The novel secretome ST266 activates Akt and protects against oxidative stress-mediated injury in human RPE and Muller cells

Oxidative stress-mediated retinal pigment epithelial (RPE) cell damage is associated with age-related macular degeneration (AMD). ST266 is the biological secretome produced by a novel population of amnion-derived multipotent progenitor cells. Herein, we investigated the effect of ST266 on RPE cell injury induced by hydroquinone (HQ), a cigarette smoke related oxidant, hydrogen peroxide (H2O2) and all-trans retinal (atRal), a pro-oxidant component of the retinoid cycle. We additionally investigated its effect on M & uuml;ller cell injury induced by H2O2. Cultured human RPE cells were pre-treated for 1 h in the presence or absence of MK-2206, a protein kinase B (Akt) inhibitor, then treated with varying concentrations of HQ, H2O2, or atRal for 1.5 h. Cultured human M & uuml;ller cells (MIO-M1) were pre-treated for 1 h in the presence or absence of MK-2206, then treated with varying concentrations of H2O2 for 1.5 h. Media were then replaced with STM100 (control media into which the ST266 secretome proteins were collected) or ST266 at various times. Cell viability was determined with WST-1 reagent. Mitochondrial membrane potential (Delta psi m) was quantified by a fluorescence plate reader. The protein phosphorylation levels of Akt, glycogen synthase kinase 3 beta (GSK-3 beta), and p70 ribosomal S6 kinase (p70S6K) were measured by Western blot. ST266 significantly improved RPE and MIO-M1 cell viability that was reduced by oxidant exposure and improved oxidant-disrupted Delta psi m. In both cell types, ST266 induced phosphorylation of Akt, GSK-3 beta, and p70S6K. MK-2206 significantly eliminated ST266-mediated protein phosphorylation of Akt, GSK-3 beta, and p70S6K and abolished the ST266-protective effect on cell viability. In conclusion, ST266 activates Akt, protects against oxidative stress-mediated cell injury in an Akt-dependent manner, and improves Delta psi m, suggesting a potential role for ST266 therapy in treating retinal diseases such as AMD.