title: Global gene expression profile of RAW264.7 cells with or without SWCNT exposure

status: Public on Jul 08 2009

first: Bin

last: Wan

address: 18 shuangqing road, Haidian District

institute: Research centre for eco-environmental sciences, CAS

summary: Carbon nanotubes (CNTs) are one of the most attractive nanomaterials that have many potential applications in industrial, medical, and environmental sectors. To investigate the toxic effect of single-walled CNTs (SWCNTs) on the immune system, we examined the influences of SWCNTs (depleted of metal impurities) on viability and global gene expression in murine macrophages. RAW264.7 Cells were exposed to various concentrations of SWCNTs, and cell viability was analyzed at various time points using WST-1 assay. SWCNT cytotoxicity in macrophages was concentration and time dependent, with significant viability loss occurring within 4 hr at 50μg/ml SWCNT exposure. As low as 1μg/ml SWCNTs reduced cell viability at 12 hr, whereas 50μg/ml SWCNT exposure completely abolished cell proliferation. Laser confocal microscopy indicated that SWCNTs could enter and aggregate in cell cytoplasm and nuclear areas. Microarray, real-time reverse-transcriptase polymerase chain reaction and enzyme-linked immunosorbent assay revealed that SWCNT altered gene and protein expression pattern. The most differentially expressed genes were related to proinflammatroy cytokines and chemokine, such as CCL-3 and CCL-4, apoptosis (e.g., Caspase-1, and -3), protective response such as Hmox-1and GSTM3) and cell surface molecules such as ICAM-1, Itgb2, indicating a possible involvement of SWCNT in inflammation and pulmonary granuloma formation, carcinogenesis and Th2 polarization of T cells. Furthermore, a substantial amount of down-regulated genes were related to ribosomal assembly and mitochondrial respiration chain, implicating a possible mechanism of SWCNT-induced oxidative stress and gene expression inhibition. The results are a first step in identifying molecular targets for the toxicity of SWCNTs and the elucidating of the molecular mechanisms for the immune modulation ability of SWCNTs.

overall_design: Cells were exposure to 50ug/ml of acid-functionzalized SWCNTs for 24 hours before gene expression profiling, cells without SWCNT exposure were used as control.


title: Specificity of the zebrafish host transcriptome response to acute and chronic mycobacterial infection

status: Public on Jul 31 2009

city: Leiden

first: Anna

middle: Magdalena

last: Zakrzewska

address: Wassenaarseweg 64

department: IBL

country: Netherlands

institute: University of Leiden

summary: Pathogenic mycobacteria have the ability to survive within macrophages and persist inside granulomas composed of host immune cells. The complex host-pathogen interactions that determine the outcome of a mycobacterial infection process result in marked alterations of the host gene expression profile. Here we used the zebrafish model to investigate the specificity of the host response to infections with two mycobacterium strains that give distinct disease outcomes: an acute disease with early lethality or a chronic disease with granuloma formation, caused by Mycobacterium marinum strains Mma 20 and E11, respectively. We performed a microarray study of different stages of disease progression in adult zebrafish and found that the acute and the chronic strains evoked partially overlapping host transcriptome signatures, despite that they induce profoundly different disease phenotypes. Both strains affected many signaling cascades, including Wnt and Tlr pathways. Interestingly, the strongest differences were observed at the initial stage of the disease. The immediate response to the acute strain was characterized by higher expression of genes encoding MHC class I proteins, matrix metalloproteinases, transcription factors, cytokines and other common immune response proteins. In contrast, small GTPase and histone gene groups showed higher expression in response to the chronic strain. We also found that nearly 1,000 mycobacterium-responsive genes overlapped between the expression signatures of infected zebrafish adults and embryos at different stages of granuloma formation. Since adult zebrafish possess an adaptive immune system similar to mammals and zebrafish embryos rely solely on innate immunity, this overlap indicates a major contribution of the innate component of the immune system in the response to mycobacterium infection. Taken together, our comparison of the transcriptome responses involved in acute versus chronic infections and in the embryonic versus adult situation provides important new leads for investigating the mechanism of mycobacterial pathogenesis.

overall_design:
Zebrafish were handled in compliance with the local animal welfare regulations and maintained according to standard protocols (http://ZFIN.org). Infection experiments were approved by the local animal welfare committee (DEC) of the VU University medical center and of Leiden Univeristy. Infection experiments with adult fish were performed on young males selected from a wild type laboratory-breeding colony and acclimated to their new environment for one week in a quarantine area. These fish were kept at 28˚C on a 12:12 h light/dark rhythm throughout the experiment. Groups of 30 fish, infected with the same dose and strain of mycobacteria, were kept in small fish tanks (10 l) with their own separate filtering system (Eheim Ecco). Zebrafish were inoculated intraperitoneally as previously described (Meijer et al., 2004) with approximately 10000 bacteria or with phosphate-buffered saline (PBS) as a control. For the acute infection study with E11 and Mma20 strains, 3 fish per group were sacrificed at 1 and 6 days post infection (dpi) and used for microarray analysis. For comparison with the end stage of chronic E11 infection we used RNA samples from our previously published chronic infection study (Meijer et al., 2005; control fish c2 and infected fish i2) and additional RNA samples (2 controls, 2 infected) from a similar infection experiment. All chronically infected fish showed overt signs of fish tuberculosis, including lethargy and skin ulcers. Histological examination of fish from the same experiments confirmed that the pathology of infected fish corresponded to fish tuberculosis (Van der Sar et al., 2004) and that no characteristics of the disease were present in the control fish.
Infection experiments at the embryonic stage were performed using mixed egg clutches from different pairs of AB strain zebrafish. Embryos were grown at 28,5 -30 °C in egg water (60µg/ml Instant Ocean see salts) and for the duration of bacterial injections embryos were kept under anaesthesia in egg water containing 0.02% buffered 3-aminobenzoic acid ethyl ester (tricaine, Sigma). Embryos were staged at 28 hours post fertilization (hpf) by morphological critera (Kimmel et al.) and approximately 50 cfu of E11 bacteria were injected into the caudal vein close to the urogenital opening. As a control an equal volume of PBS was likewise injected. Infection experiments were carried out in triplicate on separate days and pools of 15-20 embryos were taken at 2, 24 and 120 hours post infection (hpi).

first: Astrid

middle: M

last: van der Sar

first: Herman

middle: P

last: Spaink

first: Wilbert

last: Bitter

first: Annemarie

middle: H

last: Meijer


title: Gene expression imprinting of N2a sublines upon infection with the prion strain 22L

status: Public on Jul 07 2009

city: Paris

first: Jean-Louis

last: Laplanche

address: 4 avenue de l'Observatoire

department: Biologie Cellulaire

country: France

institute: Faculté de Pharmacie Université Paris5

summary:
Cell culture models allow prion propagation studies ex vivo after contact with infectious brain homogenates. To date, among the neural cell lines, the mouse neuroblastoma-derived cell line N2a has been one of the most widely used model and has yet provided interesting insights into cell biology of prion propagation. Remarkably, persistently-infected N2a sublines have been set up and replicate prions without exhibiting any pathological changes. One further interesting feature of N2a is the possibility to establish by subcloning, sublines with a range of susceptibility to prions. Indeed, susceptible sublines propagate prions and accumulate the pathogenic isoform of the prion protein, PrPSc, at the opposite of resistant sublines. The aim of our study was to apply large-scale expression analysis using microarrays combined with quantitative real-time PCR to examine the gene expression profile in a persistently-infected N2a cell line, N2a58, infected with the mouse-adapted prion strain 22L, to seek for prion-specific gene transcription. We also questioned if we could observe identical variations of expression of these genes in three other 22L-infected N2a sublines. Finally, we examined the transcriptional state of a N2a subline considered as resistant when exposed to prions. Common pathways of gene transcription would disclose information on the molecular basis of the cell infection and help to identify potential therapeutic targets.
Keywords: other

overall_design: 18 DNA-chip hybridizations of cDNAs were made

first: Stephanie

last: Chasseigneaux

first: Manuela

last: Pastore

first: Luce

last: Dauphinot

first: Sylvain

last: Lehmann

first: Jacques

last: Callebert

first: Alexander

last: Burkle

first: Nicolas

last: Sevenet


title: Investigating H. Influenzae Species Diversity by Comparative Genomic Hybridization

status: Public on Sep 09 2009

city: Rockville

first: Chun-Hua

state: MD

address: 9704 Medical Center Dr

department: Pathogen Functional Genomics Resources Center (PFGRC)

country: USA

institute: The J. Craig Venter Institute

summary:
Haemophilus influenzae(Hi) is a gram-negative rod shaped bacterium that lives symbiotically in the upper respiratory tract of humans. Capsulated Hi, as well as non-capsulated strains are known to cause a number of significant infections. Recognizing and understanding the links between the phenotypic traits and the genetic background has paramount epidemiological and clinical importance. To date, a panoply of microbiological and molecular biology tools have been developed and utilized by researchers aimed at identifying the evolutionary links among strains and isolates. Comparative genomic hybridization (CGH) has been shown to be a useful tool for screening strains for their genetic content. However, there is a major limitation when CGH is conducted on a microarray based on a single reference genome. CGH results report which genes are present or absent relative to the genome. Hence the information about novel genetic content that the query strain possesses remains obscure. We report here the construction of the first Hi pan-genome microarray representing ca. 4600 features by 70-mers. In addition to those from the Rd strain, new features originate from the unfinished genome sequences present in NCBI database and from our novel gene discovery project efforts using strain HK1212. Genomes of 20strains belonging to different phylogenetic lineages were screened for their gene loss and gain utilizing the species microarray.  The results obtained by employing the multistrain species microarray provide comprehensive information about the genomic content of uncharacterized strains. The trees generated by CGH, in general, do not reproduce the phylogeny of a species in terms of vertical evolution, but instead represents the overall relatedness of genomes to one another and provide an assessment about the species genome evolution.
Keywords: Comparative Genomic Hybridization

overall_design: Twenty query strains (denoted HKxxxx) were investigated in this study, with each query strain hybridized against the reference strain, KW20.  Two dye-swap experiments were performed, for a total of four hybridizations per query strain.  Each 70mer oligo spotted on the H. influenzae species microarray is replicated six times.  Positive controls on the array consist of oligos designed from the sequenced reference genome, KW20, and negative controls on the array consist of oligos designed from the thale cress plant, Arabidopsis thaliana.

first: L.

last: Papazisi

first: S.

last: Ratnayake

first: B.

middle: G

last: Remortel

first: G.

last: Bock

first: Q.

last: Sun

first: M.

last: Kilian

first: R.

middle: D

last: Fleischmann

middle: N

last: Peterson


title: Subtractive Hybridization Reveals Novel Genes Associated with D. melanogaster Early Embryogenesis.

status: Public on Jul 01 2009

city: Santiago

first: Alejandro

last: Zúñiga

address: Macul 5540

country: Chile

institute: INTA - Universidad de Chile

summary:
Background: The early stages of D. melanogaster embryogenesis involve cell migration and pattern formation, and lead to the formation of three germ layers (the ectoderm, mesoderm and endoderm). These developmental events are controlled by differential gene activity. In the current study we used a suppressive subtractive hybridization (SSH) procedure to identify a group of genes potentially involved in D. melanogaster early embryogenesis and to study the temporal activity of developmentally regulated genes at five different intervals covering 12 stages of embryogenesis.
Results: Macroarrays were constructed to confirm induction of expression and determine the temporal profile of isolated subtracted cDNAs during embryo development. We identified a set of 118 genes that significantly increased their expression levels at least at one developmental interval compared with the reference interval. 53.4% of them have a phenotype and/or molecular function reported in the literature, whereas 46.6% are essentially uncharacterized.  Clustering analysis revealed demarcated transcript groups with maximum gene activity at distinct developmental intervals. In situ hybridization assays were carried out on 30 of the transcripts and of these, 19 (63%) proved to have restricted expression patterns. 11 of the uncharacterized genes that exhibited temporal and spatially restricted patterns of expression in developing embryos encode putative secreted and transmembrane proteins. For three of them we validated our protein sequence predictions by expressing their cDNAs in S2R+ cells and analyze the subcellular distribution of recombinant proteins.
Conclusions: Our data provides a list of developmentally regulated D. melanogaster genes and their expression profiles during embryogenesis, including novel information on the temporal and spatial expression patterns of several previously uncharacterized genes. In particular, we recovered a significant number of novel genes encoding putative secreted and transmembrane proteins, suggesting new components of signalling pathways that might be incorporated within the existing regulatory networks controlling Drosophila embryogenesis; they are also good candidates for more functional targeted analyses.

