Influence of pre-pregnancy body mass index (p-BMI) and gestational weight gain (GWG) on DNA methylation and protein expression of obesogenic genes in umbilical vein

Autoři: Erika Chavira-Suárez aff001;  Angélica Jazmín Ramírez-Mendieta aff002;  Sofía Martínez-Gutiérrez aff002;  Paola Zárate-Segura aff003;  Jorge Beltrán-Montoya aff004;  Nidia Carolina Espinosa-Maldonado aff001;  Juan Carlos de la Cerda-Ángeles aff005;  Felipe Vadillo-Ortega aff001
Působiště autorů: Departamento de Bioquímica, Facultad de Medicina, Universidad Nacional Autónoma de México, Mexico City, Mexico aff001;  Unidad de Vinculación Científica de la Facultad de Medicina, Universidad Nacional Autónoma de México en el Instituto Nacional de Medicina Genómica, Mexico City, Mexico aff002;  Laboratorio de Medicina Traslacional, Escuela Superior de Medicina, Instituto Politécnico Nacional, Mexico City, Mexico aff003;  Instituto Nacional de Perinatología Isidro Espinosa de los Reyes, Mexico City, Mexico aff004;  Hospital General Dr. Enrique Cabrera, Secretaría de Salud CDMX, Mexico City, Mexico aff005
Vyšlo v časopise: PLoS ONE 14(12)
Kategorie: Research Article


Understanding the regulatory mechanisms that affect obesogenic genes expression in newborns is essential for early prevention efforts, but they remain unclear. Our study aimed to explore whether the maternal p-BMI and GWG were associated with regulatory single-locus DNA methylation in selected obesogenic genes. For this purpose, DNA methylation was assayed by Methylation-Sensitive High Resolution Melting (MS-HRM) technique and Sanger allele-bisulfite sequencing in fifty samples of umbilical vein to evaluate glucosamine-6-phosphate deaminase 2 (GNPDA2), peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC1α), and leptin receptor (LEPR) genes. Correlations between DNA methylation levels and indicators of maternal nutritional status were carried out. Western blotting was used to evaluate protein expression in extracts of the same samples. Results indicated that GNPDA2 and PGC1α genes have the same level of DNA methylation in all samples; however, a differential DNA methylation of LEPR gene promoter was found, correlating it with GWG and this correlation is unaffected by maternal age or unhealthy habits. Furthermore, leptin receptor (Lep-Rb) was upregulated in samples that showed the lowest levels of DNA methylation. This study highlights the association between poor GWG and adjustments on obesogenic genes expression in newborn tissues with potential consequences for development of obesity in the future.

Klíčová slova:

DNA methylation – leptin – Obesity – Pregnancy – Sequence alignment – Sequence databases – Umbilical veins


1. Affenito SG, Franko DL, Striegel-Moore RH, Thompson D. Behavioral determinants of obesity: Research findings and policy implications. J Obes. 2012;2012. doi: 10.1155/2012/150732 22957214

2. Poddar M, Chetty Y, Chetty VT. How does obesity affect the endocrine system? A narrative review. Clin Obes. 2017;7: 136–144. doi: 10.1111/cob.12184 28294570

3. Cani PD. Human gut microbiome: Hopes, threats and promises. Gut. 2018;67: 1716–1725. doi: 10.1136/gutjnl-2018-316723 29934437

4. Goodarzi MO. Genetics of obesity: what genetic association studies have taught us about the biology of obesity and its complications. Lancet Diabetes Endocrinol. 2018;6: 223–236. doi: 10.1016/S2213-8587(17)30200-0 28919064

5. Papathanasiou AE, Nolen-Doerr E, Farr OM, Mantzoros CS. Geoffrey Harris Prize lecture 2018: Novel pathways regulating neuroendocrine function, energy homeostasis and metabolism in humans. 2018; 1–29.

6. Vickers MH, Breier BH, Cutfield WS, Hofman PL, Gluckman PD. Fetal origins of hyperphagia, obesity, and hypertension and postnatal amplification by hypercaloric nutrition. Am J Physiol Metab. 2000;279: E83–E87. doi: 10.1152/ajpendo.2000.279.1.E83 10893326

7. Vickers MH, Krechowec SO, Breier BH. Is later obesity programmed in utero? Curr Drug Targets. 2007;8: 923–934. doi: 10.2174/138945007781386857 17691929

8. Abu-Saad K, Fraser D. Maternal nutrition and birth outcomes. Epidemiol Rev. 2010;32: 5–25. doi: 10.1093/epirev/mxq001 20237078

9. Scientific Advisory Committee on Nutrition. The influence of maternal, fetal and child nutrition on the development of chronic disease in later life. London, UK: London: TSO; 2011.

