Improvement of P23H rod opsin traffic after metformin treatment accelerates the rate of retinal degeneration in P23H retinitis pigmentosa

Abstract

Purpose: Metformin, the most commonly prescribed anti-diabetic drug, can alter the rate of protein production in the cells through the activation of AMP-activated protein kinase (AMPK). Activation of AMPK could potentially alter the folding and/or degradation of misfolded proteins, such as P23H rhodopsin that causes retinitis pigmentosa (RP). Therefore, we investigated the effect of metformin on cell culture and transgenic models expressing P23H rhodopsin.

Methods: SK-N-SH neuroblastoma cells were treated with metformin and the effect of treatment on P23H-GFP on localisation was assessed by immunofluorescence and confocal microscopy. Cell survival was assessed by lactate dehydrogenase (LDH) assay. The folding of P23H rod opsin in the presence of Metformin was assessed by absorption spectroscopy of purified rhodopsin to monitor pigment formation. P23H-1 rats were treated with either 300 mg/Kg metformin or vehicle administered daily via intraperitoneal injection and were subjected to electroretinogram (ERG) and spectral domain optical coherent tomography (SD-OCT) analysis, after which rats were sacrificed and tissue was taken for biochemical studies, histology and electron microscopy.

Results: Metformin treatment in SK-N-SH cells improved the traffic of P23H rod opsin from the ER to the plasma membrane, suggesting an improvement in mutant rod opsin folding, which was confirmed by an enhanced yield of purified P23H rhodopsin with a lambda max at 490 nm. Furthermore, similar to pharmacological chaperones metformin treatment reduced P23H-related cell death. Metformin was tested in the P23H-1 rat model in three independent drug treatments. Systemic metformin treatment increased APMK activation in the retina. Surprisingly, we observed that metformin treatment led to decreased photoreceptor function and survival. Metformin treated rats showed significantly lower ERG responses and thinner retinas compared to the vehicle-treated rats.

Conclusions: The improvement in P23H folding and traffic after metformin treatment correlated with an accelerated rate of retinal degeneration. This suggests that therapies aimed at rhodopsin RP might be better focused on enhancing degradation than improving folding. In addition, these data could be important for some RP patients, as metformin could potentially worsen their RP if prescribed for type II diabetes.