The role of adenosine monophosphate-activated protein kinase (AMPK) in the action of modified recombinant extracellular wt (wild-type) ASYN in vitro

• Main Authors: Jeremić Marija, Radeta Ratko, Jovanović Maja
• Format: Article
• Language: English
• Published: University of Belgrade, Medical Faculty 2016-01-01
• Series: Medicinski Podmladak
• Subjects: modified recombinant wt ASYN; AMPK; autophagy
• Online Access: http://scindeks-clanci.ceon.rs/data/pdf/0369-1527/2016/0369-15271601051J.pdf

Introduction: Alpha-synuclein (ASYN) accumulation is considered to be one of the key factors in Parkinson's disease (PD) pathogenesis. Posttranslational modifications, including oxidation and nitration, can increase aggregation of ASYN. Alterations of adenosine monophosphate-activated protein kinase (AMPK) signaling have been shown in several neurodegeneration models, including PD. Aim: The aim of this study was to investigate the role of AMPK in the action of modified recombinant wt (wild-type) ASYN on differentiated SH-SY5Y cells. Materials and methods: All experiments were conducted in all-trans retinoic acid-differentiated human neuroblastoma SH-SY5Y cells, treated with 3 different types of modified recombinant wt ASYN (oligomeric, nitrated and dopamine-modified ASYN). Activation of AMPK and Raptor, together with conversion of LC3-I to LC3-II, were monitored using immunoblotting. Cell viability was assessed using crystal violet assay. Results: Treatment with modified recombinant wt ASYN (oligomeric, nitrated and dopamine-modified ASYN) led to substantial decrease in number of viable differentiated SH-SY5Y cells. Western blot analysis showed decreased levels of AMPK and its downstream kinase Raptor, together with an increase in the conversion of LC3-I to LC3-II, upon treatment with recombinant ASYN. Pharmacological activator of AMPK (AICAR) significantly increased the number of viable cell in the presence of all 3 types of modified recombinant wt ASYN. Conclusion: Modified forms of recombinant wt (wildtype) ASYN (oligomeric, nitrated and dopamine-modified ASYN) induced decrease in number of viable differentiated SH-SY5Y cells, accompanied by inactivation of AMPK/Raptor signalling pathway and autophagy induction. Pharmacological activation of AMPK improved cell survival, indicating a protective role of AMPK in neurotoxicity of extracellular ASYN.