Keywords: Drosophila melanogaster, gastrulation, macroarray, developmental time course

overall_design: We generated a subtracted stage specific cDNA library representing genes differentially expressed between Drosophila melanogaster synciyial blastoderm and gastrula stages. Clones were spotted onto nylon membranes and hybridized against cDNA probes representing mRNA from embryos at five different developmental intervals, in order to obtain time course expression data covering the first 12 stages of Drosophila development. We designed macroarrays containing either 302 non-redundant cDNAs, including protein-coding sequences, ncRNA, transposons, introns and intergenic regions (membranes A, B and C) and the complete subtracted library (579 cDNAs, membrane D). Each clone was spotted in duplicate. The following controls were spotted onto membranes: (1) a fragment of the vector pBluescript II obtained by amplification with the T7 and SP6 universal primers; (2) several spots of DMSO 50% (solvent used); (3) PCR amplified fragment of genes serendipity α, twist, tinman, fog and snail as positive controls, and (4) actin, tubulin, and RP49 as housekeeping, and (5) four dilutions of a PCR amplified fragment from a B. subtilis dap cDNA (ATCC; number 87486), which hybridizes to an in vitro synthesized poly(A)-RNA that was added to the embryo RNA samples (dilution 1/200) prior to the labelling process and used as spike mRNA (Kane et al., 2000).

first: Christian

last: Hödar

first: Patricia

last: Hanna

first: Freddy

last: Ibáñez

first: Luis

last: Pastenes

first: Pablo

last: Moreno

first: Mauricio

last: González

first: Verónica

last: Cambiazo


title: Interleukin-10 is a negative regulatory factor of CAWS-vasculitis in CBA/J mice

status: Public on Jun 30 2009

city: Ibaraki

first: Naoki

last: Osada

state: Osaka

address: 7-6-8 Saito-Asagi

department: Division of Biomedical Research Resources

country: Japan

institute: National Institute of Biomedical Innovation

summary: The gene expression difference between CBA/J and DBA/2 mice after the injection of CAWS (Candida albicans water-soluble fraction) was compared using an oligo-DNA microarray.

overall_design: CAWS was injected into CBA/J and DBA/2 mice. 21 days after the injection, RNA was extracted from the splenocytes and measured using the mouse oligo-DNA 10K micoarray.

first: Noriko

last: Miura

first: Motohiko

last: Komai

first: Yoshiyuki

last: Adachi

first: Yosuke

last: Kameoka

first: Kazuo

last: Suzuki

first: Naohito

last: Ohno


title: Gene expression signature involved in estrogen receptor status and prognosis of breast cancers.

city: Toronto

first: Dong-Yu

last: Wang

address: 610 University Ave

country: Canada

institute: UHN-PMH-OCI

summary:
Fine needle aspiration biopsies (FNABs) of breast cancers were taken before and after surgeries from 16 patients. The cDNA microarray data were used to determine the gene expression profile responding to patient's clinical finding and tumor's pathological changes. A gene profile was generated as Estrogen Receptor Gene Signature (ERGS). The ERGS was verified in a reference dataset and correlated with patient's prognosis significantly.

Keywords: Breast cancer, fine needle biopsy, estrogen receptor, prognostic gene signature

overall_design: Dye-swap technical replicates were included both FNABs taken before and after surgeries for every patient, then the four replicated array data per patient were combined for analysis.

first: Dong

middle: Y

first: Vietty

last: Wong

first: Wey

middle: L

last: Leong


title: Expression defined classification of subtypes of diffuse large B cell lymphoma

status: Public on Jul 10 2009

city: Nagoya

first: Masao

last: Seto

state: Aichi

address: Kanokoden 1-1

department: Dept. of molecular medicine

institute: Aichi Cancer Certer Res. Ins.

summary: Diffuse large B-cell lymphoma (DLBCL) comprises molecularly distinct subgroups such as activated B-cell-like (ABC) and germinal center B-cell-like (GCB) DLBCLs. We previously reported that CD5(+) and CD5(-)CD10(+) DLBCL constitute clinically relevant subgroups. To determine whether these 2 subgroups are related to ABC and GCB DLBCLs, we analyzed the genomic imbalance of 99 cases (36 CD5(+), 19 CD5(-)CD10(+), and 44 CD5(-)CD10(-)) using array-based comparative genomic hybridization (CGH). Forty-six of these cases (22 CD5(+), 9 CD5(-)CD10(+), 1 CD5 (-), and 14 CD5(-)CD10(-)) were subsequently subjected to gene-expression profiling, resulting in their division into 28 ABC (19 CD5(+) and 9 CD5(-)CD10(-)) and 18 GCB (3 CD5(+), 7 CD5(-)CD10(+), and 8 CD5(-)CD10(-)) types. A comparison of genome profiles of distinct subgroups of DLBCL demonstrated that (1) ABC DLBCL is characterized by gain of 3q, 18q, and 19q and loss of 6q and 9p21, and GCB DLBCL is characterized by gain of 1q, 2p, 7q, and 12q; (2) the genomic imbalances characteristic of the CD5(+) and CD5(-)CD10(+) groups were similar to those of the ABC and GCB types, respectively. These findings suggest that CD5(+) and CD5(-)CD10(+) subgroups are included, respectively, in the ABC and GCB types. Finally, when searching for genomic imbalances that affect patients' prognosis, we found that 9p21 loss (p16(INK4a) locus) marks the most aggressive type of DLBCL.

overall_design: Expression profiles in 46 patients with DLBCL and 8 for normal lymph node biopsy samples.

first: Miyuki

last: Katayama

first: Hiroyuki

last: Tagawa

first: Keiichiro

last: Honma

last: Nakagawa

first: Yoshio

last: Miki


title: H3K64me3 and H3K9me3 profiling at promoters in mouse embryonic stem cells

status: Public on Jun 28 2009

city: Basel

first: Fabio

last: Mohn

address: Maulbeerstrasse 66

country: Switzerland

institute: Friedrich Miescher Institute for Biomedical Research

type: Genome binding/occupancy profiling by genome tiling array

summary:
Histone modifications are central to the regulation of all DNA-dependent processes but the repertoire of known modifications is far from complete. Lysine 64 of Histone H3 (H3K64) lies within the globular domain of H3 at a structurally important position within the nucleosome. We identify tri-methylation of H3K64 (H3K64me3) as a modification enriched in pericentric heterochromatin and associated with repeated sequences and transcriptionally inactive genomic regions. Interestingly, the pericentric enrichment of H3K64me3 depends on the Suv39h methyltransferases. Further, we show that this newly identified mark is dynamically regulated during the two major epigenetic reprogramming events in mammals. In primordial germ cells (PGCs) H3K64me3 is present at the time of specification, but disappears transiently during germline reprogramming. In the early mouse embryo it is inherited exclusively through the maternal germline and specifically enriched in heterochromatic regions of the maternal pronucleus. Subsequently the modification is rapidly removed from the maternal chromatin suggesting an important role for H3K64me3 turnover in development. Taken together, our findings firmly establish H3K64me3 as a novel histone modification mark of repressive chromatin, which displays crosstalk to the Suv39 pathway and is dynamically reprogrammed during development. We hypothesize that methylation of this lysine helps to 'secure' the nucleosome and perhaps the surrounding heterochromatin, in an appropriately repressed state during development.

Keywords: ChIP-chip, embryonic stem cells, H3K64me3, H3K9me3, heterochromatin

overall_design: ChIP-chip experiments were performed with four independent biological replicates for H3K64me3 and two independent replicates for H3K9me3. For each condition hybridizations include a dye-swap experiment.

first: Sylvain

last: Daujat

first: Thomas

last: Weiss

first: Ulrike

middle: C

last: Lange

first: Céline

last: Ziegler-Birling

last: Zeissler

first: Michael

last: Lappe

first: Dirk

last: Schübeler

first: Maria-Elena

last: Torres-Padilla

first: Robert

last: Schneider


title: Normal fertile Spermatozoal RNA Profiles

city: Detroit

first: Stephen

middle: A

last: Krawetz

state: MI

address: 275 E. Hancock, CS Mott Center

department: Obstetrics and Gynecology

institute: Wayne State University

summary: Numerous studies have shown the potential of spermatozoal RNAs to delineate failures of spermatogenic pathways in infertile samples.  However, the RNA contribution of normal fertile samples still needs to be established in relation to transcripts consistently present in human spermatozoa.  We report here the spermatozoal transcript profiles characteristic of 24 normally fertile individuals. RNA was extracted from the purified sperm cells of ejaculate and hybridized to Illumina Human-8 BeadChip Microarrays

overall_design: Spermatozoal RNAs were prepared from the semen samples of 24 fertile individuals who fathered at least one child. On each array, 18 normal fertile samples were hybridized along with somatic samples which were used as a hybridization control.  Four samples of whole testis RNA were also used as references.

first: Claudia

last: Lalancette

first: Adrian

middle: E

last: Platts

first: Graham

last: Johnson

first: Ben

last: Emery

first: Douglas

middle: T

last: Carrell


title: DRG co-culture microarray

status: Public on Jun 24 2009

city: New York City

first: prabhjot

last: dhadialla

state: NY

address: 1200 York Avenue

institute: Rockefeller University

summary: Gene regulatory interactions that shape developmental processes can often can be inferred from microarray analysis of gene expression, but most computational methods used require extensive datasets that can be difficult to generate. Here, we show that maximumentropy network analysis allows extraction of genetic interactions from limited microarray datasets. Maximum-entropy networks indicated that the inflammatory cytokine TNF-_ plays a pivotal role in Schwann cell–axon interactions, and these data suggested that TNF mediates its effects by orchestrating cytoplasmic movement and axon guidance. In vivo and in vitro experiments confirmed these predictions, showing that Schwann cells in TNF_/_ peripheral sensory bundles fail to envelop axons efficiently, and that recombinant TNF can partially correct these defects. These data demonstrate the power of maximum-entropy network-based methods for analysis of microarray data, and they indicate that TNF-_ plays a direct role in Schwann cell–axon communication.

