Unknown,Transcriptomics,Genomics,Proteomics

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Transcription profiling of brown adipose tissue in a glycerol kinase (Gyk) knockout mouse model to define GyKs role in insulin resistance

ABSTRACT: Glycerol kinase (GK) is at the interface of fat and carbohydrate metabolism and has been implicated in insulin resistance and type 2 diabetes mellitus (T2DM). To define GK's role in insulin resistance, we examined gene expression in brown adipose tissue in a glycerol kinase (Gyk) knockout (KO) mouse model using microarray analysis. Global gene expression profiles of KO mice were distinct from wild type (WT) with 668 genes that were differentially expressed. These included genes involved in lipid metabolism, carbohydrate metabolism, insulin signaling, and insulin resistance. Real-Time (RT) PCR analysis confirmed the differential expression of selected genes involved in lipid and carbohydrate metabolism. PathwayAssist analysis confirmed direct and indirect connections between GK and genes in lipid metabolism, carbohydrate metabolism, insulin signaling, and insulin resistance. Network Component Analysis (NCA) showed that the transcription factors, peroxisome proliferator-activated receptor gamma (PPAR-Î³), sterol regulatory element binding factor 1 (SREBP-1), SREBP-2, signal transducer and activator of transcription 3 (STAT3), STAT5, trans-acting transcription factor 1 (SP1), CCAAT/enhancer binding protein alpha (CEBP-Î±), cAMP responsive element binding protein 1 (CREB), glucocorticoid receptor (GR), and PPAR-Î± have altered activity in the KO mice. NCA also revealed the individual contribution of these transcription factors on the expression of genes altered in the microarray data. This study elucidates the transcription network of Gyk and further confirms a role for Gyk, a simple Mendelian disorder, in insulin resistance and T2DM, a common complex genetic disorder. Experiment Overall Design: 7 samples are analyzed; 3 wildtype and 4 KO. the WT samples are used as control and KO samples are used as the experimental group.

ORGANISM(S): Mus musculus

SUBMITTER: Lola Rahib

PROVIDER: E-GEOD-4656 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Glycerol kinase deficiency alters expression of genes involved in lipid metabolism, carbohydrate metabolism, and insulin signaling.

Rahib Lola L MacLennan Nicole K NK Horvath Steve S Liao James C JC Dipple Katrina M KM

European journal of human genetics : EJHG 20070404 6

Glycerol kinase (GK) is at the interface of fat and carbohydrate metabolism and has been implicated in insulin resistance and type 2 diabetes mellitus. To define GK's role in insulin resistance, we examined gene expression in brown adipose tissue in a glycerol kinase knockout (KO) mouse model using microarray analysis. Global gene expression profiles of KO mice were distinct from wild type with 668 differentially expressed genes. These include genes involved in lipid metabolism, carbohydrate met ...[more]

PMID: 17406644

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Project description:Glycerol kinase deficiency (GKD) is an X-linked inborn error of metabolism with metabolic and neurologic crises. Liver shows the highest level of glycerol kinase (GK) activity in humans and mice. Absence of genotype-phenotype correlations in patients with GKD indicate the involvement of modifier genes, including other network partners. To understand the molecular pathogenesis of GKD, we performed microarray analysis on liver mRNA from neonatal glycerol kinase (Gyk) knockout (KO) and wild type (WT) mice. Unsupervised learning revealed the overall gene expression profile of the KO mice was different from that of WT. Real time PCR confirmed differences for selected genes. Functional gene enrichment analysis was used to find 56 increased and 37 decreased gene functional categories. Pathway Assist analysis identified changes in gene expression levels of genes involved in organic acid metabolism indicating that GK was part of the same metabolic network which correlates well with the patients with GKD having metabolic acidemia during their episodic crises. Network component analysis (NCA) showed that transcription factors SREBP-1c, ChREBP, HNF-4alpha, and PPAR-alpha, had increased activity in the Gyk KO mice compared with WT mice; while SREBP-2 was less active in the Gyk KO mice. These studies show that Gyk deletion causes alterations in gene expression of genes in several regulatory networks and is the first time NCA has been used to expand on microarray data from a mouse knockout model of a human disease. Male WT and KO mouse pups were sacrificed on day of life (dol) 3 and each liver was harvested. Total RNA from 4 KO and 4 WT livers was isolated individually. cDNA was synthesized from the poly(A)+ mRNA in the total RNA, Biotin-tagged and fragmented to an average strand length of 100 bases (range 35-200 bases). Ten µg of each cRNA was hybridized onto an Affymetrix mus 430 2.0 GeneChip to analyze differences in liver gene expression between KO and WT mice. Day of life three was chosen because the mice are phenotypically symptomatic with statistically different parameters for hypoglycemia, acidosis; low bicarbonate and decreased base excess. On day of life 2 they are not significantly different from wild type in all of these important clinical phenotypes.

