Aging affects gene expression in pathways essential for energy metabolism, DNA repair, cell cycle regulation, and antioxidant defenses, directly affecting oocyte quality and viability. Single-cell RNA deep sequencing studies of aged versus young human MII oocytes revealed many differentially expressed genes. In addition, single human oocyte transcriptome analysis at both germinal vesicle (GV) and MII stages revealed distinct stage-dependent pathways impacted by aging, with a decrease in mitochondrial-related transcripts from GV to MII oocytes, and a much greater reduction in MII oocytes with advanced age. Our aim was to investigate the age-related differences in gene expression of germinal vesicle (GV) oocytes between young and advanced age patients. Immature GV oocytes were donated by 6 patients, divided into two age groups: The "Young" group (ages 16-29) had three participants (mean age: 23.3 ± 6.6 years), and the "Elderly" group (ages 38-40) included three participants (mean age: 39 ± 1 year). After retrieval, oocytes were denuded and donated GV oocytes were cryopreserved at -1960C until analysis. For library preparation, we used the NEBNext® Single Cell/Low Input RNA Library Prep Kit for Illumina, Sect. 1 (cat no. E6420S, New England Biolabs (NEB), USA), strictly adhering to the manufacturer's instructions. Gene expression quantification was performed using feature Counts from the Subread package (v1.5.3), and comprehensive quality control reports were generated using MultiQC (v1.25.1). To further corroborate the differential expression of hub genes associated with oocyte aging identified in our preliminary analysis, quantitative real-time PCR (qPCR) was performed for four selected hub genes (MYL4, POMZP3, and LINC002087). Of top 10 significantly differently expressed genes 7 (LINC02087, POMZP3, LINC02749, MYL4, AGPAT2, GCA, and LIMK1) were downregulated and 3 (CLEC3A, ARPP21, and CITED2) showed significant upregulation in young versus old oocytes. These genes underscore the impact of aging on critical oocyte pathways, including chromosomal stability, epigenetic regulation, mitochondrial function, immune response, structural integrity, and calcium signaling. Moreover, among these genes, LINC02087 was the most downregulated (log2FC = -7.66), while CITED2 showed the strongest upregulation (log2FC = 3.43) in young versus old oocytes. Following the RNA extraction of pooled GV oocytes of 8 elderly and 9 young donors' GV oocytes. We observed significant differences in gene expression levels between the two age groups, in line with the single-cell RNASeq. Understanding the effects of aging on the oocyte transcriptome could identify biomarkers that characterize good MII oocyte quality. The different genes expressions in aged oocytes highlight their potential contributions to oocyte quality and development. Moreover, by elucidating age-related changes across diverse cellular functions, this preliminary study opens avenues for therapeutic interventions aimed at extending reproductive longevity and optimizing outcomes in assisted reproductive technologies.
Orvieto et al. (Tue,) studied this question.