title: Methylation Changes During Soybean Seed Development

status: Public on Dec 22 2011

first: Bob

last: Goldberg

summary: What methylation changes are occurring during seed development largely remains unknown. To uncover the possible role of DNA methylation throughout all of seed development - from fertilization through dormancy and post-germination in soybean, we characterized the methylome of whole seeds representing the differentiation (GLOB and COT stages), maturation (early- [EM], mid- [B1] and late- [AA1] maturation stages), dormancy (DRY stage), and post-germination (seedling) phases of soybean seed development using Illumina sequencing. In addition, we characterized the methylome of the cotyledons of germinated seedling to examine methylation differences before and after germination.

overall_design: Illumina sequencing of bisulfite-converted genomic DNA from globular stage (GLOB), cotyledon-stage (COT), early-maturation stage (EM), mid-maturation stage (B1), and late-maturation stage (AA1), pre-dormancy stage 1 (AA2), pre-dormancy stage 2 (AA3), dormancy stage (DRY) seeds, and post-germination seedling (SDLG), seedling cotyledons (SDLG-COT).

first: Robert

first: Min

last: Chen

first: Jer-Young

last: Lin

first: Brandon

last: Le


title: A Transcriptome—Targeting EcoChip for Assessing Functional Mycodiversity

status: Public on Nov 14 2011

city: Bayreuth

first: Alfons

middle: R

last: Weig

address: Universitaetsstrasse 30

department: Genomics and Bioinformatics

country: Germany

institute: University of Bayreuth

summary: A functional biodiversity microarray (EcoChip) prototype has been developed to facilitate the analysis of fungal communities in environmental samples with broad functional and phylogenetic coverage and to enable the incorporation of nucleic acid sequence data as they become available from large-scale (next generation) sequencing projects. A dual probe set (DPS) was designed to detect a) functional enzyme transcripts at conserved protein sites and b) phylogenetic barcoding transcripts at ITS regions present in precursor rRNA. Deviating from the concept of GeoChip-type microarrays, the presented EcoChip microarray phylogenetic information was obtained using a dedicated set of barcoding microarray probes, whereas functional gene expression was analyzed by conserved domain-specific probes. By unlinking these two target groups, the shortage of broad sequence information of functional enzyme-coding genes in environmental communities became less important. The novel EcoChip microarray could be successfully applied to identify specific degradation activities in environmental samples at considerably high phylogenetic resolution. Reproducible and unbiased microarray signals could be obtained with chemically labeled total RNA preparations, thus avoiding the use of enzymatic labeling steps. ITS precursor rRNA was detected for the first time in a microarray experiment, which confirms the applicability of the EcoChip concept to selectively quantify the transcriptionally active part of fungal communities at high phylogenetic resolution. In addition, the chosen microarray platform facilitates the conducting of experiments with high sample throughput in almost any molecular biology laboratory.

overall_design: In this study, two independent RNA samples from a pine forest soil were labelled and hybridised to a custom-made EcoChip microarray consisting of about 9000 probes targeting expressed fungals genes and about 5000 probes targeting the precursor-rRNA of different fungal lineages

first: Derek

last: Persoh

first: Gerhard

last: Rambold


title: Allelic imbalance in Drosophila hybrid heads: exons, isoforms and evolution.

status: Public on Dec 21 2011

city: Gainesville

first: Martin

last: McCrory

state: FL

address: 2033 Mowry Road

country: USA

institute: University of Florida

type: Expression profiling by high throughput sequencing

summary: Using RNA-seq technology, a comprehensive assessment of cis regulatory divergence in interspecific hybrid female heads was conducted and patterns of sequence evolution (Begun et al. 2007) within causal loci were examined. Genotype specific references were shown to virtually eliminate the map bias plaguing this technology. A novel Bayesian model, which uses allele representation in F1 hybrid DNA sequence reads as a prior, was used to estimate allele frequencies in RNA sequences.

sample_organism: Drosophila melanogaster x Drosophila simulans

overall_design: There are three independent replicate samples of D. melanogaster - D. simulans F1 hybrid interspecific RNA from head tissues. RNA samples were sequenced (one lane per replicate) on a GAIIX with 54 bp paired end reads. One gDNA sample was collected for the same genotype and 3 technical replicates were run on a GAIIX with 36 bp paired end reads. The DNA data was deposited to SRA (accession SRA048616).

first: Rita

middle: M

last: Graze

first: Luis

middle: L

last: Novelo

first: Victor

last: Amin

first: Justin

last: Fear

first: George

last: Casella

first: Sergey

middle: V

last: Nuzhdin

first: Lauren

last: McIntyre


title: Mycobacterium tuberculosis H37Rv strains: Control vs. Lupulone

status: Public on Aug 30 2011

city: Changchun

first: Lu

last: Yu

address: Xi'an Road 5333

country: China

institute: Jilin university

summary: Transcriptional profiling of Mycobacterium tuberculosis H37Rv strains comparing control DMSO treated strains with Lupulone treated strains. Goal was to determine the effects of Lupulone against Mycobacterium tuberculosis H37Rv strains.

overall_design: Two-condition experiment,control DMSO treated strains vs. Lupulone treated strains. Biological replicates: 2 control replicates, 2 Lupulone replicates.

first: Junchao

last: Liang


title: A quantitative analysis of CLIP methods for identifying binding sites of RNA-binding proteins

status: Public on May 15 2011

city: Rehovot

first: Jean

last: Hausser

address: Herzl St.

country: Israel

institute: Weizmann Institute of Science

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

overall_design: Refer to individual Series


title: A quantitative analysis of CLIP methods for identifying binding sites of RNA-binding proteins (CLIP)

summary: Crosslinking and immunoprecipitation (CLIP) is increasingly used to map transcriptome-wide binding sites of RNA-binding proteins (RBPs). We developed a method for CLIP data analysis and applied it to compare 254 nm CLIP with PAR-CLIP, which involves crosslinking of photoreactive nucleotides with 365 nm UV light. We found small differences in the accuracy of these methods in identifying binding sites of HuR, a protein that binds low-complexity sequences and Argonaute 2, which has a complex binding specificity. We show that crosslink-induced mutations lead to single-nucleotide resolution for both PAR-CLIP and CLIP. Our results confirm the expectation from original CLIP publications that RNA-binding proteins do not protect sufficiently their sites under the denaturing conditions used during the CLIP procedure, and we show that extensive digestion with sequence-specific ribonucleases strongly biases the set of recovered binding sites. We finally show that this bias can be substantially reduced by milder nuclease digestion conditions.

overall_design: We performed duplicate experiments for each variant of the CLIP protocol (CLIP, PAR-CLIP), each protein (HuR, Ago2), and enzymatic digestion (complete T1 digestion, mild MNase digestion). In addition, we performed a single PAR-CLIP experiment with mild T1 digestion.

first: Shivendra

last: Kishore

first: Lukasz

last: Jaskiewicz

first: Lukas

last: Burger

first: Mohsen

last: Khorshid

first: Mihaela

last: Zavolan


title: Effect of acute ethanol on medidal prefrontal cortex across BXD genetic mapping panel and progenitors.

status: Public on Dec 01 2011

city: Richmond

first: Michael

last: Miles

state: VA

department: Pharmacology and Toxicology

institute: Virginia Commonwealth Univ.

summary: In order to elucidate the molecular mechanisms underlying individual variation in sensitivity to ethanol we profiled the prefrontal cortex transcriptomes of two inbred strains that exhibit divergent responses to acute ethanol, the C57BL6/J (B6) and DBA/2J (D2) strains, as well as 27 members of the BXD recombinant inbred panel, which was derived from a B6 x D2 cross. With this dataset we were able to identify several gene co-expression networks that were robustly altered by acute ethanol across the BXD panel. These ethanol-responsive gene-enriched networks were heavily populated by genes regulating synaptic transmission and neuroplasticity, and showed strong genetic linkage to discreet chromosomal loci. Network-based measurements of node importance identified several hub genes as established regulators of ethanol response phenotypes, while other hubs represent novel candidate modulators of ethanol responses.

overall_design: Animals were injected intraperitoneally (IP) with saline or 1.8 g/kg of ethanol. As part of a parallel study of ethanol induced anxiolysis, all mice underwent behavioral testing that included 15 minutes of restraint in a 50 mL conical tube followed by 10 minutes in a light-dark chamber. Mice were killed by cervical dislocation four hours following IP injection. Immediately thereafter, brains were extracted and chilled for 1 minute in iced phosphate buffer before being microdissected into 8 constituent regions, including the medial prefrontal cortex. Samples were randomly assigned to batch groups prior to total RNA extraction, cRNA synthesis and hybridization. Each microarray represent a pooling of 4-5 animals.

middle: F

first: Alex

middle: H

last: Putman

first: Paul

middle: J

last: Vorster

first: Aaron

last: Wolen


title: Salmonella Typhimurium: wild type vs. ∆scsA mutant

status: Public on Jul 26 2011

city: Norwich

first: Neil

last: Shearer

state: Norfolk

address: Colney Lane

department: Gut Health and Food Safety

country: United Kingdom

institute: Institute of Food Research

summary: Transcriptional profiling of log and stationary phases of S. Typhimurium, comparing wild type with the ∆scsA strain.

platform_organism: Salmonella enterica subsp. enterica serovar Typhimurium str. DT104

platform_organism: Salmonella enterica subsp. enterica serovar Typhimurium str. LT2

platform_organism: Salmonella enterica subsp. enterica serovar Enteritidis

platform_organism: Salmonella enterica subsp. enterica serovar Typhimurium str. SL1344

platform_organism: Salmonella enterica subsp. enterica serovar Gallinarum str. 287/91

overall_design: Each array used labelled cDNA against a common genomic DNA reference. Triplicate arrays were carried out for each of the 4 conditions: wild type log phase, wild type stationary phase, ∆scsA log phase and ∆scsA stationary phase.

first: Arthur

last: Thompson

first: Elin

last: Verbrugghe


title: Genome-Wide Transcript Profiling During Soybean Seed Development and Throughout the Soybean Life Cycle

status: Public on May 10 2011

city: Los Angeles

first: Bob

last: Goldberg

state: CA

address: 610 Charles E Young Drive East

department: Molecular, Cell and Developmental Biology

institute: University of California, Los Angeles

summary: We report the genome-wide transcriptome of soybean seeds across several stages of seed development and the entire life cycle using Illumina high-throughput sequencing technology. Specifically, we profiled whole seeds containing globular-stage, heart-stage, cotyledon-stage, early maturation-stage, mid-maturation-stage, and late-maturation-stage embryos. We also profiled dry soybean seeds, and vegetative and reproductive tissues including leaves, roots, stems, seedlings, and floral buds.

overall_design: IIllumina sequencing of transcripts from whole seeds at five stages of seed development (globular, heart, cotyledon, early-maturation, mid-maturation, late-maturation, dry), and vegetative (leaves, roots, stems, seedlings) and reproductive (floral buds) tissues.

