Genome-wide mRNA expression profiles of FACS-purified rat beta cells freshly isolated from control and 24h-fasted rats
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ABSTRACT:
ORGANISM(S): Rattus norvegicus
SUBMITTER: Geert Martens
PROVIDER: E-GEOD-30964 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:The study was designed to capture the in vivo adaptations of nutrient-sensing pancreatic beta cells to fed or fasted (24h) state. Short periods of fasting are thought to mediate the beta cells’ competence for insulin synthesis and secretion. We examined whether it influenced their typical mRNA expression. After 24h of fasting, a 1.5 fold change was detected in 1.5% of all assayed transcripts (p<0.05), with 95% of altered genes (430 of 452) being down-regulated (Fig.6A). Suppression was more marked for a panel of genes with beta cell-specific expression, of which 12% were 1.5-fold suppressed. Suppressed beta cell marker genes were statistically enriched in gene clusters of endoplasmic reticulum (ER), ER to Golgi transport, protein folding and secretory vesicle-mediated transport, i.e. the pathways of protein synthesis and transport. (See Martens G.A. et al. PLoS One 2011)
2011-09-01 | GSE30964 | GEO
Project description:The lower glucose-responsiveness of neonatal beta cells is generally considered a sign of endocrine immaturity. We compared mRNA profiles of neonatal and 10-weeks old rat beta cells to see how their gene expression changes with functional maturation. Neonatal beta cells showed a lower glucose-inducible increment in insulin production than adult cells. This was in part explained by basal protein synthetic hyperactivity of neonatal cells: while at 2.5mM glucose 80% of neonatal beta cells were recruited into active protein synthesis, 10 mM glucose was required to achieve a similar fraction of active adult beta cells. Besides this progressive recruitment, glucose exerted in both age groups an additional amplifying effect in the recruited cells, but clearly more so in adult beta cells that showed a higher maximal synthetic capacity/cell. Neonatal beta cells balanced an advanced endocrine differentiation as judged by their mRNA expression of conserved beta cell marker genes, with higher expression of genes involved in cell cycle and development. One example, Delta-like 1 homolog (DLK1) was used to investigate if neonatal beta cells with basal hyperactivity corresponded to a more immature subset, as marked by high DLK1. Neonatal pancreas contained distinct subsets of DLK1high and DLK1low insulin-expressing cells, but both showed equal hyperactivity. We conclude that neonatal beta cells combine advanced endocrine maturation with traits of residual developmental immaturity. If DLK1 is used as marker for the latter, the basal hyperactivity which proved to be a cardinal feature of neonatal beta cells is not a direct reflection of their residual immaturity. A genome-wide view on postnatal maturation/differentiation of the rat beta cells The study compares Affymetrix RG230.20 expression profiles of 3 cell subsets: (1) HSSC: a population of cells isolated from post natal day 2-3 rat pancreas that is enriched (65-70%) in insulin cells (n=3), (2) LSSC: a population of cells isolated from postnatal day 2-3 rat pancreas that is mostly devoid of endocrine cells, and selected by its low side scatter (n=3); and (3) adult beta: a reference population of FACS-purified adult rat beta cells (>90% insulin+) freshly isolated from pancreas of 10 weeks-old rats (n=5). RNA was extracted from the cells immediately after isolation, without further treatment or culture. Details of the isolation procedure and extensive microscopic characterization of the preps are described in the accompanying scientific paper by Martens GA et al.
