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Alzheimer’s disease (AD) and Parkinson’s disease (PD) are characterized by brain accumulation of aggregated proteins, leading to neuronal and glial dysfunction. Compelling data indicate that changes in vimentin expression, structure, and localization reflect alterations in cellular homeostasis and may correlate with disease progression. Yet, the involvement of abnormal vimentin in AD and PD remains unclear. Here, we have thoroughly characterized the distribution of modified, disease-associated vimentin in the AD and PD brain parenchyma, as well as in cerebrospinal fluid (CSF), hiPSC-derived astrocytes and organoids. For this purpose, we used the form-specific vimentin antibody [84-1], originally generated to recognize abnormal vimentin on the surface of sarcoma cells. The 84-1 vimentin reactivity pattern was characterized through immunohistochemistry and proximity ligation assay. Moreover, proteomic analysis, Western blot, and ELISA were performed to quantify 84-1 vimentin levels and gain insights into its molecular features. Our data demonstrate that 84-1 can identify a distinct population of vimentin in the human brain that shows low affinity to commonly used vimentin antibodies. The 84-1 vimentin is enriched in disease conditions and forms distinct deposits in the affected regions of the AD and PD brain, often colocalizing with pathological protein aggregates. Interestingly, the 84-1 vimentin pool consists mainly of cleaved proteoforms and reduction-resistant aggregates. Moreover, 84-1 vimentin levels are elevated in the CSF of AD and PD patients as well as in the culture medium of human astrocytes exposed to αSyn or Aβ fibrils. Taken together, our data highlights the importance of modified vimentin in neurodegeneration and presents 84-1 vimentin as a potential biomarker and future treatment target for AD and PD.

Background Alpha-synucleinopathies are common disorders that are expected to become increasingly prevalent in the future along with the longer life expectancy. However, their diagnosis is problematic as they are mainly based on clinical criteria without the support of disease-specific biomarkers. This leads to frequent misdiagnoses, as underlined by autopsy studies, and an imprecise selection of patients for clinical trials, preventing progress in the development of disease-modifying treatments. In recent years important advances have been made regarding the development of specific biomarkers for the detection of pathological alpha-synuclein (alpha-syn), which may improve the diagnosis of patients affected by alpha-synucleinopathies. Results In this review, we describe in detail the most promising techniques to detect pathological alpha-syn in patient-derived samples. In particular, we describe the diagnostic accuracy of each individual cerebrospinal fluid (CSF), plasma and skin alpha-syn biomarker in differentiating alpha-synucleinopathies from controls, from other neurodegenerative disorders and between different alpha-synucleinopathies. Furthermore, we underline the main advantages and limitations of these techniques for clinical practice. Finally, we provide our suggestions for further development considering both technical aspects and large-scale standardization. Conclusions and Relevance We conclude that immunofluorescence on biopsied skin tissue and the seed amplification assay on CSF show the best diagnostic accuracy and reliability in the studies that have been performed to date. We discuss the opportunities of these techniques as well as the main current limitations and technical problems that need to be considered before they can be adopted for clinical use.

Background: Idiopathic achalasia (IA) is characterised by the degeneration of neurons in the myenteric plexus leading to an irreversible impaired oesophageal function. Although immune-mediated mechanisms have been proposed, the underlying aetiopathology of IA remains poorly understood.

Objective: This study aimed to uncover the genetic risk architecture of IA.

Design: We carried out the first genome-wide association study (GWAS) on 4602 European patients with IA and 10 766 ethnically-matched controls.

Results: A single nucleotide polymorphism (SNP) in HLA-DQB1 leading to an 8-amino acid insertion on the protein level conferred strongest IA risk (PQGPPPAG: p=3.27x10^-68, OR=2.45). Conditional analyses within the HLA locus revealed a complex genetic risk architecture. Three additional amino acid positions showed independent IA association (Omnibus p<5x10^-8). These refer to positions 41 and 130 in HLA-DQα1, position 45 in HLA-DQβ1 and position 86 in HLA-DRβ1. Together, these findings highlight the pivotal role of class II HLA genetic variation in IA pathogenesis. Outside HLA, three independent variants showed IA association (p<5x10^-8). One leads to an amino acid substitution with functional effect in PTPN22. Another risk variant leads to a downregulated expression of TNFSF8, TNFSF15 and TNC in immune cells. The third risk SNP is located near ZNF365, but the exact underlying cellular mechanism remains unknown. Beyond the single marker level, polygenic risk scores revealed that patients with IA can be stratified based on their genetic risk. In addition, IA shows a shared aetiopathology with Crohn's disease (r_g=0.335). Integrating GWAS and single-cell RNA-sequencing data from the myenteric plexus showed that the memory T-cell type FOS⁺Tc4⁺CD8⁺ plays a central role in IA development (p=2.50x10^-19).

Conclusion: This GWAS led to the identification of SNPs, cellular mechanisms and cell types that are involved in IA aetiopathology.

