Associations Between Prenatal Vitamin D and Placental Gene Expression

Abstract

Background: Vitamin D is a hormone that regulates gene transcription. Prenatal vitamin D has been linked to immune and vascular function in the placenta, a key organ of pregnancy. Transcriptome-wide RNA sequencing can provide a more complete representation of the placental effects of vitamin D.

Objectives: We investigated the association between prenatal vitamin D concentrations and placental gene expression in a large, prospective pregnancy cohort.

Methods: Participants were recruited from Shelby County, TN, United States, in the Conditions Affecting Neurocognitive Development and Learning in Early childhood (CANDLE) study. Vitamin D (plasma total 25-hydroxyvitatmin D, [25(OH)D]) was measured at midpregnancy (16-28 wk) and delivery. RNA was sequenced from placental samples collected at birth. We identified differentially expressed genes (DEGs) using adjusted linear regression models. We also conducted weighted gene coexpression network analysis.

Results: The median 25(OH)D of participants was 21.8 ng/mL at midpregnancy (N = 774; IQR: 15.4-26.5 ng/mL) and 23.6 ng/mL at delivery (n = 753; IQR: 16.8-29.1 ng/mL). Placental expression of 17 DEGs was associated with 25(OH)D at midpregnancy, but only 1 DEG was associated with 25(OH)D at delivery. DEGs were related to energy metabolism, cytoskeletal function, and transcriptional regulation. We identified 2 weighted gene coexpression network analysis gene modules whose expression was associated with 25(OH)D at midpregnancy and 1 module associated with 25(OH)D at delivery. These modules were enriched for genes related to mitochondrial and cytoskeletal function and were regulated by transcription factors including ARNT2 and FOSL2. We also identified 12 modules associated with 25(OH)D in females and 1 module in males.

Conclusions: 25(OH)D during midpregnancy, but not at delivery, is associated with placental gene expression at birth. Future research is needed to investigate a potential role of vitamin D in modulating placental mitochondrial metabolism, intracellular transport, and transcriptional regulation during pregnancy.

Keywords: 25-hydroxyvitamin D; developmental origins of health and disease; placenta; transcriptomics; vitamin D.

FIGURE 1

Vitamin D concentrations at midpregnancy and delivery. Density plots of vitamin D concentrations measured as plasma 25(OH)D concentrations at (A) enrollment during midpregnancy (n = 774) and (B) at delivery (n = 753). Tertile cutoffs are indicated by dotted vertical lines. (C) A scatter plot of vitamin D concentrations at midpregnancy and delivery (n = 752) shows high correlation. The identity line (black) indicates equal 25(OH)D concentrations at both time points, although some participants’ 25(OH)D concentrations increased (red) or decreased (blue) from midpregnancy to delivery. 25(OH)D, 25-hydroxyvitamin D.

FIGURE 2

Weighted gene coexpression network analysis (WGCNA) modules associated with measures of 25(OH)D during pregnancy. The point size corresponds to −log(p) in each panel. (A) Associations between modules and maternal 25(OH)D concentrations at midpregnancy and delivery, as well as the change in 25(OH)D from midpregnancy to delivery were assessed with multiple linear regression adjusted for covariates (P < 0.05). Negative associations are indicated with downward-pointing triangles and positive associations are indicated with upward-pointing triangles. (B) Gene coexpression modules can be coregulated by transcription factors (TFs). TFs that were differentially expressed with 25(OH)D and whose gene targets were significantly enriched in the modules from panel A were identified using overrepresentation analysis (false discovery rate < 0.05). (C) KEGG pathways that were significantly enriched for module genes for modules in panel A were identified through an overrepresentation analysis (false discovery rate < 0.05). 25(OH)D, 25-hydroxyvitamin D.

FIGURE 3

Proposed mechanisms of differential gene expression associated with midpregnancy and delivery 25(OH)D. The placenta and conceptus develop and grow throughout gestation, as the placental builds increasingly complex vasculature [51,52]. Gene regulation and expression is dynamic to respond to changing conditions, including an increase in oxygen tension after the placenta invades maternal circulation and proinflammatory states both at implantation and in late gestation in preparation for labor [53,54]. Thus, midpregnancy exposures, like circulating 25(OH)D, could influence placental development and growth, leading to indirect impacts on placental gene expression at birth. Low vitamin D has been linked to reduced placental weight, impaired vascular development, and changes in the epigenetic landscape [14,17,[55], [56], [57]]. In turn, these changes influence placental functions like nutrient transport and alter gene accessibility, which can influence placental gene expression measured at delivery. At delivery, 25(OH)D concentrations are more relevant to concentrations of the active hormone, 1,25(OH)₂D, which directly affects gene expression by activating the VDR. 25(OH)D, 25-hydroxyvitamin D; VDR, vitamin D receptor.