Background: Oocytes may undergo two types of aging. The first is induced by exposure to an aged ovarian microenvironment before being ovulated, known as 'reproductive or maternal aging', and the second by either a prolonged stay in the oviduct before fertilization or in vitro aging prior to insemination, known as 'postovulatory aging'. However, the molecular mechanisms underlying these aging processes remain to be elucidated. As telomere shortening in cultured somatic cells triggers replicative senescence, telomere shortening in oocytes during reproductive and postovulatory aging may predict developmental competence. This study aimed to ascertain the mechanisms underlying altered telomere biology in mouse oocytes during reproductive and postovulatory aging.
Methods: We studied Tert expression patterns, telomerase activity, cytosolic reactive oxygen species (ROS) production, and telomere length in fresh oocytes from young versus reproductively-aged female mice retrieved from oviducts at 14 h post-human chorionic gonadotropin (hCG), in vivo or in vitro postovulatory-aged mouse oocytes at 23 h post-hCG. Oocytes were collected from super-ovulated C57BL/6 J mice of 6-8 weeks or 42-48 weeks of age. mRNA and protein expressions of the Tert gene were quantified using real-time quantitative reverse transcriptase polymerase chain reaction (Q-PCR) and immunochemistry. Telomerase activity was measured by a telomeric repeat amplification protocol assay, while telomere length was measured by Q-PCR and quantitative fluorescence in situ hybridization analyses.
Results: The abundance of Tert expression in oocytes significantly decreased during reproductive and postovulatory aging. Immunofluorescent staining clearly demonstrated an altered pattern and intensity of TERT protein expression in oocytes during reproductive aging. Furthermore, relative telomerase activity (RTA) in oocytes from reproductively-aged females was significantly lower than that in oocytes from young females. In contrast, RTA in postovulatory-aged oocytes was similar to that in fresh oocytes. Oocytes from reproductively-aged females and postovulatory-aged oocytes showed higher ROS levels than oocytes from young females. Relative telomere length (RTL) was remarkably shorter in oocytes from reproductively-aged females compared to oocytes from young females. However, postovulatory aging had no significant effect on RTL of oocytes.
Conclusions: Long-term adverse effects of low telomerase activity and increased ROS exposure are likely associated with telomere shortening in oocytes from reproductively-aged female mice.