Brain banking enables the study of human neural tissue and is essential for research on disease, physiology, and neuroanatomy. An essential step for whole brain studies is the extraction of the brain from the skull. Yet detailed technical descriptions of brain removal are rare, perhaps contributing to the artifactual tissue disruption sometimes observed as a result of the procedure. Here, we describe a method for whole brain extraction that could be implemented in the context of a whole-body donation, focused on the use case of fixed tissue banking. The method involves a sequential craniectomy that uses both circumferential and midline sagittal cuts, followed by a posterior approach to cutting the dura. We report the application of this protocol across n = 105 human whole-body donors. We document the time required for each procedural step and the frequency of craniectomy artifacts and skull edge artifacts at the brain surface. When the brain tissue is particularly soft, we also describe the potential use of in situ immersion fixation to increase tissue stiffness before removal, finding however that this also slows the diffusion of chemicals into the interior of the brain. Our experience suggests that the effectiveness of brain extraction can be improved via procedural optimizations. Technicians can become comfortable with the method after approximately 5-10 cases. We anticipate that improving brain extraction quality may support downstream work in the development of diagnostics and treatments for neurological and psychiatric disease.

Long-term storage of aldehyde-fixed brain tissue is commonly performed in the fluid state. This has the potential to maintain morphology for many decades, but has been found to cause progressive loss of antigenicity over time for some biomolecules, motivating interest in alternative long-term preservation strategies, such as cryopreservation. While cryoprotection and subzero storage has been successfully used for brain tissue sections or blocks, methods for preserving whole brains using this approach have not been widely characterized. Here we present a protocol for preserving fixed whole brains using graded immersion cryoprotection followed by subzero temperature storage. We refer to this general strategy - aldehyde fixation followed by cryoprotectant loading and subzero storage - as aldehyde-based cryopreservation (ABC). Our method uses a gradual ramp-up of the osmotic concentration of cryoprotectants, leading to a final solution containing 50% (v/v) ethylene glycol and 30% (w/v) sucrose in fixative. We used CT imaging to track cryoprotectant penetration, finding that approximately 9 months is required for the CT signal to stabilize throughout whole human brains. In our initial validation experiment, insufficient equilibration time prior to freezing led to ice crystal artifacts in the white matter. After refining the protocol to allow adequate diffusion time, light and electron microscopy showed preserved cellular architecture and ultrastructure. Our approach may be valuable for laboratories seeking a method for long-term subzero storage of fixed whole brain specimens.

The Tau Global Conference 2025, hosted by the Alzheimer's Association, CurePSP, and the Rainwater Charitable Foundation, convened international experts from academia, industry, government, and philanthropy to explore advances and challenges in tauopathy research. The meeting highlighted progress across tau biology, including emerging models of tau regulation, degradation, and propagation; advances in biomarker development for the diagnosis and staging of tauopathies; and evolving therapeutic strategies targeting diverse aspects of tau pathophysiology. Discussions also emphasized the importance of cross-sector collaboration, and global initiatives to address disparities in tau research. This report synthesizes key insights from the conference and underscores the critical role of interdisciplinary, biomarker-driven, and globally inclusive approaches in accelerating the translation of tau research into effective clinical applications.

Primary age-related tauopathy (PART) is a β-amyloid-independent tauopathy, thought by some to be a distinct process from Alzheimer disease neuropathologic change (ADNC). Two categories of PART have been defined: definite PART (those with complete absence of β-amyloid deposition) and possible PART (those with minimal and restricted β-amyloid distribution). It is unclear whether there is any significant cognitive effect of "isolated" or "pure" PART, as opposed to ADNC, in which cognitive decline mirrors pathologic progression. We evaluated the effects of neurodegenerative pathologies on longitudinal cognitive decline using a combination of univariate analysis, multivariable logistic regression analysis, and variance decomposition in patient cohorts with neuropathologically confirmed definite PART (n = 174) and possible PART (n = 182). ADNC-related pathologies did not significantly contribute to cognitive impairment in either cohort. Cognitive decline in definite PART was instead dependent on the presence and severity of TDP-43 pathology/limbic-predominant age-related TDP-43 encephalopathy (LATE) stage, Lewy body pathology, and arteriolosclerosis, while cognitive impairment in possible PART was primarily affected by hippocampal sclerosis and infarcts. Additionally, 67.8%-75.7% of variance in cognitive decline was unaccounted for by these neurodegenerative pathologies. These results indicate that PART pathology in isolation does not significantly impair cognition, which is instead primarily influenced by comorbid neuropathologic features.

