Lewy bodies, the defining pathological feature of Parkinson's disease, are intraneuronal inclusions enriched in aggregated alpha-synuclein (αSyn). We used correlative light and electron microscopy to selectively investigate phosphorylated αSyn (αSynpS129)-positive inclusions in the substantia nigra of end-stage postmortem Parkinson's disease brain. Here we show that somatic αSynpS129 inclusions in nigral dopaminergic neurons are consistently fibrillar, whereas the membranous-type inclusions are restricted to neuritic processes. These neuritic inclusions displayed marked ultrastructural heterogeneity, ranging from predominantly membranous to mixed membranous-fibrillar forms. The selective targeting of defined inclusions enabled detailed structural characterization of Lewy pathology, rather than quantitative or disease-stage comparisons. Our findings highlight clear ultrastructural differences between somatic and neuritic αSynpS129 pathology and demonstrate the structural complexity and heterogeneity of Lewy pathology in human Parkinson's disease brain.
Methotrexate (MTX) is a well-known medication for the treatment of different cancer types and autoimmune diseases. The current study target was to measure the capability of human placental extract (HPE) to ameliorate the nephrotoxicity induced by MTX in male albino rats. In the present study, rats were distributed into four groups; a control group (each rat was intraperitoneally injected with 0.5 ml of 0.9% NaCl daily for five days), HPE-treated group (HPE, 10.08 mg/Kg b.w/day, was subcutaneously injected for two weeks), MTX-treated group (MTX, 5 mg/Kg b.w/day, was intraperitoneally injected for five days) and MTX and HPE-treated group (Both MTX and HPE were injected to rats at the same time with the same doses, duration and injection routes in MTX and HPE groups). During the experimental period, clinical observations and body weights of rats were recorded. Rats were dissected after twenty-four hours from the last dose of each group, blood samples were collected for relative blood viscosity measurements and kidneys were also collected for biochemical, ultrastructural and dielectric properties (dielectric constant, dielectric loss and conductivity) investigations. MTX treatment resulted in a highly significant decrease in rat body weights, a highly significant decrease in glutathione (GSH) level and catalase (CAT) activity, a significant decrease in superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities and a highly significant increase in the malondialdehyde (MDA) level and relative blood viscosity compared to the control group. Besides, obvious ultrastructural changes and pronounced decrease in the dielectric properties of kidney tissues were noticed. While HPE treatment with MTX improved body weight, biochemical, ultrastructural and biophysical changes comparing to the MTX group. Human placental extract can reduce MTX-induced nephrotoxicity in rats through boosting oxidative stress/anti-oxidant balance as it is rich with essential elements.
Inflammation within the central nervous system (CNS) plays a pivotal role in neuronal survival and degeneration. Lipopolysaccharide (LPS) is a widely used agent for inducing systemic and localized inflammation in mammals, providing a model for studying neurodegenerative processes. While previous research has documented neuronal loss due to LPS-induced neurodegeneration, the progressive morphological changes in neurons remain insufficiently characterized, particularly in retinal tissues. This study addresses this gap by establishing acute and chronic retinal inflammation models in mice using single and repeated intraperitoneal LPS injections. Through ultrastructural analyses using electron microscopy, we observed significant pathological changes in retinal neurons, glial cells, and blood-retinal barrier (BRB) components. Acute LPS exposure resulted in lipid droplet accumulation and membrane disruption in retinal pigment epithelium (RPE), as well as abnormal neuronal and vascular ultrastructures. Chronic LPS exposure amplified these effects, causing more pronounced damage to neurons and exacerbating BRB dysfunction. This study provides, for the first time, detailed ultrastructural insights into LPS-induced acute and chronic retinal inflammation. These findings advance our understanding of retinal pathology in inflammatory conditions and support the development of novel therapeutic strategies for retinal and CNS neurodegenerative diseases.
