BACKGROUND AND OBJECTIVES:Heavy alcohol consumption is a major global health concern linked to increased morbidity and mortality. However, the long-term impact of excessive alcohol consumption on cognitive abilities and dementia-related neuropathology is unclear. The aim of this study was to analyze the association between alcohol consumption and age-related neuropathologic outcomes in a population-based autopsy study. METHODS:This cross-sectional study used data from the Biobank for Aging Studies, classifying participants as never, moderate, heavy, or former drinkers. Alzheimer disease pathology (neuritic plaques, amyloid deposition, and neurofibrillary tangles), Lewy body pathology, transactive DNA-binding protein 43, lacunar infarcts, hyaline arteriolosclerosis, and cerebral amyloid angiopathy were evaluated following international criteria using immunohistochemistry and hematoxylin and eosin staining. Cognitive abilities were evaluated using the Clinical Dementia Rating Scale Sum of Boxes, and the brain mass ratio was calculated by dividing the brain weight by the participant's height. Logistic and linear regression models were used to investigate the associations between alcohol consumption and neuropathology while we used mediation analysis to evaluate the direct and indirect effects of alcohol on cognition through neuropathologic lesions. RESULTS:We included 1,781 participants (mean age 74.9 ± 12.5 years, mean education 4.8 ± 4.0 years, 49.6% women, and 64.1% White). Compared with participants who never consumed alcohol, moderate (odds ratio [OR] 1.60, 95% CI 1.19-2.15, p = 0.001), heavy (OR 2.33, 95% CI 1.50-3.63, p < 0.001), and former heavy (OR 1.89, 95% CI 1.41-2.54, p < 0.001) alcohol consumptions were associated with hyaline arteriolosclerosis while only heavy (OR 1.41, 95% CI 1.10-2.30, p = 0.012) and former heavy (OR 1.31, 95% CI 1.02-1.68, p = 0.029) alcohol consumptions were associated with neurofibrillary tangles. Former heavy drinking was associated with a lower brain mass ratio (β -4.45, 95% CI -8.55 to -0.35, p = 0.033) and worse cognitive abilities (β 1.31, 95% CI 0.54-2.09, p < 0.001). The association between impaired cognitive abilities and alcohol consumption was fully mediated by hyaline arteriolosclerosis (β 0.13, 95% CI 0.02-0.22, p = 0.012). DISCUSSION:Moderate, heavy, and former heavy alcohol consumptions were associated with hyaline arteriolosclerosis and neurofibrillary tangles. Former heavy alcohol consumption was associated with reduced brain mass and cognitive abilities. The association between alcohol and cognitive abilities was fully mediated by hyaline arteriolosclerosis. The lack of longitudinal data on alcohol consumption duration restricts the interpretation of our findings.

ABSTRACT The APOE gene is a critical determinant of human healthspan and longevity, with the rare ε2 allele traditionally viewed as a universal protective factor against Alzheimer s disease (AD) and a driver of exceptional lifespan. However, this protective paradigm is predominantly derived from European-centric cohorts, leaving the evolutionary and clinical impacts of ε2 across diverse, highly admixed populations largely unknown due to a lack of local ancestry (LA) resolution. To investigate how local genomic backgrounds modulate APOE survival dynamics we analyzed two Brazilian sample collection of older adults from São Paulo city: the Biobank for Aging Studies (BAS, n = 716), a post-mortem autopsy study of naturally deceased individuals; and the Health, Well-being and Aging Study (SABE, n = 952), a census-based elderly sample collection. We evaluated deviations from Hardy-Weinberg equilibrium (HWE) using robust permutation-based models to capture ongoing selective and mortality pressures at the APOE locus. While global APOE frequencies adhered to HWE, integrating LA unveiled striking, mirrored ancestral deviations. Our findings reveal that APOE ε2 homozygotes with African ancestry significantly contribute to deviations from HWE in the BAS, with an excess of ε2AFR/ε2AFR homozygotes observed (p = 0.0196). These distinct HWE deviations demonstrate that an African LA background acts as a genetic buffer, attenuating the phenotypic extreme effects of APOE alleles. Furthermore, we observed an excess of the ε4 European haplotypes in the BAS, which is consistent with a mortality pressure allelic effect in the European LA context. Conversely, the ε4 AFR/ ε4 AFR combination was overrepresented in the SABE. While this buffering mechanism mitigates ε4 toxicity, it simultaneously dampens the exceptional longevity advantage typically conferred by the ε2 allele, leading to its neutral accumulation in the post-mortem cohort. Our study challenges the “one-size-fits-all” assumption of APOE biomarkers, demonstrating that ε2 protective mechanisms are context-dependent and modulated by local genomic backgrounds in admixed populations.