first: Robert

middle: B

first: Brandon

last: Le

first: Kelli

last: Henry

first: Anhthu

middle: Q

last: Bui

first: Xiaomeng

last: Wu

first: Min

last: Chen


title: Discovery of salivary transcriptomic biomarkers for ovarian cancer

status: Public on May 12 2011

first: Yu-Hsiang

last: Lee

address: 10833 Le Conte Avenue

department: Dental Research Institute

institute: UCLA

summary:
Ovarian cancer is the leading cause of death in gynecological diseases, and has been considered as one of the most fatal cancers due to lack of reliable detection strategy in the early stage. Therefore the capability to detect the morbidity initiation with an sensitive and effective approach is one of the most desirable goals for curing ovarian cancer. In this study, we used microarray technology for salivary mRNA biomarkers discovery, and evaluated the performance and translational utilities of discovered markers from a clinical study using an independent sample cohort .
We used microarrays to profile and compare the gene expressions between ovairan cancer patient and matched controls, and identified seven down-regulated genes after the validation study.

overall_design: To find salivary transcriptomic biomarkers for ovarian cancer, salivary transcriptomes in 11 ovarian cancer patients and 11 matched controls were profiled using Affymetrix HG-U133-Plus-2.0 array, and followed by t-test and fold-change analyses. The biomarker candidates selected from the microarray results were subjected to clinical validation using an independent sample cohort by RT-qPCR. The prediction power of biomarkers was analyzed by logistic regression approach

first: Jae

last: Kim

first: Hui

last: Zhou

first: David

middle: T

last: Wong


title: AEG-1 silencing induced gene expression alteration in glioma cells

status: Public on Jul 12 2011

city: Guangzhou

first: Mengfeng

last: Li

state: Guangdong

address: 74# zhongshan 2 Rd.

department: Zhongshan school of medicine

institute: Sun Yat-sen University

summary: This experiment is designed to evaluate gene expression alteration following AEG-1 (MTDH) silencing in glioma cells. AEG-1 is found to be a potential regulator through interaction with other transcription factor. We intended to investigate the enriched transcription factor dependent downstream gene sets regulated by AEG-1 form the differential expressed genes we obtained from AEG-1 silencing.

overall_design: Total RNA were extracted from U87MG glioma cells stably silencing AEG-1 and correspondent pSuper Retro vector cells respectively. Passage 5 of the indicated cells after infection and selection were harvested for RNA extraction.

first: Zhe

last: Ying

first: Jun


title: Epigenetic Regulation of the Transition from Seed Maturation to Germination in Soybean

summary: How epigenetics is involved in the transition from seed maturation to seed germination largely remains elusive. To uncover the possible role of epigenetics in gene expression during the transition from seed maturation to seed germination in soybean, the transcriptome of cotyledons from four stages of soybean seed maturation and germination, including mid-late maturation, late maturation, seed dormancy and seed germination, were profiled by Illumina sequencing. For the genes that are quantitatively regulated at the four stages, two antagonistic epigenetic marks, H3K4me3 and H3K27me3, together with the binding of RNA polymerase II, were investigated at the four stages by chromatin immunoprecipitation (ChIP). For 10 out of 16 genes examined, the relative enrichment of histone modification marks (H3K4me3 and H3K27me3) and RNA polymerase II binding on their promoter regions correlates well with their relative expression levels at four stages, suggesting the involvement of epigenetics in transcriptional regulation. A striking finding is that seed germination-specific genes start to show open chromatin (H3K4me3) during late seed maturation although their transcripts do not accumulate, which is further supported by RNA polymerase II binding. Together, our results provide the first evidence that seed germination genes can be primed for transcription (open chromatin and RNA polymerase II binding) during seed maturation, highlighting that the transition from seed maturation to seed germination starts at late seed maturation stages at both the genetic and epigenetic levels.

overall_design: Illumina sequencing of transcripts from cotyledons of mid-maturation (B1 stage) and late maturation (AA1 stage) seeds, whole dry seeds, and cotyledons of seedlings six days after imbibition.

first: Zidian

last: Xie

first: Tomokazu

last: Kawashima


title: Gene profiling during symbiosis between the actinorhizal tree Alnus glutinosa and the actinobacteria Frankia alni ACN14

status: Public on Apr 11 2011

city: Villeurbanne Cedex

first: Nicole

last: ALLOISIO

address: 43 bd du 11 Novembre 1917

country: France

institute: CNRS / Univ Lyon 1

summary:
Alnus glutinosa belongs to a family of angiosperms called actinorhizal plants because they can develop nitrogen-fixing nodules in association with the soil bacteria Frankia. The aim of this transcriptomic study was to get a global view of the plant symbiotic genetic program and to identify new key plant genes that control nodulation during symbiosis in A. glutinosa.
Symbiosis between A. glutinosa and Frankia was obtained after inoculation of young plant with a concentrated culture of the bacteria. Inoculation was performed in a medium depleted in nitrogen  which favors the induction of nitrogen fixing symbiosis. For this study we considered two stages of symbiosis:  - an early stage where inoculated roots were harvested 7 days after inoculation with the bacteria and compared to two controls (non-inoculated roots grown with or without nitrogen and harvested at the same time)  - a late stage where nodules (nitrogen-fixing specific organs) were harvested 21 days after inoculation and compared to non-inoculated roots harvested on the day of inoculation (which is our reference time 0d).  Three biological replicates were used for each condition.

overall_design: Microarrays were designed by Imaxio (Clermont Ferrand, France ; http://www.imaxio.com/index.php) which has been accredited by Agilent Technologies (Palo Alto, CA, USA; http://www.home.agilent.com/agilent/home.jspx) as a certified service provider for microarray technologies. Based on 14327 annotated unigenes for A. glutinosa, 60mers probes were designed using eArray software (1 probe per unigene) and custom 8 x 15K Oligo Microarrays were manufactured by Agilent

last: Alloisio

first: P

last: Fournier

last: Pujic

first: Philippe

last: Normand


title: Global changes in gene expression of buffalo cow corpus luteum during prostaglandin F2alpha- induced luteolysis

status: Public on Mar 16 2011

city: BANGALORE

first: KUNAL

last: SHAH

state: KARNATAKA

address: C.V.RAMAN AVENUE, MALLESHWARAM

department: Molecular Reproduction Development and Genetics

country: India

institute: Indian Institute of Science, Bangalore

summary: In monoovulatory species like bovines, a striking diversity in regulation of the corpus luteum (CL) function exists within species during different stages of the luteal phase.  The function and life span of CL appears to be regulated by the interplay of both luteotrophic and luteolytic factors, whose roles and actions are varied from one species to another. Although, studies continue to expand our current understanding of the cellular and molecular actions of prostaglandin F 2alpha (PGF2α - a physiological luteolysin)-induced luteolysis, knowledge of the mechanism whereby, PGF2α mediates its luteolytic actions remains poorly understood. The mechanism of PGF2α-induced luteolysis is a complex process involving changes in expression of multiple genes encoding gene products that regulate steroidogenesis, stress related or apoptotic genes, factors involved in immune response and enzymes that stimulate PGF2α production. Thus, in the present study, efforts were made to identify the temporal changes in the global gene expression profile in the CL of buffalo cows in response to PGF2α treatment. The molecular signature of genome-wide transcriptional changes in the CL tissue subjected to PGF2α-induced luteolysis will expand our knowledge on basic mechanism/s that regulates the luteolytic process. The results obtained in this study provide insight into the mechanisms underlying the PGF2α- induced regression of CL.Keywords: CL, PGF2α,  gene expression

overall_design: The objective of this study was to identify the PGF2α-induced temporal changes in transcriptome of bovine CL on day 11 of estrous cycle. In the experiments with buffalo cows, animals with history of normal estrous cyclicity were treated with Cloprostenol (an PGF2α analogue; 500 µg i.m.) on day 11 of the estrous cycle to induce luteolysis and CL tissues were collected at (control; n=4 amimals/time point), 3 (n=2 animals/time point), 6 and 18 (n=3 animals/time point) h post PGF2α administration  to examine gene expression changes associated with immediate early and delayed changes in the CL, respectively. Following retrieval of CL from the ovary, flash frozen in liquid nitrogen and stored at -70 °C for the purpose of RNA isolation. The isolated RNA was labeled and hybridized to Affymetrix GeneChip Bovine Genome Arrays as per the manufacturer’s instructions.

first: B

middle: S

last: Kunal

first: T

last: Sudeshna

first: S

last: Hepziba

last: Jayaram

first: R

last: Medhamurthy


title: differential gene expression in Sensitive and Resistant Pig to Escherichia coli F18

status: Public on Jan 26 2011

city: Yangzhou

first: Lan

last: Ye

state: Jiangsu

address: Daxue Road  NO 88

department: animal science and technology

institute: Yangzhou university

summary: An M307 G/A point mutation of FUT1 gene has been considered as a usful marker to select the piglets that are sensitive(GG/AG genotype) or resistant(AA) to Escherichia coli F18 in foreign pig breeds.However,it is not suitable to Chinese native breeds. Duodenal tissues were collected from 8 full-sib pairs of Sutai pigs (a new hybrid between the Duroc and Taihu breeds) at the age of 28-day differing in adhesion phenotype to find the differential genes that can be used as the candidate genes fit for Chinese native breeds.

overall_design: piglets at the age of 28-day sensitive (GG/AG genotype) to Escherichia coli F18 vs resistant(AA) ones.Biological replicates:8 full-sib pairs of Sutai pigs (4 pairs for GG/AA genotype of FUT1 gene, 4 pairs for AG/AA genotype)

first: Wenbin

last: Bao


title: The DNA damage response transcriptome of the moss Physcomitrella patens

status: Public on Nov 10 2010

city: San Diego

first: John

middle: William

last: Whitaker

state: Ca

address: Gliam Drive

institute: UCSD

summary: The moss Physcomitrella patens is remarkable for the ease with which mutant alleles of any gene can be generated by highly efficient homologous recombination-mediated gene targeting. Targeted transgene integration is believed to be mediated through the capture of transforming DNA by the homologous recombination DNA repair pathway. To identify components of this pathway in P. patens we have undertaken a transcriptomic analysis of the response to the sublethal induction of bleomycin-induced DNA double-strand breaks using massively parallel (Illumina) cDNA sequencing. Transcripts significantly increased in bleomycin-treated tissue include a number encoding conserved DNA-DSB components in both the homology-dependent pathway (including Rad51, CTiP, DNA ligase 1, Replication protein A, ATR) and the non-homologous end-joining pathway (including Xrcc4, DNA ligase 4, Ku70, Ku80,  PARP). Differentially regulated cell-cycle components include up-regulated Rad9 and Hus1 DNA-damage-related checkpoint proteins and down-regulated D-type cyclins and B-type CDKs, commensurate with the imposition of a checkpoint in the G2 stage of the cell cycle characteristic of homology-dependent DNA-DSB repair.  Comparison of the DNA damage transcriptome of P. patens with that of A. thaliana reveals significant up-regulation of a number of P. patens genes encoding ATP-dependent chromatin remodelling helicases of the SNF-2 class. These represent candidates for investigation of their role in mediating efficient gene targeting in P. patens.

