title: Reconstruction of the dynamic regulatory network that controls Th17 cell differentiation by systematic perturbation in primary cells (Th17 differentiation timecourse)

overall_design: Time course microarray data for Th17 differentiation, including Th0 control


title: Met synergizes with p53 loss to induce mammary tumors that possess features of claudin-low breast cancer (mRNA expression data).

status: Public on Mar 04 2013

city: St. Louis

first: Robert

last: Lesurf

state: MO

address: 4444 Forest Park Ave

department: McDonnell Genome Institute

country: USA

institute: Washington University in St. Louis

summary: Transcriptional profiling of normal and tumorigenic mouse mammary tissue. Mouse genotypes consist of wildtype, MMTV-Met, MMTV-Met;Trp53fl/+;Cre, and Trp53fl/+;Cre.

overall_design: Two-color common reference design. Samples consist of 11 normal tissues, 8 MMTV-Met tumors, 14 MMTV-Met;Trp53fl/+;Cre tumors, and 8 Trp53;Cre tumors.

first: Jennifer

middle: F

last: Knight

first: Dushanthi

last: Pinnaduwage

first: Ryan

last: Davis

first: Sadiq

last: Saleh

first: Dongmei

last: Zuo

first: Hong

last: Zhao

first: Monica

last: Naujokas

first: Naila

last: Chugthai

first: Jason

middle: I

last: Herschkowitz

first: Aleix

last: Prat

first: Anna

middle: M

last: Mulligan

first: William

middle: J

last: Muller

middle: D

last: Cardiff

first: Jeffrey

last: Gregg

first: Irene

last: Andrulis

first: Michael

last: Hallett

first: Morag

last: Park


title: Identification of stem cell populations in sweat glands and ducts: roles in homeostasis and wound repair

status: Public on Jan 02 2013

city: New York

first: Catherine

middle: P

last: Lu

state: NY

address: 1230 York Ave

institute: Rockefeller University

summary: Sweat glands are abundant glands of our body and essential for thermoregulation. Like mammary glands, they originate from epidermal progenitors. However, they display few signs of cellular turnover, and whether they have stem cells and tissue regenerative capacity remain largely unexplored. Here we address these issues. Using lineage-tracing, we identify multipotent progenitors in sweat duct that transition to unipotency after developing the sweat gland. In characterizing four adult stem cell populations of glandular skin, we show that they display distinct regenerative capabilities and remain unipotent when healing epidermal, myoepithelial-specific and luminal-specific injuries. We devise purification schemes, isolate and transcriptionally profile progenitors. Exploiting molecular differences between sweat and mammary glands, we show that only some progenitors regain multipotency to produce de novo ductal and glandular structures, but that these can retain their identity even within certain foreign microenvironments. Our findings provide new concepts about glandular stem cells and sweat gland biology.

overall_design: 10 samples from mouse paw pads were analyzed

first: Elaine

last: Fuchs


title: Master Transcription Factors and Mediator Establish Super-Enhancers at Key Cell Identity Genes [gene expression]

status: Public on Feb 13 2013

city: Cambridge

first: Richard

middle: A

last: Young

state: MA

address: 9 Cambridge Center

institute: Whitehead Institute for Biomedical Research

summary: The master transcription factors Oct4, Sox2 and Nanog bind enhancer elements and recruit the Mediator co-activator to activate much of the gene expression program of embryonic stem cells (ESCs). We report here that the ESC master transcription factors and Mediator form “super-enhancers” at most genes known to control the pluripotent state, including those encoding the master transcription factors themselves. These super-enhancers consist of extraordinarily large genomic domains occupied by exceptional amounts of Oct4, Sox2, Nanog, Klf4, Esrrb and Mediator.  Super-enhancers stimulate considerably higher transcription than typical enhancers in vivo and in reporter vectors.  Reduced levels of Oct4 or Mediator cause preferential loss of expression of super-enhancer-associated genes relative to other genes, suggesting how changes in gene expression programs might be accomplished during development.  In other more differentiated cells, super-enhancers containing cell-type-specific master transcription factors are also found at genes that define cell identity.  These results implicate super-enhancers in the control of mammalian cell identity and differentiation.

overall_design: Time-course of gene expression following shRNA knockdown of Oct4 and Med12.

middle: H

last: Kagey

first: Warren

last: Whyte

first: David

last: Orlando

first: Brian

last: Abraham

first: Denes

last: Hnisz


title: Met synergizes with p53 loss to induce mammary tumors that possess features of claudin-low breast cancer

first: Robert

last: Lesurf

type: Genome variation profiling by genome tiling array

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

overall_design: Refer to individual Series


title: Expression profile of adult Drosophila melanogaster expressing a self-replicating RNA of Sindbis virus

status: Public on Jan 31 2013

city: Bloomington

last: Hardy

state: Indiana

address: Simon Hall MSBI, Room 220E 212 S. Hawthorne Drive

department: Biology

institute: Indiana University

summary:
Alphaviruses establish a persistent infection in arthropod vectors, which is essential for effective transmission of the virus to vertebrate hosts. The development of persistence in insects is not well understood, although it is thought to involve the innate immune response. Using a transgenic fly system (SINrep) expressing a self-replicating viral genome, we have previously demonstrated the antiviral response of the Drosophila Imd (Immune Deficiency) and Jak-STAT innate immunity pathways.
In the current study, microarray analysis of SINrep flies in comparison to control GFP flies aims to detect genes that are sensitive to Sindbis viral RNA replication.

overall_design: Both SINrep and GFP adult flies were harvested 3 days post eclosion for RNA extraction and hybridization on Affymetrix microarrays. We look for genes significantly altered in the presence of viral RNA replication.

first: Megan

middle: B

last: Kingsolver

first: Zhijing

last: Huang

middle: W


title: Expression data from RAW264.7 murine macrophages infected with Toxoplasma gondii and subsequently stimulated with interferon-gamma

status: Public on Feb 19 2013

first: Emily

middle: E

last: Rosowski

address: 77 Massachusetts Ave, 68-258d

institute: MIT

summary:
Toxoplasma strains are known to inhibit the expression of several interferon-gamma induced genes, and a type II strain was shown to dysregulate genome-wide responses to interferon-gamma in human fibroblasts (Kim et al., 2007, J Immunol.).  In this study we aimed to determine the effect of infection with three clonal lineages of Toxoplasma, type I, II, and III strains on genome-wide interferon-gamma induced transcription in murine macrophages.  We also assessed the effect of the two main Toxoplasma modulators of mouse macrophage transcription, ROP16 and GRA15 (Jensen et al., 2011, Cell Host Microbe).
We used Affymetrix microarrays to analyze host cell transcription after Toxoplasma infection and interferon-gamma stimulation.

overall_design: RAW264.7 murine macrophages were left uninfected or infected with type I (RH), type I ∆rop16 (RH ∆rop16), type II (Pru), type II ∆gra15 (Pru ∆gra15), or type II (CEP) parasites at an MOI ~5 for 18 hours and subsequently stimulated with murine IFN-γ for six hours.  Plaque assays were done to assess parasite viability.  Total RNA was isolated and hybridized to Affymetrix Mouse 430A 2.0 gene chips.

first: Jeroen

last: Saeij


title: Gene expression profiles of NIH3T3 influenced by MARVELD1

status: Public on Mar 01 2013

city: Harbin

first: Shan

last: Wang

address: 2 Yikuang Street, Nangang District

country: China

institute: Harbin Institute of Technology

summary: To investigate biological function of MARVELD1, we have employed whole genome microarray expression profiling as a platform to analyze gene expression alternation influenced by MARVELD1. MARVELD1 stably transfected NIH3T3 and control vector cells were used to compare their gene expression profiling.

overall_design: Total RNA from MARVELD1 overexpressing NIH3T3 cells and control cells were extracted to analyze their gene expression profiles by using mouse whole genome expression  microarray.

first: Yu

last: Li


title: Quercetin induces hepatic lipid omega-oxidation and lowers serum lipid levels in mice

status: Public on Feb 05 2013

city: Wageningen

first: Evert

middle: M.

last: van Schothorst

address: De Elst 1

country: Netherlands

institute: Wageningen University

summary: Elevated circulating lipid levels are known risk factors for cardiovascular diseases (CVD). In order to examine the effects of quercetin on hepatic lipid metabolism and detailed serum lipid profiles, mice received a mild-high-fat diet without (control) or with supplementation of 0.33% (w/w) quercetin for 12 weeks. Gas chromatography and 1H nuclear magnetic resonance were used to measure quantitatively serum lipid profiles and whole genome microarray analysis was used to identify the responsible mechanisms in liver. There were no significant differences found in mean body weight, energy intake and hepatic lipid accumulation between the quercetin and control group.  In serum of quercetin-fed mice, TG levels were decreased with 15%, poly unsaturated fatty acids (PUFA) were increased with 14% and saturated fatty acids were decreased. Palmitic acid, oleic acid, and linoleic acid were all decreased in quercetin-fed mice by 9-15%. Both palmitic acid and oleic acid can be oxidized by omega-oxidation. Indeed, gene expression profiling showed that quercetin increased hepatic lipid metabolism, especially omega-oxidation. At the gene level, this was reflected by the up regulation of cytochrome P450 (Cyp) 4a10, Cyp4a14, Cyp4a31 and Acyl-CoA thioesterase 3 (Acot3). Two relevant regulators, Cytochrome P450 oxidoreductase (Por, rate limiting for cytochrome P450s) and the transcription factor Constitutive androstane receptor (Car; official symbol Nr1i3) were also up regulated in the quercetin-fed mice. We conclude that quercetin intake increased hepatic lipid omega-oxidation and lowered corresponding circulating lipid levels, a process that may involve Por and Car, and results in a potential beneficial CVD preventive effect.

overall_design: Liver samples were obtained from 36 C57BL/6J male adult mice. All mice started with a three week adaptation phase, in which they were fed a mild-high-fat diet. 12 mice were sacrificed immediately after the adaptation phase (t=0). The other 24 mice received the mild-high-fat diet without (HF) or with supplementation of 0.33% (w/w) quercetin (HF-Q) for 12 weeks.

first: Elise

last: Hoek-van den Hil

first: Jaap

last: Keijer

first: Annelies

last: Bunschoten

first: Ivonne

last: Rietjens

first: Peter

middle: C

last: Hollman


title: Mamld1 knockdown with siRNA in mouse Leydig tumor cells

status: Public on Jan 26 2014

city: Tokyo

first: Michiko

last: Nakamura

address: 2-10-1 Ohkura Setagaya-ku

department: Molecular Endocrinology

country: Japan

institute: National Research Institute for Child Health and Development

summary: MAMLD1 (mastermind-like domain containing 1, alias CXorf6) on human chromosome Xq28 is a causative gene for hypospadias, a mild form of 46,XY disorders of sex development. To date, multiple mutations have been identified in patients with various types of hypospadias. In this regard, the mouse homologous gene Mamld1 is transiently expressed in fetal Sertoli and Leydig cells around the critical period for sex development, and transient Mamld1 knockdown using small interfering RNAs (siRNAs) reduces testosterone production in cultured mouse Leydig tumor cells (MLTCs). We performed detailed analyses in Mamld1 knockdown experiments using MLTCs.   We used microarrays to display the global gene expression profile change, after knockdown of Mamld1 expression in MLTCs, to find the gene set regulated by Mamld1. To compare changes in gene expression profile in MLTCs, MLTCs were transfected with control siRNA or Mamld1 siRNA, 48h later, total RNA was subjected to Q-PCR to confirm a significantly decreased Mamld1 mRNA, and then these total RNA samples were for microarray analysis. The microarray data was analyzed to explore gene sets/pathways, which were significantly impacted by knockdown of Mamld1.  Real-time RT-PCR and microarray analyses showed significantly decreased Cyp17a1 expression (~70%) in both siRNA1- and siRNA2-transfected MLTCs. The siRNAs knockdown did not affect the expressions of Nr5a1 (Sf1), Star, Por, and Insl3.   47 genes including a Notch-related gene Hey1 were significantly up-regulated (fold change >2.0) and 38 genes were significantly down-regulated (fold change <0.5) in both siRNA1- and siRNA2-transfected MLTCs (Table S1 and Table S2). However, Mamld1 knockdown had no discernible effect on the Hes3 expression level (siRNA1: fold change 0.92, P=0.80; siRNA2: fold change 1.43, P=0.35).

overall_design:
MLTCs were maintained in RPMI 1640 supplemented with 10% fetal bovine serum, and were transiently transfected with two siRNAs against Mamld1 or with non-targeting control RNA. To compare changes in gene expression profile in MLTCs, 48h later, total RNA was subjected to Q-PCR to confirm a significantly decreased Mamld1 mRNA, and then these total RNA samples were for microarray analysis.