Expansion microscopy (ExM) physically enlarges biological specimens to enable ultrastructural imaging with conventional fluorescence microscopes. However, its performance in postmortem human brain tissue is unclear. Here, we evaluated how a previously established ExM protocol performs on cortical tissue from eight banked brains with postmortem intervals ranging from 40 minutes to 91 hours, and compared these data with electron microscopy (EM) on a subset of matched samples. Both techniques revealed similar patterns of ultrastructural features, including irregular, rounded, unstained spaces as postmortem artifacts that increased with longer postmortem intervals. While EM provided superior resolution for synaptic details, ExM enabled more high-throughput volumetric imaging of neural circuits. ExM also enabled molecular annotation of ultrastructure through immunofluorescence, as demonstrated by SMI-312 neurofilament and γ-protocadherin labeling. Our findings show that ExM can visualize aspects of ultrastructure in routinely banked brain tissue. While EM provides better resolution for fine synaptic detail, ExM offers a complementary approach that combines nanoscale imaging with molecular specificity and more accessible high-throughput volumetric capabilities.

BACKGROUND:XK disease is a multisystem neurodegenerative disorder caused by mutations in the XK gene that codes for the lipid scramblase XK. OBJECTIVE:The aim was to describe the lipidomic spectrum in postmortem brain tissue from XK patients. METHODS:We measured the levels of 593 lipid species in the caudate nucleus (CN), putamen, and dorsolateral prefrontal cortex (DLPFC) from postmortem tissues of 5 XK patients and 6 controls. RESULTS:In XK patients, we observed increased levels of triacylglycerol, monoacylglycerol, phosphatidylserine, and ceramide in the CN. Acylated phosphatidylglycerol levels were reduced in both the CN and putamen. Acyl carnitine, dihydrosphingomyelin, and monosialodihexosylganglioside were reduced, whereas N-acyl phosphatidylethanolamine was increased in the DLPFC. N-Acyl serine was reduced in all three regions. CONCLUSIONS:Our findings provide initial evidence of abnormal sphingolipid and phospholipid concentrations in the brains of XK patients and may provide insights into mechanisms of neurodegeneration in this disease. © 2026 International Parkinson and Movement Disorder Society.

Companion animal brain banking has been recognized as a valuable approach for translational aging and dementia research. However, realizing the full value of canine brain banks depends on optimizing the methods that are used to collect and preserve the tissue. Whole brain perfusion fixation is one promising approach, but it is not yet well described in dogs. Here we describe the development of methods for a canine brain bank (currently n = 55), including whole brain perfusion fixation via aortic cannulation and brain extraction. We assessed perfusion quality using gross examination, post-perfusion CT, and histological clearance of blood vessels. We found that body weight and average flow rate per body weight were each significantly correlated with perfusion quality in our cohort. To illustrate the kind of analysis the bank could facilitate, we next performed a preliminary study of brain aging, one of our primary planned research applications. Using a pixel classifier applied to whole slide images, we quantified lipofuscin burden, and in this preliminary cohort found that it increased strongly with age in both the thalamus and hippocampus. In the hippocampus, lipofuscin burden was also elevated in dogs with owner-reported cognitive dysfunction, although the current cohort is too small to determine to what extent this association is independent of age. Preliminary electron microscopy studies also confirmed that perfusion fixed tissue from the bank is amenable to ultrastructural analysis. This work describes one approach for canine brain perfusion fixation and introduces a brain tissue resource that may help support future neuroscience research.

