The Mitochondrial-Derived Peptide MOTS-c Improves Bioenergetics in Aging-Associated Granulosa Cells and Oocytes

Ovarian aging compromises granulosa cell function, mitochondrial homeostasis, metabolism, and oocyte quality. We investigated whether the mitochondrial-derived peptide MOTS-c mitigates these defects using an aging-associated HGL5 granulosa cell model, ex vivo oocytes from young and aged mice, and public granulosa cell transcriptomic datasets. In HGL5 cells, MOTS-c increased mitochondrial mass and ATP-associated fluorescence without increasing cellular or mitochondrial reactive oxygen species, while mass-normalized membrane potential remained unchanged. Seahorse analysis showed increased ATP-linked and maximal respiration and spare respiratory capacity, reduced proton leak, and unchanged basal respiration. MOTS-c also remodeled expression of enzymes involved in glycolysis, lactate–pyruvate handling, and pyruvate–TCA metabolism. Although fission markers were largely unchanged, MOTS-c increased proteins associated with antioxidant defense, mitochondrial transport, cell junctions, adhesion signaling, and granulosa cell function, including NRF2, phosphorylated AIFM1, RHOT1, KIF5B, ZO-1, E-cadherin, phosphorylated FAK, and AMH. Aged oocytes exhibited reduced polar body extrusion, increased DCFDA fluorescence, and decreased CMXRos intensity. Ex vivo MOTS-c treatment increased post-culture first polar body positivity and mitochondrial activity-associated fluorescence while reducing oxidative stress-associated fluorescence, although outcomes remained below young-reference levels. Reanalysis of human folliculogenesis and non-human primate ovarian aging datasets further revealed a young-enriched trend in the MOTS-c-associated metabolic program. These findings suggest that MOTS-c improves metabolic, mitochondrial, and structural features of aging-associated granulosa cells and enhances maturation-associated outcomes in aged oocytes, supporting further investigation of MOTS-c-based strategies for reproductive aging. Lay summary As women age, the quality of their eggs declines, which is a major contributor to age-related infertility. Eggs are supported by neighboring "helper" cells called granulosa cells, which act like a support crew that supplies the egg with the energy it needs to mature. These helper cells contain mitochondria, tiny energy-generating structures often called the cell's "powerhouses," and with age these powerhouses become less efficient, leaving eggs with less support than they need. This study tested whether MOTS-c, a small molecule naturally produced by mitochondria, could restore the function of these aging helper cells. In laboratory experiments using aged human granulosa cells and eggs from aged mice, MOTS-c boosted energy production and helped maintain cell structure, and was associated with improved egg maturation compared with untreated aged eggs, although treated eggs did not fully reach the maturation levels seen in young mice. These findings are from laboratory and animal experiments, not from women, and this study did not test whether MOTS-c affects fertilization, embryo development, or live birth. Further research is needed before any conclusions can be drawn about its relevance to human fertility.