Supplementary file: t-test unpaired [i1] Vs [control], [i2] Vs [control], P <= 0.05 FC >=2.0

first: M

last: Miyado

last: Fukami

first: T

last: Ogata


title: Gene expression changes in the microdissected colon epithelium are similar to those of intact colon in the Il10-/- mouse model of IBD

city: Palmerston North

first: Anna

last: Moore

address: Tennent Drive

country: New Zealand

institute: AgResearch Ltd

summary:
Epithelial cells play an important role in the protection of the colon mucosa from the resident microbiota and are involved in the initiation and maintenance of intestinal inflammation. LMD is a technique that allows the extraction of specific cell types, such as colonic epithelial cells, to analyse gene expression. LMD of colon epithelial cells followed by microarray analysis could be of more value than microarray analysis of intact colon for determining which pathways are active in the colon mucosa in the early and late stages of inflammation due to increased sensitivity to changes in specific cell populations. An experiment was performed using microarray analysis of intact colon samples and microdissected colon epithelial cell samples from Il10-/- and C57BL/6J mice at 6 and 12 weeks of age to study the molecular changes that occur in early and late inflammation stages in colon epithelium of a mouse model of colitis.
Results showed that intact colon and colon epithelial cell gene expression profiles were similar in terms of pathways between Il10-/- and C57BL/6J mice at 12 weeks of age and between Il10-/- mice at 12 and 6 weeks of age. More immune-related pathways were identified at 6 weeks of age in epithelial cells than intact colon. This suggests that LMD and targeting of specific cell types may be of particular use when studying the early stages of inflammation before the intestinal morphology is detectably altered, by increasing analysis sensitivity to mucosal gene expression changes.

overall_design: 2x2 factorial with two tissue types analysed. Two strains of mouse (Il10 knockout mouse and the background strain C57BL/6J) were sampled at 2 timepoints (6 and 12 weeks of age) and intact proximal colon and colon epithelium harvested from each mouse (6 mice per group except for group colon epithelium C57 mouse 12 weeks where only 5 samples reached quality control standards).

last: Russ

first: Nicole

last: Roy

first: Gordon

last: Reynolds

first: Jason

last: Peters

first: Rachel

last: Anderson

first: Matthew

last: Barnett

first: Warren

last: McNabb

first: Kelly

last: Armstrong

first: Paul

last: MacLean

first: Zaneta

last: Park


title: In vitro expansion of single adult Lgr5+ liver cells induced by Wnt-driven regeneration

status: Public on Jan 28 2013

city: London

first: Vivian

last: Li

address: 1 Midland Road

country: United Kingdom

institute: The Francis Crick Institute

summary: The Wnt target gene Lgr5 marks actively dividing stem cells in Wnt-driven, self-renewing tissues such as small intestine and colon, stomach and hair follicles. A 3D culture system allows long-term clonal expansion of single Lgr5+ stem cells into transplantable organoids that retain many characteristics of the original epithelial architecture. A crucial component of the culture medium is the Wnt agonist Rspo, the recently discovered ligand of Lgr5. Here we show that Lgr5-LacZ is not expressed in healthy adult liver, yet that small Lgr5-LacZ+ cells appear near bile ducts upon damage, coinciding with robust activation of Wnt signaling. As shown by lineage tracing using a novel Lgr5-ires-CreERT2 knock-in allele, damage-induced Lgr5+ cells generate hepatocytes and bile ducts in vivo.  Single Lgr5+ cells from damaged liver can be clonally expanded as organoids in Rspo1-based culture medium over multiple months. Such clonal organoids can be induced to differentiate in vitro and to generate functional hepatocytes upon transplantation into FAH-/- mice. These findings imply that previous findings on Lgr5+ stem cells in actively self-renewing tissues extend to damage-induced stem cells in a tissue with a low rate of spontaneous self-renewal.

overall_design: We first generated arrays from multiple wildtype tissues including muscle, white adipose tissues, brown adipose tissues, liver and pancreas. Then we generated arrays from liver derived cultures maintained in different conditions, and compared the expression profile with the corresponding parental tissues and other non-related tissues.

middle: S

first: Meritxell

last: Huch

first: Hans

last: Clevers


title: Sox9 mediated differential gene expression was assessed in prostaspheres derived from urogenital sinus epithelial cells

status: Public on Jan 01 2013

city: Baltimore

first: Luigi

last: Marchionni

state: MD

address: 1550 Orleans St., CRB2, Room 1M52

department: Oncology

institute: Johns Hopkins University

summary: Prostate cancer is one of the most common malignancies and the second leading cause of death from cancer in men. The molecular mechanisms driving prostate carcinogenesis are complex; with several lines of evidence suggesting that the re-expression of conserved developmental programs play a key role. Through conditional gene targeting and organ grafting, we describe conserved roles for the transcription factor Sox9 in the initiation of both prostate organogenesis and prostate carcinogenesis in murine models. Abrogation of Sox9 expression prior to the initiation of androgen signaling blocks the initiation of prostate development. Similarly, Sox9 deletion in two genetic models of prostate cancer (TRAMP and Hi-Myc) blocks cancer initiation. Expression profiling of Sox9-null prostate epithelial cells reveals that the role of Sox9 in the initiation of prostate development may relate to its regulation of multiple cytokeratins and/or calcium-related proteins. Due to its essential role in cancer initiation, manipulation of Sox9 targets in at-risk men may prove useful in the chemoprevention of prostate cancer.

overall_design: Sox9 differential gene expression in prostaspheres derived from the urogenital sinus epithelium was assessed by tow-colors direct comparisons of labeled moieties. Hybridizations were performed on the Agilent (Santa Clara, CA) Whole Mouse Genome DNA microarray (mgug4122a). Sox9 targets were assessed in murine prostate epithelial cells with targeted deletion.

first: Zhenhua

last: Huang

first: Brian

last: Simons

first: Ashley

last: Ross

first: Paula

last: Hurley

first: David

last: Berman

first: Edward

last: Schaeffer


title: Eye imaginal disc expressing transgenic human proinsulin in Drosophila eye under regulation of GMR-GAL4

status: Public on Dec 23 2012

city: Chicago

first: Soo-Young

state: IL

address: 5841 S. Maryland Ave.

department: Medicine

institute: The University of Chicago

summary: We used microarrays to detail the global program of gene expression in response to expression of either mutant (C96Y) or wild-type human proinsulin and identified distinct classes of up-regulated genes. Results provides insight into the molecular mechanisms underlying a form of neonatal diabetes.

overall_design: Drosophila eye imaginal discs were selected at the wandering stage of 3rd instar larval development for RNA extraction and hybridization on Affymetrix microarrays.

first: Michael

last: Ludwig

first: Graeme

last: Bell

first: Martin

last: Kreitman


title: Early immunologic events at the tick-host interface

status: Public on Dec 21 2012

city: Galveston

first: Dar

last: Heinze

state: Texas

address: 301 University Blvd

department: Pathology

institute: University of Texas Medical Branch

summary: Ixodes species ticks are competent vectors of tick-borne viruses including tick-borne encephalitis and Powassan encephalitis.  Tick saliva has been shown to facilitate and enhance viral infection.  This likely occurs by saliva-mediated modulation of host responses into patterns favorable for viral infection and dissemination.  Because of the rapid kinetics of tick-borne viral transmission, this modulation must occur as early as tick attachment and initiation of feeding.  In this study, the gene expression profile of cutaneous bite-site lesions created by uninfected ticks were analyzed at 1, 3, 6, and 12 hours after Ixodes scapularis nymphal tick attachment to discover host pathways or responses potentially important in tick-borne viral establishment.

overall_design: Four milimeter ear biopsies from BALB/cJ mice infested with Ixodes scapularis nymphs were assayed using Affymetrix genechip 430A 2.0 arrays at 1, 3, 6, and 12 hours after infestation during a primary exposure. 3 mice were measured at each time point. Controls were 3 similarly housed but tick-free mice.

first: D

first: J

middle: R

last: Carmical

last: Aronson

first: S

last: Thangamani


title: Defining a molecular roadmap of cellular reprogramming into iPS cells [mRNA profiling]

status: Public on Dec 20 2012

city: Boston

first: Ben

middle: S.

last: Wittner

address: 149 13th Street

department: Center for Cancer Research

institute: Massachusetts General Hospital

summary: Factor induced reprogramming is a slow and inefficient process with only rare cells progressing towards induced pluripotent stem cells (iPSCs). Owing to these restraints, mechanistic studies have been limited to analyses of heterogeneous bulk populations undergoing reprogramming and partially reprogrammed cell lines. Here, by combining surface markers (Thy1, SSEA1) and an Oct4-GFP fluorescent reporter allele, we analyzed defined intermediate cell populations poised to becoming iPSCs at the transcriptional and epigenetic levels using genome-wide and single cell technologies. We found that factor-induced reprogramming elicits two discernible transcriptional waves that are characterized by the initial extinction of the somatic gene expression program and the concomitant acquisition of an ESC-like proliferative and metabolic state, followed by the activation of an embryonic pluripotent state primed for differentiation. The first wave is mostly driven by gene activation through c-Myc and gene repression by Klf4, whereas the second wave is a result of gradually activated Oct4/Sox2 targets in cooperation with Klf4 targets and other downstream regulators. While microRNA expression and enrichment for individual histone modifications (H3K4me3 or H3K27me3 enriched promoters) mirrored the observed biphasic transcriptional pattern, the establishment of bivalent domains (H3K4me3/H3K27me3 enriched promoters) occurred more gradually. In contrast, changes in DNA methylation took place predominantly at the end of reprogramming when cells assumed a stable pluripotent state. Cells that became refractory to reprogramming activated the first but failed to initiate the second transcriptional wave. However, introduction of additional copies of the reprogramming transgenes into these cells rescued their ability to form iPSCs, indicating that suboptimal transcription factor levels are a limiting factor for efficient iPSC formation. This integrative analysis allowed us to identify novel genes and microRNAs that enhance reprogramming and surface markers that further subdivide intermediate cell populations. Collectively, our data offer new mechanistic insights into the nature and sequence of molecular events inherent to cellular reprogramming and provide a valuable resource of molecules that may act as roadblocks during iPSC formation.

overall_design: Time series design with samples sorted into subpopulations according to surface markers.

