Mutated Toll-like receptor 9 increases Alzheimer's disease risk by compromising innate immunity protection.

The development of Alzheimer's disease (AD) involves central and peripheral immune deregulation. Gene identification and studies of AD genetic variants of peripheral immune components may aid understanding of peripheral-central immune crosstalk and facilitate new opportunities for therapeutic intervention. In this study, we have identified in a Flanders-Belgian family a novel variant p.E317D in the Toll-like receptor 9 gene (TLR9), co-segregating with EOAD in an autosomal dominant manner. In human, TLR9 is an essential innate and adaptive immune component predominantly expressed in peripheral immune cells. The p.E317D variant caused 50% reduction in TLR9 activation in the NF-κB luciferase assay suggesting that p.E317D is a loss-of-function mutation. Cytokine profiling of human PBMCs upon TLR9 activation revealed a predominantly anti-inflammatory response in contrast to the inflammatory responses from TLR7/8 activation. The cytokines released upon TLR9 activation suppressed inflammation and promoted phagocytosis of Aß42 oligomers in human iPSC-derived microglia. Transcriptome analysis identified upregulation of AXL, RUBICON and associated signaling pathways, which may underline the effects of TLR9 signaling-induced cytokines in regulating the inflammatory status and phagocytic property of microglia. Our data suggest a protective role of TLR9 signaling in AD pathogenesis, and we propose that TLR9 loss-of-function may disrupt a peripheral-central immune crosstalk that promotes dampening of inflammation and clearance of toxic protein species, leading to the build-up of neuroinflammation and pathogenic protein aggregates in AD development.

Biomarker A+T-: is this Alzheimer's disease or not? A combined CSF and pathology study.

The biological definition of Alzheimer's disease using CSF biomarkers requires abnormal levels of both amyloid (A) and tau (T). However, biomarkers and corresponding cutoffs may not always reflect the presence or absence of pathology. Previous studies suggest that up to 32% of individuals with autopsy-confirmed Alzheimer's disease show normal CSF p-tau levels in vivo, but these studies are sparse and had small sample sizes. Therefore, in three independent autopsy cohorts, we studied whether or not CSF A+T- excluded Alzheimer's disease based on autopsy. We included 215 individuals, for whom ante-mortem CSF collection and autopsy had been performed, from three cohorts: (i) the Amsterdam Dementia Cohort (ADC) [n = 80, 37 (46%) Alzheimer's disease at autopsy, time between CSF collection and death 4.5 ± 2.9 years]; (ii) the Antwerp Dementia Cohort (DEM) [n = 92, 84 (91%) Alzheimer's disease at autopsy, time CSF collection to death 1.7 ± 2.3 years]; and (iii) the Alzheimer's Disease Neuroimaging Initiative (ADNI) [n = 43, 31 (72%) Alzheimer's disease at autopsy, time CSF collection to death 5.1 ± 2.5 years]. Biomarker profiles were based on dichotomized CSF Aß1-42 and p-tau levels. The accuracy of CSF AT profiles to detect autopsy-confirmed Alzheimer's disease was assessed. Lastly, we investigated whether the concordance of AT profiles with autopsy diagnosis improved when CSF was collected closer to death in 9 (10%) DEM and 30 (70%) ADNI individuals with repeated CSF measurements available. In total, 50-73% of A+T- individuals and 100% of A+T+ individuals had Alzheimer's disease at autopsy. Amyloid status showed the highest accuracy to detect autopsy-confirmed Alzheimer's disease (accuracy, sensitivity and specificity in the ADC: 88%, 92% and 84%; in the DEM: 87%, 94% and 12%; and in the ADNI cohort: 86%, 90% and 75%, respectively). The addition of CSF p-tau did not further improve these estimates. We observed no differences in demographics or degree of Alzheimer's disease neuropathology between A+T- and A+T+ individuals with autopsy-confirmed Alzheimer's disease. All individuals with repeated CSF measurements remained stable in Aß1-42 status during follow-up. None of the Alzheimer's disease individuals with a normal p-tau status changed to abnormal; however, four (44%) DEM individuals and two (7%) ADNI individuals changed from abnormal to normal p-tau status over time, and all had Alzheimer's disease at autopsy. In summary, we found that up to 73% of A+T- individuals had Alzheimer's disease at autopsy. This should be taken into account in both research and clinical settings.

Patients carrying the mutation p.R406W in MAPT present with non-conforming phenotypic spectrum.

