Inhibition of sortilin reduces neuronal and vascular damage after ischemia/reperfusion through reduced inflammatory and autophagy actions in retinal Müller cells.

High-glucose culturing conditions significantly increased inflammatory markers TNFα and NLRP3 in both primary human Müller cells and rat Müller cells (rMC-1).

• Cells were grown in normal (5 mM) or high (25 mM) glucose conditions.
• Both tumor necrosis factor α (TNFα) and NOD-like receptor protein 3 (NLRP3) protein levels were measured via western blotting.
• The increase in inflammatory markers was observed in both the primary human Müller cell line and the rat Müller cell line (rMC-1).

High-glucose culturing conditions significantly increased autophagic marker LAMP2 in both primary human Müller cells and rat Müller cells.

• LAMP2 (lysosome-associated membrane glycoprotein 2) was used as a marker of autophagy.
• Protein levels were assessed by western blotting in cells grown in 5 mM versus 25 mM glucose.
• The increase in LAMP2 was observed in both primary human Müller cells and rMC-1 cells under high-glucose conditions.

High-glucose culturing conditions significantly increased apoptotic marker cleaved caspase 3 in both primary human Müller cells and rat Müller cells.

• Cleaved caspase 3 was used as a marker of apoptosis and measured by western blotting.
• Elevated cleaved caspase 3 was found in both primary human Müller cells and rMC-1 cells grown in 25 mM glucose compared to 5 mM glucose.
• This indicates high glucose promotes apoptotic signaling in retinal Müller cells.

Treatment with AF38469, a small-molecule inhibitor of sortilin, reduced elevated inflammatory, autophagic, and apoptotic markers in Müller cells grown in high glucose.

• AF38469 was applied to both primary human Müller cells and rMC-1 cells cultured in high glucose (25 mM).
• All measured markers — TNFα, NLRP3, LAMP2, and cleaved caspase 3 — were reduced by AF38469 treatment.
• Reductions were observed in both the primary human and rat Müller cell lines.

AF38469 eye drops significantly reduced ischemia/reperfusion-induced neuronal damage in mice at 2 days post-I/R.

• The study used a mouse model of retinal ischemia/reperfusion (I/R) injury.
• Neuronal damage was assessed at 2 days post-I/R.
• AF38469 was delivered as eye drops, representing a non-invasive topical route of administration.

AF38469 eye drops significantly reduced ischemia/reperfusion-induced vascular damage in mice at 10 days post-I/R.

• Vascular damage was assessed at 10 days post-I/R in the mouse I/R model.
• AF38469 was administered as eye drops.
• Both neuronal and vascular endpoints were significantly reduced by sortilin inhibition.

Sortilin regulates inflammatory, autophagic, and apoptotic pathways in retinal Müller cells under high-glucose conditions.

• This conclusion is supported by the significant reduction of TNFα, NLRP3, LAMP2, and cleaved caspase 3 upon sortilin inhibition with AF38469.
• The findings were consistent across both primary human Müller cells and the rat rMC-1 Müller cell line.
• The authors state that 'sortilin regulates the inflammatory, autophagic, and apoptotic pathways in Müller cells grown in high glucose.'