Drp1-dependent mitochondrial fission mediates corneal injury induced by alkali burn.

Corneal alkali burn, one of the most serious ophthalmic emergencies, is difficult to be cured by conservative treatments. It is well known that oxidative stress, inflammation and neovascularization are the main causes of corneal damage after alkali burn, but its underlying mechanism remains to be elucidated. Here, we reported that the expression and phosphorylation (Ser616) of mitochondrial fission protein Drp1 were up-regulated at day 3 after alkali burn, while mitochondrial fusion protein Mfn2 was down-regulated. The phosphorylation of ERK1/2 in corneas was increased at day 1, 3, 7 and peaked at day 3 after alkali burn. In human corneal epithelial cells (HCE-2), NaOH treatment induced mitochondrial fission, intracellular ROS production and mitochondrial membrane potential disruption, which was prevented by Drp1 inhibitor Mdivi-1. In corneas, Mdivi-1 or knockdown of Drp1 by Lenti-Drp1 shRNA attenuated alkali burn-induced ROS production and phosphorylation of IκBα and p65. In immunofluorescence staining, it was detected that Mdivi-1 also prevented NaOH-induced nuclear translocation of p65 in HCE-2 cells. Moreover, the expression of NADPH oxidase NOX2 and NOX4 in corneas peaked at day 7 after alkali burn. Mdivi-1, Lenti-Drp1 shRNA or the mitochondria-targeted antioxidant mito-TEMPO efficiently alleviated activation of NF-κB, expression of NOX2/4 and inflammatory cytokines including IL-6, IL-1β and TNF-α in corneas after alkali burn. In pharmacological experiments, both Mdivi-1 and NADPH oxidases inhibitor Apocynin protected the corneas against alkali burn-induced neovascularization. Intriguingly, the combined administration of Mdivi-1 and Apocynin had a synergistic inhibitory effect on corneal neovascularization after alkali burn. Taken together, these results indicate that Drp1-dependent mitochondrial fission is involved in alkali burn-induced corneal injury through regulating oxidative stress, inflammatory responses and corneal neovascularization. This might provide a novel therapeutic target for corneal injury after alkali burn in the future.