The missense mutation p.R406W in microtubule-associated protein tau leads to frontotemporal lobar degeneration with an amnestic, Alzheimer's disease-like phenotype with an autosomal dominant pattern of inheritance. In 2003, we described the pedigree of a Belgian family, labelled ADG, with 28 p.R406W patients. Over 18 years follow-up, we extended the family with 10 p.R406W carriers and provided an in-depth clinical description of the patients. Additionally, genetic screening was used to identify p.R406W carriers in Belgian cohorts of frontotemporal dementia and Alzheimer's disease patients and to calculate p.R406W frequency. In the frontotemporal dementia cohort, we found four p.R406W carriers (n = 647, 0.62%) and three in the Alzheimer's disease cohort (n = 1134, 0.26%). Haplotype sharing analysis showed evidence of a shared haplotype suggesting that they are descendants of a common ancestor. Of the p.R406W patients, we describe characteristics of neuropsychological, imaging and fluid biomarkers as well as neuropathologic examination. Intriguingly, the phenotypic spectrum among the p.R406W patients ranged from typical behavioural variant frontotemporal dementia to clinical Alzheimer's disease, based on CSF biomarker analysis and amyloid PET scan. Heterogeneous overlap syndromes existed in between, with highly common neuropsychiatric symptoms like disinhibition and aggressiveness, which occurred in 100% of frontotemporal dementia and 58% of clinical Alzheimer's disease patients. This was also the case for memory problems, 89% in frontotemporal dementia and 100% in clinical Alzheimer's disease patients. Median age at death was significantly lower in patients with frontotemporal dementia (68 years) compared to clinical Alzheimer's disease patients (79 years), although the sizes of the sub-cohorts are limited and do not allow prognostic predictions. Post-mortem brain analysis of one p.R406W patient with behavioural variant frontotemporal dementia revealed frontotemporal lobar degeneration with tau pathology. Notably, neuropathological investigation showed only 3R tau isoforms in the absence of 4R tau reactivity, an unusual finding in microtubule-associated protein tau-related frontotemporal lobar degeneration. No traces of amyloid pathology were present. Prevalence of the p.R406W mutation was relatively high in both frontotemporal dementia and Alzheimer's disease Belgian patient cohorts. These findings grant new insights into genotype-phenotype correlations of p.R406W carriers. They may help in further unravelling of the pathophysiology of this tauopathy and in facilitating the identification of patients with p.R406W-related frontotemporal lobar degeneration, both in clinical diagnostic and research settings.

Associating Alzheimer's disease pathology with its cerebrospinal fluid biomarkers.

Alzheimer's disease CSF biomarkers 42 amino acid long amyloid-ß peptide (Aß1-42), total tau protein (T-tau), and tau protein phosphorylated at threonine 181 (P-tau181) are considered surrogate biomarkers of Alzheimer's disease pathology, and significantly improve diagnostic accuracy. Their ability to reflect neuropathological changes later in the disease course is not well characterized. This study aimed to assess the potential of CSF biomarkers measured in mid to late stage Alzheimer's disease to reflect post-mortem neuropathological changes. Individuals were selected from two autopsy cohorts of Alzheimer's disease patients in Antwerp and Amsterdam. Neuropathological diagnosis was performed according to the updated consensus National Institute on Aging-Alzheimer's Association guidelines, which includes quantification of amyloid-ß plaque, neurofibrillary tangle, and neuritic plaque load. CSF samples were analysed for Aß1-42, T-tau, and P-tau181 by ELISA. One hundred and fourteen cases of pure definite Alzheimer's disease were included in the study (mean age 74 years, disease duration 6 years at CSF sampling, 50% females). Median interval between CSF sampling and death was 1 year. We found no association between Aß1-42 and Alzheimer's disease neuropathological change profile. In contrast, an association of P-tau181 and T-tau with Alzheimer's disease neuropathological change profile was observed. P-tau181 was associated with all three individual Montine scores, and the associations became stronger and more significant as the interval between lumbar puncture and death increased. T-tau was also associated with all three Montine scores, but in individuals with longer intervals from lumbar puncture to death only. Stratification of the cohort according to APOE ε4 carrier status revealed that the associations applied mostly to APOE ε4 non-carriers. Our data suggest that similar to what has been reported for Aß1-42, plateau levels of P-tau181 and T-tau are reached during the disease course, albeit at later disease stages, reducing the potential of tau biomarkers to monitor Alzheimer's disease pathology as the disease progresses. As a consequence, CSF biomarkers, which are performant for clinical diagnosis of early Alzheimer's disease, may not be well suited for staging or monitoring Alzheimer's disease pathology as it progresses through later stages.

Mutations in RNU7-1 Weaken Secondary RNA Structure, Induce MCP-1 and CXCL10 in CSF, and Result in Aicardi-Goutières Syndrome with Severe End-Organ Involvement.