Importance:Sex and racial or ancestral disparities in Alzheimer disease remain incompletely understood; autopsy studies that examine amyloid, tau, and genetic factors are scarce. Objective:To test whether neuritic plaque burden and cognitive outcomes differ by sex and whether sex modifies the effects of apolipoprotein E ε4 (APOEε4), informant-reported race, and African ancestry. Design, Setting, and Participants:This was a cross-sectional study using postmortem neuropathological data from the Biobank for Aging Studies, University of São Paulo, São Paulo, Brazil. A total of 2268 autopsies from a population-based, diverse clinicopathological sample were collected between April 2004 and March 2025. Exposures:Sex, informant-reported race (Black, White), African ancestry proportion, and APOEε4 carrier status. Main Outcomes and Measures:Neuritic plaque burden (Consortium to Establish a Registry for Alzheimer's Disease [CERAD] score), and cognitive function (Clinical Dementia Rating-Sum of Boxes [CDR-SB]). Ordinal logistic regression examined association of sex with CERAD scores and 2- and 3-way interactions among sex, race, ancestry, and APOEε4; adjusting for age, education, vascular factors, and Braak stages. Linear models related pathology to CDR-SB, adding copathologies. Results:The analysis included 2268 autopsies (median [IQR] age, 74.8 [63.8-83.3] years; 1152 [51% male] and 1116 [49%] female; 802 [35%] Black and 1466 [65%] White; other race groups not included owing to small numbers); female individuals were older than male individuals and more likely to exhibit cognitive impairment (CDR global score ≥0.5). Female individuals had higher plaque burden than male individuals (unadjusted odds ratio [OR], 1.97; 95% CI, 1.67-2.29; P < .001), and this association remained significant in adjusted models for sociodemographic and vascular factors and APOEε4 status (adjusted OR, 1.65; 95% CI, 1.33-2.20; P < .001). APOEε4 carriers of both sexes had an approximately 4-fold greater odds of plaques. Significant 2-way interactions were found between sex, APOEε4 status, race, and ancestry on CERAD scores. Black noncarriers (OR, 0.47; 95% CI, 0.34-0.67) and noncarriers of African ancestry (OR, 0.57; 95% CI, 0.43-0.76) were least likely to have high plaque burden, whereas this protection was weakened in ε4 carriers. No significant 3-way interaction was detected. Among individuals with a CERAD score of 2 or higher, female individuals were more likely than male individuals to reach Braak stage V-VI than male individuals (probability ratio, 1.25; 95% CI, 1.13-1.38; P = .002). Adding Braak stage to multivariable models attenuated the female-male difference in plaques and interaction of sex and plaque on CDR-SB was no longer significant. Conclusions and Relevance:The findings indicate that female sex, APOEε4, and both race and African ancestry were jointly associated with amyloid in this study population. Excess amyloid among women may partly explain their greater tau burden and steeper cognitive decline. These findings highlight the importance of incorporating sex, race, and ancestry into biomarker thresholds, risk stratification, and the design of preventive or disease-modifying trials for Alzheimer disease.

INTRODUCTION:Sex differences in dementia-related neuropathology are understudied in diverse populations. METHODS:We analyzed sex differences in neuropathological and cognitive data from the Brazilian Biobank for Aging Studies. Cognitive performance was evaluated with the Clinical Dementia Rating-Sum of Boxes (CDR-SOB). Linear and logistic regression models were conducted, including interaction terms for age, race, and education. RESULTS:In 2229 participants (50.7% female, mean age ± SD 75.4 ± 12.3 years, 62.1% White), female sex was associated with a higher odds of AD pathology (Braak: odds ratio [OR] = 1.42, 95% confidence interval [CI]  = 1.17-1.72; Consortium to Establish a Registry for Alzheimer's Disease (CERAD): OR = 1.58, 95% CI = 1.27-1.98), and trans-activation response (TAR) DNA-binding protein 43 (TDP-43) (OR = 1.77, IC 95% = 1.22-2.59), and lower odds of Lewy body disease (OR = 0.69, 95% CI = 0.51-0.95). Female participant had worse cognitive performance (β = 1.56, 95% CI = 1.03-2.10). Sex modified associations of Braak, CERAD, TDP-43, and cerebral amyloid angiopathy with cognition. Age also interacted with sex and pathology on CDR-SOB. DISCUSSION:Sex differences in the associations between sex and neuropathology suggest the need for sex-informed dementia research.

Hippocampal sclerosis (HS) is characterized by neuronal loss and gliosis in the cornu Ammonis (CA) region of the hippocampus and is associated with epilepsy, hypoxia, and neurodegenerative diseases. In dementia, HS is increasingly recognized as a potential biomarker for Limbic-predominant Age-related TDP-43 Encephalopathy neuropathological change (LATE-NC). However, LATE-NC currently lacks a validated in vivo biomarker. Since HS is detectable by MRI, a key question is whether HS could serve as a reliable proxy for underlying LATE-NC pathology. This study examines the prevalence and pathological associations of HS across neurodegenerative diseases in a large population-based brain bank to assess its potential as a biomarker for LATE-NC. Data were analyzed from the Biobank for Aging Study (BAS-GEROLAB) in São Paulo, Brazil. Clinical and epidemiological information was obtained from next of kin using validated protocols, including the Clinical Dementia Rating (CDR) scale. Neuropathological assessments included morphological, vascular, and immunohistochemical analyses for beta- amyloid, tau, alpha-synuclein, and TDP-43. HS was defined as ≥70% neuronal loss in CA1 (unilateral). Individuals with a history of epilepsy were excluded. Among 1,307 individuals, 36.9% were non-White and 49.7% were women. HS was identified in 41 participants (3.1%), with higher prevalence among older individuals, women, and those with lower education levels ( p  < 0.001). HS was also significantly associated with hyaline arteriolosclerosis ( p  = 0.001) and diabetes mellitus ( p  = 0.003) (Table 1). Multivariate analysis confirmed associations between HS and LATE-NC ( p  < 0.001) as well as lacunar infarcts ( p  = 0.031) (Table 2). The positive predictive value (PPV) of HS for TDP- 43 pathology was 48.8%, increasing to 59.4% among cognitively impaired individuals ( n  = 432) (Table 3). Among individuals with HS but no TDP-43 pathology ( n  = 21), 48.4% had Braak-AD stage ≥3, and 64.5% exhibited arteriolosclerosis. HS is relatively uncommon in this population-based brain bank but is strongly associated with TDP-43 pathology. However, vascular pathology also plays a significant role, particularly in populations with high cardiovascular risk. This limits its predictive value for LATE-NC, even among cognitively impaired individuals. Given the absence of an in vivo biomarker for LATE-NC, further research is needed to determine whether MRI-detectable HS can serve as a reliable surrogate marker for LATE-NC pathology, particularly in genetically diverse populations with multiple comorbidities.

