title: Particulate Matter effect on Mouse Model of Cardiac Failure: Lung and Heart Left Ventricle

status: Public on Aug 10 2009

city: Charlottesville

first: Yong

last: Huang

address: 1340 Jefferson Park Ave

department: Medicine

institute: University of Viginia

summary:
Particulate Matter Triggers Carotid Body Dysfunction, Respiratory Dysynchrony and Cardiac Arrhythmias in Mice with Cardiac Failure

The mechanistic link between human exposure to airborne particulate matter (PM) pollution and the increased cardiovascular morbidity and mortality observed in people with congestive heart failure (CHF) is unknown. We now show that exposure of genetically-engineered mice with CHF (expressing a cardiac-specific CREB mutant transcription factor) to ambient PM (collected in Baltimore, mean aerodynamic diameter 1.9 um) unmasks severe autonomic morbidities manifested as significant reductions in heart rate variability, respiratory dysynchrony and increased frequency of serious ventricular arrhythmias, features not observed in PM-challenged wild type mice without CHF. PM exposure in CREB mice with CHF reflexly triggers autonomic dysfunction via heightened carotid body function as evidenced by pronounced afferent nerve responses to hypoxia and marked depression of breathing by hyperoxia challenge. Genomic analyses of lung and ventricular tissues revealed PM-induced molecular signatures of inflammation and oxidative stress. These findings in a murine model of cardiac failure provide the first direct assessment of autonomic function in response to PM challenge and are highly consistent with current epidemiologic findings on cardiovascular morbidity in susceptible PM-exposed human populations.
We utilized a murine model of dilated cardiomyopathy to address potential mechanistic links between PM exposure and the development of life-threatening cardiac dysrhythmias.

overall_design: four group (n=3) of animals were treated by PBS or particulate matter (20mg/kg 1.9µm particulate matter) in Wild type or CD-1 dominate negative mice

first: Gabriel

middle: D

last: Lang

first: Liliana

last: Moreno-Vinasco

first: Ting

last: Wang

first: Sascha

middle: N

last: Goonewardena

first: Ying

last: Peng

first: Eric

last: Svensson

first: Viswanathan

last: Natarajan

first: Roberto

first: John

last: Beshai

first: Patrick

last: Breysse

first: Allison

last: Geyh

first: Jonathan

last: Samet

first: Yves

last: Lussier

first: Nanduri

last: Prabhakar

first: Joe

last: Garcia


title: Distinct Self-renewal and Differentiation Phases in the Niche of Infrequently Dividing Hair Follicle Stem Cells

status: Public on Aug 07 2009

city: Ithaca

first: Tudorita

last: Tumbar

state: NY

address: 260 Biotechnology Building

department: Molecular Biology & Genetics

country: USA

institute: Cornell University

summary:
In homeostasis of adult vertebrate tissues, stem cells are thought to self-renew by infrequent and asymmetric divisions that generate another stem cell daughter and a progenitor daughter cell committed to differentiate.  This model is based largely on in vivo invertebrate or in vitro mammal studies.  Here we examine the dynamic behaviour of adult hair follicle stem cells in their normal setting by employing mice with repressible H2B-GFP expression to track cell divisions and Cre inducible mice to perform long-term single cell lineage tracing. We provide direct evidence for the infrequent stem cell division model in intact tissue.  Moreover, we find that differentiation of progenitor cells occurs at different times and tissue locations than self-renewal of stem cells.  Distinct fates of differentiation or self-renewal are assigned to individual cells in a temporal-spatial manner. We propose that large clusters of tissue stem cells behave as populations, whose maintenance involves unidirectional daughter-cell fate decisions.
We used microarrays to expression profile the stem cell progeny generated at distinct differentiation and self-renewal stages.

overall_design: We employed double transgenic mice, K5tTA x pTRE-H2B-GFP in which an epithelial Keratin 5 (K5) promoter drove repressible histone H2B-GFP expression. Repression is achieved by feeding the mice doxycycline for a period of time (chase), when the H2B-GFP dilutes in cells by 2-fold at division. This allowed us to quantify precise proliferation history in vivo, from the amount of H2B-GFP fluorescence retained in cells after chase. After various chase periods we sacrificed mice at different ages corresponding to distinct phases of hair cycle. We isolated skin cell suspensions from these mice and stained them for surface expression of stem cell niche bulge(Bu) markers CD34 and alpha 6-integrin.  Fluorescence-activated cell sorting (FACS) revealed histograms with distinct peaks of 2-fold median H2B-GFP intensity corresponding to distinct divisions. For microarray, we FACS sorted bulge(Bu) and non-bulge(NonBu) cells based cell surface marker staining and their proliferation history with H2B-GFP intensity, after two doxycycline chase schemes (Postnatal Day 18-PD21 or PD22-PD25), then extract total RNA from each cell fractions. For microarrays 5 ng of high quality RNA (Bioanalyzer, Agilent) were amplified (Ovation Amplification; Nugene) in the Cornell Microarray Core Facility. GeneChip IVT labelling was followed by Gene Chip MOE 430 2.0 hybridization, GeneArray 3000 scanning, GCOS generation of present calls and signal values (Affymetrix).

middle: V

last: Zhang


title: Affymetrix gene expression AID-GFP-positive vs AID-GFP-negative

status: Public on Aug 15 2009

city: Los Angeles

first: Markus

last: Müschen

state: CA

address: 4650 Sunset Blvd

department: Childrens Hospital Los Angeles

institute: University of Southern California


title: Foxp3-deficient Treg cells do not revert into conventional Effector CD4+ T cells but constitute a unique cell subset

status: Public on Aug 28 2009

city: Augusta

first: Piotr

last: Kraj

state: GA

address: 1120 15th Street

department: Centr. for Biotech. and Gen. Medicine

institute: Medical College of Georgia

summary:
Gene expression profiles were compared between regulatory T cells (Treg) and Effector CD4+ T cells in healthy B6 mice and sick mice with scurfy mutation.
We used microarrays to elucidate the molecular mechanisms underlying the suppression function of the Treg cells.

Keywords: Cell type comparison

overall_design: To identify candidate genes that might be related to the suppressive activity, the Treg cells expressing functional and mutant Foxp3 transcription factor were compared


title: Liver Transcriptome Profiles Associated with Strain-Specific Ehrlichia chaffeensis–induced Hepatitis

status: Public on Aug 17 2009

city: Columbus

first: Koshiro

last: Miura

state: OH

address: 1900 Coffey Rd

department: Veterinary Biosciences

institute: The Ohio State University

summary: Infection of humans with Ehrlichia chaffeensis, the etiologic agent of human monocytic ehrlichiosis, can cause hepatitis of varying severity. When the three human isolates of E. chaffeensis, each belongs to different geno-groups, are inoculated into severe combined immunodeficiency mice, the severity of clinical signs and bacterial burden detected in the liver are strain Wakulla>Liberty>Arkansas. Disseminated and granulomatous inflammation is evident in the liver of mice infected with strains Wakulla and Arkansas, respectively, but not in mice infected with strain Liberty. In this paper, we used microarray analysis to define transcriptional profiles characteristic to the histopathological features in the mouse liver. Cytokine and chemokine profiles were strikingly different among three strains of E. chaffeensis: IFN-γ, CCL5, CXCL1, CXCL2, CXCL7 and CXCL9 were highly up-regulated with strain Arkansas, TNF-α, CCL2, CCL3, CCL5, CCL6, CCL12, CCL20, CXCL2, CXCL7, CXCL9 and CXCL13 were highly up-regulated with strain Wakulla. With strain Liberty, only CXCL13 was highly up-regulated.  In the livers infected with the Arkansas strain, monocytes/macrophages and NK cells were enriched in the granulomas and increase of NK cell-marker mRNAs was detected. Livers infected with the Wakulla strain displayed infiltration of significantly more neutrophils and increase of neutrophil-marker mRNAs. Genes up-regulated commonly in the liver infected with the three stains are other host innate immune and inflammatory response genes including several acute phase proteins. Genes down-regulated commonly are related to host physiologic functions. The results suggest that marked modulation of host cytokine and chemokine profiles by E. chaffeensis strains underlie the distinct host liver disease.

overall_design:
E. chaffeensis strains Arkansas, Wakulla and Liberty were propagated in DH82 cells.  Three to four week-old male ICR-scid mice were infected intraperitoneally with 10^6 E. chaffeensis-infected DH82 cells (typically >95% infected) containing approximately the same number of bacteria. At day 15 post infection (PI) mice were euthanized and livers were harvested.
Total RNAs were extracted from E. chaffeensis-infected or mock-infected (uninfected) mouse liver. About 8 ug of each RNA sample was applied to double-strand cDNA preparation. The labeled cRNA was hybridized to GeneChip Mouse Genome 430 2.0 Array (Affymetrix). After washing and staining with streptoavidin phycoerythrin, the array was scanned.
Three mice were examined for each of the 4 groups.

first: Yasuko

last: Rikihisa


title: HSF4 deficient lens of newborn mice

status: Public on Aug 11 2009

city: Shanghai

first: Xiangyin

last: kong

address: No.225 yueyang road

department: molecular genetics

country: China

institute: Institute of health science

summary: Microarray Analyses of Newborn Mouse lens lacking HSF4. Hsf4 is essential for lens development.

overall_design: Newborn Mouse lens expression pattern of HSF4-/- and wildtype.


title: mRNA expression in Zfp36L2 knockout E14.5 fetal liver

status: Public on Aug 06 2009

city: RTP

first: NIEHS

last: Microarray Core

state: NC

address: 111 T.W. Alexander Drive

department: DIR

institute: NIEHS

summary: ZFP36L2, zinc finger protein 36, C3H type-like 2 (also known as Brf2, Erf2, Tis11D) is a member of the tristetraprolin (TTP; Zfp36) family of tandem CCCH zinc finger proteins that can bind to AU-rich elements (AREs) in the 3'-untranslated region of mRNAs, leading to their deadenylation and subsequent degradation.  We have generated Zfp36l2 knockout mice.  Knockout mice were born at the expected Mendelian frequency, but within several weeks of birth they died rather suddenly with pallor and frequent intestinal hemorrhage.  These mice exhibited pancytopenia, decreased hematopoietic progenitor cells from fetal liver and yolk sac, and ineffective hematopoietic stem cells.  Since ZFP26L2 is likely to function as an ARE-containing mRNA destabilizing protein, we were interested in identifying any abnormally stabilized transcripts in fetal livers from the Zfp36l2 knockout mice whose protein product may directly or indirectly affect hematopoietic stem cell function.

overall_design: Total RNA was isolated from individual embryonic day 14.5 fetal livers.  Five wild type and five knockout fetal livers were used to generate 5 wild type replicates and 5 knockout replicates.  RNA expression from knockout fetal livers was compared to wild type fetal livers.

first: Deborah

middle: J

last: Stumpo

first: Hal

middle: E

last: Broxmeyer

first: Toni

last: Ward

first: Scott

last: Cooper

first: Giao

last: Hangoc

first: Peter

last: Aplan

first: Yang

last: Chung

first: Mervin

middle: C

last: Yoder

first: William

last: Shelley

middle: K

last: Richfield

first: Manas

last: Ray

first: Perry

last: Blackshear


title: Gene expression in murine acute lymphoblastic leukemia in vivo after allogeneic or syngeneic bone marrow transplantation

city: Rochester

first: Craig

middle: A.

last: Mullen

address: 601 Elmwood Ave., Box 777

department: Pediatrics

institute: University of Rochester

summary: This study compared gene expression in murine bcr-abl positive acute lymphoblastic leukemia cells in vivo in allogeneic BMT recipients compared to syngneneic BMT recipients.