overall_design: Gene expression profiling monitored by transcript abundance in control tissue and tissue treated with the DNA-DSB inducing agent, bleomycin

middle: W

first: Yasuko

last: Kamisugi

first: Andrew

middle: C

last: Cuming


title: Therapeutics of Ebola hemorrhagic fever: Whole-genome transcriptional analysis of successful disease mitigation

status: Public on Nov 30 2010

city: Boston

first: Judy

last: Yen

state: MA

address: 620 Albany St., Rm 401H

department: NEIDL/Microbiology

institute: Boston University School of Medicine

summary: Ebola (EBOV) virus causes severe and often lethal hemorrhagic fever in humans and nonhuman primates (NHP), and has been classified as a Category A bioweapon agent.  There are currently no approved preventive vaccines or postexposure treatments for EBOV hemorrhagic fever.  The mechanisms of EBOV pathogenesis are only partially understood, but the dysregulation of normal host immune responses (including destruction of lymphocytes, increases in levels of circulating proinflammatory cytokines, and development of coagulation abnormalities) is thought to play a major role.  Accumulating evidence suggests that much of the observed pathology is not the direct result of virus-induced structural damage but rather is due to the release of soluble immune mediators from EBOV-infected cells.  It is therefore essential to understand how the candidate therapeutic may be interrupting the disease process and/or targeting the infectious agent.  Identification of effective treatment strategies may greatly benefit based on identification of molecular features of the host response to infection and treatment.  In order to identify these gene signatures related to correlates of protection, we used a DNA microarray-based approach to compare the host genome-wide responses of EBOV-infected NHP responding to candidate therapeutics.  With this approach, we have identified genes that appear to correlate with survival, including chemokine ligand 8 (CCL8/MCP-2), and revealed a subset of distinctly differently expressed genes that may provide possible targets for future diagnostics or therapeutics.  These analyses will assist us in understanding the pathogenic mechanisms of EBOV infection as well as identify improved therapeutic strategies.

sample_organism:
Macaca mulatta
Homo sapiens

overall_design: Transcriptional analysis of global gene expression changes in Zaire Ebola Virus (ZEBOV)-infected rhesus macaques that were treated with either recombinant nematode anticoagulant protein c2 (rNAPc2) or recombinant human activated protein C (rhAPC).  Animals were infected with 1000pfu ZEBOV, then subsequently treated with rNAPc2 or rhAPC.  Four animals were left untreated for controls.  Blood samples were taken at specified days post-infection and PBMCs were isolated from the samples and inactivated in TRIzol reagent. Total RNA was isolated from the samples, then linearly amplified and hybridized to a whole genome long-oligonucleotide microarray in a two color comparative format with a commercially available human reference RNA from Stratagene as a consistent control in dataset comparisons.

middle: Y

first: Sara

last: Garamszegi

first: Joan

last: Geisbert

first: Kathleen

last: Rubins

first: Thomas

first: Yu

last: Xia

last: Connor

first: Lisa

middle: E

last: Hensley


title: Transcriptional effects of 1,25 dihydroxi-vitamin D3 physiological and supra-physiological concentrations in breast cancer organotypic culture

status: Public on Feb 11 2011

city: São Paulo

first: Maria Lucia

middle: Hirata

last: Katayama

state: São Paulo

address: Av Dr Arnaldo

department: Radiology and Oncology

country: Brazil

institute: Faculty of Medicine University São Paulo

summary: Vitamin D may have anti-tumorigenic actions by influencing the gene expression profile of target tissues, which possess vitamin D receptors.  We used a more physiological in vitro model, represented by short-term culture of breast cancer tissue slices, to study the transcriptional effects of a near physiological concentration of calcitriol. Methods. Breast cancer fragments were sliced and maintained in culture for 24 hours in the presence or absence (control) of calcitriol 0.5nM or 100nM (called physiological or supra-physiological concentrations). Five and 16 samples were included in a test or validation group and gene expression was analyzed by microarray (using SAM paired analysis) or qPCR, respectively. Results. Nine genes were regulated by calcitriol 0.5nM including CYP24A1, DPP4, EFTUD1 (FDR£0.01), which were up-modulated. However, using a less stringent FDR value (0.25) 61 genes were differentially expressed. Among the down-regulated transcripts were members of the MHC class II complex (HLA-DPA1, HLA-DQA1, HLA-DQA2, HLA-DQB1, HLA-DRA, HLA-DRB1, HLA-DRB3) and IgG binding (FCGR2A, FCGR2B, FCGR2C). There was an enrichment of genes presenting transcription factor binding sites for vitamin D, among the up-regulated genes and for interferon regulatory factor IRF1, among the down-regulated genes. Analyzing calcitriol supra-physiological effects, a more impressive transcriptional modulation was identified and 136 and 60 genes (FDR 0.1) were more and less expressed in treated samples. Up-regulated genes were involved in vitamin metabolic process, regulation of leukocyte-mediated immunity and positive regulation of alpha-beta T cell activation. Many of the induced genes were already reported as vitamin D responsive genes, including CD14, which was also up-regulated in another set of 16 samples. Genes modulated by both calcitriol concentrations were CYP24A1, DPP4, CA2 (these three in both sets of samples), EFTUD1, TKTL1, KCNK3. Conclusion. Small increments in calcitriol concentration, within the physiological range, for a relatively short period of time may exert transcriptional effects in breast cancer samples. Further studies employing physiological concentrations of vitamin D for longer periods of time may help to elucidate the hormone effects in breast cancer treatment and prevention.

overall_design: Breast cancer fragments were sliced and maintained in culture for 24 hours in the presence or absence (control) of calcitriol 0.5nM or 100nM (called physiological or supra-physiological concentrations). Five samples were included and were categorized according to treatment in three groups: control (A), 1,25(OH)2D3 0.5nM (B) and 1,25(OH)2D3 100nM (C).Total RNA was isolated using RNeasy kit and than was carried out according to microarray Affymetrix protocol.      Data was then assessed with GeneSpring X software for background correction, normalization and summarization of raw data (CEL files) using the Robust Multi-Array Average (RMA). Five samples were included in a test and gene expression was analyzed by microarray (using SAM paired analysis). To establish a differential gene expression profile between vitamin D treated and untreated samples, SAM two class paired, provided on MEV (MultiExperiment Viewer – Boston, MA, USA) was used. Unsupervised hierarchical clustering based on Euclidean distance and average linkage was used to verify association patterns. The reliability of the clustering was assessed by the Bootstrap technique using MEV (MultiExperiment Viewer – Boston, MA, USA). Samples (16) were included in a validation group and was analyzed by qPCR.

first: Cintia

last: Milani

first: Maria

middle: A

last: Folgueira


title: mRNA analysis in different mouse tissues

status: Public on Nov 18 2010

city: Leuven

first: Katleen

last: Lemaire

address: Herestraat 49, bus 901

country: Belgium

type:
Expression profiling by array
Third-party reanalysis

summary: The functioning of a specific tissue depends on the expression pattern of the different genes. We used microarrays to compare gene expression across different murine tissues, to get a better understanding in the expression pattern and functioning of the different tissues. With this analysis, we were not only able to identify genes that were specifically expressed in a spicific tissue but, as important, we also identified genes that were specifically repressed in a tissue, compared to al the other analysed tissues.

overall_design:
C57Bl6/j6 (Janvier) mice between 10 and 12 weeks old were used to isolate 22 different tissues. Mainly male mice were used, except for placenta and ovary, where female mice were used. Also the mouse insulinoma cellline MIN6 was analysed in this study.
For every tissue, at least 3 biological replicates were used.

first: Leentje

last: Van Lommel

first: Frans

last: Schuit

first: Anica

last: Schraenen

first: Lieven

last: Thorrez


title: Transactivation-deficient p53 Mutants in Ras-induced Cellular Senescence

status: Public on May 13 2011

city: Stanford

first: Dadi

last: Jiang

address: 269 Campus Drive, CCSR-South 1240

institute: Stanford University

summary: As a critical cellular stress sensor, p53 mediates a variety of defensive processes including cell-cycle arrest, apoptosis, and senescence to prevent propagation of hyperproliferative cells or cells with a damaged genome, hence the formation of neoplasia. Transactivation of downstream genes plays an important while sometimes controversial role in regulating these cellular processes. To evaluate the dependence on transcriptional activation in p53’s activities, we generated genetically-modified mouse lines carrying mutations in the transactivation domains (TADs) of p53. These transactivatio-deficient mutants serve as unique reagents to probe the dependence on robust transactivation in p53-mediated cellular functions, as well as the underneath mechanisms. To identify genes differentially regulated by these p53 mutants, we performed gene expression profiling analysis on mouse embryonic fibroblast cells (MEFs) from these mice in the context of oncogenic Ras-induced premature cellular senescence.

overall_design: Mouse embryonic fibroblasts (MEFs) with different p53 genotypes were infected with retroviral H-Ras V12, which induces premature cellular senescence in p53 wild-type MEFs but not in p53 null MEFs. 5 genotypic groups of MEFs were used in the study: (i) p53L25Q/W26S, or "25,26", in which the first TAD (transactivation domain) of p53 is disrupted by the mutation, 5 biological samples; (ii) p53W53Q/F54S, or "53,54", in which the second TAD of p53 is disrupted by the mutation, 3 biological samples; (iii) p53L25Q/W26S/W53Q/F54S, or "QM", in which both TADs or p53 are disrupted, 3 biological samples; (iv) p53 wild-type, or "WT", 6 biological samples; (v) p53 null, or "Null", 6 biological samples.


title: Fine mapping of androgen regulated genes in LNCaP cells

status: Public on Sep 01 2010

city: Cambridge

first: Charlie

middle: Edward

last: Massie

state: CAMBS

address: CRUK CRI, Robinson Way

department: Oncology

institute: Cambridge University and Cancer Research UK

summary:
Detailed analysis of androgen regulated gene expression in the LNCaP prostate cancer cell line.  Since androgens and the AR are known to be important for prostate cancer cell proliferation and invasion we aimed to identify androgen receptor (AR) regulated genes by combining this detailed Illumina beadarray study of androgen regulated gene expression with AR ChIP-sequencing data.
LNCaP cells were grown in RPMI medium supplemented with 10% charcoal dextran stripped (steroid depleted) FBS for 72h, before treatment with 1nM R1881 or vehicle control. Total RNA was harvested every 30min for 4h and then every hour up to 24h using trizol (Sigma), quantified using a Nanodrop spectrophotometer (ND-1000). RNA samples were prepared for analysis on Illumina Human 6 v2 BeadArrays and Biotrove Realtime PCR panels according to the manufacturers protocols (see Supplementary Methods for details).  Expression array data were normalised in R using the beadarray software and BASH (see Supplementary Methods for details) (Cairns et al., 2008; Dunning et al., 2007).

overall_design:
48 total RNA samples from LNCaP cells grown for 72h in RPMI supplemented with 10% charcoal dextran stripped FBS, comprising: 3 time zero samples; 10 vehicle (ethanol) control samples taken at 2h, 4h, 8h, 12h, 24h in duplicate; 36 androgen (R1881) treated samples taken every 30min for 4h then every hour until 24h following treatment (with replicates at 1h, 2h, 4h, 8h, 12h, 16h, 20h, 24h)

The Illumina HumanWG v2 BeadArrays consist of two replicate sections that we treat as technical replicate arrays for the purposes of this analysis due to small but systematic shifts between sections that need to be addressed in the normalization. Data were analysed from the raw bead-level using the beadarray software, with spatial artefacts identified and removed automatically (BASH) and curated manually (Cairns et al., 2008; Dunning et al., 2007)

first: Andy

last: Lynch


title: Genome-wide assessment of imprinted expression in human cells

status: Public on Jan 21 2011

city: Montreal

first: Tomi

middle: Markku

last: Pastinen

state: Quebec

address: 740, Dr. Penfield Avenue

country: Canada

institute: McGill University and Génome Québec Innovation Centre

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

summary: Parent-of-origin dependent expression of alleles, imprinting, has been suggested to impact a substantial proportion of mammalian genes. Its discovery requires allele-specific detection of expressed transcripts, but in some cases detected allelic expression (AE) bias has been interpreted as  imprinting without demonstrating compatible transmission patterns and excluding heritable variation. Therefore, we utilized a genome-wide tool exploiting high density genotyping arrays in parallel measurements of genotypes in RNA and DNA to determine AE across the transcriptome in lymphoblastoid cell lines (LCLs) and skin fibroblasts derived from families.

