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Abstract Objective Perineuronal nets (PNN) are specialized extracellular matrix (ECM) components of the central nervous system, frequently accumulating at the surface of inhibitory GABAergic interneurons. While an altered distribution of PNN has been observed in neurological disorders including Alzheimer's disease, schizophrenia and epilepsy, their anatomical distribution also changes during physiological brain maturation and aging. Such an age‐dependent shift was experimentally associated also with hippocampal engram formation during brain maturation. Our aim was to histopathologically assess PNN in the hippocampus of adult and pediatric patients with temporal lobe epilepsy (TLE) compared to age‐matched post‐mortem control subjects and to compare PNN‐related changes with memory impairment observed in our patient cohort. Methods Sixty‐six formalin‐fixed and paraffin‐embedded tissue specimens of the human hippocampus were retrieved from the European Epilepsy Brain Bank. Twenty‐nine patients had histopathologically confirmed hippocampal sclerosis (HS), and eleven patients suffered from TLE without HS. PNN were immunohistochemically visualized using an antibody directed against aggrecan and manually counted from hippocampus subfields and the subiculum. Results PNN density increased with age in both human controls and TLE patients. However, their density was significantly higher in all HS patients compared to age‐matched controls. Intriguingly, TLE patients presented presurgically with better memory when their hippocampal PNN density was higher (p

Abstract Focal Cortical Dysplasia (FCD) is a frequent cause of drug-resistant focal epilepsy in children and young adults. The international FCD classifications of 2011 and 2022 have identified several clinico-pathological subtypes, either occurring isolated, i.e., FCD ILAE Type 1 or 2, or in association with a principal cortical lesion, i.e., FCD Type 3. Here, we addressed the DNA methylation signature of a previously described new subtype of FCD 3D occurring in the occipital lobe of very young children and microscopically defined by neuronal cell loss in cortical layer 4. We studied the DNA methylation profile using 850 K BeadChip arrays in a retrospective cohort of 104 patients with FCD 1 A, 2 A, 2B, 3D, TLE without FCD, and 16 postmortem specimens without neurological disorders as controls, operated in China or Germany. DNA was extracted from formalin-fixed paraffin-embedded tissue blocks with microscopically confirmed lesions, and DNA methylation profiles were bioinformatically analyzed with a recently developed deep learning algorithm. Our results revealed a distinct position of FCD 3D in the DNA methylation map of common FCD subtypes, also different from non-FCD epilepsy surgery controls or non-epileptic postmortem controls. Within the FCD 3D cohort, the DNA methylation signature separated three histopathology subtypes, i.e., glial scarring around porencephalic cysts, loss of layer 4, and Rasmussen encephalitis. Differential methylation in FCD 3D with loss of layer 4 mapped explicitly to biological pathways related to neurodegeneration, biogenesis of the extracellular matrix (ECM) components, axon guidance, and regulation of the actin cytoskeleton. Our data suggest that DNA methylation signatures in cortical malformations are not only of diagnostic value but also phenotypically relevant, providing the molecular underpinnings of structural and histopathological features associated with epilepsy. Further studies will be necessary to confirm these results and clarify their functional relevance and epileptogenic potential in these difficult-to-treat children.

Abstract Biomedical ontologies are widely used to harmonize heterogeneous data and integrate large volumes of clinical data from multiple sources. This study analyzed the utility of ontologies beyond their traditional roles, that is, in addressing a challenging and currently underserved field of feature engineering in machine learning workflows. Machine learning workflows are being increasingly used to analyze medical records with heterogeneous phenotypic, genotypic, and related medical terms to improve patient care. We performed a retrospective study using neuropathology reports from the German Neuropathology Reference Center for Epilepsy Surgery at Erlangen, Germany. This cohort included 312 patients who underwent epilepsy surgery and were labeled with one or more diagnoses, including dual pathology, hippocampal sclerosis, malformation of cortical dysplasia, tumor, encephalitis, and gliosis. We modeled the diagnosis terms together with their microscopy, immunohistochemistry, anatomy, etiologies, and imaging findings using the description logic-based Web Ontology Language (OWL) in the Epilepsy and Seizure Ontology (EpSO). Three tree-based machine learning models were used to classify the neuropathology reports into one or more diagnosis classes with and without ontology-based feature engineering. We used five-fold cross validation to avoid overfitting with a fixed number of repetitions while leaving out one subset of data for testing, and we used recall, balanced accuracy, and hamming loss as performance metrics for the multi-label classification task. The epilepsy ontology-based feature engineering approach improved the performance of all the three learning models with an improvement of 35.7%, 54.5%, and 33.3% in logistics regression, random forest, and gradient tree boosting models respectively. The run time performance of all three models improved significantly with ontology-based feature engineering with gradient tree boosting model showing a 93.8% reduction in the time required for training and testing of the model. Although, all three models showed an overall improved performance across the three-performance metrics using ontology-based feature engineering, the rate of improvement was not consistent across all input features. To analyze this variation in performance, we computed feature importance scores and found that microscopy had the highest importance score across the three models, followed by imaging, immunohistochemistry, and anatomy in a decreasing order of importance scores. This study showed that ontologies have an important role in feature engineering to make heterogeneous clinical data accessible to machine learning models and also improve the performance of machine learning models in multilabel multiclass classification tasks.

The human brain has a highly complex structure both on the microscopic and on the macroscopic scales. Increasing evidence has suggested the role of mechanical forces for cortical folding – a classical hallmark of the human brain. However, the link between cellular processes at the microscale and mechanical forces at the macroscale remains insufficiently understood. Recent findings suggest that an additional proliferating zone, the outer subventricular zone (OSVZ), is decisive for the particular size and complexity of the human cortex. To better understand how the OSVZ affects cortical folding, we establish a multifield computational model that couples cell proliferation in different zones and migration at the cell scale with growth and cortical folding at the organ scale by combining an advection-diffusion model with the theory of finite growth. We validate our model based on data from histologically stained sections of the human fetal brain and predict 3D pattern formation. Finally, we address open questions regarding the role of the OSVZ for the formation of cortical folds. The presented framework not only improves our understanding of human brain development, but could eventually help diagnose and treat neuronal disorders arising from disruptions in cellular development and associated malformations of cortical development.