overall_design: The goal of the experiment was to compare gene expression in leukemia cells that were in either an allogeneic transplant (5 samples) or syngeneic transplant (5 samples) immune environment in vivo. The bone marrow transplant model involved preparation of C57BL/6 recipients with total body irradiation and 5-fluoruracil. These recipients were then infused IV with either allogeneic C3.SW marrow and spleen cells or syngeneic C57BL/6 marrow and spleen cells. The leukemia cells were mixed in with the marrow and spleen cells.  After a few weeks (2-3) the C57BL/6 leukemia cells were purified by flow cytometry and passed into other freshly transplanted animals. After three serial transplants the leukemia cells were flow purified and RNA prepared. Leukemia was C57BL/6 background (cells contain the human p210 bcr/abl oncogenic fusion gene and also have a deletion in the Ink4a/Arf locus, see: Biology of Blood and Marrow Transplantation. 14:622-630, 2008).

middle: A


title: Expression data from mouse ES and iPS cells

status: Public on Aug 03 2009

first: Fanyi

last: Zeng

address: 24/1400 West Beijing Road

department: Institute of Medical Genetics

institute: Shanghai Jiao Tong University

summary: Induced pluripotent stem (iPS) cells were produced from reprogramming of somatic cells, and they are shown to possess pluripotent properties similar to embryonic stem (ES) cells. Here we used microarrays to detail the global expression pattern among the ES cells and iPS cells, as well as the original mouse embryo fibroblast (MEF), to identify important players involved in the reprogramming process.

overall_design: Mouse ES cell cultures, as well as selected iPS cell lines and the original MEF cells they were derived from, were used for RNA extraction and hybridization on Affymetrix microarrays. Three biological replicates for each sample were processed.  GeneChips were processed and data were analyzed as previously described (Zeng et al., Dev Biol 20004).

first: Qi

last: Zhou


title: Reduced levels of protein tyrosine phosphatase CD45 protect mice from the lethal effects of Ebola virus infection

status: Public on Aug 20 2009

city: Frederick

first: Brian

middle: D.

last: Peyser

state: MD

address: 1425 Porter St

department: Bacteriology

institute: USAMRIID

summary: To gain insight into the changes in gene expression pattern upon Ebola infection, CD45+/+ (100% protein level) and CD45+/- (62% protein level) mice were challenged with mouse adapted Ebola virus. At time-points day 0, 1, 3, 5, 7, 9, 11 and 13, spleen tissue was harvested and splenocytes isolated. Total RNA was isolated for mRNA expression analysis. The mouse genome 430 2.0 array (Affymetrix, Inc.), which consists of over 39,000 genes in a single array, was used.  Based on gene expression patterns, the variable genes were grouped into sixteen clusters. Each cluster contained genes associated with cellular immune processes, signaling, cell-cycle, complement coagulation cascade, biosynthesis/metabolism, ubiquitous genes involved in several cascades, and genes of unknown function.  Interestingly, gene expression in clusters 2 and 3 were significantly downregulated by day 1 following EBOV challenge in CD45100% mice.  In contrast, at day 1 following EBOV infection, the CD45 62% mice maintained gene expression patterns similar to day 0.  The differences in gene expression patterns between the CD45 100% and CD45 62% splenocytes were less apparent at day 3 following infection and by days 5 and 7 they became very similar.  At day 9, when wild-type mice had succumbed to the disease, the pattern in CD45 62% mice remained similar to the day 7 patterns of CD45 100% and CD45 62% mice.  The pattern at days 11 and 13 in the CD45 62% mice had returned to that of day 0 CD45 100% or CD45 62% mice.  These results suggested that in CD45 100% mice, subversion of the cell transcriptional machinery during the early stages of EBOV infection (day 1) might represent a major factor leading to death of the mice.  In CD45 62% mice, early control of gene regulation likely provided the appropriate antiviral responses leading to regulated inflammation, immune co-stimulation, and survival.

overall_design: RNA expression in CD45 100% and 62% mice were compared at each time point: days 0, 1, 3, 5, and 7, using an empirical Bayes procedure. From 3 to 8 biological replicates were in each group. Samples for days 9, 11, and 13 (4 biological replicates each) were also available for only CD45 62%. Median expression values for highly variable genes within each group were clustered using agglomerative nesting over all available time points.

first: Rekha

middle: G

last: Panchal

first: Steven

middle: B

last: Bradfute

first: Kelly

middle: L

last: Warfield

first: Gordon

last: Ruthel

first: Douglas

last: Lane

first: Tara

last: Kenny

first: Arthur

middle: O

last: Anderson

last: Raschke

first: Sina

last: Bavari


title: Human liver stem cells-derived microvesicles accelerate hepatic regeneration in partially hepatectomized rats

status: Public on Aug 05 2009

city: Torino

first: Raffaele

last: Calogero

state: To

address: Via Nizza 52

department: Molecular Biotechnology Center

country: Italy

institute: University of Torino

summary: Several studies indicate that adult stem cells may improve the recovery from acute tissue injury. It has been suggested that they may contribute to tissue regeneration by the release of paracrine factors promoting proliferation of tissue resident cells. However, the factors involved remain unknown. In the present study we found that microvesicles (MV) derived from human liver stem cells (HLSC) were able to stimulate in vitro proliferation and apoptosis resistance of human and rat hepatocytes. These effects required internalization of MV in the hepatocytes by an alpha4 integrin-dependent mechanism. However, when treated with RNase, MV despites their internalization were unable to induce hepatocyte proliferation and apoptosis resistance, suggesting an RNA dependent effect. Microarray analysis and quantitative RT-PCR demonstrated that MV were shuttling a specific subset of cellular mRNA, such as mRNA associated in the control of transcription, translation, proliferation and apoptosis. When administered in vivo, MV were found to accelerate the morphological and functional recovery of liver in a model of 70% hepatectomy in rats by inducing an hepatocytes proliferation that was abolished by RNase treatment. Using human AGO2 gene, which is shuttled by MV, as a reporter gene, we found the expression of human AGO2 mRNA and protein in the liver of hepatectomized rats treated with MV. This suggest a translation of the MV shuttled mRNA within hepatocytes of treated rats. Conclusion: these results suggest that MV derived from HLSC may activate a proliferative program in remnant hepatocytes after hepatectomy by a horizontal transfer of specific mRNA subsets.

overall_design: MV contained mRNA was submitted to microarray analysis not to define the amount of mRNA but only to define which transcripts were present. Total RNA was prepared from two independent preparation of vesicles. 250, 500 and 1000 ngs from each preparation were transformed in biotin-labeled cRNA. A simple statistical linear model was used to identify transcript signals linearly correlated to the increment of total RNA concentration used to prepare cRNA.

first: Maria

last: Herrera

first: Valentina

last: Fonsato

first: Stefano

last: Gatti

last: Deregibus

first: Andrea

last: Sordi

first: Daniela

last: Cantarella

first: Benedetta

last: Bussolati

first: Ciro

last: Tetta

first: Giovanni

last: Camussi


title: whole lungs: TAZ-deficient mice and their littermates

status: Public on Aug 01 2009

city: Tokyo

first: Akihisa

last: Mitani

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

department: Graduate School of Medicine

country: Japan

institute: University of Tokyo

summary:
TAZ-deficient mice have the abnormalities in the lung development. We expect the comparison of the gene expression profiles of TAZ-deficient and wild-type lungs would reveal the underlying mechanisms.
Keywords: genetic modification

overall_design: Four total RNA samples from E15.5 lungs were mixed for each group.

first: Ryosuke

last: Makita

first: Hiroyuki

last: Aburatani

first: Takahide

last: Nagase

first: Hiroki

last: Kurihara


title: Integration of ERG gene mapping and gene expression profiling identifies distinct categories of human prostate cancer

city: Sutton

first: Ian

last: Giddings

state: Surrey

address: 15 Cotswold Road

department: Section of Molecular Carcinogenesis

country: United Kingdom

institute: Institute of Cancer Research

summary:
OBJECTIVE: Previous expression microarray analyses have failed to take into consideration the genetic heterogeneity and complex patterns of ERG gene alteration frequently found in cancerous prostates. The objective of this study is for the first time, to integrate the mapping of ERG gene alterations with the collection of expression microarray data.
PATIENTS AND METHODS: We have deterimined genome-wide expression levels with Affymetrix GeneChip Human Exon 1.0 ST arrays using RNA prepared from 35 specimens of prostate cancer from 28 prostates.
RESULTS: The expression profiles exhibit clustering, in unsupervised hierarchical analyses, into two distinct prostate cancer categories, with one group strongly associated with indicators of poor clinical outcome. The two categories are not tightly linked to ERG gene status. Through analysis of the data we identify a subgoup of cancers lacking ERG gene rearrangements that exhibit an outlier pattern of SPINK1 mRNA expression. We also show that a major distinction between ERG gene rearranged and non-rearranged cancers involves the levels of expression of genes linked to exposure to beta-estradiol, and to retinoic acid.
CONCLUSIONS: Our studies show that expression profiling of prostate cancer samples containing single patterns of ERG alterations can provide novel insights into the mechanism of prostate cancer development, and support the view that factors other than ERG gene status are the major determinants of poor clinical outcome.

Keywords: disease state analysis

overall_design: Genome-wide expression levels were obtained from Affymetrix GeneChip Human Exon 1.0 ST arrays using RNA prepared from 35 specimens of prostate cancer from 28 prostates.  Additionally, experiments on three normal specimens were performed. 1 technical replicate was also run.

first: Sameer

last: Jhavar

first: Daniel

last: Brewer

first: Sandra

last: Edwards

first: Zsofia

last: Kote-Jarai

first: Gerhardt

last: Attard

first: Jeremy

last: Clark

first: Penny

last: Flohr

first: Timothy

last: Christmas

first: Alan

last: Thompson

first: Matthew

last: Parker

first: Christopher

last: Shepherd

first: Ulf-Hakan

last: Stenman

first: Tania

last: Marchbank

first: Raymond

last: Playford

last: Woodhouse

last: Ogden

first: Cyril

last: Fisher

first: Gyula

last: Kovacs

first: Cathy

last: Corbishley

first: Charles

last: Jameson

first: Andy

last: Norman

first: Johann

last: De-Bono

first: Anders

last: Bjartell

first: Rosalind

last: Eeles

first: Colin

middle: S

last: Cooper


title: Expression data from Bmi1-overexpressing Ink4a-Arf-null hepatoblasts

city: Chiba

first: Tetsuhiro

last: Chiba

address: 1-8-1 Inohana, Chuo-ku

institute: Graduate School of Medicine, Chiba University

summary: Forced expression of Bmi1 accelerated the self-renewal of hepatic stem/progenitor cells and eventually induced their transformation in an in vivo transplant model. The Ink4a/Arf locus, which encodes a cyclin-dependent kinase inhibitor, p16Ink4a, and a tumor suppressor, p19Arf, is a pivotal target of Bmi1. Therefore, it would be of importance to understand the contribution of the Ink4a/Arf locus to Bmi1 oncogenic functions in cancer and search for as-yet-unknown Bmi1 target genes other than Ink4a/Arf. We used microarrays to explore novel candidate downstream targets for Bmi1 in hepatic stem/progenitor cells

overall_design: Purified Dlk-positive hepatoblasts at day 28 of culture were subjected to RNA extraction and hybridization on Affymetrix microarrays. Data were obtained for quadrant samples from four independent experiments.

first: Atsushi

last: Iwama

first: Hitoshi

last: Ichikawa


title: Gene expression analysis of a podocyte specific PTIP deletion in mouse glomerular preparations at 1 month of age

status: Public on Aug 19 2009

city: Ann Arbor

first: Greg

last: Dressler

state: MI

address: 109 Zina Pitcher Drive

department: Pathology

institute: University of Michigan

summary:
Glomerular RNA comparison between wild-type and podocyte specific deletion of the PTIP gene in 1 month old kidneys. The PTIP gene was deleted using a floxed allele and a Podocin-Cre driver strain.

