Heterogeneity of cortical pTDP-43 inclusion morphologies in amyotrophic lateral sclerosis.

BACKGROUND: Despite the presence of significant cortical pTDP-43 inclusions of heterogeneous morphologies in patients diagnosed with amyotrophic lateral sclerosis (ALS), pathological subclassification is routinely performed in the minority of patients with concomitant frontotemporal dementia (FTD). OBJECTIVE: In order to improve current understanding of the presence and relevance of pathological pTDP-43 subtypes in ALS, the present study examined the pattern of cortical pTDP-43 aggregates in 61 ALS cases without FTD. RESULTS: Based on the presence, morphology and composition of pTDP-43 pathology, three distinct ALS-TDP subtypes were delineated: (1) A predominant pattern of pTDP-43 granulofilamentous neuronal inclusions (GFNIs) and grains that were immuno-negative for p62 was identified in 18% of cases designated ALS-TDP type E; (2) neuronal cytoplasmic inclusions (NCIs) that were immuno-positive for both pTDP-43 and p62 were observed in 67% of cases assigned ALS-TDP type B; and (3) scarce cortical pTDP-43 and p62 aggregates were identified in 15% of cases coined ALS-TDP type SC (scarce cortical). Quantitative analyses revealed a significantly greater burden of pTDP-43 GFNI and grains in ALS-TDP type E. Principal component analysis demonstrated significant relationships between GFNIs, grains and ALS-TDP subtypes to support the distinction of subtypes E and B. No significant difference in age at death or disease duration was found between ALS-TDP subgroups to suggest that these subtypes represent earlier or later stages of the same disease process. Instead, a significantly higher ALS-TDP stage, indicating greater topographical spread of pTDP-43, was identified in ALS-TDP type E. Alzheimer's disease neuropathological change (ABC score ≥ intermediate) and Lewy body disease (Braak stage ≥ IV) was more prevalent in the ALS-TDP type SC cohort, which also demonstrated a significantly lower overall cognitive score. CONCLUSION: In summary, the present study demonstrates that ALS-TDP does not represent a single homogenous neuropathology. We propose the subclassification of ALS-TDP into three distinct subtypes using standard immuno-stains for pTDP-43 and p62 in the motor cortex, which is routinely sampled and evaluated for diagnostic neuropathological characterisation of ALS. We propose that future studies specify both clinicopathological group and pTDP-43 subtype to advance current understanding of the pathogenesis of clinical phenotypes in pTDP-43 proteinopathies, which will have significant relevance to the development of targeted therapies for this heterogeneous disorder.

Survival in sporadic ALS is associated with lower p62 burden in the spinal cord.

The autophagy marker p62 appears as a consistent component of pathological aggregates in amyotrophic lateral sclerosis (ALS) and its modulation to facilitate protein degradation has been proposed as a potential therapeutic target. Importantly, recent studies have implicated diffuse phosphorylated TDP-43 inclusions that are immuno-negative for p62 in more rapid disease, highlighting the need for better understanding of p62 involvement in ALS pathogenesis. The present study set out to assess p62 pathology in the motor neurons of 31 patients with sporadic ALS that had either a short (<2 years) or longer (4-7 years) disease duration to determine its association with pTDP-43 pathology, motor neuron loss, and survival in sporadic disease. Our results identified significantly more cytoplasmic p62 aggregates in the spinal cord of patients with a shorter survival. Disease duration demonstrated a negative association with p62 burden and density of remaining motor neurons in the spinal cord, suggesting that survival in sporadic ALS is associated with the successful clearance of lower motor neurons with p62 aggregates. These findings implicate the autophagy pathway in ALS survival and provide support for further study of p62 as a potential prognostic biomarker in ALS.

Treatment with the copper compound CuATSM has no significant effect on motor neuronal pathology in patients with ALS.