Alzheimer disease neuropathologic change (ADNC) is considered to be the most common cause of cognitive decline and dementia worldwide. ADNC level is determined using the density of neuritic plaques in combination with the topographical distribution of β-amyloid (Aβ) plaques and hyperphosphorylated tau (p-tau)-positive neurofibrillary tangles (NFTs). While cognitive decline correlates with the level of ADNC, there remains a great deal of variation in cognitive outcomes between individuals that is unaccounted for by current neuropathologic evaluation metrics. We leveraged quantitative computer-assisted positive pixel assessments to establish the neocortical p-tau burden in the middle frontal and superior temporal gyri of 61 individuals with Braak NFT stage V who had a wide range of cognitive outcomes and trajectories. Frontal and temporal neocortical p-tau burden varied between 0.2

Myelin oligodendrocyte basic protein (MOBP) is an abundant oligodendrocyte gene implicated in multiple neurodegenerative diseases. Genetic variation at the MOBP locus has been associated with risk for progressive supranuclear palsy (PSP), amyotrophic lateral sclerosis (ALS), frontotemporal lobar degeneration (FTD), corticobasal degeneration (CBD), Alzheimer's disease (AD), Lewy body dementia (LBD), and Creutzfeldt-Jakob disease (CJD). Epigenetically, MOBP promoter hypermethylation and reduced expression have been reported in multiple system atrophy (MSA). Although MOBP is thought to play a role in oligodendrocyte morphology and myelin structure, how genetic and epigenetic variation at this locus influences gene regulation and contributes to disease risk remains poorly understood across neurodegenerative disorders. Here, we investigated whether shared or disease-specific genetic mechanisms at MOBP converge on altered DNA methylation and expression across neurodegenerative disorders. We analysed MOBP variants using summary statistics from recent GWAS for ALS, PSP, FTD, LBD, PD, MSA, AD, and CJD. Colocalisation (COLOC and SuSiE-coloc) was used to test whether disease-associated variants overlapped between diseases, and with oligodendrocyte expression quantitative trait loci (eQTLs) and bulk brain methylation quantitative trait loci (mQTLs). To further investigate mQTL effects at this locus, rs1768208, a variant previously associated with PSP, was genotyped in an overlapping brain methylation cohort, allowing direct testing of genotype-methylation associations in frontal white matter tissue. ALS and PSP GWAS demonstrated strong association at MOBP, with most strongly associated SNPs (e.g. rs631312, rs616147, rs1768208) shared between both disorders. Colocalisation analyses indicated high posterior probability that ALS and PSP share the same causal variant, with weaker overlap with FTD. mQTL colocalisation highlighted cg15069948, located near an exon junction within MOBP, as strongly colocalising with the ALS/PSP risk variants. In complementary tissue analyses, rs1768208-T carriers showed hypomethylation at cg15069948 in PSP brains. No genotype-methylation effects were detected in MSA or Parkinson's disease. Together with prior evidence of promoter hypermethylation and reduced expression in MSA, our findings identify cg15069948 as a regulatory methylation site linking ALS/PSP risk variants to altered MOBP methylation, and support MOBP dysregulation as a shared feature of neurodegeneration. However, the underlying mechanisms appear disease-specific, highlighting the complexity of involvement of this gene across neurodegenerative disorders.

Abstract Long-term storage of aldehyde-fixed brain tissue is commonly performed in the fluid state. This has the potential to maintain morphology for many decades, but has been found to cause progressive loss of antigenicity over time for some biomolecules. While cryoprotection and subzero storage has been successfully used for brain tissue sections or blocks, methods for preserving whole brains using this approach have not been widely characterized. Here we present a protocol for the preservation of fixed whole brains using graded immersion cryoprotection and subzero temperature storage, which is one type of a more general approach that we refer to as aldehyde-based cryopreservation (ABC). Our method uses a gradual ramp-up of the osmotic concentration of cryoprotectants, leading to a final solution containing 50% (v/v) ethylene glycol and 30% (w/v) sucrose. We used CT imaging to track cryoprotectant penetration, finding that with the use of our protocol, approximately 10 months is required to reach equilibration throughout whole human brains. In our initial histological validation, we found that insufficient equilibration time prior to freezing led to apparent ice crystal artifacts seen on ultrastructural imaging of the white matter. After refining the protocol to allow adequate diffusion time, histologic data at both the light and electron microscopic levels showed preserved cellular architecture and ultrastructure after the process of cryoprotectant loading, freezer storage, and unloading. This protocol can be implemented using laboratory freezers or freezer rooms and provides a degree of resilience against freezer failures because the morphology of the fixed tissue is expected to remain preserved long-term in the fluid state even if rewarmed. Our approach may be valuable for laboratories seeking to enhance the long-term preservation of antigenicity in large brain tissue specimens for future research applications.

