Amyloidogenic α-synuclein seeds do not invariably induce rapid, widespread pathology in mice

Sacino, Amanda N.; Brooks, Michael C.; Thomas, M. Albert; McKinney, Alex B.; McGarvey, Nicholas H.; Rutherford, Nicola J.; Ceballos‐Diaz, Carolina; Robertson, Janice; Golde, Todd E.; Giasson, Benoit I.

doi:10.1007/s00401-014-1268-0

Cited by 109 publications

(149 citation statements)

References 106 publications

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“…Our findings also highlight the fact, as we have noted elsewhere (51,52), that there seem to be significant differences in the susceptibility of different models to induction of pathology. Mice expressing A53T αS are susceptible to rapid onset of a neurodegenerative phenotype associated with motor neuron death and a decrease in NMJ integrity in the affected limbs.…”

Section: Discussionsupporting

confidence: 83%

“…Previous experimental studies have provided evidence for in vivo prion-like spread of αS pathology (21-26); however, several findings here, and in other recent studies from our group, suggest that additional nonexclusive mechanisms inducing or promoting αS inclusion pathology formation should be considered (33,(51)(52)(53). Indeed, (i) the distribution of αS pathology in M83 Tg mice IM injected with various forms of αS was identical to that seen in untreated, aged M83 Tg mice, (ii) IM injection of a nonamyloidogenic form of αS (Δ71-82) induced CNS αS inclusion pathology, albeit less efficiently than fibrillar αS, and (iii) there were significant increases of both astrogliosis and microgliosis associated with the regions where abundant αS pathology was induced.…”

Section: Discussionmentioning

confidence: 46%

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Intramuscular injection of α-synuclein induces CNS α-synuclein pathology and a rapid-onset motor phenotype in transgenic mice

Sacino¹,

Brooks²,

Thomas³

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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It has been hypothesized that α-synuclein (αS) misfolding may begin in peripheral nerves and spread to the central nervous system (CNS), leading to Parkinson disease and related disorders. Although recent data suggest that αS pathology can spread within the mouse brain, there is no direct evidence for spread of disease from a peripheral site. In the present study, we show that hind limb intramuscular (IM) injection of αS can induce pathology in the CNS in the human Ala53Thr (M83) and wild-type (M20) αS transgenic (Tg) mouse models. Within 2-3 mo after IM injection in αS homozygous M83 Tg mice and 3-4 mo for hemizygous M83 Tg mice, these animals developed a rapid, synchronized, and predictable induction of widespread CNS αS inclusion pathology, accompanied by astrogliosis, microgliosis, and debilitating motor impairments. In M20 Tg mice, starting at 4 mo after IM injection, we observed αS inclusion pathology in the spinal cord, but motor function remained intact. Transection of the sciatic nerve in the M83 Tg mice significantly delayed the appearance of CNS pathology and motor symptoms, demonstrating the involvement of retrograde transport in inducing αS CNS inclusion pathology. Outside of scrapie-mediated prion disease, to our knowledge, this findiing is the first evidence that an entire neurodegenerative proteinopathy associated with a robust, lethal motor phenotype can be initiated by peripheral inoculation with a pathogenic protein. Furthermore, this facile, synchronized rapid-onset model of α-synucleinopathy will be highly valuable in testing disease-modifying therapies and dissecting the mechanism(s) that drive αS-induced neurodegeneration.amyloid | Parkinson disease S ynucleinopathies are a group of diseases defined by the presence of amyloidogenic α-synuclein (αS) inclusions that can occur in neurons and glia of the central nervous system (CNS) (1-4). In Parkinson disease (PD), a causative role for αS has been established via the discovery of mutations in the αS gene SNCA resulting in autosomal-dominant PD (4-11). Although αS inclusions (e.g., Lewy bodies) are the hallmark pathology of PD, how they contribute to disease pathogenesis remains controversial (1,3,4,12).Postmortem studies have suggested that αS pathology may spread following neuroanatomical tracts (13-15) and between cells (16-18). αS pathology has also been found in the peripheral nervous system (PNS): for example, in the enteric and pelvic plexus (19,20). And it has been suggested that αS pathology might originate in the nerves of the PNS and spread to the CNS (14). Experimentally, it has been reported that intracerebral injections of preformed amyloidogenic αS fibrils in nontransgenic (nTg) and αS transgenic (Tg) mice induce the formation of intracellular αS inclusions that appear to progress from the site of injection (21-26). Collectively, these studies support the notion that αS inclusion pathology may propagate via a prion-like conformational self-templating mechanism (27, 28). A caveat of the direct intracerebral injection of αS is tha...