These mice develop protein urea by 3 months of age. This study was designed to find gene expression differences prior to the onset of the phenotype.

overall_design:
A ptip floxed allele was crossed into the Podocin-Cre expressing strain to generate ptip fl/fl homozygous mice or ptip fl/- transheterozygous mice with or without the Cre expressing transgene. Animals were genotyped and assigend into two groups: Mutants with PTIP deletion (PTIP-) with the genotype ptip fl/fl:Pod-Cre or ptip fl/-:Pod-Cre or control mice (PTIP+) with either ptip fl/fl or ptip fl/-. Note that the floxed allele produced normal levels of wild-type PTIP protein.

Kidneys were excised at 1 month of age. Glomeruli enriched fractions were generated by sieving of the tissue homogenates. RNA was prepared from glomerular enriched fractions.

first: Gregory

middle: R


title: Expression Data from in vivo follicular helper CD4 T cells (TFH) versus non follicular helper CD4 T cells (non-TFH)

status: Public on Jul 28 2009

city: San Diego

first: Shane

last: Crotty

address: 9420 Athena Circle

department: Vaccine Discovery

institute: La Jolla Institute for Allergy and Immunology

summary: Analysis of in vivo antigen-specific (LCMV-specific, SMARTA TCR transgenic) follicular helper CD4 T cells (CXCR5high),versus non-follicular helper CD4 T cells (CXCR5low), eight days after viral infection. A paper including data analysis of these experiments has been accepted for publication (Robert J. Johnston et al. Bcl6 and Blimp-1 are reciprocal and antagonistic regulators of follicular helper CD4 T cell differentiation).

overall_design: Analysis of in vivo antigen-specific (LCMV-specific, SMARTA TCR transgenic) follicular helper CD4 T cells (CXCR5high), versus non-follicular helper CD4 T cells (CXCR5low), eight days after viral infection.


title: The SWI/SNF protein ATRX co-regulates pseudoautosomal genes that have translocated to autosomes in the mouse genome

city: London

first: Nathalie

last: Berube

state: Ontario

address: 800 Commissioners Road East

country: Canada

institute: Children's Health Research Institute

summary: Pseudoautosomal regions (PAR1 and PAR2) in eutherians retain homologous regions between the X and Y chromosomes that play a critical role in the obligatory X-Y crossover during male meiosis.  Genes that reside in the PAR1 are exceptional in that they are rich in repetitive sequences and undergo a very high rate of recombination.  Remarkably, murine PAR1 homologs have translocated to various autosomes, reflecting the complex recombination history during the evolution of the mammalian X chromosome.  We now report that the SNF2-type chromatin remodeling protein ATRX controls the expression of eutherians ancestral PAR1 genes that have translocated to autosomes in the mouse.  In addition, we have identified two potentially novel mouse PAR1 orthologs.  We propose that the ancestral PAR1 genes share a common epigenetic environment that allows ATRX to control their expression.

overall_design:
At E13.5, n = 3 biological replicates of littermate-matched wt/ko pairs.  RNA from 2 forebrains were pooled to generate enough RNA for each sample.
At P0.5, n = 4 biological replicates of littermate-matched wt/ko pairs (for pair #2 there is one wt and 2 Atrx-null samples (2A & 2B) and we count this as 2 pairs).

first: Michael

last: Levy

first: Deanna

last: Tremblay


title: Fat Treg cells

status: Public on Jul 27 2009

city: Boston

first: CBDM

last: Lab

state: MA

address: 77 Avenue Louis Pasteur

department: Microbiology and Immunobiology

institute: Harvard Medical School

summary:
Comparisons of global gene-expression profiles revealed a greater distinction between CD4+ Treg cells and CD4+ conventional (Tconv) T cells  residing in abdominal (epidydimal) fat versus in more standard locations such as the spleen, thymus and LN.
Keywords: Cell population comparison

overall_design: All gene expression profiles were obtained from highly purified T cell populations sorted by flow cytometry. To reduce variability, cells from multiple mice were pooled for sorting, and three replicates were generated for all groups. RNA from 3-5 x 104 cells was amplified, labeled, and hybridized to Affymetrix M430v2 microarrays. Raw data were preprocessed with the RMA algorithm in GenePattern, and averaged expression values were used for analysis.

first: Markus

last: Feuerer

first: Christophe

last: Benoist

first: Diane

last: Mathis


title: RNA profiling of skeletal muscle in RP58 knockout mice

status: Public on Jul 31 2009

city: Setagaya

first: Hiroshi

last: Asahara

state: Tokyo

department: Department of Systems BioMedicine

institute: National Research Institute for Child Health and Development

summary:
To predict RP58-regulated genes involved in skeletal myogenesis, RNA profiling experiments were performed, comparing RNA derived from skeletal muscle tissue of a RP58+/+ mouse to that from a RP58 knockout (KO) mouse at E18.5. Importantly, well-known dominant-negative inhibitors of muscle differentiation, the Id family of genes (Id1/Id2/Id3), were upregulated in the RP58 KO muscle. On the contrary, a number of muscle differentiation-related genes, such as Ckm, troponin and Myosin, were downregulated in the same sample. These results indicate that the repressor protein RP58 is important for muscle terminal differentiation, possibly suppressing the gene expression of muscle differentiation genes such as the Ids.

Keywords: Knockout tissue

overall_design: Skeletal muscles of WT and RP58 KO mice (E18.5 diaphragm) were dissected under a microscope and RNA was extracted using ISOGEN (Nippongene). After EtOH precipitation, RNA was dissolved into DEPC-DW and its concentration was determined.   Microarray analysis - cRNA was synthesized. 10 ug of cRNA were hybridized to Affymetrix mouse 430A 2.0 arrays. Signal intensities were calculated using the RMA method. MAPPFinder (www.genmapp.org) was used to integrate expression data with known pathways.

first: Shigetoshi

last: Yokoyama

first: Satoshi

last: Yamashita

first: Yoshiaki

last: Ito


title: Expression data from mouse liver after Acetaminophen intoxication

status: Public on Aug 14 2009

city: Palo Alto

first: Hong-Hsing

last: Liu

address: S3-115E, 3431 Hillview Ave.

institute: Roche Palo Alto

summary:
Acetaminophen (APAP) is the most widely used analgesic in the United States. Its acute overdose causes liver damage by inducing localized centrilobular cell death. Because of widespread use, APAP toxicity has become the most frequent cause of acute liver failure. Many factors have been associated with the susceptibility of APAP-induced liver injuries, however, few of them have been confirmed and used in the clinical setting.
We tried to identify the subset of factors that could affect susceptibility to APAP-induced liver injury by an integrative genetic, transcriptional and 2-D-NMR-based metabolomic analysis across a panel of inbred mouse strains.

overall_design: After a single administration of high dose (300 mg/kg i.p.) APAP, liver and blood samples were extracted from 3 sensitive (C57B6, DBA/2, and SmJ) and 1 resistant (SJL) mice strains at 0, 3 and 6 hour after APAP exposure. Endogenous metabolites from liver samples were analyzed by 1H-13C 2-dimensional-NMR and gene expression changes occurring within these liver samples were simultaneously analyzed using Affymetrix microarrays. The transcriptional and metabolomic data was jointly analyzed, and functional information within the Gene Ontology database was used to identify the subset of genes that could affect susceptibility to APAP-induced liver injury in the early phase response.

first: Peng

last: Lu

first: Erin

last: Farrell

first: Kimberly

last: Dolim

first: Jimmy

last: Jung

first: Zhaomei

last: Zhang


title: Expression data from MCF-7 cells transfected with miR-26a and treated or not with estradiol

status: Public on Aug 04 2009

city: Toulouse

first: Stephan

last: Vagner

address: 20-24 Rue du Pont St Pierre

department: Institut Claudius Regaud

country: France

institute: INSERM U563

summary: Altered expression of microRNAs (miRNAs), an abundant class of small non-protein-coding RNAs that mostly function as negative regulators of protein-coding gene expression, is common in cancer. Here we analyze the regulation of miRNA expression in response to estrogen, a steroid hormone that is involved in the development and progression of breast carcinomas and that is acting via the estrogen receptors (ER) transcription factors. We set out to thoroughly describe miRNA expression, by using miRNA microarrays and real time RTPCR experiments, in various breast tumor cell lines in which estrogen signaling has been induced by 17β-estradiol (E2). We show that the expression of a broad set of miRNAs decreases following E2 treatment in an ER-dependent manner. We further show that enforced expression of several of the repressed miRNAs reduces E2-dependent cell growth, thus linking expression of specific miRNAs with estrogen-dependent cellular response. In addition, a transcriptome analysis revealed that the E2-repressed miR-26a and miR-181a regulate many genes associated with cell growth and proliferation, including the progesterone receptor gene, a key actor in estrogen signaling. Strikingly, miRNA expression is also regulated in breast cancers of women who had received antiestrogen neoadjuvant therapy thereby showing an estrogen-dependent in vivo regulation of miRNA expression. Overall, our data indicates that the extensive alterations in miRNA regulation upon estrogen signalling pathway plays a key role in estrogen-dependent functions and highlights the utility of considering miRNA expression in the understanding of antiestrogen resistance of breast cancer.

overall_design: 9 samples analyzed. Triplicates were done. MCF-7+miR26a+E2 (n=1 to 3) ; MCF7+miRctrl+E2 (n=1 to 3) ; MCF7+miRctrl+vehicle (n=1 to 3). We generated pairwise comparisons using EASANA from GenoSplice technology: MCF-7+miR26a+E2 versus MCF7+miRctrl+E2 and MCF-7+miRctrl+E2 versus MCF7+miRctrl+vehicle. Fold change ≥1.5 were selected.

first: G

last: Maillot

first: L

last: Gratadou

first: D

last: Auboeuf

first: S


title: Toward a Better Understanding of Potential Roles of Astrocytes in HIV-1-associated Neurocognitive Disorders

city: New York

first: Alejandra

last: Borjabad

address: 432 West 58th, Street

department: Medicine

institute: St.Luke's-Roosevelt Hospital Center

summary: We present a microarray analysis of primary mouse astrocytes exposed to HIV-1 in culture. Results are compared with previous genomic studies of HIV-1 effect in human astrocytes and human and macaque brains.

overall_design: Day-two neonatal mouse astrocytes were exposed in triplicates to cell-free HIV-1/NL4-3 at m.o.i. of 1 and cultured for 24 hours.  As control, cells were treated in triplicates with a mock virus concentrate at the same dilution (vol/vol) as the wild-type virus.  As an example of large scale gene expression analysis we shall provide some details of the method; full description is in Kim et al. (S.-Y. Kim et al., 2004).  Total RNA was prepared from cell cultures using the RNAeasy total RNA extraction kit (Qiagen, CA).  RNA quality was assessed by electrophoresis and spectrophotometric analysis; between 1 and 10 µg of total RNA was used to generate a cDNA, and then 1 µg of cDNA product was used in an in vitro transcription reaction that contained biotinylated UTP and CTP.  Twenty µg of full-length cRNA was fragmented and was subjected to gene expression analysis on the Affymetrix Mouse Genome 430 2.0 Array chip.  Affymetrix software was used to generate CHP files. Significance analysis was performed using ArrayAssist software (Stratagene).

