No abstract

Objective TDP-43 is deposited as cytoplasmic and intranuclear inclusions in brains of subjects with frontotemporal lobar degeneration with ubiquitinated inclusions (FTLD-U) and amyotrophic lateral sclerosis (ALS). Previous studies reported that abnormal phosphorylation takes place in deposited TDP-43. The aim of this study was to identify the phosphorylation sites and responsible kinases, and to clarify the pathological significance of phosphorylation of TDP-43. Methods We generated multiple antibodies specific to phosphorylated TDP-43 by immunizing phosphopeptides of TDP-43, and analyzed FTLD-U and ALS brains by immunohistochemistry, immunoelectron microscopy and immunoblots. Additionally, we performed investigations aimed at identifying the responsible kinases and we assessed the effects of phosphorylation on TDP-43 oligomerization and fibrillization. Results We identified multiple phosphorylation sites in carboxyl-terminal regions of deposited TDP-43. Phosphorylation-specific antibodies stained more inclusions than antibodies to ubiquitin and, unlike existing commercially-available anti-TDP-43 antibodies, did not stain normal nuclei. Ultrastructurally, these antibodies labeled abnormal fibers of 15 nm diameter, and on immunoblots recognized hyperphosphorylated TDP-43 at 45 kDa, with additional 22–28 kDa fragments in sarkosyl-insoluble fractions from FTLD-U and ALS brains. The phosphorylated epitopes were generated by casein kinase 1 and 2, and phosphorylation led to increased oligomerization and fibrillization of TDP-43. Interpretation These results suggest that phosphorylated TDP-43 is a major component of the inclusions, and that abnormal phosphorylation of TDP-43 is a critical step in the pathogenesis of FTLD-U and ALS. Phosphorylation-specific antibodies will be powerful tools for the investigation of these disorders.

We assessed the clinicopathological features of 92 patients with primary Sjögren's syndrome-associated neuropathy (76 women, 16 men, 54.7 years, age at onset). The majority of patients (93%) were diagnosed with Sjögren's syndrome after neuropathic symptoms appeared. We classified these patients into seven forms of neuropathy: sensory ataxic neuropathy (n = 36), painful sensory neuropathy without sensory ataxia (n = 18), multiple mononeuropathy (n = 11), multiple cranial neuropathy (n = 5), trigeminal neuropathy (n = 15), autonomic neuropathy (n = 3) and radiculoneuropathy (n = 4), based on the predominant neuropathic symptoms. Acute or subacute onset was seen more frequently in multiple mononeuropathy and multiple cranial neuropathy, whereas chronic progression was predominant in other forms of neuropathy. Sensory symptoms without substantial motor involvement were seen predominantly in sensory ataxic, painful sensory, trigeminal and autonomic neuropathy, although the affected sensory modalities and distribution pattern varied. In contrast, motor weakness and muscle atrophy were observed in multiple mononeuropathy, multiple cranial neuropathy and radiculoneuropathy. Autonomic symptoms were often seen in all forms of neuropathy. Abnormal pupils and orthostatic hypotension were particularly frequent in sensory ataxic, painful, trigeminal and autonomic neuropathy. Unelicited somatosensory evoked potentials and spinal cord posterior column abnormalities in MRI were observed in sensory ataxic, painful and autonomic neuropathy. Sural nerve biopsy specimens (n = 55) revealed variable degrees of axon loss. Predominantly large fibre loss was observed in sensory ataxic neuropathy, whereas predominantly small fibre loss occurred in painful sensory neuropathy. Angiitis and perivascular cell invasion were seen most frequently in multiple mononeuropathy, followed by sensory ataxic neuropathy. The autopsy findings of one patient with sensory ataxic neuropathy showed severe large sensory neuron loss paralleling to dorsal root and posterior column involvement of the spinal cord, and severe sympathetic neuron loss. Degrees of neuron loss in the dorsal and sympathetic ganglion corresponded to segmental distribution of sensory and sweating impairment. Multifocal T-cell invasion was seen in the dorsal root and sympathetic ganglion, perineurial space and vessel walls in the nerve trunks. Differential therapeutic responses for corticosteroids and IVIg were seen among the neuropathic forms. These clinicopathological observations suggest that sensory ataxic, painful and perhaps trigeminal neuropathy are related to ganglioneuronopathic process, whereas multiple mononeuropathy and multiple cranial neuropathy would be more closely associated with vasculitic process.

Synucleinopathies are human neurodegenerative diseases that include multiple system atrophy (MSA), Parkinson's disease, Parkinson's disease dementia (PDD) and dementia with Lewy bodies (DLB) (1). Existing treatments are at best symptomatic. These diseases are characterised by the presence in brain cells of filamentous inclusions of αsynuclein, the formation of which is believed to cause disease (2,3). However, the structures of α-synuclein filaments from human brain are not known. Here we show, using electron cryo-microscopy, that α-synuclein inclusions from MSA are made of two types of filaments, each of which consists of two different protofilaments. Non-proteinaceous molecules are present at the protofilament interfaces. By two-dimensional class averaging, we show that α-synuclein filaments from the brains of patients with MSA and DLB are different, suggesting that distinct conformers (or strains) characterise synucleinopathies. As was the case of tau assemblies (4-9), the structures of α-synuclein filaments extracted from the brains of individuals with MSA differ from those formed in vitro using recombinant proteins, with implications for understanding the mechanisms of aggregate propagation and neurodegeneration in human brain. These findings have diagnostic and potential therapeutic relevance, .

Ordered assembly of the tau protein into filaments characterizes multiple neurodegenerative diseases, which are called tauopathies. We previously reported that by electron cryo-microscopy (cryo-EM), tau filament structures from Alzheimer's disease (1,2), chronic traumatic encephalopathy (CTE) (3), Pick's disease (4) and corticobasal degeneration (CBD) (5) are distinct. Here we show that the structures of tau filaments from typical and atypical progressive supranuclear palsy (PSP), the most common tauopathy after Alzheimer's disease, define a previously unknown, three-layered fold. Moreover, the tau filament structures from globular glial tauopathy (GGT, Types I and II) are similar to those from PSP. The tau filament fold of argyrophilic grain disease (AGD) differs from the above and resembles the four-layered CBD fold. The majority of tau filaments from agingrelated tau astrogliopathy (ARTAG) also have the AGD fold. Surprisingly, tau protofilament structures from inherited cases with mutations +3/+16 in intron 10 of MAPT, the microtubule-associated protein tau gene, are identical to those from AGD, suggesting that a relative overproduction of four-repeat tau can give rise to the AGD fold. Finally, tau filament structures from cases of familial British dementia (FBD) and familial Danish dementia (FDD) are the same as those from Alzheimer's disease and primary age-related tauopathy (PART). These structures provide the basis for a classification of tauopathies that also allows identification of new entities, as we show here for a case diagnosed as PSP, but with abundant spherical 4R tau inclusions in limbic and other brain areas. The structures of the tau fold of this new disease (Limbic-predominant Neuronal inclusion body 4R Tauopathy, LNT) were intermediate between those of GGT and PSP.