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Section: Discussionsupporting

confidence: 83%

Section: Discussionmentioning

confidence: 46%

Intramuscular injection of α-synuclein induces CNS α-synuclein pathology and a rapid-onset motor phenotype in transgenic mice

Sacino¹,

Brooks²,

Thomas³

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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286

343

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“…Taken together with the in vitro data reported earlier, these findings are consistent with the concept of amyloid fibril structural polymorphism (5, 29) (i.e., that aggregates of the same peptide can exhibit a variety of conformational structures). This observation is interesting in light of recent reports demonstrating that fibrillar mouse explants do not induce seeding in all cases in nontransgenic mice (30) and in other animal models of disease (31)(32)(33). Furthermore, our results indicate that in addition to fibrillar AS species, soluble AS can readily aggregate following uptake by cells, a conclusion similar to the conclusion drawn from experiments with Tau in a cellular model of Alzheimer's disease (21).…”

Section: Exogenously Added Monomeric As Triggers the Nucleation Ofsupporting

confidence: 87%

Nanoscopic insights into seeding mechanisms and toxicity of α-synuclein species in neurons

Pinotsi

Michel

Buell

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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New strategies for visualizing self-assembly processes at the nanoscale give deep insights into the molecular origins of disease. An example is the self-assembly of misfolded proteins into amyloid fibrils, which is related to a range of neurodegenerative disorders, such as Parkinson's and Alzheimer's diseases. Here, we probe the links between the mechanism of α-synuclein (AS) aggregation and its associated toxicity by using optical nanoscopy directly in a neuronal cell culture model of Parkinson's disease. Using superresolution microscopy, we show that protein fibrils are taken up by neuronal cells and act as prion-like seeds for elongation reactions that both consume endogenous AS and suppress its de novo aggregation. When AS is internalized in its monomeric form, however, it nucleates and triggers the aggregation of endogenous AS, leading to apoptosis, although there are no detectable cross-reactions between externally added and endogenous protein species. Monomer-induced apoptosis can be reduced by pretreatment with seed fibrils, suggesting that partial consumption of the externally added or excess soluble AS can be significantly neuroprotective.T he proliferation of α-synuclein (AS) aggregates (1-6), including the existence of distinct "prion-like" strains (7, 8) as well as their spatial propagation throughout the brain (9), has been proposed to occur in the brains of patients suffering from Parkinson's disease, but their links to pathology and to neuronal death have remained elusive (10-13). In this study, we use optical nanoscopy (14-18) to observe neurons directly and to assay AS internalization, as well as fibril-induced templating reactions involving endogenous AS at the molecular level. We further correlate this information with toxic phenotypes.We have previously shown that the presence of AS short fibrils (seed fibrils or seeds) preformed in vitro (Materials and Methods and Fig. 1 and Fig. S1) favors elongation reactions over spontaneous nucleation (term used for seed-independent aggregation hereafter) in vitro (18). Furthermore, the seed fibrils were found to display highly inhomogeneous growth kinetics, with a significant fraction showing little or no growth at all (18). Here, we use two-color direct stochastic optical reconstruction microscopy (dSTORM), a superresolution imaging method (14), to investigate how such processes may be modified in the cellular environment. The results show the potential of this technique for studying the mechanisms of aggregation events in vivo and provide evidence for the neuroprotective role of reducing the concentration of free AS in the cellular environment. Results and DiscussionExternally Added Seed Fibrils of AS Act as Templates for Exogenously Added Monomeric AS and Prevent Its Nucleation. We first incubated neuronal cells with AS seed fibrils [50 nM, 5% covalently labeled with Alexa Fluor 568 (AF568), green] and AS monomers [500 nM, 10% covalently labeled with Alexa Fluor 647 (AF647), red], either each individually or both in sequence. Either seed fibrils and...