Astrocytes are key regulators of neuronal, metabolic, and vascular homeostasis, yet their morphological diversity and involvement in alcohol-related brain pathology remain incompletely characterized. In this study, we investigated astrocytic morphology in the human striatum of control individuals and subjects with short- and long-term alcohol exposure using immunohistochemistry combined with Sholl-based morphometric analysis, and ultrastructural assessment. GFAP immunohistochemistry was used to identify astrocytes, assess their morphology, and manually quantify GFAP+ cells in gray and white matter, followed by Sholl-based morphometric analysis to characterize astrocytic branching architecture and spatial organization. The number of GFAP+ astrocytes differed between tissue compartments, with a significant increase in white matter in alcohol-exposed individuals and no detectable change in gray matter. Morphometric analysis revealed pronounced astrocytic heterogeneity across all study groups. Sholl-derived metrics supported the distinction of six recurrent astrocytic morphometric profiles in the human striatum, distinguished by soma size, branching complexity, process length, and cell territory size. These profiles were present across gray and white matter, indicating intrinsic astrocytic structural diversity. Ultrastructural analysis further revealed alcohol-associated alterations at the astrocyte-vascular interface, including swelling of perivascular astrocytic endfeet, accumulation of intermediate filaments, and focal reductions in vascular wall coverage. Together, these findings demonstrate substantial astrocytic structural diversity in the human striatum accompanied by alcohol-related gliovascular remodeling.
Autism spectrum disorder (ASD) is a neurodevelopmental disorder that typically appears in early childhood, and there is a link between ASD and zinc deficiency. Neither the potential protective effects of the trace element zinc on the cerebellum against ultrastructural damage in animal models of ASD, nor its ability to protect against the modulation of the oxidative stress/gliosis/neuronal loss axis implicated in autism, have been previously investigated. Young rats were injected with the short-chain fatty acid produced by gut bacteria that is known to induce ASD, propionic acid (PPA) (500 mg/kg) for 5 days (model group) and the protection group was treated with 2 mg/kg zinc sulfate for two weeks following PPA injections. Cerebellum damage and behavioral impairment developed in the model group of rats as demonstrated by profound cerebellar ultrastructural alterations such as degeneration of Purkinje neurons and fragmented endoplasmic reticulum, as well as anxiety-like behavior and social interaction deficits. PPA also significantly (p < .05) modulated the cerebellar protein levels of the gliosis biomarker GFAP and the neuronal loss biomarker calbindin D28K, as well as the cerebellar tissue levels of zinc, glutathione, and the oxidative stress biomarker malondialdehyde (MDA). All these parameters were significantly (p < .05) protected by zinc treatment. Additionally, we observed a significant (p < .001) correlation between the degree of Purkinje cell damage and these parameters associated with autism. In conclusion, PPA-induced autistic behaviors in rats are associated with alteration to the cerebellar ultrastructure, modulation of the oxidative stress/gliosis/neuronal loss axis of ASD, and a decline in zinc and glutathione, which were protected by zinc treatment.
Sperm cryopreservation is a common procedure in assisted reproduction (ART) laboratories. Although generally regarded as an efficient and safe technique, a decrease in the fertilization potential has been described. Hence, a detailed ultrastructural analysis of human spermatozoa might provide important insights into cellular alterations occurring during cryopreservation. To address this question, the ultrastructure of human spermatozoa was analysed, before and after cryopreservation, for the time periods of one month (1-Mo) and seven months (7-Mo), using paired, repeated measures. Semen samples were cryopreserved with a rapid freezing protocol, using a glycerol- and raffinose-containing cryopreservation medium, allocated into cryo-straws and stored in liquid nitrogen. Ultrastructural results indicate a significant increase in anomalous sperm, with changes in the head membrane and acrosomal vesicle, after both cryopreservation periods (fresh:26.8% vs. 1-Mo:52.2%, 7-Mo:57.4%, P < 0.001, partial η²=0.849), suggesting a concurrent cryo-damage to these structures. Concerning the sperm head membrane, a significant increase in total detachment was observed, after 1 and 7 months of cryopreservation (fresh:9.3%, 1-Mo:26.3%, 7-Mo:34.5%, P < 0.001, partial η²=0.727); for the acrosomal vesicle, a significant increase was observed in apical detachment after 1 month (fresh:11.4%, 1-Mo:22.9%, 7-Mo:17.1%, P = 0.006, partial η²=0.433) and in total detachment after 7 months (fresh:24.9%, 1-Mo:31.8%, 7-Mo:43.6%, P < 0.001, partial η²=0.636). In conclusion, our results indicate that both the sperm head membrane and the acrosomal vesicle are affected by cryopreservation, with both potentially decreasing the functional competence of the spermatozoa. These data adds information about cryopreservation, highlighting the importance of detailed inspection for sperm selection during ART. (244/250).
