Autophagy as a pathomechanism in Parkinson’s disease: Neuroprotective effect of autophagy enhancer in in vitro model of Parkinson's disease

Abstract

Background and objectives: Recently, autophagy pathway is regarded as important mechanism in pathogenesis of Parkinson's disease (PD). Modulation of autophagy may be a novel strategy for the treatment of PD and autophagy could play a pivotal role in PD and modulation of autophagy may be a novel strategy for the treatment of PD. Among various autophagy enhancer, we selected 3 substrates including calcitriol, erythropoietin, and rosuvastatin, which has not yet been investigated in association with PD model. In the current study, we investigated the concentration-dependent effects of calcitriol, erythropoietin, and rosuvastatin on autophagy markers in SH-SY2Y cells and whether these substrates protect SH-SY2Y cells from rotenone-induced neurotoxicity together with the underlying mechanisms of protection, including autophagy. Material and methods: An in vitro model of Parkinson's disease, the rotenone-induced neurotoxicity model in SH-SY5Y cells was adapted. We measured cell viability using an MTT assay, Annexin V/propidium iodide assay, and intracellular reactive oxygen species levels and analyzed autophagy-associated intracellular signaling proteins by Western blotting. To detect the expression of LC3 and α-synuclein, immnunoflorescence analysis was performed in experiment with erythropoietin and rosuvatatin. Result: In all three substrates, mono-treatment increased levels of mTOR independent/upstream autophagy markers, including beclin-1, AMPK, and ULK-1. Rotenone treatment of SH-SY5Y cells reduced their viability, increased reactive oxygen species levels, induced apoptosis and α-synuclein expression; simultaneous exposure to these substrates significantly restored these effects. Rotenone enhanced mTOR expression and suppressed beclin-1 expression, indicating suppression of autophagic system. In rosuvastatin treatment, chaperone mediated autophagy marker, LAMP-2 also was decreased with rotenone treatment, but restored with combined treatment of rosuvastatin. Conclusions: All substrates protects against rotenone-induced neurotoxicity in SH-SY5Y cells by enhancing multiple steps of autophagy signaling pathways. These neuroprotective effects of these substrates against rotenone-induced dopaminergic neurotoxicity provide an experimental basis and significant impact on future PD treatment strategy. Keyword: Calcitriol; Eyrthropoietin; Statin; Parkinson's disease; Autophagy; Neuroprotection