BACKGROUND:Cerebrovascular disease is prevalent in older adults and is associated with cognitive impairment. Although the association between brain infarcts and cognition has been widely investigated, the contribution of vascular disease to cognitive impairment has been less studied, particularly in non-White populations. We investigated the epidemiological and clinical characteristics of vascular disease phenotypes and their association with cognitive abilities in a diverse population. METHODS:In a Brazilian population-based clinicopathological study (recruitment between 2004 and 2024), inclusion criteria were age at death ≥18 years and the availability of an informant. Clinical information and cognitive abilities were assessed through informant interviews. Hyaline arteriolosclerosis (HA) was evaluated in 13 brain areas and categorized as absent, mild, moderate, or severe. Moderate/severe HA defined microvascular disease (Micro[+]). Intracranial atherosclerosis in the circle of Willis was similarly classified, with moderate/severe cases defining macrovascular disease (Macro[+]). Four vascular profiles were compared: Micro[+]/Macro[+], Micro[-]/Macro[+], Micro[+]/Macro[-], and Micro[-]/Macro[-]. Linear regression models adjusted for sociodemographic, clinical, and cerebrovascular/neurodegenerative lesions evaluated the association of cognition with vascular profiles, and microvascular and macrovascular diseases separately. RESULTS:Of 2418 participants, 834 with missing data were excluded, yielding a final sample of 1584 individuals (mean age, 74.3±13.4 years; 776 [49%] women; 982 [62%] White). Microvascular disease was slightly more frequent (31%) than macrovascular (29%). Regarding vascular profiles, 788 (50%) participants were Micro[-]/Macro[-], 334 (21%) Micro[+]/Macro[-], 301 (19%) Micro[-]/Macro[+], and 161 (10%) Micro[+]/Macro[+]. Micro[+] participants were older and more frequently women, and had worse cognitive abilities. Vascular groups were similar regarding most clinical comorbidities. HA (β, 0.81 [95% CI, 0.25-1.36]; P=0.004) and Micro[+]/Macro[+] phenotype (β, 1.25 [95% CI, 0.38-2.12]; P=0.005) were associated with worse cognitive abilities compared with HA-negative and Micro[-]/Macro[-] participants, respectively. CONCLUSIONS:HA was as frequent as intracranial atherosclerosis. HA and the vascular phenotype with both microvascular and macrovascular diseases were associated with worse cognition.

Clinicopathological studies suggest a role of minor cerebrovascular changes in the cognitive decline of individuals with a low neurodegenerative burden. However, it remains unclear whether small vascular brain lesions can impact cognition in middle aging individuals. Additionally, recent clinicopathological studies have shown that even a low Alzheimer’s disease (AD) neuropathological burden can significantly impact neuropsychiatric function. This study aims to evaluate the presence of AD neuropathological changes and their relationship with cognitive impairment, neuropsychiatric symptoms and risk factors in a multiethnic group under the age of 65. A post-mortem study evaluating individuals aged 30-64 years from the Biobank for Aging Studies at the University of Sao Paulo, Brazil. Neuropathological examinations were carried out based on accepted criteria, using immunohistochemistry. The clinical diagnosis was established through a postmortem interview with an informant using the Cognitive Dementia Rating (CDR). We compared the frequency of neurofibrillary tangles, neuritic plaques, lacunar infarct, hyaline arteriolosclerosis, cerebral amyloid angiopathy, synucleinopathy, and siderocalcinosis between groups. We also described the frequency of neuropathological diagnoses and neuropsychiatric symptoms. Among participants with neuropathological disease (n = 61), vascular dementia (VaD) was the most frequent diagnosis (42,7%), followed by AD (11,5%) (Figure 1). A higher frequency of vascular pathology ( p <0.001) was found in individuals with cognitive impairment (Table 2). Some degree of neurofibrillary pathology was found in 54.2% of the individuals, while neuritic plaques were found in 9.7% of the cases, regardless of the presence of cognitive impairment or neuropsychiatric symptoms. AD-type pathology burden was higher in individuals with cognitive impairment. The frequency of NPS was higher in CI individuals with significant difference in almost all subitems of the NPI, except in appetite changes. High frequency of cerebrovascular risk factors such as hypertension (60.7%) and smoking (61.1%) was also observed in the entire sample (Table 1). Our data show that AD neuropathological changes can begin in mid-age and corroborate findings from other series predominantly involving older Caucasians with high educational attainment, which have demonstrated the role of vascular pathology in cognitive impairment. As vascular risks are preventable, implementing aggressive measures to reduce these factors may impact the prevalence of cognition-related dysfunction.