Project description:Development of insulin resistance is a key pathogenic component underlying metabolic syndrome and Type 2 diabetes (T2DM). Despite its importance, the molecular mechanisms underlying insulin resistance are poorly understood. Genome-wide association studies for T2DM and other metabolic traits have led to the identification of many candidate SNPs, but the majority of these SNPs are noncoding and determination of associated causal genes and/or specific tissue sites of action have been difficult. Adipocytes are critical regulators of mammalian metabolic homeostasis, with important effects on appetite, satiety, glucose and lipid homeostasis, energy expenditure, blood pressure, and immune function. Although insulin resistance (IR) in skeletal muscle, the major source of glucose disposal, is responsible for the bulk of the hyperglycemia observed in T2DM, muscle IR KO mice are generally healthy, IR also occurs in adipocytes, and inflammation within adipose tissue has been proposed as a primary mediator of IR, resulting in excess release of free fatty acids as well as alterations in adipokine release. Absence of adipose tissue, as observed in various lipodystrophies, or adipocyte-specific knockout of genes such as GLUT4 or insulin receptor, also alter systemic IR and T2DM risk. Transcriptional changes in adipocytes associated with metabolic disease. Despite the importance of adipocytes to metabolic disease, we have a poor understanding of how the adipocyte transcriptome changes in the disease state. Studies investigating transcriptional changes in whole adipose tissue have shown decreases in genes involved in adipogenesis, as well as alterations in inflammation, mitochondrial metabolism, lipid metabolism, detoxification, and insulin signaling. However, the vast majority of these studies use total adipose tissue samples, which does not allow for the exclusive evaluation of gene expression in mature adipocytes due to the substantial number of nonadipocyte cells (immune cells, fibroblasts, endothelial cells, pre-adipocytes, and mesenchymal cells) that also reside in adipose tissue. This is particularly relevant when studying metabolic disorders related to obesity-associated insulin resistance because these conditions are characterized by an increased influx of inflammatory cells into the adipose tissue.

Project description:Skeletal muscle mitochondrial dysfunction is secondary to T2DM and can be improved by long-term regular exercise training Mitochondrial dysfunction has long been implicated to play a causative role in development of type 2 diabetes (T2DM). However, a growing number of recent studies provide data that mitochondrial dysfunction is a consequence of T2DM development. The aim of our study is to clarify in further detail the causal role of mitochondrial dysfunction in T2DM by a comprehensive ex vivo analysis of mitochondrial function combined with global gene expression analysis in muscle of pre-diabetic newly diagnosed untreated T2DM subjects and long-standing insulin treated T2DM subjects compared with age- and BMI-matched controls. In addition, we assessed the impact of long-term interval exercise training on physical activity performance, mitochondrial function and glycemic control in long-standing insulin-treated T2DM subjects. Ex vivo mitochondrial density, quality and functioning was comparable between pre-diabetic subjects and matched controls, however, gene expression analysis showed a switch from carbohydrate toward lipids as energy source in pre-diabetes subjects. In contrast, long-term insulin treated T2DM subjects had slightly decreased mitochondrial density and ex vivo function. Expression of Krebs cycle and OXPHOS related genes were decreased, indicating a decreased capacity to use lipids as an energy source. The insulin-treated T2DM subjects had a lower physical activity level than pre-diabetic and normoglycemic subjects. A 52 weeks exercise training of these subjects increased submaximal oxidative efficiency, increased in vivo PCr recovery rate, as well as mildly increased in vitro mitochondrial function. Gene expression of β-oxidation, Krebs cycle and OXPHOS-related genes was increased. Our data demonstrate that mitochondrial dysfunction is rather a consequence than a causative factor in T2DM development as it was only detected in overt diabetes and not in early diabetes. Regular exercise training stabilized exogenous insulin requirement and improved mitochondrial functioning, fatty acid oxidation and general physical work load capacity in long-standing insulin-treated T2DM subjects. As such, the present study shows for the first time that long-term exercise interventions are beneficial in this group of complex diabetes patient and may prevent further metabolic deterioration. Insulin-treated T2DM subjects before and after 52 weeks of exercise training (T2DM_0 and T2DM_52), normoglycemic controls (NGT) and pre-diabetes subjects (IGT) and were selected. RNA was extracted from skeletal muscle biopsies and hybridized on Affymetrix microarrays.

Dataset Information

Transcription profiling of brown adipose tissue in a glycerol kinase (Gyk) knockout mouse model to define GyKs role in insulin resistance

Publications

Glycerol kinase deficiency alters expression of genes involved in lipid metabolism, carbohydrate metabolism, and insulin signaling.

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