SRI Supplement to Reproductive Sciences - Volume 25 Number 1 - March 2018 - 86A

86A

Reproductive Sciences Vol. 25, Supplement 1, March 2018

RESULTS: PNA mice weights were like controls but GTT showed
elevated glucose (p=0.004; glycemia AUC 2121±40 in PNA vs 1891±54
in C, mmol/L*180min) and insulin (p=0.012; Insulin AUC 16462±1261
in PNA vs 10843±1484 in C, pmol/L*180min) levels. Fasting glucose,
insulin and Hb1Ac levels were similar. PNA mice had longer estrus
cycles (5.1±0.2 in PNA vs 4.4±0.2 in C; p=0.046) with irregular cycles.
AA mice were obese (AA: 27.6±0.6g vs. C:22.6±0.4. p<0.0001 at
16 weeks). GTT showed elevated glucose (p=0.03; glycemia AUC
3075±282 mmol/L*180min in AA vs. 2062±93 mmol/L*180min in C)
and insulin (p<0.0001; Insulin AUC 29941±1498 pmol/L*180min in AA
vs. 13298±1621 pmol/L*180min in C) levels. Fasting insulin was ~3 fold
higher in the AA (p<0.01; 120±10pmol/L in AA vs. 44±13pmol/L in C),
but fasting glucose was similar. HbA1c was higher in AA (4.8±0.1 in AA
vs 4.2±0.05 in C, p=0.0003). AA mice had irregular estrous with more
time spent in diestrus. Mitochondrial analysis of oocytes from PNA mice
showed decreased IMM (red/green ratio 1.2±0.09 in PNA vs 1.6±0.1
in C; p=0.016) and increased ROS (18.5±3.2 in PNA vs 9.6±1.6 in C;
p=0.04), but no difference in LP. Oocytes from AA showed reduced IMM
(red/green ratio 1.3±0.07 in AA vs. 1.7±0.2 in C; p<0.05); however, no
differences in ROS or LP.
CONCLUSION: Our results show that in PCOS, PNA exposure
adversely affects oocyte mitochondrial function compared to AA exposure,
despite AA exposure resulting in obesity and a more impaired metabolic
phenotype.
*Figure(s) will be available online.

O-087
Steroidogenic Factor 1 (SF-1) is Essential for Sertoli Cell Survival
in the Developing Fetal Testis. Prashanth Anamthathmakula†, Chandra
S Miryala, Jennifer C Condon, Pancharatnam Jeyasuria. Wayne State
University, Detroit, MI, United States.
INTRODUCTION: High levels of SF-1 in the developing Sertoli cell at
the peak of SRY (sex determining region of the Y chromosome) expression
implies a vital role for SF-1 in male differentiation and/or function of
the Sertoli cell. In this study we conditionally ablated SF-1 in the Sertoli
cell (SC-SF-1-/-) of the developing testis post sex determination using the
Amh-Cre mouse model, which ablates SF-1 function at E14.5.
METHODS: SF-1 conditional knockout (SC-SF-1-/-) mice were generated
by mating male homozygous Sf-1Lox/Lox mice with Amh-Cre/Sf-1Lox/Lox
females. Sf-1Lox/Lox male embryos served as controls. Immunostaining for
SF-1, SOX9, VASA, HSD3B2, GATA4 were performed on embryonic
testes at E15-E18.5. Apoptosis and cell proliferation were assessed by
TUNEL assay and BRDU incorporation respectively. Immunoblotting was
performed in the whole cell lysates of embryonic testes to evaluate the
expression of p-p53, MDM2, SOX9 and AMH. Histological analyses in
E15.5-E18.5 and adult (6 week) testis samples were performed to assess
gross anatomical changes.
RESULTS: BRDU staining when overlapped with SOX9 showed
definitive loss of proliferation of Sertoli cells at E15.5 in SC-SF-1-/testis. Immunoblot analysis of E15.5 SC-SF-1-/- testes also showed a
marked decline in SOX9 protein expression (p<0.001). SF-1 ablation at
E14.5 resulted in increased apoptosis in the SC-SF-1-/- testis as early as
E15, a half-day post Cre ablation. To understand the apoptotic trigger
we examined the anti-apoptotic MDM2 regulated pathway as we have
discovered that the MDM2 promoter contains multiple SF-1 response
elements. MDM2 expression decreased significantly at E15 and there
was a concomitant rise in p-p53 levels (p<0.01) in the SC-SF-1-/- testis
suggesting that apoptosis was mediated through the loss of MDM2. There
was considerable loss of cord structure seen post SF-1 ablation and loss of
germ cells as visualized by Vasa IHC. Sox9 and Amh, two genes controlled
by Sry and Sf-1, are greatly diminished in the SC-SF-1-/- testis by E15.5.
Interestingly, E18.5 testis demonstrated normal HSD3B2 expression in
SC-SF-1-/- mice suggesting intact Leydig cell androgen production. Testis
weights of 6-week-old adult SC-SF-1-/- mice were drastically lower than
controls (p<0.05), however, seminal vesicles weights in SC-SF-1-/- mice
were similar.
CONCLUSION: Apoptosis of Sertoli cells observed in SC-SF-1-/- mouse
recapitulates the apoptotic gonadal dysgenesis phenotype observed in

