Rapid, high efficiency isolation of pancreatic ß-cells
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE64508 | GEO | 2015/09/04
SECONDARY ACCESSION(S): PRJNA271154
REPOSITORIES: GEO
Similar Datasets
Project description:The ability to isolate pure pancreatic ß-cells would greatly aid multiple areas of diabetes research. We developed an exendin-4-like neopeptide conjugate for the rapid purification and isolation of functional pancreatic ß-cells. By targeting the glucagon-like peptide-1 receptor, ß-cells were isolated within an hour and were >99% pure. These studies were confirmed by immunostaining, confocal microscopy and microarray analysis on isolated cells. Gene expression profiling studies of the cytofluorometrically sorted ß-cells provided new insights into the genetic programs at play of different ages and stages during type-1 diabetes development. The described isolation method should have broad applicability to the ß-cell field. Microarray profile of beta cells from isolated islets from 4 and 12 week old NOD mice. Cells were stained with 50 nM Ex4+ probe and sorted on FACS ARIA. DAPI and CD45+ cells were excluded.
2015-09-04 | E-GEOD-64508 | biostudies-arrayexpress
Project description:Glucagon and insulin are counter-regulatory pancreatic hormones that precisely control blood glucose homeostasis1. Type 2 diabetes mellitus (T2DM) is characterized by inappropriately elevated blood glucagon2-5 levels as well as insufficient glucose stimulated insulin secretion (GSIS) by pancreatic ß-cells6. Early in the pathogenesis of T2DM, hyperglucagonemia is observable antecedent to ß-cell dysfunction7-9; and in mice, liver-specific activation of glucagon receptor-dependent signaling results in impaired GSIS10. However, the mechanistic relationship between hyperglucagonemia, hepatic glucagon action, and ß-cell dysfunction remains poorly understood. Here we show that glucagon action stimulates hepatic production of the neuropeptide kisspeptin1, which acts in an endocrine manner on ß-cells to suppress GSIS. In vivo glucagon administration acutely stimulates hepatic kisspeptin1 production, and kisspeptin1 is increased in livers from humans with T2DM and from mouse models of diabetes mellitus. Synthetic kisspeptin1 potently suppresses GSIS in vivo and in vitro from normal isolated islets, which express the kisspeptin1 receptor Kiss1R. Administration of a Kiss1R antagonist in diabetic Leprdb/db mice potently augments GSIS and reduces glycemia. Our observations indicate in the pathogenesis of T2DM an endocrine mechanism sequentially linking hyperglucagonemia via hepatic kisspeptin1 production to impaired insulin secretion. In addition, our findings suggest Kiss1R antagonism as a therapeutic avenue to improve ß-cell function in T2DM. Total RNA from L-Δprkar1a KO mice compared to control D-glucose mice
2014-04-01 | E-GEOD-48815 | biostudies-arrayexpress
Project description:Glucagon and insulin are counter-regulatory pancreatic hormones that precisely control blood glucose homeostasis1. Type 2 diabetes mellitus (T2DM) is characterized by inappropriately elevated blood glucagon2-5 levels as well as insufficient glucose stimulated insulin secretion (GSIS) by pancreatic ß-cells6. Early in the pathogenesis of T2DM, hyperglucagonemia is observable antecedent to ß-cell dysfunction7-9; and in mice, liver-specific activation of glucagon receptor-dependent signaling results in impaired GSIS10. However, the mechanistic relationship between hyperglucagonemia, hepatic glucagon action, and ß-cell dysfunction remains poorly understood. Here we show that glucagon action stimulates hepatic production of the neuropeptide kisspeptin1, which acts in an endocrine manner on ß-cells to suppress GSIS. In vivo glucagon administration acutely stimulates hepatic kisspeptin1 production, and kisspeptin1 is increased in livers from humans with T2DM and from mouse models of diabetes mellitus. Synthetic kisspeptin1 potently suppresses GSIS in vivo and in vitro from normal isolated islets, which express the kisspeptin1 receptor Kiss1R. Administration of a Kiss1R antagonist in diabetic Leprdb/db mice potently augments GSIS and reduces glycemia. Our observations indicate in the pathogenesis of T2DM an endocrine mechanism sequentially linking hyperglucagonemia via hepatic kisspeptin1 production to impaired insulin secretion. In addition, our findings suggest Kiss1R antagonism as a therapeutic avenue to improve ß-cell function in T2DM.
2014-04-01 | GSE48815 | GEO
Project description:Diabetes mellitus results from an inadequately functioning beta-cell mass. In the adult pancreas, beta-cell mass is dynamic, increasing to meet metabolic demands and decreasing with metabolic or injury insults. Exendin-4 (Ex-4) is a glucagon-like peptide-1 receptor agonist that augments beta-cell mass by increasing beta-cell neogenesis and proliferation and by reducing apoptosis. We utilized a cDNA microarray approach to identify genes that are differentially regulated during islet growth after Ex-4 treatment or a partial pancreatectomy (Ppx). Mice underwent 50% Ppx or sham operation and received Ex-4 or vehicle every 24 hours. cDNA prepared from total pancreatic RNA isolated at 12, 24 and 48 hrs after surgery was hybridized to the PancChip 4.0 microarray.
