ABSTRACT: Growth to mid exponential phase in M9 minimal media supplemented with 0.5% maltose versus growth in rich LB media Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc. Media: M9 minimal media with 0.5% maltose Computed
Project description:ABSTRACT Background. Acute Kawasaki disease (KD) is difficult to distinguish from other acute rash/fever illnesses, in part because the etiologic agent(s) and pathophysiology remain poorly characterized. As a result, diagnosis and critical therapies may be delayed. Methods. We used DNA microarrays to identify possible diagnostic features of KD. We compared gene expression patterns in the blood of 23 children with acute KD and 18 age-matched febrile children with three illnesses that resemble KD. Results. Genes associated with platelet and neutrophil activation were expressed at higher levels in KD patients than in patients with acute adenovirus infections or systemic adverse drug reactions but not in patients with scarlet fever; genes associated with B cell activation were also expressed at higher levels in KD patients than in controls. A striking absence of interferon-stimulated gene expression in the KD patients was confirmed in an independent cohort of KD subjects. We successfully predicted the diagnosis in 21 of 23 KD patients and 7 of 8 adenovirus patients using a set of 38 gene transcripts. Conclusions. These findings provide insight into the molecular features that distinguish KD from other febrile illnesses, and support the feasibility of developing novel diagnostic reagents for KD based on the host response. A disease state experiment design type is where the state of some disease such as infection, pathology, syndrome, etc is studied. Disease State: One of Kawasaki Disease (KD) or control (C) of Scarlet fever (C-sf), adenovirus infection (C-ai) or drug reaction (C-dr) disease_state_design
Project description:Pancreatobiliary cancers have among the highest mortality rates of any cancer type. Discovering the full spectrum of molecular genetic alterations may suggest new avenues for therapy. To catalogue genomic alterations, we carried out array-based genomic profiling of 31 exocrine pancreatic cancers and 6 distal bile duct cancers, expanded as xenografts to enrich the tumor cell fraction. We identified numerous focal DNA amplifications and deletions, including in 19% of pancreatobiliary cases gain at cytoband 18q11.2, a locus uncommonly amplified in other tumor types. The smallest shared amplification at 18q11.2 included GATA6, a transcriptional regulator previously linked to normal pancreas development. When amplified, GATA6 was overexpressed at both the mRNA and protein level, and strong immunostaining was observed in 25 of 54 (46%) primary pancreatic cancers compared to 0 of 33 normal pancreas specimens surveyed. GATA6 expression in xenografts was associated with specific microarray gene-expression patterns, enriched for GATA binding sites and mitochondrial oxidative phosphorylation activity. siRNA mediated knockdown of GATA6 in pancreatic cancer cell lines with amplification led to reduced cell proliferation, cell cycle progression, and colony formation. Our findings indicate that GATA6 amplification and overexpression contribute to the oncogenic phenotypes of pancreatic cancer cells, and implicate GATA6 as a lineage-specific oncogene in pancreatobiliary cancer, with implications for novel treatment strategies. Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc. Tissue type: cancer xenograft from diff tissues Keywords: Logical Set cDNA microarrays from the Stanford Functional Genomics Facility were used to perform gene expression profiling on 28 out of 31 of the exocrine pancreatic cancer and all 6 distal bile duct cancer xenografts. Using regression correlation
Project description:With the goal of identifying changes in gene expression in CD4(+) T cells during the development of diabetes in the nonobese diabetic (NOD) mouse, we used DNA microarrays to analyze gene expression in CD4(+) T cells from the pancreatic draining lymph nodes of NOD/BDC 2.5 T cell receptor transgenic and WT NOD mice at different ages. At 4 and 6 weeks of age, we found up-regulation of a number of genes that are known to be induced by IFN-alpha. IFN-alpha levels and IFN-alpha-producing plasmacytoid dendritic cells were increased in the PLNs of 3- to 4-week-old NOD mice. Moreover, blockade of IFN-alpha receptor 1 in NOD mice by a neutralizing antibody at 2-3 weeks of age significantly delayed the onset and decreased the incidence of type 1 diabetes, increased the relative number of immature dendritic cells in the PLNs, and enhanced the ability of spleen CD4(+) T cells to produce IL-4 and IL-10. These findings demonstrate that IFN-alpha in the PLNs is an essential initiator in the pathogenesis of type 1 diabetes in NOD mice. An all pairs experiment design type is where all labeled extracts are compared to every other labeled extract.
