Placental imprinting variation associated with assisted reproductive technologies and subfertility

Abstract

Infertility affects one in 6 couples in developed nations, resulting in an increasing use of assisted reproductive technologies (ART). Both ART and subfertility appear to be linked to lower birth weight outcomes, setting infants up for poor long-term health. Prenatal growth is, in part, regulated via epigenetically-controlled imprinted genes in the placenta. Although differences in DNA methylation between ART and control infants have been found, it remains unclear whether these differences are due to the ART procedures or to the underlying parental subfertility and how these methylation differences affect imprinted gene expression. In this study, we examined the expression of 108 imprinted genes in placental tissues from infants born to subfertile parents (n = 79), matched naturally-conceived controls (n = 158), and infants conceived using in vitro fertilization (IVF, n = 18). Forty-five genes were identified as having significantly different expression between the subfertile infants and controls, whereas no significant differences were identified between the IVF and control groups. The expression of 4 genes-IGF2, NAPIL5, PAX8-AS1, and TUBGCP5-was significantly downregulated in the IVF compared with the subfertile group. Three of the 45 genes significantly dysregulated between subfertile and control placentae-GRB10, NDN, and CD44 -were found to have a significant positive correlation between expression and birth weight. Methylation levels for these 3 genes and 4 others-MKRN3, WRB, DHCR24, and CYR61-were significantly correlated with expression. Our findings indicate that epigenetic differences in placentas resulting from IVF pregnancies may be related to the underlying subfertility in parents using IVF rather than the IVF procedure itself.

Keywords: ART; IVF; birth weight; imprinted genes; placenta; subfertility.

Figure 1.

Volcano plots of the unadjusted P-value (y-axis) and fold change for gene expression (x-axes) of the 3 comparisons: subfertile vs. control placentae (A), IVF vs. subfertile placentae (B) and IVF vs. control placentae (C). Genes represented by purple dots were significant for both Bonferroni and FDR corrections, while genes represented by blue dots were significant at only the FDR threshold. Genes considered significantly differentially expressed are labeled with their gene name.

Figure 2.

Relationships between groups of genes significant in both comparisons. A) Venn diagram showing all genes that were significantly differentially expressed in any of the comparisons performed. The genes within the blue circle were found to have significantly different RNA expression between subfertile and control placentae, while the genes within the red circle were found to have significantly different expression between IVF and subfertile placentae. Those in both circles were significant in both comparisons. B) Differences in expression between the subfertile vs. control and subfertile vs. IVF comparisons for the 4 genes that were significant for both comparisons. For each gene, the log fold change for each comparison is plotted (dot) with the lower and upper confidence interval (horizontal line).

Figure 3.

Results of a linear model of the relationship between gene expression and birth weight in grams for the 3 genes identified as having a significant association between RNA expression and birth weight. The model included BMI, infant sex, and gestational age as potential confounders. A positive estimate indicates that an increase in expression is correlated with an increase in birth weight. Dots indicate the change in birth weight for 1-fold change of increased gene expression.