Effects of miR-199a on autophagy by targeting glycogen synthase kinase 3β to activate PTEN/AKT/mTOR signaling in an MPP+ in vitro model of Parkinson’s disease

ABSTRACT

Objectives: miR-199a can regulate autophagy, its underlying mechanisms remain unknown. The purpose of this study was to investigate the mechanisms of miR-199a involved in regulating autophagy in a 1-methyl-4-phenylpyridine (MPP⁺)-induced in vitro model of PD.

Methods: PC12 cells were incubated in MPP⁺, and the expression levels of miR-199a were bidirectionally regulated via either transfection of an miR-199a mimic or incubation in miR-199a inhibitors. The experimental manipulations were divided into four groups, including the control group, MPP⁺ group, MPP⁺ + miR-199a mimic group, and MPP⁺ + miR-199a inhibitor group. MTT, CCK-8, qRT-PCR, Western blotting and linear correlation analysis were performed to evaluate various experimental indicators.

Results: At increasing MPP⁺ concentrations, the following results were found: the expression levels of miR-199a, phosphorylated AKT and mTOR proteins expression decreased; the expression levels of phosphatase and tensin homologue (PTEN), GSK3β, Beclin1, and LC3II increased; PC12 autophagy increased; and cellular viability and survival rates decreased. Transfection of an miR-199a mimic increased miR-199a expression and induced all of the following: the expression levels of PTEN, GSK3β, Beclin1, and LC3II decreased; the expression levels of phosphorylated AKT and mTOR proteins expression increased; PC12 autophagy decreased; and cellular viability and survival rates increased.

Discussion: In this in vitro study, we found that increasing miR-199a expression in PC12 cells reduced protein levels of Beclin1 and LC3II, decreased autophagy, enhanced cellular viability, increased survival rate, and ameliorated MPP⁺-induced parkinsonian-like cellular pathologies by targeting pro-autophagic pathways and GSK3β to activate PTEN/AKT/mTOR signaling.