overall_design: Samples for microarray analysis were obtained in triplicate from separate coculture slips at 0, 0.5, 1, 6, 12, 24, 36, and 48 h after ascorbic acid addition and were prepared for use on Illumina Mouse-8 chips (Illumina) by the Rockefeller Microarray Core Facility (New York, NY).

first: Prabhjot

middle: S

last: Dhadialla

first: Ifije

last: Ohiorhenuan

first: Andrew

last: Cohen

first: Sidney

last: Strickland


title: Expression of survivin in lung eosinophils is associated with pathology in a mouse model of allergic asthma 2

status: Public on Jun 15 2009

city: Brisbane

first: Ashley

last: Connolly

address: St Lucia

country: Australia

institute: University of Queensland

summary:
Humans vary markedly in their propensity to develop asthma, despite often being exposed to similar environmental stimuli. Similarly, mouse strains vary in susceptibility to airways pathology in experimental asthma. Sensitization and aerosol challenge with ovalbumin (OVA) induces eosinophil accumulation, mucus production and airways obstruction in BALB/c and C57BL/6 mice. In contrast, CBA/Ca mice show relatively little pathology. Allergen-induced production of IL-4, IL-5, IL-10 and IFN-g was detected in all three strains, with BALB/c mice generating the highest levels of IL-4, IL-5 and IL-10. Microarray analysis was used to identify genes differentially regulated in lung tissue after OVA challenge. Differentially regulated genes in the lungs of the asthma-susceptible C57BL/6 and BALB/c strains numbered 242 and 145, respectively, whereas only 42 genes were differentially expressed in the resistant CBA/Ca strain. In C57BL/6 mice, transcripts were enriched for adhesion molecules and this was associated with high levels of eosinophil recruitment. Differentially regulated genes in the lungs of only the asthma-susceptible strains numbered 64 and several of these have not previously been associated with asthma. Many of the genes differentially regulated in the susceptible strains were enzymes involved in inflammation. Using network analysis, mRNA for the anti-apoptotic protein survivin was found to be up-regulated in the lungs following allergen challenge. Survivin mRNA and protein were also expressed at high levels in eosinophils recovered by bronchoalveolar lavage from BALB/c and C57BL/6 mice. We propose that rapid apoptosis of lung eosinophils due to low expression of survivin contributes to the limitation of pathology in CBA/Ca mice
Humans vary markedly in their propensity to develop asthma, despite often being exposed to similar environmental stimuli. Similarly, mouse strains vary in susceptibility to airways pathology in experimental asthma. Sensitization and aerosol challenge with ovalbumin (OVA) induces eosinophil accumulation, mucus production and airways obstruction in BALB/c and C57BL/6 mice. In contrast, CBA/Ca mice show relatively little pathology. Allergen-induced production of IL-4, IL-5, IL-10 and IFN-g was detected in all three strains, with BALB/c mice generating the highest levels of IL-4, IL-5 and IL-10. Microarray analysis was used to identify genes differentially regulated in lung tissue after OVA challenge. Differentially regulated genes in the lungs of the asthma-susceptible C57BL/6 and BALB/c strains numbered 242 and 145, respectively, whereas only 42 genes were differentially expressed in the resistant CBA/Ca strain. In C57BL/6 mice, transcripts were enriched for adhesion molecules and this was associated with high levels of eosinophil recruitment. Differentially regulated genes in the lungs of only the asthma-susceptible strains numbered 64 and several of these have not previously been associated with asthma. Many of the genes differentially regulated in the susceptible strains were enzymes involved in inflammation. Using network analysis, mRNA for the anti-apoptotic protein survivin was found to be up-regulated in the lungs following allergen challenge. Survivin mRNA and protein were also expressed at high levels in eosinophils recovered by bronchoalveolar lavage from BALB/c and C57BL/6 mice. We propose that rapid apoptosis of lung eosinophils due to low expression of survivin contributes to the limitation of pathology in CBA/Ca mice

overall_design: Changes in gene expression in the lungs of 4 individual of BALB/c mice challenged with OVA were monitored using the lungs of 4 individual BALB/c mice challenged with PBS as the control. The microarray analysis was performed in quadruplicate.  Changes in gene expression in the lungs of 4 individual of CBA/Ca mice challenged with OVA were monitored using the lungs of 4 individual CBA/Ca mice challenged with PBS as the control. The microarray analysis was performed in quadruplicate.  Changes in gene expression in the lungs of 4 individual of BALB/c mice challenged with OVA were monitored using the lungs of 4 individual CBA/Ca mice challenged with OVA as the control. The microarray analysis was performed in quadruplicate.  Twelve dual channel microarray slides were used in the overall design of this experiment.

first: Damon

middle: J

last: Tumes

middle: R

first: Lindsay

last: Dent


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

first: Chris

last: Ottolenghi

address: rue des Saint-Peres

institute: Inserm/Univ.Paris Descartes

summary:
Partial loss of function of the transcription factor FOXL2 leads to premature ovarian failure in women. In animal models, Foxl2 is required for maintenance, and possibly induction, of female sex determination independently of other critical genes, i.e., Rspo1 and Wnt4. Here we report expression profiling of mouse ovaries that lack Foxl2 alone or in combination with Wnt4 or Kit/c-Kit, to identify ovarian targets of Foxl2 that, along with some testis genes, were dysregulated during embryonic development. Loss of one copy of Foxl2 revealed strong gene dosage sensitivity, with molecular anomalies that were milder but resembled ovaries lacking both Foxl2 alleles. Furthermore, a Foxl2 transgene disrupted embryonic testis differentiation and increased the levels of key female markers. The results, including a comprehensive principal component analysis of published microarray datasets 1) support the proposal of dose-dependent Foxl2 function and anti-testis action throughout ovary differentiation; and 2) identify candidate genes for a role in sex determination independent of FOXL2 (notably, the transcription factor, ZBTB7C) and in the generation of the ovarian reserve downstream of it (e.g., the cadherin-domain protein CLSTN2, or the sphingomyelin synthase, SGMS2). The gene inventory provides a framework to analyze the genetic bases of ovarian development and female fertility.

Keywords: reference design

overall_design: Comparison of Foxl2+/+,+/- and -/- whole ovaries at 2 timepoints delineating follicle formation

first: Jose-Elias

last: Garcia-Ortiz

first: Emanuele

last: Pelosi

first: Shakib

last: Omari

first: Timur

last: Nedorezov

first: Yulan

last: Piao

first: Jesse

last: Karmazin

first: Manuela

last: Uda

first: Antonio

last: Cao

first: Steve

last: Cole

first: Antonino

last: Forabosco

first: David

last: Schlessinger


title: Definition, conservation and epigenetics of housekeeping and tissue-enriched genes

status: Public on Jun 10 2009

city: Seattle

first: Amit

middle: V

last: Kulkarni

state: WA

address: 401 Terry Ave N

department: Genomics

institute: Rosetta Inpharmatics / Merck Pharmaceuticals

summary:
Housekeeping genes (HKG) are constitutively expressed in all tissues while tissue-enriched genes (TEG) are expressed at a much higher level in a single tissue type than in others.  HKGs serve as valuable experimental controls in gene and protein expression experiments, while TEGs tend to represent distinct physiological processes and are frequently candidates for biomarkers or drug targets.  The genomic features of these two groups of genes expressed in opposing patterns may shed light on the mechanisms by which cells maintain basic and tissue-specific functions.

Here, we generate gene expression profiles of 42 normal human tissues on custom high-density microarrays to systematically identify 1,522 HKGs and 975 TEGs and compile a small subset of 20 housekeeping genes which are highly expressed in all tissues with lower variance than many commonly used HKGs.  Cross-species comparison shows that both the functions and expression patterns of HKGs are conserved. TEGs are enriched with respect to both segmental duplication and copy number variation, while no such enrichment is observed for HKGs, suggesting the high expression of HKGs are not due to high copy numbers. Analysis of genomic and epigenetic features of HKGs and TEGs reveals that the high expression of HKGs across different tissues is associated with decreased nucleosome occupancy at the transcription start site as indicated by enhanced DNase hypersensitivity.  Additionally, we systematically and quantitatively demonstrated that the CpG islands' enrichment in HKGs transcription start sites (TSS) and their depletion in TEGs TSS. Histone methylation patterns differ significantly between HKGs and TEGs, suggesting that methylation contributes to the differential expression patterns as well.We have compiled a set of high quality HKGs that should provide higher and more consistent expression when used as references in laboratory experiments than currently used HKGs. The comparison of genomic features between HKGs and TEGs shows that HKGs are more conserved than TEGs in terms of functions, expression pattern and polymorphisms.  In addition, our results identify chromatin structure and epigenetic features of HKGs and TEGs that are likely to play an important role in regulating their strikingly different expression patterns.

overall_design: We performed microarray experiment on more tissues and probesets in additional to the previous GEO submission (Series GSE11863). In brief, PolyA+ purified RNA pooled from multiple donors of a single human tissue type (e.g. cerebellum) were amplified with random primers and hybridized on a two-color ink-jet oligonucletodie microarray against a common reference pool, comprising ~20 normal adult tissue pools, on custom microarray patterns containing probes to monitor every exon and exon-exon junction in transcript databases, patent databases, and predicted from mouse transcripts. Data were analyzed for gene expression (the average of multiple probes), exon and junction expression, and splice form proportionality.


title: Non-radiated and Radiated lymphocytes from breast cancer patients treated with high-dose hyperfractioned radiotherapy

status: Public on Jun 05 2009

city: Las Palmas de Gran Canaria

first: Luis

last: Henriquez

state: Las Palmas

address: C/ Dr. Pasteur s/n

country: Spain

institute: University of Las Palmas de Gran Canaria

summary:
The aim of this study was to assess the relationship between gene expression profiles of ex vivo 2 Gy radiated lymphocytes and the development of acute and late toxicity due to hyperfractionated dose-escalation radiation therapy schedule in patientes with advanced breast cancer.  The gene expression profiles of ex vivo non-radiated lymphocytes were also examined.

Keywords: Toxicity grade analysis.

overall_design: Twelve patients were analyzed. No replicates of each patient was assesed. Human RNA universal control (Stratagene) was used as reference.

first: Amilcar

last: Flores

last: Henríquez


title: Cardiac muscle ring finger-1 increases susceptibility to heart failure by inhibiting creatine kinase activity in vivo

status: Public on Jun 03 2009

city: Chapel Hill

first: Monte

last: Willis

state: NC

address: 103 Mason Farm Road

department: Pathology & Laboratory Medicine

institute: UNC

summary:
Muscle ring finger-1 (MuRF1) is a muscle-specific protein implicated in the regulation of cardiac myocyte size and contractility. MuRF2, a closely related family member, redundantly interacts with protein substrates, and hetero-dimerizes with MuRF1. Mice lacking either MuRF1 or MuRF2 are phenotypically normal whereas mice lacking both proteins develop a spontaneous cardiac and skeletal muscle hypertrophy indicating cooperative control of muscle mass by MuRF1 and MuRF2. In order to identify the role that MuRF1 plays in regulating cardiac hypertrophy in vivo, we created transgenic mice expressing increased amounts of cardiac MuRF1. Adult MuRF1 transgenic (Tg+) hearts exhibited a non-progressive thinning of the left ventricular wall and a concomitant decrease in cardiac function. Experimental induction of cardiac hypertrophy by trans-aortic constriction (TAC) induced rapid failure of MuRF1 Tg+ hearts. Microarray analysis identified that the levels of genes associated with metabolism (and in particular mitochondrial processes) were significantly altered in MuRF1 Tg+ hearts, both at baseline and during the development of cardiac hypertrophy. Surprisingly, ATP levels in MuRF1 Tg+ mice did not differ from wild type mice despite the depressed contractility following TAC. To explain this discrepancy between the ongoing heart failure and maintained ATP levels in MuRF1 Tg+ hearts, we compared the level and activity of creatine kinase (CK) between wild type and MuRF1 Tg+ hearts. Although mCK and CK-M/B protein levels were unaffected in MuRF1 Tg+ hearts, total CK activity was significantly inhibited. We conclude that MuRF1’s inhibition of CK activity leads to increased susceptibility to heart failure following TAC, demonstrating for the first time that MuRF1 regulates cardiac energetics in vivo.