Abstract Focal malformations of cortical development (MCD) are linked to somatic brain mutations occurring during neurodevelopment. Mild malformation of cortical development with oligodendroglial hyperplasia in epilepsy (MOGHE) is a newly recognized clinico-pathological entity associated with pediatric drug-resistant focal epilepsy, and amenable to neurosurgical treatment. MOGHE is histopathologically characterized by clusters of increased oligodendroglial cell densities, patchy zones of hypomyelination, and heterotopic neurons in the white matter. The molecular etiology of MOGHE remained unknown so far. We hypothesized a contribution of mosaic brain variants and performed deep targeted gene sequencing on 20 surgical MOGHE brain samples from a single-center cohort of pediatric patients. We identified somatic pathogenic SLC35A2 variants in 9/20 (45%) patients with mosaic rates ranging from 7 to 52%. SLC35A2 encodes a UDP-galactose transporter, previously implicated in other malformations of cortical development (MCD) and a rare type of congenital disorder of glycosylation. To further clarify the histological features of SLC35A2-brain tissues, we then collected 17 samples with pathogenic SLC35A2 variants from a multicenter cohort of MCD cases. Histopathological reassessment including anti-Olig2 staining confirmed a MOGHE diagnosis in all cases. Analysis by droplet digital PCR of pools of microdissected cells from one MOGHE tissue revealed a variant enrichment in clustered oligodendroglial cells and heterotopic neurons. Through an international consortium, we assembled an unprecedented series of 26 SLC35A2-MOGHE cases providing evidence that mosaic SLC35A2 variants, likely occurred in a neuroglial progenitor cell during brain development, are a genetic marker for MOGHE.

Abstract Objective To comprehensively describe the pathological features of neurons in patients with ovarian teratomas and paraneoplastic anti-NMDAR encephalitis (anti-NMDARE), emphasizing on NMDA-receptor expression and infiltrating lymphocytes. Methods A retrospective study was performed in a large series of 159 patients from the West China Hospital. We retrospectively identified 12 patients with paraneoplastic anti-NMDARE (11 case with ovarian teratomas and 1 case with mixed germ cell tumor), which were compared to 35 patients with teratomas and no encephalitis and to 147 patients with anti-NMDARE and no evidence for tumors. Patient history and outcome were reviewed from the clinical charts and compared between all three groups. Histopathological examination, including double-immunofluorescence of NMDAR subunits and IgG was performed in all teratoma tissues. Magnetic Luminex Assay Human Premixed Multi-Analyte Kit was performed to investigate cytokines profile of CSF. Results Patients with paraneoplastic anti-NMDARE had a more severe clinical presentation, i.e. they required more mechanical ventilation and intensive care (p

Abstract Brain tumors represent the second most frequent etiology in patients with focal seizure onset before 18 years of age and submitted to epilepsy surgery. Hence, this category of brain tumors, herein defined as low-grade, developmental, epilepsy-associated brain tumors (LEAT) is different from those frequently encountered in adults as (A): 77% of LEAT occur in the temporal lobe; (B): the vast majority of LEAT are of low malignancy and classified as WHO I°; (C): LEAT are often composed of mixed glial and neuronal cell components and present with variable growth patterns including small cysts or nodules; (D): LEAT do not share common gene driving mutations, such as IDH1 or 1p/19q co-deletions. Characteristic entities comprise the ganglioglioma (GG), the dysembryoplastic neuroepithelial tumor (DNT), the angiocentric glioma (AG), the isomorphic diffuse glioma (IDG) and the papillary glio-neuronal tumor (PGNT), representing 73.2% of 1680 tumors collected in a large German series of 6747 patients submitted to epilepsy surgery. In the realm of exciting discoveries of genetic drivers of brain tumors new genes have been also reported for LEAT. BRAF V600E mutations were linked to GG with CD34 expression, FGFR1 mutations to DNT, MYB alterations to AG and also IDG and PRKCA fusions to PGNT, suggesting the possibility to also develop a genetically driven tumor classification scheme for LEAT. Rare availability of LEAT in a single center is a challenging obstacle, however, to systematically unravel the neurobiological nature and clinical behavior of LEAT. Other challenges in need of clarification include malignant tumor progression of LEAT entities, seizure relapse in patients following bulk tumor resection and the controversial issue of associated focal cortical dysplasia as additional pathomechanism. In order to advance our understanding and promote reliable diagnostic work-up of LEAT, we recommend, therefore, international collaboration to achieve our goals.

Abstract Introduction We investigated the frequency, neuropathology, and phenotypic characteristics of spastic paraplegia (SP) that precedes dementia in presenilin 1 (PSEN1) related familial Alzheimer's disease (AD). Methods We performed whole exome sequencing (WES) in 60 probands with hereditary spastic paraplegia (HSP) phenotype that was negative for variants in known HSP‐related genes. Where PSEN1 mutation was identified, brain biopsy was performed. We investigated the link between HSP and AD with PSEN1 in silico pathway analysis and measured in vivo the stability of PSEN1 mutant γ‐secretase. Results We identified a PSEN1 variant (p.Thr291Pro) in an individual presenting with pure SP at 30 years of age. Three years later, SP was associated with severe, fast cognitive decline and amyloid deposition with diffuse cortical plaques on brain biopsy. Biochemical analysis of p.Thr291Pro PSEN1 revealed that although the mutation does not alter active γ‐secretase reconstitution, it destabilizes γ‐secretase‐amyloid precursor protein (APP)/amyloid beta (Aβn) interactions during proteolysis, enhancing the production of longer Aβ peptides. We then extended our analysis to all 226 PSEN1 pathogenic variants reported and show that 7.5% were associated with pure SP onset followed by cognitive decline later in the disease. We found that PSEN1 cases manifesting initially as SP have a later age of onset, are associated with mutations located beyond codon 200, and showed larger diffuse, cored plaques, amyloid‐ring arteries, and severe CAA. Discussion We show that pure SP can precede dementia onset in PSEN1‐related familial AD. We recommend PSEN1 genetic testing in patients presenting with SP with no variants in known HSP‐related genes, particularly when associated with a family history of cognitive decline.