The early diagnosis of Parkinson's disease (PD) represents a challenge, and novel accurate biomarkers are therefore needed urgently. Detection of phosphorylated alpha-synuclein (p-alpha-syn) in skin nerve fibres has shown promise as such a marker. However, its accuracy for the identification of PD among patients with early signs of parkinsonism has not been explored thoroughly. In this blinded, multicentre, prospective and follow-up study, 151 patients diagnosed with early-stage parkinsonism (<18 months duration) were enrolled at three tertiary movement disorder centres. Clinical scales were performed, and initial diagnoses were reassessed at 18- and 46-month follow-up visits. Skin biopsy, with analysis for neuronal and glial p-alpha-syn deposits, and olfactory testing were performed at the baseline visit and repeated in 44 patients at the 18-month visit. After the follow-up period, a final diagnosis was reached in 140 patients: PD (n = 101; 67% of all screened patients), tauopathies (n = 22; 15%), multiple system atrophy (n = 5; 3%), vascular parkinsonism (n = 4; 3%), essential tremor (n = 3; 2%), dystonic tremor (n = 2; 1%) and no neurological illness (n = 3, 2%). Eleven patients did not fit any clinical criteria and were therefore classified as undefined. Baseline skin intraneural p-alpha-syn showed a robust diagnostic accuracy (81% sensitivity and 100% specificity) for identifying PD. Importantly, in 30 of 44 patients not diagnosed with PD until the follow-up, intraneural p-alpha-syn was already positive at baseline. Moreover, the analyses showed a large degree of consistency over time, in that the same results at baseline and follow-up were obtained in 42 (96%) of the patients tested. Finally, although olfactory testing at baseline showed a more abnormal score in PD compared with the other groups, its predictive accuracy was more modest (72%). In conclusion, given its high sensitivity, specificity and reproducibility, skin intraneural p-alpha-syn could become a valuable tool for diagnosing PD at early stages, potentially years before the diagnostic criteria are met, and for differentiating PD from atypical parkinsonism. In contrast, olfactory function, although more impaired in PD than in non-PD patients, seems to have only a more limited diagnostic accuracy.

Background and objectives: Idiopathic normal pressure hydrocephalus (iNPH) is a progressive but treatable neurological disorder. Yet, diagnosis is often confounded by overlapping symptoms and biomarker profiles with Alzheimer's disease (AD), mild cognitive impairment (MCI), and frontotemporal dementia (FTD). We aimed to determine whether cerebrospinal fluid (CSF) metabolomic profiling, combined with uncertainty-aware machine learning using conformal prediction (CP), could improve diagnostic differentiation of iNPH.

Methods: CSF samples were collected from 120 patients with iNPH, 44 healthy controls, and 152 individuals with AD, MCI, or FTD. Targeted metabolomics of 59 metabolites was performed using liquid chromatography-high-resolution mass spectrometry. Group differences were assessed using age- and sex-adjusted regression models. Multivariate classification with partial least squares discriminant analysis (PLS-DA) incorporated metabolites, demographics, and conventional biomarkers (amyloid-β42, tau, phosphorylated tau). CP was applied to address individual-level diagnostic uncertainty.

Results: Eight metabolites (proline, threonine, histidine, tyrosine, tryptophan, isobutyrylcarnitine, citric acid, and dehydroascorbic acid) were consistently reduced in iNPH (q < 0.05), independent of ventricular volume and cortical tau or amyloid-β pathology. An integrated PLS-DA model combining metabolomic, demographic, and AD-biomarker data achieved excellent discrimination (AUC = 0.97). CP provided calibrated case-level confidence, identifying clear-cut and uncertain cases while maintaining high accuracy (94% for iNPH, 97% for not-iNPH).

Discussion: iNPH exhibits a distinct CSF metabolomic signature reflecting altered amino acid metabolism, mitochondrial function, and oxidative stress. Integrating metabolomic data with established biomarkers enhances diagnostic accuracy, while CP adds individualized uncertainty estimates to improve diagnostic confidence and guide treatment decisions.

Progressive supranuclear palsy (PSP) is a neurodegenerative disease characterized by abnormal accumulation of the protein tau in the brain, leading to motor and cognitive symptoms that vary between individuals. The reasons for this clinical heterogeneity are unknown. Here we show that distinct molecular forms of tau, particularly high molecular weight (HMW) assemblies, differ in abundance and biological activity across PSP brains. By combining biochemical examination, seed amplification assays, proteomic profiling, and spatial transcriptomics, we identify that HMW tau species drive the strongest aggregation activity in the primary motor cortex. Cases with high tau seeding activity display molecular signatures of altered immune and metabolic pathways. These findings reveal that tau seeding activity reflects underlying molecular heterogeneity in PSP and suggest that measuring 4R-tau seeding capacity could help stratify patients and guide the development of targeted therapeutic approaches.