Perfusion fixation is widely used in neuroscience to prepare mammalian brain tissue for histological and ultrastructural analysis. Perfusion protocols are commonly assessed using macroscopic indicators such as gross appearance and neuroimaging, which assess the extent to which perfusate has been distributed throughout the brain. There is a critical need to determine to what extent these metrics can accurately predict high-quality ultrastructural preservation, particularly as new perfusion protocols are developed for connectomics. In this technical report, we describe evidence that these two measures can be decoupled by the addition of dehydrating agents to the perfusate solution. In three human brain donors and one canine brain donor perfused with a fixative solution containing 10% mannitol and 10% polyethylene glycol 35 kDa, macroscopic and radiological indicators of perfusion quality appeared adequate or favorable. However, electron microscopy revealed expanded extracellular space, shrunken cellular processes, and distorted cell membranes, consistent with an osmotic shock artifact resulting from severe hyperosmotic dehydration. Similar ultrastructural artifacts were observed in a canine brain donor perfused with 20% mannitol in 20% neutral buffered formalin without PEG. We compare these ultrastructural findings with findings from previously reported cases perfused with standard neutral buffered formalin without osmotic additives. These findings illustrate a risk of optimizing brain perfusion protocols designed to preserve neural circuitry based on macroscopic or radiological perfusion quality metrics alone, since these metrics can be satisfied while the ultrastructure is severely compromised.

Alzheimer’s disease (AD) is characterized by neocortical dissemination of neurofibrillary tangles (NFTs) while primary age-related tauopathy (PART) has NFTs largely confined to the hippocampus and adjacent structures. Thus, PART and AD represent two extremes of a spectrum of NFT spread. We investigated epigenetic mechanisms of interindividual variation in NFT spread. We evaluated DNA methylation (DNAm) in frontal cortex by Infinium EPIC BeadChip array in the multi-center PART Working Group cohort (PWG, N = 398), controlling for age, sex, and batch effects. Using SeSAMe and a false discovery rate of p<0.05, we identified differentially methylated positions (DMPs) associated with PART pathology quantitated from immunohistochemically-stained sections. To assess amyloid effects, we compared this set with DMPs associated with neuritic amyloid plaque using the CERAD score. We identified novel DMPs not previously implicated in AD. We validated our findings in the Religious Orders Study and Memory and Aging Project cohort (ROSMAP, N = 707). We then related DMPs to differential expression of linked genes and performed enrichment analyses using KnowYourCG. We identified DMPs associated with NFTs [ PWG : 8 novel / 8 total; ROSMAP : 31 novel / 68 total]. In the PWG cohort, of the 550 total [539 novel] DMPs that associated with CERAD score, there was no overlap with NFT-associated DMPs. In the ROSMAP cohort, 57 total [3 novel] DMPs are associated with both NFTs and diagnosis of PART vs. AD and are linked to genes related to T-cells and axonal transport. In contrast, the 11 total [10 novel] DMPs associated with NFTs alone are linked to genes related to synaptic signaling, heparin sulfate proteoglycan biosynthesis, and microtubule architecture. DNA methylation distinguishes PART and AD brain. We identify tau-specific DMPs that account for variance in NFT burden among aging individuals, both in the absence and presence of amyloid plaques in PART and AD, respectively. In PART, tau-DMPs are fully orthogonal to the set of amyloid-DMPs. In contrast, in AD, the tau-DMPs have both amyloid-associated and amyloid-independent subsets with separate gene enrichment profiles. Therefore, distinct epigenetic events may drive pathology within the limbic system compared to those that drive tau spread to neocortex.