Project description:This SuperSeries is composed of the following subset Series: GSE12166: TJ411(fer-15(b26)II; age-1(hx542)II) vs. BA713(fer-15(b26)II) GSE12167: TJ1081)fer-15(b26)II; daf-16(m26)I) vs. BA713(fer-15(b26)II) GSE12168: Aging Time course Refer to individual Series
Project description:This SuperSeries is composed of the following subset Series: GSE13206: Human shSIRT6 TNF-alpha timecourse GSE13207: Mouse Sirt6-/- TNF-alpha timecourse GSE13208: Mouse Sirt6-/- tissues GSE13209: Mouse Sirt6-/- RelA+/- tissues Refer to individual Series
Project description:Treatment of PBMC with 0.6 pM cytokine from 0-24h Abstract: Background Interferons are key modulators of the immune system, and are central to the control of many diseases. The response of immune cells to stimuli in complex populations is the product of direct and indirect effects, homotypic and heterotypic cell interactions. Dissecting the global transcriptional profiles of immune cell populations may provide insights into this regulatory interplay. The host transcriptional response may also be useful in discriminating between disease states, and in understanding pathophysiology. The transcriptional programs of cell populations in health therefore provide a paradigm for deconvoluting disease-associated gene expression profiles. Results: We used human cDNA microarrays to (1) compare the gene expression programs in human peripheral blood mononuclear cells (PBMCs) elicited by 6 major mediators of the immune response: interferons a, b, w and g, IL12 and TNFa; and (2) characterise the transcriptional responses of purified immune cell populations (CD4+ and CD8+ T cells, B cells, NK cells and monocytes) to IFNg stimulation. We defined a highly stereotyped response to type I interferons, while responses to IFNg and IL12 were largely restricted to a subset of type I interferon-inducible genes. TNFa stimulation resulted in a distinct pattern of gene expression. Cell type-specific transcriptional programs were identified, highlighting the pronounced response of monocytes to IFNg, and emergent properties associated with IFN-mediated activation of mixed cell populations. Conclusions: This information provides a detailed view of cellular activation by immune mediators, and contributes an interpretive framework for the detection and definition of host immune responses in a variety of disease settings. PBMC extraction and stimulation Human primary peripheral blood mononuclear cells (PBMCs) were purified from whole blood of healthy donors using Ficoll-Paque PLUS (GE Healthcare) according to manufacturers instructions. PBMCs were incubated at 1.5-2.0 x 106 cells/well for 24 h before stimulation in RPMI 1640 medium supplemented with 10% heat-inactivated foetal calf serum and 2 mM L-glutamine (Invitrogen) at 37C, 5% CO2. Cells were treated with 0.006 pM, 0.6 pM or 60 pM recombinant IFNg (R&D Systems), and sampled at time intervals from 0.5 h to 12 h after stimulation. Additionally, cells were treated with 0.1 % BSA/PBS alone and used for untreated (mock) control time courses. RNA extraction and amplification Total RNA was extracted in TRIzol LS (Invitrogen) followed by standard chloroform purification and isopropanol precipitation. RNA was resuspended in RNase-free water, quantitated on the NanoDrop ND-1000 spectrophotometer (NanoDrop Technologies) and stored at -80C. 500 ng total RNA was amplified using the MessageAmp modified Eberwine linear amplification procedure (Applied Biosystems). All samples to be compared were extracted and amplified together. Microarray analysis 4 ug amplified RNA was labelled with Cy5-dUTP (GE Healthcare) and combined with 3 ug of Cy3-labelled reference cDNA derived from a pool of RNA derived from a panel of 11 human cell lines (Stratagene Universal Human Reference RNA). The samples were washed and concentrated using MinElute columns (Qiagen) and competitively hybridised to custom printed cDNA microarrays containing 37,632 elements for approximately 18,000 unique human genes. The hybridised slides were scanned using a GenePix 4000A microarray scanner (Axon Instruments). Comparative spot intensities were calculated from the images, and areas of poor quality excluded from further analysis using GenePix Pro 6.0 (Axon Instruments). Data were deposited in the Stanford Microarray Database. Analysis was restricted to cDNA elements with a regression correlation of > 0.6, fluorescence intensities of > 2.5 fold signal/background in Cy3 or Cy5 channels and a minimum signal intensity of > 100 in both channels for at least 80% of the arrays. The expression ratios were normalised for array variation, and the data zero-transformed using a custom-designed Microsoft Excel macro (C. Liu, Stanford). The statistical package SAM (Significance Analysis of Microarrays, version 1.15) was used to identify genes significantly differentially expressed in the normalised data sets by pairwise comparison with a minimum 2 fold cutoff at a false discovery rate of < 1% of the median. The transformed datasets were then hierarchically clustered using Cluster 2.11 and the results displayed using Treeview 1.60. A compound treatment design type is where the response to administration of a compound or chemical (including biological compounds such as hormones) is assayed.
