mRNA EXPRESSION OF APOPTOTIC MARKERS AND ANTIOXIDANT ENZYMES IN CUMULUS AND GRANUOLSE CELLS FROM PERIOVULATORY

FOLLICLES OF YOUNG WOMEN WITH LOW OVARIAN RESERVE

R. NUÑEZ-CALONGE1, S.CORTES1, L.RANCAN2, E.VARA2, L.ORTEGA1, P.CABALLERO1, J.FERNANDEZ-TRESGUERRES2.

1 Clínica Tambre, Madrid , SPAIN2 Dpto. Fisiología, Universidad Complutense Madrid

Introduction

Introduction

Objective

To investigate apoptosis and oxidative stress in granulose (GC) and cumulus cells (CC) from periovulatory follicles of young women with low response to COH. To this end, we compared the levels of mRNA coding for antioxidant enzymes and apoptotic markers in women with low response in in vitro fertilization (IVF) and fertile oocyte donors as controls.

Methods

20 healthy,

fertile, Donors(< 35 years old)

15 low responders

Patients(< 35 years old)

COS

rFSH+ antagonist GnRH

Patients enrolled in this study were selected on the basis of their ovarian response (with normal FSH and/or AMH levels) and only patients with at least a previous cycle with low response were included.

Methods

Superoxide dismutase (SOD) Catalase Antioxidant enzymes

Caspase 3 and Caspase 9 Bcl-2 Apoptotic markers BAX

We carried out RT-PCR analysis on CGs and CCs from pooled follicular aspirates of each pacient:

Results

Results

Mean ± SD Donors Patients P value

Catalase 1,47±0,11 0,87±0,06 .005

SOD 1,55±0,07 1,42±0,02 NS

Caspase 3 0,34 ±0,01 0,71±0,049 .0006

Caspase 9 0,81±0,06 1,37±0,1 .009

BAX 1,19±0,02 2,11±0,2 .0018

Bcl2 1,02±0,02 1,46±0,09 .006

Levels of mRNA coding for antioxidant enzymes and apoptotic markers in GCs of low responders patients compared with controls.

Results

Mean ± SD Donors Patients P value

Catalase 1,66±0,14 0,99±0,06 .0037

SOD 1,19±0,07 1,2±0,1 NS

Caspase 3 1,008 ±0,12 1,01±0,006 NS

Caspase 9 1,05±0,06 1,1±0,07 NS

BAX 1,26±0,06 2,95±0,19 0.08

Bcl2 1,35±0,05 1,37±0,03 NS

Levels of mRNA coding for antioxidant enzymes and apoptotic markers in CCs of low responders patients compared with controls.

Results

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Catalase mRNA expression levels in cumulus and granulosa cells from donors and patients. Values are means ± SEM

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Results

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BAX mRNA expression levels in cumulus and granulosa cells from donors and patients. Values are means ± SEM

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Conclusions

• Our results suggest a lower antioxidant capacity and an increased apoptosis level in CC and GC of women with low response to COH compared with fertile oocyte donors.

• Otherwise, mRNAs coding for antioxidant enzymes and apoptotic markers are differentially expressed in granulose and cumulus cells.

• In addition, in this study, we demonstrate a significant difference between women with poor and normal responses to COH and we found there were a good correlation between increased GCs oxidative stress, apoptosis and low ovarian response, suggesting that oxidative stress could influence oocyte production.

Conclusions

• The relations of some antioxidant enzymes like SOD, and

apoptotic markers in CC with OE and its implication in ovarian reserve needs to be addressed in further studies on a larger sample.

• Identifying optimal biomarkers that reliably predict ovarian response in the clinical setting could lead to developing antioxidant therapies for these poorer prognosis women undergoing IVF treatment.

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