Sleep dysfunction is commonly seen in Alzheimer’s disease (AD) and Progressive Supranuclear Palsy (PSP), potentially worsening these conditions. Investigating early neuropathological changes in human sleep-promoting neurons, which often precede cognitive decline, is crucial for understanding the basis for sleep dysfunction as possible treatments yet remain underexplored. We used postmortem brains of AD and PSP patients to quantify neuronal numbers and tau burden in the intermediate nucleus of the hypothalamus (IntN), VLPO analog, known for its role in sleep maintenance. Postmortem human brain tissues from three groups were used: healthy controls, progressive AD stages, and PSP. Formalin-fixed, celloidin-embedded hypothalamic tissue was cut at 30-micron. Slides were immunostained for galanin (GAL), an IntN neuronal marker, phosphorylated-tau (p-tau) antibodies, and counterstained with Gallocyanin. The total number of GAL-expressing neurons and/or p-tau deposition were estimated using the stereology. In a cohort of 7 cases (Table 1), we found a significant decrease in neuronal population and volume in the IntN in both AD and PSP patients, with a more pronounced decline in PSP. However, the neuronal density was not changed in any of the AD or PSP conditions, reflecting a simultaneous neuronal and volume loss. GAL-positive neurons were more resilient in AD, showing better preservation than in PSP. In both AD and PSP, p-tau-positive neurons increased compared to the controls. Notably, in AD, tau aggregates were present in both GAL-positive and negative neurons, while in PSP, GAL neuronal population decreased more than the others, and tau aggregates were only observed in GaL-negative neurons (Fig. 1). In both AD and PSP, our research reveals substantial neuropathological changes in the IntN, with PSP exhibiting more pronounced alterations. These changes align with observed sleep dysfunction in both conditions. PSP features severe insomnia, although wake-promoting neurons remain relatively intact, thus a significant reduction in neurons controlling NREM-sleep could account for its clinical sleep disturbances. Conversely, AD displays a decline in wake-promoting neurons and a gradual, moderate loss of NREM neurons, both leading to excessive daytime sleepiness and sleep fragmentation, yet overall preserved total sleep time. These findings deepen our understanding of NREM sleep dysfunction in tauopathies, offering insights for targeted interventions.

Identifying the molecular mechanisms underlying selective neuronal vulnerability and resilience is crucial for developing effective treatments for Alzheimer's disease (AD). Our group has shown that RORB-positive excitatory neurons are selectively vulnerable and preferentially accumulate p -tau inclusions in the entorhinal cortex (EC) at early Braak stages. However, not all RORB-positive neurons are vulnerable. By leveraging single-nucleus RNA sequencing (snRNA-seq) data, we aim to identify the molecular pathways that differentiate vulnerable and resilient RORB subtypes. We hypothesize that these biological differences may be key to understanding the mechanisms of selective neuronal vulnerability in AD. We analyzed two snRNA-seq datasets of isolated nuclei extracted from the EC of postmortem brain tissue from healthy controls and AD patients (Leng et al. 2021 [ n  = 10] and Mathys et al. 2024 [ n  = 48]). Focusing on RORB neurons, Monocle3 trajectory analysis mapped cell state progression across Braak stages. In neurons from Braaks 0-2, we performed pseudobulk and DESeq2 analysis on the first (Q1) and fourth (Q4) quartiles of pseudotime to identify differentially expressed genes (DEGs). Q1 (pseudotime = 0-5) represented resilient/healthy transcriptomes, while Q4 (pseudotime = 15-20) represented vulnerable/stressed transcriptomes. After merging datasets and subsetting excitatory neuronal nuclei ( n  = 53284), UMAP clustering identified two RORB-positive populations in the EC (Figure 1A and B). Within a RORB population (cluster 4), the trajectory analysis revealed a progression of cell states represented by an increase in pseudotime (Figure 1C). Preliminary results comparing resilient and vulnerable RORB molecular signatures revealed 537 DEGs (Figure 2A). Vulnerable neurons significantly upregulated pathways related to supramolecular fiber organization and downregulated pathways related to protein translation, targeting, and quality control (Figure 2B and C). Our findings suggest that transcriptomic signatures of vulnerable and resilient RORB excitatory neurons in the EC, in early Braak stages, may reveal key pathways leading to tau accumulation and neuronal loss. Notably, vulnerable RORB neurons upregulated cytoskeletal organization pathways and downregulated proteostasis pathways. Ongoing analyses on this RORB subpopulation and trajectory analyses on additional excitatory neuronal subpopulations in the EC will refine our understanding of the pathways that potentially lead to selective vulnerability in AD.