overall_design: To investigate imprinting, transmission patterns were analyzed in 2 LCL trios, 1 LCL 3-generation pedigree and 9 FB trios. To investigate random monoallelic expression, 2 cell lines were treated with three concentrations of 5-azadeoxycytidine (AZA), and one cell line was treated for two time periods.

first: Lisanne

last: Morcos

first: Bing

last: Ge

first: Vonda

last: Koka

first: Kevin

last: Lam

first: Dmitry

middle: K

last: Pokholok

middle: L

last: Gunderson

first: Alexandre

last: Montpetit

first: Dominique

last: Verlaan


title: Silencing of mrhl non coding RNA in mouse spermatoginial cells GC1-Spg

status: Public on May 03 2010

city: Bangalore

first: Gayatri

last: Ganesan

address: Jakkur

department: MBGU

institute: JNCASR

summary: Mrhl is a  non coding RNA identified from mouse chromosome 8. It is a 2.4kb poly adenylated, nuclear restricted RNA expressed in multiple tissues. The 2.4 kb RNA also undergoes a nuclear processing event mediated through Drosha that generates an 80nt intermediate RNA. This study was aimed at understanding the functiion of mrhl by silencing the mrhl RNA in the mouse spermatogonial cells using a pool of siRNAs targeted against the mrhl and analyse the global gene expression change using  Affymetrix mouse expression array. The mRNAs that showed significant change in expression in mrhl siRNA treated cells against control were studied further for their biological significance with respect to mrhl silencing.

overall_design: The mouse spermatogonial cells that were either mock silenced using scrambled siRNA or silenced using siRNAs against mrhl were harvested at 44hrs post transfection.  The silencing efficiency was  checked by realtime quantitative PCR and was found be almost 80% in all three  independent experiments.The total RNA was extracted using Trizol and was taken further for microarray expriment. All the subsequent steps were according to the Affymetrix protocol. Gene expression was profiled for control and mrhl silenced cells.

first: M

middle: R

last: Rao


title: Effect of 1-methylcyclopropene (1-MCP) on preclimacteric nectarine (Prunus persica cv Stark Red Gold) fruit.

status: Public on Apr 01 2010

city: Padova

first: Livio

last: Trainotti

address: Via Ugo Bassi 58/B

department: Biology

country: Italy

institute: University of Padova

summary:
Peach is a climacteric fruit whose ripening is largely dependent on ethylene. Recently, it has been shown that auxin is able to autonomously regulate the expression of ripening related genes, and that a cross-talk between ethylene and auxin occurs during the ripening process.
By using the non-destructing index of absorbance difference (IAD), mature nectarines were homogeneously grouped into three classes (0, 1 and 2) according to their ripening phase and treated with 1-methylcyclopropene (1-MCP), known to block ethylene receptors. 1-MCP responses differed in the three classes that were thus used to molecularly investigate the auxin-ethylene cross-talk during the transition from maturation to ripening and the accompanying switch of ethylene biosynthesis from System-1 to System-2. Microarray experiments showed that 1-MCP modified the expression of 121 genes (63 were up-regulated and 59 down-regulated). Besides inducing ethylene-, auxin- and ripening-repressed genes and repressing ethylene-, auxin- and ripening-induced genes, also genes induced by ripening, auxin and 1-MCP were discovered. The up-regulating effect of 1-MCP was observed for ctg134, similar to signalling peptides, for 1-aminocyclopropane-1-carboxylic acid synthase 1 (ACS1) and, interestingly, for many auxin-regulated genes, such as GH3, whose expression is widely used as a marker for free auxin. The latter finding, together with the induction of an IAA amidohydrolase, suggests that the 1-MCP application could have caused a rise in free auxin, thus leading to an increase in ethylene biosynthesis through the induction of the tightly regulated ACS1 gene.

overall_design:
A "direct comparison" approach has been used to investigate the effect at the transcriptional level of 1-methylcyclopropene (1-MCP) on preclimacteric nectarine (Prunus persica cv Stark Red Gold) fruit.
RNA has been extracted from either 1-MCP treated fruit or from fruit kept in air (control).
Four hybridization have been conducted for a total of four technical replicates (with dye-swap).

first: Alice

last: Tadiello


title: Gene expression in LSK cells of C57B/L6, DBA/2J, B.D(Chr5), & D.B(Chr5) mice

status: Public on May 21 2010

first: Amanda

last: Waterstrat

state: KY

address: 521 Lancaster Ave

department: Biological Sciences

institute: Eastern Kentucky University

summary:
These mouse strains differ in absolute numbers of hematopoietic stem cells but differ genetically only at the Chr 5 congenic locus.
We used microarray analysis to identify candidate regulators of hematopoietic stem cells based on differential gene expression patterns.

overall_design: Triplicate RNA samples, except for duplicate DBA/2J samples,  were isolated from 80,00-137,000 LSK cells from each strain. Each of the 11 samples were analyzed on individual microarrays on two Illumina Sentrix-Mouse 6 Whole Genome Expression Beadhips.

first: Gary

last: Van Zant


title: Cardiac Resynchronization Therapy (1-color design)

status: Public on Jan 05 2010

city: Baltimore

first: Andreas

middle: Sebastian

last: Barth

state: MD

address: 720 Rutland Ave.

department: Medicine, Division of Cardiology

institute: Johns Hopkins University

summary:
Objectives: To test whether (1) electromechanical dyssynchrony induces region-specific alterations in the myocardial transcriptome and (2) dyssynchrony-induced gene expression changes can be corrected by cardiac resynchronization (CRT).
Background: To date, CRT is the only heart failure treatment that can both acutely and chronically increase systolic function and prolong survival, something not yet achieved by a drug therapy. However, the mechanisms underlying the benefits of CRT remain elusive.
Methods: Adult dogs underwent left bundle branch ablation (LBBB) and right atrial pacing at 200 bpm for either 6 weeks (dyssynchronous heart failure, DHF, n=12) or 3 weeks followed by 3 weeks of resynchronization by bi-ventricular pacing at the same pacing rate (CRT, n=10). Control animals without LBBB were not paced (NF, n=14). Echocardiography and invasive hemodynamic measurements were performed at 3 and 6 weeks. At 6 weeks, RNA was isolated from the anterior and lateral LV walls and hybridized onto canine-specific 44K microarrays.
Results: In DHF, transcriptional changes consistent with re-expression of a fetal gene program were primarily observed in the anterior LV, resulting in increased regional heterogeneity of gene expression within the left ventricle. Dyssynchrony-induced region-specific expression changes in 1050 transcripts were reversed by CRT to levels of NF hearts (false discovery rate <5%). CRT remodeled transcripts with metabolic and cell signaling function and greatly reduced regional heterogeneity of gene expression compared with DHF.
Conclusions: Our results demonstrate a profound effect of electromechanical dyssynchrony on the regional cardiac transcriptome, causing gene expression changes primarily in the anterior LV wall. CRT corrected the alterations in gene expression in the anterior wall by reversing the fetal gene expression pattern, supporting a global effect of biventricular pacing on the ventricular transcriptome that extends beyond the pacing site in the lateral wall.

overall_design: Designed as a 1-color experiments, samples from anterior and lateral left ventricular myocardium from non-failing, DHF and CRT animals were labeled with Cy3 and hybridized onto Agilent 44K long oligonucleotide arrays.

first: Wayne

last: Yu

first: David

last: Kass

first: Gordon

middle: F

last: Tomaselli


title: Cardiac Resynchronization Therapy (2-color design)

overall_design: Complementary study to GSE14327. While GSE14327 was designed as a 1-color microarray experiment, this series was carried out following a 2-color design (anterior and lateral LV wall labeled with Cy3 and Cy5, respectively, including dye swaps).


title: MAQC-II Project: Multiple myeloma (MM) data set

status: Public on Sep 11 2010

city: Shanghai

first: Leming

last: Shi

address: 2005 Songhu Road

department: School of Life Sciences

institute: Fudan University

summary: The multiple myeloma (MM) data set (endpoints F, G, H, and I) was contributed by the Myeloma Institute for Research and Therapy at the University of Arkansas for Medical Sciences (UAMS, Little Rock, AR, USA). Gene expression profiling of highly purified bone marrow plasma cells was performed in newly diagnosed patients with MM. The training set consisted of 340 cases enrolled on total therapy 2 (TT2) and the validation set comprised 214 patients enrolled in total therapy 3 (TT3). Plasma cells were enriched by anti-CD138 immunomagnetic bead selection of mononuclear cell fractions of bone marrow aspirates in a central laboratory. All samples applied to the microarray contained more than 85% plasma cells as determined by 2-color flow cytometry (CD38+ and CD45-/dim) performed after selection. Dichotomized overall survival (OS) and eventfree survival (EFS) were determined based on a two-year milestone cutoff. A gene expression model of high-risk multiple myeloma was developed and validated by the data provider and later on validated in three additional independent data sets.


title: Cardiac Resynchronization Therapy Corrects Dyssynchrony-induced Regional Gene Expression Changes on a Genomic Level

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

overall_design: Refer to individual Series


title: Identification of COX-2 as a major actor of transcriptomic adaptation of endothelial and tumor cells to cyclic hypoxia

status: Public on Dec 24 2009

city: Brussels

first: Olivier

last: Feron

address: Avenue E. Mounier, 52 - FATH5349

department: IREC

institute: UCL

summary: Tumor hypoxia is not a stable phenomenon but cycles between periods of deep hypoxia and reoxygenation. Cyclic hypoxia originates from heterogeneities in red blood cell flux and from the permanent remodelling of the angiogenic vascular network. Endothelial cells lining tumor blood vessels are therefore also influenced by cyclic hypoxia. The gene expression pattern promoted by cyclic hypoxia differs from those observed under normoxia and even continuous hypoxia. PTGS2 is one gene exquisitely up-regulated in endothelial cells (and tumor cells) in response to cyclic hypoxia. Elevated COX-2 (the PTGS2 gene product) expression and activity account for cyclic hypoxia-driven increase in endothelial cell survival and angiogenesis.

overall_design: In vitro culture of HUVEC submitted to hypoxia (oxygen rate less than 0.5%)

first: Géraldine

last: Daneau

first: Romain

last: Boidot

first: Philippe

last: Martinive


title: Effect of BMPR2 mutation on vascular smooth muscle gene expression

status: Public on Aug 25 2010

city: Nashville

first: James

last: West

state: TN

address: 4200 E 9TH Ave, Box B133

department: Medicine

institute: Vanderbilt University

summary: BMPR2 mutation is the cause of most hereditary pulmonary arterial hypertension, but the common molecular consequence of different types of BMPR2 mutation is still not known. The goal of this study was to determine the common molecular consequences of three different classes of patient-derived BMPR2 mutation in vascular smooth muscle gene expression.

overall_design: Three different classes of BMPR2 mutation, wild-type BMPR2, or empty vector were stably transfected into A7R5 vascular smooth muscle cells, and expression compared.

