The invasion of activated fibroblasts represents a key pathomechanism in fibrotic diseases, carcinogenesis and metastasis. Here, invading fibroblasts contribute to fibrotic extracellular matrix (ECM) formation and the initiation, progression, or resistance of cancer, respectively. To construct a transcriptome-wide signature of fibroblast invasion, we used a multiplex phenotypic 3D invasion assay using murine lung fibroblasts. Microarray-based gene expression profiles of invading and non-invading fibroblasts were highly distinct: 1049 genes were differentially regulated (>1.5-fold). An unbiased pathway analysis (Ingenuity) identified a significant enrichment for the functional clusters invasion of cells, idiopathic pulmonary fibrosis (IPF) and metastasis. Particularly, matrix metalloprotease13 (MMP13), transforming growth factor (TGF)1, Caveolin1 (Cav1), Phosphatase and Tensin Homolog (Pten), and secreted frizzled-related protein1 (Sfrp1) were among the highest regulated genes. In silico analysis by Ingenuity predicted TGF1, epidermal growth factor (EGF), fibroblast growth factor2 (FGF2), and platelet-derived growth factor (PDGF)-BB to induce invasion. As such, these growth factors were tested in the 3D invasion assay and displayed a significant induction of invasion, thus validating the transcriptome profile. Accordingly, our transcriptomic invasion signature describes the invading fibroblast phenotype in unprecedented detail and provides a tool for future functional studies of cell invasion and therapeutic modulation thereof.
Validated prediction of pro-invasive growth factors using a transcriptome-wide invasion signature derived from a complex 3D invasion assay.
Sex
View SamplesFibrogenic processes instigate fatal chronic diseases leading to organ failure and death. Underlying biological processes involve induced massive deposition of extracellular matrix (ECM) by aberrant fibroblasts. We subjected diseased primary human lung fibroblasts to an advanced 3D phenotypic high-content assay and screened a library of FDA/EMA approved small molecules for inhibiting ECM deposition. Fibrotic Pattern Detection by Artificial Intelligence (FANTAIL) identified Tranilast as an effective inhibitor, however, by structure-activity relationship studies we found N-(2-butoxyphenyl)-3-(phenyl)acrylamides (N23Ps) as a novel and highly potent compound class. N23Ps suppressed myofibroblast transdifferentiation, ECM deposition, cellular contractility, and altered cell shapes, thus advocating a unique mode of action. Mechanistically, transcriptomics identified SMAD (de)ubiquitination/Smurf2 as a potential therapeutic target network. Antifibrotic activity of N23Ps was verified by proteomics in a human ex vivo tissue fibrosis disease model, suppressing profibrotic markers SERPINE1/PAI1 and CXCL8/IL8. Conclusively, these data suggest N23Ps as a novel class of highly potent compounds with implications for inhibiting organ fibrosis in patients.
Phenotypic drug screening in a human fibrosis model identified a novel class of antifibrotic therapeutics.
Specimen part, Treatment
View SamplesHigh-fat diets are associated with increased obesity and metabolic disease in mice and humans. Here we used analysis of variance (ANOVA) to scrutinize a microarray data set consisting of 10 inbred strains of mice from both sexes fed atherogenic high-fat and control chow diets. An overall F-test was applied to the 40 unique groups of strain-diet-sex to identify 15,288 genes with altered transcription. Bootstrapping k-means clustering separated these changes into four strain-dependent expression patterns, including two sex-related profiles and two diet-related profiles. Sex-induced effects correspond to secretion (males) or fat and energy metabolism (females), whereas diet-induced changes relate to neurological processes (chow) or immune response (high-fat). The full set of pairwise contrasts for differences between strains within sex (90 different statistical tests) uncovered 32,379 total changes. These differences were unevenly distributed across strains and between sexes, indicating that strain-specific responses to high-fat diet differ between sexes. Correlations between expression levels and 8 obesity-related traits identified 5,274 associations between transcript abundance and measured phenotypic endpoints. From this number, 2,678 genes are positively correlated with total cholesterol levels and associate with immune-related categories while 2,596 genes are negatively correlated with cholesterol and connect to cholesterol synthesis.
Practical applications of the bioinformatics toolbox for narrowing quantitative trait loci.