Background: Recent findings have demonstrated that hippocampal subfields can be selectively affected in different disease states, which has led to efforts to segment the human hippocampus with in vivo magnetic resonance imaging (MRI). However, no studies have examined the histological accuracy of subfield segmentation protocols. The presence of MRI-visible anatomical landmarks with known correspondence to histology represents a fundamental prerequisite for in vivo hippocampal subfield segmentation. In the present study, we aimed to: 1) develop a novel method for hippocampal body segmentation, based on two MRI-visible anatomical landmarks (stratum lacunosum moleculare [SLM] & dentate gyrus [DG]), and assess its accuracy in comparison to the gold standard direct histological measurements; 2) quantify the accuracy of two published segmentation strategies in comparison to the histological gold standard; and 3) apply the novel method to ex vivo MRI and correlate the results with histology. Methods: Ultra-high resolution ex vivo MRI was performed on six whole cadaveric hippocampal specimens, which were then divided into 22 blocks and histologically processed. The hippocampal bodies were segmented into subfields based on histological criteria and subfield boundaries and areas were directly measured. A novel method was developed using mean percentage of the total SLM distance to define subfield boundaries. Boundary distances and subfield areas on histology were then determined using the novel method and compared to the gold standard histological measurements. The novel method was then used to determine ex vivo MRI measures of subfield boundaries and areas, which were compared to histological measurements. Results: For direct histological measurements, the mean percentages of total SLM distance were: Subiculum/CA1 = 9.7%, CA1/CA2 = 78.4%, CA2/CA3 = 97.5%. When applied to histology, the novel method provided accurate measures for CA1/CA2 (ICC = 0.93) and CA2/CA3 (ICC = 0.97) boundaries, but not for the Subiculum/CA1 (ICC = −0.04) boundary. Accuracy was poorer using previous techniques for CA1/CA2 (maximum ICC = 0.85) and CA2/CA3 (maximum ICC = 0.88), with the previously reported techniques also performing poorly in defining the Subiculum/CA1 boundary (maximum ICC = 0.00). Ex vivo MRI measurements using the novel method were linearly related to direct measurements of SLM length (r2 = 0.58), CA1/CA2 boundary (r2 = 0.39) and CA2/CA3 boundary (r2 = 0.47), but not for Subiculum/CA1 boundary (r2 = 0.01). Subfield areas measured with the novel method on histology and ex vivo MRI were linearly related to gold standard histological measures for CA1, CA2, and CA3/CA4/DG. Conclusions: In this initial proof of concept study, we used ex vivo MRI and histology of cadaveric hippocampi to develop a novel segmentation protocol for the hippocampal body. The novel method utilized two anatomical landmarks, SLM & DG, and provided accurate measurements of CA1, CA2, and CA3/CA4/DG subfields in comparison to the gold standard histological measurements. The relationships demonstrated between histology and ex vivo MRI supports the potential feasibility of applying this method to in vivo MRI studies.

Neuropathology-based studies in neurosurgically resected brain tissue obtained from carefully examined patients with focal epilepsies remain a treasure box for excellent insights into human neuroscience, including avenues to better understand the neurobiology of human brain organization and neuronal hyperexcitability at the cellular level including glio-neuronal interaction. It also allows to translate results from animal models in order to develop personalized treatment strategies in the near future. A nice example of this is the discovery of a new disease entity in 2017, termed mild malformation of cortical development with oligodendroglial hyperplasia in epilepsy or MOGHE, in the frontal lobe of young children with intractable seizures. In 2021, a brain somatic missense mutation of the galactose transporter SLC35A2 leading to altered glycosylation of lipoproteins in the Golgi apparatus was detected in 50 % of MOGHE samples. In 2023, the first clinical trial evaluated galactose supplementation in patients with histopathologically confirmed MOGHE carrying brain somatic SLC35A2 mutations that were not seizure free after surgery. The promising results of this pilot trial are an example of personalized medicine in the arena of epileptology. Besides this, neuropathological studies of epilepsy samples have revealed many other fascinating results for the main disease categories in focal epilepsies, such as the first deep-learning based classifier for Focal Cortical Dysplasia, or the genomic landscape of cortical malformations showing new candidate genes such as PTPN11, which is associated with ganglioglioma and adverse clinical outcome. This update will also ask why common pathogenic variants accumulate in certain brain regions, e.g., MTOR in the frontal lobe, and BRAF in the temporal lobe. Finally, I will highlight the ongoing discussion addressing commonalities between temporal lobe epilepsy and Alzheimer's disease, the impact of adult neurogenesis and gliogenesis for the initiation and progression of temporal lobe seizures in the human brain as well as the immunopathogenesis of glutamic acid decarboxylase antibody associated temporal lobe epilepsy as a meaningful disease entity. This review will update the reader on some of these fascinating publications from 2022 and 2023 which were selected carefully, yet subjectively, by the author.