BACKGROUND:The 17q21.31 region with various structural forms characterized by the H1/H2 haplotypes and three large copy number variations (CNVs) represents the strongest risk locus in progressive supranuclear palsy (PSP). OBJECTIVE:To investigate the association between CNVs and structural forms on 17q.21.31 with the risk of PSP. METHODS:Utilizing whole genome sequencing data from 1684 PSP cases and 2392 controls, the three large CNVs (α, β, and γ) and structural forms within 17q21.31 were identified and analyzed for their association with PSP. RESULTS:We found that the copy number of γ was associated with increased PSP risk (odds ratio [OR] = 1.10, P = 0.0018). From H1β1γ1 (OR = 1.21) and H1β2γ1 (OR = 1.24) to H1β1γ4 (OR = 1.57), structural forms of H1 with additional copies of γ displayed a higher risk for PSP. The frequency of the risk sub-haplotype H1c rises from 1% in individuals with two γ copies to 88% in those with eight copies. Additionally, γ duplication up-regulates expression of ARL17B, LRRC37A/LRRC37A2, and NSFP1, while down-regulating KANSL1. Single-nucleus RNA-seq of the dorsolateral prefrontal cortex analysis reveals γ duplication primarily up-regulates LRRC37A/LRRC37A2 in neuronal cells. CONCLUSIONS:The copy number of γ is associated with the risk of PSP after adjusting for H1/H2, indicating that the complex structure at 17q21.31 is an important consideration when evaluating the genetic risk of PSP. © 2025 The Author(s). Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

Foundation models have transformed computational pathology by providing generalizable representations from large-scale histology datasets. However, existing models are predominantly trained on surgical pathology data, which is enriched for non-nervous tissue and overrepresents neoplastic, inflammatory, metabolic, and other non-neurological diseases. Neuropathology represents a markedly different domain of histopathology, characterized by unique cell types (neurons, glia, etc.), distinct cytoarchitecture, and disease-specific pathological features including neurofibrillary tangles, amyloid plaques, Lewy bodies, and pattern-specific neurodegeneration. This domain mismatch may limit the ability of general-purpose foundation models to capture the morphological patterns critical for interpreting neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, and cerebellar ataxias. To address this gap, we developed NeuroFM, a foundation model trained specifically on whole-slide images of brain tissue spanning diverse neurodegenerative pathologies. NeuroFM demonstrates superior performance compared to general-purpose models across multiple neuropathology-specific downstream tasks, including mixed dementia disease classification, hippocampal region segmentation, and neurodegenerative ataxia identification encompassing cerebellar essential tremor and spinocerebellar ataxia subtypes. This work establishes that domain-specialized foundation models trained on brain tissue can better capture neuropathology-specific features than models trained on general surgical pathology datasets. By tailoring foundation models to the unique morphological landscape of neurodegenerative diseases, NeuroFM enables more accurate and reliable AI-based analysis for brain disease diagnosis and research, setting a precedent for domain-specific model development in specialized areas of digital pathology.

Immersing the brain in a solution containing formaldehyde is a commonly used method for preserving the structure of human brain tissue in brain banking. However, there are questions about the quality of preservation using this method, as formaldehyde takes a relatively long period of time to penetrate a large organ such as the human brain. As a result, there is a critical need to determine whether immersion fixation is an adequate initial preservation method. To address this, we present exploratory histologic findings from our brain bank following the immersion fixation of hemi-sectioned brain specimens under refrigeration. Using light microscopy, we found that there was no significant change in the size of pericellular or perivascular rarefaction areas based on the postmortem interval (PMI) or on the progression from the outer (frontal cortex) to the inner (striatum) brain regions. Additionally, we did not identify any significant number of ghost cells - a state of late-stage cellular necrosis - in the light micrographs analyzed. Using transmission electron microscopy of tissue from the frontal cortex, we found that synapses could still be visualized, but there was vacuolization and variable degrees of myelin disbanding identified. Using serial section transmission electron microscopy, we found that identified synapses could be traced from one section to the next. Using serial block face scanning electron microscopy, we also found that myelinated axons on 2D images can be traced with high fidelity from one image to the next, even at PMIs of up to 27 hours. Collectively, our data corroborate previous findings that immersion fixation is effective for prevention of cellular necrosis and for visualizing many ultrastructural features in at least the surface areas of the brain. However, how structural preservation quality should best be assessed in brain banking is an open question that depends on the intended research applications.

