Dose effect of gestational ethanol exposure on placentation and fetal growth

doi:10.1016/j.placenta.2015.02.010

Placenta

Volume 36, Issue 5, May 2015, Pages 523-530

https://doi.org/10.1016/j.placenta.2015.02.010 Get rights and content

Highlights

•
Prenatal ethanol exposure impairs placentation in a dose-responsive manner.
•
Severity of fetal growth impairment correlates with increasing doses of ethanol.
•
Ethanol alters branching morphogenesis in labyrinthine zone regardless of dose.
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Ethanol inhibits trophoblastic cell adhesion and motility.

Abstract

Introduction

Prenatal ethanol exposure compromises fetal growth by impairing placentation. Invasive trophoblastic cells, which mediate placentation, express the insulin-IGF regulated gene, aspartyl-asparaginyl β-hydroxylase (ASPH), which has a critical role in cell motility and invasion. The aims of this study were to characterize effects of ethanol on trophoblastic cell motility, and assess ethanol dose-dependent impairments in placentation and fetal development.

Methods

Pregnant Long Evans dams were fed with isocaloric liquid diets containing 0%, 8%, 18% or 37% ethanol (caloric content) from gestation day (GD) 6 to GD18. Fetal development, placental morphology, density of invasive trophoblasts at the mesometrial triangle, as well as placental and mesometrial ASPH and Notch-1 protein expression were evaluated. Directional motility of control and ethanol-exposed HTR-8/SVneo cells was assessed by ATP Luminescence-Based assay.

Results

Severity of fetal growth impairment correlated with increasing doses of ethanol. Ethanol exposure produced dose-dependent alterations in branching morphogenesis at the labyrinthine zone, and inhibited physiological transformation of maternal arteries. ASPH and Notch-1 protein expression levels were reduced, corresponding with impairments in placentation.

Discussion

Prenatal ethanol exposure compromises fetal growth and placentation in a dose-responsive manner. Ethanol's adverse effects on placental development are mediated by: 1) altered branching morphogenesis in labyrinthine zone; 2) suppression of invasive trophoblastic precursor cells; and 3) inhibition of trophoblastic cell adhesion and motility, corresponding with reduced ASPH and Notch-1 protein expression.

Introduction

Fetal alcohol syndrome (FAS) is the leading preventable cause of birth defects and developmental disorders in the United States [1]. The severity of alcohol-induced physical defects, and cognitive and behavioral disabilities varies by the timing, dose and duration of the alcohol exposure, nutritional status, genetic polymorphisms, maternal characteristics (gravity, parity, body mass index), and additional exposures (smoking/drugs); collectively, the phenotype is termed “Fetal Alcohol Spectrum Disorders” (FASD) [2], [3], [4], [5].

Intrauterine growth restriction (IUGR) is a key feature of FASD. Previously, we showed that prenatal ethanol exposure impairs fetal growth and placentation [6]; placentation is crucial for establishing the maternal–fetal interface needed for nutrient delivery and waste removal. A critical step in establishing the maternal–fetal interface is that invasive trophoblasts must invade maternal spiral arteries, and thereby disrupt the media and replace the endothelial cells [7], [8]. This process transforms the normally small muscular arteries into distended, flaccid, high-flow, low-resistance vessels, enabling continuous nutrient supply through placenta to the fetus. Prenatal ethanol exposure disrupts this process [6]. Mechanistically, ethanol inhibits insulin/insulin-like growth factor (IGF) signaling and expression of downstream target genes, including aspartyl-asparaginyl β-hydroxylase (ASPH) [6], [9].

ASPH is an insulin/IGF responsive gene whose hydroxylase activity regulates cell motility and invasiveness via activation and enhancement of Notch signaling [10], [11], [12]. ASPH is a positive regulator of trophoblastic cell motility. Previous in vitro and in vivo experiments demonstrated that siRNA inhibition of ASPH expression inhibits trophoblastic cell motility and alters signaling through Notch-1 leading to decreased expression levels of Notch's downstream target gene Hairy and Enhancer of Split 1 (Hes-1) [13]. The present study was designed to examine ethanol dose-effects on trophoblastic cell motility in relation to ASPH expression and fetal development.

Section snippets

In vivo model

Pregnant Long Evans rats were fed isocaloric liquid diets (BioServ, Frenchtown, NJ) containing 0%, 8%, 18%, or 37% ethanol by caloric content [14]. The liquid diets were initiated on gestation day (GD) 6 and ended on GD18. After mating, onset of pregnancy (gestation day 0; G0) was confirmed by both the presence of sperm and characteristic metestrous stage cellular changes in the vaginal smears [15]. The dams were weighed on GD 0 and weekly to monitor weight gain. Fetuses, placentas, and

Prenatal ethanol exposure alters fetal growth in a dose-responsive manner

On GD18, we detected pregnancy loss in 5 of 10 (50%) dams in 8%, 3 of 9 (33%) dams in 18% and 5 of 14 (36%) dams in 37% ethanol exposure groups. There was no pregnancy loss in the control group. We harvested the placentas and fetuses of the remainder dams: 0% (control, N = 7), 8% (N = 5), 18% (N = 6), or 37% (N = 9) ethanol by caloric content. The information regarding weight gain and blood alcohol levels of dams (mean ± S.E.M.) and litter size is summarized in Table 1. The blood alcohol levels

