SRI Supplement to Reproductive Sciences - Volume 25 Number 1 - March 2018 - 103A

Scientific Abstracts

provided by our ultrasound system. Variations of SWS with gestational
age, cervix side, and parity were statistically assessed with a linear mixed
effects model.
RESULTS: Pregnancy lasted between 23.3 and 25.6 weeks among
subjects. SWS decreased by 6% per week (a multiplicative factor of 0.94
per week; 95% CI 0.93-0.95, p<0.0001). This corresponds to an 80%
reduction in SWS over the duration of pregnancy from the initial value of
4.60 (95% CI 3.93-5.37) m/s. SWS was 7% lower in the posterior cervix.
Variations with parity were not significant.
CONCLUSION: Our SWEI implementation demonstrated an 80%
reduction of SWS during pregnancy in the Rhesus Macaque cervix. It
also exhibited significant differences in SWS in anterior versus posterior
cervix - particularly in early pregnancy. This SWS reduction during
pregnancy is larger than the previously reported in ewes (54%, Peralta
et al, PLoS ONE, 2015) and humans (21%, Muller et al, UMB, 2015),
probably due to the stiffer nature of the Rhesus macaque cervix and our
optimized SWEI implementation. We are extending our analysis to extract
information about cervix viscosity to further advance our understanding
of the dynamic and complex mechanical properties of the pregnant cervix.

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O-139
A Biomechanical Signature to Assess Cervical Extracellular
Components in Term and Preterm Birth. Charles Jayyosi†,1 Nicole
Lee,1 Shanmugasundaram Nallasamy,2 Alexandra Willcockson,2 Mala
Mahendroo*,2 Kristin Myers*.1 1Columbia University, New York, NY,
United States; 2UT Southwestern Medical Center, Dallas, TX, United
States.
INTRODUCTION: Characterizing cervical mechanical property changes
in normal and abnormal pregnancy is key to understanding preterm birth
(PTB) and develop successful therapies. The microstructural analysis of
mouse PTB models have identified unique modifications to the collagen
and elastic fiber networks that are distinct from term ripening. Hence,

103A

we developed a specific biomechanical test to tease out the mechanical
contribution of these components and assess the tissue's overall ability to
recover from loading in term and premature cervical ripening.
METHODS: Cyclic biomechanical tensile tests (Fig.1a) were conducted
on isolated cervices from normal mice at gestation day 15 and 18 and
two PTB models: a progesterone withdrawal model using mifepristone
(RU486) and an infection mediated model using lipopolysaccharide (LPS)
with associated sham group. We assessed the tangent modulus E [MPa]
at high and low stretch (associated respectively with the stiffness of the
collagen and elastic fiber networks), the stretch shift (associated with tissue
damage and its inability to recover from loading), and the hysteresis area
(associated with total dissipated energy) (Fig.1a).
RESULTS: The E at low stretch decreases significantly from 3.85e-4
MPa for d15 to 2.34e-4 MPa for d18 and 2.19e-4 for RU486 (one way
ANOVA, p<0.05, n=5) (Fig.1b). The LPS, RU486 and d18 groups present
a higher stretch shift (0.25, 0.23 and 0.19 respectively) compared to d15
and sham (0.14 and 0.10) which demonstrates a reduced ability to recover
from loading. LPS treated samples broke earlier before reaching the last
load level (20% reached the end), while other groups usually withstood
the entire loading regimen (60% of RU486 samples, 80% of d15 and
sham, 100% of d18).
CONCLUSION: Our data suggest elastic fibers play a critical role in
the evolution of cervical mechanical properties in late pregnancy as
seen with the significant drop of E at low stretch for d18 and RU486.
The previously noted differences in ECM architecture between the PTB
models are supported by the distinct mechanical signatures identified here.
The LPS group presents a weaker structure unable to withstand loading
while RU486 is relatively similar to term cervical ripening. These findings
demonstrate that in addition to the integrity of collagen and elastic fibers
in the ECM, the particular fibers organization within the network is a main
contributor to the mechanical function of the tissue.
*Figure(s) will be available online.