Cerebral toxoplasmosis is a common opportunistic parasitic infection of the CNS caused by the Toxoplasma gondii parasite. Host immunosuppression can affect disease outcomes. To explore the changes in the cerebral cortical ultrastructure accompanying the infection in different immune-altered models and to find an effective treatment against the infection, we tested the possible therapeutic effect of clofazimine (CFZ) (the FDA-approved antimycobacterial drug) against the infection using 60 male CD1 Swiss Albino mice divided into 6 groups: 3 dexamethasone (DEX)- treated groups (DEX-only, DEX-infected, and DEX-infected-treated), and 3 streptozotocin (STZ)-induced type 1 diabetic groups (STZ-only, STZ-infected, STZ-infected-treated). The worst ultrastructural changes were observed in the diabetic and diabetic-infected groups, characterized by a significant increase in neuronal apoptotic and necrotic nuclei (P < 0.05) and changes in the numbers and structure of glial cells compared to the DEX and DEX-infected groups. CFZ (at a dose of 10 mg/kg/day for 3 days starting on 45th day post infection) significantly improved cortical neuronal ultrastructural changes in both models (P < 0.05), reduced microglial numbers, increased astrocyte numbers, and restored brain capillary integrity and axonal growth, in addition to significantly reducing mature cyst numbers in both models (P < 0.05). However, the drug didn't reduce the number of atrophic and necrotic cysts in the infected-treated groups. So, in our study, CFZ showed preclinical promise in treating experimental cerebral toxoplasmosis and reducing the parasitic cyst burden, highlighting the adverse impact of the host's altered immune status on brain tissue and the course of the infection, especially in diabetes.
Desmin-related cardiomyopathy is a rare disorder caused by pathogenic variants in desmin and associated protein genes. We aimed to describe the clinical, histopathological, immunohistochemical, ultrastructural, and molecular features in a pediatric cohort. This retrospective descriptive study reviewed pediatric cases diagnosed over a 30-year period. Clinical data were extracted from electronic medical records. Archived histologic slides (H&E, desmin immunostains, and special stains) and electron photomicrographs were re-evaluated. Key features were documented and summarized. Four pediatric patients (3 males and 1 female) presented between ages 2 and 14 years with restrictive cardiomyopathy and conduction abnormalities. Endomyocardial biopsies and/or explanted heart specimens showed eosinophilic cytoplasmic inclusions that were desmin-immunoreactive, PAS-negative, and highlighted by toluidine blue. Electron microscopy revealed dense cytoplasmic granulofilamentous aggregates that were frequently continuous with the sarcoplasmic membrane, Z-bands, and intercalated discs. Two patients harbored the same pathogenic variant: DES c.1360T>C (p.Arg454Trp). Desmin-related cardiomyopathy should be considered in any child presenting with restrictive cardiomyopathy and/or rhythm disturbances. Distinctive light microscopic and ultrastructural features can aid in confirming the diagnosis. Certain pathogenic variants are increasingly linked to more severe phenotypes, highlighting the importance of genetic evaluation and its implications for family counseling.
Lung fibrosis is an irreversible life-threatening condition. So, this study directed to investigate the structure of lung tissue following bleomycin-induced pulmonary injury and to evaluate the potential protective effects of pirfenidone, granulocyte colony-stimulating factor (GCSF), and their combined administration. Fifty-six adult rats were allocated into control, bleomycin-treated, pirfenidone-treated, GCSF-treated, and combination groups. Lung specimens were processed and examined using light & transmission electron microscopes beside biochemical examination for MDA and GSH. Bleomycin administration caused marked structural alterations. Both Pirfenidone and GCSF alone resulted in partial improvement of pneumocyte ultrastructure and reduced fibrotic features. Notably, the combined therapy group demonstrated the most evident protection, with near-normal pneumocyte morphology and reduced degenerative changes. These findings suggest that combined pirfenidone and GCSF therapy exerts a synergistic effect in ameliorating bleomycin-induced lung injury at the ultrastructural level.
