Transcription profiling by array of mouse epidermis and pancreas following MycERTAM activation at 4 time points
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
SUBMITTER: Sam Robson
PROVIDER: E-MEXP-2952 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:GLP-1 analogues, such as exendin-4, preserve functional β-cell mass in various model systems and are revolutionising management of type 2 diabetes. Yet, comparatively little is known about effectiveness in the face of severe β-cell depletion. Moreover, direct and sequential effects of exendin-4 on islet-specific gene expression over time in vivo are not well characterised. To address these issues and others, we have examined the time-dependent effects of exendin-4 treatment on β-cell mass regulation alongside accompanying changes in islet gene expression in vivo. Context-dependent actions were assessed by comparing effects on normal islets and also following massive toxigenetic β-cell ablation in pIns-MYCERTAM transgenic mice in vivo. Despite over 90% loss of β-cell mass, exendin-4 treatment normalised blood glucose and insulin levels in hyperglycaemic mice, though benefits rapidly waned on withdrawal of treatment. As exendin-4 did not arrest the decline in β-cell mass or turnover in this study, we could directly isolate effects on function of surviving β-cells. Improved glucose homeostasis was associated with dynamic changes in multiple islet genes and pathways in vivo favouring glucose-stimulated insulin secretion, such as Irs2, Pdx1, Sox4, glucokinase, and glycolysis pathway. Several key growth pathways and epigenetic regulators were also differentially expressed. Thus, even in the face of extensive β-cell loss exendin-4 can markedly improve hyperglycaemia by differential gene expression in surviving islet cells. Activation of MYCERTAM was achieved through administration of 1mg of 4 hydroxytamoxifen (4OHT; Sigma-Aldrich, St. Louis, MO) by daily intraperitoneal injection. To assess the effect of exendin-4 on MYCER-induced hyperglycaemia, mice were given either twice-daily subcutaneous (sc) injections of exendin-4 (50ug/kg dissolved in 5mls water), or equivalent volumes of water vehicle, starting 2 days prior to 4OHT injections. For microarray analyses parallel mouse experiments were set up using 8-12 week old pIns-MYCERTAM male mice either treated with 4OHT or vehicle (peanut oil) and exendin-4 or vehicle, as described, for 4, 8, 16, 32 and 72 hours (n=3 for each time point and for each of four conditions; 4OHT and exendin-4 treated, peanut oil and exendin-4 treated, 4OHT and water treated, peanut oil and water treated). !Sample_data_processing = After the quality control step, the following 8 samples out of 60 showing poor reproducibility were excluded from our further study: GSM930242, GSM930247, GSM930251, GSM930263, GSM930264, GSM930289, GSM930291, GSM930298.
2014-04-11 | E-GEOD-37936 | biostudies-arrayexpress
Project description:It is known that the stresses encountered during islet isolation have deleterious effects on beta-cell physiology. The nature of these effects, however, is incompletely known, partly due to the heterogeneity of islet preparations. The objective was to assess the genome-wide effect of islet isolation and in-vitro culture on beta-cell transcriptome. Beta cells identified by their intrinsic autofluorescence, were captured from donor pancreas and isolated islets using Laser Capture Microdissection. Beta cells from donor pancreas serve as the control. Our results indicated induction of a strong inflammatory response induced by islet isolation which continues during in vitro culture manifested by upregulation of several cytokines, chemokines and their receptors. IL-8 was induced by 3.6-fold and 56-fold in fresh and 3-days cultured islets respectively. Concordantly, several pancreatic progenitor cell-specific transcription factors like were upregulated in cultured islets, suggesting progressive transformation of mature beta-cell phenotype toward an immature endocrine cell phenotype. There are three sample types in our experiment; 1) beta-cells captured from the frozen sections of donor pancreas n=8, 2) beta cells extracted from islets frozen immediately after isolation (d0 islets) n=8 and, 3) beta cells extracted from isolated islets frozen after culturing for 3 days (d3 islets) n=5. For the analysis, beta cells from pancreas were considered as the reference point.
2012-02-12 | E-GEOD-29113 | biostudies-arrayexpress
Project description:We established a model system in human DLD1 colon cancer cells to study the transcriptional crosstalk between FOXO3A and beta-Catenin. Thereby, translocation to the nucleus of a AKT-insensitive mutant (T32A, S253A, S315A) of human FOXO3A fused to the ligand binding domain of human estrogen receptor can be induced by exposure to 4-hydroxy-tamoxifen. Furthermore, expression of a stable mutant (S33Y) of human beta-catenin is doxycycline inducible. Addition of those drugs separately or in combination allows identification of common or excusive target gene sets.