first: Andrew

middle: I

last: Brooks

first: David

last: Volsky


title: Common regulatory variation impacts gene expression in a cell type dependent manner

city: Geneva

first: Emmanouil

middle: T

last: Dermitzakis

address: 1 Rue Michel-Servet

department: Department of Genetic Medicine and Development

country: Switzerland

institute: University of Geneva Medical School

summary: Studies correlating genetic variation to gene expression facilitate the interpretation of common human phenotypes and disease. As functional variants may be operating in a tissue-dependent manner, we performed gene expression profiling and association with genetic variants (SNPs) on three cell types of 85 individuals. After excluding 10 outlier individuals detected through principal component analysis, we detected cell type-specific genetic effects, with 69 - 80% of regulatory variants operating in a cell type-specific manner and identified multiple eQTLs per gene, unique or shared among cell types and positively correlated with the number of transcripts per gene. Cell type specific eQTLs were found at larger distances from genes and lower effect size similar to known enhancers. These data suggest that the complete regulatory variant repertoire can only be uncovered in the context of cell type specificity.

overall_design: Total RNA was extracted from fibroblasts, LCLs, and T-cells of the 85 unrelated individuals of the GenCord. Two one-quarter scale Message Amp II reactions (Ambion, Foster City, CA) were performed for each RNA extraction with 200 ng of total RNA. 1.5 μg of cRNA was hybridized to a commercial whole-genome expression array (WG-6 v3 Expression BeadChip, Illumina) to quantify transcript abundance. In total there were two technical replicates (labeling and hybridization) for each RNA sample.

first: Antigone

last: Dimas

first: Samuel

last: Deutsch

first: Barbara

last: Stranger

first: Stephen

last: Montgomery

first: Christelle

last: Borel

first: Homa

last: Attar-Cohen

first: Catherine

last: Ingle

first: Claude

last: Beazley

first: Maria

last: Arcelus

first: Magdalena

last: Sekowska

first: Marilyne

last: Gagnebin

first: James

last: Nisbett

first: Panos

last: Deloukas

first: Stylianos

last: Antonarakis


title: Gene expression regulated by G-actin switch

status: Public on Jul 15 2009

city: Martinsried

first: Guido

last: Posern

address: Am Klopferspitz 18

department: Dept. Molecular Biology

country: Germany

institute: Max-Planck-Institute of Biochemistry

summary: We analysed the G-actin regulated transcriptome by gene expression analysis using previously characterised actin binding drugs. We found many known MAL/MRTF-dependent target genes of serum response factor (SRF) as well as unknown directly regulated genes.

overall_design: Three conditions were used for the microarray analysis: control NIH 3T3 cells, cells treated with cytochalasin D, and cells treated with latrunculin B and cytochalasin D. For each condition, three independent biological replicates were analysed.

first: Arnaud

last: Descot

first: Reinhard

last: Hoffmann


title: Expression profiling of mouse dorsal skin during hair follicle cycling

status: Public on Jul 17 2009

city: Irvine

first: Kevin

last: Lin

address: 250 Sprague Hall

department: Biological Chemistry & Medicine

institute: University of California, Irvine

summary: Hair follicles undergo recurrent cycling of controlled growth (anagen), regression (catagen), and relative quiescence (telogen) with a defined periodicity. Taking a genomics approach to study gene expression during synchronized mouse hair follicle cycling, we discovered that, in addition to circadian fluctuation, CLOCK-regulated genes are also modulated in phase with the hair growth cycle. During telogen and early anagen, circadian clock genes are prominently expressed in the secondary hair germ, which contains precursor cells for the growing follicle. Analysis of Clock and Bmal1 mutant mice reveals a delay in anagen progression, and the secondary hair germ cells show decreased levels of phosphorylated Rb and lack mitotic cells, suggesting that circadian clock genes regulate anagen progression via their effect on the cell cycle. Consistent with a block at the G1 phase of the cell cycle, we show a significant upregulation of p21 in Bmal1 mutant skin. While circadian clock mechanisms have been implicated in a variety of diurnal biological processes, our findings indicate that circadian clock genes may be utilized to modulate the progression of non-diurnal cyclic processes.

overall_design: To investigate the molecular control of hair follicle cycling, we profiled mRNA expression in mouse dorsal skin at multiple representative time points in the synchronized second postnatal hair growth cycle and in a depilation-induced hair growth cycle. For profiling of second synchronized and depilation-induced hair growth cycle, the same upper-mid region of dorsal skin was excised from C57BL/6 mice at representative postnatal days (P). The time points for second hair growth cycle are classified into different phases of the hair growth cycle based on established morphological guidelines as follow: early anagen (P23, P25), mid anagen (P27), late anagen (P29, P34), early catagen (P37, P39), mid catagen (P41), and telogen (P44). Depilation-induced hair growth cycle by applying wax/rosin mixture on the dorsal skin of seven-week old mice (all follicles in telogen) was performed on mice. The corresponding phases of the hair growth cycle at number of days following depilation (D) is as follow: early anagen (D3), mid anagen (D5), late anagen (D8, D12), and early catagen (D17). For each time point, multiple biological replicates were profiled, with each mouse dorsal skin separately hybridized to an Affymetrix array.

first: Vivek

last: Kumar

first: Mikhail

last: Geyfman

first: Darya

last: Chudova

first: Alexander

last: Ihler

first: Padhraic

last: Smyth

first: Ralf

last: Paus

first: Joseph

last: Takahashi

first: Bogi

last: Andersen


title: Expression profiling of ntTS and TS cells

first: Mayumi

last: Oda

address: 35 Shinanomachi, Shinjuku

institute: Keio University

summary: Placental abnormalities occur frequently in cloned animals.  To investigate gene expression profile of trophoblast cell lineage of somatic cell nuclear transferred (NT) embryos, we established TS cells from blastocysts produced by NT at the blastocyst stage.

overall_design: Five independent TS cell lines derived from NT embryos were used in this study.  Those included three lines from BDF1 background NT embryos and two lines from EGFP-Tg CD-1 background NT embryos.  Two each of control lines, derived from native blastocysts of each background, were also used.

first: Satoshi

last: Tanaka

first: Misa

last: Iwatani

first: Kunio

last: Shiota


title: Gene expression profiles of normal human mammary cell subpopulations FACS sorted based on expression of CD49f and EpCAM

status: Public on Jul 09 2009

city: Parkville

first: Jane

last: Visvader

state: Victoria

address: 1G Royal Parade

department: Victorian Breast Cancer Research Consortium

country: Australia

institute: The Walter and Eliza Hall Institute of Medical Research

summary: To delineate epithelial subpopulations in human mammary tissue, hematopoietic and endothelial cells were depleted from freshly isolated cell suspensions derived from reduction mammoplasties by fluorescence-activated cell sorting. The resultant Lin- population was fractionated into four distinct subpopulations using CD49f (α6-integrin) and epithelial cell adhesion molecule (EpCAM; also referred to as CD326 and ESA). Based on the immunohistochemical phenotype, and in vivo and in vitro functional assays, these subpopulations were identified as fibroblast-enriched stromal (CD49f -EpCAM-), mammary stem cell (MaSC)-enriched (CD49f hiEpCAM-), luminal progenitor (CD49f +EpCAM+), and mature luminal (CD49f –EpCAM+) cell subpopulations. Microarray profiling was used to derive gene expression signatures representative of these subpopulations using freshly sorted cells (>90% purity) from normal breast tissue. The four mammary cell subpopulations were found to have distinct gene expression profiles.

overall_design: Four mammary cell subpopulations from three individual patient samples were analyzed.

first: Gordon

first: Geoffrey

last: Lindeman


title: ConA-induced fulminant hepatitis in a mouse model

status: Public on Jul 21 2009

city: HangZhou

first: Dong Cheng

last: Liu

address: Qingchun Road 79, Hangzhou, Zhejiang 310003, China

institute: State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, affiliated to Zhejiang University

summary:
The goal of  this experiment was to investigate the early mechanisms of human fulminant hepatitis through ConA-induced hepatitis model.Early diagnosis and interventions are important for patients with fulminant hepatitis and gene expression may be pivotal in the early diagnosis.

Keyword :ConA-induced hepatitis model

overall_design: ConA was injected through the mouse caudal vein at one of 4 time points (0 hr, 1 hr, 3 hr, 6 hr). The effects of ConA treatment on hepatic gene expression at these time points were analyzed .There are 3 replicates  at each timepoint  then  4*3=12 samples in all.

first: Feng

last: Chen

first: Hai-Hong

last: Zhu

first: Lin-Fu

last: Zhou

first: Li-Ying

last: Zhao

first: Shan-Shan

last: Wu

first: Jing

last: Wang

first: Zhi


title: Retinoic Acid Delivery within Embryoid Bodies Induces an Early Streak Phenotype in vitro

city: Atlanta

first: Nathan

middle: J.

last: Bowen

address: 223 James P Brawley Drive

department: Center for Cancer Research and Therapeutic Development

institute: Clark Atlanta University

summary: During embryogenesis, cell specification and tissue formation is directed by the concentration and temporal presentation of morphogens, and similarly, pluripotent embryonic stem cells differentiate in vitro into various phenotypes in response to morphogen treatment.  Embryonic stem cells are commonly differentiated as three dimensional spheroids called embryoid bodies (EBs); however, differentiation within EBs is typically heterogeneous and disordered.  Here we show that spatiotemporal control of microenvironmental cues embedded directly within EBs enhances the homogeneity, synchrony and organization of differentiation.  Degradable polymer microspheres releasing retinoic acid within EBs induce the formation of cystic spheroids closely resembling the early streak mouse embryo, with an exterior of visceral endoderm enveloping an epiblast layer.  These results demonstrate that controlled morphogen presentation to stem cells more efficiently directs cell differentiation and tissue formation, thereby improving developmental biology models and enabling the development of regenerative medicine therapies and cell diagnostics.

overall_design:
ESCs (D3 line) were maintained in an undifferentiated state on gelatin-coated tissue culture plates in LIF-containing media as described previously.  EBs were formed from 2x106 ESCs inoculated in LIF-free media and cultured under rotary conditions.  EBs containing microspheres were produced by coating CellTracker Red (CONTROLs) or RA-loaded (EXPERIMENTs) microspheres in 0.1% gelatin solution for 3 hours prior to mixing with ESCs in various microsphere to cell ratios and a range of rotary speeds.  EB media was exchanged every 1-2 days as needed.
Triplicate control and experiment EBs were processed for microarray.

first: Richard

last: Carpenedo

first: Adres

last: Bratt-Leal

first: Ross

last: Marklein

first: Scott

last: Seaman

first: John

middle: F

last: McDonald

first: Todd

last: McDevitt


title: Influence of type I Interferons on function of splenic conventional dendritic cells.

status: Public on Jul 07 2009

city: Munich

first: Jacek

last: Puchałka

address: Lindwurmstrasse 2a

department: University Children's Hospital

institute: Munich University Hospital

summary: Type I Interferons encompasses a large family of closely related cytokines comprising of at least 13 IFN-α isotypes and single IFN-β. Both IFN-α and IFN-β exert their activity through a common receptor IFNAR. Type I Interferons have broad regulatory effects and various subtypes of dendritic cells are influenced by this cytokines. In our study we asked question whether the low, constitutive levels of type I Interferons produced under steady state conditions are important for proper function of splenic conventional dendritic cells.

overall_design: In this approach we sorted out two populations (CD8α+ and CD8α-) of splenic dendritic cells (DCs) from untreated WT, IFN-β-/- and IFNAR-/- C57Bl/6 mice. All mice were between 8-10 weeks old. Further we isolated RNA and performed microarray analysis. Each DCs population was repeated twice.

first: Natalia

last: Zietara

first: Marcin

last: Lyszkiewicz

last: Puchalka


title: Normal human mammary tissues from BRCA1 mutation carriers, non-BRCA1/2 mutation carriers and normal women

first: Jane

last: Visvader

summary:
Gene expression profiles of normal human mammary tissue from BRCA1 mutation carriers, non-BRCA1/2 mutation carriers and normal women.

