Introduction
The term α-synucleinopathy unites a group of neurodegenerative diseases which share the pathological hallmark of fibrillary inclusions in which α-synuclein protein is the major component. The three most common members of this group are Parkinson’s disease (PD), dementia with Lewy bodies (DLB) and multiple system atrophy (MSA). PD and DLB have common neuropathological features including deposition of fibrillar α-synuclein in Lewy bodies (LB) and Lewy neurites (LN). In MSA, α-synuclein is aggregated in oligodendrocytes forming the hallmark lesion, the glial cytoplasmic inclusion (GCI), and also in neuronal cytoplasmic inclusions (NCIs), cell processes and to a lesser extent in neuronal and glial nuclei [
1,
28].
Several missense mutations of the
SNCA gene have been identified in families with autosomal dominant forms of PD. No
SNCA mutation has been linked with MSA, however, polymorphisms of the gene have been associated with increased risk of the disease in Caucasian populations, although these results have not been replicated in all populations [
2,
70,
71,
83]. The
SNCA gene, which encodes the 140-amino acid protein α-synuclein, is located on chromosome 4q21-23. In Parkinson’s disease, the first
SNCA mutation to be described was A53T in a Greek-Italian family [
62] and this was subsequently identified in families of Asian, Swedish and Polish origin [
5,
12,
42,
49,
59,
61,
66,
77]. Two further missense mutations of
SNCA were identified, A30P [
43] and E46K [
84] in German and Basque families, respectively. Very recently, we have reported the novel H50Q
SNCA mutation [
65]. Missense mutations in the N-terminal region of α-synuclein are reported to have a direct impact on α-synuclein conformation and function. The A53T and E46K mutant forms of α-synuclein exhibit faster fibrillisation kinetics than wild-type protein [
12], while fibrillisation of the A30P mutant protein is slower and results in fewer complex fibrils in LBs [
47]. Duplication and triplication of
SNCA have been discovered in a small number of families and sporadic cases of levodopa-responsive PD and, where described, pathological features of PD, in addition to GCIs similar to those of MSA, are observed [
10,
22,
27,
31,
32,
55,
56]. The number of
SNCA locus replicates is known to influence disease progression, such that triplication causes earlier onset and a more rapid clinical course than
SNCA duplication [
31,
75].
Pathological inclusions of fibrillar α-synuclein have distinct morphologies and distribution depending on disease type. For example, in PD, the characteristic LBs and LNs occur in brainstem nuclei and usually exhibit a hierarchical spread to involve limbic and neocortical regions with disease progression [
8], although not all cases conform to the proposed pattern of disease progression [
36]. In MSA, GCIs are the most abundant form of fibrillar α-synuclein inclusion and together with neuronal cytoplasmic or nuclear inclusions are distributed widely in the striatonigral, olivopontocerebellar and other regions [
1,
4,
33,
60].
We report a family with young-onset PD and a mutation in
SNCA that segregates with the disease. We describe an α-synucleinopathy with both PD and MSA-like neuropathological features together with involvement of the striatum and severe CA2/3 neuronal loss. The distribution of neuronal and oligodendroglial inclusions immunoreactive for α-synuclein, ubiquitin and p62 is described. The phosphorylation state of α-synuclein within inclusions and the relationship of α-synuclein to intracellular accumulation of tau and TDP-43 are also investigated. Together the evidence reveals neuropathological similarities to both the A53T
SNCA mutation and multiplication cases with additional unique striatal and neocortical pathology [
27,
48].
Discussion
We provide detailed clinical, genetic and neuropathological characterisation of an α-synucleinopathy in a patient carrying a G51D α-synuclein mutation, clinically diagnosed with juvenile parkinsonism. Neuropathological analysis revealed a combination of the characteristic profile of SN and LC neuronal loss together with neuronal α-synuclein immunoreactive inclusions consistent with PD as well as severe hippocampal, cortical and striatal α-synuclein pathology. An additional feature was the presence of GCI-like oligodendroglial inclusions with a distribution similar to that found in MSA. This case shows some similarity to reports of A53T mutation and multiplication of
SNCA (Table
2) and also to a Japanese kindred reported in an abstract as carrying a G51D
SNCA mutation, but without detailed segregation data [
50,
51]. However, features including dense accumulation of α-synuclein-positive inclusions in the striatum and very severe neocortical α-synuclein pathology affecting both superficial and deep cortical laminae distinguish this case from other reported cases with
SNCA mutations.