first: Kirk

last: Lane

first: Tom

last: Blackwell


title: Genetic Determinants for Susceptibility to Staphylococcus aureus Infection in A/J and C57BL/6J

status: Public on Aug 01 2010

city: Durham

first: Lindsay

last: Cowell

state: NC

address: 2424 Erwin Road

institute: Duke University

summary: Although it has recently been shown that A/J mice are highly susceptible to Staphylococcus aureus sepsis as compared to C57BL/6J, the specific genes responsible for this differential phenotype are unknown. Using chromosome substitution strains (CSS), we found that factors on chromosomes (chr) 8, 11, and 18 are responsible for susceptibility to S. aureus sepsis in A/J mice. F1 mice from C57BL/6J X CSS8 cross (C8A) and C57BL/6J X CSS18 (C18A) were also susceptible to S. aureus (median survival < 48 h), whereas F1 mice from C57BL/6J X CSS11 cross (C11A) were resistant (median survival > 120 h) to S. aureus. Bacterial loads in the kidney were consistent with F1 median survivals, with higher bacterial counts in susceptible mice. No sexlinked  associations with susceptibility were noted in F1 intercrosses. Using whole genome transcription profiling, we identified a total of 192 genes on chromosomes 8, 11, and 18 which are differentially expressed between A/J and C57BL/6J in the setting of S. aureus infection. Of these, 28 genes had Gene Ontology annotations indicating a potential immune response function. These 28 genes are associated with susceptibility to S. aureus in A/J mice, and are potential determinants of susceptibility to S. aureus infection in humans.

overall_design: To identify genes for which differential expression between A/J and C57BL/6J mice could contribute to host susceptibility to S. aureus infection, we compared the gene expression profiles between uninfected A/J and C57BL/6J mice and between infected A/J and C57BL/6J mice at 2, 4, 6, and 12 hours after infection.

first: SunHee

last: Ahn

middle: G


title: NSCLC metastasis: K-ras/p53 mutant and syngeneic mouse models

status: Public on Nov 10 2009

city: Houston

first: Chad

last: Creighton

state: TX

address: One Baylor Plaza, Mail Stop: BCM305

department: Biostatistics, Ducan Cancer Center

institute: Baylor College of Medicine


title: Over expression of mRNA for multiple genes including insulin in the PLN of NOD is associated with Islet Autoimmunity

status: Public on Oct 02 2009

city: Paris

first: Evie

last: Melanitou

address: 25 rue du Dr Roux

department: Parasitology & Mycology

institute: Institut Pasteur

summary:
The aim of this study is to identify genes implicated in the early steps of the autoimmune process, prior to inflammation in type 1 diabetes. Early Insulin AutoAntibodies (E-IAA) have been used as subphenotypic marker to select individual animals as type 1 diabetes prone and to compare gene expression patterns with insulin autoantibody negative NOD.
Variation of gene expression patterns in the pancreatic lymph nodes (PLN) issued from E-IAA positive and negative mice have been analyzed by DNA microarrays

overall_design: PLN were used from 5 weeks old NOD mice positive for Insulin Autoantibodies and compared with animals negative for Insulin AutoAntibodies

first: Béatrice

last: Regnault

first: José

last: Osorio y Fortea

first: Dongmei

last: Miao

first: George

last: Eisenbarth


title: A compendium of myeloma associated chromosomal copy number abnormalities and their prognostic value

status: Public on Oct 18 2010

city: Sutton

first: Brian

last: Walker

address: 15 Cotswold Road

department: Haemato-Oncology Research Unit

institute: The Institute of Cancer Research

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

summary: To obtain a comprehensive genomic profile of presenting multiple myeloma cases we performed high resolution single nucleotide polymorphism (SNP) mapping array analysis in 114 samples alongside 258 samples analysed by U133 Plus 2.0 expression array (Affymetrix).  We examined DNA copy number alterations and loss of heterozygosity (LOH) in order to define the spectrum of minimally deleted regions in which relevant genes of interest can be found.  The most frequent deletions are located at 1p (30%), 6q (33%), 8q (25%), 12 (22%), 13q (59%), 14q (39%), 16q (35%), 17p (7%), 20 (12%) and 22 (18%).  In addition, copy number-neutral LOH, or uniparental disomy, was also prevalent on 1q (8%), 16q (9%), and X (20%), and was associated with regions of gain and loss.  Based on fluorescent in situ hybridisation (FISH) and expression quartile analysis, genes of prognostic importance were found to be located at 1p (FAF1, CDKN2C), 1q (ANP32E), and 17p (TP53).  In addition, we identified common homozygously deleted genes which have functions relevant to myeloma biology.  Taken together, the dysregulated genes from the myeloma genome indicate that the crucial pathways in myeloma pathogenesis include the NF-?B pathway, apoptosis, cell-cycle regulation and Wnt signalling.

overall_design:
SNP data: 114 tumour samples (MM) analyzed by Affymetrix 500K array set (Nsp+Sty), of which 80 samples have matched peripheral blood (PB) (non-tumour) DNA performed on the same array types.  Matched tumour and non-tumour samples have the same ID number, e.g. MM400 and PB400

Expression data: 258 expression samples from CD138+ cell selection

first: Gareth

middle: J

last: Morgan


title: Genome-wide analysis of skin biopsies during the mucocutaneous HSV-2 reactivation

status: Public on Oct 15 2009

city: Seattle

first: Tao

last: Peng

state: WA

address: 1100 Fairview Ave N/FHCRC/E5-110

institute: University of Washington

summary: Analysis of host response at the gene expression level during the genital HSV-2 reactivation. We would like to get a comprehensive overview on host gene expression and correlate them with the infiltrated immune cell types during HSV-2 reactivation. The hypothesis is that HSV-2 may evade some aspects of host immune response during the recurrent infection.

overall_design: Total RNA obtained from lesion and post healing skin biopsies as compared to contralateral control skin biopsies


title: Spatial mapping of thymic stromal microenvironments reveals unique features influencing T lymphoid differentiation

status: Public on Nov 24 2009

city: jupiter

first: howard

middle: t

last: petrie

state: FL

address: 130 scripps way

department: cancer biology

institute: the scripps research instititute

summary: Interaction of hematopoietic progenitors with the thymic stromal microenvironment induces them to proliferate, adopt the T cell fate, and asymmetrically diverge into multiple T lineages. Progenitors at various developmental stages are stratified among different regions of the thymus, implying that the corresponding microenvironments differ from one another, and provide unique sets of signals to progenitors migrating between them. The nature of these differences remains undefined. Here we use novel physical and computational approaches to characterize these stromal subregions, distinguishing gene expression in microdissected tissues from that of their lymphoid constituents. Using this approach, we comprehensively map gene expression in functionally distinct stromal microenvironments, and identify clusters of genes that define each region. Quite unexpectedly, we find that the central cortex lacks distinctive features of its own, and instead appears to function by sequestering unique microenvironments found at the cortical extremities, and modulating the relative proximity of progenitors moving between them.

overall_design: 4 to 6 weeks old male C57bl6/J were used for microdissection of 3 thymic cortical subregions and thymic medulla or for sorting cortical and medullary thymocytes.  These samples were used for subsequent RNA purification, labeling and hybridization to Affymetrix arrays

first: Howard

middle: T

last: Petrie

first: Ann

middle: V

last: Griffith

first: Mohammad

last: Fallahi

first: Hiroshi

last: Nakase


title: Divergence of macrophage phagocytic and antimicrobial programs in leprosy

status: Public on Oct 21 2009

city: Los Angeles

first: Rosane

last: Teles

address: 10833 Le Conte Ave, CHS 52-121

institute: UCLA

summary: Effective innate immunity against many microbial pathogens requires macrophage programs that upregulate phagocytosis and direct antimicrobial pathways, two functions generally assumed to be coordinately regulated.  Here the regulation of these key functions was investigated in human blood-derived macrophages.  IL-10 induced the phagocytic pathway, including CD209 and scavenger receptors, resulting in phagocytosis of mycobacteria and oxLDL.  IL-15 induced the vitamin D-dependent antimicrobial pathway and CD209, yet the cells were less phagocytic.  The differential regulation of macrophage functional programs was confirmed by analysis of the spectrum of leprosy lesions: the macrophage phagocytosis pathway was prominent in the clinically progressive, multibacillary form, whereas the vitamin D-dependent antimicrobial pathway predominated in the self-limited form of the disease and in patients undergoing reversal reactions from the multibacillary to the self-limited form.  These data indicate that macrophage programs for phagocytosis and antimicrobial responses are distinct and differentially regulated in innate immunity in bacterial infections.

overall_design: 7 LL lesions, 10 BT lesions, 7 RR lesions

first: Dennis

last: Montoya

first: Daniel

last: Cruz

middle: M

first: Delphine

last: Lee

first: Maria

last: Ochoa

first: Stephan

last: Krutzik

first: Rene

last: Chun

first: Mirjam

last: Schenk

first: Xiaoran

last: Zhang

first: Benjamin

last: Ferguson

first: Anne

last: Burdick

first: Euzenir

middle: N

last: Sarno

first: Thomas

middle: H

last: Rea

first: Martin

last: Hewison

first: John

middle: S

last: Adams

first: Genhong

last: Cheng

first: Robert

last: Modlin


title: Gene expression profiles of 344SQ lung adenocarcinoma cells with high metastatic potential (syngeneic mouse model)

first: Chad

last: Creighton

summary:
The biologic basis for NSCLC metastasis is not well understood. Here we addressed this deficiency by transcriptionally profiling tumors from a genetic mouse model of human lung adenocarcinoma that develops metastatic disease owing to the expression of K-rasG12D and p53R172H. As a tool to investigate the biologic basis for metastasis in this model and to query the roles of specific genes in this signature, we isolated adenocarcinoma cell lines from these mice and used them to develop a syngeneic tumor model in wild-type littermates. Transcriptional profiling of the highly metastatic subcutaneous tumors revealed genes that regulate, among other processes, epithelial-to-mesenchymal transition and intra-tumoral inflammation and angiogenesis, whereas the non-metastatic tumors did not.

Keywords: two group comparison

overall_design:
Cell lines from p53R172H∆g/+ K-rasLA1/+ mice were derived from tumor tissues removed at autopsy from two different mice (#344 and #393). The tissues were minced, placed in culture, and passed serially in RPMI 1640 supplemented with 10% fetal bovine serum (FBS), which yielded mass populations of tumor cells derived from primary lung tumors (344P and 393P), mediastinal lymph nodes (344LN and 393LN), and a subcutaneous site (344SQ).  Syngeneic tumors were isolated, carefully dissected to remove the adjacent tissue, snap-frozen in liquid nitrogen and stored at -80° until use. Part of each dissected tumor was histologically evaluated by a board-certified pathologist. Snap-frozen samples were processed and analyzed on Affymetrix Mouse Expression Array 430A 2.0 chips.

