Project description:For the purpose of mechanism-based risk assessment, we investigated temporal concentration-dependent responses in cultures of PHH and HepaRG cells exposed to three cosmetics ingredients, 2,7-naphthalenediol (NPT), triclosan (TCS) and butylated hydroxytoluene (BHT), that are suspected to have liver injury liability. To facilitate the identification of early KEs and visualisation of their development over time, samples were collected at 4 time points, namely 8 and 24 hours (single exposure), 48 and 72 hours (daily repeated exposure), after exposure to a broad concentration range. Concentrations were selected based on the estimated Cmax of the cosmetic ingredients. The selected maximum concentration was set at approximately 100x Cmax, whilst the minimum tested concentration was approximately 0.2x Cmax. The large concentration range allowed to apply benchmark concentration (BMC) modelling on individual genes as well as gene co-expression networks to derive in vitro transcriptomics benchmark concentrations (BMCs) and assess their suitability to be used as PoD in chemical risk assessment.
Project description:For the purpose of mechanism-based risk assessment, we investigated temporal concentration-dependent responses in cultures of PHH and HepaRG cells exposed to three drugs, acetaminophen (APAP), cyclosporine A (CSA) and valproic acid (VPA), that have a high liability for drug-induced liver injury. To facilitate the identification of early KEs and visualisation of their development over time, samples were collected at 4 time points, namely 8 and 24 hours (single exposure), 48 and 72 hours (daily repeated exposure), after exposure to a broad concentration range. Concentrations were selected based on the reported total Cmax of each drug. As these are approved drugs currently on the market, they are not expected to induce overt adverse effects around Cmax. Therefore, the selected maximum concentration was set at approximately 30x Cmax, whilst the minimum tested concentration was approximately 0.1x Cmax. The large concentration range allowed to apply benchmark concentration (BMC) modelling on individual genes as well as gene co-expression networks to derive in vitro transcriptomics benchmark concentrations (BMCs) and assess their suitability to be used as PoD in chemical risk assessment.
Project description:The aim of this study was to perform the multiscale correlation between quantitative texture features phenotype of pre-biopsy biparametric MRI (bpMRI) and targeted sequence-based RNA expression for hypoxia-related genes. Images from pre-biopsy 3T bpMRI scans in clinically localised prostate cancer (PCa) patients of various risk categories (n=15) were used to extract textural features. The genomic landscape of hypoxia-related genes expression was obtained using post-radical prostatectomy tissue for targeted RNA expression profiling using the TempO-sequence method. The nonparametric Games Howell test was used to correlate the differential expression of the three important hypoxia-related genes with 28 radiomic texture features. Following this, cBioportal was accessed and a gene-oriented query was conducted to extract Oncoprint genomic output graph of the selected hypoxia-related genes from The Cancer Genome Atlas (TCGA). Correlation analysis using Pearson's coefficients calculated against each selected gene profile; survival analysis using Kaplan-Meier estimators were carried out. We found the quantitative bpMR imaging textural features, including histogram and grey level co-occurrence matrix (GLCM), correlated with hypoxia related genes (ANGPTL4, VEGFA, and P4HA1) seen on RNA sequencing using TempO-Seq method. Further radiogenomic analysis, including data accessed on cBioportal genomic database, confirmed that overexpressed hypoxia-related genes significantly correlated with a poor survival outcome, with a median survival of 81.11: 133.00 months in those with and without alterations of genes respectively. In summary, radiomic texture features of bpMRI in localised PCa correlate with the expression of hypoxia-related genes expression in prostate cancer. The expression data analysis showed that hypoxia-related genes are associated with poor survival.
Project description:The experiment investigates the effects of five well-known chemicals on the transcriptome of the HepaRG cell line, a metabolically competent hepatic cell line. The cells were treated individually with an increasing concentrations of aflatoxin B1, benzo[a]pyrene, cyclosporine A, rotenone or trichostatin A at five exposure time points followed by targeted RNA-seq using TempO-Seq technology (the panel of human whole transcriptome). The aim of the study was to explore how and to what extent the point-of-departure (POD) obtained from an in vitro transcriptomics study varied as a function of exposure time.
Project description:Gamma-delta (gd) T cells from pooled mouse lymph nodes and spleens were isolated and FACS-sorted for Vg1+Ly6C-, Vg1+Ly6C+, Vg4+Ly6C- and Vg4+Ly6C+ subsets of bulk CD27+ gdT cells.
Project description:The application of RNA-seq profiling of cell lines following compound treatment has been highlighted as a line of evidence for Next Generation Risk Assessment of ingredients. We demonstrate as part of a use case the evaluation of a targeted RNAseq using Tempo-Seq platform as a general unbiased approach to complement more targeted approaches. We show impact of the use of different cell lines to the point of departure.
Project description:Title: Regulation of gene expression through mitogen-activated protein kinase cascades in cardiac myocytes.<br/> Description: The aim of this study is to identify the changes in gene expression induced in rat neonatal <br/> ventricular myocytes, a well-established cell culture model, by endothelin-1, <br/> a known hypertrophic agonist, and to determine which of the changes are <br/> mediated through the ERK cascade. This continues TKAC^Ys previous study of the <br/> effects of oxidative stress, which induces cardiac myocyte apoptosis.<br/>
Project description:This experiment was carried out to investigate the responses of the human liver model (HepG2) cells upon the exposure of various DILI compounds (second set) in multiple severity scenarios by varying the concentration. The cells were seeded in 384 well plate format containing 8000 cell per well with 2 days of resting before exposure. The compounds were exposed for 4, 8, and 24 hours. The cells were lysed and the lysate was collected for high throughput targeted RNA-seq with the human whole genome library. The gene expression profiles were analyzed to identify the modulated responses of the cells. This experiment is part of the TransQST project.
Project description:Comparison of luminal and basal breast cancer cells under acute normoxia and hypoxia. Cells were plated in 96-well plates and incubated 24 hrs under normoxia or hypoxia after which the wells were washed once with cold PBS and lysed using TempO-Seq lysis buffer for 15 min at room temperature. Samples were stored at −80 °C before shipping to BioClavis for whole genome TempO-Seq analysis. For normoxia (NX) 21% oxygen was used and for hypoxia (HX) 1%oxygen was used. Each condition has 3 biological replicates.
Project description:Comparison of luminal and basal breast cancer cells under chronic normoxia and hypoxia. Cells were plated in 96-well plates and incubated 5 days under normoxia or hypoxia after which the wells were washed once with cold PBS and lysed using TempO-Seq lysis buffer for 15 min at room temperature. Samples were stored at −80 °C before shipping to BioClavis for whole genome TempO-Seq analysis. For normoxia (NX) 21% oxygen was used and for hypoxia (HX) 1%oxygen was used. Each condition has 3 biological replicates.