2005-12-01 | E-CBIL-5 | biostudies-arrayexpress
Project description:Pancreatic ß-cells adapt to compensate for the increased metabolic demand during insulin resistance. While the microRNA pathway has an essential role in the expansion of ß-cell mass, the extent of its contribution is unclear. Here we show that miR-184 is silenced in the pancreatic islets of several insulin-resistant mouse models and in the islets of type-2 diabetic human subjects. Reduction of miR-184 promotes the expression of its target Argonaute2 (Ago2), a component of the microRNA-induced silencing complex. While over-expression of Ago2 increased ß-cell proliferation, conditional deletion decreased ß-cell number. Moreover, restored expression of miR-184 in leptin-deficient ob/ob mice decreased Ago2 and prevented compensatory ß-cell expansion. Loss of Ago2 expression during insulin resistance blocked ß-cell growth and relieved the regulation of miR-375-targeted genes including the growth suppressor Cadm1. This study identifies the regulation of Ago2 by miR-184 as an essential component of the compensatory response to promote proliferation during insulin resistance.
2014-02-06 | GSE46623 | GEO
Project description:Rapid, high efficiency isolation of pancreatic ß-cells
| PRJNA271154 | ENA
Project description:Insulin-secreting β cells and glucagon-secreting α cells maintain physiological blood glucose levels, and their malfunction drives diabetes development. Using ChIP sequencing and RNA sequencing analysis, we determined the epigenetic and transcriptional landscape of human pancreatic α, β, and exocrine cells. We found that, compared with exocrine and β cells, differentiated α cells exhibited many more genes bivalently marked by the activating H3K4me3 and repressing H3K27me3 histone modifications. This was particularly true for β cell signature genes involved in transcriptional regulation. Remarkably, thousands of these genes were in a monovalent state in β cells, carrying only the activating or repressing mark. Our epigenomic findings suggested that α to β cell reprogramming could be promoted by manipulating the histone methylation signature of human pancreatic islets. Indeed, we show that treatment of cultured pancreatic islets with a histone methyltransferase inhibitor leads to colocalization of both glucagon and insulin and glucagon and insulin promoter factor 1 (PDX1) in human islets and colocalization of both glucagon and insulin in mouse islets. Thus, mammalian pancreatic islet cells display cell-type–specific epigenomic plasticity, suggesting that epigenomic manipulation could provide a path to cell reprogramming and novel cell replacement-based therapies for diabetes. Pancreatic islets were collected post-mortem from 6 human donors and subjected to FACS to separate populations of alpha, beta, and exocrine cells. Depending on the availability of resulting material, sorted islet cell populations were used for H3K4me3, H3K27me3 ChIP-seq, or RNA-seq analysis. All ChIP-seq samples have a corresponding input from the same sample.
2013-08-28 | E-GEOD-50386 | biostudies-arrayexpress
Project description:We define a pathogenic role for a ß-catenin-activated genetic pathway in murine renal dysplasia. Cre-mediated stabilization of ß-catenin in the ureteric cell lineage prior to the onset of kidney development increased ß-catenin levels and caused renal aplasia or severe hypodysplasia. A genome-wide analysis of mRNA expression in dysplastic tissue identified down-regulation of genes required for ureteric branching and up regulation of Tgfß2 and Dkk1. Hoxb7-Cre:EGFP mice ( Zhao, et al. (2004) Dev Biol 276:403-415) were crossed with mice containing loxP sites flanking exon 3 of the ß-catenin allele (ß-catdelta3/delta3) (Harada,et al. (2002) Cancer Res 62:1971-1977) to generate ß-catenin gain-of-function mutant mice specific to the uteric bud, termed ß-catGOF-UB .Eighteen ß-catGOF-UB mutant kidneys and 9 WT kidneys were micro-dissected at E12.5. Mutant kidneys were divided into three random pools (n=3) consisting of 6 kidneys each and mRNA expression assessed by microarray.
2011-03-28 | E-GEOD-20325 | biostudies-arrayexpress
Project description:Glucagon serves as an important regulatory hormone for regulating blood glucose concentrations with tight feedback control with insulin and glucose. There are critical gaps in our understanding of glucagon kinetics, pancreatic α cell function and intra-islet feedback network that are disrupted in type 1 diabetes. This is important for translational research applications of evolving dual-hormone (insulin+glucagon) closed-loop artificial pancreas algorithms and their usage in type 1 diabetes. Thus, it is important to accurately measure glucagon kinetics in vivo and to develop robust models of glucose-insulin-glucagon interplay that could inform next generation of artificial pancreas algorithms.
2022-06-09 | PXD028715 | Pride
Project description:Insulin-secreting β cells and glucagon-secreting α cells maintain physiological blood glucose levels, and their malfunction drives diabetes development. Using ChIP sequencing and RNA sequencing analysis, we determined the epigenetic and transcriptional landscape of human pancreatic α, β, and exocrine cells. We found that, compared with exocrine and β cells, differentiated α cells exhibited many more genes bivalently marked by the activating H3K4me3 and repressing H3K27me3 histone modifications. This was particularly true for β cell signature genes involved in transcriptional regulation. Remarkably, thousands of these genes were in a monovalent state in β cells, carrying only the activating or repressing mark. Our epigenomic findings suggested that α to β cell reprogramming could be promoted by manipulating the histone methylation signature of human pancreatic islets. Indeed, we show that treatment of cultured pancreatic islets with a histone methyltransferase inhibitor leads to colocalization of both glucagon and insulin and glucagon and insulin promoter factor 1 (PDX1) in human islets and colocalization of both glucagon and insulin in mouse islets. Thus, mammalian pancreatic islet cells display cell-type–specific epigenomic plasticity, suggesting that epigenomic manipulation could provide a path to cell reprogramming and novel cell replacement-based therapies for diabetes.
2013-08-28 | GSE50386 | GEO