10. Golding J, Iles-Caven Y, Ellis G, Gregory S, Nowicki S. The relationship between parental locus of control and adolescent obesity: a longitudinal pre-birth cohort. Int J Obes. 2018; 1–11. doi: 10.1038/s41366-018-0141-y 29983415

11. Sarr O, Yang K, Regnault TRH. In utero programming of later adiposity: The role of fetal growth restriction. J Pregnancy. 2012;2012. doi: 10.1155/2012/134758 23251802

12. Brett KE, Ferraro ZM, Holcik M, Adamo KB. Placenta nutrient transport-related gene expression: The impact of maternal obesity and excessive gestational weight gain. J Matern Neonatal Med. 2016;29: 1399–1405. doi: 10.3109/14767058.2015.1049522 26067267

13. Cordero P, Li J, Nguyen V, Pombo J, Maicas N, Novelli M, et al. Developmental programming of obesity and liver metabolism by maternal perinatal nutrition involves the melanocortin system. Nutrients. 2017;9: 1–11. doi: 10.3390/nu9091041 28930194

14. Lomas-Soria C, Reyes-Castro LA, Rodríguez-González GL, Ibáñez CA, Bautista CJ, Cox LA, et al. Maternal obesity has sex-dependent effects on insulin, glucose and lipid metabolism and the liver transcriptome in young adult rat offspring. J Physiol. 2018;19: 4611–4628. doi: 10.1113/JP276372

15. Fox CS, Heard-Costa N, Cupples LA, Dupuis J, Vasan RS, Atwood LD. Genome-wide association to body mass index and waist circumference: The Framingham Heart Study 100K project. BMC Med Genet. 2007;8: 1–7. doi: 10.1186/1471-2350-8-1 17227582

16. Willer CJ, Willer CJ, Speliotes EK, Speliotes EK, Loos RJF, Loos RJF, et al. Six new loci associated with body mass index highlight a neuronal influence on body weight regulation. Nat Genet. 2009;41: 25. doi: 10.1038/ng.287 19079261

17. Turcotte M, Abadi A, Peralta-Romero J, Suarez F, Reddon H, Gomez-Zamudio J, et al. Genetic contribution to waist-to-hip ratio in Mexican children and adolescents based on 12 loci validated in European adults. Int J Obes. 2018; 1–10. doi: 10.1038/s41366-018-0055-8 29777226

18. Godfrey KM, Sheppard A, Gluckman PD, Lillycrop KA, Burdge GC, McLean C, et al. Epigenetic Gene Promoter Methylation at Birth Is Associated Width Child’s Later Adiposity. Diabetes. 2011;60: 1528–1534. doi: 10.2337/db10-0979 21471513

19. Saffery R, Novakovic B. Epigenetics as the mediator of fetal programming of adult onset disease: What is the evidence? Acta Obstet Gynecol Scand. 2014;93: 1090–1098. doi: 10.1111/aogs.12431 24835110

20. Sharp GC, Lawlor DA, Richmond RC, Fraser A, Simpkin A, Suderman M, et al. Maternal pre-pregnancy BMI and gestational weight gain, offspring DNA methylation and later offspring adiposity: Findings from the Avon Longitudinal Study of Parents and Children. Int J Epidemiol. 2015;44: 1288–1304. doi: 10.1093/ije/dyv042 25855720

21. Soubry A, Murphy SK, Wang F, Huang Z, Vidal AC, Fuemmeler BF, et al. Newborns of obese parents have altered DNA methylation patterns at imprinted genes. Int J Obes. 2015;39: 650–657. doi: 10.1038/ijo.2013.193 24158121

22. Sureshchandra S, Wilson RM, Rais M, Marshall NE, Purnell JQ, Thornburg KL, et al. Maternal Pregravid Obesity Remodels the DNA Methylation Landscape of Cord Blood Monocytes Disrupting Their Inflammatory Program. J Immunol. 2017; ji1700434. doi: 10.4049/jimmunol.1700434 28887432