AIMS: Although the orally available brain-penetrant copper compound CuATSM has demonstrated promising effects in SOD1-linked mouse models, the impact of CuATSM on disease pathology in patients with amyotrophic lateral sclerosis (ALS) remains unknown. METHODS: The present study set out to address this deficit by performing the first pilot comparative analysis of ALS pathology in patients that had been administered CuATSM and riluzole [N = 6 cases composed of ALS-TDP (n = 5) and ALS-SOD1 (n = 1)] versus riluzole only [N = 6 cases composed of ALS-TDP (n = 4) and ALS-SOD1 (n = 2)]. RESULTS: Our results revealed no significant difference in neuron density or TDP-43 burden in the motor cortex and spinal cord of patients that had received CuATSM compared with patients that had not. In patients that had received CuATSM, p62-immunoreactive astrocytes were observed in the motor cortex and reduced Iba1 density was found in the spinal cord. However, no significant difference in measures of astrocytic activity and SOD1 immunoreactivity was found with CuATSM treatment. DISCUSSION: These findings, in this first postmortem investigation of patients with ALS in CuATSM trials, demonstrate that in contrast to that seen in preclinical models of disease, CuATSM does not significantly alleviate neuronal pathology or astrogliosis in patients with ALS.

Coexisting Lewy body disease and clinical parkinsonism in amyotrophic lateral sclerosis.

BACKGROUND: Amyotrophic lateral sclerosis (ALS) is associated with a range of clinical phenotypes and shows progressive degeneration of upper and/or lower motor neurons, and phosphorylated 43 kDa TAR DNA-binding protein (pTDP-43) inclusions in motor and non-motor pathways. Parkinsonian features have been reported in up to 30% of ALS patients, and Lewy bodies, normally associated with Lewy body disease (LBD), have been reported in a small number of ALS cases, with unknown clinical relevance. This study investigates the prevalence of clinically relevant LBD in a prospectively studied ALS cohort to determine whether concomitant pathology contributes to the clinical heterogeneity. METHODS: All ALS cases held by the New South Wales Brain Bank (n = 97) were screened for coexisting LBD consistent with clinical disease (Braak ≥ stage IV). Relevant clinical and genetic associations were determined. RESULTS: Six cases had coexisting LBD Braak ≥ stage IV pathology. The age at symptom onset (69 ± 7 years) and disease duration (4 ± 3 years) in ALS cases with coexisting LBD did not differ from ALS cases. Three patients had lower limb onset and two patients had bulbar onset. Two patients developed the clinical features of Parkinson's disease, with one receiving a dual diagnosis. All cases had no known relevant family history or genetic abnormalities. CONCLUSION: The prevalence of clinically relevant LBD pathology in ALS is higher than in the general population, and has implications for clinical and neuropathological diagnoses and the identification of biomarkers.

A Practical Approach to Differentiate the Frontotemporal Tauopathy Subtypes.

This study proposes a practical approach, using the minimum number of brain regions and stains, to consolidate previously published neuropathological criteria into one operationalized schema to differentiate subtypes of frontotemporal lobar degeneration with tau-immunopositive inclusions (FTLD-tau). This approach uses the superior frontal and precentral cortices and hippocampus stained for phosphorylated-tau, p62 and modified Bielschowsky silver, and the midbrain stained only for modified Bielschowsky silver. Accuracy of interrater reliability was determined by 10 raters in 24 FTLD-tau cases (Pick disease = 4, corticobasal degeneration = 9, progressive supranuclear palsy = 5, globular glial tauopathy = 6) including 4 with a mutation in MAPT collected with consent by Sydney Brain Bank. All brain regions and stains assessed proved informative for accurate pathological subtyping, and many neuropathological features were identified as common across the FTLD-tau subtypes. By identifying subtype-specific neuropathological features in the sections selected, 10 independent observers assigned the cases to a FTLD-tau subtype with almost perfect agreement between raters, emphasizing the requirement for the assessment of subtype-specific features for the accurate subtyping of FTLD-tau. This study consolidates current consensus diagnostic criteria for classifying FTLD-tau subtypes with an efficient, simple and accurate approach that can be implemented in future clinicopathological studies.

TDP-43 levels in the brain tissue of ALS cases with and without C9ORF72 or ATXN2 gene expansions.

