Objective We aimed to investigate the effect of roofplate-specific spondin 2 (RSPO2) on oxidative stress and apoptosis in ovarian granulosa cells, using human ovarian granulosa tumor cells as a cellular model. Methods We transiently transfected COV434 human granulosa cells with RSPO2 overexpression plasmid and small interfering RNA (siRNA). The efficiencies of the overexpression and interference were detected by quantitative real-time polymerase chain reaction (qPCR) and Western blot. Intracellular reactive oxygen species (ROS) were detected using a ROS assay kit, and apoptosis was detected by flow cytometry. Expression levels of oxidative stress- and apoptosis-related genes were determined by qPCR and Western blot, and interacting proteins were detected by co-immunoprecipitation. Results RSPO2 overexpression and interference were successfully realized. Overexpression of RSPO2 significantly inhibited ROS levels in COV434 cells (P<0.05) and the early apoptosis of granulosa cells (P<0.01), while RSPO2 interference had the opposite result. Overexpression of RSPO2 significantly upregulated the mRNA and protein levels of superoxide dismutase (SOD) 1, SOD2, and catalase (CAT) (P<0.05), and significantly downregulated the levels of Caspase 3 and Caspase 8 (P<0.05). In contrast, RSPO2 interference significantly downregulated SOD1 and CAT (P<0.05) and significantly upregulated Caspase 3 and Caspase 8 (P<0.05). We found protein interactions between RSPO2, SOD1, CAT, and Caspase 3. Conclusions Interfering with RSPO2 promotes the accumulation of ROS within cells and downregulates the expression of SOD1 and CAT, thereby enhancing oxidative stress. Additionally, interfering with RSPO2 upregulates the expression of Caspase 3 and Caspase 8, promoting cell apoptosis. These findings suggest that RSPO2 plays a crucial role in ovarian granulosa cells.