Frailty, characterized by increased physical vulnerability, is associated with a higher incidence and severity of cognitive impairment and also a higher burden of neurodegenerative and cerebrovascular diseases. This study investigates the association between frailty and neurodegenerative and cerebrovascular pathologies. Cross-sectional analysis using clinical and neuropathological data from individuals aged 60 or older, enrolled in the Biobank for Aging Studies between 2004 and 2023. A 42-item frailty index was constructed. Cognitive impairment was defined as a clinical dementia rating score (CDR) of 0.5 or over and participants were stratified according to cognitive status. Linear regression models, adjusting for age, sex, education and race, explored the association between frailty and neuropathology, including Alzheimer´s disease (AD), argyrophilic grain disease (AGD), Lewy-type pathology (LBP), hippocampal sclerosis, cerebral amyloid angiopathy (CAA), lacunar infarcts, hyaline arteriosclerosis, TDP-43 pathology and a neuropathological comorbidity score (NPC). We examined data from 1.343 subjects. The group with cognitive impairment was older, predominantly female, had lower education, a higher frailty index, and no race differences (Table 1). This group also exhibited a higher prevalence of all neuropathologies previously described (Table 2). In adjusted analyses, frailty was associated with AD Braak staging (β = 0.022, 95% CI=0.017; 0.028, p<0.001 ), CERAD score (β = 0.021, CI 95% = 0.013; 0.029 p<0.001), CAA (β = 0.065, 95% CI = 0.039; 0.092 p<0.001), LBD (β = 0.051, 95% CI = 0.027; 0.075 p<0.001), hippocampal sclerosis (β= 0.053, CI 95% = 0.008; 0.100 p=0.022), lacunar infarcts (β=0.071, CI 95%= 0.046; 0.097 p<0.001), siderocalcinosis (β=0.033, CI 95%= 0.015; 0.051 p<0.001), hyaline arteriosclerosis (β=0.065, CI 95%= 0.050-0.081 p<0.001) and NPC score (Table 3) (β=0.032, CI 95%= 0.024; 0.040 p<0.001). Frailty was not associated with AGD and TDP-43. Frailty was associated with several neuropathological markers of neurodegenerative and cerebrovascular diseases. More studies are warranted to investigate how this association relates to relevant outcomes such as cognitive impairment.

Excessive daytime sleepiness is a common and early symptom of Alzheimer’s disease (AD). The subcortical wake-promoting neurons in the lateral hypothalamic area, tuberomammillary nucleus (TMN), and locus coeruleus synchronize to maintain wakefulness/arousal. Although significant neuronal decline occurs in wake-promoting regions, the TMN histaminergic neurons remain relatively more intact than orexinergic and nor-adrenergic neurons. The preserved histaminergic neurons could be a potential target for addressing sleep dysfunction when neurons in the wake-promoting system degenerate. We aimed to map neuropathological and molecular events in poorly understood histaminergic neurons across AD progression in the human TMN to help understand pathogenic features and guide therapeutic strategies. We used unbiased stereology and double-immunohistochemistry to quantify pTau (CP13) accumulation, the number of histaminergic, and total neurons in the TMN in subjects across progressive Braak stages (n = 20). Data were analyzed using the Wilcoxon signed-rank test. We used a customized Neuropathology nCounter® (Nanostring) panel for proteomic analysis. Wald statistical test was used to compare the groups, and the genes were considered differentially expressed when the p-value was <0.05. TMN total neuronal count remained constant across Braak groups (BG), underscoring TMN’s resilience to AD compared to other wake-promoting neurons. Histaminergic (HDC+CP13-) neurons declined between BG 0-2 and 5-6 (p = 0.013). The number, proportion, and density of pTau inclusions in histaminergic and total TMN neurons increased across BG (p<0.05) (Fig. 1, Table 1). In Braak 5-6 over 0-2, we found 284 differentially expressed genes in TMN, of which 171 were upregulated. Gene ontology analysis demonstrated upregulation of cytokine-cytokine receptor interaction pathways (p = 0.015). In Braak 5-6 over 0-2, histamine decarboxylase (HDC) expression was downregulated (lfc = -0.819, p = 0.37) with increased expression of histamine receptors HRH1 (lfc = 1.26, p = 0.028) and HRH2 (lfc = 1.76, p = 0.019) (Fig. 2). Stereological data revealed that the decline in histaminergic neurons is associated with pTau accumulation and reduced histamine synthesis rather than neuronal loss. This is corroborated by a decrease in HDC expression and an upregulation of histaminergic receptor expression (potentially compensatory) in late Braak stages. Interventions focused on pTau removal may succeed in reinstating histaminergic neuro-transmission/-modulation, improving cognition, and restoring sleep-wake dysfunction in AD patients.