Adult neural stem cells (NSCs) are located in the sub-granular zone (SGZ) of the dentate gyrus (DG) of the hippocampus, one of the prominent regions of neurogenesis in the adult brain. Activation of endogenous stem cells may provide a therapeutic approach in demyelinating diseases. Due to their minuscule size, nanoparticles (AuNPs) can cross the blood-brain barrier. To evaluate the effect of AuNPs on adult NSCs and the progression of demyelination in the hippocampus in a model of demyelination, thirty-two adult mice were divided into: control group I, AuNPs group II (10 mg/kg BW/day), cuprizone (CPZ) group III (0.2% orally for 8 weeks to induce demyelination), and CPZ & AuNPs group IV. Hippocampus was prepared for light microscopy, nestin immunohistochemical staining (NSCs' immune marker), and electron microscopy. In groups I and II, spindle-shaped adult NSCs in the SGZ of DG showed a positive nestin immunoreaction. In group III, most of the nerves were demyelinated. Pyramidal and granular neurons showed degenerative changes. Nestin-immune-positive cells were significantly decreased. In group IV, preservation of the myelin, less degenerative changes, and a significant increase in nestin-positive cells were detected. In conclusion, AuNPs activated adult NSCs in the mouse hippocampus, partially ameliorated degenerative changes, and enhanced partial remyelination in the CPZ demyelination model.
C3 glomerulopathy (C3G) is a rare type of glomerular disease characterized by predominant deposits of C3 complement. Although dominant C3 immunofluorescent (IF) staining has become well known as a diagnostic criterion, the findings of electron microscopy (EM) are not well specified. The goal of this study was to scrutinize the characteristic features of C3 deposits on EM and to correlate with C3 IF staining patterns. We examined the EM images of a cohort of 24 cases of C3G (22 C3 dominant glomerulonephritis [C3GN] and 2 dense deposit disease [DDD]) to determine their characteristic features when compared to a cohort of patients with immune complex-mediated glomerulonephritis. In our patients with C3G, the C3 deposits were present in all three glomerular compartments (mesangial, subendothelial, and subepithelial spaces) and they had the following features: (1) Smear pattern of C3 deposits along glomerular basement membranes (GBM) and subepithelial spaces; (2) C3 deposits were mostly lighter in gray colors as opposed to the dark black appearance of immune complex deposits (ICD); (3) C3 aggregates revealed smooth contours with a homogeneous fine granular appearance when compared to the humpy and bumpy appearance of ICD, and (4) C3 deposits rarely showed either retraction artifacts around the deposits or vacuolization within the GBM or mesangial areas. In our cohort, C3 deposits exhibit reproducible EM features that are well correlated with clinical data and dominant C3 staining by IF and are different from ICD upon securitized review.
Ischemic stroke triggers severe disruption of the neurovascular unit (NVU), with pathology evident at the ultrastructural level. Hypoxia-inducible factor-1α (HIF-1α) activators such as Prolyl hydroxylase inhibitors e.g., dimethyloxalylglycine (DMOG) and histone deacetylase inhibitors (e.g., Apicidin) are promising therapeutic candidates, but their specific impact on subcellular architecture remains poorly characterized. Endothelin-1 (ET-1) model was used to induce ischemic stroke. Animals were treated intraperitoneally with either DMOG (40 mg/kg), Apicidin (1 mg/kg), or vehicle at reperfusion onset. After 7 days, behavioral analysis was conducted, Cytochrome-C immunohistochemistry and cortical tissue was processed for transmission electron microscopy (TEM) to assess ultrastructural integrity. Severe ultrastructural pathology, such as pyknotic neuronal soma, broken axons, enlarged mitochondria, synaptic disintegration, and reactive microglia and oligodendrocytes, was seen in model animals. With relatively slight improvements in synaptic and glial function and ongoing somatic, axonal, and mitochondrial damage, DMOG therapy offered little protection for neurons. With intact soma and axons, decreased mitochondrial swelling, increased synaptic vesicle density, and enhanced oligodendrocyte and microglial morphology, Apicidin therapy, on the other hand, showed strong neuronal preservation. While Apicidin is more effective at maintaining the integrity of neurons and synapses, DMOG mainly stabilizes the vascular compartment. These results imply that better neuroprotection may be provided by combination therapy that targets both vascular and neuronal components.
