Regulatory mechanism of miR-20a-5p in neuronal damage and inflammation in lipopolysaccharide-induced BV2 cells and MPTP-HCl-induced Parkinson's disease mice

Abstract

Background: Parkinson's disease (PD) is a prevailing neurodegenerative disorder increasingly affecting the elderly population. The involvement of microRNAs (miRNAs) in PD has been confirmed. We sought to explore the molecular mechanism of miR-20a-5p in PD.

Methods: Lipopolysaccharide (LPS)-induced BV2 cell model and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine hydrochloride (MPTP-HCl)-induced PD mouse model were established. miR-20a-5p, inducible nitric oxide synthase (iNOS), interleukin (IL)-6, tumour necrosis factor (TNF)-α, transforming growth factor (TGF)-β1, and IL-10 expression in BV2 cells was examined by reverse transcription - quantitative polymerase chain reaction. Cell viability was assessed by MTT assay. The apoptotic rate and levels of Bcl-2, Bax, cleaved caspase-3, and signal transducer and activator of transmission (STAT)3 were examined by flow cytometry and Western blot. Bioinformatics software predicted the potential binding sites of miR-20a-5p and STAT3. Dual-luciferase experiment verified the binding relationship. Iba1-positive and tyrosine hydroxylase (TH)-positive cell numbers in substantia nigra pars compacta were detected by immunohistochemistry. The effect of miR-20a-5p on motor function in MPTP-induced PD mice was detected by Rota-rod test, Pole test, Traction test and Beam-crossing task.

Results: miR-20a-5p was under-expressed in LPS-induced BV2 cells. Overexpression of miR-20a-5p increased the viability of LPS-induced BV2 cells and reduced apoptosis rates. Moreover, overexpression of miR-20a-5p reduced cleaved caspase-3, Bax, iNOS, IL-6, and TNF-α and increased Bcl-2 and TGF-β1, and IL-10. miR-20a-5p targeted STAT3. STAT3 overexpression partially reversed miR-20a-5p overexpression-mediated effects on LPS-induced BV2 cell viability, apoptosis, and inflammatory responses. miR-20a-5p overexpression inhibited MPTP-induced STAT3 and α-synuclein levels, microglia activation, and inflammatory response, and reduced the loss of TH-positive cells in mice. miR-20a-5p overexpression ameliorated MPTP-induced dyskinesia in PD model mice.

Conclusion: miR-20a-5p alleviates neuronal damage and suppresses inflammation by targeting STAT3 in PD.

Keywords: Parkinson's disease; STAT3; apoptosis; inflammation; miR‐20a‐5p; neuronal damage.