RESUMO
INTRODUCTION: The aim of this study is to determine whether the gene expression and associated DNA methylation regulation of H19 and IGF2 are altered in placentas conceived by assisted reproductive technologies (ART) compared to natural conceptions. METHODS: 113 pregnancies were recruited resulting in 119 placentas (83 singletons and 36 twins), where 56 were conceived via in vitro fertilization (IVF), 41 via intracytoplasmic sperm injection (ICSI), and 22 naturally. Regulation of imprinting of H19 and IGF2 was determined by the DNA methylation status at three CpG sites within the H19 imprinting control region 1 (ICR1) using bisulphite pyrosequencing. Expression of H19 and IGF2 in 45 of these placentas (17 IVF, 14 ICSI, and 14 NC) was measured by determining the relative mRNA transcript levels using RT-qPCR in placental villi. RESULTS: Placental weight and birth weight were not significantly different between groups. H19 expression was significantly increased in both IVF and ICSI placentas when compared to controls (1.8 and 1.9 fold higher, respectively). Conversely, IGF2 was significantly decreased in both ART groups (0.8 and 0.7 fold lower, respectively). Mean DNA methylation at ICR1 was found to be similar between all groups. No correlation was found between DNA methylation at ICR1 and expression of either gene. However, a significant inverse relationship was found between H19 and IGF2 expression. CONCLUSION: We provide evidence of altered H19 and IGF2 expression in ART placentas. The altered expression pattern may suggest a loss of imprinting on the paternal allele. Furthermore, these alterations may not be entirely associated with DNA methylation at ICR1. We show further indirect evidence of the H19-IGF2 inverse expression pattern.