P-259 Cumulus cells mitochondrial membrane potential as a marker of oocyte competence in assisted reproduction
Authors: Reis S , Almeida-Santos T , Ramalho-Santos J , Sousa A.P 
Published: June 2022

Abstract

Study question
Is mitochondrial function of human Cumulus-oophurus Cells (hCCs) an important non-invasive biomarker for oocyte competence and a predictor of in vitro fertilisation (IVF) outcomes?

Summary answer
Advanced maternal age of women undergoing IVF is characterized by an impairment in cumulus cell mitochondrial function.

What is known already
The increasing delay of childbearing has led to urgent development of new non-invasive biomarkers of oocyte competency. Aging is a complex process that includes impairment of mitochondrial function and metabolic activity, increase of oxidative stress, alteration in the activity of several cell-signaling systems.
A great candidate to monitoring oocyte quality is the hCCs. During folliculogenesis there is an intense bidirectional communication between oocyte and its follicular cells. During oogenesis, CCs supply the oocyte with energy substrates and essential nutrients. Age-related mitochondrial dysfunction in hCCs was described to adversely affect oocyte growth and development.

Study design, size, duration
The objective of this study was to characterise the mitochondrial membrane potential (Δψm) of hCCs provided from patients undergoing ICSI (intracytoplasmic sperm injection) or oocyte cryopreservation in the Reproductive Medicine Unit of CHUC (Centro Hospitalar e Universitário de Coimbra, Coimbra, Portugal) between July and December 2021, to establish a predictive biomarker of oocyte competency and successful IVF outcomes.

Participants/materials, setting, methods
hCCs, collected from cumulus-oocyte complexes of 53 patients, were incubated with tetramethylrhodamine methyl ester (TMRM - 50 nM), MitoTracker Green (MTG 100 nM) and Hoechst (5µg/mL). Briefly, TMRM, due to its positive charge, accumulates within the mitochondria in a ΔΨm-depend manner, and emits red fluorescence. On the other hand, MTG is green-fluorescent mitochondrial dye which appears to localize in mitochondria regardless of Δψm. Therefore, the red/green fluorescence ratio can be measured as mitochondrial activity outcome.

Main results and the role of chance
The 53 patients studied were divided in two age groups (27 patients with &lt;35 and 26 with ≥ 35 years old). A decrease in hCCs Δψm of women with advanced maternal age (≥35 y) compared with hCCs recovered from women with &lt;35 was observed (P = 0.001). No differences were found in CCs Δψm in terms of infertility cause, body mass index and social habits (e.g., smoking) (P &gt; 0.05). We also investigated if Δψm can be an effective indicator of clinical outcomes in IVF techniques; however, no correlation was observed between Δψm and oocyte maturation, fertilization, D3 embryo development and blastocyst development rates.
Our preliminary data strongly support the idea that advanced reproductive age of women undergoing IVF is accompanied by impaired cumulus cell mitochondrial function.

Limitations, reasons for caution
More data is needed to be collected to support our preliminary results, such as increasing the sample size, as well as better characterisation of the mitochondrial function in hCCs.

Wider implications of the findings
This knowledge opens a new window of opportunities in the development of new strategies to mitigate the effects of oocyte aging on (in)fertility, through clinical therapies, such as enhancing the mitochondrial function in hCCs.

Trial registration number
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