Effects of palmitate on genome-wide mRNA expression and DNA methylation patterns in human pancreatic islets

BMC Medicine

Charlotte Ling²  Marloes Dekker Nitert¹  Tina Rönn²  Karl Bacos²  Tasnim Dayeh²  Petr Volkov²  Elin Hall² 

[1] School of Medicine, Royal Brisbane Clinical School, The University of Queensland, Herston, QLD 4006, Australia;Epigenetics and Diabetes Unit, Department of Clinical Sciences, Lund University Diabetes Centre, CRC, Lund University, Scania University Hospital, Malmö, Sweden

关键词: Epigenetics;    Insulin secretion;    mRNA expression;    DNA methylation;    Lipotoxicity;    Type 2 diabetes;    Human pancreatic islets;    Palmitate;   

Others  :  840797
DOI  :  10.1186/1741-7015-12-103

 received in 2013-12-04, accepted in 2014-04-25,  发布年份 2014

【 摘 要 】

Background

Circulating free fatty acids are often elevated in patients with type 2 diabetes (T2D) and obese individuals. Chronic exposure to high levels of saturated fatty acids has detrimental effects on islet function and insulin secretion. Altered gene expression and epigenetics may contribute to T2D and obesity. However, there is limited information on whether fatty acids alter the genome-wide transcriptome profile in conjunction with DNA methylation patterns in human pancreatic islets. To dissect the molecular mechanisms linking lipotoxicity to impaired insulin secretion, we investigated the effects of a 48 h palmitate treatment in vitro on genome-wide mRNA expression and DNA methylation patterns in human pancreatic islets.

Methods

Genome-wide mRNA expression was analyzed using Affymetrix GeneChip® Human Gene 1.0 ST whole transcript-based array (n = 13) and genome-wide DNA methylation was analyzed using Infinium HumanMethylation450K BeadChip (n = 13) in human pancreatic islets exposed to palmitate or control media for 48 h. A non-parametric paired Wilcoxon statistical test was used to analyze mRNA expression. Apoptosis was measured using Apo-ONE® Homogeneous Caspase-3/7 Assay (n = 4).

Results

While glucose-stimulated insulin secretion was decreased, there was no significant effect on apoptosis in human islets exposed to palmitate. We identified 1,860 differentially expressed genes in palmitate-treated human islets. These include candidate genes for T2D, such as TCF7L2, GLIS3, HNF1B and SLC30A8. Additionally, genes in glycolysis/gluconeogenesis, pyruvate metabolism, fatty acid metabolism, glutathione metabolism and one carbon pool by folate were differentially expressed in palmitate-treated human islets. Palmitate treatment altered the global DNA methylation level and DNA methylation levels of CpG island shelves and shores, 5′UTR, 3′UTR and gene body regions in human islets. Moreover, 290 genes with differential expression had a corresponding change in DNA methylation, for example, TCF7L2 and GLIS3. Importantly, out of the genes differentially expressed due to palmitate treatment in human islets, 67 were also associated with BMI and 37 were differentially expressed in islets from T2D patients.

Conclusion

Our study demonstrates that palmitate treatment of human pancreatic islets gives rise to epigenetic modifications that together with altered gene expression may contribute to impaired insulin secretion and T2D.

【 授权许可】

   
2014 Hall et al.; licensee BioMed Central Ltd.

【 图 表 】

Figure 1.

Figure 2.

Figure 3.

Figure 4.

Figure 5.

Figure 6.

Figure 7.