Natural products have historically been integral to pharmacotherapy, attributed to their remarkable structural diversity and evolutionary refinement for biological interactions. Nonetheless, traditional natural product-based drug discovery has faced challenges such as mechanistic ambiguity, scalability limitations, and inadequate translational predictability. Concurrently, reductionist single-target approaches have been insufficient for addressing complex diseases characterized by network-level dysregulations. Recent advancements in analytical chemistry, genomics, and data-driven methodologies have rejuvenated natural product research by facilitating rapid structural elucidation, systematic exploration of biosynthetic diversity, and rational prioritization of bioactive compounds. Notably, many natural products exhibit multitarget effects that necessitate interpretation beyond isolated molecular interactions. Systems pharmacology offers a quantitative framework to analyze such network-level perturbations by integrating omics data, computational modeling, and experimental validation. However, molecular and computational predictions alone do not suffice to establish therapeutic relevance. Experimental pathology, encompassing histopathology, immunohistochemistry, spatial analysis, and ultrastructural evaluation, remains essential for validating efficacy and safety at tissue and organ levels. This review synthesizes technological innovation, systems pharmacology, and pathological validation to reposition natural products as mechanistically grounded and translationally robust resources for contemporary drug discovery.
The blood-brain barrier (BBB) plays a critical role in maintaining brain homeostasis by tightly regulating molecular transport. However, its integrity is often compromised with aging and in neurodegenerative diseases, contributing to disease pathology. Studying the biological consequences of BBB dysfunction independent of concomitant pathology remains challenging, largely due to the absence of reliable and inducible animal models that avoid unintended side effects such as osmotic effects, neuroinflammation, or vascular damage. In this study, we evaluated the use of Power Doppler ultrasound (PDUS) combined with microbubbles to induce widespread, bilateral BBB opening in the mouse brain. Mice received intravenous infusions of SonoMAC microbubbles during transcranial PDUS application. BBB permeability was assessed via Evans Blue dye extravasation and immunofluorescence analysis of extravasated immunoglobulins. Vessel integrity was evaluated at the ultrastructural level using transmission electron microscopy (TEM). PDUS combined with microbubbles successfully induced widespread BBB opening, as evidenced by diffuse Evans Blue staining and immunoglobulin extravasation in coronal sections. Immunoglobulin leakage was detected in all analyzed brain regions, with lower levels in white matter, likely reflecting its lower vascular density. Leakage appeared to primarily originate from capillaries while TEM analysis revealed no overt vascular damage. These findings support PDUS with microbubbles as a non-destructive, reproducible method to model widespread BBB dysfunction. This approach offers an in vivo platform to study BBB-related pathophysiological processes such as impaired clearance, protein aggregation, and neurotoxicity, as well as for investigation of therapeutic delivery to the brain parenchyma.