The neuromodulatory subcortical system (NSS) is vital for homeostatic balance and is among the earliest regions to accumulate tau pathology in Alzheimer's disease (AD), undergoing marked degeneration by late-stage AD. Within the NSS, orexinergic neurons (Orx N ) regulate wakefulness, appetite, and sleep-wake transitions. Experimental studies suggest that orexin dysfunction exacerbates AD, and recent clinical trials targeting the orexin pathway show improvements in AD biomarkers. However, the extent and timing of Orx N loss in human AD patients, as well as the molecular mechanisms driving early Orx N degeneration, remain unclear. We hypothesize that elucidating Orx N vulnerability at early AD stages is essential for advancing orexin-targeted therapies. We used unbiased stereology on postmortem human brains across Braak stages 0–VI ( n  = 38). Hypothalamic sections were immunostained for orexin-A and phosphorylated tau (CP13) antibodies, then counterstained with Nissl. We performed RNA sequencing of the lateral hypothalamic area (LHA) ( n  = 38), which houses all Orx N -using the NanoString nCounter ® platform, employing panels for neurotransmitter-related genes, glial profiling, and neuropathology. Genes with padj<0.05 were deemed differentially expressed. Stereological counts showed a 43% reduction in Orx N at Braak I versus 0 (padj=0.0117), signifying the earliest massive neuronal loss in AD. After a plateau at Braak II-IV, Orx N declined by 70% at Braak V-VI versus Braak 0 (padj=0.0062). A 50% drop in Orx N at Braak II correlated with differential expression of genes related to lysosomal function, glial reactivity, oxidative stress, and phosphorylation. GO/KEGG analyses revealed enrichment of T-cell-mediated inflammatory, synaptic vesicle, and organelle dysfunction pathways. Despite extensive Orx N loss, orexin-related genes HCRT (padj=0.015) and HCRTR1 (padj=0.0166) were upregulated at Braak III-IV. Orexinergic neurons are the earliest neuronal population to degenerate in AD, starting at Braak I. Early p -tau accumulation in the LHA, independent of major β-amyloid pathology, triggers T-cell-mediated inflammation, lysosomal dysregulation, and oxidative stress while surviving Orx N attempts compensatory gene upregulation. Overall, this highlights Orx N as a previously underrecognized target for early intervention, potentially offering both symptomatic and disease-modifying benefits. Given the availability of orexin-targeting pharmacotherapies, our data support expanded exploration of orexinergic modulation as a viable AD therapeutic strategy.

Selective neuronal vulnerability is an underappreciated but critical factor in understanding Alzheimer's disease (AD) pathogenesis, disease progression, and neuroprotective mechanisms. Given the unique characteristics of the human brain, studying selective vulnerability in postmortem human tissue offers invaluable insights. Our group has demonstrated a direct link between subcortical neuronal degeneration in sleep-wake and circadian-modulating regions and clinical function, highlighting these structures as key to investigating selective vulnerability. Circadian control is primarily governed by the anterior hypothalamus, where the suprachiasmatic nucleus (SCN) acts as the brain's master clock. The SCN and neighboring structures, including the paraventricular nucleus (PVN) and supraoptic nucleus (SON), share a predominant population of arginine vasopressin-expressing (AVP+) neurons, which are integral to circadian function. To assess whether these three nuclei provide a platform to study selective vulnerability in AD, we quantified neuronal populations and AD pathology burden in postmortem human brains. We analyzed postmortem anterior hypothalamic tissue, including the SCN, PVN, and SON, from 12 controls (Braak stage 0) and 28 AD cases spanning mild to severe pathology (Braak stages I, II, and VI). Using fluorescence in situ hybridization, we probed AVP+ neurons in these nuclei and quantified neuronal estimates and AVP+ neuronal area. A custom 2D image registration method was used to quantify amyloid plaques and neurofibrillary tangles in adjacent sections. AVP+ neuronal loss was evident in the SCN as early as Braak stage II, whereas AVP+ neurons remained stable in the PVN and SON. Both the SCN and PVN exhibited phosphorylated tau inclusions, while amyloid plaques were detected in the PVN but absent in the SCN and SON. The SON displayed no evidence of amyloid plaques or tau accumulation, suggesting resistance to AD pathology. Our findings reveal that among these three AVP+ nuclei, the SCN is particularly vulnerable to early AD-related neurodegeneration, whereas the PVN accumulates tau and amyloid pathology without significant neuronal loss. The SON appears relatively resistant to AD pathology. These differential patterns suggest that intrinsic properties of neuronal populations and local microenvironments may contribute to selective vulnerability, providing a unique model to explore resilience mechanisms in AD.

Alzheimer’s Disease(AD) patients experience circadian rhythm disorder. The circadian rhythm is synchronized by a master clock, the suprachiasmatic nucleus(SCN), which is spatially well-conserved but a tiny nucleus in the hypothalamus. Little is known about the molecular and pathological changes that occur in the SCN during AD progression. We examined postmortem brains of 12 controls without AD neuropathological changes (Braak0) and 39 subjects with progressive ADNC stages. We quantified neuronal numbers-arginine vasopression(AVP) and vasoactive intestinal protein(VIP) neurons- plus ADNC burden in SCN. We performed analysis in adjacent hypothalamic nuclei as regional controls: supraoptic nucleus(SON) and paraventricular nucleus(PVN) both sites of AVP+ neurons. Moreover, we performed In-situ proteomics using GeoMx Digital Spatial Profiling(DSP) in the three nuclei (total of 897 ROIs), including probes for 39 proteins commonly dysregulated in AD. SCN neurons in Braak 6 had sixteen times higher p-tau levels than Braak0. Neurofibrillary tangles were found exclusively in SCN(Fig. 1). However, there was not significant p-tau upregulation in SON and PVN in all the same stage. Additionally, the SCN showed increases in glial proteins already in Braak1(Fig. 2), whereas these proteins remained unaltered in the other nuclei, including that they displayed a milder pattern of protein dysregulation in Braak1, consistent with its lower tau expression observed in histology and DSP. Our study sheds light on the previously unexplored molecular and pathological changes occurring in SCN during AD progression. The SCN is vulnerable to AD-tau pathology and show immune dysregulation even at Braak1 but protected against beta-amyloid accumulation. This vulnerability pattern in SCN supports the idea that SCN dysfunction contributes to circadian rhythm disturbances in AD, observed even in the stage before the onset of cognitive disorder. Furthermore, the preservation of SON, a neighboring nucleus with AVP neurons but not directly connected in a neuronal circuit with the SCN, corroborates that SCN is early vulnerable to AD. PVN neurons exhibited similar but milder pattern of protein dysregulation in the early stages to the SCN, implying the effect may be caused by the efferent projection from the SCN. These results open up opportunities for tailored interventions to alleviate circadian rhythm disruptions in AD.