2013-08-20 | E-GEOD-47174 | biostudies-arrayexpress
Project description:Islet transplantation exposes beta cells to mild hyperglycemia and to the abnormal environment of the transplant site. These conditions may affect beta cells and induce the expression of genes involved in beta cell damage. Gene expression profile of human beta cells exposed to mild hyperglycemia by transplantation into ICR-SCID mice was evaluated and compared with the gene profile of beta cells obtained from non-diabetic subjects. We found that the transplanted beta cells showed an unfolded protein response (UPR). There was upregulation of many genes of the IRE-1 pathway that provide protection against the deleterious effects of ER stress. Among them, increased expression of genes coding XBP-1; the chaperone proteins PDIA4, Bip, and Grp94; and the ER degradation proteins EDEM1 and EDEM2. ERdj4 and DNA-JC3 were also upregulated. JUK had downregulated expression. The PERK and ATF-6 arms of the ER stress response had many downregulated genes in transplanted islets. The PERK's substrates, EIF2A and NRF2, showed markedly reduced expression, as downregulated was the expression of CReP. Other downregulated genes included HERP2, IRS-2, CHOP and C/EBP-beta. In the transplanted beta cells there was significantly decreased expression of ATF-6, as well as the downstream gene products CHOP and HERP2. There was increased expression of HRD1, which exerts an antiapoptotic effect by degrading unfolded proteins. In conclusion human beta cells in a transplant site had UPR changes in gene expression that protect against the proapoptotic effects of unfolded proteins. Frozen sections were obtained from pancreases of non-diabetic subjects at surgery and from human islets transplanted into ICR-SCID mice for 4 weeks. β cell enriched samples were obtained by laser capture microdissection. RNA was extracted, amplified and subjected to microarray analysis.
2010-06-08 | E-GEOD-22097 | biostudies-arrayexpress
Project description:Aims/hypothesis: Duct cells isolated from adult human pancreas can be reprogrammed to express islet beta cell genes by adenoviral transduction of the developmental transcription factor neurogenin3 (Ngn3). In this study we aimed to fully characterize the extent of this reprogramming and intended to improve it. Methods: The extent of the Ngn3-mediated duct-to-endocrine cell reprogramming was measured employing genome wide mRNA profiling. By modulation of the Delta-Notch signaling or addition of pancreatic endocrine transcription factors Myt1, MafA and Pdx1 we intended to improve the reprogramming. Results: Ngn3 stimulates duct cells to express a focused set of genes that are abundant in islet endocrine cells and/or neural tissues. This neuro-endocrine shift, however, covers a minor fraction (5%) of the estimated genome-wide transcriptome difference between duct and islet endocrine cells. Interestingly, transduction of exogenous Ngn3 activates endogenous Ngn3 suggesting auto-activation of this gene. Furthermore, pancreatic endocrine reprogramming of human duct cells can be moderately enhanced by inhibition of Delta-Notch signaling as well as by co-expressing the transcription factor Myt1, but not MafA and Pdx1. Conclusions/interpretation: The results provide further insight into the plasticity of adult human duct cells and suggest measurable routes to enhance Ngn3-mediated in vitro reprogramming protocols for regenerative beta cell therapy in diabetes. We used Affymetrix HG133A and HG133B to get a comprehensive view on the reprogramming potential in vitro of human pancreatic duct cell cultures (n=3-4) at 3 and 14/20 days after ectopic adenoviral expression of murine neurogenin 3 as compared to GFP-expressing control vectors. The microarray analysis was performed on 3 independent samples that each contained RNA extracted from a pool of 3 independent donor pancreata. The total number of non-selected donor organs is 9. Transcripts were considered as differentially regulated by Ngn3 when 1.5 fold (LCB, unpaired P < 0.05) up- or down-regulated in AdGFP-Ngn3 versus AdGFP controls, at 3 and/or 14 dpi. Transcripts that showed differential expression between day 3 and day 14 in AdGFP-Ngn3 duct cells but not in AdGFP control cells, were also considered Ngn3-regulated
2012-07-10 | E-GEOD-30802 | biostudies-arrayexpress
Project description:In cell models, changes in the “accessible” pool of plasma membrane (PM) cholesterol are linked with the regulation of ER sterol synthesis and metabolism by the Aster family of nonvesicular transporters. However, the relevance of such nonvesicular transport mechanisms for lipid homeostasis in vivo has not been defined. Here we reveal two physiological contexts that generate accessible PM cholesterol and engage the Aster pathway in liver: fasting and reverse cholesterol transport (RCT). During fasting, adipose tissue–derived fatty acids activate hepatocyte sphingomyelinase to liberate sequestered PM cholesterol. Aster-dependent cholesterol transport during fasting facilitates cholesteryl ester (CE) formation, cholesterol movement into bile, and VLDL production. During RCT, HDL delivers excess cholesterol to the hepatocyte PM through SR-BI. Loss of hepatic Asters impairs cholesterol movement into feces, raises plasma cholesterol levels, and causes cholesterol accumulation in peripheral tissues. These results reveal fundamental mechanisms by which Aster cholesterol flux contributes to hepatic and systemic lipid homeostasis.