Although Alzheimer disease neuropathologic change (ADNC) is the most common pathology underlying clinical dementia, the presence of multiple comorbid neuropathologies is increasingly being recognized as a major contributor to the worldwide dementia burden. We analyzed 1051 subjects with specific combinations of isolated and mixed pathologies and conducted multivariate logistic regression analysis on a cohort of 4624 cases with mixed pathologies to systematically explore the independent cognitive contributions of each pathology. Alzheimer disease neuropathologic change and limbic-predominant age-related TDP-43 encephalopathy neuropathologic change (LATE-NC) were both associated with a primary clinical diagnosis of Alzheimer disease (AD) and were characterized by an amnestic dementia phenotype, while only ADNC associated with logopenic variant primary progressive aphasia (PPA). In subjects with ADNC and comorbid LATE-NC, Lewy body disease, and/or cerebrovascular disease, the clinical phenotype was usually diagnosed during life as "Probable AD." Conversely, the combination of ADNC with frontotemporal lobar degeneration with TDP-43, progressive supranuclear palsy (PSP), or corticobasal degeneration (CBD) resulted in a mixed clinical picture, with variable features of amnestic dementia, PPA subtypes, behavioral variant FTD, PSP syndrome, and CBD syndrome. These findings elucidate the cumulative effects of mixed pathologies and provide insights into interactions between neurodegenerative pathologies contributing to a variety of clinical dementia presentations.

INTRODUCTION:The presence and interaction of multiple comorbid neuropathologies are a major contributor to the worldwide dementia burden. METHODS:We analyzed 1183 subjects from the National Alzheimer's Coordinating Center dataset with various combinations of isolated and mixed neurodegenerative pathologies and conducted mixed-effects multiple linear regression modeling to comprehensively compare the neurocognitive and neuropsychological trajectories between groups over time. RESULTS:In combination with Alzheimer's disease neuropathologic change, various combinations of limbic-predominant age-related transactive response DNA-binding protein 43 encephalopathy, Lewy body dementia, and cerebrovascular disease further impair global cognition and specific neurocognitive domains; however, they do not appear to extensively affect the rate of decline with time across these domains, suggesting an additive but not synergistic effect. DISCUSSION:These findings corroborate the known cumulative effects of mixed pathologies on cognition and add nuance to our understanding of their specific interactions, which is crucial for the development of biomarkers and effective therapeutics. HIGHLIGHTS:Mixed neurodegenerative pathologies are common in the elderly population. The most common neurodegenerative pathologies were Alzheimer's disease neuropathologic change (ADNC), cerebrovascular disease (CVD), Lewy body dementia (LBD), and limbic-predominant age-related transactive response DNA-binding protein 43 encephalopathy (LATE). The addition of various combinations of comorbid CVD, LBD, and LATE to ADNC worsened overall performance on cognitive and neuropsychological testing across time. In general, the addition of multiple comorbid neurodegenerative pathologies did not influence the rate of decline across the evaluated time period.

