Effects of Maternal Hypertension on Cord Blood Arginine Vasopressin Receptor Expression

Abstract

Arginine vasopressin (AVP) signaling is altered in preeclampsia and physiologic stress. AVP is implicated in fluid homeostasis and cardiovascular (CV) function, which is disrupted in some progeny from preeclamptic pregnancies. However, whether altered fetal AVP signaling occurs in preeclampsia is unknown. Here, we measured CV-related transcripts (e.g., AVP receptors) in cord blood via quantitative PCR. Chronic hypertension decreased AVPR1b, AVPR2, OXTR, LNPEP, and CUL5. AVPR1a, AVPR1b, and AVPR2 were decreased while OXTR was increased in preeclamptic cord blood. In sum, we found prenatal exposure to hypertension in pregnancy alters fetal AVP signaling and may thereby prime offspring CV disease risk.

Introduction

Preeclampsia is a common and deadly hypertensive disease of pregnancy which primes lifelong maternal and fetal risk for cardiovascular disease and other pathologies. For example, women with a history of preeclampsia have double or more increased risk for future cardiovascular disease including ischemic heart disease, while their children have increased lifetime stroke risk (RR: 1.9)[1–3]. The mechanisms mediating this increased risk are, however, mostly unknown.

Arginine vasopressin (AVP) is a key player in preeclampsia pathogenesis and is sufficient to cause endophenotypes of preeclampsia in a mouse model, as previously described by our lab [4–6]. AVP is a nonapeptide that regulates blood pressure and body fluid homeostasis and hemodynamics via binding receptors including AVPR1a, AVPR1b, and AVPR2, as well as the oxytocin receptor (OXTR) and CUL5 (aka VACM-1), a calcium-mobilizing protein activated by AVP. These targets have varying expression across tissues and functions; for instance, AVPR1a is highly expressed in smooth muscle and enables vasoconstriction, while AVPR1b is highly expressed in the pituitary and regulates adrenocorticotropic hormone release. Furthermore, maternal AVP and AVPR expression is disrupted by maternal hypertension in pregnancy, suggesting a mechanism by which gestational hypertension may influence life-long increased risk for cardiovascular disease [4, 5]. However, whether these same mechanisms are implicated in fetal cardiovascular programming is unclear. To address this gap, we assessed how cord blood expression of AVP receptors and LNPEP, which codes for a zinc-dependent aminopeptidase that degrades vasopressin, are impacted by gestational and fetal factors including chronic hypertension in pregnancy, infant sex, and gestational age at delivery.

Methods

Umbilical cord blood cells were obtained from the Iowa Maternal Fetal Tissue Bank (IRB# 200910784), which is maintained by the Department of Obstetrics and Gynecology at the University of Iowa. Biosamples were uniformly processed and frozen, as described previously[7]. All participants in the Iowa Maternal Fetal Tissue Bank (IRB# 200910784) provide informed consent prior to their participation in the biobank. Coded clinical information was provided with coded biological samples. The protocol used existing coded samples and coded clinical data and is therefore not human subjects research (exempt category 4). Samples were coded for fetal sex (n=10 male, 10 female), maternal chronic hypertension diagnosis (n=20 hypertensives, 20 controls), gestational age at delivery (n=10 preterm delivery <37 weeks, 10 term), and preeclampsia (n=10 preeclamptics, 10 controls). Maternal and fetal characteristics/diagnoses are described in supplementary table 1 and were confirmed by electronic medical record review by the Maternal Fetal Tissue Bank.

Expression of AVPR1a, AVPR1b, and AVPR2, OXTR, CUL5, and LNPEP were assessed in umbilical cord blood samples. A qPCR approach (mirVANA miRNA isolations, Superscript III reverse transcription, and PowerSYBR Green assay) described previously [5] was employed and assays were performed with the Iowa Institute of Human Genetics. qPCR results were normalized to 18S rRNA to calculate ΔCts. Change in average expression (ΔΔCt) for each receptor and fold change were then calculated. Comparisons were made by two-sided t-tests with unequal variance. P<0.05 was significant.

Results and Discussion

Fetal expression of any receptor did not differ significantly on the basis of infant sex (Figure 1a). Expression of AVPR1b (3.73-fold, term vs preterm, p=0.001), AVPR2 (2.83-fold, p=0.001), and OXTR (1.97-fold, p=0.001) were significantly decreased with preterm birth. Fetal AVPR1a, LNPEP, and CUL5 were unchanged (Figure 1b). Furthermore, AVPR1b (3.1-fold, hypertensive vs control, p=0.001), AVPR2 (8.2-fold, p<0.001), OXTR (4.3-fold, p<0.001), LNPEP (12.9-fold, p<0.001), and CUL5 (13.5-fold, p<0.001) were significantly decreased in cord blood cells from chronically hypertensive mothers while AVP1a expression was unchanged. Preeclampsia impacts on cord blood cell expression were also investigated. There were significant decreases in AVPR1a (4.7-fold, preeclampsia vs control, p<0.001), AVPR1b (2.5-fold, p=0.045), and AVPR2 (2.5-fold, 0.001), while OXTR (2.8-fold, p<0.001) was significantly increased in preeclamptics. LNPEP and CUL5 were not significantly changed.

Figure 1:

Comparisons of mRNA expression in cord blood cells between a) males and female infant, b) preterm and term pregnancies, c) chronically hypertensive and normotensive pregnancies, and d) preeclamptic and non-preeclamptic pregnancies.

*P<0.05 by two-sided t-test.

These results demonstrate that in utero exposures (e.g. to maternal hypertension) and fetal factors (e.g., prematurity) cause significant changes in transcripts involved in vasopressin signaling. Our results are not without precedent—other fetal insults, for example hypoxia or teratogens, similarly disrupt fetal vasopressin signaling and receptor expression [8, 9]. Some of the specific expression changes reported here may also help to explain the known developmental programming outcomes associated with preeclampsia. For example, the most changed transcript in preeclamptic pregnancies, AVPR1a (4.7-fold decrease), and OXTR are also implicated in stress-responsiveness, social behavior, and autism spectrum disorder [10, 11]. These are psychiatric disruptions which have previously been linked to prenatal preeclampsia exposure [12]. AVPR1a also regulates congenital cardiac function, which is known to be disrupted in offspring from preeclamptic pregnancies [13, 14].

The changes documented here demonstrate the prenatal programming of AVP signaling dysfunction by preeclamptic pregnancies. These disruptions may persist to cause life-long risk for cardiovascular and other hemoregulatory disease states. An underlying AVP signaling dysregulation mechanism may drive vulnerability for adult-onset cardiovascular conditions, which are increased among the children of preeclamptic women [15]. Furthermore, given that these changes are evident even at birth, they may be prime candidates for biomarker discovery. Future work will need to determine tractability of these changes across the lifespan and their interaction with real-world disease risk.

Highlights.

• Arginine vasopressin (AVP) signaling changes in preeclampsia and regulates fluid

• AVP receptor expression in cord blood may reflect fetal signaling and physiology

• In preeclamptic cord blood AVPR1a, AVPR1b, and AVPR2 decreased and OXTR increased

• Changes also occurred on the basis of prematurity and hypertension but not fetal sex

• Complex pregnancy exposure alters AVP signaling and may prime offspring risk