Abstract Focal cortical dysplasia type II (FCDII) is the most common cause of drug-resistant focal epilepsy in children. Herein, we performed a deep histopathology-based genotype–phenotype analysis to further elucidate the clinico-pathological and genetic presentation of FCDIIa compared to FCDIIb. Seventeen individuals with histopathologically confirmed diagnosis of FCD ILAE Type II and a pathogenic variant detected in brain derived DNA whole-exome sequencing or mTOR gene panel sequencing were included in this study. Clinical data were directly available from each contributing centre. Histopathological analyses were performed from formalin-fixed, paraffin-embedded tissue samples using haematoxylin–eosin and immunohistochemistry for NF-SMI32, NeuN, pS6, p62, and vimentin. Ten individuals carried loss-of-function variants in the GATOR1 complex encoding genes DEPDC5 (n = 7) and NPRL3 (n = 3), or gain-of-function variants in MTOR (n = 7). Whereas individuals with GATOR1 variants only presented with FCDIIa, i.e., lack of balloon cells, individuals with MTOR variants presented with both histopathology subtypes, FCDIIa and FCDIIb. Interestingly, 50% of GATOR1-positive cases showed a unique and predominantly vacuolizing phenotype with p62 immunofluorescent aggregates in autophagosomes. All cases with GATOR1 alterations had neurosurgery in the frontal lobe and the majority was confined to the cortical ribbon not affecting the white matter. This pattern was reflected by subtle or negative MRI findings in seven individuals with GATOR1 variants. Nonetheless, all individuals were seizure-free after surgery except four individuals carrying a DEPDC5 variant. We describe a yet underrecognized genotype–phenotype correlation of GATOR1 variants with FCDIIa in the frontal lobe. These lesions were histopathologically characterized by abnormally vacuolizing cells suggestive of an autophagy-altered phenotype. In contrast, individuals with FCDIIb and brain somatic MTOR variants showed larger lesions on MRI including the white matter, suggesting compromised neural cell migration.

Abstract Extracellular matrix (ECM) deposition after central nervous system (CNS) injury leads to inhibitory scarring in humans and other mammals, whereas it facilitates axon regeneration in the zebrafish. However, the molecular basis of these different fates is not understood. Here, we identify small leucine-rich proteoglycans (SLRPs) as a contributing factor to regeneration failure in mammals. We demonstrate that the SLRPs chondroadherin, fibromodulin, lumican, and prolargin are enriched in rodent and human but not zebrafish CNS lesions. Targeting SLRPs to the zebrafish injury ECM inhibits axon regeneration and functional recovery. Mechanistically, we find that SLRPs confer mechano-structural properties to the lesion environment that are adverse to axon growth. Our study reveals SLRPs as inhibitory ECM factors that impair axon regeneration by modifying tissue mechanics and structure, and identifies their enrichment as a feature of human brain and spinal cord lesions. These findings imply that SLRPs may be targets for therapeutic strategies to promote CNS regeneration.

IntroductionFocal cortical dysplasia (FCD) is a common cause of pharmacoresistant epilepsy. According to the 2022 International League Against Epilepsy classification, FCD type II is characterized by dysmorphic neurons (IIa and IIb) and may be associated with balloon cells (IIb). We present a multicentric study to evaluate the transcriptomes of the gray and white matters of surgical FCD type II specimens. We aimed to contribute to pathophysiology and tissue characterization.MethodsWe investigated FCD II (a and b) and control samples by performing RNA-sequencing followed by immunohistochemical validation employing digital analyses.ResultsWe found 342 and 399 transcripts differentially expressed in the gray matter of IIa and IIb lesions compared to controls, respectively. Cholesterol biosynthesis was among the main enriched cellular pathways in both IIa and IIb gray matter. Particularly, the genes HMGCS1, HMGCR, and SQLE were upregulated in both type II groups. We also found 12 differentially expressed genes when comparing transcriptomes of IIa and IIb lesions. Only 1 transcript (MTRNR2L12) was significantly upregulated in FCD IIa. The white matter in IIa and IIb lesions showed 2 and 24 transcripts differentially expressed, respectively, compared to controls. No enriched cellular pathways were detected. GPNMB, not previously described in FCD samples, was upregulated in IIb compared to IIa and control groups. Upregulations of cholesterol biosynthesis enzymes and GPNMB genes in FCD groups were immunohistochemically validated. Such enzymes were mainly detected in both dysmorphic and normal neurons, whereas GPNMB was observed only in balloon cells.DiscussionOverall, our study contributed to identifying cortical enrichment of cholesterol biosynthesis in FCD type II, which may correspond to a neuroprotective response to seizures. Moreover, specific analyses in either the gray or the white matter revealed upregulations of MTRNR2L12 and GPNMB, which might be potential neuropathological biomarkers of a cortex chronically exposed to seizures and of balloon cells, respectively.

The impact of a precise histopathology diagnosis and molecular workup for surgical patient management remains a controversial issue in epileptology with a lack of diagnostic agreement as root cause. Very recent advances in genotype-phenotype characterization of epilepsy-associated developmental brain lesions, including the first diagnostically useful DNA methylation studies, opened new avenues and will help to finally resolve these issues. A series of most recent articles were decisively selected by the author to exemplify the areas of improvement in neuropathology and epilepsy surgery. These topics include the progress in genotype-phenotype association studies of Focal Cortical Dysplasia (FCD) leading to the discovery of new molecularly defined entities, i.e. mild malformation of cortical development with oligodendroglial hyperplasia in epilepsy (MOGHE), SLC35A2 altered. These studies also triggered the first update of the international FCD consensus classification scheme from 2011, which will hopefully support diagnostic agreement in clinical practice and research. The dilemma of new tumor entities proposed by the 5th edition of the WHO classification primarily associated with early seizure onset yet not well introduced to the epileptology community will also be discussed in the light of emerging experimental evidence when transfecting the developing murine brain with the single most important genetic alteration for both carcino- and epileptogenesis, i.e. BRAF V600E.

