Unknown,Transcriptomics,Genomics,Proteomics

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PPARM-NM-4 activation in human myotubes increases mitochondrial fatty acid oxidative capacity and reduces glucose utilization by a switch in substrate preference.

ABSTRACT: The role of peroxisome proliferator-activated receptor M-NM-4 (PPARM-NM-4) activation on global gene expression and mitochondrial fuel utilization were investigated in human myotubes. Only 21 genes were up-regulated and 3 genes were down-regulated after activation by the PPARM-NM-4 agonist GW501516. Pathway analysis showed up-regulated mitochondrial fatty acid oxidation, TCA cycle and cholesterol biosynthesis. GW501516 increased oleic acid oxidation and mitochondrial oxidative capacity by 2-fold. Glucose uptake and oxidation were reduced, but total substrate oxidation was not affected, indicating a fuel switch from glucose to fatty acid. Cholesterol biosynthesis was increased, but lipid biosynthesis and mitochondrial content were not affected. This study confirmed that the principal effect of PPARM-NM-4 activation was to increase mitochondrial fatty acid oxidative capacity. Our results further suggest that PPARM-NM-4 activation reduced glucose utilization through a switch in mitochondrial substrate preference by up-regulating pyruvate dehydrogenase kinase isozyme 4 and genes involved in lipid metabolism and fatty acid oxidation. Keywords: Expression profiling by array Human myotubes from four donors were exposed to a PPARM-NM-4 agonist or control for 96 h after which gene expression was profiled.

ORGANISM(S): Homo sapiens

SUBMITTER: Guido Hooiveld

PROVIDER: E-GEOD-40789 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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PPARδ activation in human myotubes increases mitochondrial fatty acid oxidative capacity and reduces glucose utilization by a switch in substrate preference.

Feng Yuan Z YZ Nikolić Nataša N Bakke Siril S SS Boekschoten Mark V MV Kersten Sander S Kase Eili T ET Rustan Arild C AC Thoresen G Hege GH

Archives of physiology and biochemistry 20130830 1

The role of peroxisome proliferator-activated receptor δ (PPARδ) activation on global gene expression and mitochondrial fuel utilization were investigated in human myotubes. Only 21 genes were up-regulated and 3 genes were down-regulated after activation by the PPARδ agonist GW501516. Pathway analysis showed up-regulated mitochondrial fatty acid oxidation, TCA cycle and cholesterol biosynthesis. GW501516 increased oleic acid oxidation and mitochondrial oxidative capacity by 2-fold. Glucose uptak ...[more]

PMID: 23991827

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Project description:Fatty acids comprise the primary energy source for the heart and are mainly taken up via hydrolysis of circulating triglyceride-rich lipoproteins. While most of the fatty acids entering the cardiomyocyte are oxidized, a small portion is involved in altering gene transcription to modulate cardiometabolic functions. So far, no in vivo model has been developed enabling study of the transcriptional effects of specific fatty acids in the intact heart. In the present study, mice were given a single oral dose of synthetic triglycerides composed of one single fatty acid. Hearts were collected 6h thereafter and used for whole genome gene expression profiling. Experiments were conducted in wild-type and PPARM-NM-1M-bM-^HM-^R/M-bM-^HM-^R mice to allow exploration of the specific contribution of PPARM-NM-1. It was found that: 1) linolenic acid (C18:3) had the most pronounced effect on cardiac gene expression. 2) The largest similarity in gene regulation was observed between linoleic acid (C18:2) and C18:3. Large similarity was also observed between the synthetic PPARM-NM-1 agonist Wy14,643 and docosahexaenoic acid (C22:6). 3) Many genes were regulated by one particular treatment only. Genes regulated by one particular treatment showed large functional divergence. 4) The majority of genes responding to fatty acid treatment were regulated in a PPARM-NM-1-dependent manner, emphasizing the importance of PPARM-NM-1 in mediating transcriptional regulation by fatty acids in the heart. 5) Several genes were robustly regulated by all or many of the fatty acids studied, mostly representing well-described targets of PPARs (e.g. Acot1, Angptl4, Ucp3). 6) Deletion and activation of PPARM-NM-1 had a major effect on expression of numerous genes involved in metabolism and immunity. Our analysis demonstrates the marked impact of dietary fatty acids on gene regulation in the heart via PPARM-NM-1. To study the long term transcriptional effects of PPARM-NM-1 activation, wild type and PPARM-NM-1M-bM-^HM-^R/M-bM-^HM-^R mice were fed a chow diet (RMH-B diet Arie Block, Woerden, the Netherlands) containing 0.1% (w/w) of the specific PPARM-NM-1 agonist Wy14,643. After 5days hearts were collected and used for whole genome gene expression profiling.