Expression profiling performed on 344SQ, 393P, and 393LN

first: Jon

last: Kurie

first: Dror

last: Berel

first: Don

last: Gibbons

first: Wei

last: Lin


title: Effects of glucose transporter expression on VSMC

status: Public on Oct 23 2009

city: Ann Arbor

first: Eileen

middle: Danger

last: Vesely

state: MI

address: 1150 W. Medical Center Drive MSRB I. Room 5518

department: Nephrology

institute: University of Michigan

summary: Hypothesis:  Overexpression of the GLUT1 facilitative glucose transporter, in A7r5 vascular smooth muscle cells, is sufficient and/or necessary to induce alterations in gene expression which influence apoptosis, growth, and proliferation.

overall_design:
Scientific Approach:  A7r5 rat embryonic vascular smooth muscle cells (VSMCs) (ATCC, CRL-1444) were infected for 72 hours with adenoviral vectors containing human GLUT1 or empty vector controls at a multiplicity of infection of 10 (MOI of 1 = 100 particles/cell) and grown in both high glucose (25mM) and low glucose (5.5mM) media.  The human GLUT1 cDNA adenoviral vector and empty vector controls were a gift from Dr Arno Kumagai (University of Michigan).  Total RNA was isolated using TRIzol (Invitrogen Life Technologies, Carlsbad, CA), followed by extraction with phenol/chloroform until the interface was clear.  Total RNA was further purified using Qiagen RNeasy Mini Kit (Qiagen 74106).  Immunoblot analysis was conducted to confirm GLUT1 overexpression at the time of Total RNA isolation (72 hours post-infection) for each condition.
Number of chips:  Total RNA was isolated from each of 2 conditions (control cells in low glucose (5.5mM), and GLUT1 overexpressing cells in low glucose), on three separate days and pooled to obtain one sample for each condition.  In total we submitted two samples.

first: Frank

middle: C

last: Brosius

middle: D


title: Gene expression down-regulation in prostate tumor-associated stromal cells involves organ-specific genes

status: Public on Sep 09 2009

city: Ribeirao Preto

first: Ricardo

last: Vêncio

state: SP

address: Av. Bandeirantes, 3900

department: Computing and Mathematics Department

institute: Universidade de São Paulo

summary: The prostate stroma is a key mediator of epithelial differentiation and development, and potentially plays a role in the initiation and progression of prostate cancer. Isolation and characterization of viable populations of the constituent cell types of prostate tumors could provide valuable insight into the biology of cancer. The CD90+ stromal fibromuscular cells from tumor specimens were isolated by cell-sorting and analyzed by DNA microarray. Dataset analysis was used to compare gene expression between normal and tumor-associated reactive stromal cells. Reactive stroma is characterized by smooth muscle differentiation, prostate down-regulation of SPOCK3, MSMB, CXCL13, and PAGE4, bladder down-regulation of TRPA1, HSD17B2, IL24, and SALL1, and an up-regulation of CXC-chemokines. This study identified a group of differentially expressed genes in CD90+ reactive stromal cells that are potentially involved in organ development and smooth muscle cell differentiation.

overall_design:
A total of 15 arrays were run for the following sample types obtained from 10 patients:

2 CD90+ prostate tumor-associated stromal:
Patient 1: CP_Str_08-028_CD90posi
Patient 2: 08-032_CP_strom_CD90posi

2 CD13+ normal bladder stromal:
Patient 3: 06-125_NB_CD13posi
Patient 4: 06-070_NB_str_CD13posi

1 CD13+ bladder tumor-associated stromal:
Patient 5: 07-008_CB_str_CD13posi

5 whole tissue prostate cancer and 5 normal tissue from matched pairs:
Patient 6: 05-206_CaP, 05-206_NP
Patient 7: 05-213_CaP, 05-213_NP
Patient 8: 05-214_CaP, 05-214_NP
Patient 9: 05-218_CaP, 05-218_NP
Patient 10: 05-220_CaP, 05-220_NP

Additionally, 8 arrays were run for the following sample types obtained from 7 patients:

5 CD49a+ normal prostate stromal (PMID 16638148):
Patient 1: CD49a_01-26-04
Patient 2: CD49a_03-23-04
Patient 3: CD49a_03-04-04
Patient 4: CD49a-1_06-02-04
Patient 5: CD49a-4_06-02-04

3 CD26+ prostate cancer (2 biological replicates, 1 sample run twice):
Patient 6: 05-179_CD26t
Patient 6: 05-179_CD26t_2
Patient 7: 08-032_CP_epi_CD26posi

The following two prostate cancer samples were also included in the analyses:

CD26+ cancer cell, replicate 1
CD26+ cancer cell, replicate 2

The tissue samples consisted of prostate tissue specimens obtained from patients undergoing radical prostatectomy under approval by the University of Washington Institutional Review Board. The same approach was used for both cancer-free and cancer-enriched (where at least 85% of the cells in the corresponding frozen section were of cancer) samples. To obtain bladder stromal cells for analysis, tissue specimens were obtained from cystoprostatectomy surgeries. For cell sorting, the collected specimens were processed within hours. Cell types were sorted using monoclonal antibodies specific for tumor-associated prostate stromal cells (CD90), tumor-associated bladder stromal cells (CD13) and normal bladder stromal cells (CD13) with MACS.


title: WAGO pathway

status: Public on Oct 01 2009

city: Worcester

first: Craig

last: Mello

address: 368 Plantatoin Street, Suite AS5-2047

department: Program in Molecular Medicine

institute: University of Massachusetts Medical School

type: Non-coding RNA profiling by genome tiling array

type: Non-coding RNA profiling by high throughput sequencing


title: Expression Data from Ovarian Surface Kinome

status: Public on Apr 07 2009

city: Portland

first: Solange

last: Mongoue

state: OR

address: 3181 SW Sam Jackson Park Road, CR-145

department: Cancer Institute

institute: Oregon Health & Science University

summary:
The human kinome is incolved in multiple function in the life cycle of cells, and ther differntial expression in cacner suggests that protein kinases play an important role in tumor progression and proliferation.
To delineate pathways that may be important for neoplastic change in women at high risk for ovarian cancer, we compared the expression signature of surface kinases in normal ovarian surface epithelium with ovarian epithelium from patients at high risk, and epithelial ovarian cancer using Affymetrix expresion array HG U133Plus2.

overall_design:
A total of 18 ovarian samples were collected. The number of samples and subjects for each group were: 5 cancer samples (5 subjects), 6 normal samples (4 patients) and 7 high-risk samples (5 subjects).
After the initial inspection, one cancer sample (Sample 9) was excluded from the statistical analysis due to a poor RNA quality.

first: Tanja

last: Pejovic


title: 1Day Post Amputation Vehicle or Beclomethasone Exposed

status: Public on Jan 31 2009

city: Corvallis

last: Tanguay

department: EMT

institute: Oregon State University

summary:
Zebrafish caudal fin regeneration is an established model to study tissue regeneration. In order to identify novel molecular signaling pathways critical for regeneration, we developed a rapid throughput in vivo regeneration assay. We screened a 2000 member structurally diverse small molecule library, followed by assessment of regenerative progression at three days post amputation. A cluster of glucocorticoids was identified among the “positive hits”. To identify the molecular targets of the activated glucocorticoid receptor, microarray analysis was performed using RNA isolated from the regenerates of control and glucocorticoid exposed zebrafish. We identified 673 transcripts that were differentially regulated. The level of expression and spatial expression pattern of select genes were completed by qPCR and by in situ hybridization, respectively. Altogether, these studies demonstrate the power of chemical genetics to identify chemical probes and their targets which will provide a path towards defining conserved regenerative pathways.
Keywords: genomics

overall_design: The caudal fin of zebrafish larvae at 2days post fertilization were amputated and exposed to vehicle control alone or Beclomethasone . Regenerating fins were isolated at 1days post amputation. Three replicates were collected at each time point. 150 fins were pooled to comprise one replicate.

first: Sumitra

last: Sengupta

first: Lijoy

middle: K

last: Mathew

first: Jane

last: LaDu


title: Comparative transcriptional profiling of rice undergoing infection by X. oryzae pv. oryzae or by X. oryzae pv. oryzicola

status: Public on Jun 25 2009

city: Ames

first: PLEXdb Curator

middle: Sudhansu

state: IA

address: 1023  Crop Genomes Informatics Lab (Iowa State University)

department: Virtual Reality Application Center

institute: Iowa State University

summary: RESEARCH SUMMARY     Bacterial blight and bacterial leaf streak, two of the most serious diseases of rice, are caused by Xanthomonas oryzae pathovar (pv.) oryzae (Xoo) and X. oryzae pv. oryzicola (Xoc) respectively. Xoo is a vascular pathogen that invades the xylem, whereas Xoc is non-vascular and colonizes the mesophyll apoplast. These distinctive types of infection represent the two most common ways plant pathogenic bacteria produce disease. As closely related pathogens of a model plant host, Xoo and Xoc constitute a powerful comparative system. We hypothesize that tissue-specific susceptibility to pathogenic bacteria can be explained partially by differential host responses during infection. To establish the global structure of susceptible responses to infection by Xoo and Xoc, we used the Affymetrix Rice Genome Array to analyze the abundance of transcripts in rice plants over four days after inoculation with virulent strains.      MATERIAL AND METHODS    - Plant Material: Oryza sativa ssp. japonica cv. Nipponbare plants were grown in pots containing LC-1 soil mixture, and arranged in flats of 20 pots, in growth chambers under a 12-h, 28ºC light and a 12-h, 25ºC dark cycle. To allow inversion of the flats for inoculation (modified dip-inoculation method, Niño-Liu et al, 2005, Journal of Phytopathology) seeds were sown through 1-cm perforations in a fibreglass tray resting on top of the pots. Peters Professional Fertilizer and iron chelate micronutrient were applied with watering every two days at rates of 0.25 and 4.5 g/L, respectively, until the day before inoculation.    - Bacterial Strains and preparation of inoculum: Xoo strain PXO99A and Xoc strain BLS256 were used. For each, a single colony from a glucose yeast extract (GYE) agar plate was transferred to 5 ml of GYE liquid medium and incubated for 24 h at 28ºC with constant shaking at 250 rpm. Subsequently, 2 ml of this culture was transferred to 300 ml of fresh GYE liquid medium and incubated as above for an additional 18 h. Cells were pelleted by centrifugation at 4000 rpm. for 10 min, washed twice and resuspended in sterile 10 mm MgCl2 to an OD600 of 0.05, yielding 7 liters of inoculum. Tween-20 was added to a final concentration of 0.5%.    - Modified dip-inoculation method: Fourteen days after sowing, and 2 h after the beginning of the light period, seedlings were inverted into inoculum, one flat each for Xoo, Xoc and Mock inoculum consisting of sterile 10 mM MgCl2 with 0.5% Tween-20, and vaccuum infiltrated at 500mm Hg for two minutes, two consecutive times, in a custom-made chamber. Following inoculation, plants were returned to the growth chamber. During growth, plants were cycled weekly in the same order through three growth chambers. In this way all plants could be incubated in the same final chamber following inoculation. For each replicate, position of the flats in the growth chamber (from left to right, used also for the order of inoculation and harvest) was determined by a split-plot design.       - Timepoints, tissue collection: Within flats, assignment of pots for the harvest of inoculated tissue at 2, 4, 8, 24 and 96 h was made using a separate randomized complete block design. Tissue was harvested from plants in four pots (approximately 2g fresh weight) for each timepoint per inoculum per replication. Harvested tissue was immediately frozen in liquid nitrogen and stored at -80ºC until processing.    - RNA isolation, purification, labeling, hybridizations, scanning: Total RNA was initially isolated from frozen whole plant tissue using the hot TRIzol reagent protocol for barley as described by Caldo et al (Plant Cell, 2004). The RNA was purified using Qiagen RNeasy spin columns, and adjusted to a final concentration of 1 ug/uL. Probe synthesis, labeling, and hybridization were performed at the Iowa State University GeneChip Core facility.    ACKNOWLEDGEMENTS    This work was supported by a grant (0227357) from the Plant Genome Research Program of the National Science Foundation.   ****[PLEXdb(http://www.plexdb.org) has submitted this series at GEO on behalf of the original contributor,  David O Niño-Liu.  The equivalent experiment is OS3 at PLEXdb.]

