The objective of the study was to evalute the changes in gene expression associated to UCP2 invalidation in colon tumors from AOM/DSS-treated mice
UCP2 Deficiency Increases Colon Tumorigenesis by Promoting Lipid Synthesis and Depleting NADPH for Antioxidant Defenses.
Specimen part
View SamplesZinc finger nucleases (ZFN) are powerful tools for editing genes in cells. Here we use ZFNs to interrogate the biological function of human ADPGK, which encodes an ADP-dependent glucokinase (ADPGK), in tumour cell lines. The hypothesis tested is that ADPGK utilises ADP to phosphorylate glucose under conditions where ATP becomes limiting, such as hypoxia. We characterised two ZFN knockout clones in each of two tumour cell lines (H460 and HCT116). All four lines had frameshift mutations in all alleles at the target site in exon 1 of ADPGK, and were ADPGK-null by immunoblotting. ADPGK knockout had little or no effect on cell proliferation, but compromised the ability of H460 cells to survive siRNA silencing of hexokinase-2 under oxic conditions, with clonogenic survival falling from 213% for the parental line to 6.40.8% (p=0.002) and 4.30.8% (p=0.001) for the two knockouts. A similar increased sensitivity to clonogenic cell killing was observed under anoxia. No such changes were found when ADPGK was knocked out in HCT116 cells, for which the parental line was less sensitive than H460 to anoxia and to hexokinase-2 silencing. While knockout of ADPGK in HCT116 cells caused few changes in global gene expression, knockout of ADPGK in H460 cells caused notable up-regulation of mRNAs encoding cell adhesion proteins. Surprisingly, we could discern no effect on glycolysis as measured by glucose consumption or lactate formation under oxic or anoxic conditions, or extracellular acidification rate (Seahorse XF analyser) under oxic conditions in a variety of media. However, oxygen consumption rates were generally lower in the ADPGK knockouts, in some cases markedly so. Collectively, the results demonstrate that ADPGK can contribute to tumour cell survival under conditions of high glycolytic dependence, but the phenotype resulting from knockout of ADPGK is cell line dependent and appears to be unrelated to priming of glycolysis.
Expression and role in glycolysis of human ADP-dependent glucokinase.
Cell line
View SamplesIn the developing embryo, haematopoietic stem cells (HSCs) emerge from the aorta-gonad-mesonephros (AGM) region but the molecular regulation of this process is poorly understood. Recently, the progression from E9.5 to E10.5 and polarity along the dorso-ventral axis have been identified as clear demarcations of the supportive HSC niche. To identify novel secreted regulators of HSC maturation, we performed RNA-sequencing over these spatio-temporal transitions in the AGM region, and supportive OP9 cell line. Overall design: RNA-sequencing profiles of the aorta-gonad-mesonephros region from E9.5 embryos and E10.5 embryos sub-dissected into dorsal (AoD), ventral (AoV) and urogenital ridges (UGR) and pooled from between 15 and 34 embryos in three separate experiments.
A molecular roadmap of the AGM region reveals BMPER as a novel regulator of HSC maturation.
Specimen part, Subject
View SamplesIn the developing embryo, haematopoietic stem cells (HSCs) emerge from the aorta-gonad-mesonephros (AGM) region but the molecular regulation of this process is poorly understood. Recently, the progression from E9.5 to E10.5 and polarity along the dorso-ventral axis have been identified as clear demarcations of the supportive HSC niche. To identify novel secreted regulators of HSC maturation, we performed RNA-sequencing over these spatio-temporal transitions in the AGM region, and supportive OP9 cell line. Overall design: RNA-sequencing profiles of OP9 cells grown in flat, submersed culture or reaggregate and cultured at the liquid-gas interface were compared.
A molecular roadmap of the AGM region reveals BMPER as a novel regulator of HSC maturation.
Specimen part, Cell line, Subject
View SamplesHere we describe a method for fabricating a primary human Small Intestine-on-a-Chip (Intestine Chip) containing epithelial cells isolated from healthy regions of intestinal biopsies. The primary epithelial cells are expanded as 3D organoids, dissociated, and cultured on a porous membrane within a microfluidic device with human intestinal microvascular endothelium cultured in a parallel microchannel under flow and cyclic deformation. In the Intestine Chip, the epithelium forms villi-like projections lined by polarized epithelial cells that undergo multi-lineage differentiation similar to that of intestinal organoids, however, these cells expose their apical surfaces to an open lumen and interface with endothelium. Transcriptomic analysis also indicates that the Intestine Chip more closely mimics whole human duodenum in vivo when compared to the duodenal organoids used to create the chips. Because fluids flowing through the lumen of the Intestine Chip can be collected continuously, sequential analysis of fluid samples can be used to quantify nutrient digestion, mucus secretion and establishment of intestinal barrier function over a period of multiple days in vitro. The Intestine Chip therefore may be useful as a research tool for applications where normal intestinal function is crucial, including studies of metabolism, nutrition, infection, and drug pharmacokinetics, as well as personalized medicine.
Development of a primary human Small Intestine-on-a-Chip using biopsy-derived organoids.
Specimen part
View SamplesWe report application of RNA-seq to quantify gene expression changes in fasted mouse livers compared to re-fed controls. Overall design: RNA-seq from livers of re-fed and 48h fasted mice.
Histone propionylation is a mark of active chromatin.
Sex, Specimen part, Treatment, Subject
View SamplesThe molecular role of iron in gene expression remains poorly characterized. Moreover, the alterations in global gene expression after iron chelation remains unclear and are important to assess for understanding the molecular pathology of iron-depletion and the biological effects of iron chelators. We assessed the effect on whole genome gene expression of two iron chelators (desferrioxamine and Dp44mT). These studies are important for understanding the molecular and cellular effects of iron-depletion.
Cellular iron depletion stimulates the JNK and p38 MAPK signaling transduction pathways, dissociation of ASK1-thioredoxin, and activation of ASK1.
Cell line
View SamplesWe have characterized gene expression changes in HeLa cells following long term depletion of Cyclin T2 or Cyclin T1 using shRNA
Limited redundancy in genes regulated by Cyclin T2 and Cyclin T1.
Cell line, Treatment
View SamplesAT6.1 cells transfected to over-express Ndrg-1 were compared with AT6.1 vector control cells in a microarray analysis. The aim of the study was to identify differentially expressed genes between the two cell lines, as these may be modulated by Ndrg-1.
The iron-regulated metastasis suppressor, Ndrg-1: identification of novel molecular targets.
Cell line
View SamplesRNA-seq data of crwn1, crwn2, crwn4, crwn1 crwn2 and crwn1 crwn4
Loss of CRWN Nuclear Proteins Induces Cell Death and Salicylic Acid Defense Signaling.
Age, Specimen part
View Samples