Role of the 820 A/G variant in the IGF-2 gene and recurrent spontaneous abortion in southern Iran: A cross-sectional study
Background: Insulin-like growth factor-2 (IGF-2) is a polypeptide growth factor and one of the first genes expressed prior to the implantation of the embryo, with its highest expression in the placental cells. Its activity strongly depends on the genomic imprinting, and the result of the loss of genetic imprinting is the termination of the early stages of embryonic development, which can lead to recurrent spontaneous abortion.
Objective: This cross-sectional study aimed to investigate the role of 820A/G variant of the IGF-2 gene and the probability to recurrent spontaneous abortion (RSA) in southern Iran.
Materials and Methods: In this study, 50 aborted fetuses tissue for the study group and blood samples umbilical-cord from newborns as control group (n = 50) were collected from Shiraz-Iran (2017). The genotyping of the target point in the IGF-2 gene was performed by Real-time Polymerase Chain Reaction and analyzed through highresolution melting (HRM) curve.
Results: Based on the collected data (AA genotype = reference), allele “A” frequency in aborted fetus was 51% and control 68% as well as allele G 49% and 32%, respectively. Moreover, 27 aborted embryos (54%) were heterozygous (A/G) (OR = 3.274, 95% CI = 1.015-10.561, p = 0.04), while 18 cases (36%) in control sample showed heterozygosity. Considering the phenotypic status, the G allele had a dominant effect on the incidence of RSA (p = 0.008, OR = 3.167).
Conclusion: Based on the present study, the risk of abortion due to loss of heterozygosity or quantitative decline of the IGF-2 is about three-fold in the southern Iran.
Key words: Variant, IGF-2, Spontaneous abortion, Genomic imprinting, Gene expression.
 Matthiesen L, Kalkunte S, Sharma S. Multiple pregnancy failures: an immunological paradigm. Am J Reprod Immunol 2012; 67: 334–340.
 McNamee K, Dawood F, Farquharson R. Recurrent miscarriage and thrombophilia: An update. Curr Opin Obstet Gynecol 2012; 24: 229–234.
 Roland L, Gagne A, Belanger MC, Boutet M, Julien P, Bilodeau JF. Plasma interleukin-18 (IL- 18) levels are correlated with antioxidant vitamin coenzyme Q (10) in preeclampsia. Acta Obstet Gynecol Scand 2010; 89: 360–366.
 Huang HY. The cytokine network during embryo implantation. Chang Gung Med J 2006; 29: 25–36.
 Frequently asked questions: pregnancy. repeated miscarriages. The American College of Obstetricians and Gynecologists. Available at: URL: https://m.acog. org/Patients/FAQs/Repeated-Miscarriages.
 Cui H, Onyango P, Brandenburg S, Wu Y, Hsieh CL, Feinberg AP. Loss of imprinting in colorectal cancer linked to hypomethylation of H19 and IGF2. Cancer Res 2002; 62: 6442–6446.
 Gurrieri F, Accadia M. Genetic imprinting: the paradigm of prader-willi and angelman syndromes. Endocr Dev 2009; 14: 20–28.
 Moore GE, Oakey R. The role of imprinted genes in humans. Genome Biol 2011; 12: 106.
 Ulaner GA, Vu TH, Li T, Hu JF, Yao XM, Yang Y, et al. Loss of imprinting of IGF2 and H19 in osteosarcoma is accompanied by reciprocal methylation changes of a CTCF-binding site. Hum Mol Genet 2003; 12: 535–549.
 Tycko B, Morison IM. Physiological functions of imprinted genes. J Cell Physiol 2002; 192: 245–258.
 Horike SI, Ferreira JCP, Meguro-Horike M, Choufani S, Smith AC, Shuman C, et al. Screening of DNA methylation at the H19 promoter or the distal region of its ICR1 ensures efficient detection of chromosome 11p15 epimutations in Russell-Silver syndrome. Am J Med Genet A 2009; 149: 2415– 2423.
 Netchine I, Rossignol S, Dufourg MN, Azzi S, Rousseau A, Perin L, et al. 11p15 imprinting center region 1 loss of methylation is a common and specific cause of typical Russell-Silver syndrome: clinical scoring system and epigenetic-phenotypic correlations. J Clin Endocrinol Metab 2007; 92: 3148–3154.
 Filippova GN, Fagerlie S, Klenova EM, Myers C, Dehner Y, Goodwin G, et al. An exceptionally conserved transcriptional repressor, CTCF, employs different combinations of zinc fingers to bind diverged promoter sequences of avian and mammalian c-myc oncogenes. Mol Cell Biol 1996; 16: 2802–2813.