overall_design:
XOC - time: 2 hai (4-replications)
XOC - time: 4 hai (4-replications)
XOC - time: 8 hai (4-replications)
XOC - time: 24 hai (4-replications)
XOC - time: 96 hai (4-replications)
XOO - time: 2 hai (4-replications)
XOO - time: 4 hai (4-replications)
XOO - time: 8 hai (4-replications)
XOO - time: 24 hai (4-replications)
XOO - time: 96 hai (4-replications)
MOCK - time: 2 hai (4-replications)
MOCK - time: 4 hai (4-replications)
MOCK - time: 8 hai (4-replications)
MOCK - time: 24 hai (4-replications)
MOCK - time: 96 hai (4-replications)

first: David

middle: O

last: Niño-Liu


title: p53 Regulates Hematopoietic Stem Cell Quiescence

status: Public on Nov 07 2008

city: New York

first: Agnes

last: Viale

state: NY

address: 1275 York Avenue

department: Molecular Biology

institute: Memorial Sloan Kettering Cancer Center

summary: The importance of the p53 protein in the cellular response to DNA damage is well known, but its function during steady-state hematopoiesis has not been established. We have defined a critical role of p53 in regulating hematopoietic stem cell quiescence, especially in promoting the enhanced quiescence seen in HSCs that lack the MEF/ELF4 transcription factor. Transcription profiling of HSCs isolated from wild type and p53 null mice identified Gfi-1 and Necdin as p53 target genes and using lentiviral vectors to upregulate or knockdown the expression of these genes, we show their importance in regulating HSC quiescence. Establishing the role of p53 (and its target genes) in controlling the cell cycle entry of HSCs may lead to therapeutic strategies capable of eliminating quiescent cancer (stem) cells.

overall_design: RNAs isolated from wild type, p53 -/- and p53 -/- Mef -/-  LSK cells were used in oligonucleotide arrays (Affymetrix)

first: Yan

last: Liu

first: Steven

last: Nimer


title: High throughput sequencing of mutants in the WAGO pathway

summary: Diverse naturally-occurring small RNA species interact with Argonaute proteins to mediate sequence-specific regulation in animals. In addition to micro-RNAs (miRNAs), which collectively regulate thousands of target mRNAs, other endogenous small RNA species include the Piwi-associated piRNAs that are important for fertility and a less well-characterized class of small RNAs often referred to simply as endo-siRNAs. Here we have utilized deep-sequencing technology and C. elegans genetics to explore the biogenesis and function of endo-siRNAs. We describe conditional alleles of the dicer-related helicase, drh-3, that implicate DRH-3 in both the response to foreign dsRNA as well as the RNA-dependent RNA Polymerase (RdRP)-dependent biogenesis of a diverse class of endogenous small RNAs, termed 22G-RNAs. We show that 22G-RNAs are abundantly expressed in the germline and maternally inherited and are the products of at least two distinct 22G-RNA systems. One system is dependent on worm-specific AGOs, including WAGO-1, which localizes to germline nuage-related structures termed P-granules. The WAGO 22G-RNA system silences transposons, pseudogenes and cryptic loci as well as a number of genes. Finally, we demonstrate that components of the nonsense-mediated decay pathway function in at least one of the multiple, distinct WAGO surveillance pathways. These findings broaden our understanding of the biogenesis and diversity of 22G-RNA species and suggest potential novel regulatory functions for these small RNAs.

overall_design: 18 samples examined. Small RNA libraries generated from: C. elegans animals with mutations in the WAGO pathway and a WAGO-1 immunopercipitate.

first: Weifeng

last: Gu


title: Effects of PMN-Ectosomes on human macrophages

status: Public on Jan 16 2009

city: Basel

first: Ceylan

last: Eken

address: Hebelstrasse 20

department: Biomedicine Department

country: Switzerland

institute: University Hospital Basel

summary: In order to define the inhibitory activity of PMN-Ectosomes, we investigated the early gene expression profiles of resting and zymosan A-stimulated human monocyte-derived macrophages in the absence or presence of PMN-Ectosomes.

overall_design: We cultured monocytes isolated from fresh buffy coats of 4 healthy donors (2 males and 2 females between the ages of 20 to 45) for 7 days at 37°C. At day 7, macrophages were cultured for another 3 hours alone, in presence of PMN-Ectosomes, with zymosan A, and with zymosan A and PMN-Ectosomes.

first: Jürg

middle: A

last: Schifferli


title: Validation Set for Prediction of Outcome in Adult Male Germ Cell Tumors

status: Public on Sep 30 2008

first: James

last: Korkola

address: 1275 York Ave

department: Cell Biology

institute: Memorial Sloan-Kettering

summary:
This series represents expression profiles of 34 non-seminoma germ cell tumors (NSGCTs) from patients who received cisplatin based chemotherarpy for treatment of their disease for whom full clinical follow-up information was available. These specimens were used as a validation set to test outcome prediction models using a subset of previously profiled GCT specimens (see GEO accession #GSE3218).
Keywords: Disease state analysis (good vs. poor patient outcome)

overall_design: Tumor tissues were collected under IRB-approved protocols at the Memorial Sloan-Kettering Cancer Center, New York, between 1987 and 2003. The tumor samples consist of 34 NSGCT specimens. RNA was isolated, labeled, and hybridized to Affymetrix U133A and U133B microarrays using standard protocols. Data were processed by the RMA method.

middle: E

last: Houldsworth

first: Darren

last: Feldman

first: Victor

last: Reuter

first: George

last: Bosl

last: Chaganti


title: Gene expression analysis of macrophages from ankylosing spondylitis patients reveals interferon-gamma dysregulation

status: Public on Jun 27 2008

city: Cincinnati

first: Michael

last: Barnes

state: OH

address: 3333 Burnet Ave

department: Rheumatology

institute: Cincinnati Childrens Hospital Medical Center

summary:
OBJECTIVE: To determine whether macrophages, a type of cell implicated in the pathogenesis of ankylosing spondylitis (AS), exhibit a characteristic gene expression pattern. METHODS: Macrophages were derived from the peripheral blood of 8 AS patients (median disease duration 13 years [range <1-43 years]) and 9 healthy control subjects over 7 days with the use of granulocyte-macrophage colony-stimulating factor. Cells were stimulated for 24 hours with interferon-gamma (IFNgamma; 100 units/ml), were left untreated for 24 hours, or were treated for 3 hours with lipopolysaccharide (LPS; 10 ng/ml). RNA was isolated and examined by microarray and real-time quantitative reverse transcription-polymerase chain reaction analysis. RESULTS: Microarray analysis revealed 198 probe sets detecting the differential expression of 141 unique genes in untreated macrophages from AS patients compared with healthy controls. Clustering and principal components analysis clearly distinguished AS patients and controls. Of the differentially expressed genes, 78 (55%) were IFN-regulated, and their relative expression indicated a reverse IFN signature in AS patient macrophages, where IFNgamma-up-regulated genes were underexpressed and down-regulated genes were overexpressed. Treatment of macrophages with exogenous IFNgamma normalized the expression of these genes between patients and controls. In addition, the messenger RNA encoded by the IFNgamma gene was approximately 2-fold lower in AS patient macrophages at baseline (P = 0.004) and was poorly responsive to LPS (P = 0.018), as compared with healthy controls. CONCLUSIONS: Our findings reveal consistent differences in gene expression in macrophages from AS patients, with evidence of a striking reverse IFN signature. Together with poor expression and responsiveness of the IFNgamma gene, these results suggest that there may be a relative defect in IFNgamma gene regulation, with autocrine consequences and implications for disease pathogenesis.

Keywords: Disease state analysis with treatment effect

overall_design: Macrophages were derived from the peripheral blood of 8 AS patients (median disease duration 13 years [range <1–43 years]) and 9 healthy control subjects over 7 days with the use of granulocyte– macrophage colony-stimulating factor. Cells were stimulated for 24 hours with interferon- (IFN; 100 units/ ml), were left untreated for 24 hours, or were treated for 3 hours with lipopolysaccharide (LPS; 10 ng/ml). RNA was isolated and examined by microarray and real-time quantitative reverse transcription–polymerase chain reaction analysis.

first: Judy

last: Smith

middle: G

first: Dihua

last: Hong

first: Monica

middle: L

last: Delay

first: Robert

last: Inman

last: Colbert


title: Identification of Signature Molecule-Marked Native Mesenchymal Stem Cells

status: Public on Oct 28 2008

city: Hiroshima

first: Masakazu

last: SHIMIZU

state: Hiroshima

address: 1-2-3 Kasumi-cho, Minami-ku

department: Departments of Dental and Medical Biochemistry

country: Japan

institute: Hiroshima University

summary:
Background. The in vivo distribution status and molecular signature of bone marrow mesenchymal stem cells (MSC) remain unknown, although ex vivo expanded MSC have been used in numerous studies.
Methods/Results. To address these issues, we identified human MSC-characteristic genes, including 9 transcription factor genes - using DNA microarray analysis: Most (75 %) of the MSC-characteristic genes were down-regulated by knockdown of the transcription factors. Furthermore, we used an antibody set for the MSC-characteristic molecules in immunohistochemistry, which revealed that, in embryonic/neonatal mice, elongated MSC were abundant near the endosteum and in the interior of bone marrow, perichondrium and periosteum, whereas in adult mice round/compact MSC were found only in the interior of bone marrow: No MSC-like cells were found in any other skeletal or non-skeletal tissues using this antibody set.
Conclusion. The identification of native MSC in developing and adult skeletal tissues should be useful in basic and clinical studies on MSC.
Keywords: cell type comparison

overall_design:
Cell Cultures
Human iliac MSC were purchased from BioWhittaker Inc (Walkersville, MD) or obtained from iliac crest according to protocol approved by ethical authorities at Hiroshima University. Human skin fibroblasts were purchased from KURABO (Osaka, Japan), and human gingival fibroblasts were isolated as described previously (Kawahara K, Shimazu A (2003) Expression and intracellular localization of progesterone receptors in cultured human gingival fibroblasts. J Periodontal Res 38: 242-246.). Culture of MSC and their differentiation from the osteoblast, chondrocyte or adipocyte lineage were performed as described (Matsubara T, Suardita K, Ishii M, Sugiyama M, Igarashi A, et al. (2005) Alveolar bone marrow as a cell source for regenerative medicine: differences between alveolar and iliac bone marrow stromal cells. J Bone Miner Res 20: 399-409.) : We used FGF-2 at 1 ng/mL to expand MSC, because MSC expanded ex vivo with FGF-2 maintained their multidifferentiation potential throughout many mitotic divisions (Tsutsumi S, Shimazu A, Miyazaki K, Pan H, Koike C, et al. (2001) Retention of multilineage differentiation potential of mesenchymal cells during proliferation in response to FGF. Biochem Biophys Res Commun 288: 413-419.). To avoid direct action of FGF-2 on gene expression, FGF-2 was removed from the culture medium of MSC or fibroblasts 72 h before isolation of RNA.

