Glycosylation spectral signatures for glioma grade discrimination using Raman spectroscopy.

BACKGROUND: Gliomas are the most common brain tumours with the high-grade glioblastoma representing the most aggressive and lethal form. Currently, there is a lack of specific glioma biomarkers that would aid tumour subtyping and minimally invasive early diagnosis. Aberrant glycosylation is an important post-translational modification in cancer and is implicated in glioma progression. Raman spectroscopy (RS), a vibrational spectroscopic label-free technique, has already shown promise in cancer diagnostics. METHODS: RS was combined with machine learning to discriminate glioma grades. Raman spectral signatures of glycosylation patterns were used in serum samples and fixed tissue biopsy samples, as well as in single cells and spheroids. RESULTS: Glioma grades in fixed tissue patient samples and serum were discriminated with high accuracy. Discrimination between higher malignant glioma grades (III and IV) was achieved with high accuracy in tissue, serum, and cellular models using single cells and spheroids. Biomolecular changes were assigned to alterations in glycosylation corroborated by analysing glycan standards and other changes such as carotenoid antioxidant content. CONCLUSION: RS combined with machine learning could pave the way for more objective and less invasive grading of glioma patients, serving as a useful tool to facilitate glioma diagnosis and delineate biomolecular glioma progression changes.

Molecular changes in the absence of severe pathology in the pulvinar in dementia with Lewy bodies.

BACKGROUND: Dementia with Lewy bodies is characterized by transient clinical features, including fluctuating cognition and visual hallucinations, implicating dysfunction of cerebral hub regions, such as the pulvinar nuclei of the thalamus. However, the pulvinar is typically only mildly affected by Lewy body pathology in dementia with Lewy bodies, suggesting additional factors may account for its proposed dysfunction. METHODS: We conducted a comprehensive analysis of postmortem pulvinar tissue using whole-transcriptome RNA sequencing, protein expression analysis, and histological evaluation. RESULTS: We identified 321 transcripts as significantly different between dementia with Lewy bodies cases and neurologically normal controls, with gene ontology pathway analysis suggesting the enrichment of transcripts related to synapses and positive regulation of immune functioning. At the protein level, proteins related to synaptic efficiency were decreased, and general synaptic markers remained intact. Analysis of glial subpopulations revealed astrogliosis without activated microglia, which was associated with synaptic changes but not neurodegenerative pathology. DISCUSSION: These results indicate that the pulvinar, a region with relatively low Lewy body pathological burden, manifests changes at the molecular level that differ from previous reports in a more severely affected region. We speculate that these alterations result from neurodegenerative changes in regions connected to the pulvinar and likely contribute to a variety of cognitive changes resulting from decreased cortical synchrony in dementia with Lewy bodies. © 2018 International Parkinson and Movement Disorder Society.

Doublecortin expression in CD8+ T-cells and microglia at sites of amyloid-ß plaques: A potential role in shaping plaque pathology?

INTRODUCTION: One characteristic of Alzheimer's disease is the formation of amyloid-ß plaques, which are typically linked to neuroinflammation and surrounded by inflammatory cells such as microglia and infiltrating immune cells. METHODS: Here, we describe nonneurogenic doublecortin (DCX) positive cells, DCX being generally used as a marker for young immature neurons, at sites of amyloid-ß plaques in various transgenic amyloid mouse models and in human brains with plaque pathology. RESULTS: The plaque-associated DCX+ cells were not of neurogenic identity, instead most of them showed coexpression with markers for microglia (ionized calcium-binding adapter molecule 1) and for phagocytosis (CD68 and TREM2). Another subpopulation of plaque-associated DCX+ cells was negative for ionized calcium-binding adapter molecule 1 but was highly positive for the pan-leukocyte marker CD45. These hematopoietic cells were identified as CD3-and CD8-positive and CD4-negative T-cells. DISCUSSION: Peculiarly, the DCX+/ionized calcium-binding adapter molecule 1+ microglia and DCX+/CD8+ T-cells were closely attached, suggesting that these two cell types are tightly interacting and that this interaction might shape plaque pathology.