Statins are a class of drugs commonly prescribed to lower cholesterol and low-density lipoprotein (LDL) cholesterol levels, as well as to reduce morbidity and mortality associated with cardiovascular illnesses. However, statins are associated with potential side effects, such as muscle pain, liver problems, and neurological or psychiatric disturbances. Therefore, a protective medication is required to mitigate the harmful effects of statins. Thymoquinone (THQ), found in Nigella sativa, has several medicinal benefits and protective effects. The current study aimed to investigate the effects of simvastatin on cerebellar and hepatic tissues, focusing on cellular apoptosis, using light and electron microscopy, morphometric analysis, and immunohistochemical techniques. Additionally, the study evaluates whether thymoquinone can protect against simvastatin-induced histological and ultrastructural damage in hepatic and cerebellar tissues. Thirty adult male Wistar albino rats were divided into three equal groups. Group I was the control group (Ctrl group). Group II (statin group) received simvastatin (20 mg/kg/day) orally for 8 weeks. Group III (statin + THQ group) received concomitant treatment with simvastatin (20 mg/kg/day) and thymoquinone (10 mg/kg/day) for 8 weeks. Tissue specimens were taken from the liver and cerebellum of all animal groups and processed for light and electron microscopic examination. An immunohistochemical investigation for caspase-3 was performed. The molecular and granular layer thickness, number of Purkinje cells, central vein diameter, hepatocyte nuclear diameter, and optical density of anti-caspase-3-positive cells were measured in both the cerebellum and the liver in all experimental groups. Results showed that cerebellum from simvastatin-treated animals showed perinuclear halos in cells within the molecular layer. Purkinje cells appeared disorganized, irregularly arranged, shrunken, with irregular outlines, hyperchromatic nuclei, and vacuolated cytoplasm. Moreover, the granular cell layer appeared shrunken with irregular hyperchromatic nuclei. In addition, decreased thickness of both the molecular and granular layers, as well as a reduction in the number and diameter of Purkinje cells, was observed and confirmed by morphometric analysis. Liver tissue showed shrunken hepatocytes, widened blood sinusoids, widened central veins, and inflammatory cell infiltration. Furthermore, strong positive expression of caspase-3 was observed in both the cerebellum and liver. However, all these histological, ultrastructural, and morphometric changes were markedly decreased after the addition of thymoquinone. It can be concluded that simvastatin induces significant degeneration in both the cerebellar cortex and liver tissue, primarily due to increased oxidative stress and reduced antioxidant protection. Thymoquinone exerts protective effects against this oxidative damage in both organs and alleviates degenerative changes, suggesting that it may be recommended for hypercholesterolemic patients on statins.
Diabetic kidney disease (DKD) is a leading cause of end-stage renal disease, with proximal tubule fibrosis being a key pathological feature. Our previous study identified significant upregulation of cytosolic nonspecific dipeptidase 2 (CNDP2) in the renal tubules of a DKD mouse model, yet its functional role remains unclear. This study aimed to investigate the role of CNDP2 in renal tubular fibrosis during DKD, focusing specifically on the "sulfur-containing amino acids (SAAs) metabolism-mammalian target of rapamycin (mTOR)" signaling axis. We employed integrated in vivo and in vitro models. Kidney- specific Cndp2 knockdown was achieved using an adeno-associated virus vector, and the CNDP2 inhibitor bestatin was used for pharmacological intervention. Renal function, fibrosis, the Ragulator-Rag GTPase-mTOR signaling pathway, amino acid profiles, and ultrastructural changes were assessed. A dietary intervention restricting SAAs was also applied. CNDP2 was specifically upregulated in renal tubules under DKD conditions. Both genetic knockdown and pharmacological inhibition of CNDP2 significantly improved renal function and attenuated fibrosis. Mechanistically, CNDP2 hydrolyzes dipeptides, leading to elevated levels of SAAs (cysteine/cystine). This promotes the activation of the Ragulator-Ras-related GTPase (Rag) complex, resulting in subsequent hyperactivation of mTOR signaling and driving tubular fibrosis. Notably, dietary restriction of SAAs similarly ameliorated DKD pathology. CNDP2 drives renal tubular fibrosis in DKD by activating mTOR signaling through disruption of SAAs metabolism. Our findings reveal a novel "CNDP2-SAAs-mTOR" pathway, identifying CNDP2 as a promising therapeutic target for DKD intervention.
