P–563 Assessing ovarian age: Could we use leukocyte telomere length as a surrogate marker of cumulus cells telomere content?
Authors: Lar. Molina E E , Franasiak J M , Tao X , Florensa M , Martin M , Molla-Zaragoza P , Díaz-Gimeno P , Ballesteros A , Seli E , Pellicer A 
Published: August 2021

Abstract

Study question
Is leukocyte telomere length (LTL) correlated with cumulus cells telomere length (CCTL) in an age-heterogeneous women population?

Summary answer
LTL showed a positive correlation with CCTL in the studied population. Hence, its potential value as indicator of ovarian age would deserve further evaluation.

What is known already
Progressive telomere shortening has been related to ovarian aging and genomic instability during early development. A positive correlation between short telomere length of the first polar body and aneuploidy rate has been reported. CCTL has shown to be a biomarker of oocyte and embryo quality, but its assessment is impractical. LTL has been proposed as a surrogate of TL of follicular cells, but telomere lengthening through folliculogenesis could be controlled by different mechanisms. Thus, we aimed to determine if LTL in an age-heterogeneous population is correlated with CCTL and therefore considered an accurate surrogate for telomere length in the ovary.

Study design, size, duration
In this prospective non-interventional cohort study, 35 egg donors and 17 women undergoing Preimplantation Genetic Testing for Aneuploidy (PGT-A) treatment were included during sixteen months. Following controlled ovarian stimulation determined by treating physicians, oocyte retrieval was performed 36 hours after final maturation induction. Cumulus cells (CC) for telomere length (TL) measurement were obtained after the pick-up and oocyte stripping. A blood sample was collected through peripheral venous access for LTL measurement.

Participants/materials, setting, methods
Genomic DNA of CC and leukocytes from the 52 subjects was isolated. Average delta cycle threshold (ΔCt) was determined using a SYBR green quantitative real-time PCR protocol for relative TL. For normalization of measurements, a Taqman assay for the multicopy gene Alu was performed. ΔCtL and ΔCtCC were compared by a paired t-test analysis and the fold change was calculated. Additionally, the association between them and patient age was analyzed by a Pearson correlation test.

Main results and the role of chance
Mean participant’s age was 29.94 ± 7.55 years and mean values for ΔCtL and ΔCtCC were 7.99 ± 0.53 and 7.46 ± 0.75, respectively. A positive significant correlation was found between age and ΔCt (ΔCtL: R2=0.71, p-value=5.18e–09; ΔCtCC: R2=0.47, p-value=0.00049). Since ΔCt values are inversely proportional to the amount of nucleic acids amplified and, therefore, to the telomere length, this correlation means that TL in both cell types decreases as women age. Additionally, ΔCtL was significantly higher than ΔCtCC (ΔCt fold change: 0.93, p-value=9e–07), meaning that CC showed significantly longer telomeres than leukocytes, thus supporting our previous published results in young egg donors. When analyzing the ΔCtL and ΔCtCC in these age-heterogeneous sample, a positive moderate and significant correlation was observed (R2=0.42, p-value=0.002). Thus, LTL could be suggested as a potential indicator of CCTL and therefore as a candidate for a biological marker of ovarian aging.

Limitations, reasons for caution
The sample size of this study was moderate and perhaps increasing the number of subjects might give additional strength to our findings. In addition, although relative telomere length allowed for adequate comparison between subjects, this method did not allow for absolute TL measurement.
Wider implications of the findings: While reproductive implications of LTL measurement need to be further studied, our results support the potential usefulness of LTL measurement as an indicator of CCTL and ovarian aging when analyzing an age-heterogeneous population. Further, our findings suggest that CC could possess different mechanisms to cope against telomere length shortening.

Trial registration number
Not applicable