Abstract: OBJECTIVE: To culture human granulosa cells in vitro,detect the cytotoxic effect of CuInS₂/ZnS quantum dots and analyze its molecular mechanism. METHODS: Hyaluronidase was used to remove granulosa cells outside of human superovulation,and primary cultures were carried out. Cells were treated with medium (10 μg/mL) and high (100 μg/mL) CuInS₂/ZnS quantum dot doses. Changes of cell viability after treatment were detected by the CCK-8 method and the median lethal dose (IC₅₀) was calculated. Apoptosis was detected by flow cytometry with the Annexin V/PI double staining method;and release and oxidation of sex hormones were detected by kits stress changes. RESULTS: Human granulosa cells were successfully cultured in vitro. The IC₅₀of CuInS₂/ZnS quantum dots was 66.72 μg/mL. Compared with the PBS control group,cell viability was significantly inhibited and apoptosis was induced in the high- dose group (P<0.01). In the two treatment groups,release of progesterone (P) and estradiol (E2) were obviously increased (P<0.05 or P<0.01),levels of reactive oxygen species (ROS) in granulosa cells were significantly elevated (P<0.05 or P<0.01),and malondialdehyde (MDA),total antioxidative capacity (T-AOC) and total superoxide dismutase (T-SOD) activity were significantly decreased (P<0.01) compare with the control group. In addition, intracellular glutathione(GSH) activity was remarkedly increased in the high-dose treated group (P<0.01). CONCLUSION: Human granulosa cell lines were successfully cultured in vitro,and CuInS₂/ZnS quantum dots above 10 μg/mL induced oxidative damage by invading granulosa cells and promoted apoptosis. Our results provide valuable information for the biosafety assessment of CuInS₂/ZnS quantum dots in clinical applications.

Key words: human ovarian granulosa cells, oxidative damage, cytotoxic effects, CuInS₂/ZnS, quantum dots