BACKGROUND: Aicardi-Goutières syndrome (AGS) is a type I interferonopathy usually characterized by early-onset neurologic regression. Biallelic mutations in LSM11 and RNU7-1, components of the U7 small nuclear ribonucleoprotein (snRNP) complex, have been identified in a limited number of genetically unexplained AGS cases. Impairment of U7 snRNP function results in misprocessing of replication-dependent histone (RDH) pre-mRNA and disturbance of histone occupancy of nuclear DNA, ultimately driving cGAS-dependent type I interferon (IFN-I) release. OBJECTIVE: We performed a clinical, genetic, and immunological workup of 3 unrelated patients with uncharacterized AGS. METHODS: Whole exome sequencing (WES) and targeted Sanger sequencing of RNU7-1 were performed. Primary fibroblasts were used for mechanistic studies. IFN-I signature and STAT1/2 phosphorylation were assessed in peripheral blood. Cytokines were profiled on serum and cerebrospinal fluid (CSF). Histopathology was examined on brain and kidney tissue. RESULTS: Sequencing revealed compound heterozygous RNU7-1 mutations, resulting in impaired RDH pre-mRNA processing. The 3' stem-loop mutations reduced stability of the secondary U7 snRNA structure. A discrete IFN-I signature in peripheral blood was paralleled by MCP-1 (CCL2) and CXCL10 upregulation in CSF. Histopathological analysis of the kidney showed thrombotic microangiopathy. We observed dysregulated STAT phosphorylation upon cytokine stimulation. Clinical overview of all reported patients with RNU7-1-related disease revealed high mortality and high incidence of organ involvement compared to other AGS genotypes. CONCLUSIONS: Targeted RNU7-1 sequencing is recommended in genetically unexplained AGS cases. CSF cytokine profiling represents an additional diagnostic tool to identify aberrant IFN-I signaling. Clinical follow-up of RNU7-1-mutated patients should include screening for severe end-organ involvement including liver disease and nephropathy.

Family-based exome sequencing identifies RBM45 as a possible candidate gene for frontotemporal dementia and amyotrophic lateral sclerosis.

Neurodegenerative disorders like frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) are pathologically characterized by toxic protein deposition in the cytoplasm or nucleus of affected neurons and glial cells. Many of these aggregated proteins belong to the class of RNA binding proteins (RBP), and, when mutated, account for a significant subset of familial ALS and FTD cases. Here, we present first genetic evidence for the RBP gene RBM45 in the FTD-ALS spectrum. RBM45 shows many parallels with other FTD-ALS associated genes and proteins. Multiple lines of evidence have demonstrated that RBM45 is an RBP that, upon mutation, redistributes to the cytoplasm where it co-aggregates with other RBPs into cytoplasmic stress granules (SG), evolving to persistent toxic TDP-43 immunoreactive inclusions. Exome sequencing in two affected first cousins of a heavily affected early-onset dementia family listed a number of candidate genes. The gene with the highest pathogenicity score was the RBP gene RBM45. In the family, the RBM45 Arg183* nonsense mutation co-segregated in both affected cousins. Validation in an unrelated patient (n = 548) / control (n = 734) cohort identified an additional RBM45 Arg183* carrier with bvFTD on a shared 4 Mb haplotype. Transcript and protein expression analysis demonstrated loss of nuclear RBM45, suggestive of a loss-of-function disease mechanism. Further, two more ultra-rare VUS, one in the nuclear localization signal (NLS, p.Lys456Arg) in an ALS patient and one in the intrinsically disordered homo-oligomer assembly (HOA) domain (p.Arg314Gln) in a patient with nfvPPA were detected. Our findings suggest that the pathomechanisms linking RBM45 with FTD and ALS may be related to its loss of nuclear function as a mediator of mRNA splicing, cytoplasmic retention or its inability to form homo-oligomers, leading to aggregate formation with trapping of other RBPs including TDP-43, which may accumulate into persisted TDP-43 inclusions.

A case series of verbal semantic processing in primary progressive aphasia: Evidence from the N400 effect.

