Oral Abstracts

Rachel Waller1, Paul R Heath1, Julie E Simpson1, Yoshiki Hase2, Rajesh N Kalaria2, Stephen B Wharton1 1The University of Sheffield, Sheffield, United Kingdom; 2Newcastle University, Newcastle upon Tyne, United Kingdom Background: Stroke doubles the risk of dementia in the over 65s and around a quarter of stroke survivors develop progressive cognitive decline more than 3 months after the stroke, eventually leading to vascular dementia. Impaired executive function is a characteristic fea‐ ture of vascular dementia, including post stroke dementia (PSD). PSD is associated with pathology of the anterior cognitive circuits, but the mechanisms leading to PSD are unclear. This study aims to find novel pathways by which cell damage in the brain can give rise to PSD. Methods: Dorsal lateral prefrontal cortex (DLPFC) of 10 control, 10 PSD, and 10 post‐ stroke non‐ demented (PSND) cases from the Cognitive Function After STROKE (CogFAST) study were sub‐ jected to laser capture microdissection to obtain RNA enriched for; astrocytes, endothelium and neurons. Microarray analysis was performed to identify differentially expressed genes in cortical neurons and the frontal white matter cells of the gliovascular unit. Results: We identified altered gene expression associated with PSD. Pathway analysis demonstrated a decrease in communication and signalling pathways in astrocytes, whilst endothelium showed an in‐ crease in inflammatory pathways and altered metabolism. Neurons showed reduction in pathways for metabolism and endocytosis. These changes will be confirmed at both the cellular (immunohisto‐ chemistry) and molecular (NanoString) level. Conclusions: Within the anterior cognitive circuit, cortical neurons and cells of the gliovascular unit showed alterations in their ability to function normally and communicate with each other. These findings provide new information about how stroke can lead to dementia, and may identify novel targets for future studies.

expressed genes (DEGs) were assessed by microarray analysis and altered cellular pathways determined using Integrated Molecular Pathway Level Analysis. Gene expression changes in neurones were validated by a second method, the NanoString nCounter platform.
Neurones showed 912 DEGs between samples from diabetic and non-diabetic individuals. Cell pathway changes included alterations in insulin signalling, cell cycle, cellular senescence, inflammatory mediators, and components of the mitochondrial respiratory electron transport chain. There were 2,202 and 1,227 DEGs in astrocytes and endothelial cells respectively. There was little overlap in individual DEGs in different cell types, but pathway analysis demonstrated that impaired insulin signalling was shared by neurovascular unit cells.
Additionally, apoptotic pathway changes in astrocytes and dysregulation of advanced glycation end-product signalling in endothelial cells were identified. Transcriptomic analysis identified changes in key cellular pathways associated with T2D that may contribute to neuronal damage and dysfunction. These alterations potentially contribute to a diabetic dementia, and may provide novel avenues for investigation.

O3 | The inflammatory environment in glioblastoma
Jeanette L. Norman 1 , Kastytis Sidlauskas 1,2 , Sean Pellow 1 , Reece Savage 1 , David S. Chatelet 3 , Mark Fabian 4 , Jeng Ching 5 , Paul axonal injury can be detected in patients with near instantaneous/ rapid death. Forty-nine deaths following road traffic collision (RTC) where we have reliable information concerning the time between the incident and death, were retrospectively studied. Thirty seven patients (group 1) died virtually instantaneously or very rapidly at the scene. A further three died between 30 minutes to 11 hours (group 2) and eight between 2 and 31 days (group 3). Brains from four patients who died instantaneously due to sudden unexpected death in epilepsy were used as a non-traumatic control group. The brains were comprehensively examined, and βAPP immunohistochemistry carried out on sections from a number of brain areas. βAPP immunoreactivity was demonstrated in in 35/37 brains in group 1, albeit with a low frequency and in a variable pattern and with more intensity and frequency in all brains of group 2 and 7/8 brains from group 3, compared with no βAPP immuno reactivity in any of the control group.
The results suggest axonal injury can be detected even in those who died near instantaneously following fatal road traffic collision. Although DNA methylation changes have been consistently associated with neurodegenerative diseases, little is known in FTLD. The main goal of this study was to explore FTLD-TDP DNA methylation signatures.

Methods:
We dissected frontal cortex grey matter tissue of FTLD-TDP cases and heathy controls. We generated genome-wide DNA methylation profiles using the Illumina EPIC arrays (>850,000 methylation sites). We performed differential methylation analysis and weighted gene correlation network analysis to identify FTLD-TDP methylation signatures. We compared these signatures with available FTLD-TDP proteomics data.

Results:
We found a DNA methylation signature inversely correlated with the disease age at onset (r = −0.7, p = 3 × 10 −4 ), and nominally correlated with the FTLD-TDP status (r = 0.44, p = 0.04), and another signature inversely correlated with the disease duration (r = −0.67, p = 7 × 10 −4 ). These signatures contain genetic risk factors for FTLD (e.g. SORT1 and ATXN2), and genes encoding for proteins that are dysregulated in FTLD-TDP. Functional enrichment analysis of these signatures suggests that higher levels of methylation in genes involved in axon guidance and RNA metabolism pathways may lead to an earlier appearance of the disease, and that with increasing disease duration there is a decrease in DNA methylation levels at genes involved in infection and transcription related pathways.

Conclusions:
Our data suggests that DNA methylation plays a role in the pathogenesis of FTLD-TDP.  We have validated these events using PCR amplification and gel electrophoresis in both SH-SY5Y cells with hnRNP K knockdown and post-mortem human brain tissue, where a cryptic exon within FAM1602B was found to be more common in FTLD cases exhibiting hnRNP K mislocalistion compared to controls. Therefore, the mislocalisation of hnRNP K from the nucleus to the cytoplasm in FTLD is associated with the inclusion of cryptic exons in multiple gene targets. This suggests that similarly to TDP-43, hnRNP K plays a crucial role in RNA processing and splicing regulation which when impaired could lead to transcriptional dysregulation and disease pathogenesis. We focused on (I) astrocytic tau inclusions in striatum, frontal and occipital regions, and (II) neuronal and oligodendroglia tau inclusions in globus pallidus, subthalamic nucleus, and cerebellum. Thirty-two cases (91%) readily conformed to the new staging system, ranging from stage 2 to 6 and across clinical phenotypes of PSP. Neuropathology stages correlated with clinical severity at death using both PSPRS and ACE-R, weighted for the interval between last assessment and donation. Our study supports the proposed sequential distribution of tau-aggregates in PSP pathology, and the hypothesised relationship between clinical and neuropathological severity. For future studies, in order to standardise rating between centres, we propose a set of operational criteria for region-specific thresholds of tau burden, and a visual guide. For example, the serotonergic pathway originates from the dorsal raphe nucleus (DRN) in the rostral pons, projecting across the entire brain to regulate a wide range of physiological functions, such as appetite, vomiting, body temperature, sleep-wake cycle, as well as emotional state and cognition, specifically learning and memory.

O12 | Interneuron vulnerability in Lewy body dementia
In this study, we have examined the serotonergic projections from the DRN to the frontal cortex in a cohort of patients with DLB, PDD or PD without dementia. Immunohistochemical and image analysis techniques have been performed using a neuronal marker, tryptophan hydroxylase-2 (TPH2), to enable a comparison between neuronal quantity in each of the clinical subgroups. Hence, the serotonergic pathway may be implicated in the pathogenesis of LBDs and potentially correlate with cognitive fluctuations and dysfunctions in emotional state.