Project description:The transcription factor Peroxisome Proliferator-Activated Receptor M-NM-1 (PPARM-NM-1) is an important regulator of hepatic lipid metabolism. While PPARM-NM-1 is known to activate transcription of numerous genes, no comprehensive picture of PPARM-NM-1 binding to endogenous genes has yet been reported. To fill this gap, we performed ChIP-chip in combination with transcriptional profiling on HepG2 human hepatoma cells treated with the PPARM-NM-1 agonist GW7647. We found that GW7647 increased PPARM-NM-1 binding to 4220 binding regions. GW7647-induced binding regions showed a bias around the transcription start site and most contained a predicted PPAR binding motif. Several genes known to be regulated by PPARM-NM-1, such as ACOX1, SULT2A1, ACADL, CD36, IGFBP1 and G0S2, showed GW7647-induced PPARM-NM-1 binding to their promoter. A GW7647-induced PPARM-NM-1-binding region was also assigned to SREBP-targets HMGCS1, HMGCR, FDFT1, SC4MOL, and LPIN1, expression of which was induced by GW7647, suggesting cross-talk between PPARM-NM-1 and SREBP signaling. Our data furthermore demonstrate interaction between PPARM-NM-1 and STAT transcription factors in PPARM-NM-1-mediated transcriptional repression, and suggest interaction between PPARM-NM-1 and TBP and C/EBPM-NM-1 in PPARM-NM-1-mediated transcriptional activation. Overall, our analysis leads to important new insights into the mechanisms and impact of transcriptional regulation by PPARM-NM-1 in human liver and highlight the importance of cross-talk with other transcription factors. HepG2 cells were grown in phenol red-free DulbeccoM-bM-^@M-^Ys modified medium (DMEM) supplemented with 10% fetal bovine serum (FBS), 2 mM L-glutamine, 0.1 mg/ml streptomycin and 100 U/ml penicillin. Cells were split the day before experiments. Cells were kept at 37 M-BM-0C and 5% CO2. The following day, cells were treated with either 100 nM of the PPARM-NM-1 agonist GW7647 or control vehicle (DMSO). Cells used for gene expression analysis were harvested after 6 h or 24 h of GW7647 treatment. This submission represents the gene expression component of the study.

Project description:Dietary polyunsaturated fatty acids (PUFA) act as potent natural hypolipidemics and are linked to many health benefits in humans and in animal models. Mice fed long-term a high fat diet, in which medium-chain alpha linoleic acid (ALA) was partially replaced by long-chain docosahexaenoic (DHA) and eicosapentaenoic (EPA) fatty acids, showed reduced accumulation of body fat and prevention of insulin resistance, besides increased mitochondrial beta-oxidation in white adipose tissue and decreased plasma lipids. ALA, EPA and DHA all belong to PUFA of n-3 series. The intestine is a gatekeeper organ for ingested lipids. To examine the potential contribution of the intestine in the beneficial effects of EPA and DHA, this study assessed gene expression changes using whole genome microarray analysis on small intestinal scrapings. The main biological process affected was lipid metabolism. Fatty acid uptake, peroxisomal and mitochondrial beta-oxidation, and omega-oxidation of fatty acids were all increased. Quantitative real time PCR and intestinal fatty acid oxidation measurements ([14C(U)]-palmitate) confirmed significant gene expression differences in a dose-dependent manner. Furthermore, no major changes in the expression of lipid metabolism genes were observed in colonic scrapings. In conclusion, we show that marine n-3 fatty acids regulate small intestinal gene expression patterns. Since this organ contributes significantly to whole organism energy use, this adaptation of the small intestine may contribute to the complex and observed beneficial physiological effects of these natural compounds under conditions that will normally lead to development of obesity and diabetes. Experiment Overall Design: Male 4-month-old C57BL/6J mice were maintained for 4 weeks on semisynthetic high-fat (20% wt/wt) diets differing in the composition of n-3 PUFA. Two isocaloric diets were used (n=12): control sHFf diet which contained flax-seed oil (rich in ALA) as the only lipid source, or the sHFf-F2 diet, which had the same composition except that 44 % of the lipids were replaced by a n-3 PUFA concentrate (EPA&DHA) containing 6 % EPA and 51 % DHA (EPAX 1050TG; EPAX AS, Lysaker, Norway). Quality control of RNA samples showed that four samples did not pass quality thresholds, and these samples were excluded from array analysis (two control small intestine samples and one EPA&DHA small intestine sample). The remaining RNA samples were pooled per tissue per diet group. Thus; the flax-seed (control) array was hybridized with RNA pooled from of 10 mice, the PUFA array was hybridized with RNA pooled from 11 mice.