Discussion

This study demonstrates the inhibitory effects of ethanol on trophoblastic cell motility and adhesion, and dose-dependent effect on fetal growth and placentation. The severity of clinical symptoms of FASD has been attributed to variations in timing, duration and dose of maternal alcohol consumption, nutritional status, genetic polymorphisms, maternal characteristics (gravity, parity, body mass index), and additional exposures (smoking/drugs) [2], [3], [4], [5]. Herein, we demonstrate increased

Conflict of interest

The authors report no conflict of interest.

Acknowledgments

We are grateful to Terry Pasquariello for performing immunohistochemistry. This work was supported by National Institutes of Health grant K08AA-016783 to F.G. and K24-16126, AA-12908, AA-11347 to S.D.L.M.

References (39)

F. Gundogan et al.
Impaired placentation in fetal alcohol syndrome
Placenta
(2008)
C. Dunk et al.
A novel in vitro model of trophoblast-mediated decidual blood vessel remodeling
Lab Invest
(2003)
F. Gundogan et al.
siRNA inhibition of aspartyl-asparaginyl beta-hydroxylase expression impairs cell motility, Notch signaling, and fetal growth
Pathol Res Pract
(2011)
J. Xu et al.
Ethanol impairs insulin-stimulated neuronal survival in the developing brain: role of PTEN phosphatase
J Biol Chem
(2003)
L. Vercruysse et al.
Interstitial trophoblast invasion in the decidua and mesometrial triangle during the last third of pregnancy in the rat
Placenta
(2006)
C.H. Graham et al.
Establishment and characterization of first trimester human trophoblast cells with extended lifespan
Exp Cell Res
(1993)
S.M. de la Monte et al.
Ethanol inhibition of aspartyl-asparaginyl-beta-hydroxylase in fetal alcohol spectrum disorder: potential link to the impairments in central nervous system neuronal migration
Alcohol
(2009)
T. Urso et al.
Blood ethanol levels in sober alcohol users seen in an emergency room
Life Sci
(1981)
M. Gasperowicz et al.
The notch signalling pathway in the development of the mouse placenta
Placenta
(2008)
J.C. Cross et al.
Branching morphogenesis during development of placental villi
Differentiation
(2006)

D.G. Simmons et al.

Determinants of trophoblast lineage and cell subtype specification in the mouse placenta

Dev Biol

(2005)

P. Bischof et al.

The human cytotrophoblastic cell, a mononuclear chameleon

Int J Biochem Cell Biol

(2005)

F. Gundogan et al.

Role of aspartyl-(asparaginyl) beta-hydroxylase in placental implantation: relevance to early pregnancy loss

Hum Pathol

(2007)

B.A. Bailey et al.

Pregnancy and alcohol use: evidence and recommendations for prenatal care

Clin Obstet Gynecol

(2008)

H.M. Barr et al.

Identifying maternal self-reported alcohol use associated with fetal alcohol spectrum disorders

Alcohol Clin Exp Res

(2001)

C. Guerri et al.

Foetal alcohol spectrum disorders and alterations in brain and behaviour

Alcohol Alcohol

(2009)

K.R. Warren et al.

Genetic polymorphisms: impact on the risk of fetal alcohol spectrum disorders

Birth Defects Res A Clin Mol Teratol

(2005)

P.A. May et al.

Maternal risk factors for fetal alcohol spectrum disorders: not as simple as it might seem

Alcohol Res Health

(2011)

R.B.J. Pijnenborg et al.

The pattern of interstitial trophoblastic invasion of the myometrium in early human pregnancy

Placenta

(1981)

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Dose effect of gestational ethanol exposure on placentation and fetal growth

Highlights

Abstract

Introduction

Methods

Results

Discussion

Introduction

Section snippets

In vivo model

Prenatal ethanol exposure alters fetal growth in a dose-responsive manner

Discussion

Conflict of interest

Acknowledgments

Placenta

Lab Invest

Pathol Res Pract

J Biol Chem

Placenta

Exp Cell Res

Alcohol

Life Sci

Placenta

Differentiation

Dev Biol

Int J Biochem Cell Biol

Hum Pathol

Pregnancy and alcohol use: evidence and recommendations for prenatal care

Clin Obstet Gynecol

Identifying maternal self-reported alcohol use associated with fetal alcohol spectrum disorders

Alcohol Clin Exp Res

Foetal alcohol spectrum disorders and alterations in brain and behaviour

Alcohol Alcohol

Genetic polymorphisms: impact on the risk of fetal alcohol spectrum disorders

Birth Defects Res A Clin Mol Teratol

Maternal risk factors for fetal alcohol spectrum disorders: not as simple as it might seem

Alcohol Res Health

The pattern of interstitial trophoblastic invasion of the myometrium in early human pregnancy

Placenta