23. Kühnen P, Handke D, Waterland RA, Hennig BJ, Silver M, Fulford AJ, et al. Interindividual Variation in DNA Methylation at a Putative POMC Metastable Epiallele Is Associated with Obesity. Cell Metab. 2016;24: 502–509. doi: 10.1016/j.cmet.2016.08.001 27568547

24. Gordon L, Joo JE, Powell JE, Ollikainen M, Novakovic B, Li X, et al. Neonatal DNA methylation profile in human twins is specified by a complex interplay between intrauterine environmental and genetic factors, subject to tissue-specific influence. Genome Res. 2012;22: 1395–1406. doi: 10.1101/gr.136598.111 22800725

25. Murphy SK, Huang Z, Hoyo C. Differentially methylated regions of imprinted genes in prenatal, perinatal and postnatal human tissues. PLoS One. 2012;7. doi: 10.1371/journal.pone.0040924 22808284

26. Cheung CYY, Tso AWK, Cheung BMY, Xu A, Ong KL, Fong CHY, et al. Obesity susceptibility genetic variants identified from recent genome-wide association studies: Implications in a Chinese population. J Clin Endocrinol Metab. 2010;95: 1395–1403. doi: 10.1210/jc.2009-1465 20061430

27. Takeuchi F, Yamamoto K, Katsuya T, Nabika T, Sugiyama T, Fujioka A, et al. Association of genetic variants for susceptibility to obesity with type 2 diabetes in Japanese individuals. Diabetologia. 2011;54: 1350–1359. doi: 10.1007/s00125-011-2086-8 21369819

28. León-Mimila P, Villamil-Ramírez H, Villalobos-Comparán M, Villarreal-Molina T, Romero-Hidalgo S, López-Contreras B, et al. Contribution of Common Genetic Variants to Obesity and Obesity-Related Traits in Mexican Children and Adults. PLoS One. 2013;8. doi: 10.1371/journal.pone.0070640 23950976

29. Tyrrell J, Wood AR, Ames RM, Yaghootkar H, Beaumont RN, Jones SE, et al. Gene-obesogenic environment interactions in the UK Biobank study. Int J Epidemiol. 2017;46: 559–575. doi: 10.1093/ije/dyw337 28073954

30. Mejía-Benítez A, Klünder-Klünder M, Yengo L, Meyre D, Aradillas C, Cruz E, et al. Analysis of the contribution of FTO, NPC1, ENPP1,. 2013; 1–6.

31. Vázquez-Del Mercado M, Guzmán-Ornelas MO, Meraz FIC, Ríos-Ibarra CP, Reyes-Serratos EA, Castro-Albarran J, et al. The 482Ser of PPARGC1A and 12Pro of PPARG2 Alleles Are Associated with Reduction of Metabolic Risk Factors even Obesity in a Mexican-Mestizo Population. Biomed Res Int. 2015;2015. doi: 10.1155/2015/285491 26185753

32. Rojano-Rodriguez ME, Beristain-Hernandez JL, Zavaleta-Villa B, Maravilla P, Romero-Valdovinos M, Olivo-Diaz A. Leptin receptor gene polymorphisms and morbid obesity in Mexican patients. Hereditas. 2016;153: 1–5. doi: 10.1186/s41065-015-0005-6

33. Hellmuth C, Lindsay KL, Uhl O, Buss C, Wadhwa PD, Koletzko B, et al. Association of maternal prepregnancy BMI with metabolomic profile across gestation. Int J Obes. 2017;41: 159–169. doi: 10.1038/ijo.2016.153 27569686

34. Gondwe A, Ashorn P, Ashorn U, Dewey KG, Maleta K, Nkhoma M, et al. Pre-pregnancy body mass index (BMI) and maternal gestational weight gain are positively associated with birth outcomes in rural Malawi. PLoS One. 2018;13: 1–15. doi: 10.1371/journal.pone.0206035 30352100

35. Chávez-Lizárraga D, Zárate-Segura P, Beltrán-Montoya J, Canchola-Sotelo C, Vadillo-Ortega F, Chavira-Suárez E. DNA Methylation Variability in a Single Locus of the RXRα Promoter from Umbilical Vein Blood at Term Pregnancy. Biochem Genet. 2018;56. doi: 10.1007/s10528-017-9838-1 29305749