Epilepsy surgery is an effective treatment option for drug-resistant focal epilepsy patients with associated structural brain lesions. However, little epidemiological data are available regarding the number of patients with these lesions. We reviewed data regarding (1) the prevalence and incidence of epilepsy; (2) the proportion of epilepsy patients with focal epilepsy, drug-resistant epilepsy, and drug-resistant focal epilepsies; and (3) the number of epilepsy presurgical evaluations and surgical resections. We also assessed the relative proportion of brain lesions using post-surgical histopathological findings from 541 surgical patients from the Cleveland Clinic and 9,523 patients from a European multi-center cohort. Data were combined to generate surgical candidate incidence and prevalence estimates and the first lesion-specific estimates for hippocampal sclerosis (HS), low-grade epilepsy-associated brain tumors (LEAT), malformations of cortical development (MCD), glial scars, vascular malformations, and encephalitis. The most frequently diagnosed brain lesions were HS (incidence = 2.32 ± 0.26 in 100,000, prevalence = 19.40 ± 2.16 in 100,000) for adults and MCD (incidence = 1.15 ± 0.34 in 100,000, prevalence = 6.52 ± 1.89 in 100,000) for children. Our estimates can guide patient advocacy groups, clinicians, researchers, policymakers in education, development of health care strategy, resource allocation, and reimbursement schedules.

The treatment options for neuropathic pain caused by lumbar disc herniation have been debated controversially in the literature. Whether surgical or conservative therapy makes more sense in individual cases can hardly be answered. We have investigated whether a machine learning-based prediction of outcome, regarding neuropathic pain development, after lumbar disc herniation treatment is possible. The extensive datasets of 123 consecutive patients were used to predict the development of neuropathic pain, measured by a visual analogue scale (VAS) for leg pain and the Oswestry Disability Index (ODI), at 6 weeks, 6 months and 1 year after treatment of lumbar disc herniation in a machine learning approach. Using a decision tree regressor algorithm, a prediction quality within the limits of the minimum clinically important difference for the VAS and ODI value could be achieved. An analysis of the influencing factors of the algorithm reveals the important role of psychological factors as well as body weight and age with pre-existing conditions for an accurate prediction of neuropathic pain. The machine learning algorithm developed here can enable an assessment of the course of treatment after lumbar disc herniation. The early, comparative individual prediction of a therapy outcome is important to avoid unnecessary surgical therapies as well as insufficient conservative therapies and prevent the chronification of neuropathic pain.

Objective: The recent FDA approval of the first 7T MRI scanner for clinical diagnostic use in October 2017 will likely increase the utilization of 7T for epilepsy presurgical evaluation. This study aims at accessing the radiological and clinical value of 7T in patients with pharmacoresistant focal epilepsy and 3T-visible lesions.Methods: Patients with pharmacoresistant focal epilepsy were included if they had a lesion on pre-operative standard-of-care 3T MRI and also a 7T research MRI. An epilepsy protocol was used for the acquisition of the 7T MRI. Prospective visual analysis of 7T MRI was performed by an experienced board-certified neuroradiologist and communicated to the patient management team. The clinical significance of the additional 7T findings was assessed by intracranial EEG (ICEEG) ictal onset, surgical resection, post-operative seizure outcome and histopathology. A subset of lesions were demarked with arrows for subsequent, retrospective comparison between 3T and 7T by 7 neuroradiologists using a set of quantitative scales: lesion presence, conspicuity, boundary, gray-white tissue contrast, artifacts, and the most helpful sequence for diagnosis. Conger's kappa for multiple raters was performed for chance-adjusted agreement statistics.Results: A total of 47 patients were included, with the main pathology types of focal cortical dysplasia (FCD), hippocampal sclerosis, periventricular nodular heterotopia (PVNH), tumor and polymicrogyria (PMG). 7T detected additional smaller lesions in 19% (9/47) of patients, who had extensive abnormalities such as PMG and PVNH; however, these additional findings were not necessarily epileptogenic. 3T−7T comparison by the neuroradiologist team showed that lesion conspicuity and lesion boundary were significantly better at 7T (p < 0.001), particularly for FCD, PVNH and PMG. Chance-adjusted agreement was within the fair range for lesion presence, conspicuity and boundary. Gray-white contrast was significantly improved at 7T (p < 0.001). Significantly more artifacts were encountered at 7T (p < 0.001).Significance: For patients with 3T-visible lesions, 7T MRI may better elucidate the extent of multifocal abnormalities such as PVNH and PMG, providing potential targets to improve ICEEG implantation. Patients with FCD, PVNH and PMG would likely benefit the most from 7T due to improved lesion conspicuity and boundary. Pathologies in the antero–inferior temporal regions likely benefit less due to artifacts.

The COVID-19 pandemic has significantly changed the way we treat patients and educate healthcare professionals (HCPs). In summer 2020, the International League against Epilepsy (ILAE) implemented a virtual CME program with three integrated program elements addressing challenges in patient treatment as well as challenges caused by the forced transition to a virtual environment. Despite the highly competitive environment with exponential increase of webinars offered to HCPs, the program achieved high participation and satisfaction rates. Over 60% of participants indicated a change in their clinical practice after the interventions. With our outcomes evaluation, we aimed to better understand how well such an integrated program resonates with the learner and if it can make a difference in a highly competitive environment by supporting educators to become more adaptive and responsive to learner needs. Our pilot project was shown to be well accepted, achieving high satisfaction and perceived impact by the learner. In the light of an upcoming “digital fatigue” and a wish to return to face-to-face, we reiterate the value of the digital approach and recommend continuing along this successful path as we believe that taking a learner on a digital educational journey has been successful in a highly competitive and challenging environment.