Specific patterns of neuronal loss in the pulvinar nucleus in dementia with lewy bodies.

BACKGROUND: Complex visual hallucinations occur in 70%-80% of dementia with Lewy bodies patients and significantly affect well-being. Despite the prevalence of visual hallucinations in dementia with Lewy bodies, the neuropathological basis of this phenomenon is poorly understood. The pulvinar nucleus of the thalamus has not previously been neuropathologically examined, but has been linked to visual hallucinations in dementia with Lewy bodies. The objective of this study was to investigate whether neuropathological or morphometric changes occur in the pulvinar nucleus in dementia with Lewy bodies cases that may contribute to visual hallucinations. METHODS: Postmortem pulvinar tissue was acquired from 8 individuals with dementia with Lewy bodies, 8 with Alzheimer's disease, and 8 control cases and was analyzed using stereological and quantitative neuropathological techniques. RESULTS: Lewy body pathology was present throughout the pulvinar in dementia with Lewy bodies but was most severe in the medial pulvinar. Neuronal loss was found in the lateral pulvinar in dementia with Lewy bodies and Alzheimer's disease but was more severe in dementia with Lewy bodies. CONCLUSIONS: The pulvinar has an important role in visual attention, visual target selection and affective visual perception. These functions are thought to be deficient in dementia with Lewy bodies and may contribute a vulnerability to visual hallucinations. Therefore, this study has demonstrated neuropathological changes that may promote the manifestation of visual hallucinations in dementia with Lewy bodies. © 2017 International Parkinson and Movement Disorder Society.

Quantitative neuropathology: an update on automated methodologies and implications for large scale cohorts.

A tissue microarray (TMA) has previously been developed for use in assessment of neurodegenerative diseases. We investigated the variation of pathology loads in semi-quantitative score categories and how pathology load related to disease progression. Post-mortem tissue from 146 cases were used; Alzheimer's disease (AD) (n = 36), Lewy body disease (LBD) (n = 56), mixed AD/dementia with Lewy bodies (n = 14) and controls (n = 40). TMA blocks (one per case) were constructed using tissue cores from 15 brain regions including cortical and subcortical regions. TMA tissue sections were stained for hyperphosphorylated tau (HP-T), ß amyloid and α-synuclein (αsyn), and quantified using an automated image analysis system. Cases classified as Braak stage VI displayed a wide variation in HP-T pathology in the entorhinal cortex (interquartile range 4.13-44.03%). The interquartile range for ß amyloid in frontal cortex in cases classified as Thal phase 5 was 6.75-17.03% and for αsyn in the cingulate in cases classified as McKeith neocortical LBD was 0.04-0.58%. In AD and control cases, HP-T load predicted the Braak stage (p < 0.001), ß amyloid load predicted Thal phase (p < 0.001) and αsyn load in LBD cases predicted McKeith type of LBD (p < 0.001). Quantitative data from TMA assessment highlight the range in pathological load across cases classified with 'severe' pathology and is beneficial to further elucidate the heterogeneity of neurodegenerative diseases. Quantifying pathology in multiple brain regions may allow identification of novel clinico-pathological phenotypes for the improvement of intra vitam stratification of clinical cohorts according to underlying pathologies.

Neuronal Loss and Α-Synuclein Pathology in the Superior Colliculus and Its Relationship to Visual Hallucinations in Dementia with Lewy Bodies.