2012-05-01 | E-MEXP-3262 | biostudies-arrayexpress
Project description:The mammalian FoxO transcription factors - FoxO1, FoxO3, FoxO4 - function in the nucleus to direct transcription of specific gene targets governing cellular survival, proliferation, metabolism, differentiation and oxidative defense. Activation of PI3K by extracellular growth factors leads to AKT-mediated phosphorylation of FoxO1, FoxO3 and FoxO4, resulting in their sequestration in the cytoplasm such that they are unable to regulate their gene targets. Our study identified FoxOs as novel tumor suppressors in kidney cancer (Gan et al, 2010, Cancer Cell). To understand the tumor suppression function of FoxOs in kidney cancer cells, we performed gene expression profiling in human kidney cancer cells upon FoxO1 or FoxO3 reactivation in order to identify the key transcriptomic alterations mediating FoxO tumor suppression function in kidney cancer cells. We generated RCC4 and UMRC2 cell lines (two human kidney cancer cells with low endogenous FoxO1 and FoxO3 expression) with stable expression of FoxO1(TA)ERT2 or FoxO3(TA)ERT2 construct, which expressed a fusion protein consisting of FoxO(TA) (containing three Ser/Thr AKT phosphorylation sites mutated to alanine) fused to the T2-modified estrogen receptor (ERT2) moiety. We documented that the FoxO(TA)ERT2 fusion protein sequestered FoxO(TA) in the cytoplasm and that 4OHT treatment resulted in rapid translocation of FoxO(TA)ERT2 into the nucleus. We also established stable cell lines with ERT2 expression as control cell lines. (For simplicity, ERT2, FoxO1(TA)ERT2 and FoxO3(TA)ERT2 cell lines will be referred to as EV (empty vector), FoxO1 and FoxO3, respectively, hereafter). We then conducted comparative transcriptome analysis (using the Human Genome U133 Plus 2.0 Array) of EV, FoxO1, or FoxO3-expressing RCC4 and UMRC2 cells at 12 hours with or without 100 nm 4OHT treatment (cultured in DMEM+10% FBS with puromycin selection). To enrich for more proximal actions of FoxO, we selected the 12 hour time point as time course studies revealed dramatic transcriptional changes of known FoxO targets (such as Cyclin D1), yet no discernable cellular phenotypes (apoptosis and cell cycle arrest). We generated 4 transcriptome datasets: FoxO1 RCC4, FoxO3 RCC4, FoxO1 UMRC2, and FoxO3 UMRC2 (by comparing transcriptome data with or without 4OHT treatment), and normalized these transcriptome data against 4OHT-treated EV cells, which show modest 4OHT-induced transcriptional changes.
2011-10-20 | E-GEOD-23926 | biostudies-arrayexpress
Project description:Peroxisome proliferator-activated receptor beta/delta protects against obesity by reducing dyslipidemia and insulin resistance via effects in various organs, including muscle, adipose tissue, liver, and heart. However, nothing is known about the function of PPAR-beta in pancreas, a prime organ in the control of glucose metabolism. To gain insight into so far hypothetical functions of this PPAR isotype in insulin production, we specifically ablated Ppar-beta in pancreas. The mutated mice developed a chronic hyperinsulinemia, due to an increase in both beta-cell mass and insulin secretion. Gene expression profiling indicated a broad repressive function of PPAR-beta impacting the vesicular compartment, actin cytoskeleton, and metabolism of glucose and fatty acids. Analyses of insulin release from the islets revealed an increased second-phase glucose-stimulated insulin secretion. Higher levels of PKD, PKC-delta and diacyglycerol in mutated animals lead to an enhanced formation of trans-Golgi network (TGN)-to-plasma-membrane transport carriers in concert with F-actin disassembly, which resulted in increased insulin secretion and its associated systemic effects. Taken together, these results provide evidence for PPAR-beta playing a repressive role on beta-cell growth and insulin exocytosis, which shed new light on its anti-obesity action. Pancreas-specific knock-out animals were generated by breeding mice harbouring a floxed Ppar-beta (PPARbetafl/fl) to mice expressing the Cre transgene under the control of the Pdx1 promoter (Pdx1Cre). Islets from 3 different animals from the knock-out group Pdx1Cre;PPARbetafl/fl and the control littermates group PPARbetafl/fl were compared.
2012-08-30 | E-GEOD-16048 | biostudies-arrayexpress
Project description:Hyperinsulinemia often precedes type 2 diabetes but its role in disease progression is unknown. Palmitoylation, a protein modification implicated in regulated exocytosis, is reversed by acyl-protein thioesterase 1 (APT1). We found altered APT1 biology in pancreatic islets from humans with type 2 diabetes, and APT1 knockdown in nondiabetic human islets caused insulin hypersecretion. Chow fed global and islet specific APT1 knockout mice had enhanced glucose tolerance due to islet autonomous increased glucose-stimulated insulin secretion. APT1 deficiency did not affect islet calcium dynamics but prolonged insulin granule fusion. Using palmitoylation proteomics, we identified Scamp1 as an APT1 substrate that localized to insulin secretory granules. Knockdown of Scamp1 caused insulin hypersecretion. APT1 deficient insulinoma cells subjected to nutrient excess had increased apoptosis, and expression of a mutated Scamp1 incapable of being palmitoylated in APT1 deficient cells rescued insulin hypersecretion and nutrient induced apoptosis. Compared to APT1 sufficient controls, high fat fed islet specific APT1 knockout mice and global APT1 deficient db/db mice showed increased -cell failure. These findings suggest that the depalmitoylation enzyme APT1 is regulated in human islets, and that APT1 deficiency causes insulin hypersecretion leading to -cell failure, modeling the evolution of some forms of human type 2 diabetes.
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Project description:This SuperSeries is composed of the SubSeries listed below. Refer to individual Series
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2014-06-11 | E-GEOD-58360 | biostudies-arrayexpress
Project description:Expression of deltaNB-cateninER allows the time and duration of B-catenin activation to be controlled precisely through the application of 4OHT. We have shown that B-catenin activation in adult mouse epidermis stimulates de novo hair follicles and the formation of a new bulge and melanocyte niche. We utilize Affymetrix Mouse Genome 430A 2.0 oligonucleotide arrays to investigate the differential regulation of RNA in skin from 7 week old female deltaNB-cateninER transgenic mice following 0, 1, or 7 days B-catenin activation in the epidermis. RNA from age and sex matched wild-type littermates was used as a control.
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