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Oxytocin Inhibits the Potassium Channel SLO2.1: A Novel
Mechanism of Regulating Uterine Excitability. Celia M Santi*, Juan J
Ferreira†, Alice Buttler†, Richard Stewart†, Ana Laura González-Cota†,
Pascale Lybaert†, Chinwendu Amazu†, Erin L Reinl†, Monali WaklePrabagaran†, Lawrence B Salkoff*, Sarah K England*. Washington
University, Saint Louis, MO, United States.
INTRODUCTION: At the end of pregnancy, the uterus must transition
from quiescence to contraction. An important contributor to this transition
is the membrane potential of myometrial smooth muscle cells (MSMCs).
Depolarization of the MSMC membrane leads to influx of calcium, which
activates myosin and leads to contraction. However, the ion channels
controlling the membrane potential and their mode of regulation have
not been fully defined. Here, we investigated the role and regulation of
the previously overlooked potassium channel SLO2.1. This channel is
of interest because it is predicted to be open at the resting membrane
potential of MSMCs and thus promote quiescence.
METHODS: We performed electrophysiological measurements in
primary MSMCs isolated from term non-laboring women and in
the transformed MSMC line hTERT. Additionally, we performed
immunofluorescence, PCR, and proximity ligation assays to determine
SLO2.1 expression and protein-protein interactions in these cells.
RESULTS: PCR and immunofluorescence provided the first evidence
that primary MSMCs express SLO2.1 channels. Electrophysiological
experiments showed that SLO2.1 channels express sodium-dependent
potassium currents in primary MSMCs, and that these currents can be
inhibited by limiting sodium ion entry and by lowering intracellular
sodium ion concentrations. Additionally, we show that SLO2.1 currents
are inhibited by oxytocin in both hTERTs and primary MSMCs. This
inhibition occurs via oxytocin binding to the oxytocin receptor and
activating protein kinase C.
CONCLUSION: These results reveal a new pathway by which oxytocin
could promote uterine contraction at the end of pregnancy. Oxytocin could
bind to its receptor, whose expression increases at the end of pregnancy,

Saturday Orals

Proliferative and Migratory Properties Reveals Metastate of Amnion
Cells Mediated by Epithelial to Mesenchymal and Mesenchymal to
Epithelial Transitions. Lauren Richardson†, Jayshil Trivedi, Ramkumar
Menon*. UTMB, Galveston, TX, United States.
INTRODUCTION: Fetal membranes develop microfractures throughout
gestation but they are resealed to maintain membrane integrity and to
support pregnancy. The mechanisms that reseal membrane microfractures
are unclear. We determined that amniotic membrane cells (AEC) can
proliferate, migrate and remodel through epithelial to mesenchymal
transitions (EMT) followed by a reversal, mesenchymal to epithelial
transition (MET), to seal microfractures promoting healing in utero.
METHODS: Human primary AECs were grown to 80-90% confluence
and a scratched was created mimicking wound with 200ul pipette tip.
Following treatments (for 48 hours) were performed; 1) normal cell culture
conditions (control) 2) term-not-in-labor amniotic fluid (TNIL AF), 3)
oxidative stress induction by cigarette smoke extract (CSE) to mimic term
labor, or 4) CSE plus antioxidant N-acetyl-l-cysteine (NAC). Images were
collected with bright field and confocal microscopy at frequent intervals up
to 48 hours to track migration rates and to determine cytokeratin (CK)18/
vimentin ratios of migrating cells to verify EMT/MET changes. One- Way
ANOVA was used to determine statistical significance greater than 0.05.
RESULTS: In our scratch assay, AECs proliferated, migrated, and sealed
the scratch site within 48 hrs. Treatments with TNIL AF sealed scratch
edges faster than controls (p=0.0132). CSE treatment did not seal the
scratch (p<0.0001), whereas co-treatment with NAC improved scratch
healing (p=0.0005). Vimentin+ cells with epithelial morphology was
predominant prior to scratch and at sealed edges whereas CK-18+ cells
with mesenchymal morphology was seen at leading edges.
CONCLUSION: AECs are pluripotent cells with capacity to proliferate,
migrate and transition. Their natural ability to seal microfractures in fetal
membranes is mediated through EMT-MET. Nourishment by amniotic
fluid helps with sealing whereas OS prevents this process. We postulate
that at term or in response to OS inducing risk factors in preterm, reduction
of microfracture sealing capacity can predispose membranes to rupture.

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