Abstract
PURPOSE: To investigate how mitochondria and telomeres evolve during early follicle growth in human preantral follicles, and how these changes relate to reproductive age. METHODS: This prospective study included ovarian cortex and peripheral blood samples from 21 women, categorized into young (20-26, n = 7), mid (27-36, n = 6) and advanced (37-46, n = 8) reproductive age groups. Ovarian biopsies were processed for (i) follicle isolation for mitochondrial marker assessment (MitoTracker Deep Red and MitoSOX live staining) and telomerase subunit expression (hTERT, hTERC) by droplet digital PCR; and (ii) histological evaluation of the ovarian follicle pool, mitochondrial mass (TOMM20) and telomere length in different ovarian compartments (oocytes, granulosa cells and stroma) by fluorescence in situ hybridization (FISH). Leukocyte telomere length was measured by flow-FISH. RESULTS: Mitochondrial mass (TOMM20) in the follicle pool was stable across age groups, but increased significantly during early follicle growth in younger patients (p < 0.01). Active mitochondria did not decline with age, but reactive oxygen species (ROS) generation showed complex age-related patterns, with younger subjects exhibiting lower ROS levels during follicle growth (p < 0.01) than other age groups. Telomere length was stable in granulosa cells and oocytes within primordial follicles, but increased during growth in younger age groups. Telomerase subunit expression was detected in preantral follicles without any significant age or size differences. CONCLUSIONS: These results point to greater mitochondrial mass, mitochondrial ability to limit ROS generation, and telomere dynamics changing with increasing age in early growing follicles, potentially guiding the development of strategies to enhance fertility outcomes during the early stages of folliculogenesis.