scholarly journals Characterisation of the invasive tumour niche using astrocyte-glioblastoma organoids and decellularised human brain

2019 ◽  
Vol 21 (Supplement_4) ◽  
pp. iv7-iv7
Author(s):  
Mohammed Diksin ◽  
Jonathan Rowlinson ◽  
Alexandar Kondrashov ◽  
Chris Denning ◽  
Jamie Hughes ◽  
...  
Keyword(s):  
Human Brain ◽  
Cross Talk ◽  
Secondary Ion Mass Spectrometry ◽  
Brain Parenchyma ◽  
Brain Extract ◽  
Malignant Cells ◽  
3D Scaffold ◽  
Glioma Invasion ◽  
Naive Patient ◽  
Invasive Margin

Abstract Glioblastoma therapeutic challenges are in considerable part due to myriad survival adaptations and mechanisms, which allow malignant cells to repurpose signalling pathways within discreet microenvironments. These Darwinian adaptations facilitate invasion into brain parenchyma and perivascular space or promote evasion from repressive factors that represent anti-cancer defence mechanisms. We hypothesised that pre-clinical modelling of glioma invasion by recapitulating early events occurring immediately after surgery at the glioblastoma invasive margin, could reveal the cross-talk between malignant cells and the surrounding healthy astrocytes, which facilitates tumour recurrence. We first generated transgenic H1-derived neural stem cells using CRISPR/Cas9-mediated knock-in of the YFP reporter gene under the control of the GFAP promoter. Reproducible ultrahigh-throughput AggreWells™ (19,200 micro-wells per 24-well plate) were used to create astrocyte-glioblastoma organoids, which we term ‘Gliomasphere Matrices’. YFP-labelled astrocytes were co-cultured with 10 treatment-naïve patient-derived cell lines isolated from the 5-aminolevulinic (5ALA)-determined glioblastoma invasive margin. Co-cultures were seeded upon on a sequentially constructed, time-of-flight secondary ion mass spectrometry (ToF-SIMS)-characterised 3D scaffold, composed of decellularised human brain extract with defined PEGDA hydrogel. YFP-astrocytes were purified from each of the 10 Gliomasphere Matrices using fluorescence-activated cell sorting (FACS) after 6- and 10-days co-culture. RNAseq profiling to address both putative astrocytic reprogramming by invasive glioblastoma cells and gene expression changes intrinsic to tumour cells will be discussed in relation to RNAseq data from patient-derived 5ALA FACS-purified glioblastoma invasive margin tissue. This novel multi-faceted model offers a unique opportunity to recapitulate early molecular cross-talk which facilitates glioblastoma recurrence and may be utilised for high-throughput drug screening.

scholarly journals TMOD-11. CHARACTERISATION OF THE POST-SURGICAL INVASIVE TUMOUR NICHE USING ASTROCYTE-GLIOBLASTOMA ORGANOIDS AND DECELLULARISED HUMAN BRAIN

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi264-vi265
Author(s):  
Mohammed Diksin ◽  
Jonathan Rowlinson ◽  
Alexander Kondrashov ◽  
Chris Denning ◽  
Jaime Hughes ◽  
...  
Keyword(s):  
Human Brain ◽  
Rna Sequencing ◽  
Cross Talk ◽  
Chromosome 1 ◽  
Brain Extract ◽  
Malignant Cells ◽  
Sequencing Data ◽  
Glioma Invasion ◽  
Naive Patient ◽  
Invasive Margin

Abstract Glioblastoma therapeutic challenges are in considerable part due to myriad survival mechanisms which allow malignant cells to repurpose signalling pathways within discreet microenvironments. These Darwinian adaptations facilitate invasion into brain parenchyma and perivascular space. We hypothesised that pre-clinical modelling of glioma invasion by recapitulating early events occurring immediately after surgery at the glioblastoma invasive margin, could reveal the cross-talk between malignant cells and surrounding healthy astrocytes. We first generated transgenic H1-derived neural stem cells using CRISPR/Cas9-mediated knock-in of the YFP reporter gene under the control of the GFAP promoter at the AAVS1 safe harbour locus. Reproducible ultrahigh-throughput AggreWells™ (7200 mini-wells per plate) were used to create astrocyte-glioblastoma organoids, which we term ‘Gliomasphere Matrices’. YFP-labelled astrocytes were co-cultured with 10 treatment-naïve patient-derived cell lines isolated from the 5-aminolevulinic (5ALA)-determined glioblastoma invasive margin. Co-cultures were seeded upon a sequentially constructed, time-of-flight secondary ion mass spectrometry (ToF-SIMS)-characterised decellularised human brain extract. YFP-astrocytes were purified from each of the 10 Gliomasphere Matrices using fluorescence-activated cell sorting (FACS) after 6- and 10-days co-culture. RNA-sequencing of the putatively reprogrammed YFP-astrocytes showed the characteristic expression of canonical key regulators of multiple malignant diseases including high-grade glioma such as SND1 and EFNB2 in addition to the identification of a single novel marker located at chromosome 1 (C1orf61), highly expressed in malignant glioma when compared to somatic cancers according to TCGA RNA-sequencing data. Differentiated YFP-astrocytes also overexpressed IFITM2 and IFITM10, known to be involved in priming resistance against pathogenic microorganisms. This ultimately suggests a fluctuating state between malignant transformation imposed by the highly infiltrative glioma cells and the counter-action of the normal astrocytes to these deleterious invasive cells. This multi-faceted model offers a unique opportunity to recapitulate early molecular cross-talk which facilitates glioblastoma recurrence and may be utilised for high-throughput drug screening.