Keywords: Genetic modification, physiological manipulation.

overall_design: Three-condition experiment, MuRF1 Tg+ vs. WT mice. Biological replicates: 4 WT baseline, 3 MuRF1 Tg+ baseline, cardiac overpressure hypertrophy induced by trans-aortic banding at 1 week (3 WT, 3 MurF Tg) and 4 weeks (3 WT, 3 MurF Tg), hearts harvested. One replicate per array.

first: Jonathan

last: Schisler

first: Luge

last: Li

first: Jessica

last: Rodriguez

first: Eleanor

last: Hilliard

first: Peter

last: Charles

first: Cam

last: Patterson


title: Genes modulated by miR-205 in DU145 prostate cancer cells

city: Milan

first: Paolo

last: Gandellini

address: Via Amadeo 42

department: Department of Applied Research and Technological Development

country: Italy

institute: Fondazione IRCCS Istituto Nazionale Tumori

summary:
The study was aimed at identifying genes directly or indirectly regulated by miR-205 in the prostate. To this purpose, DU145 prostate cancer cells, which express miR-205 at very low levels, were transfected with miR-205 synthetic precursor and consequent alterations of gene expression analyzed using a microarray approach.

Keywords: comparison betweed cells exposed to different miRNA precursors

overall_design: DU145 cells were transfected with Pre-miR™ miRNA Precursor Molecule miR-205 (pre205) or negative control #1 (preNeg), Ambion. Four independent experiments were carried out and gene expression profiles were analyzed at day 3 after transfection.

first: Marco

last: Folini


title: Expression of survivin in lung eosinophils is associated with pathology in a mouse model of allergic asthma

first: Ashley

last: Connolly

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

overall_design: Refer to individual Series

first: Damon

last: Tumes

first: Lindsay

last: Dent


title: MicroRNA expression analysis in Uveal melanoma from formalin fixed paraffin embedded sections

status: Public on Jul 11 2009

city: Bangalore

first: Sugandan

last: Sivamani

address: 259, Apurva

department: Microarray Data Analysis

country: India

institute: Genotypic

summary: miRNA expression analysis was done on Uveal melanoma, metastatic and non - metastatic case in formalin fixed paraffin embedded sections, identified from case registry and confirmed by Chromosome insitu hybridization harboring monosomy 3 aberration in metastaic case and no aberration in non metastatic case. 19 miRNAs were found to be expressed only in class I tumors (choroidal melanoma) and not in class II (liver metastasis) and 11 miRNAs were found to be expressed only in class II and not in class I. The tumors were found to harbor oncomirs in both choroidal melanoma and metastasizing melanoma targeting tumor suppressor genes and metastatic suppressor genes. None of the differentially expressed miRNAs in either case were found to be located on the chromosomes which have been proved to carry chromosomal abnormalities. Rather it was found that genes targeted by the miRNAs were found to be present in chromosomal regions that are often found to be deleted 8p22, 13q and 17p.

overall_design:
Organism used: Homo sapiens.
* Slides: Human miRNA V2 8x15k Arrays AMADID: 19118.
* Starting material: 18 slides with formalin fixed Paraffin embedded sections-20 μm.
* RNA Samples used: 225-94-DUP, 225-94-E, 727-07-DUP, 727-07 -E.
* Labeling kit: Agilent miRNA labeling reagent and Hybridization Kit.
* Labeling Method: Ligation of one cyanine3-pCp molecule to the 3' end of RNA molecule with greater than 90% efficiency that generates fluorescent miRNA.
* Total RNA and cRNA Purification Kit: Biorad MicroBio Spin 6 columns.
* Hybridization Kit: Agilent miRNA Hybridization kit.

Hybridization protocol See Agilent Technologies website for miRNA microarray hybridization and wash protocol: http://www.chem.agilent.com/scripts/LiteraturePDF.asp?iWHID=50500
Scan protocol Laser detection of Cyanine 3 and Cyanine 5 fluorescence is performed using a confocal scanning instrument containing two tuned lasers, which excite Cyanine dyes at the appropriate wavelengths.
Description Microarrays were scanned on an Agilent scanner (G2505C) at 100% PMT with 10% for lower intensity (XDR Scanning).Data extraction was carried out with Agilent Feature Extraction software (version 9.1), and normalization was done using linear per array algorithm according to the manufacturer’s protocol
Data processing Feature extracted data was analyzed using GeneSpring GX v 10.0.2 software from Agilent. Normalization of the data was done in GeneSpring GX using the recommended Percentile shift normalization (50th percentile) . Further quality control of normalized data was done using correlation based condition tree to eliminate bad experiments. High expressed miRNA's were clustered using gene tree to identify significant gene expression patterns.

first: Abhirami

last: Krishnan

first: Subramaniam

last: Krishnakumar


title: Functional genomic analysis of frataxin deficiency

status: Public on Jun 02 2009

city: Los Angeles

first: Giovanni

last: Coppola

state: CA

address: 1524 Gonda, 695 Charles Young Drive South

department: Psychiatry and Neurology

institute: UCLA


title: Functional genomic analysis of frataxin deficiency, Illumina data

summary:
Functional genomic analysis of frataxin deficiency reveals tissue-specific alterations and identifies the PPARγ pathway as a therapeutic target in Friedreich's ataxia

Friedreich's ataxia (FRDA), the most common inherited ataxia, is characterized by focal neurodegeneration, diabetes mellitus, and life-threatening cardiomyopathy. Frataxin, which is significantly reduced in patients with this recessive disorder, is a mitochondrial iron-binding protein, but how its deficiency leads to neurodegeneration and metabolic derangements is not known. We performed microarray analysis of heart and skeletal muscle in a mouse model of frataxin deficiency, and found molecular evidence of increased lipogenesis in skeletal muscle, and alteration of fiber-type composition in heart, consistent with insulin resistance and cardiomyopathy, respectively. Since the peroxisome proliferator-activated receptor gamma (PPARγ) pathway is known to regulate both processes, we hypothesized that dysregulation of this pathway could play a key role in frataxin deficiency. We confirmed this by showing a coordinate dysregulation of the PPARγ coactivator Pgc1a and transcription factor Srebp1 in cellular and animal models of frataxin deficiency, and in cells from FRDA patients, who have marked insulin resistance. Finally, we show that genetic modulation of the PPARγ pathway affects frataxin levels in vitro, supporting PPARγ as a novel therapeutic target in FRDA.

overall_design: To compare frataxin deficient (KIKO) mice vs. WT, heart, skeletal muscle, and liver.

first: Daniele

last: Marmolino

first: Daning

last: Lu

first: Qing

last: Wang

first: Miriam

last: Cnop

first: Myriam

last: Rai

first: Fabio

last: Acquaviva

first: Sergio

last: Cocozza

first: Massimo

last: Pandolfo

first: Daniel

last: Geschwind


title: Chromosomal duplication in the grandifolia-D mutants of Arabidopsis thaliana, expression data

city: Gent

first: Nathalie

last: Gonzalez

address: Technologiepark 927

department: Plant System Biology

country: Belgium

institute: VIB

summary: The number of cells in an organ is a major factor for the determination of organ size. However, genetic basis of cell number determination is not well understood. Three grandifolia-D (gra-D) mutants of Arabidopsis thaliana developed huge leaves containing two- to three-fold increased number of cells of the wild type. Tiling array and microarray analysis of gra-D mutants suggested that genes found in a lower part of chromosome 4 were upregulated, suggesting the occurrence of segmental chromosomal duplications in the gra-D mutants. These region contain positive regulators of cell proliferation such as  AINTEGUMENTA (ANT) and cyclin genes such as CYCD3;1.

overall_design: Total RNAs were extracted using RNeasy plant mini kit (Qiagen) from vegetative part of seedlings at stage 1.03 grown in vitro. RNAs from three biological repeats of wild type (wt, control) and the 3 grandifolia mutants (gra1-D, gra2-D and gra3-D) were submitted to ATH1 array hybridization.

first: Gorou

last: Horiguchi

first: Gerrit

last: Beemster

first: Dirk

last: Inzé

first: Hirokazu

last: Tsukaya


title: CD6 co-stimulation of human peripheral blood mononuclear cells

status: Public on May 30 2009

first: madavan

last: vasudevan

state: Karnataka

address: Kasturi Nagar

institute: Theomics International Private Limited

summary: Difference in gene expression profile between 30000 freshly isolated human PBMCs stimulated with the immobilized anti-CD3 mAb (0.25µg/ml) alone (lane 1), along with ALCAM-Fc (1µg/ml) (lane 2), IL-2 (2.5ng/ml) (lane 4) or both (lane 3) or compared to unstimulated cells (lane 5), incubated for 72 h.

overall_design:
Organism used: Human
Slides: Human 8x15k Arrays
Starting material: Human peripheral blood mononuclear cells
RNA Samples used: Pooled 1 – Unstimulated, Anti-CD3, Anti-CD3 + IL-2, Anti-CD3 + ALCAM, Anti-CD3 +ALCAM + IL-2.
Labeling kit: Agilent’s Quick-Amp labeling Kit (p/n5190-0444)
Labeling Method: T7 promoter based-linear amplification to generate labeled complementary RNA   (One-Color Microarray-Based Gene Expression Analysis)
Total RNA and cRNA Purification Kit:
Hybridization Kit: Qiagen’s RNeasy minikit Cat#74104
RNA quality was checked using Bioanalyzer.

first: Pradip

last: Nair

first: Ramakrishnan

last: Melarkode

first: David

last: Rajkumar

first: Enrique

last: Montero


title: Early Response Genes in Osteoblasts Exposed to Dynamic Fluid Flow

status: Public on May 28 2009

city: San Antonio

first: stephen

middle: e

last: harris

state: TX

address: 7703 Floyd Curl Drive

department: Periodontics

institute: U. of Texas Health Sci Center at San Antonio

summary: Cyclic mechanical loads applied to the skeleton increase bone formation. Dynamic fluid flow is a potent anabolic stimulus for cultured osteoblasts. In this study, gene profiles involved in mediating the anabolic response of MC3T3 osteoblasts to dynamic fluid flow are investigated.

overall_design: MC3T3 osteoblast RNA was harvested 30-minutes and 1-hour post-stimulation respectively as experimental samples for comparison to the control group without dynamic fluid flow.