【 参考文献 】

• [1]McCarthy MI: Genomics, type 2 diabetes, and obesity. N Engl J Med 2010, 363:2339-2350.
• [2]Voight BF, Scott LJ, Steinthorsdottir V, Morris AP, Dina C, Welch RP, Zeggini E, Huth C, Aulchenko YS, Thorleifsson G, McCulloch LJ, Ferreira T, Grallert H, Amin N, Wu G, Willer CJ, Raychaudhuri S, McCarroll SA, Langenberg C, Hofmann OM, Dupuis J, Qi L, Segrè AV, van Hoek M, Navarro P, Ardlie K, Balkau B, Benediktsson R, Bennett AJ, Blagieva R, et al.: Twelve type 2 diabetes susceptibility loci identified through large-scale association analysis. Nat Genet 2010, 42:579-589.
• [3]Ling C, Groop L: Epigenetics: a molecular link between environmental factors and type 2 diabetes. Diabetes 2009, 58:2718-2725.
• [4]Yang BT, Dayeh TA, Kirkpatrick CL, Taneera J, Kumar R, Groop L, Wollheim CB, Dekker Nitert M, Ling C: Insulin promoter DNA methylation correlates negatively with insulin gene expression and positively with HbA(1c) levels in human pancreatic islets. Diabetologia 2011, 54:360-367.
• [5]Yang BT, Dayeh TA, Volkov PA, Kirkpatrick CL, Malmgren S, Jing X, Renstrom E, Wollheim CB, Dekker Nitert M, Ling C: Increased DNA methylation and decreased expression of PDX-1 in pancreatic islets from patients with type 2 diabetes. Mol Endocrinol 2012, 26:1203-1212.
• [6]Ling C, Del Guerra S, Lupi R, Rönn T, Granhall C, Luthman H, Masiello P, Marchetti P, Groop L, Del Prato S: Epigenetic regulation of PPARGC1A in human type 2 diabetic islets and effect on insulin secretion. Diabetologia 2008, 51:615-622.
• [7]Dayeh TA, Olsson AH, Volkov P, Almgren P, Rönn T, Ling C: Identification of CpG-SNPs associated with type 2 diabetes and differential DNA methylation in human pancreatic islets. Diabetologia 2013, 56:1036-1046.
• [8]Nitert MD, Dayeh T, Volkov P, Elgzyri T, Hall E, Nilsson E, Yang BT, Lang S, Parikh H, Wessman Y, Weishaupt H, Attema J, Abels M, Wierup N, Almgren P, Jansson PA, Rönn T, Hansson O, Eriksson KF, Groop L, Ling C: Impact of an exercise intervention on DNA methylation in skeletal muscle from first-degree relatives of patients with type 2 diabetes. Diabetes 2012, 61:3322-3332.
• [9]Ribel-Madsen R, Fraga MF, Jacobsen S, Bork-Jensen J, Lara E, Calvanese V, Fernandez AF, Friedrichsen M, Vind BF, Hojlund K, Beck-Nielsen H, Esteller M, Vaag A, Poulsen P: Genome-wide analysis of DNA methylation differences in muscle and fat from monozygotic twins discordant for type 2 diabetes. PLoS One 2012, 7:e51302.
• [10]Volkmar M, Dedeurwaerder S, Cunha DA, Ndlovu MN, Defrance M, Deplus R, Calonne E, Volkmar U, Igoillo-Esteve M, Naamane N, Del Guerra S, Masini M, Bugliani M, Marchetti P, Cnop M, Eizirik DL, Fuks F: DNA methylation profiling identifies epigenetic dysregulation in pancreatic islets from type 2 diabetic patients. EMBO J 2012, 31:1405-1426.
• [11]Barres R, Osler ME, Yan J, Rune A, Fritz T, Caidahl K, Krook A, Zierath JR: Non-CpG methylation of the PGC-1alpha promoter through DNMT3B controls mitochondrial density. Cell Metab 2009, 10:189-198.
• [12]Boden G, Shulman GI: Free fatty acids in obesity and type 2 diabetes: defining their role in the development of insulin resistance and beta-cell dysfunction. Eur J Clin Invest 2002, 32:14-23.
• [13]Kahn SE, Hull RL, Utzschneider KM: Mechanisms linking obesity to insulin resistance and type 2 diabetes. Nature 2006, 444:840-846.
• [14]Maedler K, Oberholzer J, Bucher P, Spinas GA, Donath MY: Monounsaturated fatty acids prevent the deleterious effects of palmitate and high glucose on human pancreatic beta-cell turnover and function. Diabetes 2003, 52:726-733.