OBJECTIVE: To assess the amount of phosphorylated and nonphosphorylated TAR DNA-binding protein 43 (TDP-43) in the motor brain regions of cases of amyotrophic lateral sclerosis (ALS) with and without repeat expansions in the ATXN2 or C9ORF72 genes. METHODS: The 45-kDa phosphorylated form of TDP-43 and 43-kDa nonphosphorylated form of TDP-43 were quantified by immunoblot in postmortem brain tissue from the motor cortex, spinal cord, and cerebellar vermis of 23 cases with ALS with repeat expansions in the ATXN2 or C9ORF72 genes and sporadic disease and 10 controls. RESULTS: Significantly greater levels of phosphorylated TDP-43 were identified in the motor cortex of cases with ALS with C9ORF72 expansions, and significantly greater amounts of phosphorylated TDP-43 were found in the spinal cord of cases with ALS with intermediate ATXN2 expansions. In contrast, however, similar levels of nonphosphorylated TDP-43 were found in all 3 regions between ALS groups. CONCLUSION: Despite its central role in the pathogenesis of ALS and the emergence of potential targets to modify its aggregation, TDP-43 levels have not been quantified in pathologically confirmed cases with ALS. The present results demonstrating significant differences in phosphorylated but not nonphosphorylated TDP-43 levels suggest that different posttranslational modifications are involved in the generation of greater pathologic TDP-43 levels identified here in our cohort of cases with genetic expansions. These findings are consistent with emerging studies implicating distinct pathomechanisms in the generation of pathologic TDP-43 in cases with ALS with C9ORF72 or ATXN2 expansions and are of relevance to therapeutic research aimed at reducing pathologic TDP-43 in all or a subset of patients with ALS.

Von Economo Neurons in Behavioral Variant Frontotemporal Dementia with Underlying Alzheimer's Disease.

The selective loss of von Economo neurons has been linked to the behavioral deficits in behavioral variant frontotemporal dementia (bvFTD) but whether these neurons are affected in bvFTD patients with underlying Alzheimer's disease (AD) has yet to be established. The present study assesses the von Economo neurons in pathological AD cases clinically diagnosed with either AD or bvFTD. Our results demonstrate no significant loss of von Economo neurons in all pathological AD cases, irrespective of clinical diagnosis or co-existing Lewy body pathology. These results suggest that the behavioral deficits in patients with clinical bvFTD and underlying pathological AD are not driven by the loss of von Economo neurons.

Coexisting Lewy body disease and clinical parkinsonism in frontotemporal lobar degeneration.

OBJECTIVE: To investigate the prevalence of clinically relevant multiple system atrophy (MSA) and Lewy body disease (LBD) pathologies in a large frontotemporal lobar degeneration (FTLD) cohort to determine if concomitant pathologies underlie the heterogeneity of clinical features. METHODS: All prospectively followed FTLD-tau and FTLD-TDP cases held by the Sydney Brain Bank (n = 126) were screened for coexisting MSA and LBD (Braak ≥ stage IV) pathology. Relevant clinical (including family history) and genetic associations were determined. RESULTS: MSA pathology was not identified in this series. Of the FTLD cohort, 9 cases had coexisting LBD ≥ Braak stage IV and were associated with different FTLD subtypes including Pick disease (n = 2), corticobasal degeneration (n = 2), progressive supranuclear palsy (n = 2), and TDP type A (n = 3). All FTLD-TDP cases with coexisting LBD had mutations in progranulin (n = 2) or an abnormal repeat expansion in C9orf72 (n = 1). All FTLD-tau cases with coexisting LBD were sporadic. The H1H1 MAPT haplotype was found in all cases that could be genotyped (n = 6 of 9). Seven cases presented with a predominant dementia disorder, 3 of which developed parkinsonism. Two cases presented with a movement disorder and developed dementia in their disease course. The age at symptom onset (62 ± 11 years) and disease duration (8 ± 5 years) in FTLD cases with coexisting LBD did not differ from pure FTLD or pure LBD cases in the brain bank. CONCLUSION: Coexisting LBD in FTLD comprises a small proportion of cases but has implications for clinical and neuropathologic diagnoses and the identification of biomarkers.