OBJECTIVE: Patients with dementia with Lewy bodies (DLB) often experience visual hallucinations, which are related to decreased quality of life for patients and increased caregiver distress. The pathologic changes that contribute to visual hallucinations are not known, but several hypotheses implicate deficient attentional processing. The superior colliculus has a role in visual attention and planning eye movements and has been directly implicated in several models of visual hallucinations. Therefore, the present study sought to identify neurodegenerative changes that may contribute to hallucinations in DLB. METHODS: Postmortem superior colliculus tissue from 13 comparison, 10 DLB, and 10 Alzheimer disease (AD) cases was evaluated using quantitative neuropathologic methods. RESULTS: α-Synuclein and tau deposition were more severe in deeper layers of the superior colliculus. DLB cases had neuronal density reductions in the stratum griseum intermedium, an important structure in directing attention toward visual targets. In contrast, neuronal density was reduced in all laminae of the superior colliculus in AD. CONCLUSION: These findings suggest that regions involved in directing attention toward visual targets are subject to neurodegenerative changes in DLB. Considering several hypotheses of visual hallucinations implicating dysfunctional attention toward external stimuli, these findings may provide evidence of pathologic changes that contribute to the manifestation of visual hallucinations in DLB.

Cellular morphometry in late-life depression: a review of postmortem studies.

The impact of major depression in late life is considerable and set to intensify with a worldwide shift in demographic profile toward an elderly population. Although the precise neurobiological mechanisms are not fully understood, a significant body of clinical, epidemiological, and imaging data have suggested divergent pathophysiological pathways underlie depression in late life, when compared with younger patients. Neuroimaging studies have demonstrated significant increases in white matter hyperintensities in late-life depression in several key areas involved in affective circuitry. Postmortem cellular morphometry studies have played a vital role in the identification of discrete changes in the brain microstructure in depression. This review draws together such postmortem studies, which have utilized tissue from younger/mixed age and late-life depressed patients. These findings have suggested varying neuronal and glial cell pathology in depression between different age cohorts. This age-related disparity may suggest different pathophysiological basis for depression, with vascular factors playing a potentially greater role in late life.

Advancing Brain Research through Surface-Enhanced Raman Spectroscopy (SERS): Current Applications and Future Prospects.

Surface-enhanced Raman spectroscopy (SERS) has recently emerged as a potent analytical technique with significant potential in the field of brain research. This review explores the applications and innovations of SERS in understanding the pathophysiological basis and diagnosis of brain disorders. SERS holds significant advantages over conventional Raman spectroscopy, particularly in terms of sensitivity and stability. The integration of label-free SERS presents promising opportunities for the rapid, reliable, and non-invasive diagnosis of brain-associated diseases, particularly when combined with advanced computational methods such as machine learning. SERS has potential to deepen our understanding of brain diseases, enhancing diagnosis, monitoring, and therapeutic interventions. Such advancements could significantly enhance the accuracy of clinical diagnosis and further our understanding of brain-related processes and diseases. This review assesses the utility of SERS in diagnosing and understanding the pathophysiological basis of brain disorders such as Alzheimer's and Parkinson's diseases, stroke, and brain cancer. Recent technological advances in SERS instrumentation and techniques are discussed, including innovations in nanoparticle design, substrate materials, and imaging technologies. We also explore prospects and emerging trends, offering insights into new technologies, while also addressing various challenges and limitations associated with SERS in brain research.

Uncovering potential diagnostic and pathophysiological roles of α-synuclein and DJ-1 in melanoma.

BACKGROUND: Melanoma, the most lethal skin cancer type, occurs more frequently in Parkinson's disease (PD), and PD is more frequent in melanoma patients, suggesting disease mechanisms overlap. α-synuclein, a protein that accumulates in PD brain, and the oncogene DJ-1, which is associated with PD autosomal recessive forms, are both elevated in melanoma cells. Whether this indicates melanoma progression or constitutes a protective response remains unclear. We hereby investigated the molecular mechanisms through which α-synuclein and DJ-1 interact, suggesting novel biomarkers and targets in melanoma. METHODS: The Cancer Genome Atlas (TCGA) expression profiles derived from UCSC Xena were used to obtain α-synuclein and DJ-1 expression and correlated with survival in skin cutaneous melanoma (SKCM). Immunohistochemistry determined the expression in metastatic melanoma lymph nodes. Protein-protein interactions (PPIs) and molecular docking assessed protein binding and affinity with chemotherapeutic drugs. Further validation was performed using in vitro cellular models and ELISA immunoassays. RESULTS: α-synuclein and DJ-1 were upregulated in primary and metastatic SKCM. Aggregated α-synuclein was selectively detected in metastatic melanoma lymph nodes. α-synuclein overexpression in SK-MEL-28 cells induced the expression of DJ-1, supporting PPI and a positive correlation in melanoma patients. Molecular docking revealed a stable protein complex, with differential binding to chemotherapy drugs such as temozolomide, dacarbazine, and doxorubicin. Parallel reduction of both proteins in temozolomide-treated SK-MEL-28 spheroids suggests drug binding may affect protein interaction and/or stability. CONCLUSION: α-synuclein, together with DJ-1, may play a role in melanoma progression and chemosensitivity, constituting novel targets for therapeutic intervention, and possible biomarkers for melanoma.