scholarly journals Exogenous Aβ seeds induce Aβ depositions in the blood vessels rather than the brain parenchyma, independently of Aβ strain-specific information

2021 ◽  
Vol 9 (1) ◽  
Author(s):  
Tsuyoshi Hamaguchi ◽  
Jee Hee Kim ◽  
Akane Hasegawa ◽  
Ritsuko Goto ◽  
Kenji Sakai ◽  
...  
Keyword(s):  
Human Brain ◽  
Blood Vessels ◽  
Brain Homogenate ◽  
Brain Parenchyma ◽  
Brain Extract ◽  
Amyloid Angiopathy ◽  
Specific Information ◽  
Brain Extracts ◽  
Iatrogenic Transmission ◽  
The Brain

AbstractLittle is known about the effects of parenchymal or vascular amyloid β peptide (Aβ) deposition in the brain. We hypothesized that Aβ strain-specific information defines whether Aβ deposits on the brain parenchyma or blood vessels. We investigated 12 autopsied patients with different severities of Aβ plaques and cerebral amyloid angiopathy (CAA), and performed a seeding study using an Alzheimer’s disease (AD) mouse model in which brain homogenates derived from the autopsied patients were injected intracerebrally. Based on the predominant pathological features, we classified the autopsied patients into four groups: AD, CAA, AD + CAA, and less Aβ. One year after the injection, the pathological and biochemical features of Aβ in the autopsied human brains were not preserved in the human brain extract-injected mice. The CAA counts in the mice injected with all four types of human brain extracts were significantly higher than those in mice injected with PBS. Interestingly, parenchymal and vascular Aβ depositions were observed in the mice that were injected with the human brain homogenate from the less Aβ group. The Aβ and CAA seeding activities, which had significant positive correlations with the Aβ oligomer ratio in the human brain extracts, were significantly higher in the human brain homogenate from the less Aβ group than in the other three groups. These results indicate that exogenous Aβ seeds from different Aβ pathologies induced Aβ deposition in the blood vessels rather than the brain parenchyma without being influenced by Aβ strain-specific information, which might be why CAA is a predominant feature of Aβ pathology in iatrogenic transmission cases. Furthermore, our results suggest that iatrogenic transmission of Aβ pathology might occur due to contamination of brain tissues from patients with little Aβ pathology, and the development of inactivation methods for Aβ seeding activity to prevent iatrogenic transmission is urgently required.

scholarly journals CMET-14. BRAIN METASTASES: A NEW PATHOPHYSIOLOGY, A NEW TREATMENT PARADIGM, AND, PERHAPS, A NEW PROGNOSIS

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi54-vi54
Author(s):  
Richard Dowd ◽  
Tao Ouyang ◽  
Krishnamoorthy Thamburaj ◽  
Dawit Aregawi ◽  
Howard Safran ◽  
...  
Keyword(s):  
Brain Metastases ◽  
Substantial Reduction ◽  
Blood Stream ◽  
Brain Parenchyma ◽  
Neoplastic Meningitis ◽  
Normal Brain ◽  
Malignant Cells ◽  
Therapeutic Implications ◽  
Treatment Paradigm ◽  
Pre Treatment

Abstract INTRODUCTION Brain metastases are widely held to reach the CNS through the blood stream. We provide evidence that the initial site of most CNS metastases is the CSF, with subsequent invasion of brain parenchyma. We also model the therapeutic implications of this novel hypothesis. METHODS Two neuro-radiologists independently assessed whether brain metastases were contiguous with CSF spaces in 200 consecutive patients using pre-treatment MRI. CSF was examined for malignant cells in 66 newly diagnosed, previously untreated patients. We contoured normal brain MRIs of 3 patients to calculate the percentage of brain within 5 mm of a CSF space. We queried an international neoplastic meningitis database to document response of brain metastases to intra-CSF chemotherapy. RESULTS Mean age was 64.2. One hundred patients were male; 143 had lung cancer, 15 melanoma, 12 gastrointestinal, 11 breast, 9 renal, 7 bladder. Mean number of metastases was 4.63. Eighty-five percent of metastases touched a CSF space. In 67% of patients, all metastases touched a CSF space. Neither histology, number or size of metastases, nor patient age predicted contiguity with CSF spaces. In our consecutive subset of patients, 44% (10/23) with one, 46% (5/11) with two, 63% (5/8) with three, and 71% (17/24) with >3 metastases had malignant cells in the CSF (R2=0.93, p=0.037). Five of 7 patients with both brain and CSF metastases receiving only IT chemotherapy experienced a substantial reduction in the size of at least some metastases. Up to 75% of the brain parenchyma lies within 5 mm of CSF spaces. CONCLUSIONS Our data suggest that brain metastases may access the CNS through the CSF rather than the bloodstream. IT chemotherapy may treat brain metastases. We suggest that the CSF should be monitored in all patients with, or at risk for, brain metastases. True cures may require treatment of the CSF space.

scholarly journals GFAP splice variants fine-tune glioma cell invasion and tumour dynamics by modulating migration persistence.