BACKGROUND: The semantic variant of primary progressive aphasia (PPA) is typically associated with a loss of semantic knowledge. Research on the semantic processing in the other clinical variants of PPA is, however, rather sparse and limited to off-line behavioural studies. AIMS: This study aimed to investigate verbal semantic processing in patients with the three variants of PPA by the event-related potential technique. The presence, latency, amplitude and/or topographic distribution of the N400 effect may be helpful in the diagnosis of PPA and its clinical variants and it provides temporal information about semantic processing (disturbances) in the three variants of PPA. METHODS & PROCEDURES: The N400 effect was studied by a categorical word-priming paradigm and a semantic-anomaly paradigm at sentence level in eight persons with PPA(-plus) and 30 age-matched healthy controls. The mean amplitudes and onset latencies of the N400 effect were compared between each patient and the control group by two methods that are applicable in clinical practice, namely visual inspection and Z-scores. OUTCOMES & RESULTS: The N400 effect elicited by the categorical-priming paradigm was only present in the two patients with the non-fluent variant of PPA. This effect was absent in the two patients with the semantic variant(-plus), two patients with the logopenic variant(-plus), one patient with the non-fluent variant-plus, and the patient with PPA not otherwise specified. The results of the N400 effect elicited by the semantic-anomaly task at the sentence level were variable, but differences in the presence, mean amplitudes, onset latencies and/or topographic distributions of the effect were found in all patients with PPA(-plus) in comparison with the control group. CONCLUSIONS & IMPLICATIONS: The results of our study showed that the evaluation of the N400 effect might have an added value in the diagnostic process of PPA in general and in the differentiation of patients with the non-fluent variant from patients with the logopenic and semantic variants. Furthermore, our results indicate the presence of difficulties with retrieving stored semantic knowledge or semantic integration of a word in the preceding context in patients with the three variants of PPA. These findings might help the speech-language pathologist in determining individualized therapy goals and indicate that it might be helpful to focus on verbal semantic processing in language therapy in patients with the three variants of PPA and not only in patients with the semantic variant. WHAT THIS PAPER ADDS: What is already known on the subject The semantic variant of PPA is characterized by an impaired object knowledge and single-word comprehension and these functions are relatively spared in the non-fluent and logopenic variants following the guidelines of Gorno-Tempini et al. (2011). Research on the semantic processing in patients with the non-fluent and logopenic variant is, however, rather sparse and limited to off-line behavioural studies. Only four group studies investigated verbal semantic processing by the N400 effect, and these studies indicate disturbances in the three variants of PPA. What this paper adds to existing knowledge Our results indicate the presence of difficulties with retrieving stored semantic knowledge or semantic integration of a word in the preceding context during a semantic-priming paradigm in patients with the semantic and logopenic variants of PPA and during a semantic-anomaly task at the sentence level in patients with the three variants of PPA. What are the potential or actual clinical implications of this work? The results of our study showed that the evaluation of the N400 effect might have an added value in the diagnostic process of PPA in general and in the differentiation of patients with the non-fluent variant from patients with the logopenic and semantic variants. The evaluation of the N400 effect might also help the speech-language pathologist in determining individualized therapy goals and indicate that it might be helpful to focus on verbal semantic processing in language therapy in patients with the three variants of PPA and not only in patients with the semantic variant.

Loss of DPP6 in neurodegenerative dementia: a genetic player in the dysfunction of neuronal excitability.

Emerging evidence suggested a converging mechanism in neurodegenerative brain diseases (NBD) involving early neuronal network dysfunctions and alterations in the homeostasis of neuronal firing as culprits of neurodegeneration. In this study, we used paired-end short-read and direct long-read whole genome sequencing to investigate an unresolved autosomal dominant dementia family significantly linked to 7q36. We identified and validated a chromosomal inversion of ca. 4 Mb, segregating on the disease haplotype and disrupting the coding sequence of dipeptidyl-peptidase 6 gene (DPP6). DPP6 resequencing identified significantly more rare variants-nonsense, frameshift, and missense-in early-onset Alzheimer's disease (EOAD, p value = 0.03, OR = 2.21 95% CI 1.05-4.82) and frontotemporal dementia (FTD, p = 0.006, OR = 2.59, 95% CI 1.28-5.49) patient cohorts. DPP6 is a type II transmembrane protein with a highly structured extracellular domain and is mainly expressed in brain, where it binds to the potassium channel Kv4.2 enhancing its expression, regulating its gating properties and controlling the dendritic excitability of hippocampal neurons. Using in vitro modeling, we showed that the missense variants found in patients destabilize DPP6 and reduce its membrane expression (p < 0.001 and p < 0.0001) leading to a loss of protein. Reduced DPP6 and/or Kv4.2 expression was also detected in brain tissue of missense variant carriers. Loss of DPP6 is known to cause neuronal hyperexcitability and behavioral alterations in Dpp6-KO mice. Taken together, the results of our genomic, genetic, expression and modeling analyses, provided direct evidence supporting the involvement of DPP6 loss in dementia. We propose that loss of function variants have a higher penetrance and disease impact, whereas the missense variants have a variable risk contribution to disease that can vary from high to low penetrance. Our findings of DPP6, as novel gene in dementia, strengthen the involvement of neuronal hyperexcitability and alteration in the homeostasis of neuronal firing as a disease mechanism to further investigate.

Distinct [18F]THK5351 binding patterns in primary progressive aphasia variants.