Telomeres are repetitive DNA sequences at the ends of chromosomes which contribute to maintaining chromosomal stability. Telomere shortening is a hallmark of aging and shorter blood leukocyte telomere length (LTL) has been associated with increased risk for age-related diseases, however, little is understood about the biology of brain telomeres and how they may be involved in disease. Considering the increased neuropathologic burden of phosphorylated tau (ptau) with age, we investigated how shorter brain telomere length (brain-TL) may relate to increased ptau burden. We studied a cohort of 112 individuals with primary age-related tauopathy (PART), a neuropathological diagnosis characterized by mild-to-moderate tau burden (Braak=I-IV) primarily in the medial temporal lobe, with the relative absence of amyloid-beta plaques (CERAD=0). These individuals had both brain-TL (mean length by telomere qPCR, blinded) and DNA methylation measures from the frontal cortex, along with semi-quantitative Aperio ptau measures from the hippocampus. A subset ( n  = 81) had SNP genotyping data available. In an independent cohort ( n  = 10, Braak=0-VI, CERAD=0-3), we performed quantitative fluorescence in-situ hybridization (FISH) microscopy to measure the average ratio of telomere to centromere DNA content in nuclei from the frontal and visual cortices. In linear regression models, frontal cortex brain-TL did not relate to age. When age-adjusted, shorter brain-TL related to higher hippocampal ptau (β=-1.06, CI=-1.92–-0.195, p  = 0.017). A previously established DNA methylation model predictive of hippocampal ptau partially mediated the relationship between brain-TL and hippocampal ptau (proportion mediated=0.664, CI=0.246–1.33, p  = 0.012, Figure 1). A polygenic score for LTL did not relate to either age, brain-TL or hippocampal ptau. With FISH, we observed that individuals with CERAD=0 had shorter telomeres in the frontal cortex compared to individuals with CERAD=3. Within the CERAD=0 group, an individual with Braak=II had shorter telomeres than an individual with Braak=I. These patterns were not observed in the visual cortex. In a PART cohort, shorter frontal cortex brain-TL was related to higher hippocampal ptau, and this relationship was partially mediated by a DNA methylation model predictive of hippocampal ptau. A polygenic score for LTL was not predictive of brain-TL or hippocampal ptau. Together, this further emphasizes the importance of tissue-specific epigenetic modifiers of age-related ptau neuropathology.

The accumulation of abnormal tau protein in neurons and glia in the human brain is the defining feature of neurodegenerative diseases known as tauopathies. Progressive supranuclear palsy (PSP), the most common primary tauopathy, is typified by selective vulnerability of dopaminergic neurons and glia in the midbrain leading to an atypical parkinsonian movement disorder. To investigate candidate disease mechanisms underlying PSP, there is a critical need for model systems that more accurately recapitulate the cellular and molecular environment in the human brain. Human induced pluripotent stem cell (hiPSC)-derived organoid models have emerged as a powerful tool to address this gap. Skin biopsies were collected from living clinically diagnosed PSP patients or during autopsy. Fibroblasts were cultured and reprogrammed into hiPSCs using Sendai virus. HiPSCs were maintained with StemCultures FGF2 Discs to improve pluripotency and FACS was performed to confirm pluripotency marker expression. To generate midbrain organoids, hiPSCs were seeded into suspension spinner flasks, patterned using pharmacological directed differentiation, and grown for four months. Reliable patterning was confirmed with qRT-PCR, immunohistochemistry and immunoblot using a panel of cell-type specific markers. Astrocytes were extracted from mature organoids, cultured, and screened for astrocyte-specific markers. Fibroblasts have been banked from twenty-two PSP patients and seven reprogrammed into hiPSCs. Sporadic case status was determined by Sanger sequencing confirming the absence of a MAPT mutation. We found that hiPSCs grown with controlled-release FGF2 discs were 90-100% positive for pluripotency markers and negative for off-target genes. During patterning, organoids displayed morphological and cytoarchitectural patterns consistent with developing neuroectoderm and midbrain. Midbrain neural progenitor and dopaminergic markers such as FOXA2 , LMX1A , and TH were positive in a time-dependent manner, with mature dopaminergic neurons expressing NURR1 and GIRK2 detected by day 30. GFAP-positive astrocytes appeared around day 100. Astrocytes extracted from mature organoids were positive for multiple astrocyte markers. Sporadic PSP patient hiPSCs reliably differentiate into midbrain dopaminergic organoids and astrocytes, resulting in a sporadic tauopathy model containing key cell types affected in PSP. This cell collection is a valuable resource to investigate candidate mechanisms underlying tauopathy and could provide insight into cell-type specific disease drivers.