36. Rahman M, Berenson AB. Accuracy of current body mass index obesity classification forwhite, black and Hispanic reproductive-age women. Obs Gynecol. 2010;115: 982–988. doi: 10.1097/AOG.0b013e3181da9423 20410772

37. Wojdacz TK, Dobrovic A. Methylation-sensitive high resolution melting (MS-HRM): a new approach for sensitive and high-throughput assessment of methylation. Nucleic Acids Res. 2007;35: e41. doi: 10.1093/nar/gkm013 17289753

38. Liu L, Ying C, Zhao Z, Sui L, Zhang X, Qian C, et al. Identification of reliable biomarkers of human papillomavirus 16 methylation in cervical lesions based on integration status using high-resolution melting analysis. Clin Epigenetics. 2018;10: 1–12. doi: 10.1186/s13148-017-0434-3

39. Institute of Medicine and National Research Council. Weight Gain During Pregnancy: Reexaming the Guidelines. Rasmussen KM, Yaktine AL, editors. IEEE International Symposium on Information Theory—Proceedings. Washington, D.C.: National Academies Press; 2009. doi: 10.1109/ISIT.2010.5513257

40. Li LC, Dahiya R. MethPrimer: Designing primers for methylation PCRs. Bioinformatics. 2002;18: 1427–1431. doi: 10.1093/bioinformatics/18.11.1427 12424112

41. Akika R, Awada Z, Mogharbil N, Zgheib NK. Region of interest methylation analysis: a comparison of MSP with MS-HRM and direct BSP. Mol Biol Rep. 2017;44: 295–305. doi: 10.1007/s11033-017-4110-7 28676996

42. Yajnik CS. Transmission of obesity-adiposity and related disorders from the mother to the Baby. Ann Nutr Metab. 2014;64: 8–17. doi: 10.1159/000362608 25059801

43. Voisin S, Almén MS, Zheleznyakova GY, Lundberg L, Zarei S, Castillo S, et al. Many obesity-associated SNPs strongly associate with DNA methylation changes at proximal promoters and enhancers. Genome Med. 2015;7: 1–16. doi: 10.1186/s13073-014-0122-2

44. Gemma C, Sookoian S, Alvarĩas J, García SI, Quintana L, Kanevsky D, et al. Maternal pregestational BMI is associated with methylation of the PPARGC1A promoter in newborns. Obesity. 2009;17: 1032–1039. doi: 10.1038/oby.2008.605 19148128

45. Xie X, Gao H, Zeng W, Chen S, Feng L, Deng D, et al. Placental DNA methylation of peroxisome-proliferator-activated receptor-γ co-activator-1α promoter is associated with maternal gestational glucose level. Clin Sci. 2015;129: 385–394. doi: 10.1042/CS20140688 25875376

46. Volkov P, Olsson AH, Gillberg L, Jørgensen SW, Brøns C, Eriksson KF, et al. A genome-wide mQTL analysis in human adipose tissue identifies genetic variants associated with DNA methylation, gene expression and metabolic traits. PLoS One. 2016;11: 1–31. doi: 10.1371/journal.pone.0157776 27322064

47. Gardner KR, Sapienza C, Fisher JO. Genetic and epigenetic associations to obesity-related appetite phenotypes among African-American children. Pediatr Obes. 2015;10: 476–482. doi: 10.1111/ijpo.12010 25779370

48. Marzi SJ, Meaburn EL, Dempster EL, Lunnon K, Paya-Cano JL, Smith RG, et al. Tissue-specific patterns of allelically-skewed DNA methylation. Epigenetics. 2016;11: 24–35. doi: 10.1080/15592294.2015.1127479 26786711

49. Chatterjee A, Stockwell PA, Rodger EJ, Duncan EJ, Parry MF, Weeks RJ, et al. Genome-wide DNA methylation map of human neutrophils reveals widespread inter-individual epigenetic variation. Sci Rep. 2015;5: 1–16. doi: 10.1038/srep17328 26612583

50. Casanello P, Schneider D, Herrera EA, Uauy R, Krause BJ. Endothelial heterogeneity in the umbilico-placental unit: DNA methylation as an innuendo of epigenetic diversity. Front Pharmacol. 2014;5 MAR: 1–9. doi: 10.3389/fphar.2014.00001

