Oxytocin receptor DNA methylation is associated with exogenous oxytocin needs during parturition and postpartum hemorrhage

Abstract

Background: The oxytocin receptor gene (OXTR) is regulated, in part, by DNA methylation. This mechanism has implications for uterine contractility during labor and for prevention or treatment of postpartum hemorrhage, an important contributor to global maternal morbidity and mortality.

Methods: We measured and compared the level of OXTR DNA methylation between matched blood and uterine myometrium to evaluate blood as an indicator of uterine methylation status using targeted pyrosequencing and sites from the Illumina EPIC Array. Next, we tested for OXTR DNA methylation differences in blood between individuals who experienced a postpartum hemorrhage arising from uterine atony and matched controls following vaginal birth. Bivariate statistical tests, generalized linear modeling and Poisson regression were used in the analyses.

Results: Here we show a significant positive correlation between blood and uterine DNA methylation levels at several OXTR loci. Females with higher OXTR DNA methylation in blood had required significantly more exogenous oxytocin during parturition. With higher DNA methylation, those who had oxytocin administered during labor had significantly greater relative risk for postpartum hemorrhage (IRR 2.95, 95% CI 1.53-5.71).

Conclusions: We provide evidence that epigenetic variability in OXTR is associated with the amount of oxytocin administered during parturition and moderates subsequent postpartum hemorrhage. Methylation can be measured using a peripheral tissue, suggesting potential use in identifying individuals susceptible to postpartum hemorrhage. Future studies are needed to quantify myometrial gene expression in connection with OXTR methylation.

Fig. 1. OXTR gene schematic and location of CpG sites with significantly correlated methylation.

Gene schematic of OXTR. Boxes represent exons and lines represent introns. Coding regions are in white and untranslated regions are in gray. The black arrow indicates the transcription start site. The black bar below the gene denotes the CpG island. The 5’ region of the gene is enlarged to show detail. Blue lines indicate CpG sites assayed by pyrosequencing and red lines indicate CpG sites assayed by Illumina Methylation EPIC 850 K array. CpG sites with significantly correlated DNA methylation in blood and myometrium are indicated by letters and Spearman’s rho is reported below the gene schematic.

Fig. 2. DNA methylation of CpG sites in OXTR are correlated in blood and myometrium.

A Significantly correlated CpGs between tissues (n = 26 participants with matched samples) p < 0.05 after correction for multiple comparisons using the Benjamini-Hochberg method: −924 (red): ρ = 0.52, p = 0.03, −934 (light blue): ρ = 0.65, p = 0.005, cg02192228 (green): ρ = 0.62, p = 0.001, cg03257388 (dark blue): ρ = 0.66, p = 0.005, cg11171527 (apricot): ρ = 0.55, p = 0.02, cg15317815 (lavender): ρ = 0.59, p = 0.01. B Differences in methylation levels at each CpG site between myometrium and blood, stars denote Bonferroni corrected significant differences: −934 (t(25) = 10.1, p = <0.001), −924 (t(25) = 9.6, p < 0.001), cg15317815 (t(24) = 3.2, p = 0.004), cg02192228 (t(24) = −4.3, p = 0.003), cg03257388 (t(24) = 1.12, p = 0.27), cg11171527 (t(24) = 2.2, p = 0.04). Whiskers denote +/−1.5 x interquartile range.

Fig. 3. Cases of postpartum hemorrhage had lower DNA methylation when labor was not stimulated by oxytocin.

A Star denotes significant difference in total DNA methylation between cases of postpartum hemorrhage cases and controls among participants whose labor was not stimulated with oxytocin (red box plots, n = 38), z = 2.84, p = 0.003. B Difference in DNA methylation between postpartum hemorrhage cases and controls when oxytocin was used in labor (blue box plots, n = 53) z = −1.86, p = 0.06. Whiskers denote +/−1.5 x interquartile range.

Fig. 4. Increased OXTR DNA methylation is associated with increased oxytocin use and total postpartum blood loss only when oxytocin is administered during labor.

Red dots show participants without oxytocin use in labor (n = 38); blue dots denote oxytocin was used in labor (n = 53). Shaded area denotes 95% CI of regression line. A Higher −934 OXTRm is associated with higher intra/postpartum administration of oxytocin among participants receiving oxytocin (blue dots). B Postpartum bleeding volume associated with DNA methylation however, severity increased when oxytocin was used in labor.