The underacknowledged PPA-ALS: A unique clinicopathologic subtype with strong heritability.

OBJECTIVE: To assess the incidence, heritability, and neuropathology of primary progressive aphasia (PPA) with amyotrophic lateral sclerosis (ALS) in a large Australian cohort. METHODS: A total of 130 patients with a primary nonfluent variant of PPA (nfvPPA) or semantic variant of PPA (svPPA) were assessed for concomitant ALS and a strong family history of neurodegenerative diseases (Goldman score ≤3). Neuropathologic examination was carried out in 28% (n = 36) of these PPA and PPA-ALS cases that had come to autopsy. RESULTS: ALS was identified in 18% of patients with nfvPPA and 5% of patients with svPPA. PPA-ALS but not PPA was found to have a strong family history. At autopsy, frontotemporal lobar degeneration (FTLD)-TDP was identified in 100% of nfvPPA-ALS cases, 100% of svPPA-ALS cases, 24% of nfvPPA cases, and 78% of svPPA cases. Clinicopathologic assessments revealed a significant association between a strong family history and underlying FTLD-TDP pathology. Pathogenic mutations in known frontotemporal dementia (FTD)/ALS genes were identified in 100% of these familial PPA cases but only 50% of familial PPA-ALS cases, suggesting the involvement of novel genetic variants in this underacknowledged phenotype. CONCLUSION: The present study identified ALS in 12% of a large cohort of patients with nfvPPA and svPPA, which is comparable to the 10%-15% reported in FTD overall, indicating that a third of patients with FTD-ALS will have a predominant language profile. These findings highlight the importance of assessing for ALS in PPA, particularly since this is the only PPA phenotype in which a perfect clinicopathologic association has been reported in to date.

Distinct TDP-43 inclusion morphologies in frontotemporal lobar degeneration with and without amyotrophic lateral sclerosis.

The identification of the TAR DNA-binding protein 43 (TDP-43) as the ubiquitinated cytoplasmic inclusions in frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis (ALS) confirmed that these two diseases share similar mechanisms, likely to be linked to the abnormal hyperphosphorylation, ubiquitination and cleavage of pathological TDP-43. Importantly however, a quantitative analysis of TDP-43 inclusions in predilection cortical regions of FTLD, FTLD-ALS and ALS cases has not been undertaken. The present study set out to assess this and demonstrates that distinct TDP-43 inclusion morphologies exist in the anterior cingulate cortex, but not the motor cortex of FTLD and FTLD-ALS. Specifically, in the anterior cingulate cortex of FTLD cases, significant rounded TDP-43 inclusions and rare circumferential TDP-43 inclusions were identified. In contrast, FTLD-ALS cases revealed significant circumferential TDP-43 inclusions and rare rounded TDP-43 inclusions in the anterior cingulate cortex. Distinct TDP-43 inclusion morphologies in the anterior cingulate cortex of FTLD and FTLD-ALS may be linked to heterogeneity in the ubiquitination of pathological TDP-43 inclusions, with the present study providing evidence to suggest the involvement of distinct pathomechanisms in these two overlapping clinical syndromes.

Assessment of amyloid ß in pathologically confirmed frontotemporal dementia syndromes.

INTRODUCTION: The diagnostic utility of in vivo amyloid ß (Aß) imaging to aid in the clinical distinction between frontotemporal dementia (FTD) and Alzheimer's disease remains unclear without data on the prevalence and severity of Aß in pathologically confirmed FTD syndromes. METHODS: Aß was assessed in 98 autopsy-confirmed FTD and 36 control cases, and the pathological accuracy of 11C-Pittsburgh compound B (PiB)-positron emission tomography imaging was assessed in a subset of FTD cases (n = 15). RESULTS: Aß was identified in a similar proportion of FTD syndromes and age-matched controls and increases with age. Alzheimer's disease pathology was identified in all cases with high PiB retention and in one case with low PiB retention. We further demonstrate a strong regional correlation between volume fraction of histological Aß with PiB standard uptake value ratio scaled to the white matter. DISCUSSION: The present study provides a pathologic reference to assist in the interpretation of in vivo assessments in FTD syndromes.