The immunoglobulin (Ig) heavy and light chain variable gene mutational pattern of the B cell receptor (BCR) in primary central nervous system (CNS) lymphoma (PCNSL) cells suggests antigenic selection to drive pathogenesis and confinement to the CNS. This hypothesis is supported by the observation that the tumor B cell receptor (tBCR) of PCNSL is polyreactive and may be stimulated by CNS proteins. To obtain further insight into the role of the germinal center (GC) reaction on BCR reactivity, we constructed recombinant antibodies (recAb) with Ig heavy and light chain sequences of the corresponding naive BCR (nBCR) by reverting tBCR somatic mutations in 10 PCNSL. Analysis of nBCR-derived recAb reactivity by a protein microarray and immunoprecipitation demonstrated auto- and polyreactivity in all cases. Self-/polyreactivity was not lost during the GC reaction; surprisingly, tBCR significantly increased self-/polyreactivity. In addition to proteins recognized by both the nBCR and tBCR, tBCR gained self-/polyreactivity particularly for proteins expressed in the CNS including proteins of oligodendrocytes/myelin, the S100 protein family, and splicing factors. Thus, in PCNSL pathogenesis, a faulty GC reaction may increase self-/polyreactivity, hereby facilitating BCR signaling via multiple CNS antigens, and may ultimately foster tumor cell survival in the CNS.

Objective: Temporal lobe epilepsy patients (TLE) often present with hippocampal atrophy, increased T2 relaxation, and reduced magnetization transfer ratio (MTR) in magnetic resonance images (MRI). The histological correlates of the reduced hippocampal MTR are so far unknown. Since MTR is dependent on the tissue’s macromolecules, our aim was to evaluate the correlations between cellular populations, extracellular matrix molecules and the MTR in TLE patients. Methods: Patients with TLE (n = 26) and voluntaries (=20) were scanned in a 3 Tesla MRI scanner, and MTR images were calculated from 3DT1 sequences with magnetization pulse on resonance. Immunohistochemistry for neurons, reactive astrocytes, activated microglia, and extracellular matrix chondroitin sulfate were performed in formalin fixed, paraffin embedded tissues of TLE and autopsy controls (n = 10). Results were considered significant with adjusted p

Background: Epilepsy surgery for focal cortical dysplasia type II (FCD II) offers good chances for seizure freedom, but remains a challenge with respect to lesion detection, defining the epileptogenic zone and the optimal resection strategy. Integrating results from magnetic source imaging from magnetoencephalography (MEG) with magnetic resonance imaging (MRI) including MRI postprocessing may be useful for optimizing these goals. Methods: We here present data from 21 adult FCD II patients, investigated during a 10 year period and evaluated including magnetic source imaging. 16 patients had epilepsy surgery, i.e. histopathologically verified FCD II, and a long follow up. We present our analysis of epileptogenic zones including MEG in relation to structural data according to MRI data and relate these results to surgical outcomes. Results: FCD II in our cohort was characterized by high MEG yield and localization accuracy and MEG showed impact on surgical success-rates. MEG source localizations were detected in 95.2% of patients and were as close as 12.3 ± 8,1 mm to the MRI-lesion. After a mean follow up of >3 years, we saw >80% Engel I outcomes, with more favourable outcomes when the MEG source was completely resected (Fishers exact test 0,033). Conclusion: We argue for a high value of conducting a combined MEG-MRI approach in the presurgical workup and the resection strategy in patients with FCD II related epilepsy. Keywords: Focal cortical dysplasia, Magnetic source imaging, epilepsy surgery

Background: Processing whole-slide images (WSI) to train neural networks can be intricate and labor intensive. We developed an open-source library dealing with recurrent tasks in the processing of WSI and helping with the training and evaluation of neuronal networks for classification tasks. Methods: Two histopathology use-cases were selected and only hematoxylin and eosin (H&E) stained slides were used. The first use case was a two-class classification problem. We trained a convolutional neuronal network (CNN) to distinguish between dysembryoplastic neuroepithelial tumor (DNET) and ganglioglioma (GG), two neuropathological low-grade epilepsy-associated tumor entities. Within the second use case, we included four clinicopathological disease conditions in a multilabel approach. Here we trained a CNN to predict the hormone expression profile of pituitary adenomas. In the same approach, we also predicted clinically silent corticotroph adenoma. Results: Our DNET-GG classifier achieved an AUC of 1.00 for the ROC curve. For the second use case, the best performing CNN achieved an area under the curve (AUC) of 0.97 for the receiver operating characteristic (ROC) for corticotroph adenoma, 0.86 for silent corticotroph adenoma, and 0.98 for gonadotroph adenoma. All scores were calculated with the help of our library on predictions on a case basis. Conclusions: Our comprehensive and fastai-compatible library is helpful to standardize the workflow and minimize the burden of training a CNN. Indeed, our trained CNNs extracted neuropathologically relevant information from the WSI. This approach will supplement the clinicopathological diagnosis of brain tumors, which is currently based on cost-intensive microscopic examination and variable panels of immunohistochemical stainings.

Focal cortical dysplasia (FCD) and hemimegalencephaly (HME) are epileptogenic neurodevelopmental malformations caused by mutations in mTOR pathway genes. Deep sequencing of these genes in FCD/HME brain tissue identified an etiology in 27 of 66 cases (41%). Radiographically indistinguishable lesions are caused by somatic activating mutations in AKT3, MTOR, and PIK3CA and germline loss-of-function mutations in DEPDC5, NPRL2, and TSC1/2, including TSC2 mutations in isolated HME demonstrating a “two-hit” model. Mutations in the same gene cause a disease continuum from FCD to HME to bilateral brain overgrowth, reflecting the progenitor cell and developmental time when the mutation occurred. Single-cell sequencing demonstrated mTOR activation in neurons in all lesions. Conditional Pik3ca activation in the mouse cortex showed that mTOR activation in excitatory neurons and glia, but not interneurons, is sufficient for abnormal cortical overgrowth. These data suggest that mTOR activation in dorsal telencephalic progenitors, in some cases specifically the excitatory neuron lineage, causes cortical dysplasia.

