Gelatin Methacryloyl-Tannic Acid Hydrogel with Sustained Antioxidant Activity for Protecting Ovarian Granulosa Cells from Oxidative Stress

Background: Oxidative stress is a major contributor to granulosa cell dysfunction and follicular atresia, with excessive reactive oxygen species (ROS) impairing mitochondrial activity and cell survival. Hydrogels based on gelatin methacryloyl (GelMA) are biocompatible and easily photocrosslinked, but they lack intrinsic antioxidant function. Methods: A GelMA–tannic acid (TA) hydrogel was synthesized by visible light curing in the presence of TA. Structural features were confirmed by FT-IR and ¹H NMR. Photorheology was used to determine gelation kinetics, and scanning electron microscopy assessed morphology. Antioxidant performance was evaluated by DPPH, ABTS, FRAP, and H₂O₂ assays. The protective effect on human granulosa-like KGN cells under H₂O₂ stress was examined by CCK-8 viability, DCFH-DA ROS detection, JC-1 mitochondrial membrane potential, and Live/Dead staining. Results: GelMA–TA exhibited a rapid gelation time of 8.5 s and a plateau modulus of 1.8 kPa, higher than the 1.5 kPa modulus of GelMA alone. In antioxidant assays, GelMA–TA showed significant radical scavenging activity with DPPH (86.1%), ABTS (91.4%), and FRAP (1.15 mmol Fe²⁺ equivalent), as well as nearly complete H₂O₂ removal (94.8%). In KGN cells, GelMA–TA reduced intracellular ROS by 60%, restored mitochondrial membrane potential (Δψm) to 0.9 (compared to 0.5 for H₂O₂ treatment), and improved cell viability by 30%. Conclusion: These findings demonstrate that GelMA–TA forms a fast-curing, antioxidant hydrogel capable of maintaining a low-ROS microenvironment and protecting granulosa cells, offering a promising platform for ovarian tissue engineering and related regenerative applications.