treated or w/o GH

first: Peter

last: Klover

first: Weiping

first: Bing

last: Zhu

first: Lothar

last: Hennighausen


title: Identification of direct transcriptional targets of V600EBRAF/MEK in melanoma

status: Public on Aug 19 2009

city: London

first: Leisl

middle: M

last: Packer

address: 237 Fulham Rd

department: Cell and molecular biology

institute: Institute of Cancer Research

summary:
A375P melanoma cells were treated with 1uM of the MEK inhibitor PD184352 or 0.4uM of the V600EBRAF inhibitor PLX4720 for 2hr, 6hr and 24hrs.
DMSO treatment for 2hr, 6hr and 24hrs serves as the negative control

overall_design: Triplicate experiments were performed for DMSO, PD184352 and PLX4720 treatment at 3 timepoints - 2, 6, 24hrs.

first: Richard

last: Marais


title: The epithelial splicing factors ESRP1 and ESRP2 positively and negatively regulate alternative splicing events

status: Public on Aug 15 2009

city: Iowa City

first: Yi

last: Xing

address: 3294 CBRB, 285 Newton Rd

department: Internal Medicine, Biomedical Engineering

institute: University of Iowa

summary:
Epithelial Splicing Regulatory Proteins 1 and 2 (ESRP1 and ESRP2) are recently discovered epithelial-specific RNA-binding proteins that promote splicing of the epithelial variant of the FGFR2, ENAH, CD44, and CTNND1 transcripts.  To catalogue a larger set of splicing events under the regulation of the ESRPs, we profiled splicing changes induced by RNA interference-mediated knockdown of ESRP1 and ESRP2 expression in a human epithelial cell line using the splicing-sensitive Affymetrix Exon ST1.0 Arrays.  Analysis of the microarray data using the previously described MADS tool resulted in the identification of over a hundred candidate ESRP-regulated splicing events.  We were able to independently validate 37 of these targets by RT-PCR.  The ESRP-regulated events encompass all known types of alternative splicing events.  Importantly, a number of these regulated splicing events occur in gene transcripts that encode proteins with well-described roles in the regulation of actin cytoskeleton organization, cell-cell adhesion, cell polarity, and cell migration.  In sum, this work reveals a novel list of transcripts differentially spliced in epithelial and mesenchymal cells, implying that coordinated alternative splicing plays a critical role in determination of cell type identity.

Keywords: control / knockdown comparison

overall_design: Short interfering knockdown of ESRP1 and ESRP2 in human PNT2 prostatic epithelium cells was performed as described before (Warzecha et al., 2009, Molecular Cell 33:591-601). The efficiency of ESRP1 and ESRP2 knockdown was monitored by quantitative RT-PCR as described before (Warzecha et al., 2009, Molecular Cell 33:591-601).  In all cases the efficiency of the knockdown was close to 80%. We conducted Exon array profiling on RNAs from four siESRP1/2-treated samples and four siGFP controls.

first: Claude

last: Warzecha

first: Shihao

last: Shen

first: Russ

middle: P

last: Carstens


title: Involvement of 4E-BP1 in the protection induced by HDLs on pancreatic beta cells

city: RTP

first: Joan

last: Stuart

address: 5 Moore Drive

institute: GSK

summary: High-density lipoproteins (HDLs) protect pancreatic β cells against apoptosis. This property might be related to the increased risk to develop diabetes in patients with low HDL blood levels. However, the mechanisms by which HDLs protect β cells are poorly characterized. Here we use a transcriptomic approach to identify genes differentially modulated by HDLs in β cells subjected to apoptotic stimuli.

overall_design:
Tested conditions on beta-TC3 cells for 6h (in triplicates for each condition; GSK3 and GSK4 indicate two independent HDL preparations):

Complete Medium + Vehicle [V]
Complete Medium + HDL (prep. HDL GSK3 or HDL GSK4) [HDL]
Starvation medium + Vehicle [V.S]
Starvation medium + HDL (prep. HDL GSK3 or HDL GSK4) [HDL.S]

first: Christian

last: Widmann

first: Sujoy

last: Ghosh


title: Murine fracture healing

city: Phoenix

first: Winnie

last: Liang

state: AZ

address: 445 N. Fifth Street

institute: Translational Genomics

summary: Time-point expression analysis of fractures calluses at 1, 3, and 5 days post-fracture in young and old BALB/c mice.

overall_design: Femur fractures were generated on female c57BI6 mice in triplicate: 8 month old retired breeders (old mice) and 6 week old mice (young mice) were used. 1, 3, and 5 days post-fracture, fracture calluses were dissected and total RNA isolated. Expression profiling was performed using Affymetrix's Mouse Genome 430 2.0 array.

first: Aaron

last: Daluiski


title: AID-positive vs AID-negative

city: Los Angeles

first: Markus

last: Müschen

address: 4650 Sunset Blvd

department: Childrens Hospital Los Angeles

institute: University of Southern California

type:
Genome variation profiling by SNP array
Expression profiling by array

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: refer to individual Series