AMPK/MFF Activation: Role in Mitochondrial Fission and Mitophagy in Dry Eye

PURPOSE: To assess the role of mitochondrial morphology and adenosine monophosphate–activated protein kinase (AMPK)/mitochondrial fission factor (MFF) in dry eye and the underlying mechanisms. METHODS: Immortalized human corneal epithelial cells (HCECs) and primary HCECs were cultured under high osmotic pressure (HOP). C57BL/6 female mice were injected subcutaneously with scopolamine. Quantitative real-time PCR was used to measure mRNA expression. Protein expression was assessed by western blot and immunofluorescence staining. Mitochondrial morphology was observed by confocal microscopy and transmission electron microscopy. RESULTS: First, HOP induced mitochondrial oxidative damage to HCECs, accompanied by mitochondrial fission and increased mitophagy. Then, AMPK/MFF pathway proteins were increased consequent to HOP-induced energy metabolism dysfunction. Interestingly, the AMPK pathway promoted mitochondrial fission and mitophagy by increasing the recruitment of dynamin-related protein 1 (DRP1) to the mitochondrial outer membrane in the HOP group. Moreover, AMPK knockdown attenuated mitochondrial fission and mitophagy due to HOP in HCECs. AMPK activation triggered mitochondrial fission and mitophagy. Mitochondrial fission of HCECs stressed by HOP was mediated via MFF phosphorylation. MFF knockdown reversed mitochondrial fragmentation and mitophagy in HCECs treated with HOP. Inhibition of MFF protected HCECs against oxidative damage, cell death, and inflammation in the presence of HOP. Finally, we detected mitochondrial fission and AMPK pathway activation in vivo. CONCLUSIONS: The AMPK/MFF pathway mediates the development of dry eye by positively regulating mitochondrial fission and mitophagy. Inhibition of mitochondrial fission can alleviate oxidative damage and inflammation in dry eye and may provide experimental evidence for treating dry eye.