Understanding seizure development requires an integrated knowledge of different scales of organization of epileptic networks. We developed a model of “epilepsy-in-a-dish” based on dissociated primary neuronal cells from neonatal rat hippocampus. We demonstrate how a single application of glutamate stimulated neurons to generate spontaneous synchronous spiking activity with further progression into spontaneous seizure-like events after a distinct latency period. By computational analysis, we compared the observed neuronal activity in vitro with intracranial electroencephalography (EEG) data recorded from epilepsy patients and identified strong similarities, including a related sequence of events with defined onset, progression, and termination. Next, a link between the neurophysiological changes with network composition and cellular structure down to molecular changes was established. Temporal development of epileptiform network activity correlated with increased neurite outgrowth and altered branching, increased ratio of glutamatergic over GABAergic synapses, and loss of calbindin-positive interneurons, as well as genome-wide alterations in DNA methylation. Differentially methylated genes were engaged in various cellular activities related to cellular structure, intracellular signaling, and regulation of gene expression. Our data provide evidence that a single short-term excess of glutamate is sufficient to induce a cascade of events covering different scales from molecule- to network-level, all of which jointly contribute to seizure development.

Magnetic resonance imaging (MRI) provides a unique tool for in vivo visualization and tracking of stem cells in the brain. This is of particular importance when assessing safety of experimental cell treatments in the preclinical or clinical setup. Yet, specific imaging requires an efficient and non-perturbing cellular magnetic labeling which precludes adverse effects of the tag, e.g., the impact of iron-oxide-nanoparticles on the critical differentiation and integration processes of the respective stem cell population investigated. In this study we investigated the effects of very small superparamagnetic iron oxide particle (VSOP) labeling on viability, stemness, and neuronal differentiation potential of primary human adult neural stem cells (haNSCs). Cytoplasmic VSOP incorporation massively reduced the transverse relaxation time T2, an important parameter determining MR contrast. Cells retained cytoplasmic label for at least a month, indicating stable incorporation, a necessity for long-term imaging. Using a clinical 3T MRI, 1 × 103 haNSCs were visualized upon injection in a gel phantom, but detection limit was much lower (5 × 104 cells) in layer phantoms and using an imaging protocol feasible in a clinical scenario. Transcriptional analysis and fluorescence immunocytochemistry did not reveal a detrimental impact of VSOP labeling on important parameters of cellular physiology with cellular viability, stemness and neuronal differentiation potential remaining unaffected. This represents a pivotal prerequisite with respect to clinical application of this method.

Correct positioning and differentiation of neurons during brain development is a key precondition for proper function. Focal cortical dysplasias (FCDs) are increasingly recognized as causes of therapy refractory epilepsies. Neuropathological analyses of respective surgical specimens from neurosurgery for seizure control often reveal aberrant cortical architecture and/or aberrantly shaped neurons in FCDs. However, the molecular pathogenesis particularly of FCDs with aberrant lamination (so-called FCD type I) is largely unresolved. Lipoproteins and particularly low-density lipoprotein receptor-related protein 12 (LRP12) are involved in brain development. Here, we have examined a potential role of LRP12 in the pathogenesis of FCDs. In vitro knockdown of LRP12 in primary neurons results in impaired neuronal arborization. In vivo ablation of LRP12 by intraventricularly in utero electroporated shRNAs elicits cortical maldevelopment, i.e. aberrant lamination by malpositioning of upper cortical layer neurons. Subsequent epilepsy phenotyping revealed pentylenetetrazol (PTZ)-induced seizures to be aggravated in cortical LRP12-silenced mice. Our data demonstrates IUE mediated cortical gene silencing as an excellent approach to study the role of distinct molecules for epilepsy associated focal brain lesions and suggests LRP12 and lipoprotein homeostasis as potential molecular target structures for the emergence of epilepsy-associated FCDs.

In patients with temporal lobe epilepsy (TLE), presurgical magnetic resonance imaging (MRI) often reveals hippocampal atrophy, while neuropathological assessment indicates the different types of hippocampal sclerosis (HS). Different HS types are not discriminated in MRI so far. We aimed to define the volume of each hippocampal subfield on MRI manually and to compare automatic and manual segmentations for the discrimination of HS types. The T2-weighted images from 14 formalin-fixed age-matched control hippocampi were obtained with 4.7T MRI to evaluate the volume of each subfield at the anatomical level of the hippocampal head, body, and tail. Formalin-fixed coronal sections at the level of the body of 14 control cases, as well as tissue samples from 24 TLE patients, were imaged with a similar high-resolution sequence at 3T. Presurgical three-dimensional (3D) T1-weighted images from TLE went through a FreeSurfer 6.0 hippocampal subfield automatic assessment. The manual delineation with the 4.7T MRI was identified using Luxol Fast Blue stained 10-μm-thin microscopy slides, collected at every millimeter. An additional section at the level of the body from controls and TLE cases was submitted to NeuN immunohistochemistry for neuronal density estimation. All TLE cases were classified according to the International League Against Epilepsy's (ILAE's) HS classification. Manual volumetry in controls revealed that the dentate gyrus (DG)+CA4 region, CA1, and subiculum accounted for almost 90% of the hippocampal volume. The manual 3T volumetry showed that all TLE patients with type 1 HS (TLE-HS1) had lower volumes for DG+CA4, CA2, and CA1, whereas those TLE patients with HS type 2 (TLE-HS2) had lower volumes only in CA1 (p ≤ 0.038). Neuronal cell densities always decreased in CA4, CA3, CA2, and CA1 of TLE-HS1 but only in CA1 of TLE-HS2 (p ≤ 0.003). In addition, TLE-HS2 had a higher volume (p = 0.016) and higher neuronal density (p < 0.001) than the TLE-HS1 in DG + CA4. Automatic segmentation failed to match the manual or histological findings and was unable to differentiate TLE-HS1 from TLE-HS2. Total hippocampal volume correlated with DG+CA4 and CA1 volumes and neuronal density. For the first time, we also identified subfield-specific pathology patterns in the manual evaluation of volumetric MRI scans, showing the importance of manual segmentation to assess subfield-specific pathology patterns.