first: Jose

last: Polo

first: Endre

last: Anderssen

first: Ryan

last: Walsh

first: Benjamin

last: Schwarz

first: Christian

last: Nefzger

first: Sue

last: Lim

first: Marti

last: Borkent

first: Effie

last: Apostolou

first: Sara

last: Alaei

first: Jeniffer

last: Cloutier

first: Ori

last: Bar-Nur

first: Sihem

last: Cheloufi

first: Matthias

last: Stadtfeld

first: Maria

last: Figueroa

first: Daisy

last: Robinton

first: Sridaran

last: Natesan

first: Ari

last: Melnick

first: Jinfang

last: Zhu

first: Sridhar

last: Ramaswamy

first: Konrad

last: Hochedlinger


title: Growth factor independence 1 (Gfi1) regulates cell-fate decision of the bipotential granulocytic-monocytic precursor defined by the expression of CD48 as a new marker

status: Public on Dec 19 2012

city: Montréal

first: Lothar

last: Vassen

state: Quebec

address: 110, avenue des Pins Ouest

department: Hematopoiesis and Cancer

country: Canada

institute: IRCM

summary:
Using bone marrow cells of GFP:Gfi1 knock in mice, we separated Gfi1-high and Gfi1-low expressing cells in the classical CD11b+, GR1-low monocytic cell fraction. We sorted CD11b+, GR1-low GFP:Gfi1-high and low cells as well as CD11b+, GR1-high granulocytes and CD11b-high, GR1-intermediate cells from Gfi1-knock-out mice for further analysis.
We used Affymetrix Mouse Genome 430A 2.0 arrays (GPL8321)

overall_design: The study should determine how Gfi1 regulates the cell fate of monocytes/granulocytes in mice

last: Kosan

first: Hui

last: Zeng

first: Tarik

last: Möröy


title: Expression profiling in primary embryonic fibroblasts deficient for the N-terminal truncation of Lats1

status: Public on Dec 17 2012

city: Suita

first: Daisuke

last: Okuzaki

state: Osaka

address: Yamadaoka 3-1

department: Immunology Frontier Research Center

institute: Osaka univ.

summary: Lats1 and Lats2 are the mediators of the Hippo pathway that regulates tissue growth and proliferation. Lats1 and Lats2 kinases share 85% sequence identity in the kinase domain. However, their non-kinase regions at the N-terminus are distinct except for Lats conserved domain 1 (LCD1) and LCD2, suggesting that their N-terminal regions are important for Lats1/2-specific functions. In this study, we generated Lats1 knockout mice disrupting the N-terminal region containing LCD1 (Lats1ΔN/ΔN). We show that some Lats1ΔN/ΔN mice were born safely and grew normally. However, mouse embryonic fibroblasts (MEFs) from Lats1ΔN/ΔN mice displayed drastic defects in mitosis, showing enhanced centrosome overduplication, chromosomal misalignment, multipolar spindle formation, chromosome bridging, and cytokinesis failure. Moreover, they displayed accelerated cell cycle and cell growth bypassing a cell-cell contact inhibition like tumor cells, and exhibited anchorage independent growth. Indeed, Lats1ΔN/ΔN MEFs produced tumors in nude mice after subcutaneous injection, although the tumor growth rate was much slower than ordinary cancer cells. Furthermore, Yap, the key transcriptional co-activator in the Hippo pathway, was overexpressed and retained stable in Lats1ΔN/ΔN MEFs under high cell density, and expression of Lats2 mRNA were down-regulated.

overall_design: Total RNAs were extracted from MEFs under high or low cell density using miRNeasy extraction kit (Qiagen). A Dye-swapped experiment was performed by hybridizing complimentary RNA (cRNA) labeled with either Cyanine (Cy) -3 or Cy-5 (Perkin-Elmer) onto Whole Mouse Genome Oligo Microarray (G4122F; Agilent Technologies). Co-hybridizations were performed and data from Cy3 or Cy5 channels were analyzed as normalized signal intensities rather than as log ratios corresponding to each array.

first: Hiroshi

last: Nojima


title: Expression data from wild type and ttk^twk mutant whole ovaries

status: Public on Dec 06 2012

city: Seattle

first: Nathaniel

last: Peters

state: Washington

department: Genome Sciences

institute: University of Washington

summary:
Epithelial tube  morphogenesis requires precise orchestration of cell signaling, shape, migration, and adhesion. Follicle cells in the Drosophila ovary form  epithelial tubes, the lumens of which act as molds for the eggshell respiratory filaments, or dorsal appendages (DAs).  The Tramtrack69 (TTK69) transcription factor controls DA lumen volume and shape by promoting tube expansion; the tramtrack mutation twin peaks (ttk^twk) reduces TTK69 levels late in oogenesis, inhibiting DA tube expansion.
We used microarrays to identify differentially expressed genes in the ttk^twk mutant that are downstream of TTK69 and that play a functional role in DA tube expansion.

overall_design: Ovaries were dissected from female flies at 36 hours of age, to maximize the proportion of developing egg chambers undergoing DA tube expansion.  Flies were raised at a constant temperature of 25C in the presence of wet yeast, to maximize the rate of egg chamber production.

first: Celeste

last: Berg


title: Expression data of Normal versus Mutant MPS VII C3H mouse

status: Public on Dec 01 2012

city: Philadelphia

first: Michael

middle: K.

last: Parente

state: PA

address: 3615 Civic Center Blvd.

department: Neurology

institute: Children's Hospital of Philadelphia

summary:
We used microarray to detect pathway differences in the various brain regions in a monogenic in mucopolysaccharidosis type VII ( MPS VII ), a mouse model of a lysosomal storage disease
A number of changes revealed unexpected system and process alterations, such as upregulation of the immune system with few inflammatory changes (a significant difference from the closely related MPS IIIb model), down-regulation of major oligodendrocyte genes even though white matter changes are not a feature histopathologically, and a plethora of developmental gene changes.

overall_design: 94 samples, no replicates, made up of half normals and half MPS mutant mice for the MPS VII mutation backcrossed on a C3h-heouj background

middle: K

first: Ramona

last: Rozen

first: John

last: Wolfe


title: Dnmt3a regulates both cell proliferation and differentiation of mouse neural stem cells

status: Public on Dec 31 2012

city: Los Angeles

first: Kevin

last: Huang

state: CA

address: Gonda BLDG Rm. 6554, P.O.Box 957088

department: Human Genetics

institute: UCLA

summary: DNA methylation is known to regulate cell differentiation and neuronal function in vivo. Here we examined whether deficiency of a de novo DNA methyltransferase, Dnmt3a, affects in vitro differentiation of mouse embryonic stem cells (mESCs) to neuronal and glial cell lineages. We found that Dnmt3a-/- neural stem cells (NSCs) derived from mESCs have globally reduced methylcytosine levels and precociously differentiates into astrocytes and oligodendrocytes, consistent with our previous findings in the more severely hypomethylated Dnmt1-/- NSCs.  Moreover, Dnmt3a-/- NSC proliferation rate was significantly increased when compared to control. Thus, our work revealed a novel role for Dnmt3a in regulating both timing of neural cell differentiation and cell proliferation in NSCs.

overall_design: Dnmt3a KO vs. WT neural stem cells; 3 biological replicates of each.

first: Zhourui

last: Wu

first: Guoping

last: Fan


title: Expression profiling of Notch constitutive activation in the liver of mice

city: Barcelona

first: Clara

last: Alsinet

state: Catalonia

address: Rosselló 153

country: Spain

institute: IDIBAPS - Hospital Clínic

summary: Notch intracellular domain (NICD) is the active form of the Notch receptor. In this mouse model, NICD is inserted in the Rosa26 locus downstream of a loxP-STOP-LoxP (lsl) sequence and therefore NICD expression is dependant on Cre recombinase expression (Mono transgenic control Rosa26-lsl-NICD) . These mice are crossed with the AFP-Cre strain that expresses Cre in hepatoblasts due to its regulation by the AFP promoter and albumin enhancer (Double transgenic mutant AFP-Cre/Rosa26-lsl-NICD). Newborn mice at day 0 and day 2 are sacrificed and liver RNA samples from control monotransgenic Rosa26-lsl-NICD and from bitransgenic AFP-Cre/Rosa26-lsl-NICD (AFP-NICD) are obtained. Whole genome expression profiling of these samples is submitted.

overall_design: Five liver samples from Control mice and six liver samples from Mutant mice are analyzed using the Agilent Whole Mouse Genome Oligo Microarray G4122A platform. Array data was preprocessed and analyzed using GenePattern software and R.

first: Augusto

last: Villanueva

first: Kilangsungla

last: Yanger

first: Yujin

last: Hoshida

first: Yiwei

last: Zong

first: Sara

last: Toffanin

first: Leonardo

last: Rodriguez-Carunchio

first: Manel

last: Sole

first: Swan

last: Thung

first: Ben

middle: Z

last: Stanger

first: Josep

last: Llovet


title: Hepatic Leukemia Factor Reproduces Circadian Resistance To Cell Death

status: Public on Jan 15 2013

city: Richland

first: Katrina

middle: M

last: Waters

state: WA

address: 902 Battelle Blvd; MSIN J4-18

department: Biological Sciences Division

institute: Pacific Northwest National Laboratory

summary:
Here we examine the regulation of cell death by hepatic leukemia factor (HLF), which is an output regulator of circadian rhythms and is aberrantly expressed in human cancers, using an ectopic expression strategy in JB6 mouse epidermal cells and human keratinocytes.  Ectopic HLF expression inhibited cell death in both JB6 cells and human keratinocytes, as induced by serum-starvation, tumor necrosis factor alpha and ionizing radiation.
Microarray analysis indicates that HLF regulates a complex multi-gene transcriptional program encompassing upregulation of anti-apoptotic genes, downregulation of pro-apoptotic genes, and many additional changes that are consistent with an anti-death program.  Collectively, our results demonstrate that ectopic expression of HLF, an established transcription factor that cycles with circadian rhythms, can recapitulate many features associated with circadian-dependent physiological variation.

overall_design: Murine progenitor cells, JB6 cell line, were stably transformed with an HLF retroviral expression system or vector only control.  Confirmation of HLF ectopic expression was performed with western blot analysis.  Samples were collected from randomly cycling, preconfluent cells maintained in normal growth medium.

first: Thomas

last: Weber


title: Genetic ablation of Rest leads to in vitro-specific derepression of neuronal genes during neurogenesis

first: Yasuhiro

last: Yamada

address: 7-3-1 Hongo, Bunkyo-ku

department: Department of Molecular Pathology

institute: University of Tokyo

summary: Rest (RE1 silencing transcription factor, also called NRSF) is involved in the maintenance of the undifferentiated state of neuronal stem/progenitor cells in vitro by preventing precocious expression of neuronal genes.  However, the function of Rest during neurogenesis in vivo remains to be elucidated because of the early embryonic lethal phenotype of the conventional Rest knockout mice.  In the present study, we have generated Rest conditional knockout mice, and the effect of genetic ablation of Rest during the embryonic neurogenesis can be examined in vivo.  We herein show that Rest plays a role in suppressing the expression of neuronal genes in cultured neuronal cells in vitro, as well as in non-neuronal cells outside of the central nervous system, but that it is dispensable for the embryonic neurogenesis in vivo.  Our findings highlight the significance of extrinsic signals for the proper intrinsic regulation of neuronal gene expression levels in the specification of cell fate during embryonic neurogenesis in vivo.

overall_design: Total RNAs from E13.5 limbs and brains were analyzed for global gene expressions by Agilent microarray.

first: Hitomi

last: Aoki


title: Development of distinct gene expression signature in neurons from different regions of the brain

status: Public on Dec 05 2012

first: Michael

last: Katze

address: Rosen Building 960 Republican St.

department: Microbiology

summary: We have performed microarray expression profiling of mouse primary neurons (cortical neurons and granule cell neurons) to model molecular networks and define whether distinct antiviral IFN responses occurred in neurons corresponding to different brain regions.

overall_design: Primary mouse cortical neurons and granule cell neurons were left untreated, treated with 100IU/mL of IFN-β, or infected with West Nile virus (MOI of 1) and RNA was harvested after 24 hours. Three independent experiments were performed using a balanced design.