Parkinson's disease (PD) is characterized by a progressive loss of dopaminergic neurons, linked to aggregation of alpha-synuclein (αSYN) into Lewy bodies. Current treatments are symptomatic and do not halt or reverse the neurodegeneration. Immunotherapy targeting aggregated αSYN shows potential, but therapeutic efficacy is limited by poor brain penetration of antibodies. We developed a bispecific antibody, RmAb38E2-scFv8D3, based on αSYN oligomer selective RmAb38E2 fused to a transferrin receptor (TfR)-binding domain to enhance brain delivery. Both RmAb38E2 and RmAb38E2-scFv8D3 showed higher affinity for αSYN oligomers than for monomers or fibrils. In vivo, RmAb38E2-scFv8D3 exhibited higher brain and lower blood concentrations compared to RmAb38E2, suggesting a better brain uptake and reduced peripheral exposure for the bispecific antibody. Treatment over five days of 3–4 months old transgenic L61 mice, which overexpress human αSYN, with three doses of RmAb38E2-scFv8D3 reduced brain αSYN oligomer levels and increased microglial activation, as indicated by elevated soluble TREM2 levels. Treatment with the monospecific RmAb38E2, however, showed no significant effect compared to PBS. This study demonstrates that TfR-mediated delivery enhances the therapeutic potential of αSYN-targeted immunotherapy by resulting in a higher concentration and a more uniform distribution of antibodies in the brain. The use of bispecific antibodies offers a promising strategy to improve the efficacy of antibody therapies in PD and other α-synucleinopathies.

Today, 13 intra-amyloid-β (Aβ) amyloid precursor protein (APP) gene mutations are known to cause familial Alzheimer's disease (AD). Most of them are point mutations causing an increased production or a change in the conformation of Aβ. The Uppsala APP mutation (Δ690-695 in APP, Δ19-24 in Aβ) is the first known multi-codon deletion causing autosomal dominant AD. Here, we applied cryo-electron microscopy (cryo-EM) to investigate the structure of Aβ fibrils with the Uppsala APP mutation from tg-UppSwe mouse brain tissue. Murine Aβ Upp(1-42)_Δ19-24 are made of two identical S-shaped protofilaments with an ordered fibril core of S8-A42. The murine Aβ fold is almost identical to previously described human type II filaments, although the amino acid sequences differ considerably. In addition, we report the cryo-EM structure of Aβ fibrils from the temporal cortex of a patient with the Uppsala APP mutation. The observed structure of the human Aβ fold closely resembles previously described type I fibrils. Structural modeling suggests that these fibrils are composed of wild-type Aβ, which implies that AβUpp may be less soluble and thus not readily accessible for cryo-EM image processing and structure determination. Additionally, from the human sample we determined the structures of tau paired helical filaments and tau straight filaments, which are identical to those found in sporadic AD cases. Finally, we present the 3D cryo-EM structures of four dominant AβUpp(1-42)_Δ19-24 fibril polymorphs, formed in vitro. All four polymorphs differ from the observed folds of Uppsala Aβ in murine and human brain tissue, respectively.

INTRODUCTION:

Neuroinflammation, a key player in Alzheimer's disease (AD) pathogenesis, may be differentially involved in young-onset (YOAD) compared to late-onset (LOAD) AD.

METHODS:

Using proximity extension assay technology, we examined 737 inflammatory markers in the CSF of 26 healthy controls (63.9 +/- 8.7; 12 female), 57 patients with YOAD (60.8 +/- 4.9 y/o; 40 female), and 33 with LOAD (76.6 +/- 4.5 y/o; 18 female). We also assessed biomarkers of AD pathology (A beta 42, p-tau181, t-tau) and neurodegeneration (neurofilament light-chain [NfL]).

RESULTS:

Compared to controls, SCRN1 and MMP10 were increased in LOAD and YOAD, but 16 markers showed YOAD-specific increases. Forty-six markers were significantly associated with NfL. P-tau181 and t-tau mediated the association between inflammatory markers and NfL in YOAD. In LOAD we could not identify a direct or indirect relationship between neuroinflammation and neurodegeneration.

DISCUSSION:

Using a proteomics approach, we observed an exacerbation of neuroinflammatory changes and a differential contribution of neuroinflammation to AD pathology and neurodegeneration in YOAD compared to LOAD.

Alzheimer's disease (AD) is a common and increasing societal problem due to the extending human lifespan. In males, loss of chromosome Y (LOY) in leukocytes is strongly associated with AD. We studied here DNA methylation and RNA expression in sorted monocytes and granulocytes with and without LOY from male AD patients. Through multi-omic analysis, we identified new candidate genes along with those previously associated with AD. Global analyses of DNA methylation in samples with LOY vs. normal state showed that hypomethylation dominated both in granulocytes and monocytes. Our findings highlight LOY-related differences in DNA methylation that occur in gene regulatory regions. Specifically, we observed alterations in key genes involved in leukocyte differentiation: FLI1, involved in early hematopoiesis; RUNX1, essential for blood cell development; RARA, regulating gene expression in response to retinoic acid; CANX, crucial for protein folding; CEBPB, a transcription factor important for immune responses; and MYADM, implicated in cell adhesion and migration. Moreover, protein-protein interaction analysis in granulocytes identified that products of two of these genes, CANX and CEBPB, are key hub proteins. This research underscores the potential of multi-omic approach in pure hematopoietic cell populations to uncover the molecular underpinnings of AD. Finally, our results link previous analysis showing impact of LOY on leukocyte differentiation, LOY-associated transcriptional dysregulation and GWAS studies of LOY.