DNA Microarray Analysis
Total RNA was isolated from confluent cultures of 3 lines of MSC (passage 6-9), 3 lines of fibroblasts (passage 7-14), 3 lines of MSC incubated for 28 days in the differentiation-induction medium, using RNeasy Mini Kit (QIAGEN, Chatsworth, CA). DNA microarray analysis was performed by KURABO GeneChip Custom Analysis Service with Human Genome U133 Plus 2.0 chips(Affymetrix. Inc., Santa Clara, CA). The CHP data (Microarray Suite version 5.0, Scaling Factor = 1, Normalization Value = 1, Detection Call a1 = 0.05, a2 = 0.065, Tau = 0.015, Affymetrix Inc.) were standardized by the global median normalization method using GeneSpring (Silicon Genetics, Redwood City, CA). The normalization was limited by flag values, and the median was calculated using the genes that exceeded the Present or Marginal flag restriction. Gene filtering on normalized intensity followed by fold changes (>2-fold) was used to generate the list of genes for expression profiles before or after differentiation.

last: Shimizu

first: Hiroshi

last: Kubo

first: Yuji

last: Taya

first: Takeshi

last: Kawamoto

first: Emi

last: Kaneko

first: Akira

last: Igarashi

first: Masahiro

last: Nishimura

first: Kazumi

last: Segoshi

first: Yoshihito

last: Shimazu

first: Koichiro

last: Tsuji

first: Takaaki

last: Aoba

first: Yukio

last: Kato


title: Myasthenia gravis model: gene expression profile in diaphragm, extensor digitorum longus and extraocular muscles of rats

status: Public on Oct 01 2008

city: St Louis

first: Bendi

last: Gong

state: MO

address: 1402 S Grand Ave

department: Neurology & Psychiaty

institute: St Louis University

summary:
gene expression profile in diaphragm (DIA), extensor digitorum longus (EDL) and extraocular (EOM) muscles of rats with actively induced experimentally acquired MG (EAMG) using Affymetrix rat RAE230 gene chip.
Keywords: myasthenia gravis (MG), rats with actively induced experimentally acquired MG (EAMG)

overall_design:
Total RNA was obtained with TRIzol (Invitrogen Carlsbad, CA) following the manufacturers recommended protocol. Tissues from 3 animals were combined into each RNA sample to decrease inter-subject variability. RNA pellets were cleaned by RNasey kits and resuspended at 1 μg RNA/μl DEPC-treated water and 5 ug was used in a reverse transcription reaction (SuperScript II; Life Technologies, Rockville, MD) to generate first strand cDNA.  Double strand cDNA was synthesized and used in an in vitro transcription (IVT) reaction to generate biotinylated cRNA.  Fragmented cRNA (15 ug) was used in a 300 ul hybridization cocktail containing herring sperm DNA and BSA as carrier molecules, spiked IVT controls, and buffering agents.  A 200 ul aliquot of this cocktail was used for hybridization to Affymetrix rat REA230 (Santa Clara, CA) microarrays for 16 hrs at 45o C.  The manufacturer’s standard post-hybridization wash, double-stain, and scanning protocols used an Affymetrix GeneChip Fluidics Station 400 and a Hewlett Packard Gene Array scanner.

Microarray data analysis
Raw data from microarray scans were analyzed with Affymetrix GCOS 2.0.  GCOS evaluates sets of perfect match (PM) and mismatch (MM) probe sequences to obtain both hybridization signal values and present/absent calls for each transcript. Microarrays were scaled to the same target intensity and pairwise comparisons were made between experimental and control samples. Comparisons the one-sided Wilcoxon’s signed rank test to estimate “increase/no change/ decrease” difference calls for each pair-wise comparison. Transcripts defined as differentially regulated met the criteria of: (a) consistent increase/decrease call across 7 out of 9 replicate comparisons, based upon Wilcoxon’s signed rank test (algorithm assesses probe pair saturation, calculates a p value and determines increase, decrease, or no change calls) and (b) the average fold difference ≥ 2.0.  Data were visualized as a hierarchical dendrogram generayed on computer (Genespring software,ver 7.2; Silicon Genetics, Redwood city, CA). Annotation was done according to Affymetrix NetAffyx Gene Ontology database.

first: Henry

last: Kaminski


title: Expression profiling of neuroblastic tumors

status: Public on Oct 16 2008

first: Olivier

last: Delattre

address: 26 rue d'Ulm

department: Genetics

institute: Institut Curie

summary: The expression profiles of 64 neuroblastic tumors (mainly neuroblastoma) were determined on Affymetrix chips HG U133 Plus 2.0.

overall_design: 64 neuroblastic tumors


title: Genomic Profiling of Adult Male Germ Cell Tumors

status: Public on Nov 15 2007

first: James

last: Korkola

type: Genome variation profiling by genome tiling array

summary:
Genomic profiling of a panel of 74 adult male germ cell tumors of various histologies from primary and metastatic sites using high resolution BAC array CGH. See Pubmed ID # 17943972.

Completed Study: Statistical analyses were performed to identify regions that were frequently altered in all tumors, as well as in specific histologic subsets and from different tumor sites (i.e., primary gonadal, primary extra-gonadal, and metastatic). Target genes of altered regions were also identified using corresponding tumor expression data (see GEO GSE3218 for expression data).
Ongoing:
1) Analysis of high level amplifications and multiple copy number gains and losses and underlying target genes.
2) Regional alterations associated with patient outcome
Keywords: tumor profiling

overall_design: DNA was isolated from tumor specimens using standard techniques. For the reference sample, karyotypically normal human female placental DNA was used. Samples were labeled with either Cy3 or Cy5 using a Bioprime Labeling Kit, combined with a differentially expressed reference sample, then hybridized to Spectral Genomic 2600 BAC arrays according to manufacturer's recommended conditions; 5 samples done with dye swaps to ensure reproducibility; otherwise one replicate per specimen. Images were captured using an Axon Instruments 4000B scanner and data were processed by median subarray centering followed by a global loess correction. Gains and losses were called using the Circular Binary Segmentation (CBS) algorithm. Associations with histologic subtypes and tumor sites were tested with Fisher's exact test. Genes mapping to regions that were differentially altered were tested for expression differences using Significance Analysis for Microarrays (SAM) by comparing tumors with a specific alteration to those lacking the alteration.

first: Sandy

last: Heck

first: R

last: Chaganti

first: Jane

last: Houldsworth

first: George

middle: J

last: Bosl


title: Expression Data from Macrophage Maturation and Polarization Experiment

status: Public on Jul 10 2006

city: Guildford

first: Fernando

last: Martinez Estrada

state: Surrey

address: Stag Hill

department: Faculty of Health & Medical Sciences

institute: University of Surrey

summary: Monocytes were induced to mature to macrophages with M-CSF. Cells were then activated with  Interferon gamma and LPS or IL-4.

overall_design: Freshly isolated monocytes were cultured in the presence of M-CSF (100 ng/ml) for 7 days, and then activated. The study includes Monocytes at day 0, macrophages at day 3 and 7, Interferon Gamma and LPS treated macrophages and IL-4 treated macrophages.

last: Martinez


title: Effects of TGF-beta on mature macrophages

status: Public on May 15 2008

city: Mannheim

first: Alexei

last: Gratchev

department: Department of Dermatology

institute: Medical Centre Mannheim, University of Heidelberg

summary:
The goal of the study was to identify the effects of TGF-beta on primary human macrophages maturated under different conditions.
Keywords: time course, differentiation conditions, alternative activation

overall_design: For the analysis of TGF-beta effects macrophages were differentiated in the presence of IL-4 or IL-4 in combination with dexamethasone. After 5 days differentiation mature macrophages were stimulated by TGF-beta for 24 hours. In the case of IL-4/dexamethasone stimulation an additional data point was taken. In this case macrophages were also stimulated by TGF-beta for 3 hours. For every group 5 independent donors were taken.

first: Julia

last: Kzhyshkowska


title: HCC Gene Expression Profile

status: Public on Jan 26 2006

city: Bethesda

first: Snorri

last: Thorgeirsson

address: 37 Convent Dr. # 4146

department: Laboratory of Experimental Carcinogenesis

institute: National Cancer institute/NIH

summary:
The variability in the prognosis of hepatocellular carcinoma (HCC) patients suggests that HCC may comprise several distinctive biological phenotypes. These phenotypes may result from different neoplastic pathways during the tumorigenesis and/or from a different cell of origin. Here we address if the transcriptional characteristics of the HCC would provide insight into the cellular origin of the tumors. We integrated gene expression data from rat fetal hepatoblasts and adult hepatocytes, HCC from mouse models, and human HCC. The HCC patients who shared gene expression patterns with fetal hepatoblasts showed extremely poor prognosis when compared with those lacking the hepatoblast signature. The gene expression program that distinguishes this novel subtype from the rest of HCC includes well known markers of hepatic oval cells, suggesting that HCC in this subtype may arise from hepatic progenitor cells. Two independent gene network analyses of the gene expression signature characteristic for the tumors sharing the hepatoblast expression patterns revealed that activation of AP-1 transcription factors might play key roles in tumor development in the newly identified HCC subtype. In addition, by applying hepatoblast-specific and genome-wide global signatures, HCC patients were further stratified into three distinct subgroups with a significant association with overall survival and recurrence.
Keywords: individual genetic characteristic design

overall_design: Total RNAs from 19 normal livers were pooled and used as the reference for all microarray experiments. To obtain gene expression profile data from 49 human HCC, 20 µg of total RNAs from tissues were used to drive fluorescently (Cy-5 or Cy-3) labeled cDNA. At least two hybridizations were carried out for each tissue using a dye-swap strategy to eliminate dye labeling bias.


title: Human Hepatocellular Carcinoma study

status: Public on Oct 28 2004

summary:
Global gene expression patterns of 91 human hepatocellular carcinomas (HCC) were analyzed to define the molecular characteristics of the tumors and to test the prognostic value of the expression profiles. Total RNAs were isolated from frozen liver tissue using CsCl density gradient centrifugation methods. Total RNA from 18 normal livers were pooled and used as reference in entire microarray experiments. To obtain gene expression profile data from human HCC, 20 μg of total RNAs from tissues were used to drive fluorescently (Cy-5 or Cy-3) labeled cDNA. At least two hybridizations were carried out for each tissue using dye-swap strategy to eliminate dye labeling bias. Unsupervised classification methods revealed two distinctive subclasses of HCC that are highly associated with survival of the patients. This association was validated by five independent supervised learning methods. Genes most strongly associated with survival were identified by using the Cox proportional hazards survival analysis.  This approach identified a limited number of genes that accurately predicted the length of survival and provides new molecular insight into the pathogenesis of HCC. Tumors from the low survival subclass have strong cell proliferation and anti-apoptosis gene expression signatures. In addition, the low survival subclass displayed higher expression of genes involved in ubiquitination and histone modification suggesting an etiological involvement of these processes in accelerating the progression of HCC. The biological differences identified in the HCC subclasses should provide an attractive source for the development of therapeutic targets (e.g. HIF1a) for selective treatment(s) of HCC patients.

Keywords: other

first: Ju-Seog

last: Lee

first: I

last: Chu

first: J

last: Heo

first: D

middle: F

last: Calvisi

first: Z

last: Sun

first: T

last: Roskams

first: A

last: Durnez

last: Demetris