first: Giridhar

last: Shivaram

first: Chi

last: Kim

first: Nikhi

last: Batra

first: Wuchen

last: Yang

first: Stephen

last: Harris

first: Christopher

last: Jacobs


title: Gene expression profiling in SMA I and III patients

status: Public on Jun 08 2009

city: Padova

first: Gerolamo

last: Lanfranchi

address: Via U. Bassi, 58/B

department: CRIBI - Biotechnology Center and Biology Department

institute: University of Padova

summary:
Proximal spinal muscular atrophy (SMA) is a neurodegenerative disorder associated with mutations of the survival motor neuron gene SMN and characterized by progressive paralysis caused by degeneration of α–motor neurons of the spinal cord.
To better understand the effects of SMN gene defect in SMA skeletal muscle tissues we performed a transcriptional analysis of muscle specimens of patient affected by the two extreme forms of SMA: the most severe type I and the mild type III. Patients of both groups had a variable SMN2 copy number that did not correlate with phenotype severity. The two forms of SMA resulted with a distinct expression profile: in SMA III patients the reduced number of modified genes suggests a proximity to normal condition, whereas in SMA I patients there is highly deregulated picture of gene expression. Genes implicated in signal transduction were up regulated in SMA III condition whereas those typical of energy metabolism and muscle contraction were consistently down regulated in severe disease type. Microarray results were validated, and the expression pattern of gene networks involved in atrophy signaling were completed by RQ-PCR. The analysis shows that specific atrophy pathways are involved: IGF/PI3K/Akt and those regulating MNK1-2 (more precisely TNF-α/p38 MAPK and Ras/ERK pathways). The results suggest also a different picture of atrophy pathways in the two forms of SMA. In particular in SMA I muscles, p38 may be the protein synthesis regulator. Moreover SMA III profile appear to be the result of a more complex transcriptome given by the concurrent presence of atrophic and hyperthrophic fibers. This more favorable condition might be due to action of MTOR whose over expression in SMA III could lead to a compensative hypertrophy.
Keywords: disease state analysis

overall_design:
Total RNA purified from muscle samples of SMA I or III single patient were compared with total RNA from age-matched muscle control.
These samples were labeled and hybridized to muscle specific microarrays produced by our group (GPL2011). Two replicates of each experiment were performed using different microarray slides, in which sample and reference RNAs, labeled either with Cy3 or Cy5 fluorochromes, were crossed in both combinations (dye-swapping procedure).

first: Caterina

last: Millino

first: Marina

last: Fanin

first: Andrea

last: Vettori

first: Maria-Luisa

last: Mostacciuolo

first: Corrado

last: Angelini


title: Expression profiling of the forming atrioventricular node using a novel Tbx3-based node-specific transgenic reporter

city: Leiden

first: Henk

last: Buermans

state: ZH

address: Albinusdreef 2

department: Human Genetics

institute: Leiden University Medical Center

summary:
The atrioventricular (AV) node is a recurrent source of potentially life-threatening arrhythmias. Nevertheless, limited data are available on its developmental control or molecular phenotype. We used a novel AV node-specific reporter mouse to gain insight into the gene programs determining the formation and phenotype of the AV node. In the transgenic reporter, green fluorescent protein (GFP) expression was driven by 160 kbp of Tbx3 and flanking sequences. GFP was selectively expressed in the AV canal of embryos, and in the AV node of adults, while all other Tbx3+ conduction system components, including the AV bundle, were devoid of GFP expression. Fluorescent AV nodal (Tbx3BAC-Egfp) and complementary working (NppaBAC336-Egfp) myocardial cell populations of E10.5 embryos and E17.5 fetuses were purified using fluorescence-activated cell sorting, and their expression profiles were assessed by microarray analysis. We constructed a comprehensive list of sodium, calcium, and potassium channels specific for the nodal or working myocard. Furthermore, the data revealed that the AV node and the working myocardium phenotypes diverge during development, but that the functional gene classes characteristic for both compartments are maintained. Interestingly, the AV node-specific gene repertoire consisted of multiple neurotrophic factors not yet appreciated to play a role in nodal development.  These data present the first genome-wide transcription profiles of the AV node during development, providing valuable information concerning its molecular identity.

Keywords: Tbx3, AV node, working myocardium, embryonic development, cardiac development, cardiac conduction system

overall_design:
24 samples:
6x working myocardium stage E10.5 (NppaBAC336-Egfp mice),
6x AV canal myocardium stage E10.5 (Tbx3BAC-Egfp mice),
6x working myocardium stage E17.5 (NppaBAC336-Egfp mice),
6x AV node myocardium stage E17.5 (Tbx3BAC-Egfp mice)

first: Thomas

last: Horsthuis

first: Janynke

middle: F

last: Brons

first: Arie

middle: O

last: Verkerk

first: Vincent

last: Wakker

first: Danielle

last: Clout

first: Martijn

last: Bakker

first: Antoon

last: Moorman

first: Peter

last: ’t Hoen

middle: M

last: Christoffels


title: Replacing skeletal muscle alpha-actin with cardiac actin in mouse skeletal muscle

status: Public on May 25 2009

city: Nedlands

first: Kristen

middle: Jean

last: Nowak

address: WAIMR, B Block, QEII Medical Centre

department: Centre for Medical Research

institute: University of Western Australia/WA Institute for Medical Research

summary:
Skeletal muscle actin mice (Crawford et al., (2002) Mol Cell Biol 22, 5587) were crossed with cardiac actin transgenic mice (termed "ACTC^Coco" or "Coco" for short), to produce mice that had cardiac actin instead of skeletal muscle actin in their skeletal muscles (termed "ACTC^Co/KO" or for short "Coco/KO").  Microarray analysis using the Illumina mouse-6 v1.1 expression beadchip was performed on RNA extraced from the soleus muscle of Coco/KO mice and wildtype mice, to confirm the swith in actin isoform expression, and to determine what other differences might exist between wildtype mice and the Coco/KO mice.

Keywords: genetic modification

overall_design: 3 RNA samples (each being the pool of two individual samples extracted from different soleus muscles from different individual mice) per genotype (either wildtype or Coco/KO) were used.  The total 6 RNA samples were processed using an Illumina mouse-6 v1.1expression beadchip and then the differentially expressed genes determined.

first: Nigel

middle: G

last: Laing

first: Gianina

last: Ravenscroft


title: The protein disulfide isomerase AGR2 is essential for the production of intestinal mucus

status: Public on May 15 2009

city: San Francisco

last: Barczak

address: 1550 4th St Rock Hall 545

institute: UCSF Sandler Center Functional Genomics Core Facility

summary:
Protein disulfide isomerases (PDIs) aid protein folding and assembly by catalyzing formation and shuffling of cysteine disulfide bonds in the endoplasmic reticulum (ER).
Many members of the PDI family are expressed in mammals but the roles of specific PDIs in vivo are poorly understood.
A recent homology-based search for additional PDI family members identified anterior gradient homolog 2 (AGR2), a protein originally presumed to be secreted by intestinal epithelial cells, but the function of AGR2 has been obscure. Here we show that AGR2 is expressed in the ER of secretory cells and is essential for in vivo production of intestinal mucin, a large cysteine-rich glycoprotein that forms the protective mucus gel lining the intestine.
A cysteine residue within the AGR2 thioredoxin-like domain forms mixed disulfide bonds with MUC2, consistent with a direct role for AGR2 in mucin processing.
Despite a complete absence of intestinal mucin, mice lacking AGR2 appeared healthy but were highly susceptible to dextran sodium sulfate-induced experimental colitis, indicating a critical role for AGR2 in protection from environmental insults.
We conclude that AGR2 is a unique member of the PDI family that has a specialized and non-redundant role in intestinal mucus production.

Keywords: small intestine and colon gene expression profiles for Agr2-/- and littermate control mice

overall_design: DNA miocroarrays were used to analyze small intenstine and colon mRNA expression of AGR2 KO and littermate control mice. The experiment incorporated a 1 color design and used Agilent arrays that contained roughly 44,00 60mer probes that provide complete coverage of the mouse genome. 12 arrays were hybridized and represent 8 small intestine samples ( 4 each KO and WT) and 4 colon samples (2 each KO and WT)

first: David

last: Erle

first: Sung-Woo

last: Park

first: Guohua

last: Zhen


title: Profiling of filarial induced genes in lymphatic endothelial cells

city: Bethesda

first: Sasisekhar

last: Bennuru

address: 10 Center Dr, Bldg 10, Rm 11N208

department: LPD

institute: NIH/NIAID

summary: To observe the global changes in the lymphatic endothelial cells upon exposure to filarial antigens or parasites, LECs were stimulated for 24, 48, and 72hrs and the expression profiles were carried out.

overall_design: Human filarial parasites Brugia malayi and Wuchereria bancrofti habitat the lymphatics and cause lymphatic dilatation and lymphedema. In order to evaluate the effect of various stage specific effects on the lymphatic endothelial cells (LEC) and understand how they modulate the lymphatic dysfunction, LECs were stimulated in antigens derived from the Brugia malayi. These are preliminary time course data towards understanding how the filarial antigens induce lymphangiogenesis.

middle: B

last: Nutman


title: Expression profiling in lymphoblastoid cell lines of family with maternally inherited non-syndromic hearing loss.

status: Public on Jun 01 2009

first: Yelena

last: Bykhovskaya

address: 8700 Beverly Blvd

institute: Cedars-Sinai Medical Center

summary:
We analyzed samples from fourteen deaf individuals (Affected 1 through 14), fifteen hearing maternally related family members (Unaffected 1-15), six marry-in controls (Controls 1-6) from extended pedigree from Arab-Israeli village, and nine individuals from another Arab-Israeli village (Controls 7-15). All affected and unaffected maternally-related individuals carry homoplasmic mutation in the 12S rRNA gene of the mitochondrial DNA, associated with both non-syndromic and aminoglycosides-induced deafness.
Keywords: Comparison of genome-wide expression in cell lines of maternally-related individuals with mitochondrial mutation and controls carrying wild-type mitochondrial chromosome.

overall_design: Lymphoblastoid cell lines established from peripheral blood lymphocytes of study participants and immortalized with Epstein-Barr virus, were grown in suspension in T flasks in RPMI-1640 medium (Invitrogen, Inc., Carlsbad, CA), containing 2 mmol/L L-glutamine, 100 ug/ml streptomycin, and 10% fetal calf serum. Total RNA was extracted using Trizol reagent (Invitrogen, Inc.) according to the manufacturer?s protocol. cRNA amplification and labeling with biotin were performed using Illlumina® TotalPrep RNA amplification kit, manufactured by Ambion, Inc (Austin, TX) according to the manufacturer?s protocol using 100 ng of total RNA as input material. 1ug of labeled cRNAs were hybridized to the eight ?whole genome? Sentrix Human-6 v2 Expression BeadChips (Illumina, San Diego, CA). RNA sample from patient Unaffected 2 was duplicated on the same chip; RNA sample from Affected 3 was analyzed in triplicate: two on the same chip, and one on a different chip. All hybridization and scanning steps were performed according to the manufacturers? instructions using reagents and equipment purchased from Illumina, Inc

first: Y

first: E

last: Mengesha

first: N

last: Fischel-Ghodsian


title: Response of mouse liver to Adenovirus, Helper Dependent Adenovirus, and Adeno-Associated Virus

status: Public on May 13 2009

city: Richmond

first: Paul

last: Fawcett

state: VA

address: 1101 E. Marshall St.