first: Hyelim

last: Cho

first: Sean

last: Proll

first: Kristy

middle: J

last: Szretter

middle: G

last: Gale

last: Diamond


title: Pharyngeal mesoderm regulatory network controls cardiac and head muscle morphogenesis

status: Public on Nov 26 2012

city: Rehovot

first: Eldad

last: Tzahor

address: Herzel 234

country: Israel

institute: The Weizmann Institute of Science

summary: The search for developmental mechanisms driving vertebrate organogenesis has paved the way toward a deeper understanding of birth defects. During embryogenesis, parts of the heart and craniofacial muscles arise from pharyngeal mesoderm (PM) progenitors. Here, we reveal a hierarchical regulatory network of a set of transcription factors expressed in the PM that initiates heart and craniofacial organogenesis. Genetic perturbation of this network in mice resulted in heart and craniofacial muscle defects, revealing robust cross-regulation between its members. We identified Lhx2 as a novel player during cardiac and pharyngeal muscle development. Lhx2 and Tcf21 genetically interact with Tbx1, the major determinant in the etiology of DiGeorge/velo-cardio-facial/22q11.2 deletion syndrome.  Furthermore, knockout of these genes in the mouse recapitulates specific cardiac features of this syndrome. We suggest that PM-derived cardiogenesis and myogenesis are network properties rather than properties specific to individual PM members. These findings shed new light on the developmental underpinnings of congenital defects.

overall_design: Mouse embryos were selected at E9.5, 10.5 and 11.5. We used Myf5Cre;RosaYFP in order to label all the myogenic lineage. Trunk (T) and Head (H) tissues were isolated, dispersed into a single cell suspension and FACS sorted for YFP+ population. Further analysis is described in Harel et al, PNAS (2012).

first: Itamar

last: Harel

first: Yoshiro

last: Maezawa

first: Roi

last: Avraham

first: Ariel

last: Rinon

first: Hsiao-Yen

last: Ma

first: Joe

last: Cross

first: Noam

last: Leviatan

first: Julius

middle: T

last: Hegesh

first: Achira

last: Roy

first: Jasmine

last: Jacob-Hirsch

first: Gideon

last: Rechavi

first: Jaime

last: Carvajal

first: Shubha

last: Tole

first: Chrissa

last: Kioussi

first: Sue

last: Quaggin


title: Expression data from RUNX1(41-214)-expressing and Runx1-knockout mice KSL cells

status: Public on Nov 29 2012

city: La Jolla

first: Dong-Er

last: Zhang

address: 3855 Health Sciences Drive

department: Moores UCSD Cancer Center

institute: University of California San Diego

summary: Mutations of RUNX1 are detected in patients with myelodysplastic syndrome (MDS). In particular, C-terminal truncation mutations lack a transcription regulatory domain and have increased DNA binding through the runt homology domain (RHD). The expression of the RHD, RUNX1(41-214), in mouse hematopoietic cells induced progression to MDS and acute myeloid leukemia (AML). Analysis of pre-myelodysplastic animals revealed expansion of c-Kit+Sca-1+Lin- (KSL) cells and skewed differentiation to myeloid at the expense of the lymphoid lineage. These abnormalities correlate with the phenotype of Runx1-deficient animals, as expected given the reported dominant-negative role of C-terminal mutations over the full-length RUNX1. However, MDS is not observed in Runx1-deficient animals. Gene expression profiling revealed that RUNX1(41-214) KSLs have an overlapping yet distinct gene expression profile from Runx1-deficient animals. Moreover, an unexpected parallel was observed between the hematopoietic phenotype of RUNX1(41-214) and aged animals. Genes deregulated in RUNX1(41-214), but not in Runx1-deficient animals, were inversely correlated with the aging gene signature of hematopoietic stem cells (HSC), suggesting that disruption of the expression of genes related to normal aging by RUNX1 mutations contributes to development of MDS. The data presented here provide insights into the mechanisms of development of MDS in HSCs by C-terminal mutations of RUNX1.

overall_design: Gene expression analysis were performed on c-Kit+/Sca-1+/Lin-/IL7Ra- (KSL) cells sorted from RUNX1(41-214)-expressing and Runx1-knockout (Runx1floxed/floxed MxCre+/-) and control mice (Runx1floxed/floxedMxCre-/-).

first: Shinobu

last: Matsuura

first: Kristen

last: Stevenson

first: Donna

last: Neuberg


title: Expression data from hearts of wild-type C57BL/6 mice infected with T. cruzi and controls (uninfected)

status: Public on Nov 20 2012

city: Worcester MA

first: Ricardo

last: Gazzinelli

address: 364 Plantation Street, LRB

department: Infectious Diseases and Immunology

institute: University of Massachussetts Medical School

summary:
An efficient innate immune recognition of the intracellular parasite T. cruzi is crucial for host protection against development of Chagas disease, which often leads to multiple organ damage, particularly the heart leading to cardiomyopathy. Mechanisms modulated by MyD88 have been shown to be necessary for resistance against T, cruzi infection. Recently, Nod-like receptors have been shown to play an important role as innate immune sensors, particularly as they relate to inflammasome function, caspase activation, and inflammatory cytokine production. In this study, we aimed to investigate the participation of innate immune responses in general, and inflammasomes in particular, in heart inflammation and cardiac damage upon infection with the T. cruzi parasite.
We used microarrays to gain insight into gene expression in the cardiac tissue of mice infected with the causative agent of Trypanosoma cruzi, and identified distinct classes of up-regulated genes during this process, including important genes involved in inflammasome activation and innate immune responses in general.

overall_design: The hearts of C57BL/6 mice day 18 post-infection with a Y strain of the parasite T. cruzi, and uninfected controls were extraced for RNA extraction and hybridization on Affymetrix microarrays. We sought to compare gene expression among two groups of mice, and so extracted the hearts of 3 control uninfected mice, and of 3 infected mice 18 days post-infection.

first: Mariana

middle: I

last: De Niz

first: Bernardo

last: Franklin


title: Gene expression in the embryonic Lrrc10 knockout heart

status: Public on Nov 15 2012

city: Madison

first: Matt

last: Brody

state: WI

address: 1300 University Ave.

institute: UW-Madison

summary: LRRC10 is a heart-specific gene required for proper cardiac function. The effects of Lrrc10 deletion on gene expression in the embryonic mouse heart was investigated.

overall_design: Pregnant heterozygous Lrrc10 knockout mice (Lrrc10+/-) mated to male heterozygous knockouts (Lrrc10+/-), housed in 12 hour light:12 dark, ad lib feeding and drinking conditions, were sacrificed at embryonic day 15.5 (E15.5) and embryonic hearts were dissected for cardiac gene expression analysis. A two color, reference design experiment in which heart RNA from 2 Lrrc10 homozygous knockout embryos (Lrrc10-/-) were pooled and labeled with Cy5 and hybridized according to Agilent protocols against a reference pool of RNA madeup from respective tissue taken from littermate E15.5 wildtype (Lrrc10+/+) hearts and labeled with Cy3.

first: Youngsook

last: Lee


title: Annexin A2 Modulates Radiation-Sensitive Transcriptional Programming and Cell Fate

status: Public on Nov 28 2012

first: Katrina

last: Waters

summary:
Whole genome expression profiling in the presence and absence of annexin A2 [shRNA] identified fundamentally altered transcriptional programming that changes the radioresponsive transcriptome.
Bioinformatics predicted that silencing AnxA2 may enhance cell death responses to stress in association with reduced activation of pro-survival signals such as nuclear factor kappa B. This prediction was validated by demonstrating a significant increase in sensitivity toward tumor necrosis factor alpha induced cell death in annexin A2 silenced cells, relative to vector controls, associated with reduced nuclear translocation of RelA (p65) following tumor necrosis factor alpha treatment.

overall_design: Murine progenitor cells, JB6 cell line, were stably transformed with Annexin A2 shRNA or vector only control and then treated with ionizing radiation at two doses, 10cGy and 100cGy.  Samples were collected at 2hr, 6hr and 24hr for RNA isolation.  Two untreated time points were collected also: 2hr and 24hr.

first: Thomas

last: Weber


title: Transcriptome-wide miR-155 binding map reveals widespread non-canonical microRNA targeting [mRNA expression data]

status: Public on Nov 09 2012

first: Aly

middle: Azeem

last: Khan

address: 6045 S. Kenwood Ave.

institute: University of Chicago

summary:
microRNAs (miRNAs) are essential components of gene regulation, but identification of miRNA targets remains a major challenge.  Most target prediction and discovery relies on perfect complementarity of the miRNA seed to the 3’ untranslated region (UTR).  However, it is unclear to what extent miRNAs target sites without seed matches.  Here, we performed a transcriptome-wide identification of the endogenous targets of a single miRNA—miR-155—in a genetically controlled manner.  We found that approximately forty percent of miR-155-dependent Argonaute binding occurs at sites without perfect seed matches.  The majority of these non-canonical sites feature extensive complementarity to the miRNA seed with one mismatch.  These non-canonical sites confer regulation of gene expression albeit less potently than canonical sites.  Thus, non-canonical miRNA binding sites are widespread, often contain seed-like motifs, and can regulate gene expression, generating a continuum of targeting and regulation.
We used microarrays to measure changes in gene expression between activated wild type (WT) and miR-155 deficient (155KO) primary CD4 T cells.

overall_design: CD4 T cells were harvested from WT and miR-155 KO mice (Thai et al., 2007) and activated for 3 days in vitro.  Each array is from a separate biological replicate, which are cells originating from a separate mouse.

first: Gabriel

last: Loeb

middle: A

first: David

last: Canner

first: Joseph

last: Hiatt

first: Jay

last: Shendure

first: Robert

last: Darnell

first: Christina

last: Leslie

first: Alexander

middle: Y

last: Rudensky


title: Gene expression profiles of Notch signaling suppression in prostate basal cells.

status: Public on Nov 08 2012

city: Houston

first: Chad

last: Creighton

state: TX

address: One Baylor Plaza, Mail Stop: BCM305

department: Biostatistics, Ducan Cancer Center

institute: Baylor College of Medicine

summary: The role of Notch signaling in the maintenance and differentiation of adult prostate stem cells remains unclear. We found that Notch ligands are mainly expressed within the basal cell lineage, while active Notch signaling is detected in both the prostate basal and luminal cell lineages. Disrupting the canonical Notch effector RBP-J impairs the differentiation of prostate basal stem cells and increases their proliferation in vitro and in vivo, but does not affect luminal cell biology. Conversely, ectopic Notch activation in adult prostates results in basal cell depletion and luminal cell hyper-proliferation. TGFβ dominates over Notch and overrides Notch ablation-induced proliferation of prostate basal cells. In turn, Notch confers positive feedback by up-regulating a plethora of TGFβ signaling components including TGFβRI. These findings reveal crucial roles of the self-enforced positive reciprocal regulatory loop between TGFβ and Notch in maintaining prostate basal stem cell dormancy.

overall_design: We employed an in vitro prostate sphere assay to further investigate how Notch signaling regulates basal cell proliferation and differentiation. FACS isolated adult murine prostate basal cells (using FVB mice) were cultured in the prostate sphere assay with or without N-[(3,5-Difluorophenyl)acetyl]-L-alanyl-2-phenyl]glycine-1,1-dimethylethyl ester (DAPT), an inhibitor for the γ-Secretase complex. Gene expression profiles were taken of vehicle- and DAPT-treated prostate spheres.