2022-10-24 | GSE206278 | GEO
Project description:Aims: establishment of reference samples to investigate gene expression selective for endocrine or ductal-exocrine cells within the adult human pancreas. To this end, human islet endocrine cells, FACS-enriched in insulin+ cells, (n=3) and human exocrine ductal cells (n=2) are compared on Affymetrix HG133A platform with duplicate hybridizations of a panel of other primary human tissues. The microarray analysis was performed on 3 pools of human beta cell-enriched cell fractions, isolated from 10 non-selected donor organs, and 2 pools of duct cell-enriched fractions obtained from 6 non-selected donor pancreases. The cells were suspension-cultured for 2-3 weeks along standard procedures and with no specific treatment prior to FACS-sorting and RNA extraction. The average composition of human beta cell-preparations was 55 ± 13% insulin+ cells, 13±8% glucagon+ cells and 21 ±7% non-granulated cells. Pancreatic duct cells-enriched preparations contained 85 ±7% cytokeratin 19+ cells with 4 ± 1% insulin+ cells and 6 ±4% glucagon+ cells and were isolated as described by Heimberg H. et al.( Diabetes 2001,49: 571-579 ). Pancreatic cell mRNA profiles were compared to those of a panel of other human primary tissues (n=2 biological replicates). This dataset is part of the TransQST collection.
2011-07-21 | E-GEOD-30803 | biostudies-arrayexpress
Project description:Background The gut is a major energy consumer, but a comprehensive overview of the adaptive response to fasting is lacking. Gene-expression profiling, pathway analysis, and immunohistochemistry were therefore carried out on mouse small intestine after 0, 12, 24, and 72 hours of fasting. Results Intestinal weight declined to 50% of control, but this loss of tissue mass was distributed proportionally among the gut’s structural components, so that the microarrays’ tissue base remained unaffected. Unsupervised hierarchical clustering of the microarrays revealed that the successive time points separated into distinct branches. Pathway analysis depicted a pronounced, but transient early response that peaked at 12 hours, and a late response that became progressively more pronounced with continued fasting. Early changes in gene expression were compatible with a cellular deficiency in glutamine and metabolic adaptations directed at glutamine conservation, inhibition of pyruvate oxidation, stimulation of glutamate catabolism via aspartate and phosphoenolpyruvate to lactate, and enhanced fatty-acid oxidation and ketone-body synthesis. In addition, the expression of key genes involved in cell cycling and apoptosis was suppressed. At 24 hours of fasting, many of the early adaptive changes abated. Major changes upon continued fasting implied the production of glucose rather than lactate from carbohydrate backbones, a downregulation of fatty-acid oxidation and a very strong downregulation of the electron-transport chain. Cell cycling and apoptosis remained suppressed. Conclusion The changes in gene expression indicate that the small intestine rapidly looses mass during fasting to generate lactate or glucose and ketone bodies. Meanwhile, intestinal architecture is maintained by downregulation of cell turnover. Keywords: fasting response, time course
2007-11-12 | GSE8019 | GEO
Project description:The CREB binding protein inhibitor ICG-001 suppresses pancreatic cancer growth We used microarrays to detail global gene expression changes in the pancreatic cancer cell line AsPC1 following treatment with ICG-001 or siRNA-mediated knockdown of CTNNB1 (beta-catenin) AsPC1 cells were treated with 10uM ICG-001 or vehicle control (DMSO) for either 6 hours or 24 hours. AsPC-1 cells were also separately transfected with 20nM control siRNA or CTNNB1 siRNA for 48 hours. RNA was extracted at these time points for hybridization to Affymetrix microarrays
