Project comparing RNA prep and sequencing methods, as well as alignment and analysis methods.
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Sex, Age, Specimen part, Cell line
View SamplesIntact living conduit vessels (umbilical veins) were exposed to normal or high intraluminal pressure, or low or high shear stress in combination with a physiological level of the other force. We used a unique vascular ex vivo perfusion system. After six hours of perfusion endothelial cells were isolated from the stimulated vessels and RNA was extracted. RNA from 16 experiments from each stimulation were pooled and analyzed in duplicate DNA microarrays.
Differential global gene expression response patterns of human endothelium exposed to shear stress and intraluminal pressure.
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View SamplesSynaptic dysfunction is thought to underlie altered sociability in autism. However, the gene regulatory mechanisms that control synaptic protein expression in the context of social behaviour are poorly explored. Here we show that deletion of the large placental mammal specific miR379-410 cluster in mice leads to hypersocial behaviour, increased excitatory synaptic transmission and exaggerated expression of ionotropic glutamate receptor complexes in the hippocampus. Thus, interfering with miR379-410 could represent a novel therapeutic strategy for social deficits in autism.
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Sex, Age, Specimen part, Cell line
View SamplesTo identify epigenetically silenced genes in multiple myeloma (MM) cell lines and to determine the effects of 5-aza-2-deoxycytidine and trichostatin A on gene expression. We treated 3 multiple myeloma cell lines (MM1, NCI-H929, U266) with 5-aza-2-deoxycytidine and/or trichostatin A.
Genome-wide transcriptional response to 5-aza-2'-deoxycytidine and trichostatin a in multiple myeloma cells.
Specimen part, Disease, Cell line
View SamplesWe collected human aborted embryonic and fetal hearts from authorized resources with appropriate informed consents. Collected hearts were micro-dissected into 3 parts (atria, ventricle, and outflow tract), which were further digested into single cardiac cells and subject to single-cell RNA-seq analyses.
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View SamplesHuman colon mucosal biofilms, whether from tumor hosts or healthy individuals undergoing screening colonoscopy, are carcinogenic in murine models of CRC.
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Sex, Specimen part, Cell line, Treatment
View SamplesBackground: RNA-Seq is supplanting microarrays as the preferred method of transcriptome-wide identification of differentially expressed genes. However, RNA-Seq analysis is still rapidly evolving, with a large number of tools available for each of the three major processing steps: read alignment, expression modeling, and statistical determination of differentially expressed genes. Although some studies have benchmarked these tools against gold standard gene expression sets, few have evaluated their performance in concert with one another. Additionally, there is a general lack of testing of such tools on real-world, biologically relevant datasets, which often possess qualities not reflected in tightly controlled reference RNA samples or synthetic datasetsResults: Here we evaluate ten combinatorial implementations of several of the most commonly used analysis tools (RSEM, TopHat2, STAR, htseq, Cufflinks, DESeq2, edgeR, EBseq, and Cuffdiff) for their impact on differential gene expression analysis by RNA-Seq. A test dataset was generated from highly purified human classical and nonclassical monocyte subsets from a clinical cohort, allowing us to evaluate analysis workflow performance using four previously published microarray and BeadChip analyses of the same cell populations as reference datasets. We find that the choice of methodologies leads to wide variation in number of genes called significant, as well as precision and recall. In general, recall is correlated with the number of significant genes identified, whereas precision is inversely correlated with both recall and the number of significant genes identified. Additionally, we report that the choice of statistical analysis approach and read aligner exhibited stronger impacts on recall, precision, and F1 score than the choice of software for expression modeling.Conclusions: There is wide variation in the performance of RNA-Seq workflows to identify differentially expressed genes. Different workflows lead to a precision/recall tradeoff, and the ultimate choice of workflow should take into consideration how the results will be used in subsequent applications. Our analyses highlight the performance characteristics of these workflows, and the data generated in this study could also serve as a useful resource for future development of software for RNA-Seq analysis.
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Sex, Specimen part
View SamplesCardiac hypertrophy is a response to hemodynamic stress, and is associated with cardiac dysfunction and death. However, whether hypertrophy itself represents a disease process remains unclear. Hypertrophy is driven by changes in myocardial gene expression that require the MEF2 family of DNA-binding transcription factors, as well as the nuclear lysine acetyltransferase p300. In this study we sought to determine the effects of preventing MEF2 acetylation on cardiac adaptation to stress, using a small molecule designed to interfere with MEF2-co-regulator binding and acetylation. The data provided here include RNASeq analysis of left ventricular tissue from mice subjected to surgical pressure overload or a sham operation, and treated with 8MI or its vehicle for 4 weeks. We observed that 8MI transformed the transcriptional response to pressure overload, normalizing almost all 232 genes dysregulated by hemodynamic stress. We conclude that MEF2 acetylation is required for development and maintenance of pathological cardiac hypertrophy, and that blocking MEF2 acetylation can permit recovery from hypertrophy without impairing physiologic adaptation.
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Sex, Specimen part
View SamplesDespite recent advances in understanding macrophage activation, little is known regardinghow human alveolar macrophages in health calibrate its transcriptional response to canonicalTLR4 activation. In this study, we examined the full spectrum of LPS activation and determinedwhether the transcriptomic profile of human alveolar macrophages is distinguished bya TIR-domain-containing adapter-inducing interferon-ß (TRIF)-dominant type I interferon signature.Bronchoalveolar lavage macrophages were obtained from healthy volunteers, stimulatedin the presence or absence of ultrapure LPS in vitro, and whole transcriptomic profilingwas performed by RNA sequencing (RNA-Seq). LPS induced a robust type I interferon transcriptionalresponse and Ingenuity Pathway Analysis predicted interferon regulatory factor(IRF)7 as the top upstream regulator of 89 known gene targets. Ubiquitin-specific peptidase(USP)-18, a negative regulator of interferon a/ß responses, was among the top up-regulatedgenes in addition to IL10 and USP41, a novel gene with no known biological function but withhigh sequence homology to USP18.We determined whether IRF-7 and USP-18 can influencedownstream macrophage effector cytokine production such as IL-10.We show thatIRF-7 siRNA knockdown enhanced LPS-induced IL-10 production in human monocytederivedmacrophages, and USP-18 overexpression attenuated LPS-induced production ofIL-10 in RAW264.7 cells. Quantitative PCR confirmed upregulation of USP18, USP41, IL10,and IRF7. An independent cohort confirmed LPS induction of USP41 and IL10 genes. Theseresults suggest that IRF-7 and predicted downstream target USP18, both elements of a typeI interferon gene signature identified by RNA-Seq, may serve to fine-tune early cytokineresponse by calibrating IL-10 production in human alveolar macrophages.
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Sex, Age, Specimen part, Treatment
View SamplesCampylobacter jejuni promotes colorectal tumorigenesis through the action of cytolethal distending toxin
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Sex, Specimen part, Cell line
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