Abstract
OBJECTIVES: To explore the antioxidative effects of gypenosides (Gyps) in orbital fibroblasts (OFs) derived from patients with thyroid-associated ophthalmopathy (TAO) and to validate their therapeutic efficacy in vivo using an animal model. METHODS: Bioinformatics analyses were performed to screen potential genes and signalling pathways underlying the effect of Gyps in OFs. OFs were isolated from orbital connective tissues of both TAO patients and non-TAO controls. CCK-8 assay was used to detect cell proliferation. Oxidative stress was evaluated by measuring reactive oxygen species (ROS) and superoxide dismutase (SOD) levels. Quantitative reverse transcriptive-polymerase chain reaction (RT-qPCR), ELISA, and western blotting were employed to ascertain the effects of Gyps on H(2)O(2)-induced oxidative stress, inflammation, fibrosis, and autophagy. Furthermore, a TAO mouse model was established. Gyps were administered by intraperitoneal injection (50 mg/kg/day for 4 weeks) to evaluate their protective effects against oxidative stress, inflammation, and fibrosis in orbital tissues. RESULTS: Bioinformatic analysis revealed that the identified genes were primarily enriched in metabolic and oxidative stress-related pathways. In vitro experiments demonstrated that Gyps significantly reduced H(2)O(2)-induced ROS generation, increased SOD levels, and suppressed the expression of inflammation-, fibrosis-, and autophagy-related markers. These effects were associated with the activation of the nuclear factor erythroid 2-related factor 2 (Nrf2)/extracellular-regulated kinase (ERK)/heme oxygenase 1 (HO-1) pathway. In vivo animal experiments further confirmed that Gyps treatment effectively alleviated oxidative injury, inflammatory cell infiltration, and collagen deposition in the orbital tissues of TAO model mice. CONCLUSIONS: Gyps exert significant antioxidative, anti-inflammatory, and anti-fibrotic effects by activating the Nrf2/ERK/HO-1 signalling pathway in both in vitro and in vivo TAO models.