Project description:Throughout the course of early development, the reliable occurrence of stereotyped events is imperative. This careful orchestration is dependent upon regulation at the level of gene expression. While all cell types derive from a single fertilized cell and share a single genome, individual cell types differentially utilize this genome. Such differential utilization is essential for the acquisition, maintenance, and modulation of cell properties. In order to better understand the molecular basis of early development, we evaluated the mRNA expression programs of both early post-implantation mouse embryos and differentiating embryonic stem cells using a cDNA microarray platform covering 74% of the mouse genome. Mining of these datasets revealed a strong association across RNA processing, modulation of the cell cycle, and chromatin organization. Genes involved in these processes demonstrated sharp regulation during well-defined biological transitions. Dominant patterns of expression with biologically separable properties were identified. The evolutionary conservation of one cluster in particular suggested a molecular basis for the alignment of the murine and fly developmental programs. Moreover, the data suggest putative gene relationships that may also have broader implications for gene networks connecting the cell cycle to the molecular repertoire of the cell. Design: Dataset is a murine embryonic developmental timecourse consisting of morphologically staged samples from E6.25 to E9.0 (at approximately 0.25d intervals). There two replicates of each sample and there are 13 samples in each biological replicate series. A common reference design was employed. A development or differentiation experiment design type assays events associated with development or differentiation or moving through a life cycle. Development applies to organism(s) acquiring a mature state, and differentiation applies to cells acquiring specialized functions. Developmental Stage: morphologically staged samples from E6.25 to E9.0 (at approximately 0.25d intervals) Keywords: development_or_differentiation_design Using regression correlation
Project description:In response to polarization cues, cultured Caco-2 cells, a human colon adenocarcinoma-derived cell line, form a polarized epithelium resembling normal enterocytes. We investigated potential signaling mechanisms activated by Caco-2 cells that might trigger the genome-wide transcriptional reprogramming that accompanies polarization (Saaf et al, submitted-I). cDNA microarrays were used to compare the transcriptional profile of Caco-2 polarization to the gene expression profiles of normal human colon and colon tumors. The transcript profile of proliferating, non-polarized Caco-2 cells has striking parallels to the gene expression profile of human colon cancer in vivo. However, as Caco-2 cells develop polarity, the gene expression profile shifts to one more closely resembling that of normal colon tissue, suggesting that the underlying regulatory mechanisms that mediate Caco-2 cell polarization are similar to those that occur during in vivo enterocyte differentiation. We show that transcriptional re-programming of Caco-2 cells during development of cell polarity occurs in the context of signaling pathways that are regulated in a manner that is remarkably similar to those in normal intestinal development. For example, transcriptional targets of the Wnt pathway are tightly regulated during Caco-2 cell polarization, mimicking the gradient of Wnt-mediated transcription in the crypt (high expression) to villus (low expression) axis in human intestine. However, Caco-2 cells lack full-length APC necessary for normal Wnt-regulated degradation of beta-catenin. Biochemical analysis indicates that regulation of the Wnt pathway occurs in the nucleus at the level of activation of target genes by the beta-catenin-TCF complex, revealing a novel additional mechanism by which intestinal cells may regulate Wnt signaling during their maturation. In addition, other signaling pathways including Notch, BMP, Hedgehog, and growth factor, were temporally regulated during Caco-2 cell polarization. Surprisingly, modulation of these signaling pathways in Caco-2 cells occurs in the absence of morphogen gradients and interactions with stromal cells characteristic of enterocyte differentiation in situ. This dataset contains gene expression profiles of 9 normal colon samples and 15 colon tumor samples. Samples of tumor and normal colon mucosa were collected from colon cancer resection from Department of Surgery, Queen Mary Hospital University of Hong Kong. Tissue was frozen in liquid nitrogen within 30 min of resection. Nonneoplastic mucosa from colon was dissected free of muscle and histologically confirmed to be tumor free by frozen section. Total RNA was extracted using Trizol (Invitrogen, Carlsbad, CA) from each tissue sample and processed for microarray hybridization. A disease state experiment design type is where the state of some disease such as infection, pathology, syndrome, etc is studied. Disease State: Tumor/Normal colon samples Using regression correlation
Project description:A possible functional role of VCP/p97 during differentiation of U937 cells was addressed by siRNA-targeting of VCP/p97. Functional changes in VCP-siRNA-transfected cells following phorbol ester-induced monocytic differentiation have been examind by DNA microarray analysis. Cells transfected with the non-targeting AllStars negative siRNA (Qiagen) served as a reference. Computed
Project description:Human scrotum and labia majora samples treated with DHT and/or AZA A compound treatment design type is where the response to administration of a compound or chemical (including biological compounds such as hormones) is assayed. Compound Based Treatment: dihydrotestosterone (DHT), 5-aza-deoxy-cytidine (AZA), none or both Cell Line: normal scrotum or labia majora derived from complete androgen insensitivity syndrome due to inactivating mutation of the androgen receptor gene Computed