INTRODUCTION:Alzheimer's disease (AD) selectively affects certain brain regions, yet the mechanisms of selective vulnerability remain poorly understood. The neuromodulatory subcortical system, which includes nuclei exhibiting a range of vulnerability and resilience to AD-type degeneration, presents a framework for uncovering these mechanisms. METHODS:We leveraged transcriptomics and immunohistochemistry in paired samples from human post mortem tissue representing a vulnerable and resilient region-the locus coeruleus (LC) and substantia nigra (SN). These regions have comparable anatomical features but distinct vulnerability to AD. RESULTS:We identified significant differences in cholesterol homeostasis, antioxidant pathways, KRAS signaling, and estrogen signaling at a bulk transcriptomic level. Notably, evidence of sigma-2 receptor upregulation was detected in the LC. DISCUSSION:Our findings highlight pathways differentiating the LC and SN, potentially explaining the LC's selective vulnerability in AD. Such pathways offer potential targets of disease-modifying therapies for AD. HIGHLIGHTS:Intraindividual comparative RNAseq was used to study selective vulnerability. Metallothionein genes are significantly enriched in the substantia nigra. Cholesterol homeostatic genes are significantly enriched in the locus coeruleus. The locus coeruleus is likely more susceptible to toxic amyloid beta oligomers.

Alzheimer's disease (AD) neuropathological changes (ADNC) are characterized by amyloid plaques and neurofibrillary tangles. The role of nitric oxide synthase (NOS) in disease progression remains unclear. This study investigates the expression of neural, inducible, and endothelial NOS (nNOS, iNOS, eNOS) and 3-nitrotyrosine (3-NT) in the hippocampal subregions of individuals with ADNC and their association with cognitive abilities. Immunohistochemistry was performed on hippocampal samples from 10 individuals with ADNC and 10 age- and sex-matched controls to detect NOS enzymes, and 3-NT expression in CA1, CA2, CA3, and CA4. Logistic ordinal regressions evaluated associations of NOS and 3-NT expression with AD pathology, while linear regressions assessed relationships with cognitive abilities. Overexpression of nNOS was observed in all hippocampal subregions in ADNC participants. iNOS expression was elevated in CA1 and CA3, while eNOS showed increased levels only in CA3. 3-NT was significantly higher in CA3 of ADNC participants. nNOS expression in all hippocampal regions correlated with AD pathology and cognitive impairment. iNOS in CA3 was associated with AD pathology and cognitive scores. No associations were found for eNOS, and 3-NT in CA3 correlated with cognitive impairment. Associations of nNOS and iNOS with neuropathology and cognition suggest a role for NOS in AD pathophysiology.

BACKGROUND:Hippocampal sclerosis (HS) is characterized by neuronal loss and gliosis in the cornu Ammonis (CA) region of the hippocampus and is associated with epilepsy, hypoxia, and neurodegenerative diseases. In dementia, HS is increasingly recognized as a potential biomarker for Limbic-predominant Age-related TDP-43 Encephalopathy neuropathological change (LATE-NC). However, LATE-NC currently lacks a validated in vivo biomarker. Since HS is detectable by MRI, a key question is whether HS could serve as a reliable proxy for underlying LATE-NC pathology. This study examines the prevalence and pathological associations of HS across neurodegenerative diseases in a large population-based brain bank to assess its potential as a biomarker for LATE-NC. METHOD:Data were analyzed from the Biobank for Aging Study (BAS-GEROLAB) in São Paulo, Brazil. Clinical and epidemiological information was obtained from next of kin using validated protocols, including the Clinical Dementia Rating (CDR) scale. Neuropathological assessments included morphological, vascular, and immunohistochemical analyses for beta- amyloid, tau, alpha-synuclein, and TDP-43. HS was defined as ≥70% neuronal loss in CA1 (unilateral). Individuals with a history of epilepsy were excluded. RESULT:Among 1,307 individuals, 36.9% were non-White and 49.7% were women. HS was identified in 41 participants (3.1%), with higher prevalence among older individuals, women, and those with lower education levels (p < 0.001). HS was also significantly associated with hyaline arteriolosclerosis (p = 0.001) and diabetes mellitus (p = 0.003) (Table 1). Multivariate analysis confirmed associations between HS and LATE-NC (p < 0.001) as well as lacunar infarcts (p = 0.031) (Table 2). The positive predictive value (PPV) of HS for TDP- 43 pathology was 48.8%, increasing to 59.4% among cognitively impaired individuals (n = 432) (Table 3). Among individuals with HS but no TDP-43 pathology (n = 21), 48.4% had Braak-AD stage ≥3, and 64.5% exhibited arteriolosclerosis. CONCLUSION:HS is relatively uncommon in this population-based brain bank but is strongly associated with TDP-43 pathology. However, vascular pathology also plays a significant role, particularly in populations with high cardiovascular risk. This limits its predictive value for LATE-NC, even among cognitively impaired individuals. Given the absence of an in vivo biomarker for LATE-NC, further research is needed to determine whether MRI-detectable HS can serve as a reliable surrogate marker for LATE-NC pathology, particularly in genetically diverse populations with multiple comorbidities.