• [15]Morgan NG: Fatty acids and beta-cell toxicity. Curr Opin Clin Nutr Metab Care 2009, 12:117-122.
• [16]Cnop M, Hannaert JC, Hoorens A, Eizirik DL, Pipeleers DG: Inverse relationship between cytotoxicity of free fatty acids in pancreatic islet cells and cellular triglyceride accumulation. Diabetes 2001, 50:1771-1777.
• [17]Zhou YP, Grill VE: Long-term exposure of rat pancreatic islets to fatty acids inhibits glucose-induced insulin secretion and biosynthesis through a glucose fatty acid cycle. J Clin Invest 1994, 93:870-876.
• [18]Olofsson CS, Collins S, Bengtsson M, Eliasson L, Salehi A, Shimomura K, Tarasov A, Holm C, Ashcroft F, Rorsman P: Long-term exposure to glucose and lipids inhibits glucose-induced insulin secretion downstream of granule fusion with plasma membrane. Diabetes 2007, 56:1888-1897.
• [19]Carpentier A, Mittelman SD, Lamarche B, Bergman RN, Giacca A, Lewis GF: Acute enhancement of insulin secretion by FFA in humans is lost with prolonged FFA elevation. Am J Physiol 1999, 276:E1055-E1066.
• [20]Leung N, Sakaue T, Carpentier A, Uffelman K, Giacca A, Lewis GF: Prolonged increase of plasma non-esterified fatty acids fully abolishes the stimulatory effect of 24 hours of moderate hyperglycaemia on insulin sensitivity and pancreatic beta-cell function in obese men. Diabetologia 2004, 47:204-213.
• [21]Busch AK, Cordery D, Denyer GS, Biden TJ: Expression profiling of palmitate- and oleate-regulated genes provides novel insights into the effects of chronic lipid exposure on pancreatic beta-cell function. Diabetes 2002, 51:977-987.
• [22]Malmgren S, Spégel P, Danielsson AP, Nagorny CL, Andersson LE, Dekker Nitert M, Ridderstråle M, Mulder H, Ling C: Coordinate changes in histone modifications, mRNA levels and metabolite profiles in clonal INS-1 832/13 beta-cells accompany functional adaptations to lipotoxicity. J Biol Chem 2013, 288:11973-11987.
• [23]Woods SC, Lutz TA, Geary N, Langhans W: Pancreatic signals controlling food intake; insulin, glucagon and amylin. Philos Trans R Soc Lond B Biol Sci 2006, 361:1219-1235.
• [24]Igoillo-Esteve M, Marselli L, Cunha DA, Ladrière L, Ortis F, Grieco FA, Dotta F, Weir GC, Marchetti P, Eizirik DL, Cnop M: Palmitate induces a pro-inflammatory response in human pancreatic islets that mimics CCL2 expression by beta cells in type 2 diabetes. Diabetologia 2010, 53:1395-1405.
• [25]Cunha DA, Igoillo-Esteve M, Gurzov EN, Germano CM, Naamane N, Marhfour I, Fukaya M, Vanderwinden JM, Gysemans C, Mathieu C, Marselli L, Marchetti P, Harding HP, Ron D, Eizirik DL, Cnop M: Death protein 5 and p53-upregulated modulator of apoptosis mediate the endoplasmic reticulum stress-mitochondrial dialog triggering lipotoxic rodent and human beta-cell apoptosis. Diabetes 2012, 61:2763-2775.
• [26]Lefebvre B, Vandewalle B, Balavoine AS, Queniat G, Moerman E, Vantyghem MC, LeBacquer O, Gmyr V, Pawlowski V, Kerr-Conte J, Pattou F: Regulation and functional effects of ZNT8 in human pancreatic islets. J Endocrinol 2012, 214:225-232.
• [27]Natalicchio A, Labarbuta R, Tortosa F, Biondi G, Marrano N, Peschechera A, Carchia E, Orlando MR, Leonardini A, Cignarelli A, Marchetti P, Perrini S, Laviola L, Giorgino F: Exendin-4 protects pancreatic beta cells from palmitate-induced apoptosis by interfering with GPR40 and the MKK4/7 stress kinase signalling pathway. Diabetologia 2013, 56:2456-2466.