PURPOSE: To assess the binding of the PET tracer [18F]THK5351 in patients with different primary progressive aphasia (PPA) variants and its correlation with clinical deficits. The majority of patients with nonfluent variant (NFV) and logopenic variant (LV) PPA have underlying tauopathy of the frontotemporal lobar or Alzheimer disease type, respectively, while patients with the semantic variant (SV) have predominantly transactive response DNA binding protein 43-kDa pathology. METHODS: The study included 20 PPA patients consecutively recruited through a memory clinic (12 NFV, 5 SV, 3 LV), and 20 healthy controls. All participants received an extensive neurolinguistic assessment, magnetic resonance imaging and amyloid biomarker tests. [18F]THK5351 binding patterns were assessed on standardized uptake value ratio (SUVR) images with the cerebellar grey matter as the reference using statistical parametric mapping. Whole-brain voxel-wise regression analysis was performed to evaluate the association between [18F]THK5351 SUVR images and neurolinguistic scores. Analyses were performed with and without partial volume correction. RESULTS: Patients with NFV showed increased binding in the supplementary motor area, left premotor cortex, thalamus, basal ganglia and midbrain compared with controls and patients with SV. Patients with SV had increased binding in the temporal lobes bilaterally and in the right ventromedial frontal cortex compared with controls and patients with NFV. The whole-brain voxel-wise regression analysis revealed a correlation between agrammatism and motor speech impairment, and [18F]THK5351 binding in the left supplementary motor area and left postcentral gyrus. Analysis of [18F]THK5351 scans without partial volume correction revealed similar results. CONCLUSION: [18F]THK5351 imaging shows a topography closely matching the anatomical distribution of predicted underlying pathology characteristic of NFV and SV PPA. [18F]THK5351 binding correlates with the severity of clinical impairment.

Motor neuron degeneration in spastic paraplegia 11 mimics amyotrophic lateral sclerosis lesions.

The most common form of autosomal recessive hereditary spastic paraplegia is caused by mutations in the SPG11/KIAA1840 gene on chromosome 15q. The nature of the vast majority of SPG11 mutations found to date suggests a loss-of-function mechanism of the encoded protein, spatacsin. The SPG11 phenotype is, in most cases, characterized by a progressive spasticity with neuropathy, cognitive impairment and a thin corpus callosum on brain MRI. Full neuropathological characterization has not been reported to date despite the description of >100 SPG11 mutations. We describe here the clinical and pathological features observed in two unrelated females, members of genetically ascertained SPG11 families originating from Belgium and Italy, respectively. We confirm the presence of lesions of motor tracts in medulla oblongata and spinal cord associated with other lesions of the central nervous system. Interestingly, we report for the first time pathological hallmarks of SPG11 in neurons that include intracytoplasmic granular lysosome-like structures mainly in supratentorial areas, and others in subtentorial areas that are partially reminiscent of those observed in amyotrophic lateral sclerosis, such as ubiquitin and p62 aggregates, except that they are never labelled with anti-TDP-43 or anti-cystatin C. The neuropathological overlap with amyotrophic lateral sclerosis, associated with some shared clinical manifestations, opens up new fields of investigation in the physiopathological continuum of motor neuron degeneration.

Clinical features of TBK1 carriers compared with C9orf72, GRN and non-mutation carriers in a Belgian cohort.

We identified in a cohort of patients with frontotemporal dementia (n = 481) or amyotrophic lateral sclerosis (n = 147), 10 index patients carrying a TBK1 loss of function mutation reducing TBK1 expression by 50%. Here, we describe the clinical and pathological characteristics of the 10 index patients and six of their affected relatives carrying a TBK1 mutation. Six TBK1 carriers were diagnosed with frontotemporal dementia, seven with amyotrophic lateral sclerosis, one with both clinical phenotypes and two with dementia unspecified. The mean age at onset of all 16 TBK1 carriers was 62.1 ± 8.9 years (range 41-73) with a mean disease duration of 4.7 ± 4.5 years (range 1-13). TBK1 carriers with amyotrophic lateral sclerosis had shorter disease duration than carriers with frontotemporal dementia. Six of seven TBK1 carriers were diagnosed with the behavioural variant of frontotemporal dementia, presenting predominantly as disinhibition. Memory loss was an important associated symptom in the initial phase of the disease in all but one of the carriers with frontotemporal dementia. Three of the patients with amyotrophic lateral sclerosis exhibited pronounced upper motor neuron symptoms. Overall, neuroimaging displayed widespread atrophy, both symmetric and asymmetric. Brain perfusion single-photon emission computed tomography or fluorodeoxyglucose-positron emission tomography showed asymmetric and predominantly frontotemporal involvement. Neuropathology in two patients demonstrated TDP-43 type B pathology. Further, we compared genotype-phenotype data of TBK1 carriers with frontotemporal dementia (n = 7), with those of frontotemporal dementia patients with a C9orf72 repeat expansion (n = 65) or a GRN mutation (n = 52) and with frontotemporal dementia patients (n = 259) negative for mutations in currently known causal genes. TBK1 carriers with frontotemporal dementia had a later age at onset (63.3 years) than C9orf72 carriers (54.3 years) (P = 0.019). In clear contrast with TBK1 carriers, GRN carriers were more often diagnosed with the language variant than the behavioural variant, and presented in case of the diagnosis of behavioural variant, more often than TBK1 carriers with apathy as the predominant characteristic (P = 0.004). Also, TBK1 carriers exhibited more often extrapyramidal symptoms than C9orf72 carriers (P = 0.038). In conclusion, our study identified clinical differences between the TBK1, C9orf72 and GRN carriers, which allows us to formulate guidelines for genetic diagnosis. After a negative result for C9orf72, patients with both frontotemporal dementia and amyotrophic lateral sclerosis should be tested first for mutations in TBK1. Specifically in frontotemporal dementia patients with early memory difficulties, a relatively late age at onset or extrapyramidal symptoms, screening for TBK1 mutations should be considered.