Cerebellar atrophy in neurodegeneration-a meta-analysis.

INTRODUCTION: The cerebellum has strong cortical and subcortical connectivity, but is rarely taken into account for clinical diagnosis in many neurodegenerative conditions, particularly in the absence of clinical ataxia. The current meta-analysis aims to assess patterns of cerebellar grey matter atrophy in seven neurodegenerative conditions (Alzheimer's disease (AD), Parkinson's disease (PD) and Huntington's disease (HD), frontotemporal dementia (FTD), amyotrophic lateral sclerosis (ALS), multiple system atrophy (MSA), progressive supranuclear palsy (MSP)). METHODS: We carried out a systematic search in PubMed (any date: 14 July 2016) and a hand search of references from pertinent articles according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. The authors were contacted to provide missing coordinate data. Peer-reviewed studies with direct comparison of patient and control groups, and availability of coordinate data of grey matter cerebellar atrophy in patients were included. These coordinates were used in an anatomical likelihood estimation meta-analysis. RESULTS: Across 54 studies, clusters of cerebellar atrophy were found for AD, ALS, FTD, MSA, and PSP. Atrophy patterns were largely disease-specific, with overlap in certain areas of the cerebellar hemisphere, which showed marked atrophy in AD, ALS, FTD and PSP (Crus I/II), and MSA and PSP (lobules I-IV), respectively. Atrophy colocated with cerebellar areas implicated for motor (PSP, MSA) or cognitive symptoms (FTD, ALS, PSP) in the diseases. DISCUSSION: Our findings suggest that cerebellar changes are largely disease-specific and correspond to cortical or subcortical changes in neurodegenerative conditions. High clinical variability in PD and HD samples may explain the absence of findings for consistent grey matter loss across studies. Our results have clinical implications for diagnosis and cerebellar neuroimaging referencing approaches.

von Economo Neuron Density and Thalamus Volumes in Behavioral Deficits in Frontotemporal Dementia Cases with and without a C9ORF72 Repeat Expansion.

BACKGROUND: The early and selective loss of von Economo neurons in the anterior cingulate cortex has been linked to behavioral deficits in frontotemporal dementia (FTD). Importantly, whether these neurons are also targeted in patients with the C9ORF72 repeat expansion has yet to be established. This is of particular interest given the recent evidence highlighting the thalamus rather than anterior cingulate cortex as a region of significant degeneration in patients with the C9ORF72 repeat expansion. OBJECTIVE: To assess the von Economo neuron density and thalamus volumes in behavioral variant FTD (bvFTD) cases with the C9ORF72 repeat expansion, sporadic bvFTD, sporadic ALS, and controls. METHODS: Volumetric and quantitative cell counting methods were employed to assess the von Economo neuron density and thalamus volumes in 37 pathologically-confirmed cases comprised of patients with bvFTD (n = 13) cases with the C9ORF72 repeat expansion (62% with psychosis), sporadic bvFTD (n = 8), sporadic amyotrophic lateral sclerosis (n = 7) and controls (n = 9). RESULTS: von Economo neuron density was significantly reduced in sporadic bvFTD cases only. Thalamus degeneration was identified only in bvFTD cases with the C9ORF72 repeat expansion, and to a similar extent in cases with and without psychosis. No significant difference in von Economo neuron density or thalamus degeneration was seen between bvFTD cases with or without the C9ORF72 repeat expansion. CONCLUSION: The present histological findings converge with neuroimaging results to corroborate the anterior cingulate cortex as a core region involved in sporadic bvFTD, and the thalamus as a major region targeted in patients with the C9ORF72 expansion.

ALS/FTLD: experimental models and reality.