first: Li

last: Xin

last: Valdez


title: Diabetes risk gene and Wnt effector Tcf7l2/TCF4 controls hepatic response to perinatal and adult metabolic demand

status: Public on Nov 01 2012

city: London

first: Vivian

last: Li

address: 1 Midland Road

institute: The Francis Crick Institute

summary: Most studies on TCF7L2 SNP variants in the pathogenesis of type 2 diabetes (T2D) focus on a role of the encoded transcription factor TCF4 in β-cells. Here, a mouse genetics approach shows that removal of TCF4 from β-cells does not affect their function, while manipulating TCF4 levels in the liver has major effects on metabolism. In Tcf7l2-/- mice, the immediate postnatal surge in liver metabolism does not occur. Consequently, pups die due to hypoglycemia. Combining chromatin immunoprecipitation with gene expression profiling, we identify a TCF4-controlled metabolic gene program that is acutely activated in the postnatal liver. In concordance, adult liver-specific Tcf7l2 knockout mice show reduced hepatic glucose production during fasting and display improved glucose homeostasis when maintained on high-fat diet. Furthermore, liver-specific TCF4 overexpression increases hepatic glucose production. These observations imply that TCF4 directly activates metabolic genes, and that inhibition of Wnt signaling may be beneficial in metabolic disease.

overall_design: RNA was extracted from liver tissues of the Tcf7l2 wildtype or knockout mice with treatments as indicated. Microarray analysis was performed to compare the expression profile changes between Tcf7l2 knockout and wildtype mice in response to treatment.

first: Sylvia

middle: F

last: Boj

first: Hans

last: Clevers


title: Murine Kidney Tissue: Unilateral ureteral obstructed mice vs. Sham control

city: Stanford

first: Bo

last: Wu

state: Ca

address: 300 Pasteur Dr. Grant/S259

department: Urology

institute: Stanford University

summary:
INTRODUCTION AND OBJECTIVE: Loss of renal function is often the impetus for operative intervention in renal obstruction.  Obstructive nephropathy is characterized discrete morphological and physiologic changes including tubular dilatation, apoptosis, and atrophy, as well as interstitial cellular infiltration and progressive interstitial fibrosis. We hypothesize that there are gene expression alterations correlated with obstructive nephropathy that could serve as biomarkers for early intervention.
METHODS: C57BL/6 mice were subjected to Unilateral Urethral Obstruction (UUO) or sham surgery at postnatal day 21 of life, and kidneys were harvested after 1, 2, 5 and 9 days postoperatively. RNA was extracted from kidneys and comprehensive gene expression profiling was performed with Agilent microarrays. We used biological replicates: 13 UUO replicates, 9 sham replicates. 2-Channel microarrays were hybridized using universal reference in the Cy3 channel. Ingenuity pathway analysis was used to analyze the biological function and gene networks of gene expression data.
RESULTS: Microarray analysis revealed more than 1800 transcripts that were up- or down-regulated over days 1 through 9 following obstruction, and included many transcripts reported previously (FOS, CD44, CLU, SPP1 and EGF). Pathway analysis revealed significant enrichment of transcripts in cell activation/differentiation, immune/inflammatory responses, cell cycle, metabolic process and transport. Interestingly, IPA network analysis showed that transcriptional regulatory pathway involving CEBPB/HNF4A is involved in obstructive nephropathy.
CONCLUSIONS: Transcript profiling identified numerous gene expression changes following UUO and suggests a role for CEBPB/HNF4A pathway in obstructive nephropathy. This dataset provides a foundation for development of biomarkers for obstructive nephropathy.
Time: day samples were collected post operation
Somatic Modification: mice were operated on to generate urethral obstruction (Obstructed/Control)


title: Early Mechanisms of Pathobiology are Revealed by Transcriptional Temporal Dynamics in Hippocampal CA1 Neurons of Prion Infected Mice

city: Winnipeg

first: Sarah

middle: Jean

last: Medina

state: Manitoba

address: 1015 Arlington Street

department: Molecular PathoBiology

institute: Public Health Agency of Canada

summary: Prion diseases typically have long pre-clinical incubation periods during which time the infectious prion particle and infectivity steadily propagate in the brain. Abnormal neuritic sprouting and synaptic deficits are apparent during pre-clinical disease, however, gross neuronal loss is not detected until the onset of the clinical phase. The molecular events that accompany early neuronal damage and ultimately conclude with neuronal death remain obscure. In this study, we used laser capture microdissection to isolate hippocampal CA1 neurons and then determined their pre-clinical transcriptional response during infection. We found that gene expression within these neurons is dynamic and characterized by distinct phases of activity. A major cluster of genes is altered during pre-clinical disease after which expression either returns to basal levels, or alternatively undergoes a direct reversal during clinical disease. Strikingly, we show that this cluster contains a signature highly reminiscent of synaptic N-methyl-D-aspartic acid (NMDA) receptor signaling and the activation of neuroprotective pathways. Additionally, genes involved in neuronal projection and dendrite development were also altered throughout the disease, culminating in a general decline of gene expression for synaptic proteins. Similarly, deregulated miRNAs such as miR-132-3p, miR-124a-3p, miR-16-5p, miR-26a-5p, miR-29a-3p and miR-140-5p follow concomitant patterns of expression. This is the first in depth genomic study describing the pre-clinical response of hippocampal neurons to early prion replication. Our findings suggest that prion replication results in the persistent stimulation of a programmed response, at least in part mediated by synaptic NMDA receptor activity that initially promotes cell survival and neurite remodelling. However, this response is terminated prior to the onset of clinical symptoms in the infected hippocampus, seemingly pointing to a critical juncture in the disease. Manipulation of these early neuroprotective pathways may redress the balance between degeneration and survival, providing a potential inroad for treatment.

overall_design: The CA1 hippocampal region was dissected out using LCM and RNA was extracted from these samples.  In total, 6 different time points were screened for both RNA and miRNA expression levels in prion infected and control animals.  Gene expression profiles from 6 time points (n ≥ 2) were determined using whole mouse 4x44K arrays.  We successfully validated a subset of candidate genes that were deregulated during early prion disease.  We performed a similar assessment of temporal miRNAs expression levels throughout the infection using the TLDA platform which was further validated by individual real-time PCR assays.  In parallel, immunoctyochemistry was used to characterize the cellular presence of astrocytes, microglial and neurons in the CA1 region throughout the disease which correlated well with both mRNA and miRNA expression profiles.  Staining for the PrPRes and neuronal toxicity levels was also performed to determine the spatial and temporal PrPRes deposition and assess the level of neuronal death that occurs in the hippocampus, respectively.  Using bioinformatic methods, potential pathways that were implicated by our data to be deregulated during early prion disease were presented while potential miRNA regulation of some of these candidate genes implicated in these pathways was also included.

first: Anna

last: Majer

first: Yulian

last: Niu

first: Bernard

last: Abrenica

first: Kathy

last: Manguiat

middle: L

last: Frost

first: Clark

last: Phillipson

first: Debra

last: Sorensen

first: Stephanie

last: Booth


title: Mouse Dbh-/- vs. Dbh+/+ E10.5 hearts

status: Public on Nov 06 2012

city: Orlando

first: Steven

middle: N.

last: Ebert

state: Florida

address: 6900 Lake Nona Blvd

department: Biomedical Sciences

institute: Univ of Central Florida

summary: Genomic microarray analysis of adrenergic-deficient (Dbh-/-) vs. wild-type control (Dbh+/+) mouse heart expression at embryonic day 10.5 (E10.5).

overall_design: Four Dbh-/- and four Dbh+/+ E10.5 hearts were isolated, and RNA was extracted from each for genomic microarray analysis using Affymetrix 430A 2.0 arrays.  The biological samples were collected and prepared in Dr. Ebert's laboratory at the Univ of Central Florida in Orlando, FL.  The RNA samples were sent to GeneLogic (Gaithersburg, MD) for the microarray analysis.

first: Kingsley

last: Osuala

first: Ha-Long

last: Nguyen

first: David

last: Weinshenker

middle: N


title: Global analysis of Cdk9-dependence for VSV-induced genes in Drosophila cells

status: Public on Oct 18 2012

first: Sara

last: Cherry

address: 304K Lynch Labs

institute: University of Pennsylvania

summary: To determine the Cdk9 targets of VSV-induced genes in Drosophila cells at 4 hours post-infection

overall_design: Drosophila cells were treated with Bgal or Cdk9 and infected with VSV (MOI 10) for 4 hours, in two independent biological replicates. Total RNA was isolated using Trizol and microarray experiments were performed at the University of Pennsylvania Microarray Facility (U. Penn MF) following directions according to the manufacturer’s protocol (Affymetrix). In brief, 100 ng RNA were amplified with the Ovation RNA Amplification System v2 (NuGen), the Encore Biotin Module (NuGen) was used for fragmentation and labeling, Protocol FS450 002 was used for hybridization, washing, and staining. Images were scanned using the GeneChip® Scanner 3000 and image analysis was performed using the Affymetrix® GeneChip® Command Console® Software (AGCC).

first: Jie

last: Xu

first: Mathew

last: Tudor


title: Expression data from uninfected and VSV-infected Drosophila cells at 4 hours post-infection

summary: Expression profiling of rapidly-induced genes upon VSV infection at 4 hours post-infection in Drosophila cells

overall_design: Drosophila cells were treated with Bgal dsRNA and either treated as uninfected or infected with VSV (MOI 10) for 4 hours, in two independent biological replicates. Total RNA was isolated using Trizol and microarray experiments were performed at the University of Pennsylvania Microarray Facility (U. Penn MF) following directions according to the manufacturer’s protocol (Affymetrix). In brief, 100 ng RNA were amplified with the Ovation RNA Amplification System v2 (NuGen), the Encore Biotin Module (NuGen) was used for fragmentation and labeling, Protocol FS450 002 was used for hybridization, washing, and staining. Images were scanned using the GeneChip® Scanner 3000 and image analysis was performed using the Affymetrix® GeneChip® Command Console® Software (AGCC).


title: Loss of Pim1 Imposes a Hyperadhesive Phenotype on Endothelial Cells

status: Public on Oct 29 2012

city: Zürich

first: Thomas

last: Walpen

address: Gloriastrasse 30 H09

department: Research Unit Division Internal Medicine

country: Switzerland

institute: University Hospital Zürich

summary: Background: PIM1 is a constitutively active serine-threonine kinase regulating cell survival and proliferation. Increased PIM1 expression has been correlated with cancer metastasis by facilitating migration and anti-adhesion. Endothelial cells play a pivotal role in these processes by contributing a barrier to the blood stream. Here, we investigated whether PIM1 regulates mouse aortic endothelial cell (MAEC) monolayer integrity. Methods: Pim1-/-MAEC were isolated from Pim1 knockout mice and used in trypsinization-, wound closure assays, electrical cell-substrate sensing, immunostaining, cDNA transfection and as RNA source for microarray analysis. Results: Pim1-/-MAEC displayed decreased migration, slowed cell detachment and increased electrical resistance across the endothelial monolayer. Reintroduction of Pim1- cDNA into Pim1-/-MAEC significantly restored wildtype adhesive characteristics. Pim1-/--MAEC displayed enhanced focal adhesion and adherens junction structures containing vinculin and β-catenin, respectively. Junctional molecules such as Cadherin 13 and matrix components such as Collagen 6a3 were highly upregulated in Pim1-/- cells. Intriguingly, extracellular matrix deposited by Pim1-/- cells alone was sufficient to induce the hyperadhesive phenotype in wildtype endothelial cells. Conclusion: Loss of Pim1 induces a strong adhesive phenotype by enhancing endothelial cell-cell and cell-matrix adhesion by the deposition of a specific extracellular matrix. Targeting PIM1 function therefore might be important to promote endothelial barrier integrity.