• [28]Ghanaat-Pour H, Sjöholm Å: Exenatide and pioglitazone regulate fatty acid-induced gene expression in normal and diabetic human islets. Metabolomics 2011, 1:102.
• [29]Cnop M, Abdulkarim B, Bottu G, Cunha DA, Igoillo-Esteve M, Masini M, Turatsinze JV, Griebel T, Villate O, Santin I, Bugliani M, Ladriere L, Marselli L, McCarthy MI, Marchetti P: Sammeth M. Eizirik DL: RNA-sequencing identifies dysregulation of the human pancreatic islet transcriptome by the saturated fatty acid palmitate. Diabetes; 2013. [Epub ahead of print]
• [30]Sargsyan E, Bergsten P: Lipotoxicity is glucose-dependent in INS-1E cells but not in human islets and MIN6 cells. Lipids Health Dis 2011, 10:115. BioMed Central Full Text
• [31]Carlsson M, Wessman Y, Almgren P, Groop L: High levels of nonesterified fatty acids are associated with increased familial risk of cardiovascular disease. Arterioscler Thromb Vasc Biol 2000, 20:1588-1594.
• [32]Irizarry RA, Bolstad BM, Collin F, Cope LM, Hobbs B, Speed TP: Summaries of Affymetrix GeneChip probe level data. Nucleic Acids Res 2003, 31:e15.
• [33]Bibikova M, Barnes B, Tsan C, Ho V, Klotzle B, Le JM, Delano D, Zhang L, Schroth GP, Gunderson KL, Fan JB, Shen R: High density DNA methylation array with single CpG site resolution. Genomics 2011, 98:288-295.
• [34]Teschendorff AE, Menon U, Gentry-Maharaj A, Ramus SJ, Gayther SA, Apostolidou S, Jones A, Lechner M, Beck S, Jacobs IJ, Widschwendter M: An epigenetic signature in peripheral blood predicts active ovarian cancer. PLoS One 2009, 4:e8274.
• [35]Du P, Zhang X, Huang CC, Jafari N, Kibbe WA, Hou L, Lin SM: Comparison of Beta-value and M-value methods for quantifying methylation levels by microarray analysis. BMC Bioinforma 2010, 11:587. BioMed Central Full Text
• [36]Du P, Kibbe WA, Lin SM: Lumi: a pipeline for processing Illumina microarray. Bioinformatics 2008, 24:1547-1548.
• [37]Bolstad BM, Irizarry RA, Astrand M, Speed TP: A comparison of normalization methods for high density oligonucleotide array data based on variance and bias. Bioinformatics 2003, 19:185-193.
• [38]Chen C, Grennan K, Badner J, Zhang D, Gershon E, Jin L, Liu C: Removing batch effects in analysis of expression microarray data: an evaluation of six batch adjustment methods. PLoS One 2011, 6:e17238.
• [39]Smyth GK: Limma: linear models for microarray data. In Bioinformatics and Computational Biology Solutions Using R and Bioconductor. New York: Springer; 2005:397-420.
• [40]Zhang B, Kirov S, Snoddy J: WebGestalt: an integrated system for exploring gene sets in various biological contexts. Nucleic Acids Res 2005, 33:W741-W748.
• [41]WEB-based GEne SeT AnaLysis Toolkit: translating gene lists into biological insights [http://bioinfo.vanderbilt.edu/webgestalt/ webcite]
• [42]Scholz H, Lund T, Dahle MK, Collins JL, Korsgren O, Wang JE, Foss A: The synthetic liver X receptor agonist GW3965 reduces tissue factor production and inflammatory responses in human islets in vitro. Diabetologia 2009, 52:1352-1362.
• [43]National Human Genome Research Institute [http://www.genome.gov/gwastudies webcite]
• [44]Hindorff LA, Sethupathy P, Junkins HA, Ramos EM, Mehta JP, Collins FS, Manolio TA: Potential etiologic and functional implications of genome-wide association loci for human diseases and traits. Proc Natl Acad Sci U S A 2009, 106:9362-9367.
• [45]Chen YA, Lemire M, Choufani S, Butcher DT, Grafodatskaya D, Zanke BW, Gallinger S, Hudson TJ, Weksberg R: Discovery of cross-reactive probes and polymorphic CpGs in the Illumina Infinium HumanMethylation450 microarray. Epigenetics 2013, 8:203-209.
