Hydrogen Sulfide-Preconditioned Mesenchymal Stem Cells Attenuate Ferroptosis in Corneal Alkali Burn via the AMPK/Nrf2/HO-1 Pathway in an IL-10-Dependent Manner.

PURPOSE

We aimed to investigate whether hydrogen sulfide (H2S)-preconditioned mesenchymal stem cells (MSCs) could enhance the anti-ferroptotic effects in a murine model of corneal alkali burn.

METHODS

MSCs were treated with various concentrations of GYY4137 (an H2S donor) and Western blot was used to detect the expression of IL-10 and cleaved-caspase-3. Slit-lamp photography and hematoxylin and eosin (H&E) staining were used to observe corneal structure after receiving a single subconjunctival injection of H2S-MSCs. Quantitative PCR (QPCR) or Western blot were used to detect the expression of pro-inflammatory cytokines, ferroptosis-regulatory molecules (SLC7A11, GPX4, 4-HNE, and FTH1) and AMPK/Nrf2/HO-1 pathway on day 7 after corneal injury. A co-culture model of human corneal epithelial cells (HCECs) and MSCs was used to investigate the anti-ferroptotic effects of MSCs.

RESULTS

Under the stimulation of 1 µmol/L GYY4137 for 48 hours, MSCs showed enhanced expression of IL-10. Furthermore, H2S pretreatment suppressed hydrogen peroxide (H2O2)- or serum deprivation-induced MSC apoptosis in vitro. On days 7 and 14 post-treatment, corneal opacity was ameliorated and corneal structure was more regular in the H2S-MSC group. H2S-MSC effectively attenuated ferroptosis in the cornea and HCECs by upregulating the expression of SLC7A11 and GPX4, while suppressing the level of 4-HNE and FTH1. Meanwhile, H2S-MSC activated AMPK/Nrf2/HO-1 pathway in vitro and in vivo. Knockdown of IL-10 via siRNA attenuated the inhibitory effects of H2S-MSC on erastin-induced ferroptosis in HCECs.

CONCLUSIONS

H2S preconditioning augmented the anti-inflammatory and anti-ferroptotic effects of MSCs via activation of the AMPK/Nrf2/HO-1 pathway in an IL-10-dependent way, offering a novel strategy for corneal alkali burn treatment.