Rare mutations in SQSTM1 modify susceptibility to frontotemporal lobar degeneration.

Mutations in the gene coding for Sequestosome 1 (SQSTM1) have been genetically associated with amyotrophic lateral sclerosis (ALS) and Paget disease of bone. In the present study, we analyzed the SQSTM1 coding sequence for mutations in an extended cohort of 1,808 patients with frontotemporal lobar degeneration (FTLD), ascertained within the European Early-Onset Dementia consortium. As control dataset, we sequenced 1,625 European control individuals and analyzed whole-exome sequence data of 2,274 German individuals (total n = 3,899). Association of rare SQSTM1 mutations was calculated in a meta-analysis of 4,332 FTLD and 10,240 control alleles. We identified 25 coding variants in FTLD patients of which 10 have not been described. Fifteen mutations were absent in the control individuals (carrier frequency <0.00026) whilst the others were rare in both patients and control individuals. When pooling all variants with a minor allele frequency <0.01, an overall frequency of 3.2 % was calculated in patients. Rare variant association analysis between patients and controls showed no difference over the whole protein, but suggested that rare mutations clustering in the UBA domain of SQSTM1 may influence disease susceptibility by doubling the risk for FTLD (RR = 2.18 [95 % CI 1.24-3.85]; corrected p value = 0.042). Detailed histopathology demonstrated that mutations in SQSTM1 associate with widespread neuronal and glial phospho-TDP-43 pathology. With this study, we provide further evidence for a putative role of rare mutations in SQSTM1 in the genetic etiology of FTLD and showed that, comparable to other FTLD/ALS genes, SQSTM1 mutations are associated with TDP-43 pathology.

Pooled analysis of patients with inherited prion disease caused by two- to twelve-octapeptide repeat insertions in the prion protein gene (PRNP).

Inherited prion diseases caused by two- to twelve-octapeptide repeat insertions (OPRIs) in the prion protein gene (PRNP) show significant clinical heterogeneity. This study describes a family with two new cases with a 4-OPRI mutation and two asymptomatic mutation carriers. The pooled analysis summarizes all cases reported in the literature to date and describes the relation between survival, age of onset, number of OPRI and codon 129 polymorphism. MEDLINE and Google Scholar were queried from database inception up to December 31, 2022. Age of onset was compared per number of OPRI and per codon 129 polymorphism using the Kruskal-Wallis and Wilcoxon-Mann-Whitney tests, respectively. Disease duration was modeled non-parametrically by a Kaplan-Meier model and semi-parametrically by a Cox model. This study comprised 164 patients. Lower number of OPRI and presence of valine (cis-V) versus methionine (cis-M) on codon 129 were associated with an older age of onset (P < 0.001 and P = 0.025, respectively) and shorter disease duration (P < 0.001 and P = 0.003, respectively). Within patients with 5- or more OPRI codon cis-V remained significantly associated with a shorter disease duration. Codon 129 homozygosity versus heterozygosity was not significantly associated with age of onset or disease duration (P = 0.076 and P = 0.409, respectively). This study summarized the largest cohort of patients with two- to twelve-OPRI to date. Lower number of OPRI and codon 129 cis-V is associated with an older age of onset and shorter disease duration, while homozygosity or heterozygosity on codon 129 was not.

Brain age as a biomarker for pathological versus healthy ageing - a REMEMBER study.