• [46]Kankaanpaa M, Lehto HR, Parkka JP, Komu M, Viljanen A, Ferrannini E, Knuuti J, Nuutila P, Parkkola R, Iozzo P: Myocardial triglyceride content and epicardial fat mass in human obesity: relationship to left ventricular function and serum free fatty acid levels. J Clin Endocrinol Metab 2006, 91:4689-4695.
• [47]Clore JN, Allred J, White D, Li J, Stillman J: The role of plasma fatty acid composition in endogenous glucose production in patients with type 2 diabetes mellitus. Metabolism 2002, 51:1471-1477.
• [48]Dayeh T, Volkov P, Salö S, Hall E, Nilsson E, Olsson AH, Kirkpatrick CL, Wollheim CB, Eliasson L, Rönn T, Bacos K, Ling C: Genome-wide DNA methylation analysis of human pancreatic islets from type 2 diabetic and non-diabetic donors identifies candidate genes that influence insulin secretion. PLoS Genet 2014, 10:e1004160.
• [49]Rakyan VK, Beyan H, Down TA, Hawa MI, Maslau S, Aden D, Daunay A, Busato F, Mein CA, Manfras B, Dias KR, Bell CG, Tost J, Boehm BO, Beck S, Leslie RD: Identification of type 1 diabetes-associated DNA methylation variable positions that precede disease diagnosis. PLoS Genet 2011, 7:e1002300.
• [50]Toperoff G, Aran D, Kark JD, Rosenberg M, Dubnikov T, Nissan B, Wainstein J, Friedlander Y, Levy-Lahad E, Glaser B, Hellman A: Genome-wide survey reveals predisposing diabetes type 2-related DNA methylation variations in human peripheral blood. Hum Mol Genet 2012, 21:371-383.
• [51]Nogueira TC, Paula FM, Villate O, Colli ML, Moura RF, Cunha DA, Marselli L, Marchetti P, Cnop M, Julier C, Eizirik DL: GLIS3, a susceptibility gene for type 1 and type 2 diabetes, modulates pancreatic beta cell apoptosis via regulation of a splice variant of the BH3-Only protein bim. PLoS Genet 2013, 9:e1003532.
• [52]Jones PA: Functions of DNA methylation: islands, start sites, gene bodies and beyond. Nat Rev Genet 2012, 13:484-492.
• [53]Taqi MM, Bazov I, Watanabe H, Sheedy D, Harper C, Alkass K, Druid H, Wentzel P, Nyberg F, Yakovleva T, Bakalkin G: Prodynorphin CpG-SNPs associated with alcohol dependence: elevated methylation in the brain of human alcoholics. Addict Biol 2011, 16:499-509.
• [54]Shukla S, Kavak E, Gregory M, Imashimizu M, Shutinoski B, Kashlev M, Oberdoerffer P, Sandberg R, Oberdoerffer S: CTCF-promoted RNA polymerase II pausing links DNA methylation to splicing. Nature 2011, 479:74-79.
• [55]Whitaker RC, Wright JA, Pepe MS, Seidel KD, Dietz WH: Predicting obesity in young adulthood from childhood and parental obesity. N Engl J Med 1997, 337:869-873.
• [56]Köbberling J, Tillil H: Empirical risk figures for first-degree relatives of non-insulin dependent diabetics. In The Genetics of Diabetes Mellitus. London: Academic Press; 1982:201-209.
• [57]Park JH, Stoffers DA, Nicholls RD, Simmons RA: Development of type 2 diabetes following intrauterine growth retardation in rats is associated with progressive epigenetic silencing of Pdx1. J Clin Invest 2008, 118:2316-2324.
• [58]Pinney SE, Jaeckle Santos LJ, Han Y, Stoffers DA, Simmons RA: Exendin-4 increases histone acetylase activity and reverses epigenetic modifications that silence Pdx1 in the intrauterine growth retarded rat. Diabetologia 2011, 54:2606-2614.
• [59]Thompson RF, Fazzari MJ, Niu H, Barzilai N, Simmons RA, Greally JM: Experimental intrauterine growth restriction induces alterations in DNA methylation and gene expression in pancreatic islets of rats. J Biol Chem 2010, 285:15111-15118.