Brazilian porcupinepox virus (BPoPV) has been associated with fatal infections in free-ranging Brazilian porcupines (Coendou longicaudatus), raising concerns regarding its epidemiology and conservation impact. This study comprehensively investigates BPoPV infection, pathology, and mortality in a free-living population from the Federal District, Brazil. Between 2021 and 2024, 30 porcupines submitted for postmortem examination were evaluated using gross and histopathology, PCR, epidermal morphometric analysis, and transmission electron microscopy. BPoPV infection was detected in 70% of animals (21/30), including all ten fatal cases and eleven individuals that died from unrelated traumatic injuries. Six PCR-positive animals without gross lesions showed significant epidermal thickening on microscopy, revealing a previously unrecognized subclinical infection. Fatal cases consistently exhibited severe proliferative and degenerative cutaneous lesions, intracytoplasmic inclusions, marked dermal inflammation, and visceral involvement, particularly interstitial pneumonia, esophageal pustules, and lymphoid depletion, suggesting systemic dissemination. Ultrastructural analysis confirmed abundant poxvirus-like particles within keratinocytes. Traumatic injuries, mainly dog attacks and electrocution, represented the most frequent noninfectious cause of death. These findings demonstrate a high frequency of clinical and subclinical BPoPV infection, identify the virus as a major infectious threat, and provide essential baseline data to support wildlife disease surveillance and conservation strategies for C. longicaudatus.
In Alzheimer's disease (AD), endogenous tau undergoes a pathogenic transition to form paired helical filaments (PHFs), but the cellular mechanisms driving this process have been elusive. Here, we identify the neuron-specific plasma membrane proteasome ('neuroproteasome') as a critical determinant of tau proteostasis. Selective inhibition of neuroproteasome function rapidly triggers the de novo formation of endogenous, sarkosyl-insoluble tau PHFs in primary neurons and mouse brain, which share key biochemical and ultrastructural features with PHFs from human AD brains. The APOE gene has three isoforms (E2, E3 and E4), with APOE4 being the largest genetic risk factor for AD. Neuroproteasome abundance at the plasma membrane is differentially modulated by ApoE isoforms (E2 > E3 > E4) and declines with age. ApoE4 neurons accumulate tau aggregates following modest neuroproteasome disruption, whereas ApoE2 neurons remain resistant. Our findings delineate a neuron-specific mechanism linking genetic and age-related risk factors to the formation of AD-relevant tau pathology, and position neuroproteasome function as a potential target to preserve proteostasis.
Ionizing radiation impairs male fertility, but the ultrastructural lesions linking radiation to sperm dysfunction remain unclear. The sperm annulus, a ring structure essential for flagellar integrity, is a potential but unresolved target. Here, we integrate synchrotron X-ray ptychography, cryogenic soft X-ray tomography, electron microscopy, and confocal imaging for cross-scale visualization of radiation-induced injury. Ptychography enables quantitative imaging of the annulus at ∼40-nm resolution, bridging light and electron microscopy. In irradiated mice, we find annulus fracture, loss of cytomembrane coverage, and disruption at the midpiece-principal piece junction. These defects worsen during maturation, correlating with reduced motility and increased abnormalities. Protein analyses show down-regulation of SEPT12 and AKAP4, indicating compromised annulus and fibrous sheath. Cryogenic tomography confirms ultrastructural deterioration in the axoneme, fibrous sheath, and mitochondria. These findings identify the sperm annulus and fibrous sheath as primary radiation targets, defining a structural basis for radiation-induced male infertility. This multimodal approach offers a generalizable strategy for nanoscale 3D cellular architecture analysis.
Correlative microscopy integrates complementary imaging modalities to enable structural analysis of biological samples across multiple length scales. Here, we present two correlative workflows for formalin-fixed, paraffin-embedded (FFPE) human tissues that extend the utility of archived clinical specimens for high-resolution imaging. The first, an on-slide correlative light and electron microscopy (CLEM) workflow, enables direct integration of brightfield light microscopy with scanning electron microscopy to resolve ultrastructural features from standard histological sections. The second, a multiscale three-dimensional workflow, combines X-ray micro-computed tomography with serial block-face scanning electron microscopy to enable targeted, volumetric ultrastructural analysis within a defined tissue context. Using these approaches, we demonstrate that FFPE tissues retain sufficient structural integrity to resolve cellular and subcellular architecture across liver, lung, brain, and heart specimens. Finally, we apply artificial intelligence-assisted image processing and segmentation to enable scalable, quantitative analysis of large-volume datasets. Together, these workflows establish FFPE specimens as a versatile and accessible resource for multiscale correlative imaging in both research and clinical settings.