Sex
View SamplesIdentify genes that are differentially regulated as a consequence of restoration of full-length functional APC in a colorectal cancer cell lines. Overall design: Examine mRNA expression level changes between SW480 (APC defective) and SW480+APC (SW480 cells with restored functional APC) cells, whilst accounting for any non-specific expression changes by comparison to SW480+control vector.
Differential RNA-seq analysis comparing APC-defective and APC-restored SW480 colorectal cancer cells.
No sample metadata fields
View SamplesHigh quality genetic material is an essential pre-requisite when analyzing gene expression using microarray technology. Peripheral blood mononuclear cells (PBMC) are frequently used for genomic analyses, but several factors can affect the integrity of nucleic acids prior to their extraction, including the methods of PBMC collection and isolation. In this study, we compared the Ficoll-Paque density gradient centrifugation and BD Vacutainer cell preparation tube (CPT) protocols to determine if either method offered a distinct advantage in preparation of PBMC-derived immune cell subsets for their use in gene expression analysis. We compared gene expression in PBMC and individual immune cell types from Ficoll and CPT isolation protocols using Affymetrix microarrays.
Immune cell subsets and their gene expression profiles from human PBMC isolated by Vacutainer Cell Preparation Tube (CPT™) and standard density gradient.
Specimen part
View SamplesThis study compares the gene expression changes in Sus scrofa in response to two different methods for abdominal surgical incisions ; electrosurgery and harmonic blade.
Ultrasonic incisions produce less inflammatory mediator response during early healing than electrosurgical incisions.
Specimen part, Treatment
View SamplesSalicylic acid (SA) is a critical molecule mediating plant innate immunity with an important role limiting the growth and reproduction of the virulent powdery mildew (PM) Golovinomyces orontii on Arabidopsis thaliana. To investigate this later phase of the PM interaction, and the role played by SA, we performed replicated global expression profiling for wild type and SA biosynthetic mutant ics1 Arabidopsis from 0 to 7 days post infection. We found that ICS1-impacted genes comprise 3.8% of profiled genes with known molecular markers of Arabidopsis defense ranked very highly by the multivariate empirical Bayes statistic (T2 statistic ((Tai and Speed, 2006)). Functional analyses of T2-selected genes identified statistically significant PM-impacted processes including photosynthesis, cell wall modification, and alkaloid metabolism that are ICS1-independent. ICS1-impacted processes include redox, vacuolar transport/secretion, and signaling. Our data also supports a role for ICS1 (SA) in iron and calcium homeostasis and identifies components of SA crosstalk with other phytohormones. Through our analysis, 39 novel PMimpacted transcriptional regulators were identified. Insertion mutants in one of these regulators, PUX2, results in significantly reduced reproduction of the powdery mildew in a cell death independent manner. Though little is known about PUX2, PUX1 acts as a negative regulator of Arabidopsis CDC48 (Rancour et al., 2004; Park et al., 2007), an essential AAA-ATPase chaperone that mediates diverse cellular activities including homotypic fusion of ER and Golgi membranes, ER-associated protein degradation, cell cycle progression, and apoptosis. Future work will elucidate the functional role of the novel regulator PUX2 in PM resistance.
Temporal global expression data reveal known and novel salicylate-impacted processes and regulators mediating powdery mildew growth and reproduction on Arabidopsis.
Age, Specimen part
View SamplesWe report the application of ultrashort metabolic labeling of RNA for high-throughput profiling of RNA processing in Drosophila S2 cells. Overall design: Examination of 3 different labeling timepoints in Drosophila S2 cells.
The kinetics of pre-mRNA splicing in the <i>Drosophila</i> genome and the influence of gene architecture.
Cell line, Subject
View SamplesWe obtained global measurements of decay and translation rates for mammalian mRNAs with alternative 3'' untranslated regions (3'' UTRs). Overall design: 1 3P-Seq sample from 3T3 cells and 1 3P-Seq sample from mouse ES cells; 2 2P-Seq steady state and 4 2P-Seq with actinomycin D; 6 polysome fraction 2P-Seq
3' UTR-isoform choice has limited influence on the stability and translational efficiency of most mRNAs in mouse fibroblasts.
Specimen part, Treatment, Subject
View SamplesMouse FGF15 and human FGF19 are orthologous proteins that regulate bile acid metabolism. However, other hepatic functions of FGF15/19 are not well characterized.
FGF15/19 regulates hepatic glucose metabolism by inhibiting the CREB-PGC-1α pathway.
Sex, Specimen part
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