• [60]Rakyan VK, Down TA, Balding DJ, Beck S: Epigenome-wide association studies for common human diseases. Nat Rev Genet 2011, 12:529-541.
• [61]Hall E, Dayeh T, Kirkpatrick CL, Wollheim CB, Dekker Nitert M, Ling C: DNA methylation of the glucagon-like peptide 1 receptor (GLP1R) in human pancreatic islets. BMC Med Genet 2013, 14:76.
• [62]Koeck T, Olsson AH, Dekker Nitert M, Sharoyko VV, Ladenvall C, Kotova O, Reiling E, Rönn T, Parikh H, Taneera J, Eriksson JG, Metodiev MD, Larsson NG, Balhuizen A, Luthman H, Stančáková A, Kuusisto J, Laakso M, Poulsen P, Vaag A, Groop L, Lyssenko V, Mulder H, Ling C: A common variant in TFB1M is associated with reduced insulin secretion and increased future risk of type 2 diabetes. Cell Metab 2011, 13:80-91.
• [63]Butler AE, Janson J, Bonner-Weir S, Ritzel R, Rizza RA, Butler PC: Beta-cell deficit and increased beta-cell apoptosis in humans with type 2 diabetes. Diabetes 2003, 52:102-110.
• [64]Clark A, Jones LC, de Koning E, Hansen BC, Matthews DR: Decreased insulin secretion in type 2 diabetes: a problem of cellular mass or function? Diabetes 2001, 50:S169-S171.
• [65]Henquin JC, Rahier J: Pancreatic alpha cell mass in European subjects with type 2 diabetes. Diabetologia 2011, 54:1720-1725.
• [66]Leibiger IB, Leibiger B, Berggren PO: Insulin signaling in the pancreatic beta-cell. Annu Rev Nutr 2008, 28:233-251.
• [67]Hellemans KH, Hannaert JC, Denys B, Steffensen KR, Raemdonck C, Martens GA, Van Veldhoven PP, Gustafsson JA, Pipeleers D: Susceptibility of pancreatic beta cells to fatty acids is regulated by LXR/PPARalpha-dependent stearoyl-coenzyme A desaturase. PLoS One 2009, 4:e7266.
• [68]Busch AK, Gurisik E, Cordery DV, Sudlow M, Denyer GS, Laybutt DR, Hughes WE, Biden TJ: Increased fatty acid desaturation and enhanced expression of stearoyl coenzyme A desaturase protects pancreatic beta-cells from lipoapoptosis. Diabetes 2005, 54:2917-2924.
• [69]Locasale JW: Serine, glycine and one-carbon units: cancer metabolism in full circle. Nat Rev Cancer 2013, 13:572-583.
• [70]Taneera J, Lang S, Sharma A, Fadista J, Zhou Y, Ahlqvist E, Jonsson A, Lyssenko V, Vikman P, Hansson O, Parikh H, Korsgren O, Soni A, Krus U, Zhang E, Jing XJ, Esguerra JL, Wollheim CB, Salehi A, Rosengren A, Renström E, Groop L: A systems genetics approach identifies genes and pathways for type 2 diabetes in human islets. Cell Metab 2012, 16:122-134.
• [71]Watt MJ, Hoy AJ, Muoio DM, Coleman RA: Distinct roles of specific fatty acids in cellular processes: implications for interpreting and reporting experiments. Am J Physiol Endocrinol Metab 2012, 302:E1-E3.
• [72]Olsson AH, Yang BT, Hall E, Taneera J, Salehi A, Nitert MD, Ling C: Decreased expression of genes involved in oxidative phosphorylation in human pancreatic islets from patients with type 2 diabetes. Eur J Endocrinol 2011, 165:589-595.
• [73]Rönn T, Volkov P, Davegårdh C, Dayeh T, Hall E, Olsson AH, Nilsson E, Tornberg A, Dekker Nitert M, Eriksson KF, Jones HA, Groop L, Ling C: A six months exercise intervention influences the genome-wide DNA methylation pattern in human adipose tissue. PLoS Genet 2013, 9:e1003572.
• [74]Jacobsen SC, Brons C, Bork-Jensen J, Ribel-Madsen R, Yang B, Lara E, Hall E, Calvanese V, Nilsson E, Jorgensen SW, Mandrup S, Ling C, Fernandez AF, Fraga MF, Poulsen P, Vaag A: Effects of short-term high-fat overfeeding on genome-wide DNA methylation in the skeletal muscle of healthy young men. Diabetologia 2012, 55:3341-3349.