institute: Virginia Commonwealth University

summary: Understanding host responses to viral gene therapy vectors is necessary for the development of safe and efficacious in vivo gene transfer agents. We describe the use of high-density spotted complementary DNA microarrays in monitoring the in vivo host transcriptional responses in mouse liver upon administration of either a "first-generation"adenoviral (Ad) vector, a helper-dependent "gutless" adenoviral (HD) vector, or an adeno-associated viral (AAV) vector containing human factor IX (hFIX) expression cassettes. Since HD and AAV do not contain any viral genes, they allow us to assess the host response to the viral capsid and packaged nonviral DNA in whole animals. Comparison of the host response to Ad and HD helps assess the importance of leaky adenoviral gene expression. While all three vectors induced characteristic temporally sequenced programs of gene expression, the gene expression programs induced by the Ad and HD adenovirus vectors were remarkably similar, including the induction of a prominent type I interferon (IFN)-dependent cluster within 6 hours of administration. In contrast, the AAV-based vector caused far fewer alterations of host-gene expression. Our results indicate that recognition of the Ad capsid or double-stranded DNA (of nonviral origin) in the vector elicits a robust type I IFN response that is, however, not elicited by AAV-derived vector transduction.

overall_design: An all pairs experiment design type is where all labeled extracts are compared to every other labeled extract.


title: A comparison of Illumina bead chips reveals unexpected challenges in the use of newest generation microarrays

status: Public on May 12 2009

city: Bonn

first: Joachim

last: Schultze

state: NRW

address: Carl-Troll-Strasse 31

department: Genomics and Immunoregulation

country: Germany

institute: LIMES (Life and Medical Sciences Center Genomics and Immunoregulation)

summary:
The development of a comparison approach for Illumina bead chips unravels unexpected challenges applying newest generation microarrays
The MAQC project demonstrated that microarrays with comparable content show inter- and intra-platform reproducibility. However, since the content of gene databases still increases, the development of new generations of microarrays covering new content is mandatory. To better understand the potential challenges updated microarray content might pose on clinical and biological projects we developed a methodology consisting of in silico analyses combined with performance analysis using real biological samples.   Here we clearly demonstrate that not only oligonucleotide design but also database content and annotation strongly influence comparability and performance of subsequent generations of microarrays. Additionally, using human blood samples and purified T lymphocyte subsets as two independent examples, we show that a performance analysis using biological samples is crucial for the assessment of consistency and differences.   This study provides an important resource assisting investigators in comparing microarrays of updated content especially when working in a clinical or regulatory setting.

overall_design: Two datasets were analyzed in the study. The Treg dataset includes 6 samples, three Treg and three nonTreg samples.   The whole blood dataset includes 11 samples of patients with scleroderma and 7 samples of patients with bacteremia.   Both datasets were hybridized both on Illumina v1 and v2 BeadChips.

first: Daniela

last: Eggle

first: Svenja

last: Debey-Pascher

first: Marc

last: Beyer


title: Cystic leukoencephalopathy without megalencephaly

city: Goettingen

first: Gabriela

last: Salinas

state: Lower-Saxony

address: Kreuzbergring 57

department: Department of Pathology

institute: Universitaetsmedizin Goettingen

summary: The cystic leukoencephalopathy without megalencephaly has been defined as distinct autosomal-recessive syndrome of quasi-static encephalopathy with normo- or microcephaly, impaired psychomotor development and characteristic pattern on brain MRI. We identified mutations in the gene encoding the RNASET2 glycoprotein in seven affected individuals as the cause of disease. The results suggest that RNASET2 plays an important role in central nervous system myelination and development.

overall_design: 2-Color dye swap design independent for two different families including technical and biologial replicates for affected family members (disease) and not affected family members (controls).

first: Marco

last: Henneke

first: Simone

last: Diekmann

first: Andreas

last: Ohlenbusch

first: Jens

last: Kaiser

first: Volkher

last: Engelbrecht

first: Alfried

last: Kohlschuetter

first: Ralph

last: Kraetzner

first: Marcos

last: Madruga-Garrido

first: Michele

last: Mayer

first: Lennart

last: Opitz

first: Diana

last: Rodriguez

first: Franz

last: Rueschendorf

first: Johannes

last: Schumacher

first: Holger

last: Thiele

first: Sven

last: Thoms

first: Robert

last: Steinfeld

first: Peter

last: Nuernberg

first: Jutta

last: Gaertner


title: The RNA-binding Protein HuR Controls Thymocyte Development, Selection and Migration

status: Public on May 20 2009

city: Durham

first: Neelanjan

last: Mukherjee

address: 101 Science Drive

department: Institute for Genome Sciences & Policy

institute: Duke University

summary: The progressive development of T-cells in the thymus is a highly regulated process that aims to arm the body with T-cells against pathogens whilst maintaining tolerance to self. Although the role of gene transcription in this process has been extensively studied, little is known about the significance of associated post-transcriptional mechanisms. Post-transcriptional control is imposed by the dynamic interactions of RNA-binding proteins (RBPs) with RNA molecules which destine mRNA molecules towards translation or destruction. In this study we focus on the functions of an RBP called HuR. We made use of novel transgenic systems in which HuR is deleted in thymic T-cells to test whether HuR is involved in their developmental maturation. We show that HuR controls the generation and provision of T-cells competent for the proper recognition of antigens in the body and the elimination of harmfull cells. We further show that HuR controls T-cell development through its involvement in basic cellular processes like cell cycle regulation, cellular activation, death and migration which were associated to HuR’s efficacy in regulating the expression or function of key signaling components. Our studies provide important insight on the post-transcriptional regulation of fundamental cellular processes and identify HuR as an important modulator of adaptive immunity and autoimmunity.

overall_design:
Total thymocytes were isolated from female 6-8 week old LckCre+ Elavl1+, LckCre+Elavl1fl/fl
and Elavlfl/fl thymi following desegregation and pooled from n=6 littermate mice/group,
were cultured in complete RPMI-1640 medium the presence or absence of PMA (10 ng/ml) and
ionomycin (500ng/ml). Cells were collected at 0, 4 and 12 hrs post stimulation and washed
in ice cold PBS via centrifugation, prior to lysis with Trizol (Invitrogen). Total RNA was
then treated with DNase I to minimize contamination from genomic DNA. Further purification
was carried out using the RNeasy columns (Qiagen). The experiment was performed two more times
to complete a set of 3 biological replicates. In all cases, Ex vivo RNA samples were
labeled with Cy3 and reference samples were labeled with Cy5. NaOH was used to
destroy residual RNA.

first: Olympia

last: Papadaki

first: Vicky

last: Katsanou

first: Stavros

last: Milatos

first: Patrick

last: Lager

first: Jack

middle: D

last: Keene

first: Dimitris

last: Kontoyiannis


title: Affinity purification of ribosomes and associated RNAs from stress-treated cells

city: ETH Hoenggerberg, Zuerich

first: Andre

last: Gerber

address: Wolfgang Pauli Str. 10, HCI H396

department: Institute of Pharmaceutical Science

institute: Swiss Federal Institute of Technology

type:
Expression profiling by array
Other

summary:
In this study, we systematically identified ribosome associated RNAs. To identify ribosome associated RNAs, C-terminal ZZ-tagged Rpl16a, expressed under control of its native promoter, was affinity purified from whole cell extracts of cultures grown to mid-log phase. Extracts were incubated with immunoglobulin G (IgG) coupled microbeads, washed, and ribosomes were eluted by tobacco etch virus (TEV) protease treatment. To analyze how changes in steady-state mRNA levels (= transcriptome) are related to changes of respective messages in the translatome, total RNA and ribosome associated RNA from stress-treated and untreated cells were analyzed with yeast cDNA microarrays. To achieve this, fluorescently labeled (Cy5) cDNA was prepared from total RNA isolated from yeast cell extracts and from affinity-purified ribosomes, and each sample was mixed with differentially labeled (Cy3) cDNA prepared from a common reference RNA pool, and competitively hybridized on yeast cDNA microarrays. We performed five independent experiments with untreated cells grown in minimal medium (t=0 reference) and three or more independent biological replicates of treated cells. Altogether, we sampled the cell's response to five different "stress treatments" that were applied for relatively short periods of time (10 or 20 minutes) to avoid secondary effects triggered by transcriptional adaption.
A stimulus or stress experiment design type is where that tests response of an organism(s) to stress/stimulus. e.g. osmotic stress, behavioral treatment
Time: Duration of the treatment
Growth Condition: Type of treatment

first: Regula

last: Halbeisen


title: Affinity purification of ribosomes and associated RNAs from stress-treated cells using tagged Rpl16a and Rpl16b

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


title: The expression profiles of chicken ileum

status: Public on May 01 2009

city: Beijing

first: Yeshi

last: Yin

address: NO.1 West Beichen Road, Chaoyang District

country: China

institute: Institute of Microbiology

summary: We used two different chermostat products and sterilized water to inoculate chicks, and then detect the expression profiles affected by the different inoculation.

overall_design: Three probes for each of the 55366 genes(4126 Refseq genes and 51240 predicted genes) and 3259 random probes were designed and every slide have two blocks. 9 samples were used to analysis and 6 samples had two replications.

first: Baoli

last: Zhu

first: Xin

last: Wang


title: Duke-UNC-Texas-EBI ENCODE expression project

status: Public on Apr 23 2009

city: Rensselaer

first: Sridar

last: Chittur

address: One Discovery Drive, CRC 342G

department: Center for Functional Genomics

institute: SUNY-University at Albany

summary:
These samples are being analyzed by the Duke-UNC-Texas-EBI ENCODE consortium. Expression from these cell types will compared to three whole genome open chromatin methodologies: DNaseI hypersensitivity (DNase-seq), Formaldehyde-Assisted Isolation of Regulatory elements (FAIRE-seq), and Chromatin Immunoprecipitation (ChIP-seq) .

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

overall_design: These were processed on Affymetrix Exon 1.0 ST arrays to obtain expression data from all cell types being analyzed for open chromatin.

first: G

last: Crawford

first: V

last: Iyer

first: J

last: Lieb

first: E

last: Birney

first: T

last: Furey


title: Expression profiling of HCC patients with different recurrent-free survival time

status: Public on Apr 24 2009

first: Hongsong

last: Chen

address: 11 Xizhimen S St

department: People's Hospital

institute: Peking university

summary:
The prognosis of hepatocellular carcinoma (HCC) after surgery is poor due to its high recurrence rate. Oligonucleotide microarrays were used to analyze the gene expression of HCC patients with differing recurrent-free survival times following resection to see if gene expression can predict recurrence of HCC.
Keywords: Disease states analysis

overall_design: 32 HCC patients with a recurrent-free survival times that varied between 3 months to >3 years were used for microarray analysis.RNA from a mixed pool of adjacent non-tumor tissue derived from 9 patients was used as a reference pool for all microarray hybridizations.

first: Xingwang

last: Xie

first: Minghui

last: Mei

first: Ran

last: Fei

first: Liling

last: Qin

first: Henghui

last: Zhang

first: Weijia

last: Liao

first: Lai

last: Wei


title: Analysis of human myeloma cell lines expressing beta 3 integrin

status: Public on Apr 29 2009

city: Naples

first: gabriella

last: rodolico

address: Via Costantinopoli 16

institute: Second University of Naples

summary:
The aim of these experiments was designed to compare gene expression in human myeloma cell lines expressing beta 3 integrin vs counterpart cell lines that do not express beta 3 integrin.