RNA was prepared from normal breast tissue (confirmed by pathology) from reduction mammoplasties (n=5) and prophylactic mastectomies of known BRCA1 (n=7) and non-BRCA1/2 mutation carriers (n=8). non-BRCA1/2 carriers were individuals with a strong family history of breast cancer (kConFab Category 1 (http://www.kconfab.org/Collection/Eligibility.shtml) where no mutation in BRCA1 or BRCA2 has been identified in the family by high sensitivity testing (http://www.kconfab.org/Progress/Sensitivity.shtml) of any individual affected by breast or ovarian cancer. Normal breast samples refer to reduction mammoplasty specimens, where family history is generally not known.

overall_design: Microarray expression profiles were obtained from whole pre-neoplastic breast tissue from three categories of patient, as described in the summary. Samples were checked for keratin gene expression levels as a marker for epithelium tissue, and a number of samples were removed because two or more keratin genes lacked detectable expression (BeadStudio detection p-value 0.01). One other sample was removed because its expression profile appeared abnormal. Out of 36 samples initially profiled, 20 passed the quality filters and were used for the final analysis.

first: Gordon

last: Smyth

first: Geoffrey

middle: J

last: Lindeman


title: Expression profiling of Clock mutant dorsal skin at telogen

first: Kevin

last: Lin

overall_design: To gain molecular understanding of the the hair cycle delay in Clock mutant mice, we profiled the dorsal skin of Clock mutant and their wild-type littermates at P23. At P23, the skin samples are comparable because all the samples are in telogen just prior to the hair cycle delay was observed. Histological sections were used to classify each sample into specific stage of the hair growth cycle based on established morphological guidelines. RNA from each mouse dorsal skin were separately hybridized to an Affymetrix array.

first: Vivek

last: Kumar

first: Mikhail

last: Geyfman

first: Darya

last: Chudova

first: Alexander

last: Ihler

first: Padhraic

first: Ralf

last: Paus

first: Joseph

last: Takahashi

first: Bogi

last: Andersen


title: C2C12 culture: myogenesis timecourse and shRp58 treatment

first: Hiroshi

last: Asahara

summary:
To predict Rp58-regulated gene involved in myogenesis, RNA profiling experiments were performed, comparing RNA derived from C2C12 with or without expressing shRNA for Rp58. As a result, 271 genes were upregulated in C2C12 stably expressing shRNA-Rp58 cells compared with control C2C12 cells. As Rp58 is repressor in C2C12, we hypothesized that Rp58 regulates gene cluster which expression is downregulated in accordance with Rp58 expression and myogenesis progression. In this regard, we also characterized dynamic gene expression patterns during myogenesis by microarray at 4 different stage (GM, day 0, 2, 4) of C2C12 myogenesis assays and found that 399 genes expression is characterized as downregulation pattern during myogenesis. Importantly, this down regulation gene set and upregulated genes by shRNA for Rp58 were highly overlapped.

Keywords: time course, shRNA

overall_design:
C2C12 murine skeletal muscle cells were purchased from American Type
culture Collection (ATCC). These cells were mainteined in GM (DMEM
supplemented with 10% FBS). Cells were grown in GM and after reaching
counfluence, the medium was switched to DM (DMEM supplemented with 2% hourse serum) and further incubated. The medium was changed every 2 days. Culture was performed by using within five passages cells. For the experiment of shRNA for Rp58, transfection was performed by using Lipofectamin 2000 (Invitrogen). Stable transfectants were obtained by selection of the transfected C2C12 cells for two weeks.
Microarray analysis - RNA was isolated as described from C2C12, and cRNA was synthesized. 10 ug of cRNA were hybridized to Affymetrix mouse 430 2.0 arrays. Intensity values were quantified using RMA algorithm.
MAPPFinder (www.genmapp.org) was used to integrate expression data with known pathways.

first: Shigetoshi

last: Yokoyama

first: Yoshiaki

last: Ito


title: Gene expression in mouse tracheal basal cells

status: Public on Jul 02 2009

city: Durham

first: Jason

last: Rock

address: 353 Nanaline Duke, Research Drive

department: Cell Biology

institute: Duke University Medical Center

summary: Epithelial basal cells (BCs) are an important stem cell population of the airways.  We purified BCs from a KRT5-GFP transgenic mouse line and used Affymetrix arrays to compare there gene expression to that of non-BC epithelium.

overall_design: Three populations of cells were obtained by FACS: 1. KRT5-GFP-, lectin- columnar epithelium. 2. KRT5-GFP-, lectin+ BCs. 3. KRT5-GFP+, lectin+ BCs.  Three biological replicates of each sample were hybridized to Affymetrix microarray.

first: Brigid

last: Hogan


title: Expression and DNA methylation profiling of ntTS and TS cells

city: Tokyo

first: Mayumi

last: Oda

type: Methylation profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Gene expression analysis in control and HIF-2 alpha deficient murine lung endothelial cells under hypoxia

city: Philadelphia

first: Nicolas

last: Skuli

state: PA

address: 421 Curie Boulevard

institute: HHMI

summary:
Journal : Blood. 2009 Jul 9;114(2):469-77. Epub 2009 May 13.

Title : Endothelial deletion of hypoxia-inducible factor-2alpha (HIF-2alpha) alters vascular function and tumor angiogenesis.

Authors : Skuli N, Liu L, Runge A, Wang T, Yuan L, Patel S, Iruela-Arispe L, Simon MC, Keith B.

Abstract : Hypoxia-inducible factor-2alpha (HIF-2alpha) is highly expressed in embryonic vascular endothelial cells (ECs) and activates the expression of target genes whose products modulate vascular function and angiogenesis. In this report, we describe a genetic model designed to test the physiologic consequences of deleting HIF-2alpha in murine endothelial cells. Surprisingly, mice with HIF-2alpha-deficient ECs developed normally but displayed a variety of phenotypes, including increased vessel permeability, aberrant endothelial cell ultrastructure, and pulmonary hypertension. Moreover, these animals exhibited defective tumor angiogenesis associated with increased hypoxic stress and tumor cell apoptosis. Immortalized HIF-2alpha-deficient ECs displayed decreased adhesion to extracellular matrix proteins and expressed reduced levels of transcripts encoding fibronectin, integrins, endothelin B receptor, angiopoietin 2, and delta-like ligand 4 (Dll4). Together, these data identify unique cell-autonomous functions for HIF-2alpha in vascular endothelial cells.

Keywords: Murine lung endothelial cell study

overall_design: Cnt1,2 and 3 (Hif-2a floxed/floxed) cells were subjected to 0.5% oxygen treatment for 16hrs and KO1,2 and 3 (Hif-2a knockout) cells were subjected to 0.5% oxygen treatment for 16hrs.

first: Liping

last: Liu

first: Anja

last: Runge

first: Tao

last: Wang

first: Lijun

last: Yuan

first: Sunny

last: Patel

first: Luisa

last: Iruela-Arispe

first: M

last: Simon

first: Brian

last: Keith


title: Induction of Pluripotency in Adult Unipotent Germline Stem Cells

city: Aachen

first: Martin

last: Zenke

state: NRW

address: Universitatsklinikum Aachen, RWTH

institute: Institute for Biomedical Engineering

summary: Mouse and human stem cells with features similar to those of embryonic stem cells have been derived from testicular cells. Although pluripotent stem cells have been obtained from defined germline stem cells (GSCs) of mouse neonatal testis, only multipotent stem cells have been obtained so far from defined cells of mouse adult testis. In this study we describe a robust and reproducible protocol for obtaining germline-derived pluripotent stem (gPS) cells from adult unipotent GSCs. Pluripotency of gPS cells was confirmed by in vitro and in vivo differentiation, including germ cell contribution and transmission. As determined by clonal analyses, gPS cells indeed originate from unipotent GSCs. We propose that the conversion process requires a GSC culture microenvironment that depends on the initial number of plated GSCs and the length of culture time.

overall_design:
20 samples in total.
Germline stem cells (GSC):
- GSC_1
- GSC_2
- GSC_3
Germline pluripotent stem cells (gPS):
- gPSC_1
- gPSC_2
- gPSC_3
- gPSC_4
- gPSC_5
Neural stem cells (NSC):
- NSC_1
- NSC_2
- NSC_3
Mouse embryonic fibroblast (MEF):
- MEF_1
- MEF_2
- MEF_3
Embryonic stem cells (ESC):
- ESC_1
- ESC_2
- ESC_3
ESC-like cells (ESC_like)
- ESC_like_1
- ESC_like_2
- ESC_like_3

first: Kinarm

last: Ko

first: Natalia

last: Tapia

first: Guangming

last: Wu

first: Jeong

last: Kim

first: Marcos

last: Araúzo-Bravo

first: Philipp

last: Sasse

first: Tamara

last: Glaser

first: David

last: Ruau

first: Dong

middle: W

last: Han

first: Boris

last: Greber

first: Kirsten

last: Hausedörfer

first: Vittorio

last: Sebastiano

last: Stehling

first: Bernd

last: Fleischmann

first: Oliver

last: Brüstle

first: Hans

last: Schöler


title: IL-1b responses in receptor-reconstituted AcP-deficient neurons

status: Public on Jul 01 2009

city: Seattle

first: Chris

last: Russell

state: WA

address: 1201 Amgen Ct W

department: Molecular Sciences

institute: Amgen

summary: The purpose was to determine AcP- and AcPb-dependent gene responses to IL-1 by virally-reconstituting AcP-deficient mouse embryonic cortical neurons with CD25 (control), full length AcP, AcPb or the combination of both. A control population was transduced with a CD25-expressing virus. Half the samples were stimulated with IL-1-beta for four hours, RNA was analyzed by microarray.

overall_design: Each in triplicate: CD25 cells, AcP cells, AcPb cells, AcP+AcPb cells. Cultures stimulated with IL-1b (10 ng/ml) for 4 hours then RNA was collected for processing and microarray analysis.

first: Dirk

middle: E

last: Smith

first: Chen

last: Hongbo


title: Altered mouse mammary gland gene expression and tumor growth following chronic social isolation

city: Chicago

first: Suzanne

middle: Daniela

last: Conzen

state: IL

address: 5841 S. Maryland Ave, SBRI J301, MC 2115

institute: The University of Chicago

summary: Clinical studies have revealed that social support improves the outcome of cancer patients while epidemiological studies suggest that social isolation increases the risk of death associated with several chronic diseases. However, the precise biological consequences of an unfavorable social environment have not been defined. To do so, robust, reproducible pre-clinical models are needed to study the mechanisms whereby an adverse environment impacts on gene expression and cancer biology.  Because random assignment of inbred laboratory mice to well-defined social environments allows accurate and repeated measurements of behavioral and endocrine parameters, transgenic mice provide a pre-clinical framework with which to begin to determine gene-environment mechanisms.  In this study, we found that female C3(1)/SV40 T-antigen mice deprived of social interaction from weaning exhibited increased expression of genes encoding key metabolic pathway enzymes in the pre-malignant mammary gland. Chronic social isolation was associated with upregulated fatty acid synthesis and glycolytic pathway gene expression - both pathways known to contribute to increased breast cancer growth. Consistent with the expression of metabolic genes, isolated mice subsequently developed significantly larger mammary gland tumors compared to group-housed mice. Endocrine evaluation confirmed that isolated mice developed a heightened corticosterone stress response compared to group-housed mice. Together, these transdisciplinary studies show for the first time that an adverse social environment is associated with altered mammary gland gene expression and tumor growth.  Moreover, the identification of specific alterations in metabolic pathways favoring tumor growth suggests potential molecular biomarkers and/or targets (e.g. fatty acid synthesis) for preventive intervention in breast cancer.

overall_design: SV40 Tag mice we isolated or grouped at weaning. Mouse mammary glands were rapidly excised at necropsy and immediatley flash frozen to detect difference in gene expression between thoraci MG from isolated versus group-housed female mice.

middle: D

first: Masha

last: Kocherginsky


title: Molecular characterization of CD8+ T cell with suppressive capacities

city: Braunschweig

first: Robert

last: Geffers

address: Inhoffenstr. 7

department: Dep. Molecular Bacteriology

institute: HCI - Helmholtz Centre for Infection Research

summary: In the VILLIN-HA/CL4-TCR transgenic mouse model a population of CD8 T cells with suppressive capacities was indentified. For the molecular characterisation of CD8 regulatory T cells gene comparative expression profiles of naive, activated and regulatory CD8 T cells were performed.

overall_design: Three population of CD8 T cells are included in the analysis. 1. CD8 T cells 2. in vivo activated CD8 T cells 3. in vivo generated CD8 regulatory T cells

first: R

first: A

last: Westendorf


title: Comparative gene expression analysis of 2 subpopulations of dermal papilla cells.

status: Public on Jun 26 2009

city: Cambridge

first: Ryan

last: Driskell

state: Cambridgeshire

address: Tennis Court Rd.

department: Department of Genetics

institute: University of Cambridge

summary:
Different types of hair follicles can be found in the skin of mice. It is believed that the signals that control hair follicle differentiation arise from cells in a structure called the dermal papilla. Understanding the nature of those signals is of interest for the biology of the normal tissue.

