P-346 Endometriosis-associated infertility changes the microRNA profile of cumulus cells with a notably pronounced effect on oocytes that failed fertilisation

Abstract Study question Does endometriosis-associated infertility (E) change the microRNA (miRNA) profile of cumulus cells (CCs) with a differential footprint between fertilized oocytes and oocytes that failed fertilization? Summary answer The CCs miRNA profile is altered in E versus control (C) patients (male/non-hormonal causes), with a differential signature between oocytes with successful or failed fertilization. What is known already For E patients, ART are often considered as an option to achieve successful pregnancy. But the impact of E on oocyte quality and its potential to be fertilized are still under debate with very few molecular clues supporting clinical data. Endometriosis causes alterations in gene expression in oocytes and CCs, but little is known about the changes at miRNA level. MicroRNAs are small non-coding RNAs with an important role in gene regulation. They are active players in oocyte maturation and fertilization, have been pointed as diagnostic biomarkers of E and thus, making them promising biomarkers of oocyte quality in E. Study design, size, duration Thirty-six CCs from metaphase II oocytes from 33 patients, i.e., 15 E patients and 18 C patients (11 male, 7 non-hormonal factors) undergoing ART between 2019-2023 at the University Hospital Zurich were analyzed. CCs samples were retrospectively classified into: 1) E patients with fertilized oocytes (2PN) (n = 11; E2); 2) E patients with oocytes that failed fertilization (0PN) (n = 4; E0); 3) C patients with 2PN oocytes (n = 12; C2); 4) C patients with 0PN oocytes (n = 7; C0). Participants/materials, setting, methods Small RNA-sequencing was performed for 36 single CCs samples. RNA-seq data was analyzed using the Galaxy Europe server (usegalaxy.eu) and differential expression analysis was performed with the Bioconductor R package EdgeR. The differential abundance of two miRNAs, miR-143-3p and miR-10b-5p, was further validated in 24 remaining samples by qPCR. MIENTURNET was used to identify miRNA target genes and DAVID and KEGG pathway databases to examine biological functions and pathways associated to the predicted target genes. Main results and the role of chance Ninety-nine differential abundant (DA) miRNAs were identified between E and C patients (false discovery rate, FDR <5%). Regardless the disease/health status, 31 miRNAs were DA between 2PN and 0PN oocytes (FDR <20%). In E patients, 27 DA miRNAs were found between E2 and E0 oocytes (FDR <5%). In C patients, 15 DA miRNAs were found between C2 and C0 oocytes (FDR <5%). Around 60% of the DA miRNAs in CCs from E versus C comparison have been found in endometrium or plasma samples from E patients. Comparisons among identified DA miRNAs revealed three sets of miRNAs: 1) affected only by E, 2) affected by both E and the potential to be fertilized; and 3) reporting a E2 miRNA signature in CCs of E patients but still with the potential to be fertilized. Target genes of DA miRNAs were involved in known biological functions and pathways affected by E, e.g.: oxidative stress, calcium signaling, mitochondria and spindle alterations, embryo development, estrogen signaling, and TGF-beta pathway. Interestingly, 11 DA miRNAs identified in set 1 - unique to E, and 3 DA miRNAs in set 3 - reflecting E2 oocyte signature, target 39 and 32 genes involved in progesterone-mediated oocyte maturation pathway, respectively. Limitations, reasons for caution The sample size of the study cannot be considered as large, with a particular small group of E patients with 0PN oocytes compared to the other experimental groups. We cannot rule out that other unknown individual genetic or clinical factors may have influenced the reported results. Wider implications of the findings Our findings can inspire new diagnosis approaches and personalized treatments with therapeutic miRNAs for underexpressed miRNAs or with miRNA inhibitors for miRNAs overexpressed in CCs of oocytes from E patients. The potential applications can be of value to E patients with low number and poor quality of oocytes retrieved. Trial registration number No