OBJECTIVES: This study aimed to evaluate the potential clinical value of a new brain age prediction model as a single interpretable variable representing the condition of our brain. Among many clinical use cases, brain age could be a novel outcome measure to assess the preventive effect of life-style interventions. METHODS: The REMEMBER study population (N = 742) consisted of cognitively healthy (HC,N = 91), subjective cognitive decline (SCD,N = 65), mild cognitive impairment (MCI,N = 319) and AD dementia (ADD,N = 267) subjects. Automated brain volumetry of global, cortical, and subcortical brain structures computed by the CE-labeled and FDA-cleared software icobrain dm (dementia) was retrospectively extracted from T1-weighted MRI sequences that were acquired during clinical routine at participating memory clinics from the Belgian Dementia Council. The volumetric features, along with sex, were combined into a weighted sum using a linear model, and were used to predict 'brain age' and 'brain predicted age difference' (BPAD = brain age-chronological age) for every subject. RESULTS: MCI and ADD patients showed an increased brain age compared to their chronological age. Overall, brain age outperformed BPAD and chronological age in terms of classification accuracy across the AD spectrum. There was a weak-to-moderate correlation between total MMSE score and both brain age (r = -0.38,p < .001) and BPAD (r = -0.26,p < .001). Noticeable trends, but no significant correlations, were found between BPAD and incidence of conversion from MCI to ADD, nor between BPAD and conversion time from MCI to ADD. BPAD was increased in heavy alcohol drinkers compared to non-/sporadic (p = .014) and moderate (p = .040) drinkers. CONCLUSIONS: Brain age and associated BPAD have the potential to serve as indicators for, and to evaluate the impact of lifestyle modifications or interventions on, brain health.

A pan-European study of the C9orf72 repeat associated with FTLD: geographic prevalence, genomic instability, and intermediate repeats.

We assessed the geographical distribution of C9orf72 G(4) C(2) expansions in a pan-European frontotemporal lobar degeneration (FTLD) cohort (n = 1,205), ascertained by the European Early-Onset Dementia (EOD) consortium. Next, we performed a meta-analysis of our data and that of other European studies, together 2,668 patients from 15 Western European countries. The frequency of the C9orf72 expansions in Western Europe was 9.98% in overall FTLD, with 18.52% in familial, and 6.26% in sporadic FTLD patients. Outliers were Finland and Sweden with overall frequencies of respectively 29.33% and 20.73%, but also Spain with 25.49%. In contrast, prevalence in Germany was limited to 4.82%. In addition, we studied the role of intermediate repeats (7-24 repeat units), which are strongly correlated with the risk haplotype, on disease and C9orf72 expression. In vitro reporter gene expression studies demonstrated significantly decreased transcriptional activity of C9orf72 with increasing number of normal repeat units, indicating that intermediate repeats might act as predisposing alleles and in favor of the loss-of-function disease mechanism. Further, we observed a significantly increased frequency of short indels in the GC-rich low complexity sequence adjacent to the G(4) C(2) repeat in C9orf72 expansion carriers (P < 0.001) with the most common indel creating one long contiguous imperfect G(4) C(2) repeat, which is likely more prone to replication slippage and pathological expansion.

Rare exonic variant affects GRN splicing and contributes to frontotemporal lobar degeneration.

Heterozygous loss-of-function (LOF) mutations in the progranulin gene (GRN) cause frontotemporal lobar degeneration (FTLD) by a mechanism of haploinsufficiency. For most missense mutations, the contribution to FTLD is however unclear. We studied the pathogenicity of rare GRN missense mutations using patient biomaterials. We identified a new mutation in GRN, c.1178 A>C, in a patient with a diagnosis of primary progressive aphasia. Neuropathological examination of autopsied brain showed FTLD with TAR DNA-binding protein 43 (FTLD-TDP) type A pathology with concomitant Alzheimer's disease pathology. Serum progranulin protein levels were reduced to levels comparable to known LOF mutations. The mutation is in the last codon of exon 10, in the splice donor sequence. Our data provide evidence that the mutation leads to aberrant splicing, resulting in a frameshift (p.(Glu393AlafsTer31)) and consequently nonsense-mediated mRNA decay. Our finding demonstrates that carefully examining sequencing data around splice sites is needed since this mutation was annotated as a missense mutation. Unraveling the pathogenicity of variants of unknown significance is important for clinical diagnosis and genetic counseling.

CSF proteome profiling reveals biomarkers to discriminate dementia with Lewy bodies from Alzheimer´s disease.

Diagnosis of dementia with Lewy bodies (DLB) is challenging and specific biofluid biomarkers are highly needed. We employed proximity extension-based assays to measure 665 proteins in the cerebrospinal fluid (CSF) from patients with DLB (n = 109), Alzheimer´s disease (AD, n = 235) and cognitively unimpaired controls (n = 190). We identified over 50 CSF proteins dysregulated in DLB, enriched in myelination processes among others. The dopamine biosynthesis enzyme DDC was the strongest dysregulated protein, and could efficiently discriminate DLB from controls and AD (AUC:0.91 and 0.81 respectively). Classification modeling unveiled a 7-CSF biomarker panel that better discriminate DLB from AD (AUC:0.93). A custom multiplex panel for six of these markers (DDC, CRH, MMP-3, ABL1, MMP-10, THOP1) was developed and validated in independent cohorts, including an AD and DLB autopsy cohort. This DLB CSF proteome study identifies DLB-specific protein changes and translates these findings to a practicable biomarker panel that accurately identifies DLB patients, providing promising diagnostic and clinical trial testing opportunities.