We have developed a technique for isolation of dermal cells by enzymatic digestion of intact skin. We have identified two subpopulations of cells that can be separated by FACS. The Sox2-positive CD133-positive cells are found exclusively in the dermal papillae of guard/awl/auchene hairs, while Sox2-negative, CD133-positive cells are found in the other hair follicle types. We compared these populations with unfractionated dermal cells.

overall_design:
We isolated the following 3 populations of cells from the back skin of neonatal mice (P2) by Flow Cytometry:
1)	GFP-CD133- Total dermal cells
2)	GFP-CD133+ Dermal Papilla cells
3)	GFP+CD133+ Dermal Papilla cells
The yield is approximately 50,000 cells of each population.


title: Mndal, a new interferon-inducible family member, suppresses cell growth and may modify plasmacytoma susceptibility

status: Public on Jul 24 2009

city: Bethesda

first: Ke

last: Zhang

address: 37 Convent Dr.

institute: NCI

summary:
Mndal, a new interferon-inducible family member, is highly polymorphic, suppresses cell growth and may modify plasmacytoma susceptibility.
The human HIN-200 gene cluster and its mouse counterpart, the Interferon inducible-200 (Ifi200) family, both on Chr 1, are associated with several diseases, including solid tumors and lupus.  Our study was initiated to identify the modifier gene(s) encoded by the Pctm locus, in which mouse B cell plasmacytomas induced by pristane are associated with heterozygosity of Chr 1 genes near the Ifi200 cluster.
A screen for differentially expressed genes in granulomatous tissues induced by pristane in resistant and susceptible strains identified a new Ifi200 member whose expression was 1000-fold higher in the strain carrying the resistant allele of Pctm, and was the most highly expressed Ifi200 gene.  The gene, designated Mndal (for MNDA-like, myeloid nuclear differentiation antigen like), was absent in the susceptible genome, as were genomic sequences upstream of Ifi203, the gene adjacent to Mndal.  Ectopic expression of MNDAL suppressed cell growth, which, together with the disease susceptibility of heterozygotes at the Pctm locus, suggests that Mndal, perhaps with Ifi203, acts as a tumor suppressor and display(s) haplo-insufficiency.  Mndal is highly polymorphic among inbred mouse strains, as it is absent in 10/24 strains.  This polymorphism may have implications for other disease modifiers mapping to the same region.

overall_design: Using institutionally approved animal protocols, mice were given 0.5 ml TMPD intraperitoneally.  Three and 18 days later, total RNA was extracted from mesentery of experimental/control animals, using TRIzol (Invitrogen).  Labeled aRNA prepared from 1 µg of RNA (MessageAmpII aRNA Amplification kit, Ambion protocols) was hybridized to mouse genome 430 2.0 array chips, processed on Workstation 450, and analyzed with Gene Chip Operating Software (GCOS; Affymetrix).  Differential expression was assessed by ANOVA (Partek Genomics Suite).

first: Daniel

last: Kagan

first: Wendy

last: DuBois

first: Richard

last: Robinson

first: Valery

last: Bliskovsky

first: William

middle: C

last: Vass

first: Shuling

first: Beverly

middle: A

last: Mock


title: Expression data from activated bone marrow-derived dendritic cells (BMDCs)

status: Public on Jun 25 2009

first: Akihiko

last: Yoshimura

address: 35 Shinanomachi Shinjyuku-ku

department: Microbiology and Immunology

institute: Keio Medical School

summary:
genes regualted by LPS or LPS+cAMP stimulation in BMDCs
We used microarrays to identify genes that up-regulated by LPS+cAMP compared with just LPS.

overall_design: BMDCs were stimilated with LPS (10 ng/ml) in the presence or absence of cAMP (100 microM) for 3h. Specifically up-regualted gene by cMP was identified.

first: Keiko

last: Koga


title: Activation-induced cytidine deaminase accelerates clonal evolution in BCR-ABL1-driven acute lymphoblastic leukemia

status: Public on Jun 24 2009

city: Los Angeles

first: Markus

last: Muschen

state: CA

address: 4650 Sunset Blvd MS 136

department: Hematology Oncology

institute: Childrens Hospital Los Angeles

summary: Activation-Induced Cytidine Deaminase (AID) is required for somatic hypermutation and immunoglobulin (Ig) class switch recombination in germinal center B lymphocytes.  Occasionally, AID targets non-Ig genes, thereby contributing to B cell lymphomagenesis.  We recently reported aberrant expression of AID in BCR-ABL1-driven acute lymphoblastic leukemia (ALL).  To elucidate the biological significance of aberrant AID expression, we studied loss of AID function in a murine model of BCR-ABL1 ALL.  Mice transplanted with BCR-ABL1-transduced AID-/- bone marrow had prolonged survival as compared to mice transplanted with leukemia cells generated from AID+/+ bone marrow.  Consistent with a causative role of AID in genetic instability, AID-/- leukemia had a decreased frequency of amplifications, deletions and a lower frequency of mutations in non-Ig genes including Pax5 and Rhoh as compared to AID+/+ leukemias.  AID-/- and AID+/+ ALL cells showed a markedly distinct gene expression pattern as determined by principle component analysis, with 2,365 genes differentially expressed.  In contrast to AID+/+ leukemia, AID-/- ALL cells failed to downregulate a number of tumor suppressor genes such as Rhoh, Cdkn1a (p21), and Blnk (SLP65).  We conclude that AID accelerates clonal evolution in BCR-ABL1 ALL by enhancing genetic instability, aberrant somatic hypermutation, and by transcriptional inactivation of tumor suppressor genes.

overall_design: We used microarrays to detect differences in gene expression profiles between AID expressing leukemia and AID deficient leukemia

first: Tanja

last: Gruber


title: Innate immune response of murine nasal-associated lymphoid tissue (NALT) to Streptococcus pyogenes infection.

city: Pearl River

first: Ellen

last: Murphy

state: NY

address: 401 N. Middletown Road

department: Vaccines

institute: Wyeth Research

summary:
Streptococcus pyogenes is a major causative agent of tonsillitis or pharyngitis in children, which can lead to more invasive infections and noninfectious sequellae. S. pyogenes can persist in tonsils, while one-third of children treated with antibiotics continue to shed streptococci and have recurrent infections. Mouse nasal-associated lymphoid tissue (NALT) is functionally analogous to human oropharangeal lymphoid tissues. The innate immune responses of naïve cells from a mucosal site to S. pyogenes is not well described; therefore, we infected C57BL/6 mice intranasally with 108 CFU S. pyogenes. Transcriptional responses by NALT after S. pyogenes infection were analyzed by Affymetrix microarray and quantitative RT-PCR.  Wild-type S. pyogenes induces transcription of both type I and IFN-gamma-responsive genes, pro-inflammatory genes, and acute phase response plasma proteins within 24h after infection.   Invasion of NALT and the induction of the interferon response were not dependent on expression of anti-phagocytic M1 protein.  However, infection with an attenuated, less invasive mutant indicated that a robust innate response by NALT is significantly influenced by intra-NALT bacterial load.  Granulocytic populations of NALT, cervical lymph nodes and spleen were discriminated by characteristic surface and intracellular markers.  Intranasal infection induces systemic release of neutrophils and a substantial influx of neutrophils into NALT at 24h, which decline by 48h after infection.  Macrophages do not significantly increase in S. pyogenes-infected NALT.  Intranasal infection of IFN-gamma -/- (GKO) C57BL/6 mice did not lead to systemic dissemination of wild type S. pyogenes, despite reduced expression of IFN-gamma-responsive mRNAs in NALT.   Infected GKO mice had an unregulated influx of neutrophils into NALT compared to immunocompetant mice and mice treated with an anti-IFN-gamma antibody more rapidly cleared S. pyogenes from NALT.  Thus, IFN-gamma-induced responses have a suppressive influence on early clearance of this pathogen from NALT.

Keywords: disease state analysis

overall_design: C57BL/6 mice (6-10 weeks old), 4 per group, were infected intranasally with log-phase Streptococcus pyogenes, 2 to 4 x 10^8 CFU per 15 µl of pyrogen-free PBS.  Sham-infected mice were administered 15 µl of the same PBS.  Mice were infected with wild type strain 90-226 (Cue 1998), a 90-226 strain containing an in-frame deletion of M1 protein (90-226 delta emm1) (Zimmerlein 2005) or an attenuated 90-226 which lacks both M1 and SCPA proteins (90-226att).  NALT was collected from mice at 24h after infection and stored in RNAlater until RNA could be purified).

first: Kendra

last: Hyland

last: Brennan

first: Steven

last: Olmstead

first: Eduardo

last: Rojas

first: Christine

last: Herfst

first: Beinan

last: Wang

first: P

middle: P

last: Cleary


title: Discrimination of Tumorigenic Triazole Conazoles from Phenobarbital by Transcriptional Analyses of Mouse Liver

last: Ward

address: 109 TW Alexander Dr

department: NHEERL

institute: United States EPA

summary: The goal of this study was to apply transcriptional analyses to hepatic tissues from mice exposed to PB, propiconazole (Pro) or triadimefon (Tri) at tumorigenic exposure levels to reveal similarities and differences in response among these treatments.The goal of this study was to apply transcriptional analyses to hepatic tissues from mice exposed to PB, propiconazole (Pro) or triadimefon (Tri) at tumorigenic exposure levels to reveal similarities and differences in response among these treatments.

overall_design: Briefly, male CD-1 mice received propiconazole (2500 ppm), tridimefon (1800 ppm), or phenobarbital (850 ppm) in the feed or, control feed for periods of 4 and 30 days. This resulted in an average daily intake of 349.6 ± 46.7 of propiconazole, 257.1 ± 33.5  of tridimefon or 97.2 ± 5.9 mg/kg/day of phenobarbital.

first: Stephen

last: Nesnow

first: Tanya

last: Moore

first: Hongzu

last: Ren

first: Susan

last: Hester


title: Effect of v-erbA on T3-responsive genes in AML-12 hepatocytes

status: Public on Jun 30 2009

city: Jackson

first: Tereza

last: Ventura-Holman

state: MS

address: 2500 North State St.

institute: University of Mississippi Medical Center

summary: The v-erbA oncogene belongs to a superfamily of transcription factors called nuclear receptors, which includes the thyroid hormone receptors (TRs) responsible for mediating the effects of thyroid hormone (T3). Nuclear receptors bind to specific DNA sequences in the promoter region of target genes and v-erbA is known to exert a dominant negative effect on the activity of the TRs. The repressor activity of v-erbA has been linked to the development of hepatocellular carcinoma (HCC) in a mouse model. We have used microarray analysis to identify genes differentially expressed in hepatocytes in culture (AML12 cells) stably transfected with v-erbA and exposed to T3. We have found that v-erbA can negatively regulate expression of T3-responsive genes known to have a protective function against tumor development. We have also identified a number of v-erbA- (but not T3-) responsive genes that are known to be involved in carcinogenesis and which may play a role in the development of HCC.

overall_design: AML12 control cells and v-erbA-transfected AML12 cells were exposed to 10 nM T3 for 3h or 24h. Using microarray analysis, we compared gene expression in the presence and absence of v-erbA and identified T3-regulated genes differentially expressed in the presence of v-erbA.

first: Abulkhair

last: Mamoon

first: José

last: Subauste


title: Foxl2 functions in sex determination and histogenesis throughout mouse ovary development, analyzed by Affymetrix arrays

city: Paris

first: Chris

last: Ottolenghi

address: rue des Saint-Peres

institute: Inserm/Univ.Paris Descartes

summary:
Comparison of Foxl2-null ovaries to wildtype ovaries, ovaries lacking Wnt4 or Kit, or testes, throughout mouse development.
The goal of this study was to identify early Foxl2 target genes as well as other ovarian, anti-testis genes that may act independently of Foxl2.

