SRI Supplement to Reproductive Sciences - Volume 25 Number 1 - March 2018 - 187A

Scientific Abstracts

MOR was below the assay detection. Confirmatory IHC on fetal membrane
rolls (n=5 patients) confirmed the protein expression of OGFR, DOR and
KOR. However, interestingly MOR protein expression was clearly seen
in all cells of the fetal membranes, with particularly remarkable label in
AMC. Western blotting is ongoing to further confirm the expression and
activation of these receptors.
CONCLUSION: This data suggests that the fetal membrane may a
target for opioids and thus may influence the behavior of the human fetal
membranes if taken during pregnancy. Further research into the signaling
pathways of these receptors in the fetal membrane is necessary to increase
our understanding the implications of prenatal opioid exposure.

Cervicovaginal Epithelial Cells Differentially Express Acetate and
CXCL8 in Co-Culture with Preterm Birth-Associated Bacteria and
Protective Lactobacillus Species. Emmanuel Amabebe†, Haoyuan
Qiao, Craig Murdoch, Dilly Anumba*. University of Sheffield, Sheffield,
United Kingdom.
INTRODUCTION: An imbalance in the distribution of the vaginal
microflora during pregnancy may be associated with infection-associated
preterm birth (PTB). Colonization by anaerobic bacterial species as
in bacterial vaginosis (BV) may predispose to PTB if the protective
dominance by Lactobacillus spp. is lost. Having previously demonstrated
signature microbial and metabolic profiles of PTB from mid-trimester
cervicovaginal fluid samples, we investigated the expression of infectionassociated metabolite (acetate) and pro-inflammatory chemokine (CXCL8
or IL-8) by cervicovaginal epithelial cells infected with BV-associated
bacteria (BVAB) or Lactobacillus spp.
METHODS: The concentrations of acetate (enzyme-based
spectrophotometry, K-ACETGK 08/14, Megazyme) and CXCL8 (ELISA,
BD OptEIA™) were determined and compared between monolayercultured HeLa cells infected with strains of Gardnerella vaginalis (ATCC
14019), Mobiluncus curtisii (ATCC 35241) or L. crispatus (ATCC 33820)
alone or in combination with each other over a 72-hour period (MOI=10).
Uninfected HeLa cells were used as controls.
RESULTS: HeLa cells infected with G. vaginalis (p<0.01) or M. curtisii
(p=0.01) produced higher concentrations of CXCL8 compared to HeLa
cells infected with L. crispatus or uninfected cells over time. The addition
of L. crispatus to G. vaginalis or M. curtisii infection models reduced
expression levels of CXCL8 compared to G. vaginalis or M. curtisii
infection alone.
In addition, acetate concentration increased in a time-dependent
nonsignificant manner in all infected cultures compared to uninfected
controls, except those with G. vaginalis where a decrease was observed
after 72 hours only. The addition of L. crispatus to the infection models
of HeLa cells with either G. vaginalis or M. curtisii, decreased acetate
production compared to HeLa cells cultured with G. vaginalis or M.
curtisii alone.
CONCLUSION: Infection of cervicovaginal epithelial cells by BVAB
such as G. vaginalis and M. curtisii stimulate secretion of acetate and
the pro-inflammatory chemokine CXCL8. The presence of L. crispatus
attenuate these potentially deleterious effects by reducing acetate and
CXCL8 levels, consistent with a role for L. crispatus in promoting
vaginal floral health.

F-015
Flurofamide Fails to Resolve Intrauterine Ureaplasma Parvum
Infection in a Sheep Model of Pregnancy. Haruo Usuda,3 Matthew
Kemp,3 Mike Beeton,1 Matthew Payne,3 Owen Brad Spiller.2 1Cardiff
Metropolitan University, Cardiff, United Kingdom; 2Cardiff University,
Cardiff, United Kingdom; 3The University of Western Australia, Crawley
WA, Australia.
INTRODUCTION: Intrauterine infection involving Ureaplasma parvum
(UP) is commonly associated with preterm birth and adverse neonatal
outcomes. Flurofamide is a urease inhibitor with specific antimicrobial
activity for UP. We first established materno-fetal pharmacokinetics for
flurofamide, then tested the use of flurofamide to treat intrauterine UP
infection using a sheep model of pregnancy.

