Metformin Activates AMP-Activated Protein Kinase-Transcription Factor EB Signaling to Restore Lysosomal and Mitochondrial Homeostasis and Suppress Epithelial-to-Mesenchymal Transition in Oxidative Stress-Injured Retinal Pigment Epithelium.

Disruption of lysosomal homeostasis and accumulation of dysfunctional mitochondria contribute to degenerative pathologies, including age-related macular degeneration. Here, how inhibition of autophagic lysosome reformation (ALR) alters lysosomal dynamics, mitophagy, and downstream stress signaling in retinal pigment epithelial (RPE) cells was investigated, and whether these changes are pharmacologically reversible was determined. In ARPE-19 cells, ALR inhibition by nocodazole or siRNA-mediated depletion of kinesin-1 (UKHC) and dynamin-2 (DNM2) induced enlarged lysosomes with reduced degradative capacity, impaired mitophagic turnover, and accumulation of dysfunctional mitochondria. ALR blockade increased reactive oxygen species and cytosolic Ca2+, promoted activation and mitochondrial translocation of protein kinase C, and triggered phosphorylation of glycogen synthase kinase-3β with subsequent stabilization of Snail family transcriptional repressor 1 (SNAIL/SNAI1), consistent with epithelial-to-mesenchymal transition. Metformin restored lysosomal homeostasis by activating AMP-activated protein kinase and enhancing transcription factor EB-dependent lysosome biogenesis, thereby improving autophagic flux, limiting reactive oxygen species/Ca2+ accumulation, suppressing protein kinase C activation, and attenuating epithelial-to-mesenchymal transition-associated marker changes. In a sodium iodate-induced oxidative injury model, metformin preserved RPE microtubule architecture and reduced lysosomal and mitochondrial abnormalities. Although these findings rely on a prolonged monolayer culture system and an acute injury model, they support a protective role for AMP-activated protein kinase-transcription factor EB-driven lysosome restoration in RPE stress resilience and suggest lysosome-directed repurposing potential for metformin in degenerative retinal disease.