51. Heijmans BT, Tobi EW, Stein AD, Putter H, Blauw GJ, Susser ES, et al. Persistent epigenetic differences associated with prenatal exposure to famine in humans. Proc Natl Acad Sci. 2008;105: 17046–17049. doi: 10.1073/pnas.0806560105 18955703

52. Mosher, Melton, Stapleton, Schanfield, Crawford. Patterns of DNA Methylation across the Leptin Core Promoter in Four Diverse Asian and North American Populations. Hum Biol. 2016;88: 121. doi: 10.13110/humanbiology.88.2.0121 28161997

53. Kominiarek MA, Peaceman AM. Gestational Weight Gain. Am J Obstet Gynecol. 2017;217: 642–651. doi: 10.1016/j.ajog.2017.05.040 28549978

54. Ásbjörnsdóttir B, Rasmussen SS, Kelstrup L, Damm P, Mathiesen ER. Impact of restricted maternal weight gain on fetal growth and perinatal morbidity in obese women with type 2 diabetes. Diabetes Care. 2013;36: 1102–1106. doi: 10.2337/dc12-1232 23248191

55. Davis RR, Hofferth SL, Shenassa ED. Gestational weight gain and risk of infant death in the United States. Am J Public Health. 2014;104: 90–95. doi: 10.2105/AJPH.2013.301584

56. Hinkle SN, Sharma AJ, Swan DW, Schieve LA, Ramakrishnan U, Stein AD. Excess Gestational Weight Gain Is Associated with Child Adiposity among Mothers with Normal and Overweight Prepregnancy Weight Status. J Nutr. 2012;142: 1851–1858. doi: 10.3945/jn.112.161158 22955516

57. Bacart J, Leloire A, Levoye A, Froguel P, Jockers R, Couturier C. Evidence for leptin receptor isoforms heteromerization at the cell surface. FEBS Lett. 2010;584: 2213–2217. doi: 10.1016/j.febslet.2010.03.033 20347812

58. Popko K, Kucharska A, Wasik M. Leptin Receptors. 2010;15: 50–54. doi: 10.1186/2047-783X-15-S2-50

59. Akerman F, Lei ZM, Rao C V. Human umbilical cord and fetal membranes co-express leptin and its receptor genes. Gynecol Endocrinol. 2002;16: 299–306. doi: 10.1080/gye. 12396559

60. Kratzsch J, Schubring C, Stitzel B, Böttner A, Berthold A, Thiery J, et al. Inverse changes in the serum levels of the soluble leptin receptor and leptin in neonates: Relations to anthropometric data. J Clin Endocrinol Metab. 2005;90: 2212–2217. doi: 10.1210/jc.2004-1454 15671103

61. Lin R, Ju H, Yuan Z, Zeng L, Sun Y, Su Z, et al. Association of maternal and fetal LEPR common variants with maternal glycemic traits during pregnancy. Sci Rep. 2017;7: 1–9. doi: 10.1038/s41598-016-0028-x

62. Forhead AJ, Lamb CA, Franko KL, O’connor DM, Wooding FBP, Cripps RL, et al. Role of leptin in the regulation of growth and carbohydrate metabolism in the ovine fetus during late gestation. J Physiol. 2008;586: 2393–2403. doi: 10.1113/jphysiol.2007.149237 18325979

63. Matsuda J, Yokota I, Tsuruo Y, Murakami T, Ishimura K, Shima K, et al. Developmental changes in long-form leptin receptor expression and localization in rat brain. Endocrinology. 1999;140: 5233–5238. doi: 10.1210/endo.140.11.7152 10537153

64. Zhao J, Townsend KL, Schulz LC, Kunz TH, Li C, Widmaier EP. Leptin receptor expression increases in placenta, but not hypothalamus, during gestation in Mus musculus and Myotis lucifugus. Placenta. 2004;25: 712–722. doi: 10.1016/j.placenta.2004.01.017 15450389

65. Kahveci H, Laloglu F, Kilic O, Ciftel M, Kara M, Laloglu E, et al. Fasting and postprandial glucose, insulin, leptin, and ghrelin values in preterm babies and their mothers: Relationships among their levels, fetal growth, and neonatal anthropometry. J Matern Neonatal Med. 2015;28: 916–921. doi: 10.3109/14767058.2014.937693 25068948