187A

METHODS: Pharmacokinetic study: Six ewes carrying a single fetus (92
days' gestation) underwent recovery surgery to place maternal and fetal
jugular catheters and an amniotic fluid (AF) catheter. Animals received
a single maternal intravenous bolus of flurofamide (10 mg/kg). Maternal
and fetal blood and amniotic fluid samples were collected over 24 hours
for measurement of flurofamide by mass spectrometry. Efficacy study:
Flurofamide minimum inhibitory concentration (MIC) values against UP
were established using an in vitro UP susceptibility assay. Ewes carrying
a single fetus (88 days' gestation) were randomised to receive ultrasoundguided intraamniotic injections of either: sterile saline (Group 1, negative
control group); or 10^7 colour change units of UP (Groups 2, positive
control group and 3, intervention group). After 3 days, animals in Group
3 received a 5-day course of maternal intravenous flurofamide treatment,
with dosing based on pharmacokinetic study data. AF was sampled 8
days after treatment commenced. Lambs were delivered 12 days after
treatment commencing, euthanised and sampled to measure UP load and
fetal inflammation. Data were assessed for variance and distribution and
tested with ANOVA as appropriate.
RESULTS: Pharmacokinetic study: The in vitro MIC 90 for flurofamide
was 0.75±0.4 μg/mL. Maternal flurofamide readily crossed the placenta
giving maximal AF and fetal plasma concentrations of 1.4 μg/mL at 2
hours and 0.4 μg/mL at 9 hours, respectively. Efficacy study: All Group 1
animals were negative for UP. In Groups 2 and 3, there was no statistically
significant differences in UP concentrations in AF at either 8 or 12 days
post-treatment, or in fetal plasma at delivery.
CONCLUSION: In preterm lambs infected with UP, maternal treatment
with flurofamide failed to resolve infection, despite the use of a dosing
regimen achieving supra-MIC flurofamide concentrations in the AF and
fetal plasma.

F-016
Administration of Subclinical Doses of Lipopolysaccharide
Simultaneously Reverses Dysbiosis and Protects High Fat Diet
Mice from Preterm Delivery. Clarence Manuel†,2 Sandra E Reznik*.1,2
1
Montefiore Medical Center/Albert Einstein College of Medicine, Bronx,
NY, United States; 2St. John's University, Queens, NY, United States.
INTRODUCTION: Despite the clinical associations between high-fat
diet (HFD) and pre-term birth (PTB), the mechanisms of HFD triggered
PTB are unresolved. To gain insight into these issues, we designed in-vitro
and in-vivo models of HFD potentiated PTB. Previous studies reported that
maternal HFD consumption alters the diversity of commensal microbiota,
and may trigger PTB, although the links are poorly understood. We
hypothesized that HFD influences PTB by increasing systemic oxidative
stress and modifying the bacterial composition in the gut.
METHODS: Human placental HTR-8/SVneo (HTR-8) trophoblasts were
cultured for 12hrs in either 0.5 mM palmitic acid (PA) or linoleic acid (LA)
in the absence or presence of lipopolysaccharide (LPS) or lipoteichoic
acid (LTA) (1µg/ml) for an additional 24hrs. MTT Cell Proliferation
Assay was used to assess viability. Total antioxidant capacity (TAC),
lipid peroxidation, H2O2 production, DNA/RNA oxidative damage, and
Heme-Oxygenase-1 (HO-1) activity assays were used to assess oxidative
stress. C57BL6/J timed pregnant mice were maintained on either a
HFD (60% fat), or normal chow diet (NCD, 13.4% fat), beginning eight
weeks before pregnancy. A weekly subclinical dose (0.2 mg/kg) of LPS
was administered intraperitoneally (ip) to half the mice throughout the
regimen, and mice were then administered 0.3 mg/kg of LPS ip on
E15.5. We assessed the time of delivery, fetal rescue, maternal oxidative
stress response, and the diversity of maternal gut microbiota. 16S rRNA
sequencing was used to assess microbiota of fecal samples. ELISA was
used to measure serum progesterone levels.
RESULTS: PA incubation impairs cell viability (***p<0.0001), decreases
TAC (*p<0.05), and increases lipid peroxidation (*p<0.05); whereas
treatment with LA improves TAC (*p< 0.05), and increases the activity
of HO-1 (*p<0.05). Pregnant HFD mice deliver more pups (*p<0.05)
within 24hrs after LPS administration than pregnant NCD mice, which
was associated with reduced TAC (*p< 0.05), increased lipid peroxidation
(***p<0.0001) and dysbiosis of gut microbiome. LPS priming protects
HFD dams from PTB (**p-<.001), and rescues pups from fetal death.

Friday Posters

F-014

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