Rubicon, a Key Molecule for Oxidative Stress-Mediated DNA Damage, in Ovarian Granulosa Cells

Aging drives excessive ovarian oxidative stress (OS), impairing fertility and affecting granulosa cells (GCs), which are involved in folliculogenesis. This study aims to clarify the relationship between OS and autophagy in GCs and to identify compounds that enhance OS resistance. We identified Rubicon, an autophagy suppressor, as a key mediator of DNA damage in GCs under OS. Hydrogen peroxide (H₂O₂) compromised cell viability via DNA damage in the human GC cell line, HGrC1, without affecting autophagic activity. However, autophagy activation increased OS resistance in HGrC1 cells, and vice versa. Among clinically safe materials, trehalose, a disaccharide, protected cells as an autophagy activator against H₂O₂-induced cytotoxicity. Trehalose significantly increased autophagic activity, accompanied by reduced Rubicon expression, compared to other carbohydrates. It also reduced the expression of DNA damage-responsive proteins and the production of reactive oxygen species. Rubicon knockdown mitigated OS-induced DNA damage, while Rubicon overexpression enhanced DNA damage and decreased HGrC1 cell viability. Trehalose enhanced OS resistance by activating autophagy and suppressing Rubicon in a bidirectional manner. As Rubicon expression increases in aged human ovaries, trehalose may improve ovarian function in patients with infertility and other OS-related diseases.