overall_design: Pim-1-/- mouse aortic endothelial cells were compared to wildtype cells

first: Martin

last: Peier

first: Elvira

last: Haas

first: Ina

last: Kalus

first: Jürg

last: Schwaller

first: Edouard

last: Battegay

first: Rok

last: Humar


title: Expression data from mouse proximal intestinal epithelial Lgr5(hi) stem cells and differentiated villus cells (enterocytes from Atoh1 conditional knockout)

status: Public on Oct 13 2012

city: Boston

first: Amit

last: Sinha

state: MA

address: 44 Binney St

department: Pediatric Oncology

institute: Dana-Farber Cancer Institute

summary: We used microarrays to detail the differentail gene expression between intestinal Lgr5(hi) stem cells and differentiated cells

overall_design: Assay the differential gene expression using total RNA from flow cytometry sorted Lgr5(hi) cells and EDTA isolated enterocytes from Atoh1 conditional knockout

first: Li-Lun

last: Ho

first: Scott

last: Armstrong

first: Ramesh

last: Shivdasani


title: Gene expression profile after TRPV4 knockdown in 3T3-F442A adipocytes

status: Public on Oct 12 2012

first: Li

last: Ye

address: 450 brookline ave. CLSB11118

summary: Comparing gene expression profile in 3T3-F442A adipocytes with shRNA against TRPV4 or GFP. TRPV4 is an ion channel expressed in adipocytes. Results provided information that how loss-of-function of TRPV4 affects gene expression in adipocytes.

overall_design: 4 samples were analyzed as two groups: shGFP (control) and shTRPV4 (experimental). Each group has two replicates.

first: Sandra

last: Kleiner

first: Jun

last: Wu

first: Rajan

last: Sah

first: Rana

last: Gupta

first: Alexander

middle: S

last: Banks

first: Paul

last: Cohen

first: Melin

last: Khandekar

first: Pontus

last: Boström

first: Rina

last: Mepani

first: Dina

last: Laznik

first: Theodore

last: Kamenecka

first: Xinyi

last: Song

first: Wolfgang

last: Liedtke

first: Vamsi

last: Mootha

first: Pere

last: Puigserver

first: Patrick

middle: R

last: Griffin

middle: E

last: Clapham

first: Bruce

last: Spiegelman


title: stem cell therapy of murine acute hepatitis

status: Public on Oct 11 2012

city: Kanazawa

first: Yoshio

last: Sakai

state: Ishikawa

address: 13-1 Takara-machi

department: Department of Gastroenterology

institute: Kanazawa University

summary: Gene expression profiles of the liver and hepatic inflammatory cells in murine concanavalin A-induced acute hepatitis models by treatment with adipose tissue derived stem cells

overall_design: C57Bl/6 female mice were treated with adipose tissue-derived stem cells immediately or 3 hours after concanavalin A injection. Two hours later, the liver tissues or hepatic inflammatory cells were obtained and gene expression of them were examined by DNA microarray.

first: Mami

last: Higashimoto


title: Chromatin Remodeling Enzyme Smarca5/Snf2h Regulates Cell Cycle Exit, Differentiation of the Lens Epithelium, and Denucleation of Lens Fiber Cells

city: LYON

first: Jiri

last: ZAVADIL

state: Other

address: 150 COURS ALBERT THOMAS

country: France

institute: IARC WHO

summary: Genome-wide approach to identify the cell-autonomous role of Snf2h in lens fiber cell terminal differentiation. Differential gene expression was analyzed in Snf2h lens-conditional knockout and wildtype newborn mouse eyeballs, with subsequent comparison of this data with the Brg1 lens-conditional knockout mouse eyes expression data (GSE25168).

overall_design: Four biological replicate experiments were performed.

first: Shuying

last: He

first: Juraj

last: Kokavec

first: Wanda

last: Kantorow

first: Lisa

last: Brennan

first: Vojtech

last: Kulvait

first: Ruth

last: Ashery-Padan

first: Arthur

last: Skoultchi

last: Zavadil

first: Marc

first: Tomas

last: Stopka

first: Ales

last: Cvekl


title: Expression data in Hepa1c1c7 cells treated with Nrf2 activator, diethyl maleate (DEM)

status: Public on Oct 03 2012

city: Sendai

first: Masayuki

last: Yamamoto

address: 2-1 Seiryo-machi, Aoba-ku

department: Medical Biochemistry

institute: Tohoku University Graduate School of Medicine

summary: To examine the gene expression profile of Hepa1c1c7 cells in response to DEM, we performed microarray analysis.

overall_design: Two independent RNA samples from Hepa1c1c7 cells (a mouse hepatoma cell line) treated with 100-µM DEM or DMSO for 6 hours were used for the microarray analysis.

first: Yosuke

last: Hirotsu

first: Fumiki

last: Katsuoka

first: James

middle: D

last: Engel


title: A combinatorial extracellular matrix platform identifies cell-extracellular matrix interactions that correlate with metastasis

city: Cambridge

first: Charles

middle: Arthur

last: Whittaker

institute: Koch Institute

summary: Extracellular matrix interactions play essential roles in normal physiology and many pathological processes. Here, we report a novel screening platform capable of measuring phenotypic responses to combinations of ECM molecules. While the importance of ECM interactions in metastasis is well documented, systematic approaches to identify their roles in distinct stages of tumorigenesis have not been described. Using a genetic mouse model of lung adenocarcinoma, we measured the ECM-dependent adhesion of tumor-derived cells.  Hierarchical clustering of adhesion profiles generated using this platform differentially segregated metastatic cell lines from primary tumor lines. Furthermore, we uncovered that metastatic cells selectively associate with fibronectin when in combination with galectin-3, galectin-8, or laminin. These interactions appear to be mediated in part by α3β1 integrin both in vitro and in vivo. We show that these galectins also correlate with human disease at both a transcriptional and histological level. Thus, our in vitro platform allowed us to interrogate the interactions of metastatic cells with their surrounding environment, and identified ECM and integrin interactions that could lead to therapeutic targets for metastasis prevention.

overall_design: Cell lines derived from murine lung primary adenocarcinomas and their metastases (Winslow et al., 2011 Nature 473:101-104)

first: N

last: Reticker-Flynn

first: D

last: Braga Malta

first: M

middle: W

last: Winslow

first: J

middle: M

last: Lamar

last: Xu

first: G

middle: H

last: Underhill

first: R

middle: O

last: Hynes

first: T

last: Jacks

first: S

last: Bhatia


title: Role of the Cytoskeleton in muscle transcriptional response to altered use

status: Public on Oct 06 2012

first: Gretchen

last: Meyer

address: 9500 Gilman Dr

department: Bioengineering

summary: Desmin is a cytoskeletal protein in muscle involved in integrating cellular space and transmitting forces.  In this study we sought to determine the combinatory effects of desmin deletion, aging and eccentric exercise on skeletal muscle at the transcriptional level across many pathways of muscle physiology.

overall_design: RNA was isolated from the TA muscle of mice of two genotypes (wildtype (WT) and desmin knockout (KO)) and two ages (7-9 weeks (Adult) and 12-14 months (Aged)). TA muscles were subjected to a bout of 50 eccentric contractions 12 hours prior to RNA isolation.  Numbers per group are as follows: WT_Adult (5), WT_Aged (5), KO_Adult (5), KO_Aged (4).

first: Richard

last: Lieber


title: Diet-Induced Obesity Exacerbates Inflammatory and Oxidative Stress Responses in Mice Exposed to Cigarette Smoke

status: Public on Sep 15 2012

city: Corvallis

first: Susan

middle: C.

last: Tilton

state: OR

address: 1007 ALS Bldg

department: AG Envr & Molec Toxicology

institute: Oregon State University

summary:
To identify biosignatures that describe these lifestyle susceptibility factors, we performed parallel exposures of regular weight (RW) C57BL/6 and diet-induced obese (DIO) C57BL/6 mice to cigarette smoke, either mainstream (MS) or sidestream (SS), mimicking both the smoker and environmental exposure through second-hand smoke, respectively.
Transcriptional responses were measured by global microarray analysis of lung tissue.

overall_design: Groups (N=8 biological replicates) of RW and DIO C57BL/6 mice (15-weeks old at start of exposures) were exposed to either filtered air (sham controls, SC), mainstream (MS) or sidestream (SS) cigarette smoke by nose-only inhalation exposure for 5 hr/day for a total of eight exposures over two weeks as follows: 5 consecutive days of exposure, followed by 2 days with no exposure, then three days of exposure, with necropsies occurring the day following the last exposure.

middle: C

first: Katrina

last: Waters


title: The Oscillating miRNA 959-964 cluster impacts Drosophila feeding time and other circadian outputs [expression].

status: Public on Nov 02 2012

city: Waltham

first: Stefan

last: Pescatore

address: 415 South Street

department: Biology

institute: Brandeis University / HHMI

summary:
Using high throughput sequencing of Drosophila head RNA, a small set of miRNAs that undergo robust circadian oscillations in levels were discovered. We concentrated on a cluster of six miRNAs, mir-959-964, all of which peak at about ZT12 or lights-off. The data indicate that the cluster pri-miRNA is transcribed under bona fide circadian transcriptional control and that all 6 mature miRNAs have short half-lives, a requirement for oscillating. Manipulation of food intake dramatically affects the levels and timing of cluster miRNA transcription with no more than minor effects on the core circadian oscillator. This indicates that the central clock regulates feeding, which in turn regulates proper levels and cycling of the cluster miRNAs. Viable Gal4 knock-in as well as cluster knock-out and over-expression strains were used to localize cluster miRNA expression as well as explore their functions. The adult head fat body is a major site of expression, and feeding behavior, innate immunity, metabolism, and perhaps stress responses are under cluster miRNA regulation. The feeding behavior results indicate that there is a feedback circuit between feeding time and cluster miRNA function as well as a surprising role of post-transcriptional regulation in these behaviors and physiology.
To address possible functions of the cluster miRNAs, the knock-out strain (mir959-952-KO) as well as a cluster over-expression strain (UAS-cluster, tim-gal4) was assayed for mRNA changes relative to their WT counterparts on Affymetrix expression arrays. The strategy was based on the observation that miRNAs often cause a decrease in the steady-state levels of their target mRNAs (Guo et al., 2010; Lim et al., 2005). Because of the circadian regulation and the possibility of non-fat body expression, the over-expression strain was generated with the broad circadian driver tim-gal4 rather than a fat body driver. To accommodate the possibility that important mRNA changes only appear at certain circadian times, RNA was assayed from heads collected at two different times, ZT4 and ZT16.

overall_design: Analyzed expression data was assayed from total RNA from Drosophila heads in two sample groups consisting of four samples (with eight samples in total): knock-out strain at ZT4 and ZT16, and its respective wildtype control at ZT4 and ZT16; cluster over-expression strain at ZT4 and ZT16 and its respective wildtype control at ZT4 and ZT16.

first: Sadanand

last: Vodala

first: Joseph

last: Rodriguez

first: Marita

last: Buescher

first: Ya-Wen

last: Chen

first: Ruifen

last: Weng

first: Stephen

last: Cohen

first: Michael

last: Rosbash


title: Dll1 marks early secretory progenitors in gut crypts that can revert to stem cells upon tissue damage

status: Public on Sep 27 2012

city: Dresden

first: Daniel

middle: E.

last: Stange

address: Fetscherstr. 74

country: Germany

institute: University Clinic Dresden

summary: The introduction of GFP into the Dll1 locus has made it possible to isolate and study primary intestinal Dll1 positive cells, which constitute the secretory cell precursors of the intestine. Applying DNA array technology, we profiled the RNA changes of FACS-sorted Dll1_high/CD24_low, Dll1_high/CD24_medium and Dll1_high/CD24_high cells .