Table 2
Neuropathology of α-synuclein mutation cases where α-synuclein immunohistochemistry data are available
Golbe et al. [ 26] and Duda et al. [ 19] | A53T | 2 | Severe | Mild–moderate | Yes | No | PD-like | Moderate grains and threads | Deep laminae | Not described | I | Not described |
| A53T | 2 | Severe | Severe | No | Severe | PD-like | None | Deep laminae | None | 0 | Not described |
| A53T | 2 | Severe | Mild | Severe (n = 1) | CA2 (n = 1) | PD-like | Grains and threads (n = 1) | Deep laminae | Small numbers GCIs | IV (n = 1), I (n = 1) | Limbic and cortical (n = 1, with CA1 neuronal loss) |
| A30P | 1 | Severe | Not described | Not described | Not described | PD-like | LBs | Deep laminae | Small numbers CB and astrocytes of PD type | II | Not described |
| E46 K | 1 | Moderate–severe | Mild | Not described | Not described | PD-like | None | Present, distribution not described | Not described | 0 | Not described |
| Duplication | 1 | Severe | Severe | None | Moderate | Features of PD and MSA | None | Present, distribution not described | Small numbers GCIs and CB | I | Not described |
| Duplication | 1 | Severe | Not described | None | Severe | PD-like | Not described | Present, distribution not described | Few, type not specified | III | Not described |
| Triplication | 1 | Severe | Severe | None | Severe | Features of PD and MSA | LBs and neurites | Predominantly deep laminae | Small numbers GCIs and CB | 0 | Not described |
| Triplication | 1 | Severe | Severe | None | Severe | PD-likea
| Not described | Present, distribution not described | Not described | Not described | Not described |
This report | G51D | 1 | Severe | Severe | Mild | Severe | Features of PD and MSA | Frequent neuronal inclusions and threads | Superficial and deep laminae | GCIs and CB | II | Limbic |
In comparison with other reported mutations in the
SNCA gene, the age of onset in this family, clinical features and progression are most similar to the
SNCA triplication [
16] and A53T mutations, which are typically associated with young-onset PD frequently associated with cognitive impairment and hallucinations. This is similar to the family where the proband presented early at 19 years while his father and sister presented at age 39 and 40 years, respectively.
In this family, an
SNCA G > A heterozygous mutation at codon 51 causes a glycine to aspartic acid amino acid change (Fig.
1a), which segregates with the disease (Fig.
1b) and was not found in over 4,500 control individuals. The G51D mutation is located in the N-terminal domain of the protein, a region required for lipid affinity and membrane binding and may thus influence these functions (Fig.
2) [
20,
37,
68]. As the A53T and E46K mutations result in faster fibrillisation of α-synuclein [
13,
62], we postulate that the G51D mutation might have a similar effect.
The neuropathological findings included some features of both PD and MSA. In common with PD and MSA, there was severe neuronal loss in the SN. Neuronal α-synuclein pathology had a distribution compatible with PD in that brainstem, limbic and cortical regions were affected. Unusual for PD, however, was the severe neuronal loss in the CA2/3 subregions of the hippocampus and the extensive accumulation of neuronal α-synuclein in the hippocampus, including the DF, and striatum [
7,
8,
38,
52]. Involvement of the DF by neuronal α-synuclein inclusions has been described previously in the case of DLB with additional MSA-type pathology and in some cases of MSA [
73,
74,
78]. The severe neuronal loss observed in CA2/3 of this case bears similarity to that described in association with A53T
SNCA mutation [
48,
77] and in cases of multiplication of
SNCA, though this pattern of loss was suggested to be a unique feature of the latter [
21] (Table
2). As frequently observed in PD and DLB, and also reported in conjunction with A53T
SNCA mutation, α-synuclein-positive threads were frequent in CA2/3 [
17,
18,
34,
35]. The severe neuronal loss we observed is likely to be associated with the susceptibility of these hippocampal subregions to α-synuclein accumulation.