Amyotrophic lateral sclerosis is characterised by a loss of upper and lower motor neurons and characteristic muscle weakness and wasting, the most common form being sporadic disease with neuronal inclusions containing the tar DNA-binding protein 43 (TDP-43). Frontotemporal lobar degeneration is characterised by atrophy of the frontal and/or temporal lobes, the most common clinical form being the behavioural variant, in which neuronal inclusions containing either TDP-43 or 3-repeat tau are most prevalent. Although the genetic mutations associated with these diseases have allowed various experimental models to be developed, the initial genetic forms identified remain the most common models employed to date. It is now known that these first models faithfully recapitulate only some aspects of these diseases and do not represent the majority of cases or the most common overlapping pathologies. Newer models targeting the main molecular pathologies are still rare and in some instances, lack significant aspects of the molecular pathology. However, these diseases are complex and multigenic, indicating that experimental models may need to be targeted to different disease aspects. This would allow information to be gleaned from a variety of different yet relevant models, each of which has the capacity to capture a certain aspect of the disease, and together will enable a more complete understanding of these complex and multi-layered diseases.

TDP-43 in the hypoglossal nucleus identifies amyotrophic lateral sclerosis in behavioral variant frontotemporal dementia.

The hypoglossal nucleus was recently identified as a key brain region in which the presence of TDP-43 pathology could accurately discriminate TDP-43 proteinopathy cases with clinical amyotrophic lateral sclerosis (ALS). The objective of the present study was to assess the hypoglossal nucleus in behavioral variant frontotemporal dementia (bvFTD), and determine whether TDP-43 in this region is associated with clinical ALS. Twenty-nine cases with neuropathological FTLD-TDP and clinical bvFTD that had not been previously assessed for hypoglossal TDP-43 pathology were included in this study. Of these 29 cases, 41% (n=12) had a dual diagnosis of bvFTD-ALS at presentation, all 100% (n=12) of which demonstrated hypoglossal TDP-43 pathology. Of the 59% (n=17) cohort that presented with pure bvFTD, 35% (n=6) were identified with hypoglossal TDP-43 pathology. Review of the case files of all pure bvFTD cases revealed evidence of possible or probable ALS in 5 of the 6 hypoglossal-positive cases (83%) towards the end of disease, and this was absent from all cases without such pathology. In conclusion, the present study validates grading the presence of TDP-43 in the hypoglossal nucleus for the pathological identification of bvFTD cases with clinical ALS, and extends this to include the identification of cases with possible ALS at end-stage.

Cerebellar neuronal loss in amyotrophic lateral sclerosis cases with ATXN2 intermediate repeat expansions.

OBJECTIVE: Despite evidence suggesting that the cerebellum may be targeted in amyotrophic lateral sclerosis (ALS), particularly in cases with repeat expansions in the ATXN2 and C9ORF72 genes, the integrity of cerebellar neurons has yet to be examined. The present study undertakes a histopathological analysis to assess the impact of these repeat expansions on cerebellar neurons and determine whether similar cerebellar pathology occurs in sporadic disease. METHODS: Purkinje and granule cells were quantified in the vermis and lateral cerebellar hemispheres of ALS cases with repeat expansions in the ATXN2 and C9ORF72 genes, sporadic disease, and sporadic progressive muscular atrophy with only lower motor neuron degeneration. RESULTS: ALS cases with intermediate repeat expansions in the ATXN2 gene demonstrate a significant loss in Purkinje cells in the cerebellar vermis only. Despite ALS cases with expansions in the C9ORF72 gene having the highest burden of inclusion pathology, no neuronal loss was observed in this group. Neuronal numbers were also unchanged in sporadic ALS and sporadic PMA cases. INTERPRETATION: The present study has established a selective loss of Purkinje cells in the cerebellar vermis of ALS cases with intermediate repeat expansions in the ATXN2 gene, suggesting a divergent pathogenic mechanism independent of upper and lower motor neuron degeneration in ALS. We discuss these findings in the context of large repeat expansions in ATXN2 and spinocerebellar ataxia type 2, providing evidence that intermediate repeats in ATXN2 cause significant, albeit less substantial, spinocerebellar damage compared with longer repeats in ATXN2.