The diagnostic and prognostic potential of the EGFR/MUC4/MMP9 axis in glioma patients.

Glioblastoma is the most aggressive form of brain cancer, presenting poor prognosis despite current advances in treatment. There is therefore an urgent need for novel biomarkers and therapeutic targets. Interactions between mucin 4 (MUC4) and the epidermal growth factor receptor (EGFR) are involved in carcinogenesis, and may lead to matrix metalloproteinase-9 (MMP9) overexpression, exacerbating cancer cell invasiveness. In this study, the role of MUC4, MMP9, and EGFR in the progression and clinical outcome of glioma patients was investigated. Immunohistochemistry (IHC) and immunofluorescence (IF) in fixed tissue samples of glioma patients were used to evaluate the expression and localization of EGFR, MMP9, and MUC4. Kaplan-Meier survival analysis was also performed to test the prognostic utility of the proteins for glioma patients. The protein levels were assessed with enzyme-linked immunosorbent assay (ELISA) in serum of glioma patients, to further investigate their potential as non-invasive serum biomarkers. We demonstrated that MUC4 and MMP9 are both significantly upregulated during glioma progression. Moreover, MUC4 is co-expressed with MMP9 and EGFR in the proliferative microvasculature of glioblastoma, suggesting a potential role for MUC4 in microvascular proliferation and angiogenesis. The combined high expression of MUC4/MMP9, and MUC4/MMP9/EGFR was associated with poor overall survival (OS). Finally, MMP9 mean protein level was significantly higher in the serum of glioblastoma compared with grade III glioma patients, whereas MUC4 mean protein level was minimally elevated in higher glioma grades (III and IV) compared with control. Our results suggest that MUC4, along with MMP9, might account for glioblastoma progression, representing potential therapeutic targets, and suggesting the 'MUC4/MMP9/EGFR axis' may play a vital role in glioblastoma diagnostics.

Differential regulation of psychostimulant-induced gene expression of brain derived neurotrophic factor and the immediate-early gene Arc in the juvenile and adult brain.

Psychostimulant drugs are widely used in children for the treatment of attention-deficit/hyperactivity disorder. Recent animal studies have suggested that exposure to these agents in early life could be detrimental to brain development. Here, for the first time, the effect of methylphenidate (MPH) and D-amphetamine (AMPH) on the expression of two key genes for neuronal development and plasticity, brain-derived neurotrophic factor (bdnf) and the effector immediate early gene activity-regulated, cytoskeletal-associated protein (Arc), was examined in both juvenile and adult rats. Both MPH [2 mg/kg, intraperitoneal (i.p.)] and AMPH (0.5 mg/kg, i.p.) induced marked decreases of bdnf mRNA in hippocampal and cortical brain regions of juveniles, whereas effects in adults were significantly less (hippocampus) or opposite (frontal cortex). In comparison, Arc mRNA was decreased (hippocampus and parietal cortex), largely unaffected (frontal cortex) or increased (striatum) in juveniles, whereas in adults, Arc mRNA increased in most brain regions. MPH-induced locomotion was also measured, and showed a much smaller increase in juveniles than in adults. In summary, our data show that the effects of MPH and AMPH on expression of the neurodevelopmentally important genes, bdnf and Arc, differ markedly in juvenile and adult rats, with juveniles showing evidence of brain region-specific decreases in both genes. These age-dependent effects on gene expression may be linked with the reported long-term harmful effects of psychostimulants in animal models.