Scientific Abstracts

the SF-1 null mouse. We suggest that SF-1 acts as the survival factor
in vivo of the five transcription factors (GATA4, DMRT1, SOX9, WT1
and SF-1) that are minimally required for conversion of pluripotent
fibroblast to Sertoli cells, indicating the importance of SF-1 in Sertoli
cell differentiation.

O-088
Stem Cell Therapy of Polycystic Ovary Syndrome (PCOS):
Implantation of Human Bone Marrow Stem Cells Enhance Energy
Expenditure and Improves Hyperandrogenemia in an ImmuneCompetent Severe PCOS Mouse Model. Abdeljabar El Andaloussi†*,
Prosper Igboeli*, Mona Omar*, Ayman Al-Hendy*. Augusta University,
Augusta, GA, United States.
INTRODUCTION: Polycystic ovary syndrome (PCOS) is the most
common metabolic disorder affecting 5-20% of reproductive age women.
The clinical manifestations of PCOS include hyperandrogenism and
ovulatory dysfunction. Most PCOS treatment approaches aim to reverse
such metabolic challenges with lifestyle (exercise and diet) modifications,
or the use insulin-sensitizing medicines but with limited success. Human
bone marrow mesenchymal stem cells (h-MSC) secret a plethora of
important growth factors that can favorably modulate ovarian biology. In
this work, we explore the utility of h-MSC as a novel alternative approach
to ameliorate PCOS-related metabolic dysfunction and infertility.
METHODS: we established the PCOS animal model in presexual
(3 weeks) C57BL6 female mice by implanting letrozole (LET) pellet
subcutaneously in the neck area (5 mg/pellet, 90 days release). Mice were
randomly assigned to one of three groups: 1- Placebo control, untreated,
2- LET group, untreated and 3- LET group, treated with h-MSCs. The mice
weight-gain induced by LET treatment was monitored weekly. After 4
weeks of Letrozole treatment, h-MSCs (250,000 cell/ovary) were injected
into both ovaries using limited laparotomy. The control mice received
sham surgery and were injected with PBS. To study the impact of MSCs
on the metabolic criteria of PCOS, We evaluated energy expenditure in
h-MSCs-treated vs control animals by monitoring metabolic parameters
such us O2 volume, CO2 volume, respiratory exchange ratio (RER), heat
production, food intake and motility. Furthermore, gonadal fat tissues
collected after 2 weeks of treatment were examined by H&E staining
and immunohistochemistry for UCP-1 (Uncoupled protein-1, brown fat
marker expressed on mitochondria membrane, a marker for browning.
The analysis of fat mRNA markers (UCP-1, Prdm-16, and PGC-1a) was
done by Q-RT-PCR.
RESULTS: The engraftment of h-MSC for 8 weeks following PCOS
induction with LTZ, was able to significantly reduce the circulating levels
of androgen in treated PCOS mice (< 20 ng/dl) versus PCOS-untreated
group (28.1 ± 4.3)(P < 0.05). Furthermore, indirect calorimetry in opencircuit Oxymax chambers demonstrated significantly increased heat
production in PCOS mice engrafted with h-MSC compared to PCOS
placebo-treated control group (P < 0.05). Additionally, the expression
of UCP-1 was significantly increased in the white gonadal fat from
h-MSC-treated group versus placebo control both at mRNA and protein
levels (P < 0.005).
CONCLUSION: The induction of browning is a proof of beige fat
conversion to brown fat secondary to enhanced energy expenditure in
PCOS mice treated with h-MCS. Stem cell therapy might potentially be
a novel tool for effective treatment of PCOS-related morbidities.

O-089
Early-Pregnancy Inflammatory Markers and Preterm Birth
Classified According to Placental Features. Baiyang Sun†,2,3 Marnie
Bertolet*,3 Hyagriv N Simhan*,2,3 W Tony Parks*,1 Janet M Catov*.2,3
1
Dartmouth-Hitchcock Medical Center, Lebanon, NH, United States;
2
Magee-Womens Research Institute, Pittsburgh, PA, United States;
3
University of Pittsburgh, Pittsburgh, PA, United States.
INTRODUCTION: Preterm birth (PTB) is a heterogenous outcome;
early pregnancy biomarkers and placental features may identify subgroups
amenable to intervention. We hypothesized that inflammation and

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