P-543 The role of FMRP-associated microRNAs miR-125b and miR-132 and their target genes in the regulation of ovarian function
Authors: Vilkaite A , Nguyen X.P , Bender U , Hinderhofer K , Strowitzki T , Rehnitz J 
Published: June 2022

Abstract

Study question
Do FMRP (Fragile-X-Mental Retardation 1 Protein)-associated miRNA miR-125b, miR-132 and their target genes participate in the FMR1/FMRP regulatory loop and result in altered ovarian response?

Summary answer
miR-125b and its target gene TP53 (Tumor Protein P53) are downregulated in follicular fluid and granulosa cells of poor ovarian responders, respectively.

What is known already
MicroRNAs are a class of endogenous non-coding RNAs that play an important role in ovarian function and follicular development. It was found that miRNAs directly interact with FMRP and are involved in translational repression. MicroRNAs miR-125b and miR-132 have been shown to associate with FMRP in neural cells, but their relationship in the human ovary has not yet been evaluated. TP53 and LIN28A (Lin-28 Homolog A) are targets of miR-125b, the former regulating cell cycle and apoptosis, and the latter involved in germ cell formation. Investigating these molecules more thoroughly may provide insight into the pathogenesis of poor ovarian response.

Study design, size, duration
RNA from granulosa cells (GCs) and follicular fluid (FF) from poor (POR, nGC = 26, nFF = 12) and normal (NOR, nGC = 31, nFF = 16) ovarian responders was extracted and reverse transcription was carried out. RT-PCR was then performed to compare the relative expression of the targets, with NOR serving as the control group. Sample collection is ongoing and began in 2013.

Participants/materials, setting, methods
57 patients who underwent ovarian stimulation were recruited and divided into two groups based on their ovarian response according to the Bologna criteria. GCs (n = 57) and FF (n = 28) were collected during oocyte retrieval. RNA was extracted and specific or non-specific cDNA was synthesized depending on the target. RT-PCR was performed with TaqMan reagents. Statistical analysis was performed using the 2–ΔΔCt method, Student’s t-test and Pearson’s coefficient.

Main results and the role of chance
We found that miR-125b was significantly downregulated in FF of the POR group (p = 0,0190), with no similar effect observed in GCs. The miR-132 levels showed no difference between the NOR and POR groups. This finding suggests that miR-125b may play a role in the cell-to-cell communication, and its suppression may be related to poor ovarian response. We then investigated two potential miR-125b target genes, TP53 and LIN28A, and their relation to FMR1. While LIN28A is similarly expressed in both groups, TP53 is significantly downregulated in GCs of POR group (p = 0,0086). Interestingly, TP53 expression correlates moderately positively with FMR1&gt; (Pearson’s r = 0,5) in the NOR group, while no significant correlation is evident in the POR group (Pearson’s r = 0,19). These results suggest the existence of regulatory mechanisms involving miR-125b, TP53, FMRP and FMR1 that may be disrupted in case of poor ovarian response.

Limitations, reasons for caution
More patient samples are needed to confirm the current results. A possible age-related influence cannot be excluded because of the existing difference between the groups.

Wider implications of the findings
For the first time, the role of miR-125b together with its target TP53 was investigated in the human ovary, suggesting that they are involved in the FMR1/FMRP regulatory loop and thus providing more insight into the etiology and pathogenesis of poor ovarian response.

Trial registration number
not applicable