2014-07-15 | E-GEOD-57728 | biostudies-arrayexpress
Project description:Delayed access to feed following hatching has been associated with reduction in growth performance in chicks. However, the gene networks within the hypothalamus that regulate feed intake and metabolism, and the effects of fasting on those pathways are not well understood. The present experiment evaluated global hypothalamic gene expression in neonatal chicks using the Arizona Gallus gallus 20.7K Oligo Array v1.0 to elucidate genes and pathways regulated by feeding, fasting, and refeeding. Ten groups of chicks were sampled over four days post-hatch, including: control (at hatch), 24h fed, 24h fasted, 48h fed, 48h fasted, 48h fasted then 4h refed, 72h fed, 48h fasted then 24h refed, 96h fed, and 48h fasted then 48h refed. Non-esterified fatty acids were elevated, and triglycerides were decreased in fasted chicks at 24 and 48h, indicating that fasting induced physiological changes. Hypothalamic samples were collected from 16 chicks per group, and total RNA was extracted and pooled for hybridization (n=4). Expression patterns of selected genes were confirmed by quantitative real-time PCR. Two-fold differences in gene expression were detected in 1,272 genes between treatment groups, and of those, 119 genes were significantly (P<0.05) different. Assignment of gene ontology terms to the significant genes resulted in 34 different categories of biological processes, with 24% of genes participating in signal transduction, transport, or metabolic processes. Genes that were upregulated during fasting (and confirmed by qPCR) include FK506BP51, which is involved in the formation of steroid receptor complexes, and deiodinase type II, which is responsible for converting the thyroid hormone T4 into T3. Confirmed genes, downregulated due to fasting, included proopiomelanocortin and fatty acid binding protein 7. Other regulated genes were identified that play a role in feeding and obesity in other species, such as relaxin 3 and adrenergic receptor-β2. Further analysis of differentially regulated genes could provide new information regarding the role of the hypothalamus in feed intake and metabolism. Keywords: Metabolic perturbation
2008-10-18 | GSE13257 | GEO
Project description:Grainyhead genes are involved in wound healing and developmental neural tube closure. In light of the high degree of similarity between the epithelial-mesenchymal transitions (EMTs) occurring in wound healing processes and the cancer stem cell-like compartment of tumors, including TGF-β-dependence, we investigated the role of a Grainyhead gene (GRHL2) in oncogenic EMT. Grainyhead was specifically down-regulated in the claudin-low subclass of mammary tumors and in the basal-B subclass of breast cancer cell lines. Functionally, GRHL2 suppressed TGF-β-induced, Twist-induced or spontaneous EMT, enhanced anoikis-sensitivity, and suppressed mammosphere generation in mammary epithelial cells. These effects were mediated, in part, by its suppression of ZEB1 expression, through direct repression of the ZEB1 promoter. GRHL2 also inhibited Smad-mediated transcription, and up-regulated mir200b/c as well as the TGF-β receptor antagonist, BMP2. The expression of GRHL2 in the breast cancer cell line MDA-MB-231 triggered a mesenchymal-to-epithelial transition and sensitized the cells to anoikis. These results indicate that GRHL2 is a suppressor of the oncogenic EMT. 3 biologic replicates for each cell line. Comparison of HMLE+Twist-ER cells expressing GRHL2/pMIG vs. HMLE+Twist-ER cells expressing empty pMIG.
2012-02-25 | E-GEOD-36081 | biostudies-arrayexpress