SRI Supplement to Reproductive Sciences - Volume 25 Number 1 - March 2018 - 99A

Scientific Abstracts

prefrontal cortex, and dorsal striatum of juvenile and adult offspring.
Intracellular DA and norepinephrine (NE) were quantified in adult whole
brain via HPLC-EC. Offspring were weighed in juvenile and adult life.
RESULTS: Electrically-evoked DA release was quantified in 33 juvenile
and 32 adult offspring. HPLC for catecholamines was performed in 36
adult offspring. MATOB-exposed juveniles had significantly decreased
DA release (Fig 1A). 2-way ANOVA demonstrated a significant effect
of MATOB, offspring sex, and MATOB x sex interaction on juvenile
DA release. Significantly decreased DA release persisted into adult life
in Fs only (Fig 1B). MATOB-exposed adults had significantly lower
intracellular DA and NE levels than controls, suggesting deficient
synthesis (Fig 2). Only F offspring had significant weight differences,
with MATOB-exposed Fs significantly lighter as juveniles and heavier
as adults compared to controls.
*Figure(s) will be available online.
CONCLUSION: MATOB is associated with a DA and NE synthesis
deficit in exposed M and F adult offspring. DA release studies
demonstrated that only Fs had significantly decreased DA release
persisting from juvenile to adult life. This lasting decrease in DA reward
signaling may have implications for the increased vulnerability of F
offspring to overeating and obesity in the setting of MATOB exposure.

Maternal Diet Modification Restores Placental Blood Flow in a NonHuman Primate Model of Diet-Induced Obesity. Jamie Lo,1 Matthias
Schabel,1 Victoria Roberts,2 Antonio Frias*.1 1Oregon Health & Science
University, Portland, OR, United States; 2Oregon National Primate
Research Center, Beaverton, OR, United States.
INTRODUCTION: With the obesity epidemic, there is concern
of maternal nutrition influencing placental dysfunction and adverse
pregnancy outcomes. We previously demonstrated that maternal westernstyle diet (WSD) in non-human primates (NHP) disturbs utero-placental
hemodynamics using Doppler ultrasound in the early third-trimester. It
is unknown whether switching NHPs from a chronic WSD to a standard
chow diet pre-pregnancy (REV) would improve placental function. We
have developed a quantitative MRI technique, dynamic contrast enhanced
(DCE)-MRI, in the NHP that can image blood flow in the entire placental
unit simultaneously, allowing the summation of total spiral artery blood
flow across the placenta. Our study objective was to utilize DCE-MRI to
facilitate quantification of placental perfusion in response to a maternal
WSD and REV during pregnancy. We hypothesized that WSD would
reduce placental blood flow at mid to late gestation, but be restored with
diet-reversal pre-pregnancy.
METHODS: Japanese macaques were divided into 3 treatment groups:
control (CON, n=6), REV (n=5) and WSD (n=5). Animals were
maintained on standard chow (14% calories from fat), a WSD (36%
calories from fat), or REV (switched from a chronic WSD to a CON diet
3 months prior to conception). All underwent DCE-MRI at gestational
day 90 (G90) and G129 (term is G175). A novel DCE-MRI protocol was
utilized to calculate placental perfusion from maternal spiral arteries.
RESULTS: By DCE-MRI, maternal-side placental blood flow increased
from G90 to G129 in all three cohorts. Consumption of a WSD throughout
pregnancy significantly diminished blood flow compared to controls and
this was apparent at mid-gestation and persisted in the early third trimester
(*p<0.05) [Figure1]. Placental blood flow was restored to control levels
by dietary reversal.
CONCLUSION: Maternal utero-placental blood flow increased between
G90 and G129 in all groups as expected. However, overall placental
perfusion was significantly reduced in WSD compared to CON and
REV, which is apparent earlier at mid-gestation and persistent in the
third trimester on DCE-MRI. This suggests that a WSD adversely effects
placental blood flow and vascular development, but dietary reversal may
ameliorate these concerning findings.
*Figure(s) will be available online.

