Inhibition of nSMase2 Reduces the Transfer of Oligomeric α-Synuclein Irrespective of Hypoxia.

Valerie Sackmann,Maitrayee Sardar Sinha,Martin Hallbeck,Anna Ansell-Schultz,Livia Civitelli,Joakim Bergström,Christopher Sackmann

doi:10.3389/fnmol.2019.00200

Valerie Sackmann, Maitrayee Sardar Sinha + Show 5 more

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https://doi.org/10.3389/fnmol.2019.00200

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Abstract

Recently, extracellular vesicles (EVs), such as exosomes, have been proposed to play an influential role in the cell-to-cell spread of neurodegenerative diseases, including the intercellular transmission of α-synuclein (α-syn). However, the regulation of EV biogenesis and its relation to Parkinson’s disease (PD) is only partially understood. The generation of EVs through the ESCRT-independent pathway depends on the hydrolysis of sphingomyelin by neutral sphingomyelinase 2 (nSMase2) to produce ceramide, which causes the membrane of endosomal multivesicular bodies to bud inward. nSMase2 is sensitive to oxidative stress, a common process in PD brains; however, little is known about the role of sphingomyelin metabolism in the pathogenesis of PD. This is the first study to show that inhibiting nSMase2 decreases the transfer of oligomeric aggregates of α-syn between neuron-like cells. Furthermore, it reduced the accumulation and aggregation of high-molecular-weight α-syn. Hypoxia, as a model of oxidative stress, reduced the levels of nSMase2, but not its enzymatic activity, and significantly altered the lipid composition of cells without affecting EV abundance or the transfer of α-syn. These data show that altering sphingolipids can mitigate the spread of α-syn, even under hypoxic conditions, potentially suppressing PD progression.

Highlights

Parkinson’s disease (PD) is a progressive age-related movement disorder that results in the selective loss of midbrain dopaminergic neurons and widespread pathology at later stages of the disease (Braak et al, 2003)
The loss of neutral sphingomyelinase 2 (nSMase2) protein expression in nSMase2 KO cells was confirmed by immunoblotting and immunocytochemistry with N- and C-terminal specific antibodies (Figures 2A–C and Supplementary Figure S1), while Small interfering RNA (siRNA) treatment induced both a significant reduction in protein expression and gene downregulation (Figures 2A– C, 3). neutral sphingomyelinase (nSMase) enzymatic activity was significantly decreased in KO cells (0.63 ± 0.16 mU/ml) compared to control cells (1.48 ± 0.23 mU/ml) (Figure 2D)
SiRNA-treated cells (1.13 ± 0.25 mU/ml) were not significantly modified, it is possible that other nSMases are compensating from the acute transfection and are being detected in the assay

Summary

Introduction

Parkinson’s disease (PD) is a progressive age-related movement disorder that results in the selective loss of midbrain dopaminergic neurons and widespread pathology at later stages of the disease (Braak et al, 2003). The mechanisms of this spreading, including the important role of cell-surface proteins (Reyes et al, 2019) and extracellular vesicles (EVs) (Russo et al, 2012), are beginning to be elucidated. Small EVs, exosomes, are small membranous vesicles (30–150 nm) that are formed by the invagination and budding of endosomal multivesicular bodies from the membrane (Van Niel et al, 2006). These vesicles can serve as vehicles for transferring proteins, lipids, mRNA, and miRNA between cells.

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