The Role of ERK1/2 Activation in Sarpogrelate-Mediated Neuroprotection

Abstract

PurposeTo characterize the mediators of 5-HT2A serotonin receptor–driven retinal neuroprotection.MethodsAlbino mice were treated intraperitoneally with saline or sarpogrelate, a 5-HT2A antagonist, immediately before light exposure (LE). Following LE, retinas were harvested for a high-throughput phosphorylation microarray to quantify activated phosphorylated proteins in G protein–coupled receptor (GPCR) signaling. To confirm microarray results and define temporal changes, Western blots of select GPCR signaling proteins were performed. Since both methodologies implicated MAPK/ERK activation, the functional significance of sarpogrelate-mediated ERK1/2 activation was examined by inhibition of ERK1/2 phosphorylation via pretreatment with the MEK inhibitor (MEKi) PD0325901. The degree of neuroprotection was evaluated with spectral-domain optical coherence tomography (SD-OCT) and electroretinography (ERG). To determine the effects of sarpogrelate on gene expression, a qPCR array measuring the expression of 84 genes involved in oxidative stress and cell death was performed 48 hours post LE.ResultsSarpogrelate led to an activation of the MAPK/ERK pathway. Temporal analysis further demonstrated a transient activation of ERK1/2, starting with an early inhibition 20 minutes into LE, a maximum activation at 3 hours post LE, and a return to baseline at 7 hours post LE. Inhibition of ERK1/2 with MEKi pretreatment led to attenuation of sarpogrelate-mediated neuroprotection. LE caused significant changes in the expression of genes involved in iron metabolism, oxidative stress, and apoptosis. These changes were prevented by sarpogrelate treatment.ConclusionsSarpogrelate-mediated retinal protection involves a transient activation of the MAPK/ERK pathway, although this pathway alone does not account for the full effect of neuroprotection.

Monomethyl Fumarate Protects the Retina From Light-Induced Retinopathy

“…Western blot was performed as described previously 39. Proteins in the NF-kB and NRF2 pathways were measured in the neural retina, at 1, 3, 6, 12, and 24 hours after LIR.…”

Interpretation of OCT and OCTA images from a histological approach: Clinical and experimental implications

“…As an example, in a light-induced photoreceptor degeneration model, 15 mg/kg of sarpogrelate treatment (a selective 5-HT2A receptor antagonist) was enough to obtain significant improvement of retinal function in ERG recordings that cannot be observed by OCT. Nevertheless, after a higher dose (40 mg/kg) OCT imaging was able to evidence an increase in ONL thickness (Ku et al, 2018;Tullis et al, 2015). On the other hand, OCT was useful to show the attenuation of neuroprotection when same treatment is combined with ERK1/2 inhibitor, while ERG recordings were not able to show these differences (Ku et al, 2018).…”

“…Nevertheless, after a higher dose (40 mg/kg) OCT imaging was able to evidence an increase in ONL thickness (Ku et al, 2018;Tullis et al, 2015). On the other hand, OCT was useful to show the attenuation of neuroprotection when same treatment is combined with ERK1/2 inhibitor, while ERG recordings were not able to show these differences (Ku et al, 2018).…”

A Ketogenic & Low-Protein Diet Slows Retinal Degeneration in rd10 Mice

“… 38 , 39 Our work, as well as others, has shown that increases in corticosterone and activation of the MAPK pathway, as well as CREB, can elicit neuroprotective pathways and protect from light-induced retinopathy. 32 , 40 – 42 In addition to activating a stress response, early weaning can delay the triggering of satiety and modulate the circadian rhythm of food intake, which was demonstrated in male Wistar rat pups out to PD100. 43 It is feasible that changes in behavioral satiety and food consumption can impact the retina, because recent studies have linked the gut microbiota with retinal disease.…”

“…Retinal rhodopsin and β-actin protein content was measured via Western blot at PD40. 32 Total protein was extracted, and 100 µg of protein was loaded into a 4% to 20% tris-glycine gel. After transfer, membranes were blocked for one hour, incubated overnight in primary antibodies (anti-rhodopsin Rho 4D2, ab98887 and anti-actin, ab3280; Abcam, Cambridge, UK), and incubated in a secondary antibody (1:15,000, goat anti-mouse IRDye 800CW, 925-32210; LICOR, Lincoln, NE, USA).…”