99A

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Maternal Diabetes Suppresses Sox2ot Expression and CRISPR-Cas9
Editing on Sox2ot Gene Promoter Disrupts Its Expression Leading
to Neural Tube Defects. Daoyin Dong†, E Albert Reece, Wei-Bin Shen,
Peixin Yang. University of Maryland School of Medicine, Baltimore,
MD, United States.
INTRODUCTION: Pregestational maternal diabetes disrupts neurulation
leading to neural tube defects (NTDs) in the offspring. Sox2 overlapping
transcript (Sox2ot) is a long non-coding RNA (lncRNA). The Sox2ot gene
harbors one of the major neural development regulators, the Sox2 gene,
in its intronic region and they co-transcribe in the same direction. Sox2ot
is primarily expressed in the developing neuroepithelium. However, how
maternal diabetes regulates this lncRNA during neurulation and its role
in neural tube closure remain to be characterized.
METHODS: Diabetes in female mice was induced by intravenous
injection of 65 mg/kg streptozotocin over 2 days. Diabetes was defined
as a 12-hour fasting blood glucose level ≥ 14 mM. Embryos at embryonic
day 8.5 (E8.5) from both nondiabetic and diabetic dams were collected
for an lncRNA profiling assay. The expression of sox2ot in E8.5 embryos
was confirmed by quantitative real-rime PCR (RT-qPCR). To investigate
whether Sox2ot is required for normal embryonic development and its
deletion will lead to NTDs, the promoter region of Sox2ot was disrupted
by CRISPR-Cas9 genome editing technology to deplete Sox2ot gene
expression in the mouse. NTDs were examined in E9.5 embryos. E8.5
neurulation-stage embryos were used for molecular and biochemical
analyses.
RESULTS: The lncRNA profiling showed that the expression of Sox2ot
was down-regulated approximately 2.5-fold in E8.5 embryos of diabetic
dams compared to that of embryos in the nondiabetic group. RT-qPCR
confirmed the suppression of Sox2ot expression by maternal diabetes in
E8.5 embryos. In the Sox2ot gene editing experiment, we designed several
single-stranded guide RNA (sgRNA) to specifically target the Sox2ot
promoter. We detected small insertions and deletions (INDELs) in the
promoter region of the Sox2ot gene in cells transfected with Cas9 mRNA
and the three sgRNAs. Then, these three sgRNAs and Cas9 mRNA were
injected into mouse zygotes and implanted into pesudopregnant mice. We
obtained 9 founders with the Sox2ot gene promoter gene edited and there
were three of them with significant suppression of Sox2ot expression.
Furthermore, NTDs were observed in homozygous embryos of these
three mouse lines.
CONCLUSION: Maternal diabetes suppresses the lncRNA Sox2ot gene
expression which is essential for neural tube closure. CRISPR-Cas9 gene
editing in the promoter region abolishes Sox2ot gene expression and
induces NTDs in mouse. Sox2ot down-regulation may contribute to the
etiology of maternal diabetes-induced NTDs.

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Artificial Sweeteners during Pregnancy - Impact on Maternal
Metabolic Health. Jasmine F Plows†, Anna P Ponnampalam, Mark H
Vickers, Clare M Reynolds*. University of Auckland, Auckland, New
Zealand.
INTRODUCTION: Sugar-sweetened beverage (SSB) consumption
has been associated with the development of metabolic diseases such as
obesity, type 2 diabetes, and cardiovascular disease. Low- or no-calorie
artificially-sweetened beverages (ASB) are therefore often promoted as an
alternative. However, there is little evidence that substituting SSBs with
ASBs meaningfully reduces metabolic disease risk, and evidence for the
safety of ASBs during pregnancy is lacking. We examined the effects of
SSB and ASB consumption during pregnancy in mice, and hypothesized
that both SSBs and ASBs would impair maternal metabolic function.
METHODS: Pregnant female C57BL/6J mice were randomized to
receive either water (CD), high fructose corn syrup (Fr), or the artificial
sweetener acesulfame-K (AS) in their drinking water, from gestational
day (GD) 0.5 (n=8/group). Doses of Fr and AS were prepared daily to
represent the quantities consumed in 500mL of commercially-available
SSBs and ASBs, respectively. Body weight and food/water intake were
assessed every second day, an oral glucose tolerance test (OGTT) was

Saturday Orals

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Reproductive Sciences Vol. 25, Supplement 1, March 2018

Table of Contents for the Digital Edition of SRI Supplement to Reproductive Sciences - Volume 25 Number 1 - March 2018