Sex differences have been investigated in Alzheimer's disease neuropathological changes (ADNC). However, few studies analyzed the occurrence of multiple co-pathological changes. This study examined sex differences in AD neuropathology, co-pathology, and cognitive abilities in a diverse sample. Participants were from the Biobank for Aging Studies, a large population-based autopsy study in Brazil. The neuropathological assessment includes AD pathology (neuritic plaques and neurofibrillary tangles), TDP-43, Lewy body pathology, lacunar infarct, hyaline arteriolosclerosis, and cerebral amyloid angiopathy. Cognitive abilities were evaluated using the Clinical Dementia Rating Sum of Boxes (CDR-SOB). We used ordinal logistic and linear regressions to investigate the associations of sex, education, and race with neuropathological and cognitive data adjusted for sociodemographic and clinical variables. Among the 711 participants, 324 were men, and 387 were women; the mean age was 74.2±13.1 years old, and 67.4% were white. Women were older, less educated, and exhibited poor cognitive abilities compared to men in CDR-SOB (β 5.8 CI=0.0-15.0, p  <0.001). Women showed a more significant frequency of AD pathology and co-pathologies ( p  <0.001). The most frequent co-pathologies were: Poorer cognitive abilities in women were associated with the number of co-pathologies associated with ADNC, even when data was adjusted by age of death, education, race, hypertension, dyslipidemia, and diabetes. Women had poor cognitive abilities and higher association of ADNC and co-pathologies than men.

Neuropsychiatric symptoms (NPS) are common in neurodegenerative disorders and may precede cognitive decline over several years. Investigating the relationship between neuropathological lesions and NPS in young and middle-aged adults can provide critical insights into the biological underpinnings of NPS and their potential as early markers of neurodegenerative disease. This population-based post-mortem study analyzed neuropathological data from individuals aged 30–64 years, including assessments for neurofibrillary tangles (NFT), amyloid-β burden, Lewy body (LB) pathology, TDP-43, lacunar infarcts, cerebral amyloid angiopathy (CAA), and hyaline atherosclerosis. Post-mortem interviews provided information on NPS using the Neuropsychiatric Inventory and cognitive status using the Cognitive Dementia Rating (CDR). We evaluated the relationship between neuropathological substrates and seven common neuropsychiatric symptoms (depression, agitation, apathy, irritability, hallucinations, delusions, anxiety) using models adjusted for sociodemographic and clinical variables, and investigated the frequency of NPS in individuals with and without cognitive impairment. In 413 cases (mean age 56.7±6.1yo, 36.3% women, 60.3% white), NPS were more prevalent in individuals with cognitive impairment than in those without (Figure 1). Depression and anxiety were associated with Braak NFT stages, anxiety was associated with cerebral amyloid angiopathy, while agitation, irritability, and delusions were associated with the presence of lacunar infarcts (Table 1). Our findings highlight the relationship between vascular lesions and neuropsychiatric symptoms (NPS).

Previous studies suggest an association between Alzheimer’s disease and carotid artery atherosclerosis. However, the association between atherosclerotic carotid plaque composition and Alzheimer’s disease pathology (neuritic plaques and neurofibrillary tangles) has not been explored yet. Carotid arteries were dissected and the segments with the largest obstruction in the carotid bifurcation, and the common and internal carotid arteries were obtained. Each segment was immersed in paraffin and stained using Hematoxylin-Eosin (HE) and Masson’s Trichrome. Each histological slide was photographed using a stereomicroscope (SMZ 1000; Nikon). We analyzed artery obstruction, fibrous cap, the number of vasa vasorum, and plaque components, using the software Atherosclerotic Plaque Analyzer. We characterized the neuritic plaque deposition in the brain using the Consortium to Establish a Registry for Alzheimer’s Disease (CERAD) criteria and neurofibrillary tangle deposition using the Braak & Braak criteria. We performed ordinal logistic regression models adjusted for age, sex, education, hypertension, diabetes, dyslipidemia, smoking, excessive alcohol consumption, body mass index, and physical inactivity. Cross-sectional data from 237 subjects from the Biobank for Aging Studies of the University of Sao Paulo Medical School were analyzed (mean age = 79.0±9.9 yo, 51.5% women, 59.9% white, 35.9% black and 3.8% Asian). As the percentage of smooth muscle cell area increased, neurofibrillary tangles deposition decreased (OR = 0.97, 95% CI = 0.94-1.00, p = 0.03). Similarly, as the percentage of fibrous cap area increased, neurofibrillary tangles deposition increased (OR = 1.13, 95% CI = 1.00-1.25, p = 0.04). Given that the fibrous cap is typically present in advanced atherosclerosis, and considering that smooth muscle cell area was solely assessed in the absence of well-established atherosclerotic plaque, our examination of atherosclerotic plaque composition in carotid artery segments suggests a connection between atherosclerosis development and the accumulation of neurofibrillary tangles in the brain. We found an association with the fibrous cap proportion of the tunica intima area and an inverse association with the proportion of smooth muscle cells in the tunica intima area.