The genetics and neuropathology of frontotemporal lobar degeneration.

Frontotemporal lobar degeneration (FTLD) is a heterogeneous group of disorders characterized by disturbances of behavior and personality and different types of language impairment with or without concomitant features of motor neuron disease or parkinsonism. FTLD is characterized by atrophy of the frontal and anterior temporal brain lobes. Detailed neuropathological studies have elicited proteinopathies defined by inclusions of hyperphosphorylated microtubule-associated protein tau, TAR DNA-binding protein TDP-43, fused-in-sarcoma or yet unidentified proteins in affected brain regions. Rather than the type of proteinopathy, the site of neurodegeneration correlates relatively well with the clinical presentation of FTLD. Molecular genetic studies identified five disease genes, of which the gene encoding the tau protein (MAPT), the growth factor precursor gene granulin (GRN), and C9orf72 with unknown function are most frequently mutated. Rare mutations were also identified in the genes encoding valosin-containing protein (VCP) and charged multivesicular body protein 2B (CHMP2B). These genes are good markers to distinguish underlying neuropathological phenotypes. Due to the complex landscape of FTLD diseases, combined characterization of clinical, imaging, biological and genetic biomarkers is essential to establish a detailed diagnosis. Although major progress has been made in FTLD research in recent years, further studies are needed to completely map out and correlate the clinical, pathological and genetic entities, and to understand the underlying disease mechanisms. In this review, we summarize the current state of the rapidly progressing field of genetic, neuropathological and clinical research of this intriguing condition.

Therapy-Induced Electrophysiological Changes in Primary Progressive Aphasia: A Preliminary Study.

Aims: This preliminary study aimed to investigate therapy-induced electrophysiological changes in persons with primary progressive aphasia (PPA). The investigated event-related potential (ERP) components associated with language processing were the mismatch negativity, P300, N400, and P600. Methods: A linguistic ERP test battery and standardized language assessment were administered in four patients with PPA of which two received speech-language therapy (SLT) and two did not receive therapy. The battery was administered twice with approximately 6 months in between in each patient. The results of the follow-up assessments were compared to the results of the initial assessments. Results: Although the results of the behavioral language assessment remained relatively stable between the initial and follow-up assessments, changes in the mean amplitudes, onset latencies, and duration of the ERP components were found in the four patients. In the two patients that did not receive SLT, an increased delay in 50% and a decreased mean amplitude in 25% of the measured ERP components were found. The electrophysiological changes found in the patients that received SLT were variable. Interestingly, the mismatch negativity and the N400 effect elicited by the categorical priming paradigm were less delayed and had an increased mean amplitude at the follow-up assessment in the patient with the non-fluent variant who received SLT. In this patient, the P600 component was absent at the initial assessment but present at the follow-up assessment. Conclusion: Although no clear patterns in electrophysiological changes between patients who received SLT and patients who did not receive SLT were found by our preliminary study, it seems like the SLT induced improvements or compensation mechanisms in some specific language comprehension processes in the patient with the NFV. The results of this study are still preliminary because only four heterogeneous patients were included. Future studies should include larger patient groups of the three clinical variants because the therapy-induced electrophysiological changes might differ depending on the clinical variant and the underlying pathology.

Proteomic Assessment of C57BL/6 Hippocampi after Non-Selective Pharmacological Inhibition of Nitric Oxide Synthase Activity: Implications of Seizure-like Neuronal Hyperexcitability Followed by Tauopathy.

Nitric oxide (NO) is a small gaseous signaling molecule responsible for maintaining homeostasis in a myriad of tissues and molecular pathways in neurology and the cardiovasculature. In recent years, there has been increasing interest in the potential interaction between arterial stiffness (AS), an independent cardiovascular risk factor, and neurodegenerative syndromes given increasingly epidemiological study reports. For this reason, we previously investigated the mechanistic convergence between AS and neurodegeneration via the progressive non-selective inhibition of all nitric oxide synthase (NOS) isoforms with N(G)-nitro-L-arginine methyl ester (L-NAME) in C57BL/6 mice. Our previous results showed progressively increased AS in vivo and impaired visuospatial learning and memory in L-NAME-treated C57BL/6 mice. In the current study, we sought to further investigate the progressive molecular signatures in hippocampal tissue via LC-MS/MS proteomic analysis. Our data implicate mitochondrial dysfunction due to progressive L-NAME treatment. Two weeks of L-NAME treatment implicates altered G-protein-coupled-receptor signaling in the nerve synapse and associated presence of seizures and altered emotional behavior. Furthermore, molecular signatures implicate the cerebral presence of seizure-related hyperexcitability after short-term (8 weeks) treatment followed by ribosomal dysfunction and tauopathy after long-term (16 weeks) treatment.