The protective effect of antioxidants on orbital fibroblasts from patients with Graves’ ophthalmopathy in response to oxidative stress

Purpose: To investigate the biphasic effects of hydrogen peroxide (H(2)O(2)) on the orbital fibroblasts of patients with Graves’ ophthalmopathy (GO) and the relation to antioxidants and proinflammatory cytokines. METHODS: Proliferation of cultured orbital fibroblasts from patients with GO and normal controls was evaluated in response to various concentrations of H(2)O(2). The effect of low concentrations of H(2)O(2) (6.25 μM) on the cellular proliferation and induction of intracellular proinflammatory cytokines, and reactive oxygen species of orbital fibroblasts were assessed. Protective effects of N-acetylcysteine and vitamin C on GO fibroblasts in response to 6.25 μM H(2)O(2) stimulation were also investigated. RESULTS: When the GO fibroblasts were exposed to H(2)O(2) at a concentration of 50 μM or above, significant cytotoxicity was observed. In contrast, lower concentrations of H(2)O(2) (3.125–25 μM) increased the survival of GO fibroblasts with the peak cellular proliferation at 6.25 μM H(2)O(2). However, this biphasic effect of H(2)O(2) on the viability of orbital fibroblasts was not found in normal controls. In addition, 6.25 μM H(2)O(2) led to significant elevation of the levels of transforming growth factor, beta 1, interleukin-1β, and superoxide anion in GO fibroblasts, but no significant change in the normal controls. Pretreatment with N-acetylcysteine or vitamin C reversed the enhanced proliferation capacity and the induction of transforming growth factor, beta 1, interleukin-1β and superoxide anion of GO fibroblasts in response to 6.25 μM H(2)O(2). CONCLUSIONS: These findings revealed the biphasic effect of H(2)O(2) on cellular proliferation of GO orbital fibroblasts. Importantly, a low level of H(2)O(2) can stimulate proliferation of GO orbital fibroblasts and induce the production of proinflammatory cytokines, which can be inhibited by pretreatment with antioxidants. This provides a theoretical basis for the rational use of antioxidant in treating GO at an early stage.