overall_design: We used cell fractions of intestines from Dll1-EGFP-ires-CreERT2 mice, expressing GFP under the control of the Dll1 promoter. RNA was isolated from three FACS sorted cell populations: all are expressing Dll1-GFP at high levels, but are expressing different levels of Cd24 (low, medium and high). For this set two biological replicates were generated. One additional fraction was isolated from Lgr5-EGFP-ires-CreERT2 mice, expressing GFP under the control of the Lgr5 promoter. This cell fraction contains intestinal stem cells.   Differentially labelled cRNA from these cell fractions was hybridized against a reference (RNA isolated from whole intestine) on 4X44K Agilent Whole Mouse Genome dual colour Microarrays (G4122F).

first: Marc

last: van de Wetering

first: Toshiro

last: Sato

first: Johan

last: van Es

first: Hans

last: Clevers


title: Diet-Induced Obesity Reprograms the Inflammatory Response of the Murine Lung to Inhaled Endotoxin

summary:
To identify key biological pathways that define susceptibility factors for pulmonary infection during obesity, diet-induced obese (DIO) and regular weight (RW) C57BL/6 mice were exposed to 0.5 µg/L inhaled lipopolysaccharide (LPS) for 1 hr/d for 4 days over a period of 2 weeks.
Transcriptional responses were measured by global microarray analysis of lung tissue.

overall_design: Groups (N=8 biological replicates) of regular weight (RW) and diet-induced obese (DIO) C57BL/6 mice (15-weeks old at start of exposures) were exposed to either filtered air (sham controls, SC) or 0.5 µg/L LPS by nose-only inhalation exposure for 1 hr/day for 4 days over a 10-day period with necropsies occurring on the day following the last exposure (Day 11).


title: Regulation of antibacterial defense in the small intestine by the nuclear bile acid receptor

status: Public on Sep 12 2012

first: Scott

middle: Andrew

last: Ochsner

address: One Baylor Plaza

department: Molecular and Cellular Biology

summary: Decreased bile secretion in rodents by either ligation of the common bile duct or induction of cirrhosis causes changes in the small intestine, including bacterial overgrowth and translocation across the mucosal barrier. Oral administration of bile acids inhibits these effects. The genes regulated by FXR in ileum suggested that it might contribute to the enteroprotective actions of bile acids. To test this hypothesis, mice were administered either GW4064 or vehicle for 2 days and then subjected to bile duct ligation (BDL) or sham operation. After 5 days, during which GW4064 or vehicle treatment was continued, the mice were killed and their intestines were analyzed for FXR target gene expression.

overall_design: Mice were treated with or without FXR ligand GW4064 for 2 days prior to bile duct ligation surgery and for 5 days after surgery.  After 5 days the mice were sacrificed and the ileum collected and processed for gene expression analysis. Gene expression in the ilium from each sample group was assayed in duplicate using Affymetrix Mouse Genome 430A 2.0 Gene Chips.

first: Takeshi

last: Inagaki

first: Steven

middle: A

last: Kliewer


title: Microarray analysis of genes regulated by SHP in wild-type and ob/ob mice.

summary: Liver gene expression was compared in wild type, SHP knockout, ob/ob, and ob/ob SHP double mutant mice.

overall_design: A total of eight samples were analyzed using Affymetrix arrays.  Each of the four genotypes studied were assayed in duplicate.

first: Li

last: Wang

first: David

last: Moore


title: Pulmonary gene expression in MutaTMMouse orally gavaged with three doses of benzo(a)pyrene and vehicle control

city: Ottawa

first: Andrew

last: Williams

state: ON

address: 50 Columbine

institute: Health Canada

summary: In this study, we investigate mRNA profiles in lung of mice exposed to benzo(a)pyrene. Male mice were exposed to three doses (25, 50, and 75 mg/kg/day BaP) for 28 days and profiles were examined three days post-exposure. Our analyses reveal that BaP causes pulmonary specific cellular transformation indicative of carcinogenesis.

overall_design: Individual total RNA (200 ng) from 5 mice per treatment group (control, 25mg/kg/day, 50mg/kg/day, and 75mg/kg/day) and universal reference total RNA (Stratagene, Mississauga, ON, Canada) was used to synthesize double stranded cDNA.

first: Sarah

last: Labib

first: Volker

last: Arlt

first: Paul

last: White

first: Sabina

last: Halappanavar


title: Impact of Cux2 knockdown on gene expression in female mouse liver (Mus musculus)

status: Public on Sep 10 2012

middle: J.

last: Waxman

address: 5 Cummington Mall

department: Department of Biology and Bioinformatics Program

institute: Boston University

summary: We identified genes expressed in mouse liver that are regulated by Cux2, a highly female-specific liver transcription factor whose expression is regulated by sex-dependent plasma GH patterns.  Using siRNA to knockdown Cux2 expression in female liver, we show that female specific genes are predominantly repressed by Cux2 knockdown.  In contrast, similar numbers of male-biased genes are repressed as are induced by Cux2 knockdown.  A scrambled, non-specific siRNA was used as a control.  (Published in: TL Conforto et al 2012, Mol Cell Biol. 2012, 32:4611-4627. PubMed PMID: 22966202; PMCID: PMC3486175)

overall_design: Liver RNA isolated from the following 3 groups of mice was used in the present study:  (1) 8 wk old female mice treated with non-specific siRNA control (n = 13; 6 or 7 per each pool); (2) 8 wk old female mice treated with Cux2 siRNA and euthanized 5 days later (n = 5; 2 or 3 per each pool); (3) 8 wk old female mice treated with Cux2 siRNA and euthanized 8 days later (n = 4; 2 per each pool).  These RNA pools were used in two separate sets of competitive hybridization experiments: 1) 8 wk non-specific siRNA treated vs. 8 wk Cux2 siRNA treated for 5 days; 2) 8 wk non-specific siRNA treated vs. 8 wk Cux2 siRNA treated for 8 days.  Fluorescent labeling of RNA and hybridization of the Alexa 555-labeled (green) and Alexa 647-labeled (red) RNA samples to Agilent Mouse Gene Expression 4x44k v1 microarrays (Agilent Technology, Palo Alto, CA; catalog # G4122F-014868) were carried out, with dye swapping for each of the two hybridization experiments to eliminate dye bias.  Two microarrays, one for each mixed cDNA sample, were hybridized for each of the two fluorescent reverse pairs, giving a total of 4 microarrays.

first: Tara

middle: L

last: Conforto

first: Jennifer

last: Sherman

middle: J


title: Impact of Cux2 overexpression on gene expression in male and female mouse liver (Mus musculus)

summary: We identified genes expressed in mouse liver that are regulated by Cux2, a highly female-specific liver transcription factor whose expression is regulated by sex-dependent plasma GH patterns.  Using adenovirus to overexpress Cux2 (Adeno-Cux2) in male liver, we show that Cux2 represses ~35% of male-biased genes and induces/de-represses ~35% of female-biased genes. Adeno-CMV was used as a control for adenoviral infection.  (Published in: TL Conforto et al 2012, Mol Cell Biol. 2012, 32:4611-4627. PubMed PMID: 22966202; PMCID: PMC3486175)

overall_design: Liver RNA isolated from the following eight groups of mice was used in the present study:  (1) 8 wk old untreated male (M) mice (n = 10; 5 per each pool); (2) 8 wk old untreated female mice (F) mice (n = 11; 5 or 6 per each pool);  (3) 8 wk old male mice treated with Adeno-Cux2 and euthanized 5 days later (n = 12; 6 per each pool); (4) 8 wk old female mice treated with  Adeno-Cux2 and euthanized 5 days later (n = 8; 4 per each pool); (5) 8 wk old male mice treated with Adeno-CMV and euthanized 5 days later (n = 13; 6 or 7 per each pool); (6) 8 wk old female mice treated with Adeno-CMV and euthanized 5 days later (n = 7; 3 or 4 per each pool); (7) 8 wk old male mice treated with Adeno-Cux2 and euthanized 3 days later (n=11; 5 or 6 per each pool);  (8) 8 wk old  male mice treated with Adeno-CMV and euthanized 3 days later (n=11; 5 or 6 per pool).  These RNA pools were used in four separate sets of competitive hybridization experiments: 1) 8 wk untreated M vs. 8 wk untreated F; 2) 8 wk M + Ad-Cux2 (5 day) vs. 8 wk M + Ad-CMV (5 day); 3) 8 wk F + Ad-Cux2 (5 day) vs. 8 wk F +  Ad-CMV (5 day); 4) 8 wk M + Ad-Cux2 (3 day) vs. 8 wk M +  Ad-CMV (3 day).  Fluorescent labeling of RNA and hybridization of the Alexa 555-labeled (green) and Alexa 647-labeled (red) RNA samples to Agilent Mouse Gene Expression 4x44k v1 microarrays (Agilent Technology, Palo Alto, CA; catalog # G4122F-014868) were carried out, with dye swapping for each of the three hybridization experiments to eliminate dye bias.  Two microarrays, one for each mixed cDNA sample, were hybridized for each of the four fluorescent reverse pairs, giving a total of 8 microarrays.

first: Yijing

last: Zhang


title: Comparison of gene expression profiles of effector T cells from leptin-receptor-deficient mice and wild type mice

status: Public on Sep 05 2012

city: Tampere

first: Dario

last: Greco

address: Arvo ylpön Katu 34

department: Faculty of Medicine and Health Technology

country: Finland

institute: Tampere University

summary: We have employed whole genome microarray expression profiling as a discovery platform to identify genes differentially expressed during leptin receptor deficiency, which is partly responsible for the inhibition of T cell effector functions. Mouse CD4+CD25- T cells were purified from leptin-receptor-deficient mice and wild type mice, and stimulated with anti-CD3 and anti-CD28 for 12h. Six mice of each genotype were used.

overall_design: Leptin receptor deficiency-induced gene expression profiles were evaluated in mouse effector T cells from db/db mice and compared to the ones observed in their WT counterpart (db/+ mice). Two independent experiments were performed, and in each of them, 3 mice/group were pooled.

first: Claudio

last: Procaccini

first: Giuseppe

last: Matarese


title: Muscle Involvement in Preservation of Metabolic Flexibility by Treatment using n-3 PUFA or Rosiglitazone in Dietary-Obese Mice

status: Public on Sep 07 2012

city: Wageningen

first: Evert

middle: M.

last: van Schothorst

address: De Elst 1

institute: Wageningen University

summary: Impaired resistance to insulin, the key defect in type 2 diabetes (T2D), is associated with a low capacity to adapt fuel oxidation to fuel availability, i.e., metabolic inflexibility. The hampered metabolic adaptability triggers a further damage of insulin signaling. Since skeletal muscle is the main site of glucose uptake, effectiveness of T2D treatment depends in large on the improvement of insulin sensitivity and metabolic adaptability of the muscle. We have shown previously in mice fed an obesogenic high-fat diet that a combination treatment using n-3 long-chain polyunsaturated fatty acids (n-3 LC-PUFA) and thiazolidinedione (TZD) anti-diabetic drugs preserved metabolic health and synergistically improved muscle insulin sensitivity. We investigated here whether TZD rosiglitazone could elicit the additive beneficial effects on metabolic flexibility when combined with n-3 LC-PUFA. Adult male C57BL/6N mice were fed an obesogenic corn oil-based high-fat diet (cHF) for 8 weeks, or randomly assigned to various dietary treatments: (i) cHF+F, cHF with n-3 LC-PUFA concentrate replacing 15% of dietary lipids; (ii) cHF+ROSI, cHF with 10 mg rosiglitazone/kg diet; and (iii) cHF+F+ROSI, or chow-fed. Indirect calorimetry demonstrated superior preservation of metabolic flexibility to carbohydrates in response to the combination treatment. Metabolomic and gene expression analyses in the muscle suggested distinct and complementary effects of the single treatments, with rosiglitazone augmenting insulin sensitivity by the modulation of branched-chain amino acid metabolism, and n-3 LC PUFA supporting complete oxidation of fatty acids in mitochondria. These beneficial metabolic effects were associated with the activation of the switch between glycolytic and oxidative muscle fibers, especially in the cHF+F+ROSI mice. Our results further support the idea that the combination treatment using n-3 LC-PUFA and TZDs could improve the efficacy of the treatment of obese and diabetic patients.