Keywords: disease state analysis; genetic modification; developmental study

overall_design: We studied 43 samples over 15 conditions to cover a wide range of wiltype and pathological states showing highly divergent alterations of cell type composition. This was meant to identify the most specific, cell context-independent targets of Foxl2.

first: Emanuele

last: Pelosi


title: Myocardial expression data from gnotobiotic wild-type and Ppara-/- mice

status: Public on Jun 23 2009

city: St. Louis

first: Peter

last: Crawford

state: MO

address: 660 S. Euclid Ave.

institute: Washington University School of Medicine

summary:
Germ free (GF) and conventionalized (CONV-D) wild-type C57Bl/6 male mice in the CARB-fed, 24h fasted, and 30d trained states; plus GF and CONV-D CARB-fed Ppara-/- mice.  CARB-fed indicates a standard polysaccharide-rich mouse chow diet.
CONV-D mice are those that received a microbiota transplant from conventionally raised mice 2-3 weeks before experiment was initiated

Keywords: RNA Expression Array

overall_design: Hearts from six to ten week-old GF and CONV-D mice, from the following eight treatment groups (GF, CONV-D) X (CARB-fed [wild-type], 24h fast [wild-type], 30d trained [wild-type], Ppara-/- [CARB-fed])


title: Differential Cardiac Gene Regulation by Arg- and Gly389 Polymorphic Forms of the beta1-adrenergic Receptor

city: Cincinnati

first: Bruce

last: Aronow

state: OH

institute: Cincinnati Children's Hospital Medical Center

summary:
The beta1-adrenergic receptor (beta1AR; ADRB1) polymorphism Arg 389Gly is located in an intracellular loop and is associated with distinct human and mouse cardiovascular phenotypes. To test the hypothesis that beta1-Arg389 and beta1-Gly389 alleles could differentially couple to pathways beyond that of classic Gs-adenylyl cyclase (AC)/cAMP signaling, we performed comparative gene expression profile analyses on hearts from wildtype and transgenic mice that expressed either human beta1-Arg389 and beta1-Gly389 receptors, or AC5 adenyl cyclase, sampling at an early age and stage, prior to the onset of pathologic features. We observed substantial overlap of dysregulated genes across all three transgenic heart models, consistent with a shared coupling to cAMP-dependent regulation of cardiac processes and adaptive responses.  All three models up-regulated genes associated with RNA metabolism and translation, and down-regulated genes associated with mitochondria and energy metabolism, consistent with cAMP-driven increase in cardiac contractility, protein synthesis, and compensatory down-regulation of mitochondrial energy production. Both beta1AR transgenics activated additional genes associated with kinase-dependent pathways, and uniquely, beta1-Arg389 hearts caused up-regulation of genes associated with inflammation, programmed cell death, and extracellular matrix. These results substantially expand the scope of 7-transmembrane domain receptor signaling propagation beyond known cognate G-protein couplings. Moreover, they implicate alterations of a repertoire of processes evoked by a single amino acid variation in the cardiac beta1AR that might be exploited for genotype-specific heart failure diagnostics and therapeutics.
Keywords: Polymorphism, adrenergic, heart failure, beta-blocker, microarrays.

Keywords: transgenic alteration of target organ (heart)

overall_design: RNA was isolated from hearts obtained from non-transgenic mice, AC5 transgenic mice, beta1-Gly389 transgenic mice and beta1-Arg389 transgenic mice at 90 days of age. 6 biological replicates were used for each genotype.

first: S

middle: M

last: Swift

first: B

last: Gaume

first: K

last: Small

middle: B

last: Liggett


title: Foxl2 functions throughout mouse ovary development


title: Expression profiling of joint tissue from C3H and interval specific congenic mouse lines post- B. burgdorferi infection

status: Public on Jun 22 2009

city: Salt Lake City

first: Janis

last: Weis

state: UT

address: 15 North Medical Drive East

department: Pathology

institute: University of Utah

summary:
Gene expression profile of joint tissue from C3H and interval specific congenic mouse lines (ISCL) following infection with Borrelia burgdorferi
Congenic lines of mice generated between C3H and C57BL/6 with penetrant arthritis severity phenotype were infected with B. burgdorferi
Joint tissue was collected from uninfected control mice and from mice infected for 1 week, RNA was prepared and gene expressed analyzed by Affymetrix microarray.

overall_design: Each sample was comprised of pooled RNA from at least 5 female mice of each genotype.

first: Y

last: Ma

first: J

last: Miller

first: H

last: Crandall

first: E

last: Larsen

first: D

last: Dunn

first: R

last: Weiss

first: M

last: Subramanian

middle: H

last: Zachary

first: C

last: Teuscher


title: RNA species bound by deiminated and non-deiminated MA-Brent-1 (bhatt-affy-mouse-581641)

status: Public on Jun 20 2009

city: Phoenix

first: Winnie

last: Liang

state: AZ

address: 445 N. Fifth Street

institute: Translational Genomics

summary:
We have identified loss of deiminated MA-Brent-1 (an RNA and export binding protein) in the retinal ganglion cells (RGCs) in multiple sclerosis and in glaucoma eyes compared to normal controls. Deimination refers to posttranslational modification of protein bound arginine (not free arginine) in citrulline. Our preliminary studies suggest binding of different repertoire of RNA by non-deiminated and deiminated MA-Brent-1. In vitro, in neurites of cultured RGCs and hippocampal neurons, the select mRNA translation is enhanced by addition of deiminated but not non-deiminated MA-Brent-1. These observations suggest that lack of deiminated MA-Brent-1 has consequences for protein synthesis, remodeling and plasticity of RGCs/neurons. Identification of RNA species bound by deiminated and non-deiminated MA-Brent-1 will enable us there further verification and determining the role that deimination plays in biological function of MA-Brent-1 in multiple sclerosis and glaucoma. To summarize identification of RNA species bound by deiminated and non deiminated MA-Brent-1 will enable us to gain further insight into role of deimination in the overall disease process.
To identify the RNA species bound by deiminated and non-deiminated MA-Brent-1

The deiminanted MA-Brent-1 binds to a different repertoire of RNA and non-deiminated MA-Brent-1.

Corollary to this hypothesis is that the deiminated MA-Brent-1 enhances translation for bound species of RNA. Loss of deimination results in less dendritic protein synthesis and has consequences for remodeling and plasticity in multiple sclerosis and glaucoma RGCs where lack of deimination (compared to control eyes) has been observed.

About 10 microgram of purified recombinant and his-tagged non-deiminated MA-Brent-1 or deiminated MA-Brent-1 will be incubated with about 400 microgram of total RNA. The RNA bound his-tagged protein will be incubated with about 100 microliters of Ni-NTA beads and loaded onto a mini column. The column will be washed with 50 volumes of binding buffer. The bound his-tagged protein will be released using 100 mM immidazole in the binding buffer. The MA-Brent-1 bound RNA will be recovered by boiling the protein in 0.25% SDS followed by purification of RNA using columns or using a 4% PAGE-0.8% agarose composite gel followed by gel purification. The composite gel RNA purification is routine practice in the laboratory.

RNA binding experiment for each protein variant (deiminated or non-deiminated) MA-Brent-1 will be repeated twice. Identified RNA species common in both binding experiments for a given species will be candidates for further pursuit.

The recombinant purified MA-Brent-1 will be subjected to deimination by incubating with recombinant peptidylarginine deiminase type II (PAD2). PAD2 is a C-terminus GST tag fusion protein and will be purified using GST column. Deiminated and non-deiminated MA-Brent-1, C-terminal his-tag fusion proteins will be purified using Ni-NTA beads/column and immidazole containing buffer as eluents.

overall_design: in vitro RNA binding with deiminated and non-deiminated MA-Brent-1

first: Sanjoy

middle: K

last: Bhattacharya


title: In vitro modeling of primordial germ cell development

city: Boston

first: Jason

middle: Allen

last: West

address: One Blackfan Circle Room 7004B

institute: Children's Hospital, Boston

summary:
Transgenic StellaGFP ESCs were used to derive primordial germ cells during embryoid body (EB) differentiation, and microarry analysis used to compared FACS sorted Stella-positive cells of day 7 Ebs with the parental ESCs and Stella-negative cells of day 7 Ebs.
Keywords: in vitro, primordial germ cells, embryonic stem cells, stella

overall_design: Four replicates of Stella-positive day 7 cells were compared to two replicates of stella-negative day 7 cells.  Also, there were three replicated of FACS sorted Stella-negative ESCs, two replicated of FACS-sorted Stella-postive ESCs, and two replicates of unsorted StellaGFP ESCs.


title: Expression data from AtT20 mouse pituitary gland cells following overexpression or down regulation of NSBP1

status: Public on Jun 13 2009

first: Mark

last: Rochman

state: MD

address: 6000 Rockville Pike 37/3122

department: NCI

institute: NIH

summary:
Chromatin architectural protein NSBP1/HMGN5 belongs to the family of HMGN proteins which specifically interact with nucleosomes via Nucleosome Binding Domain, unfold chromatin and affect transcription. Mouse NSBP1 is a new and uncharacterized member of HMGN protein family. NSBP1 is a nuclear protein which is localized to euchromatin, binds to linker histone H1 and unfolds chromatin.
We analyzed the effect of altered expression levels of mouse NSBP1 protein on global gene expression profile in AtT20 pituitary cells. We found that NSBP1 modulates the fidelity of cellular transcription in the nucleosome binding-dependent manner.

overall_design: Stable clones overexpressing wild type mouse NSBP1 protein or mutated NSBP1SE protein which does not bind to nucleosomes were generated by retroviral infection of AtT20 cells. siRNA treatment was applied to down regulate NSBP1 in the cells. Total RNA was collected from biological triplicates and analyzed by Affymetrix expression arrays.


title: Gene expression of RonTK-/- mammary glands compared to RonTK+/+ controls during development

status: Public on Jun 16 2009

first: Susan

last: Waltz

address: 3125 Eden Ave ML0521

institute: University of Cincinnati

summary:
RON WT and RON KO at 5, 6, 7 week virgin mammary glands
In the study, we demonstrated that RON regulates mammary gland branching morphogenesis in pubertal development associated with changes in gene expression.