Analysis of primary visual cortex in dementia with Lewy bodies indicates GABAergic involvement associated with recurrent complex visual hallucinations.

Dementia with Lewy bodies (DLB) patients frequently experience well formed recurrent complex visual hallucinations (RCVH). This is associated with reduced blood flow or hypometabolism on imaging of the primary visual cortex. To understand these associations in DLB we used pathological and biochemical analysis of the primary visual cortex to identify changes that could underpin RCVH. Alpha-synuclein or neurofibrillary tangle pathology in primary visual cortex was essentially absent. Neurone density or volume within the primary visual cortex in DLB was also unchanged using unbiased stereology. Microarray analysis, however, demonstrated changes in neuropeptide gene expression and other markers, indicating altered GABAergic neuronal function. Calcium binding protein and GAD65/67 immunohistochemistry showed preserved interneurone populations indicating possible interneurone dysfunction. This was demonstrated by loss of post synaptic GABA receptor markers including gephyrin, GABARAP, and Kif5A, indicating reduced GABAergic synaptic activity. Glutamatergic neuronal signalling was also altered with vesicular glutamate transporter protein and PSD-95 expression being reduced. Changes to the primary visual cortex in DLB indicate that reduced GABAergic transmission may contribute to RCVH in DLB and treatment using targeted GABAergic modulation or similar approaches using glutamatergic modification may be beneficial.

Neuropathology of depression in Alzheimer's disease: current knowledge and the potential for new treatments.

Depression is among the most common behavioral and psychological symptoms of dementia, and leads to more rapid decline and higher mortality. Treatment for depression in dementia has centered on conventional antidepressant drug treatment based around the monoamine hypothesis of depression. However, recent major studies have suggested that conventional antidepressant treatments that aim to correct underlying deficits in monoamine neurotransmitters are not effective for depression in dementia. Postmortem studies have also suggested that depression in dementia does not arise from serotonergic or noradrenergic abnormalities, or indeed from the degenerative pathology associated with Alzheimer's disease. In contrast, considerable recent evidence has suggested that alterations in glutamatergic transmission may contribute to the pathophysiology of depression. This supports the view that treatment-resistant depressed patients, such as many dementia patients, may benefit from agents affecting glutamate transmission. This review will thus draw together the wealth of pathological data examining the basis of depression in Alzheimer's disease and relate this to current thinking on treatment, with the aim of generating discussion on potential novel therapeutic strategies.

The role of 5-hydroxytryptamine receptor subtypes in the regulation of brain-derived neurotrophic factor gene expression.

OBJECTIVES: The study aims to investigate the role of 5-hydroxytryptamine receptor subtypes in mediating the inhibitory effect of the selective serotonin reuptake inhibitor (fluoxetine on brain-derived neurotrophic factor gene (bdnf) expression in rat hippocampus. METHODS: In situ hybridization was used for regional determination of bdnf expression levels in hippocampal brain slices from normal, lesioned (5-hydroxytryptamine or noradrenaline) or adrenalectomized rats; treated with fluoxetine and/or 5-hydroxytryptamine selective ligands. KEY FINDINGS: Our study shows that the transient fluoxetine-induced down-regulation of bdnf gene expression depends on an intact 5-hydroxytryptamine but not noradrenaline system or circulating glucocorticoids. Pretreatment with the 5-hydroxytryptamine4 antagonist SB-204070 blocked the overall fluoxetine-induced inhibition of bdnf levels in hippocampus, while pretreatment with the 5-hydroxytryptamine2 antagonists ketanserin had an effect in the CA3 but not in the dentate gyrus sub-region of hippocampus. The 5-hydroxytryptamine1A antagonist WAY-100635 and the 5-hydroxytryptamine3 antagonist granisetron were both ineffective. CONCLUSIONS: Our study found strong support for a primary effect of 5-hydroxytryptamine but not noradrenaline or circulating glucocorticoids in the mediation of fluoxetine-induced down-regulation of bdnf expression. More specifically, we also show that 5-hydroxytryptamine4 receptor-stimulation seems to play a pivotal role in this effect.