overall_design: Male C57BL/6N mice had free access to water and Chow. Three-month-old mice were randomly assigned (8 animals per group) to cHF diet (lipid content ~35% wt/wt) or to the following 'treatments' by isocaloric cHF-based diets, namely (i) cHF+F, cHF diet supplemented with n-3 LC-PUFA concentrate (46% DHA, 14% EPA, wt/wt, as triglycerides; product EPAX 1050 TG), which replaced 15% wt/wt of dietary lipids; (ii) cHF+ROSI, cHF diet supplemented with 10 mg rosiglitazone/kg diet; and (iii) cHF+F+ROSI, cHF diet supplemented with both n-3 LC-PUFA concentrate and rosiglitazone. cHF+ROSI at a higher dose, namely 100 mg rosiglitazone/kg diet was also included in the study, but not in the final microarray analysis. The treatment lasted for 8 weeks, whereafter  the animals were first fasted for 10 hours during the light phase of the day (between 8.00hr and 18.00hr), than re-fed Chow (starting at 18.00hr), and killed the following day by cervical dislocation under pentobarbital anesthesia (between 9.00hr and 11.00hr); the so-called 'diet-switch protocol'. Gastrocnemius muscle was isolated and used for total RNA isolation.

first: Dasa

last: Medrikova

first: Annelies

last: Bunschoten

first: Pavel

last: Flachs

first: Jan

last: Kopecky

first: Jaap

last: Keijer


title: Rapid estrogen receptor signaling is essential for the protective effects of estrogen against vascular injury

status: Public on Sep 01 2012

first: Gavin

middle: R.

last: Schnitzler

address: 800 Washington St. Box 80

department: Molecular Cardiology Research Institute

institute: Tufts Medical Center

summary:
Background: Clinical trial and epidemiological data support that the cardiovascular effects of estrogen are complex, including a mixture of both potentially beneficial and harmful effects. In animal models, estrogen protects females from vascular injury and inhibits atherosclerosis. These effects are mediated by estrogen receptors (ERs), which when bound to estrogen can bind to DNA to directly regulate transcription. ERs can also activate several cellular kinases by inducing a “rapid” non-nuclear signaling cascade. However, the biologic significance of this rapid signaling pathway has been unclear.

Methods and Results: Here, we develop a novel transgenic mouse in which rapid signaling is blocked by over-expression of a peptide that prevents ERs from interacting with the scaffold protein, striatin (the Disrupting Peptide Mouse, DPM). Microarray analysis of ex vivo-treated mouse aortas demonstrates that rapid ER signaling plays an important role in E2-mediated gene regulatory responses.  Disruption of ER-striatin interactions also eliminates the ability of E2 to stimulate cultured endothelial cell migration and to inhibit cultured vascular smooth muscle cell growth. The importance of these findings is underscored by in vivo experiments demonstrating loss of estrogen-mediated protection against vascular injury in the DPM mouse following carotid artery wire injury.

Conclusions: Taken together, these results support that rapid, non-nuclear ER signaling contributes to the transcriptional regulatory functions of ER, and is essential for many of the vasoprotective effects of estrogen. These findings also identify the rapid ER signaling pathway as a potential target for the development of novel therapeutic agents.

overall_design: Transgenic disrupting peptide mice (DPM) express the Estrogen Receptor (ER) alpha amino acids 176-253 peptide under CMV promoter control. This peptide blocks interactions between ER and striatin, which inhibits rapid non-genomic signaling by ER to cellular kinases. Female WT and DPM mice (4 mice per sample) were overiectomized, and 1 week later aortas were harvested and treated + 10 nM beta-estradiol (E2) or + EtOH vehicle (Veh) for 4 hrs ex vivo. Total RNA was collected, reverse transcribed to cDNA and used to probe Affymetrix mouse 430.A 2.0 arrays.

first: Sophie

last: Bernelot Moens

middle: R

first: Moriah

last: Nickerson

first: Huiming

last: Guo

first: Kazutaka

last: Ueda

first: Qing

last: Lu

first: Mark

last: Aronovitz

first: Heather

first: Wendy

last: Baur

first: Ulla

last: Hansen

first: Lakshmanan

middle: K

last: Iyer

first: Richard

last: Karas


title: Muscle Involvement in Preservation of Metabolic Flexibility by a Combination Treatment using n-3 PUFA and Rosiglitazone in Dietary-Obese Mice

first: Evert

last: van Schothorst

summary: Impaired resistance to insulin, the key defect in type 2 diabetes (T2D), is associated with a low capacity to adapt fuel oxidation to fuel availability, i.e., metabolic inflexibility. The hampered metabolic adaptability triggers a further damage of insulin signaling. Since skeletal muscle is the main site of glucose uptake, effectiveness of T2D treatment depends in large on the improvement of insulin sensitivity and metabolic adaptability of the muscle. We have shown previously in mice fed an obesogenic high-fat diet that a combination treatment using n-3 long-chain polyunsaturated fatty acids (n-3 LC-PUFA) and thiazolidinedione (TZD) anti-diabetic drugs preserved metabolic health and synergistically improved muscle insulin sensitivity. We investigated here whether TZD rosiglitazone could elicit the additive beneficial effects on metabolic flexibility when combined with n-3 LC-PUFA. Adult male C57BL/6J mice were fed an obesogenic corn oil-based high-fat diet (cHF) for 8 weeks, or randomly assigned to various dietary treatments: (i) cHF+F, cHF with n-3 LC-PUFA concentrate replacing 15% of dietary lipids; (ii) cHF+ROSI, cHF with 10 mg rosiglitazone/kg diet; and (iii) cHF+F+ROSI, or chow-fed. Indirect calorimetry demonstrated superior preservation of metabolic flexibility to carbohydrates in response to the combination treatment. Metabolomic and gene expression analyses in the muscle suggested distinct and complementary effects of the single treatments, with rosiglitazone augmenting insulin sensitivity by the modulation of branched-chain amino acid metabolism, and n-3 LC PUFA supporting complete oxidation of fatty acids in mitochondria. These beneficial metabolic effects were associated with the activation of the switch between glycolytic and oxidative muscle fibers, especially in the cHF+F+ROSI mice. Our results further support the idea that the combination treatment using n-3 LC-PUFA and TZDs could improve the efficacy of the treatment of obese and diabetic patients.

overall_design: Male C57BL/6N mice had free access to water and Chow. Three-month-old mice were randomly assigned (8 animals per group) to cHF+ROSI diet (lipid content ~35% wt/wt) supplemented with  10 mg rosiglitazone/kg diet, or cHF+F+ROSI in which n-3 LC-PUFA concentrate (46% DHA, 14% EPA, wt/wt, as triglycerides; product EPAX 1050 TG) replaced 15% wt/wt of dietary lipids. The treatment lasted for 8 weeks, whereafter  the animals were first fasted for 10 hours during the light phase of the day (between 8.00hr and 18.00hr), than re-fed Chow (starting at 18.00hr), and killed the following day by cervical dislocation under pentobarbital anesthesia (between 9.00hr and 11.00hr); the so-called 'diet-switch protocol'. Gastrocnemius muscle was isolated and used for total RNA isolation.

first: Dasa


title: Muscle Involvement in Preservation of Metabolic Flexibility by Treatment using n-3 PUFA, Rosiglitazone, and a combination of both

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

overall_design: Refer to individual Series


title: Treg specific Gata3 knock out array

status: Public on Aug 28 2012

city: New York

first: Aaron

last: Arvey

state: NY

institute: Memorial Sloan-Kettering Cancer Center

summary:
The transcription factor Foxp3 is indispensible for the differentiation and function of regulatory T cells (Treg cells). To gain insights into the molecular mechanisms of Foxp3 mediated gene expression we purified Foxp3 complexes and explored their composition. Biochemical and mass-spectrometric analyses revealed that Foxp3 forms multi-protein complexes of 400-800 kDa or larger and identified 361 associated proteins ~30% of which are transcription-related. Foxp3 directly regulates expression of a large proportion of the genes encoding its co-factors.  Reciprocally, some transcription factor partners of Foxp3 facilitate its expression. Functional analysis of Foxp3 cooperation with one such partner, Gata3, provided further evidence for a network of transcriptional regulation afforded by Foxp3 and its associates to control distinct aspects of Treg cell biology.
Gene expression profile of Treg specific knock-out of Gata3 vs. their littermate controls were analyzed to gain insight into Gata3 dependendent genes in Treg cells.

overall_design: Treg cells (CD4+CD25+YFP+) sorted from Gata3F/FFoxp3YFP-Cre and Gata3F/+Foxp3YFP-Cre mice (three mice in each group) followed by microarray analyses.

first: Dipayan

last: Rudra

first: Alexander

last: Rudensky


title: Gene expression profiling along the intestinal crypt axis.

status: Public on Aug 15 2012

first: Daniel

last: Stange

summary: The identification of Lgr5 as an intestinal stem cell marker has made it possible to isolate and study primary intestinal stem cells. Transcriptional differences between intestinal stem cells can be explored by the use of the Lgr5-eGFP-ires-CreERT2 knock-in mouse. In this mouse model GFP expression is driven from the Lgr5 locus, leading to highest GFP levels in the Lgr5 positive cells. Yet, due to the stability of the GFP protein, it is distributed upon cell division to the daughter cells. Here we arbitrarily sorted crypt cells based on GFP expression into five fractions. The fraction with the lowest GFP level is named 1+, the fraction with the highest level 5+.

overall_design: We used cell fractions of intestines from Lgr5-EGFP-ires-CreERT2 mice, expressing GFP under the control of the Lgr5 promoter. RNA was isolated from five FACS sorted cell populations, with fraction 5+ expressing GFP at highest levels and 1+ expressing GFP at lowest levels. Differentially labelled cRNA from fractions 1+ to 4+ were hybridized against fraction 5+ on 4X44K Agilent Whole Mouse Genome dual colour Microarrays (G4122F) in dye swap experiments, resulting in eight individual arrays.

middle: E

first: Marc

last: van de Wetering

first: Hans

last: Clevers


title: Differentiating Mouse Colon and Rectum: Transcriptional Differences and Environmental Exposure

status: Public on Aug 20 2012

city: Madison

first: Peter

last: Geiger

address: 1400 University Ave

department: Surgery

institute: UW-Madison

summary: We have demonstrated that the colon and rectum are transcriptionally distinct in mice and that the colon can be differentiated from rectum based on microarray profiles.  This transition occurs over the first weeks of life, indicating environmental exposure is an important mediator.

overall_design: Colon and rectum RNA from E18 to 8 week old C57BL/6J mice along with 8 week old gnotobiotic C57BL/6J mice were evaluated with microarray technology.  For each array analysis a pool of RNA(2-8) from individual colons or rectums were co-hybridized with a reference pool of 4 RNA samples from mouse small intestine, rectum and colon (SIRC)) to Agilent Mouse Whole Genome 4x44k microarrays and scanned on an Agilent microarray scanner.  Files were extracted using Agilent Feature Extraction version 9.5.

first: Sean

last: Ronnekleiv-Kelly

middle: G

first: Gregory

middle: D

last: Kennedy


title: Gene expression profiles of effector T cells

first: Dario

last: Greco

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus