Artesunate protects against ocular fibrosis by suppressing fibroblast activation and inducing mitochondria‐dependent ferroptosis

Artesunate, a derivative from extracts of Artemisia annua, has recently been reported to alleviate fibrosis recently. Here, in this study, we sought to determine the anti‐fibrosis effect of artesunate in rabbit glaucoma filtration surgery (GFS) model and illuminate underlying mechanisms. Our results showed that artesunate subconjunctival injection alleviated bleb fibrosis by inhibiting fibroblast activation and inducing ferroptosis. Further mechanistic investigation in primary human ocular fibroblasts (OFs) showed that artesunate abrogated fibroblast activation by inhibiting TGF‐β1/SMAD2/3 and PI3K/Akt pathways and scavenged OFs by inducing mitochondria‐dependent ferroptosis. Mitochondrial dysfunction, mitochondrial fission, and iron‐dependent mitochondrial lipid peroxidation were observed in artesunate‐treated OFs. Besides, mitochondria‐localized antioxidants inhibited artesunate‐induced cell death, suggesting a critical role of mitochondria in artesunate‐induced ferroptosis. Our study also found that expression of mitochondrial GPX4 but no other forms of GPX4 was decreased after artesunate treatment and that mitochondrial GPX4 overexpression rescued artesunate‐induced lipid peroxidation and ferroptosis. Other cellular ferroptosis defense mechanisms, including cellular FSP1 and Nrf2, were also inhibited by artesunate. In conclusion, our study demonstrated that artesunate protects against fibrosis through abrogation of fibroblast activation and induction of mitochondria‐dependent ferroptosis in OFs, which may offer a potential treatment for ocular fibrosis. Low concentrations of artesunate could reduce fibroblast activation by suppressing TGF‐β1/SMAD2/3 and PI3K/Akt pathways. High concentrations of artesunate induce mitochondrial lipid peroxidation, ferrous iron accumulation, and mitochondrial fission; downregulate mitochondrial membrane potential; inhibit the expression of mitochondrial GPX4 and FSP1; and suppress the activation of the Nrf2/ARE pathway.