Keywords: Gene expression,cell lines, siRNA

overall_design: Phenotype characterization of both primary and cultured myeloma plasma cells was assessed by flow cytometry (FACScanto, Becton Dickinson, San Jose, CA) using a panel of MoAbs or antisera to the following markers: CD138, CD38, CD56, k/l chains, CD20, CD44, CD54, alphav and beta3 chains. The microarray analysis was performed on: myeloma bone resorbing cell lines alphav-beta3 positive and the same cell line silenced for the integrins; myeloma non bone resorbing cell lines negative for the expression of avb3.

first: Gabriella

last: Rodolico

first: Raffaele

last: De Palma

first: Marco

last: Tucci

first: Francesco

last: Silvestris


title: The IRE1α-XBP1 Pathway of the Unfolded Protein Response is Required for Adipogenesis

status: Public on Apr 25 2009

city: Ithaca

first: Ling

last: Qi

address: 307 Biotech Bldg, Cornell Univ

department: Division of Nutritional Sciences

institute: Cornell University

summary: Signaling cascades during adipogenesis culminate in the expression of two essential adipogenic factors, PPARγ and C/EBPα.  Here we demonstrate that the IRE1α-XBP1 pathway, the most conserved branch of the unfolded protein response (UPR), is indispensable for adipogenesis.  Indeed, XBP1-deficient mouse embryonic fibroblasts and 3T3-L1 cells with XBP1 or IRE1α knockdown exhibit profound defects in adipogenesis.  Intriguingly, C/EBPβ, a key early adipogenic factor, induces Xbp1 expression by directly binding to its proximal promoter region.  Subsequently, XBP1 binds to the promoter of Cebpa and activates its gene expression.  The posttranscriptional splicing of Xbp1 mRNA by IRE1α is required as only the spliced form of XBP1 (XBP1s) rescues the adipogenic defect exhibited by XBP1-deficient cells.  Taken together, our data show that the IRE1α-XBP1 pathway plays a key role in adipocyte differentiation by acting as a critical regulator of the morphological and functional transformations during adipogenesis.

overall_design: The Xbp1 deficient MEFs are subjected to adipogenesis using standard drug induction.  The cells are collected at day 0 and day 3. Total RNA are extracted from the cells and used for one channel microarray analysis. Expression levels on day 3 are compared to that on day 0.

first: Haibo

last: Sha

first: Yin

last: He

first: Hui

first: Cindy

last: Wang

first: Anna

last: Zenno

first: Shi

last: Hang

first: Xiaoyong

last: Yang

first: Xinmin


title: Host lung gene expression patterns predict the etiology of infectious pneumonias

city: Houston

first: Scott

last: Evans

address: 1400 Holcombe Blvd

department: Pulmonary Medicine - Unit 1462

institute: University of Texas-M.D. Anderson Cancer Center

summary:
Rationale: Lower respiratory tract infections continue to exact unacceptable worldwide mortality, often because the infecting pathogen cannot be identified.  The respiratory epithelia provide protection from pneumonias through organism-specific generation of antimicrobial products, offering potential insight into the identity of infecting pathogens.  Objective: This study assesses the capacity of the host gene expression response to infection to predict lower respiratory pathogens without reliance on culture data.  Methods: Mice were inhalationally challenged with S. pneumoniae, P. aeruginosa, A. fumigatus or PBS prior to whole genome gene expression microarray analysis of their pulmonary parenchyma.  Characteristic gene expression patterns for each condition were identified, allowing the derivation of prediction rules for each pathogen.  After confirming the predictive capacity of gene expression data in blinded challenges, a computerized algorithm was devised to predict the infectious conditions of subsequent subjects.  Measurements and Main Results: We observed robust, pathogen-specific gene expression patterns as early as 2 h after infection.  We were able to develop a predictive model that used a limited number of specifically regulated transcripts to discriminate perfectly among infecting pathogens in the training data.  Validation trials using an algorithmic decision tree revealed 94.4% diagnostic accuracy when discerning the presence of bacterial infection.  The model subsequently differentiated between bacterial pathogens with 71.4% accuracy and between non-bacterial conditions with 70.0% accuracy, both far exceeding the expected diagnostic yield of standard culture-based bronchoscopy with bronchoalveolar lavage.  Conclusions: These data substantiate the specificity of the pulmonary innate immune response and support the feasibility of a gene expression-based clinical tool for pneumonia diagnosis.

Keywords: pathogen- and time-dependent host lung gene expression changes in pneumonia

overall_design: Mouse lungs were removed 6 h after challenge with P. aeruginosa, S. pneumoniae, A. fumigatus or sham (PBS), RNA was collected from leukoreduced lung homogenates, then cRNA was amplified and hybridized to Illumina Sentrix mouse-6 gene expression arrays v1.1.  Gene expression data was analyzed to determine if there were distinct patterns of host trasnscription that corresponded with a specific infection.


title: Reciprocal regulation of genes expression by Ephedra herba in mouse brain

status: Public on Apr 04 2009

city: Daejon

first: Bu-Yeo

last: Kim

address: 1672 Yuseongdae-ro, Yuseong-gu

department: Herbal Medicine Research Division

country: South Korea

institute: Korea Institute of Oriental Medicine

summary: A microarray analysis was conducted using the brain tissue of mouse orally administrated aqueous extracts of Ephedra herba, in which the expression level of many genes were altered depending on the duration of treatment time. In particular, temporal changes in the expressions of genes can be classified into two major reciprocal patterns.

overall_design: Aqueous extracts of Ephedra herba were administered orally at dosage of 50 or 1,250 mg/kg once daily for 1, 3 and 10 consecutive days on mice. Expression level of genes from brain tissue was compared to that from untreated control mice.

first: Li

middle: H

last: Cao

first: Hwayong

last: Park

first: Sangkyun

last: Jeong


title: Human embryonic stem cells, differerentiated EBs and adult cells

status: Public on Apr 17 2009

city: Doha

middle: Maria

last: Marincola

address: Al Nasr Tower, AL Corniche Street, PO Box 26999

country: Qatar

institute: Sidra Medical and Research Center

type:
Expression profiling by array
Non-coding RNA profiling by array

platform_organism:
Homo sapiens
Mus musculus
Rattus norvegicus
Human gammaherpesvirus 8
Macacine gammaherpesvirus 4
Betapolyomavirus macacae

overall_design: Refer to individual Series


title: Immunomodulatory function of bone marrow-derived mesenchymal stem cells in experimental autoimmune type 1 diabetes

status: Public on Apr 02 2009

city: Boston

first: Andrea

last: Vergani

state: MA

address: 300 Longwood Ave

institute: Children's Hospital Harvard Medical School

summary: Human clinical trials in type 1 diabetes (T1D) patients are underway using mesenchymal stem cells (MSC) without prior validation in a mouse model for the disease.  In response to this void, we characterized bone marrow-derived murine MSC for their ability to modulate immune responses in the context of T1D, as represented in non-obese diabetic (NOD) mice. In comparison to NOD-, BALB/c-MSC express higher levels of the negative costimulatory molecule PD-L1 and promote a shift toward Th2-like responses in treated NOD mice. In addition, transfer of MSC from resistant strains (i.e. NOR or BALB/c), but not from NOD mice, conferred disease protection when administered to prediabetic NOD mice. The number of BALB/c-MSC trafficking to the pancreatic lymph nodes of NOD mice was higher than in NOD mice provided autologous NOD-MSC. Administration of BALB/c-MSC resulted in reversal of hyperglycemia in 90% of NOD mice (p=0.002). Transfer of autologous NOD-MSC imparted no such therapeutic benefit, and in fact soft tissue and visceral tumors were uniquely observed in this setting (i.e. no tumors were present with BALB/c- or NOR-MSC transfer). These data provide important preclinical data supporting the basis for further development of allogeneic MSC-based therapies for T1D and potentially, other autoimmune disorders.

overall_design:
To generate MSC, BM mononuclear cells were isolated from the femurs and tibiae of at least 5 mice in order to minimize cell variability. Cells are seeded in flasks at a concentration of 10x106/25 cm2 in M10 medium (DMEM medium [Cambrex, East Rutherford, New Jersey] containing 10% fetal calf serum [HyClone, Logan, Utah], 1% penicillin-streptomycin, and 1% L-glutamine [both from Cambrex]). To examine MSC in an inflammatory setting, 7.5x105 NOD- or BALB/c-MSC/well were cultured for 48h in 6-well plates in M10 medium containing 10 ng/ml recombinant murine IL-1beta (Peprotech, Rocky Hill, NJ).
To ensure that our cultured cells had multipotent potential, we tested MSC P4 cultures for their ability to undergo differentiation into chondrocytes, osteocytes, and adipocytes as previously published.10 Chondrogenic differentiation was induced by 50 ug/ml ascorbic acid and 1 ng/ml TGF?-1 (Peprotech), osteogenic differentiation was induced by 50 ug/ml ascorbic acid, 10 mM sodium alpha-glycerophosphate, and 10-8 M dexamethasone, and adipogenic differentiation was induced by 10-7 M dexamethasone and 6 ng/ml insulin.
MSC were analyzed for expression at passage 4 (P4).
We examined the gene expression profile of NOD- (samples #1 to 4) and BALB/c-MSC (samples #5 to 8), obtained from normoglycemic NOD mice and age-matched BALB/c mice. A striking concordance in the pattern of gene regulation across the entire screen was observed when BALB/c-MSC genes were compared to the NOD-MSC 4 different culture of NOD derived MSC were compared to 4 differente cell culture of Balb/c derived MSC, Balb/c cell coltures were considered reference samples.

first: Paolo

last: Fiorina

first: Reza

last: Abdi


title: Transcriptional dosage compensation in heterozygotes for deletions in Drosophila

status: Public on Apr 03 2009

city: Johnson City

first: Lev

last: Yampolsky

state: TN

address: ETSU

department: Biological Sciences

institute: East Tennessee State University

summary:
We measured gene expression in two isogenic Drosophila lines heterozygous for long deletions. We find that a majority of genes are at least partially compensated at transcription for ½-fold dosage. The degree of compensation does not vary among functional classes of genes. Compensation for deletions is stronger for highly expressed genes than for genes with low expression level. In contrast, the degree of compensation for duplications observed in Gupta et al, 2006, (J. of Biology 5, 3) data for heterozygotes for a duplication is stronger for weakly expressed genes. Thus, transcriptional compensation appears to be based on general regulatory mechanisms that insure high levels of transcription some genes and low transcription levels of other genes, instead of precise maintenance of a particular homeostatic expression level. Given the ubiquity of transcriptional compensation, dominance of wild-type alleles may be at least partially caused by of the regulation at transcription level.

Keywords: Genome-wise dosage compensation study

overall_design: Two DrosDel [34] Drosophila melanogaster isogenic lines, Df(3L)ED4475 and Df(3L)ED4543, heterozygous for long deletions on 3L chromosomal branch both maintained against the TM6C balancer were used for microarray experiment. Twenty five adult flies 2-5 days after eclosion were frozen in liquid nitrogen and used for RNA extraction by Trizol method (Invitrogen ®, Carlsbad, CA) in twelve replicates from each line. The two deletion lines served as controls to each other.

first: L

first: A

last: McAnally


title: Gene expression profiling among human embryonic stem cells, differerentiated EBs and adult cells

first: Francesco

summary:
Human embryonic stem (hES) cells have unique features: self-renewal ability and pluripotency. They can be continuously cultured in undifferentiated state and give rise to cells and tissues of all three germ layers. Thus hES cells provide a resource not only for cell replacement therapy but also for studying human developmental biology. However, much work needs to be done to understand the molecular mechanisms responsible for the maintenance of the undifferentiatiation state and the differentiation process of human embryonic stem cells.