 Rubio ED, Reiss DJ, Welcsh PL, Disteche CM, Filippova GN, Baliga NS, et al. CTCF physically links cohes in to chromatin. Proc Natl Acad Sci USA 2008; 105: 8309–8314.
 Koukoura O, Sifakis S, Soufla G, Zaravinos A, Apostolidou S, Jones A, et al. Loss of imprinting and aberrant methylation of IGF2 in placentas from pregnancies complicated with fetal growth restriction. Int J Mol Med 2011; 28: 481–487.
 IGF2 gene. Genetics Home Reference. Available at: URL: https://ghr.nlm.nih.gov/gene/IGF2.
 McKinnon T, Chakraborty C, Gleeson LM, Chidiac P, Lala PK. Stimulation of human extra villous trophoblast migration by IGF-II is mediated by IGF type 2 receptor involving inhibitory G protein (s) and phosphorylation of MAPK. J Clin Endocrinol Metab 2001; 86: 3665–3674.
 Lee OH, Bae SK, Bae MH, Lee YM, Moon EJ, Cha HJ, et al. Identification of angiogenic properties of insulin-like growth factor II in in vitro angiogenesis models. Br J Cancer 2000; 82: 385–391.
 Hills FA, Elder MG, Chard T, Sullivan MHF. Regulation of human villous trophoblast by insulin-like growth factors and insulin-like growth factor-binding protein- 1. J Endocrinol 2004; 183: 487–496.
 Herr F, Liang OD, Herrero J, Lang U, Preissner KT, Han VK, et al. Possible angiogenic roles of insulinlike growth factor II and its receptors in uterine vascular adaptation to pregnancy. J Clin Endocrinol Metab 2003; 88: 4811–4817.
 Reik W, Constancia M, Fowden A, Anderson N, Dean W, Ferguson-Smith A, et al. Regulation of supply and demand for maternal nutrients in mammals by imprinted genes. J Physiol 2003; 547: 35–44.
 Stewart CE, Rotwein P. Insulin-like growth factor- II is an autocrine survival factor for differentiating myoblasts. J Biol Chem 1996; 271: 11330–11338.
 Soejima H, Yun K. Allele-specific-polymerase chain reaction: a novel method for investigation of the imprinted insulin-like growth factor II gene. Lab Invest 1998; 78: 641–642.
 Stray-Pedersen B, Stray-Pedersen S. Etiologic factors and subsequent reproductive performance in 195 couples with a prior history of habitual abortion. Am J Obstet Gynecol 1984; 148: 140–146.
 Zastavna D, Makukh H, Tretjak B, Bilevych O, Tyrka M. Loss of Imprinting of IGF2 Gene in the Chorionic Tissues of Spontaneously 21-Eliminated Human Embryos. Genet Epigenet 2013; 5: 17–22.
 Ostojic S, Pereza N, Volk M, Kapovic M, Peterlin B. Genetic predisposition to idiopathic recurrent spontaneous abortion: contribution of genetic variations in IGF-2 and H19 imprinted genes. Am J Reprod Immunol 2008; 60: 111–117.
 Solter D, Aronson J, Gilbert SF, McGrath J. Nuclear transfer in mouse embryos: activation of the embryonic genome. Cold Spring Harb Symp Quant Biol 1985; 50: 45–50.
 Rodriguez S, Gaunt TR, O Dell SD, Chen XH, Gu D, Hawe E, et al. Haplotypic analyses of the IGF2-INSTH gene cluster in relation to cardio vascular risk traits. Hum Mol Genet 2004; 13: 715–725.
 Sandhu MS, Gibson JM, Heald AH, Dunger DB, Wareham NJ. Low circulating IGF-II concentrations predict weight gain and obesity in humans. Diabetes 2003; 52: 1403–1408.
 Chen J, Fang Q, Chen B, Zhou Y, Luo Y. Study on the imprinting status of insulin-like growth factor II (IGF-II) gene in villus during 6-10 gestational weeks. ObstetGynecolInt 2010; 2010: 965905–965908.
 Lighten AD, Hardy K, Winston RM, Moore GE. IGF2 is parentally imprinted in human preimplantation embryos. Nat Genet 1997; 15: 122–123.
 Lighten AD, Hardy K, Winston RM, Moore GE. Expression of mRNA for the insulin-like growth factors and their receptors in human preimplantation embryos. MolReprodDev 1997; 47: 134–139.