Effects of repeated 5-HT6 receptor stimulation on BDNF gene expression and cell survival.

In support of the neurotrophic hypothesis of depression chronic antidepressant drug treatment increases brain-derived neurotrophic factor (bdnf) gene expression and neurogenesis. Regarding 5-HT active drugs, the 5-HT receptor behind these effects remains unidentified. Here we report the effect of repeated 5-HT6-receptor stimulation on bdnf expression and cell survival. The previously reported acute stimulatory action of the selective 5-HT6 agonist LY-586713 on hippocampal bdnf expression was still present following sub-chronic (4 days), but not chronic (14 days), treatment. The effect on 5-HT6-mediated cell survival was also dependent on a similar length of treatment. Hence, our study found no support for a primary effect of 5-HT6 receptors in the mediation of chronic antidepressant drug-induced up-regulation of bdnf expression or neurogenesis.

Effects of GABAB ligands alone and in combination with paroxetine on hippocampal BDNF gene expression.

Brain-derived neurotrophic factor (BDNF) has been suggested as a target for antidepressant treatment and chronic antidepressant drug administration shows a 'biphasic effect' on BDNF mRNA in rat hippocampus (transient decrease followed by an increase). In comparison, following acute administration only, an inhibitory action on BDNF gene expression is detected. The present study aimed to understand the mechanism behind the acute inhibitory action on BDNF gene expression by investigating the possible involvement of γ-aminobutyric acid (GABA) receptors in mediating this effect. Rats were injected with either saline, the GABA(A) selective compound 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol (THIP), the benzodiazepine flunitrazepam or the GABA(B) selective compound baclofen. BDNF mRNA levels were measured 4h later using in-situ hybridization. Baclofen (10mg/kg, i.p.), but not THIP (10mg/kg, i.p.) or flunitrazepam (10mg/kg, i.p.), administration resulted in significant inhibition of BDNF mRNA expression in the cornu ammonis 3 and dentate gyrus but not in the cornu ammonis 1 region of the hippocampus. The inhibitory effect of baclofen on hippocampal BDNF mRNA expression was significantly attenuated by pre-treatment the selective GABA(B) antagonists, CGP 46381 and CGP 55845 (10mg/kg, i.p.). The inhibitory action by the selective serotonin re-uptake inhibitor (SSRI) paroxetine on hippocampal BDNF mRNA was also attenuated by CGP 46381. Our findings suggest a role for GABA(B), but not GABA(A), receptor-mediated mechanisms in the inhibitory regulation of basal hippocampal BDNF gene expression. Our results indicate that GABA(B) receptor activation may play a role in the antidepressant drug-induced inhibition of BDNF gene expression in the hippocampus.

Morphometric changes in early- and late-life major depressive disorder: evidence from postmortem studies.

BACKGROUND: Neuroimaging studies have revealed structural and functional changes in brain regions associated with major depressive disorder (MDD). These abnormalities appear to be more common and extensive in patients with late-life depression than in younger patients. It has therefore been hypothesized that different morphometric and pathological changes may be associated with MDD, depending on age. METHODS: This review stratifies the findings of the various studies on cell morphology in MDD according to age and assesses any possible differences in neuronal and glial cell changes in younger and older age groups. RESULTS: Recent morphological studies in postmortem tissue have revealed alterations in neuron and glial cell populations in the frontal and subcortical circuitry associated with depression. These may differ by age, with glial reduction consistently reported in younger groups in cortical areas and neuronal changes identified in studies with older subjects. CONCLUSIONS: Apparent differences in the morphological changes between younger and elderly patients may suggest a differing pathological basis in MDD, dependent on age.