Keywords: cell type comparison design

overall_design: We performed gene expression profiling on 3 separate passages from 3 different hES cell lines (WA09 (H9), TE06 (I6) and BG01v), EBs samples derived from WA09 cells at 3 different time points, and 5 types of adult cells (HUVEC, HMVEC, UASMC NHA, and LFB).


title: Transcription profile of K-ras activated murine menstruating uteri

city: Cambridge

first: Ching-wen

last: Cheng

address: Robinson Way

department: Obstetrics and Gynaecology

country: United Kingdom

institute: University of Cambridge

summary: The expression of K-ras transgene is induced in  K-rasV12+/-/Cre+/+/ROSA26R-LacZ+/+ transgenic mice uteri, which were primed with hormone treatment. Decidualization was artificially induced in these uteri and hormone injection was withdrawn to induced menstruation-like endometrial degeneration. Microarray was then used to study the molecular changes in these menstruating uteri following the K-ras activation.

overall_design: Uterine tissues were collected from K-ras induced and littermate controled menstruating uteri. Experiments were carried out using Illumina Mouse WG-6 BeadChips (n=6 in each group). Data were normalized in R environment using the Lumi Bioconductor package.

first: Diana

last: Licence

first: Feijun

last: Luo

first: Mark

last: Arends

first: Cristin

middle: C

last: Print

first: D

last: Charncock-Jones


title: Functional characterization of E- and P-cadherin in invasive breast cancer cells

status: Public on Apr 18 2009

city: Bron

first: Hector

last: Hernandez-Vargas

state: Rhone

address: 36 bis Rue de la Batterie

institute: Genomics Consulting

summary:
Alterations in the cadherin-catenin adhesion complexes are involved in tumor initiation, progression and metastasis. However, the functional implication of distinct cadherin types in breast cancer biology is still poorly understood. Methods: To compare the functional role of E-cadherin and P-cadherin in invasive breast cancer, we stably transfected these molecules into the MDA-MB-231 cell line, and investigated their effects on motility, invasion and gene expression regulation.
Expression of either E- and P-cadherin significantly increased cell aggregation and induced a switch from fibroblastic to epithelial morphology. Although expression of these cadherins did not completely reverse the mesenchymal phenotype of MDA-MB-231 cells, both E- and P-cadherin decreased fibroblast-like migration and invasion through extracellular matrix in a similar way. Moreover, microarray gene expression analysis of MDA-MB-231 cells after expression of E- and P-cadherins revealed that these molecules can activate signaling pathways leading to significant changes in gene expression. Although the expression patterns induced by E- and P-cadherin showed more similarities than differences, 40 genes were differentially modified by the expression of either cadherin type.

overall_design: Microarray gene expression analysis of MDA-MB-231 cells after expression of E- and P-cadherins using different clones for each conditions to reveal that these molecules can activate signaling pathways leading to significant changes in gene expression

first: David

last: Sarrio

first: Jose

last: Palacios

first: Marta

last: Hergueta-Redondo

first: Amaparo

last: Cano

first: Gema

last: Moreno-Bueno


title: Novel Lipogenic Enzyme ELOVL7 Involves in Prostate Cancer Growth through Saturated Fatty-acid and Steroid Metabolism

status: Public on Apr 01 2009

city: Tokyo

first: Hidewaki

last: Nakaagwa

state: Tokyo

address: 4-6-1 Shirokanedai, Minato-ku,

department: Institute of Medical Science

institute: The University of Tokyo

summary: We identified a novel lipogenic gene, ELOVL7, as an over-expressed gene in prostate cancer (PC) cells and it is involved in elongation of long-chain fatty acids and can play some important roles in prostate carcinogenesis. To investigate into downstream pathways of ELOVL7, we performed the gene expression analysis by the genome-wide cDNA microarrays. The genome–wide gene expression patterns were compared between PC cells transfected with siELOVL7 and these with siCONTROL and between the ELOVL7-overexpressing in-vivo tumors in mice bread with high fat diet (HFD) and those with normal diet (ND). HFD treatment significantly promoted the growth of ELOVL7-overexpressed PC cells in vivo. As a result, we identified five genes (IER3, HDAC8, GDF15, UGT2B11, and S1PR3) whose expressions were down-regulated with 20% and more in ELOVL7 knockdown and also were up-regulated with 20% and more in the tumors in the mice bread with HFD.

overall_design: The genome–wide gene expression patterns were compared between PC cells transfected with siELOVL7 and these with siCONTROL and between the ELOVL7-overexpressing in-vivo tumors bread with high fat diet (HFD) and those with normal diet (ND). HFD treatment significantly promoted the growth of ELOVL7-overexpressed PC cells in vivo.　The competitive hybridization was performed three times for each comparison (siELOVL7-1, -2, and-3; HFD-1, -2, and -3).

last: Nakagawa

first: Kenji

last: Tamura

first: Yusuke

last: Nakamura


title: Genomic profiling of chondrosarcoma: chromosomal patterns in central and peripheral tumors (BAC/CGH)

city: Lund

first: Karolin

last: Nord

address: BMC C13

department: Dept of Clinical Genetics

country: Sweden

institute: Lund University

summary:
DNA copy number analysis of 67 fresh frozen chondrosarcoma biopsies using 32k BAC and 244k oligo array CGH. Genomic imbalances, in most tumors affecting large regions of the genome, were found in 90% of the cases. Although rare, recurrent amplifications were found at 8q24.21-q24.22 and 11q22.1-q22.3, and homozygous deletions of loci previously implicated in chondrosarcoma development affected the CDKN2A, EXT1 and EXT2 genes.

Keywords: chondrosarcoma, array comparative genomic hybridization

overall_design: DNA copy number analysis was performed using tiling microarrays containing more than 32,000 partly overlapping BAC clones, generating complete coverage of the Homo sapiens genome. The arrays were produced at the Swegene DNA Microarray Resource Center, Department of Oncology, Lund University (http://swegene.onk.lu.se), using BAC clones mapped to the hg17 genome build. Male genomic DNA (Promega) was used as reference. In five cases (cases 2, 4, 34, 36, and 67) the DNA copy number status was assessed by oligo microarrays containing ~236,000 oligonucleotide probes (Agilent, Palo Alto, CA, USA). These samples were hybridized against gender-matched reference DNA.

first: KH

last: Hallor

first: J

last: Staaf

first: JVMG

last: Bovée

first: PCW

last: Hogendoorn

first: AM

last: Cleton-Jansen

first: S

last: Knuutila

last: Savola

first: O

last: Brosjö

first: HCF

last: Bauer

first: F

last: VultvonSteyern

first: K

last: Jonsson

first: M

last: Skorpil

first: N

last: Mandahl

last: Mertens


title: Resistance of primary ovarian cancer cells to oncolytic adenoviruses

city: Seattle

first: Robert

last: Strauss

address: 1705 NE Pacific st.

department: Medicine

institute: University of Washington


title: Resistance of primary ovarian cancer cells to oncolytic adenoviruses part2 of 2

summary:
The mechanisms of primary ovarian cancer cells for resistance to viral oncolysis were investigated using Ad5/35.IR.E1A/TRAIL on clonal cultures derived from ovc316m cells.
Part 2 of 2, 26 clonal ovc316m cultures additionally to Resistance of primary ovarian cancer cells to oncolytic adenoviruses part1 of 2

overall_design: Cells were infected for 8 days and cell survival determined by MTT assay. Uninfected control cells of each clonal culture were utilized for DNA expression arrays. SKOV3-ip1 cells were used for reference RNA in all samples. The reference RNA from SKOV3-ip1 cells for part 2 of 2 had to be re-amplified.

first: Pavel

last: Sova

first: Ying

last: Liu

first: David

last: Pritchard

first: Andre

last: Lieber


title: Resistance of primary ovarian cancer cells to oncolytic adenoviruses part1 of 2

summary:
The mechanisms of primary ovarian cancer cells for resistance to viral oncolysis were investigated using Ad5/35.IR.E1A/TRAIL on clonal cultures derived from ovc316m cells.
Part 1 of 2, initital study involving 5 clonal ovc316m cultures

overall_design: Cells were infected for 8 days and cell survival determined by MTT assay. Uninfected control cells of each clonal culture were utilized for DNA expression arrays. SKOV3-ip1 cells were used for reference RNA in all samples.


title: Expression Profile-Defined Classification of Lung Adenocarcinoma

city: Nagoya

first: Takashi

last: Takahashi

state: Aichi

address: 1-1 Kanokoden, Chikusa-ku

institute: Aichi Cancer Center

summary:
Expression profiles of 18,175 unique genes and three major genetic changes, p53, EGFR and K-ras, were investigated in 149 patients with non-small cell lung cancer (NSCLC), including 90 patients with adenocarcinomas (AD) to determine their relationships with various clinicopathologic features and Gene Ontology (GO) terms.

Keywords: Disease state analysis

overall_design: Expression profiles in 149 patients with NSCLC, 9 patients with SCLC and 5 for normal lung tissue.

first: Toshiyuki

last: Takeuchi

first: Shuta

last: Tomida

first: Yasushi

last: Yatabe

first: Takayuki

last: Kosaka

first: Hirotaka

last: Osada

first: Kiyoshi

last: Yanagisawa

first: Tetsuya

last: Mitsudomi


title: Genomic profiling of chondrosarcoma: chromosomal patterns in central and peripheral tumors

first: Karolin

last: Nord

type:
Expression profiling by array
Genome variation profiling by array

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


title: Effect of Rac1 haploinsufficiency on the gene expression profile of mouse endothelial cells

status: Public on Mar 16 2009

first: Naoki

last: Sawada

address: 65 Landsdowne Street

institute: Brigham and Women's Hospital

summary: The therapeutic potential of neurotrophic factors has been hampered by their inability to achieve adequate tissue penetration. Brain blood vessels, however, could be an alternative target for neuro-salvage therapies by virtue of their close proximity to neurons. Here we show that hemizygous deletion of Rac1 in mouse endothelial cells (ECs) attenuates brain injury and edema following focal cerebral ischemia. To explore the mechanisms whereby decrease of Rac1 in ECs provide neuroprotection, ECs were derived from Rac1+/+ and Rac1+/- mice. Microarray analysis was performed to determine the differential gene expression between Rac1+/+ and Rac1+/- ECs.

overall_design: Rac1+/+ and Rac1+/- ECs were cultured to subconfluence. Total RNA was extracted and reverse transcribed. Rac1+/+ (n = 2) and Rac1+/- (n = 2) EC cDNAs were labeled with Cy3 and Cy5, respectively, and equimolar mixtures of labeled Rac1+/+ and Rac1+/- probes were hybridized to 2 slides (sample #21648, #21649). Additionally, Rac1+/+ (n = 2) and Rac1+/- (n=2) EC cDNAs labeled with Cy5 and Cy3 (dye-swapping) were hybridized to 2 slides (sample #21650, #21651). Comparison of signal intensities between Rac1+/+ (channel 2) and Rac1+/- (channel 1) was conducted on each slide (sample), and the ratio of intensities from 4 slides were statistically assessed for each gene feature to investigate differential gene expression.

first: James

middle: K

last: Liao