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“…pSer129 clone 81A is a mouse monoclonal antibody that reacts with ␣S phosphorylated at Ser129 (pSer129) (62), but also cross-reacts with phosphorylated low-molecular-mass neurofilament subunit (63). The rabbit monoclonal antibody pSer129/EP1536Y that reacts with ␣S phosphorylated at Ser129 and does not cross-react with phosphorylated low-molecular-mass neurofilament subunit (6) was obtained from Abcam (Cambridge, MA).…”

Section: Methodsmentioning

confidence: 99%

Robust Central Nervous System Pathology in Transgenic Mice following Peripheral Injection of α-Synuclein Fibrils

Ayers

Brooks

Rutherford

et al. 2017

J Virol

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Misfolded ␣-synuclein (␣S) is hypothesized to spread throughout the central nervous system (CNS) by neuronal connectivity leading to widespread pathology. Increasing evidence indicates that it also has the potential to invade the CNS via peripheral nerves in a prion-like manner. On the basis of the effectiveness following peripheral routes of prion administration, we extend our previous studies of CNS neuroinvasion in M83 ␣S transgenic mice following hind limb muscle (intramuscular [i.m.]) injection of ␣S fibrils by comparing various peripheral sites of inoculations with different ␣S protein preparations. Following intravenous injection in the tail veins of homozygous M83 transgenic (M83 ϩ/ϩ ) mice, robust ␣S pathology was observed in the CNS without the development of motor impairments within the time frame examined. Intraperitoneal (i.p.) injections of ␣S fibrils in hemizygous M83 transgenic (M83 ϩ/Ϫ ) mice resulted in CNS ␣S pathology associated with paralysis. Interestingly, injection with soluble, nonaggregated ␣S resulted in paralysis and pathology in only a subset of mice, whereas soluble Δ71-82 ␣S, human ␤S, and keyhole limpet hemocyanin (KLH) control proteins induced no symptoms or pathology. Intraperitoneal injection of ␣S fibrils also induced CNS ␣S pathology in another ␣S transgenic mouse line (M20), albeit less robustly in these mice. In comparison, i.m. injection of ␣S fibrils was more efficient in inducing CNS ␣S pathology in M83 mice than i.p. or tail vein injections. Furthermore, i.m. injection of soluble, nonaggregated ␣S in M83 ϩ/Ϫ mice also induced paralysis and CNS ␣S pathology, although less efficiently. These results further demonstrate the prion-like characteristics of ␣S and reveal its efficiency to invade the CNS via multiple routes of peripheral administration. IMPORTANCEThe misfolding and accumulation of ␣-synuclein (␣S) inclusions are found in a number of neurodegenerative disorders and is a hallmark feature of Parkinson's disease (PD) and PD-related diseases. Similar characteristics have been observed between the infectious prion protein and ␣S, including its ability to spread from the peripheral nervous system and along neuroanatomical tracts within the central nervous system. In this study, we extend our previous results and investigate the efficiency of intravenous (i.v.), intraperitoneal (i.p.), and intramuscular (i.m.) routes of injection of ␣S fibrils and other protein controls. Our data reveal that injection of ␣S fibrils via these peripheral routes in ␣S-overexpressing mice are capable of inducing a robust ␣S pathology and in some cases cause paralysis. Furthermore, soluble, nonaggregated ␣S also induced ␣S pathology, albeit with much less efficiency. These findings further support and extend the idea of ␣S neuroinvasion from peripheral exposures.

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Amyloidogenic α-synuclein seeds do not invariably induce rapid, widespread pathology in mice

Cited by 109 publications

References 106 publications

Intramuscular injection of α-synuclein induces CNS α-synuclein pathology and a rapid-onset motor phenotype in transgenic mice

Intramuscular injection of α-synuclein induces CNS α-synuclein pathology and a rapid-onset motor phenotype in transgenic mice

Nanoscopic insights into seeding mechanisms and toxicity of α-synuclein species in neurons

Robust Central Nervous System Pathology in Transgenic Mice following Peripheral Injection of α-Synuclein Fibrils

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