Oligodendroglial pathology in the form of GCI-like inclusions in regions such as the posterior frontal white matter, pontine base and cerebellar white matter was also notable and prompts comparison with MSA. In common with GCIs in MSA, these inclusions were immunoreactive for αB-crystallin [
63]. Oligodendroglial pathology with similar morphological appearances has been described in cases with
SNCA multiplication or A53T mutation (Table
2) and, interestingly, was also reported in a sporadic DLB case [
74]. Neuronal and glial α-synuclein pathology in the striatum is also a common feature of MSA and we noted very frequent striatal neuronal α-synuclein immunoreactive inclusions coupled with severe gliosis. However, in notable contrast to MSA, in which the putamen has a gradient of pathology most severely affecting the posterior and dorsal aspects of this nucleus with less severe involvement of the caudate, there was a uniform distribution of pathology in all regions of both nuclei in this case [
1,
58]. This pattern of striatal pathology appears to be unique to our case as it has not been described in the context of other
SNCA mutations (Table
2).
Hippocampal sclerosis with severe neuronal loss affecting CA1 and the subiculum is a feature of many neurodegenerative diseases and is often associated with TDP-43 immunoreactive inclusions in residual neurons in these regions and elsewhere [
64]. TDP-43 pathology is reported to be frequent in DLB cases, but is considerably rarer in MSA and in PD [
3,
24,
30,
53]. While we observed sparse TDP-43-positive inclusions in these areas, the overall pattern of neuronal loss, being most severe in CA2/3, did not resemble typical hippocampal sclerosis. Of particular note was the abundance of TDP-43 immunoreactive NCIs in the caudate and putamen, regions also vulnerable to TDP-43 pathology in FTLD-TDP [
9]. We did not observe TDP-43-positive inclusions in neocortex or brainstem motor nuclei. A further neuropathological feature, which may be distinctive in the G51D α-synuclein mutation, is the distribution of neocortical α-synuclein pathology. In PD, MSA and other
SNCA mutations, α-synuclein pathology is predominantly found in the deep cortical laminae (Table
2) [
19,
26,
27,
48,
72,
77] compared with our observation of severe involvement of both the superficial and deep cortical layers.
We further investigated the nature of intracellular inclusions and Gallyas silver impregnation indicated the presence of fibrillar protein. Phosphorylation of α-synuclein at Ser129 is a feature of LB pathology and may promote oligomerisation, while phosphorylation at Y125 may increase protein fibrillisation of α-synuclein [
11,
29,
54]. Using immunofluorescence, we showed phosphorylation of α-synuclein at both Y125 and Ser129. Although the majority of neuronal and glial inclusions were immunoreactive for p62 and ubiquitin, a proportion remained unstained and these possibly represented an early phase of inclusion formation [
40,
41,
44]. A varying degree of tau pathology has been reported in cases of
SNCA mutation or multiplication sometimes co-localising with α-synuclein in neuronal inclusions [
19,
27,
32,
48,
57,
72,
84]. We report tau pathology in the hippocampal formation and entorhinal cortex corresponding to Braak and Braak stage II, but with additional involvement of the DF. Phosphorylated tau co-localised with a subpopulation of neuronal α-synuclein inclusions, particularly in the CA1 and DF. A relationship between tau and both the levels and aggregation state of α-synuclein, such that greater numbers of tau-positive inclusions may increase α-synuclein pathology, is well described [
14,
15,
25,
46]. More recently, it has been shown that α-synuclein oligomers can seed tau aggregation in vitro and this may explain the occurrence of tau in a proportion of α-synuclein-containing inclusions [
45].
The defining neuropathological hallmark of MSA is the presence of α-synuclein-containing GCIs coupled with neurodegeneration in the striatonigral and/or olivopontocerebellar regions [
79]. The mechanism of GCI formation is currently unknown and mature oligodendrocytes are not thought to express α-synuclein under normal circumstances [
76]. Evidence of GCI-like pathology in cases of
SNCA multiplication [
27,
57], A53T mutation [
48] and in this case of G51D mutation, provide a strong link between these mutations and the pathological mechanisms of MSA. One of the earliest stages of MSA pathogenesis may involve the overexpression or aberrant localisation of α-synuclein in oligodendrocytes, where it becomes fibrillar [
69] and forms GCIs [
39]. Greater understanding of the effect of the G51D and A53T mutations and
SNCA multiplication may shed further light on the pathological cascades, which result in GCI formation. The data presented indicate that G51D
SNCA mutation results in a neuropathological profile, which shares some neuropathological features of both PD and MSA and, therefore understanding the consequences of this mutation, has the potential to provide greater insight into the role of α-synuclein mutation or dysfunction in the pathogenesis of PD and also MSA. Understanding the biology of this G51D
SNCA mutation could help us to target pathways in PD, MSA and other synucleinopathies, which lead to neuronal and glial α-synuclein accumulation.