The aim of the present study is to differentiate subtypes of hippocampal sclerosis (HS) using ex vivo ultra-high field magnetic resonance imaging (MRI). Included were 14 surgically resected hippocampi of patients with medically intractable temporal lobe epilepsy. The resected hippocampi were histologically categorized into subtypes of hippocampal sclerosis (HS type 1 (n = 10), HS type 2 (n = 2) and no-HS (n = 2)) and subsequently scanned on a preclinical 7T MRI acquiring T2-weighted morphology, relaxometry and diffusion tensor imaging. On the morphological images, the pyramidal cell layer (PCL) of the hippocampus was segmented and the following parameters were derived: T2 signal intensity, T1-, T2- and T2*-relaxation times, apparent diffusion coefficient (ADC), fractional anisotropy (FA) and mean diffusivity (MD). Furthermore, the area of the PCL was determined, as well as the parameter product which refers to the widths of the PCL parallel and perpendicular to the stratum moleculare. Spearman correlation coefficient was used to demonstrate relationships between MR-parameters and type of sclerosis. In comparison to no-HS specimens, the PCL was significantly narrower in HS type 1 and HS type 2 hippocampi. This narrowing affected the entire cornu ammonis sector (CA) 1 in HS type 1, while it was limited to the upper half of CA1 in direction to CA2 in HS type 2. The parameter product median increased from 0.43 to 1.67 and 2.91 mm2 for HS type 1, HS type 2 and no-HS, respectively. Correlation coefficients were significant for the PCL parameters product (0.73), area (0.71), T2*-time (-0.67), FA (0.65) and ADC (0.55). Our initial results suggest that HS type 1, HS type 2 and no-HS subtypes can be distinguished from each other using ex vivo UHF MRI based on T2-weighted morphologic images and the assessment of the parameter product. Upon clinical translation, UHF-MRI may provide a promising technique for the preoperative differentiation of HS subtypes in patients.

Parkinson's disease (PD) is the second most common neurodegenerative disorder after Alzheimer's disease, affecting 0.7% of the elderly population (defined as over 65 years of age). PD is clinically characterized by resting tremor, muscular rigidity, hypokinesia and postural instability. These motor symptoms result largely from the deficiency or dysfunction of dopaminergic neurons in the substantia nigra. Histopathological analysis reveals depletion of dopaminergic neurons as well as eosinophilic intracytoplasmic inclusions (Lewy bodies) in surviving neurons of the substantia nigra and other brain regions. The molecular pathogenesis is linked to protein misfolding by compromised alpha-synuclein and/or related proteins (synucleinopathy). Therefore, successful therapy of motor symptoms aims for the restoration of dopaminergic neurotransmission. Pharmacological drug treatment is usually effective only at an early stage of the disease but cannot halt progressive neuronal degeneration. With recent developments in stem cell technology, cell repair or replacement approaches came into focus. Here, we review new therapeutic strategies resulting from the innate propensity of the adult brain to generate new neurons, either by pharmacological stimulation of endogenous adult stem cell population or exogenous cell transplantation modalities.

Today, there is increasing evidence that PPARγ agonists, including thiazolidinediones (TDZs) and nonthiazolidinediones, block the motility and invasiveness of glioma cells and other highly migratory tumor entities. However, the mechanism(s) by which PPARγ activators mediate their antimigratory and anti-invasive properties remains elusive. This letter gives a short review on the debate and adds to the current knowledge by applying a PPARγ inactive derivative of the TDZ troglitazone (Rezulin) which potently counteracts experimental glioma progression in a PPARγ independent manner.

Background and Objective: Patients with presumed nonlesional focal epilepsy-based on either MRI or histopathologic findings-have a lower success rate of epilepsy surgery compared with lesional patients. In this study, we aimed to characterize a large group of patients with focal epilepsy who underwent epilepsy surgery despite a normal MRI and had no lesion on histopathology. Determinants of their postoperative seizure outcomes were further studied., Methods: We designed an observational multicenter cohort study of MRI-negative and histopathology-negative patients who were derived from the European Epilepsy Brain Bank and underwent epilepsy surgery between 2000 and 2012 in 34 epilepsy surgery centers within Europe. We collected data on clinical characteristics, presurgical assessment, including genetic testing, surgery characteristics, postoperative outcome, and treatment regimen., Results: Of the 217 included patients, 40% were seizure-free (Engel I) 2 years after surgery and one-third of patients remained seizure-free after 5 years. Temporal lobe surgery (adjusted odds ratio [AOR]: 2.62; 95% CI 1.19-5.76), shorter epilepsy duration (AOR for duration: 0.94; 95% CI 0.89-0.99), and completely normal histopathologic findings-versus nonspecific reactive gliosis-(AOR: 4.69; 95% CI 1.79-11.27) were significantly associated with favorable seizure outcome at 2 years after surgery. Of patients who underwent invasive monitoring, only 35% reached seizure freedom at 2 years. Patients with parietal lobe resections had lowest seizure freedom rates (12.5%). Among temporal lobe surgery patients, there was a trend toward favorable outcome if hippocampectomy was part of the resection strategy (OR: 2.94; 95% CI 0.98-8.80). Genetic testing was only sporadically performed., Discussion: This study shows that seizure freedom can be reached in 40% of nonlesional patients with both normal MRI and histopathology findings. In particular, nonlesional temporal lobe epilepsy should be regarded as a relatively favorable group, with almost half of patients achieving seizure freedom at 2 years after surgery-even more if the hippocampus is resected-compared with only 1 in 5 nonlesional patients who underwent extratemporal surgery. Patients with an electroclinically identified focus, who are nonlesional, will be a promising group for advanced molecular-genetic analysis of brain tissue specimens to identify new brain somatic epilepsy genes or epilepsy-associated molecular pathways.