Parkinson’s disease and other alpha-synucleinopathies are progressive neurodegenerative diseases characterized by aggregates of misfolded alpha-synuclein spreading throughout the brain. Recent evidence suggests that the pathological progression is likely due to neuron-to-neuron transfer of these aggregates between neuroanatomically connected areas of the brain. As the impact of this pathological spreading mechanism is currently debated, we aimed to investigate the transfer and subcellular location of alpha-synuclein species in a novel 3D co-culture human cell model based on highly differentiated SH-SY5Y cells. Fluorescently-labeled monomeric, oligomeric and fibrillar species of alpha-synuclein were introduced into a donor cell population and co-cultured with an EGFP-expressing acceptor-cell population of differentiated neuron-like cells. Subsequent transfer and colocalization of the different species were determined with confocal microscopy. We could confirm cell-to-cell transfer of all three alpha-synuclein species investigated. Interestingly the level of transferred oligomers and fibrils and oligomers were significantly higher than monomers, which could affect the probability of seeding and pathology in the recipient cells. Most alpha-synuclein colocalized with the lysosomal/endosomal system, both pre- and postsynaptically, suggesting its importance in the processing and spreading of alpha-synuclein.

The intercellular transfer of alpha-synuclein (α-syn) has been implicated in the progression of Parkinson’s disease (PD) and multiple system atrophy (MSA). The cellular mechanisms underlying this process are now beginning to be elucidated. In this study, we demonstrate that the gap junction protein connexin-32 (Cx32) is centrally involved in the preferential uptake of α-syn oligomeric assemblies (oα-syn) in neurons and oligodendrocytes. In vitro, we demonstrate a clear correlation between Cx32 expression and oα-syn uptake. Pharmacological and genetic strategies targeting Cx32 successfully blocked oα-syn uptake. In cellular and transgenic mice modeling PD and MSA, we observed significant upregulation of Cx32 which correlates with α-syn accumulation. Notably, we could also demonstrate a direct interaction between α-syn and Cx32 in two out of four human PD cases that was absent in all four age-matched controls. These data are suggestive of a link between Cx32 and PD pathophysiology. Collectively, our results provide compelling evidence for Cx32 as a novel target for therapeutic intervention in PD and related α-synucleinopathies. Electronic supplementary material The online version of this article (10.1007/s00401-019-02007-x) contains supplementary material, which is available to authorized users.

No 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.

Alzheimer's disease (AD) is the most common form of dementia globally and is characterized by aberrant accumulations of amyloid-beta (Aβ) and tau proteins. oligomeric forms of these proteins are believed to be most relevant to disease progression, with oligomeric amyloid-β (oAβ) particularly implicated in AD. oAβ pathology spreads among interconnected brain regions, but how oAβ induces pathology in these previously unaffected neurons requires further study. Here, we use well characterized ipSc-derived human neurons to study the early changes to the proteome and phosphoproteome after 24 h exposure to oAβ 1-42. Using nLC-MS/MS and label-free quantification, we identified several proteins that are differentially regulated in response to acute oAβ challenge. At this early timepoint, oAβ induced the decrease of TDP-43, heterogeneous nuclear ribonucleoproteins (hnRNPs), and coatomer complex I (COPI) proteins. Conversely, increases were observed in 20 S proteasome subunits and vesicle associated proteins VAMP1/2, as well as the differential phosphorylation of tau at serine 208. These changes show that there are widespread alterations to the neuronal proteome within 24 h of oAβ uptake, including proteins previously not shown to be related to neurodegeneration. this study provides new targets for the further study of early mediators of AD pathogenesis.Alzheimer's disease (AD) is the most prevalent form of dementia globally and is expected to grow substantially along with the aging global population. Much of the research in this field has focused on the pathological hallmarks of AD, amyloid-β (Aβ) and tau, but the way in which these mediators initiate neuronal pathogenesis at the molecular level requires further understanding. An important consideration in the study of AD is the different properties of Aβ assemblies: monomers, oligomers and fibrils. In particular, there is growing evidence to suggest that low molecular weight Aβ oligomers (oAβ) and protofibrils seed the aggregation of naïve Aβ, confer neurotoxicity, and also correlate to cognitive decline, as reviewed in 1-4 . Much of our current understanding of AD comes from the study of long-term exposure to Aβ in animal models, which has provided valuable insight into the effects of chronic exposure to Aβ, but often overlook the initial steps involved in pathogenesis. In order to further our understanding of the acute effects of exposure to oAβ, we challenged human iPSC-derived neurons with oAβ for 24 h and examined the changes to the proteome and phosphoproteome elicited by this acute oAβ treatment. In this way, we aim to elucidate some of the early effects elicited by oAβ upon neurons.During AD, widespread changes occur in the proteome, including the up-and downregulation of disease-related proteins as well as the post-translational modification (PTM) of proteins. PTMs such as phosphorylation greatly affect the function of proteins and are well known to be important for the pathogenesis of neurodegenerative disorders, where detection of phosphorylated proteins are us...