Keywords: Pubertal mammary glands

overall_design: In the study, we hybridized RNA from 5, 6, 7 week old virgin female RON WT and KO mammary glands to Affymetrix GeneChip Mouse Genome 430 2.0 Array

first: Sara

last: Meyer

first: Glendon

last: Zinser

first: William

last: Stuart


title: Effector cells derived from naïve or central memory pmel-1 CD8+ T cells

status: Public on Jun 10 2009

first: Zachary

last: Borman

address: National Institutes of Health, 9000 Rockville Pike, Building 10 CRC 3W-5816

department: Surgery Branch

institute: National Cancer Institute

summary: Effector cells for adoptive immunotherapy can be generated by in vitro stimulation of naïve or memory subsets of CD8+ T cells. While the characteristics of CD8+ T cell subsets are well defined, the heritable influence of those populations on their effector cell progeny is not well understood. We studied effector cells generated from naïve or central memory CD8+ T cells and found that they retained distinct gene expression signatures and developmental programs. Effector cells derived from central memory cells tended to retain their CD62L+ phenotype, but also to acquire KLRG1, an indicator of cellular senescence. In contrast, the effector cell progeny of naïve cells displayed reduced terminal differentiation, and, following infusion, they displayed greater expansion, cytokine production, and tumor destruction. These data indicate that effector cells retain a gene expression imprint conferred by their naïve or central memory progenitors, and they suggest a strategy for enhancing cancer immunotherapy.

overall_design: Effector cells were generated from naive or central memory CD8+ T cells. The cells were then rested (unstimulated) or restimulated (stimulated). This experimental design resulted in 4 groups (Naïve-derived/stimulated, Naïve-derived/unstimulated, Central memory-derived/stimulated, Central memory-derived/unstimulated). Three replicates from independent experiments were analyzed.

first: Christian

middle: S

last: Hinrichs

first: Lydie

last: Cassard

first: Luca

last: Gattinoni

first: Rosanne

last: Spolski

first: Zhiya

last: Yu

first: Luis

last: Sanchez-Perez

first: Pawel

last: Muranski

first: Steven

last: Kern

first: Carol

last: Logun

first: Douglas

last: Palmer

first: Yun

last: Ji

first: Robert

last: Reger

first: Warren

last: Leonard

middle: L

last: Danner

last: Rosenberg

first: Nicholas

last: Restifo


title: Heterogeneity of LN resident macrophage

status: Public on Jun 05 2009

city: Sydney

first: Tri

middle: Giang

last: Phan

state: NSW

address: 384 Victoria St, Darlinghurst

department: Immunology and Inflammation

institute: Garvan Institute

summary:
LN resident macrophages lining the lymphatic sinuses play critical roles in antigen capture and presentation as well as degradation.

We used microarray to examine global gene expression profiles to compare SCS and med macrophages to determine the underlying molecular basis of their differential handling of antigens.

overall_design: Primary SCS and med macrophages were isolated and purified from LNs by flourescence-activated cell sorting and RNA extracted, amplified and hybridized for Affymetrix analysis.

first: Jason

middle: G

last: Cyster


title: Human CD28+/- tumor infiltrating lymphocytes gene expression

city: houston

first: yufeng

last: li

state: TX

address: 7455 Fannin

department: Melanoma

institute: MDACC

summary:
This analysis reveals the different gene expression profiles between human melanoma tumor infiltrating lymphocytes subsets CD8+CD28+ and CD8+CD28-.

Adoptive cell therapy (ACT) is an effective approach that removes tumor-infiltrating lymphocytes (TILs) from this suppressive tumor microenvironment and expands and activates CD8+ and CD4+ T cells in vitro, differentiating them into potent anti-tumor effector cells that can be re-introduced back into patients. One of the key problems that may limit tumor regression and long-term durable clinical responses in ACT is the persistence of TILs following infusion. Our study has indicated that ex vivo expanded TILs to be hypo-responsive to peptide re-stimulation, manifested in slow entry into cell cycle and increased apoptosis. Phenotypic and functional analysis using a number of different T-cell differentiation markers found that CD28 was markedly down-modulated in post-REP cells, while CD27 and CD57 levels showed no statistically-relevant changes in expression. Here on a global gene expression level, microarray gene chip analysis of sorted CD8+CD28+ and CD8+CD28- post-REP TILs revealed a number of key differences in their gene expression profiles; notably, an increase in KIR family member expression, loss of p53-binding proteins, and lower CD127/IL-7Ra gene expression found in the CD28- population. On top of advancing T cell phenotype, reactivation and memory, this set of data may also provide insight for ACT clinical protocol optimization to improve TIL response in vivo.

overall_design:
After isolation from tumor fragment and ex vivo expansion with IL-2, human tumor infiltrating lymphocytes were sorted by FACS into two subsets as described, followed by an immediate microarray analysis.

Specifically: The tumor samples were cut into 3-5 mm2 pieces and cultured in 2 ml of TIL culture medium (TIL-CM) consisting of RPMI 1640, 10% human AB serum, 1 mM Glutamine, 1 mM sodium pyruvate, 50 mM 2-mercaptoethanol, 1X Penicillin-Streptomcyin (Pen-Strep), and 20 mg/ml Gentmicin (Invitrogen, Carlsbad, CA) in 24-well plates.  IL-2, at 6,000 U/ml was added to all wells. The dividing TIL lines were fed with fresh TIL-CM containing 3,000 U/ml IL-2 (Proleukin® from Novartis) and sub-cultured 1:1 in TIL-CM with 6,000 U/ml IL-2 to maintain viable cell density between 0.5-2 x 10e6/ml.  These TILs were designated as “pre-REP” and were subjected to continued expansion using the “rapid expansion protocol” (REP) originally designed by Riddell and Greenberg.  Briefly, 1.3 x 10e5 pre-REP TILs in TIL-CM were added to upright T-25 flasks containing 30 ng/ml OKT3 and 26 x 10e6 allogeneic irradiated (5,000 cGy) PBMC feeder cells obtained from 4 pooled normal donor PBMC (obtained from the Gulf Coast Regional Blood Bank, Houston, TX) in 20 ml of TIL-CM.  On day 2, 10 ml of AIMV (Invitrogen) was added together with 6,000 U/ml IL-2 (final concentration). The TILs were expanded for another 12 days and diluted as needed with AIMV and IL-2 to keep the viable cell density between 1-4 x 106/ml. Using this methodology, routinely 50-100 x 10e6 “post-REP” TIL were isolated from each starting T-25 flask culture. Post-REP TILs were freshly harvested from the flasks and sorted after anti-CD4 and anti-CD28 staining into CD4-, CD28+ and CD4-, CD28- two subsets. After a 3 h rest period to shed off staining antibodies post sorting, TILs were processed with Qiagen RNeasy kit to obtain RNA for microarray experiment. 1 mg of RNA of each sample was subjected to reverse transcription and probe blotting using an Illumina kit, where human Ref6 chip was used (Illumina, Inc.).

first: Yufeng

last: Li

first: Laszlo

last: Radvanyi

first: Patrick

last: Hwu


title: Enhancing CD8 T Cell Memory by Modulating Fatty Acid Metabolism

city: Philadelphia

first: Li-San

last: Wang

address: 1424 Blockley Hall, 423 Guardian Drive

department: Pathology and Laboratory Medicine

institute: University of Pennsylvania

summary: CD8 T cells play a crucial role in immunity to infection and cancer.  They are maintained in constant numbers, but upon stimulation with antigen undergo a developmental program characterized by distinct phases encompassing the expansion and then contraction of antigen-specific populations, followed by the persistence of long-lived memory cells.  Although this predictable pattern of a CD8 T cell response is well established, the underlying cellular mechanisms regulating the transition to memory remain undefined.  Here we show that TRAF6, an adapter protein in the TNF-receptor (TNFR) and IL-1R/TLR superfamily, regulates CD8 T cell memory development following infection by modulating fatty acid metabolism. We show that mice with a T cell-specific deletion of TRAF6 mount robust primary CD8 T cell effector responses, but have a profound defect in their ability to generate memory. This defect is CD8 T cell intrinsic and is characterized by the disappearance of antigen-specific cells in the weeks following primary immunization.  Microarray analyses revealed that TRAF6-deficient CD8 T cells from early timepoints following immunization exhibit altered expression of genes that regulate fatty acid metabolism.  Consistent with this, activated CD8 T cells lacking TRAF6 are unable to upregulate mitochondrial β-oxidation in response to growth factor withdrawal in vitro.  Treatment with drugs that induce fatty acid oxidation enabled CD8 T cell memory generation in the absence of TRAF6.  Remarkably, these treatments also increased CD8 T cell memory in wild type mice, and consequently were able to significantly improve the efficacy of an experimental anti-cancer vaccine.

overall_design: CD8 T cells from two mouse strains (OTI-WT and OTI-TRAF6 knockout) at two timepoints (6d with 3 replicates and 10d with 5 replicates) after infection are used.

first: Erika

last: Pearce

first: Matthew

last: Walsh

first: Pedro

middle: J

last: Cejas

first: Gretchen

last: Harms

first: Hao

last: Shen

first: Russell

last: Jones

first: Yongwon

last: Choi


title: E2F4 cooperates with pRB in the development of extra-embryonic, not embryonic, tissues

status: Public on Jun 08 2009

first: Charles

middle: Arthur

last: Whittaker

state: MA

institute: Koch Institute

summary:
Homozygous mutation of the murine retinoblastoma tumor suppressor gene, Rb, results in embryonic lethality between E13.5 and E15.5 with defects in cellular proliferation, differentiation and apoptosis.  Many of these defects are suppressed by mutation of an activating E2F, E2f1 or E2f3, indicating that they are key downstream targets of the retinoblastoma protein, pRB. In this study, we assess how E2F4 contributes to the developmental consequences of pRb-loss.  In stark contrast to the activating E2Fs, the homozygous mutation of E2f4 shortened the lifespan of Rb-/- embryos. This resulted from an exacerbation of the placental defect of the Rb-/- mice indicating that E2F4 and pRB cooperate in the development of this tissue. Further analyses indicated that this defect reflects an increase in trophectoderm-like cells. Under conditions where the placenta was wild-type but the embryo mutant for E2f4 and pRb embryos survived to birth and exhibited all of the defects that were observed in the E2f4 and Rb single mutant embryos. Thus, while pRB and E2F4 cooperate in placental development, they play largely non-overlapping roles the development of many embryonic tissues.

Keywords: genetically modified tissue comparison

overall_design: In one experiment, embryonic Day 13.5 placentas from wild-type and Rb -/- knock-out mice were compared. In the second experiment, embronic day 11.5 placentas from wild-type and Rb -/-;E2f4 -/- double knock-out mice were compared.

first: Eunice

middle: Y

last: Lee

first: Tina

last: Yuan

first: Jacqueline

last: Lees


title: Gene expression data from gastrocnemius muscle (m.Gas) in young adult mice

first: Harvey

last: Armbrecht

address: #1 Jefferson Barracks Drive

department: Division of Geriatric Medicine & Department of Molecular Microbiology & Immunology

institute: St. Louis Veterans Affairs Medical Center , & Saint Louis University School of Medicine

summary: This study examined the effects of castration and testosterone replacement on global differential gene transcription in the gastrocnemius muscle (m.Gas) in young adult mice over 14-days.

overall_design: Each of the 3 treatment groups was composed of 3 independent biological replicates, totaling 9 arrays. Equal amounts of the highest quality total RNA (4 µg) from three animals were pooled for each biological replicate, totaling the use of 27 animals in the 9-array experiment.  The balanced design of this study enabled the use of a 1-way ANOVA, which was used to statistically determine which of the >34,000 genes (>45,000 probes) were differentially expressed between groups. The three pair-wise comparisons were: Castration vs. Sham (castration (C) effect); Testosterone vs. Castration (testosterone replacement (T) effect); and Testosterone vs. Sham (overall effects).

first: Akbar

last: Siddiqui

middle: T

last: Haren

first: H