Project description:Aims/hypothesis: While lipid deposition in skeletal muscle is considered to be involved in obesity-associated insulin resistance, neutral intramyocellular lipid (IMCL) accumulation per se does not necessarily induce insulin resistance. We previously demonstrated that overexpression of the lipid droplet coat protein perilipin 2 augments intramyocellular lipid content while improving insulin sensitivity. Another member of the perilipin family, perilipin 5 (PLIN5), is predominantly expressed in oxidative tissues like skeletal muscle. Here we investigated the effects of PLIN5 overexpression M-bM-^@M-^S in comparison with effects of PLIN2 M-bM-^@M-^S on skeletal muscle lipid levels, gene expression profiles and insulin sensitivity. Methods: Gene electroporation was used to overexpress PLIN5 in tibialis anterior muscle of rats fed a high fat diet. Eight days after electroporation, insulin-mediated glucose uptake in skeletal muscle was measured by means of a hyperinsulinemic euglycemic clamp. Electron microscopy, fluorescence microscopy and lipid extractions were performed to investigate IMCL accumulation. Gene expression profiles were obtained using microarrays. Results: TAG storage and lipid droplet size increased upon PLIN5 overexpression. Despite the higher IMCL content, insulin sensitivity was not impaired and DAG and acylcarnitine levels were unaffected. In contrast to the effects of PLIN2 overexpression, microarray data analysis revealed a gene expression profile favoring FA oxidation and improved mitochondrial function. Conclusions/interpretation: Both PLIN2 and PLIN5 increase neutral IMCL content without impeding insulin-mediated glucose uptake. As opposed to the effects of PLIN2 overexpression, overexpression of PLIN5 in skeletal muscle promoted expression of a cluster of genes under control of PPARM-NM-1 and PGC1M-NM-1 involved in FA catabolism and mitochondrial oxidation. Rats received a high fat diet for 3 weeks; 2 weeks after start of the diet intervention Plin5 (OXPAT) or Plin2 (ADRP) were overexpressed in either the right or left tibialis anterior muscle. One week later pooled tibialis anterior muscle samples were analysed on microarrays.

Project description:Red meat consumption is associated with an increased colon cancer risk. Heme, present in red meat, injures the colon surface epithelium by luminal cytotoxicity and reactive oxygen species. This surface injury is overcompensated by hyperproliferation and hyperplasia of crypt cells. Transcriptome analysis of mucosa of heme-fed mice showed, besides stress- and proliferation-related genes, many upregulated lipid metabolism-related PPARM-NM-1 target genes. The aim of this study was to investigate the role of PPARM-NM-1 in heme-induced hyperproliferation and hyperplasia. Male PPARM-NM-1 KO and WT mice received a purified diet with or without heme. As PPARM-NM-1 is proposed to protect against oxidative stress and lipid peroxidation, we hypothesized that the absence of PPARM-NM-1 leads to more surface injury and crypt hyperproliferation in the colon upon heme-feeding. Heme induced luminal cytotoxicity and lipid peroxidation and colonic hyperproliferation and hyperplasia to the same extent in WT and KO mice. Transcriptome analysis of colonic mucosa confirmed similar heme-induced hyperproliferation in WT and KO mice. Stainings for alkaline phosphatase activity and expression levels of Vanin-1 and Nrf2-targets indicated a compromised antioxidant defense in heme-fed KO mice. Our results suggest that the protective role of PPARM-NM-1 in antioxidant defense involves the Nrf2-inhibitor Fosl1, which is upregulated by heme in PPARM-NM-1 KO mice. We conclude that PPARM-NM-1 plays a protective role in colon against oxidative stress, but PPARM-NM-1 does not mediate heme-induced hyperproliferation. This implies that oxidative stress of surface cells is not the main determinant of heme-induced hyperproliferation and hyperplasia. Wild type and peroxisome proliferator-activated receptor alpha (PPARM-NM-1) knockout mice were fed a Westernized high fat diet, or the same diet supplemented with 0.5 M-BM-5mol heme/g diet. After 14 days of intervention, mice were killed and gene expression was profiled in colon.

Dataset Information

PPARM-NM-4 activation in human myotubes increases mitochondrial fatty acid oxidative capacity and reduces glucose utilization by a switch in substrate preference.

Publications

PPARδ activation in human myotubes increases mitochondrial fatty acid oxidative capacity and reduces glucose utilization by a switch in substrate preference.

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