66. Martínez-Cordero C, Amador-Licona N, Guízar-Mendoza JM, Hernández-Méndez J, Ruelas-Orozco G. Body Fat at Birth and Cord Blood Levels of Insulin, Adiponectin, Leptin, and Insulin-like Growth Factor-I in Small-for-Gestational-Age Infants. Arch Med Res. 2006;37: 490–494. doi: 10.1016/j.arcmed.2005.11.004 16624648

67. Volberg V, Heggeseth B, Harley K, Huen K, Yousefi P, Davé V, et al. Adiponectin and leptin trajectories in Mexican-American children from birth to 9 years of age. PLoS One. 2013;8. doi: 10.1371/journal.pone.0077964 24205046

68. Lazo-De-La-Vega-Monroy ML, González-Domínguez MI, Zaina S, Sabanero M, Daza-Benítez L, Malacara JM, et al. Leptin and its Receptors in Human Placenta of Small, Adequate, and Large for Gestational Age Newborns. Horm Metab Res. 2017;49: 350–358. doi: 10.1055/s-0043-103345 28351089

69. Nogues P, Dos Santos E, Jammes H, Berveiller P, Arnould L, Vialard F, et al. Maternal obesity influences expression and DNA methylation of the adiponectin and leptin systems in human third-trimester placenta. Clin Epigenetics. 2019;11: 1–18. doi: 10.1186/s13148-018-0606-9

70. Lv Y, Gao S, Zhang Y, Wang L, Chen X, Wang Y. MiRNA and target gene expression in menstrual endometria and early pregnancy decidua. Eur J Obstet Gynecol Reprod Biol. 2016;197: 27–30. doi: 10.1016/j.ejogrb.2015.11.003 26699100

71. Laganà AS, Vitale SG, Sapia F, Valenti G, Corrado F, Padula F, et al. miRNA expression for early diagnosis of preeclampsia onset: hope or hype? J Matern Neonatal Med. 2018;31: 817–821. doi: 10.1080/14767058.2017.1296426 28282763

72. Carreras-Badosa G, Bonmat A, Ortega FJ, Mercader JM, Guindo-Martnez M, Torrents D, et al. Dysregulation of placental miRNA in maternal obesity is associated with pre-and postnatal growth. J Clin Endocrinol Metab. 2017;102: 2584–2594. doi: 10.1210/jc.2017-00089 28368446

73. Chiofalo B, Laganà AS, Vaiarelli A, La Rosa VL, Rossetti D, Palmara V, et al. Do miRNAs play a role in fetal growth restriction? A fresh look to a busy corner. Biomed Res Int. 2017;2017. doi: 10.1155/2017/6073167 28466013

74. Block T, El-Osta A. Epigenetic programming, early life nutrition and the risk of metabolic disease. Atherosclerosis. 2017;266: 31–40. doi: 10.1016/j.atherosclerosis.2017.09.003 28950165

75. Münzberg H, Morrison CD. Structure, production and signaling of leptin. Metabolism. 2015;64: 13–23. doi: 10.1016/j.metabol.2014.09.010 25305050

76. McFarlane J, Parker B, Soeken K. Physical abuse, smoking, and substance use during pregnancy: prevalence, interrelationships, and effects on birth weight. J Obstet Gynecol Neonatal Nurs. 1996;25: 313–320. doi: 10.1111/j.1552-6909.1996.tb02577.x 8708832

77. Chełchowska M, Ambroszkiewicz J, Mazur J, Lewandowski L, Maciejewski TM, Ołtarzewski M, et al. Effect of tobacco smoking on the maternal and fetal adipokine axis in relation to newborn birth weight and length. Przegla̧d Lek. 2014;71: 567–571.

78. Chan JL, Blu S, Yiannakouris N, Suchard MA, Kratzsch J, Mantzoros CS. Regulation of Circulating Soluble Leptin Receptor Levels By Gender, Adiposity, Sex Steroids, and Leptin. Diabetes. 2002;51: 2105–2112. doi: 10.2337/diabetes.51.7.2105 12086939

79. Shen L, Waterland RA. Methods of DNA methylation analysis. Curr Opin Clin Nutr Metab Care. 2007;10: 576–581. doi: 10.1097/MCO.0b013e3282bf6f43 17693740

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