2021 ◽  
Author(s):  
Jacco van Rheenen ◽  
Elly Hol ◽  
Claire Vennin ◽  
Jessy van Asperen ◽  
Rebeca Uceda-Castro ◽  
...  
Keyword(s):  
Splice Variants ◽  
Ex Vivo ◽  
Glioma Cells ◽  
Brain Parenchyma ◽  
Intermediate Filament Protein ◽  
Glioma Invasion ◽  
Primary Brain Tumours ◽  
Invasion Model ◽  
Filament Protein ◽  
The Brain

Glioma is the most common form of malignant primary brain tumours in adults. Their highly invasive nature makes the disease incurable to date, emphasizing the importance of better understanding the mechanisms driving glioma invasion. Glial fibrillary acidic protein (GFAP) is an intermediate filament protein that is characteristic for astrocyte- and neural stem cell-derived gliomas. Glioma malignancy is associated with changes in GFAP alternative splicing, as the canonical isoform GFAPα is downregulated in higher-grade tumours, leading to increased dominance of the GFAPδ isoform in the network. In this study, we used intravital imaging and an ex vivo brain slice invasion model. We show that the GFAPδ and GFAPα isoforms differentially regulate the tumour dynamics of glioma cells. Depletion of either isoform increases the migratory capacity of glioma cells. Remarkably, GFAPδ-depleted cells migrate randomly through the brain tissue, whereas GFAPα-depleted cells show a directionally persistent invasion into the brain parenchyma. This study shows that distinct compositions of the GFAP-network lead to specific migratory dynamics and behaviours of gliomas.

A Comparison of Plasma and Serum Factor VII Activity and the Formation of Tissue Thromboplastin

1962 ◽  
Vol 08 (02) ◽  
pp. 286-296
Author(s):  
P Fantl ◽  
E. C Osborn
Keyword(s):  
Human Brain ◽  
Human Plasma ◽  
Factor Vii ◽  
Brain Extract ◽  
Factor X ◽  
Similar Activity ◽  
Bovine Plasma ◽  
Tissue Thromboplastin ◽  
Species Specific ◽  
Glass Surfaces

Summary1. A mixture of human serum or plasma and bovine plasma free of factors VII and X gave, with human brain extract, identical clotting times.2. An assay of factor VII in materials low in prothrombin using human plasma euglobulin was devised.3. Factor VII isolated from human plasma or serum gave similar activity with human brain extract.4. From a preparation containing factors VII and X which was added to human brain extract in the average 31% factor VII and 25% factor X was recovered. This was not dependent on the activity of factors VII and X in the original preparation. This indicates that factors VII and X are in equilibrium with tissue thromboplastin.5. Factors VII and X are not species specific but a higher concentration of these factors is required for prothrombin conversion in a heterologous reaction mixture.6. Factor VII activity is identical in silicone-coated or uncoated glass surfaces.

Investigation of Precipitins to Human Brain in Sera of Psychotic Patients

1965 ◽  
Vol 111 (479) ◽  
pp. 1003-1006 ◽  
Author(s):  
Robert T. Rubin
Keyword(s):  
Human Brain ◽  
Complement Fixation ◽  
Serum Proteins ◽  
Psychiatric Patients ◽  
Double Diffusion ◽  
Brain Extract ◽  
Monkey Brain ◽  
Functional Psychoses ◽  
Schizophrenic Patients ◽  
Altered Serum

Altered serum proteins in mental illness have been reported by workers in various parts of the world, and a review of these studies has led to the consideration of an autoimmune mechanism in the pathogenesis of some functional psychoses (Fessel, 1962a). Several reports have appeared in recent years which suggest the presence of antibodies in the serum of certain psychiatric patients to central nervous system tissue. Many of these have been reviewed by Vartanyan (1963). Fessel (1962b, 1963) demonstrated agglutination of latex particles coated with monkey brain extract and agglutination of tanned sheep red blood cells coated with human brain extract, but no precipitins in double diffusion in agar of the sera against monkey brain extract. As an alternative to antibrain antibodies he suggested a less specific physicochemical abnormality of the serum which caused the agglutination. Yokoyama, Trams, and Brady (1962), using a sheep red blood cell haemagglutination technique, showed the presence of anti-asialoganglioside antibodies in the sera of 3 of 14 schizophrenic patients and anti-ganglioside antibody in the serum of another. Kuznetzova and Semenov (1961), by complement fixation, demonstrated antibodies in the sera of 22 of 84 schizophrenics, mainly to human brain and not to other organs. The antibodies appeared more frequently in the later stages of the illness (Semenov, Morozov, and Kuznetzova, 1961). Skalickova and Jezkova (1961), also with a complement fixation technique, demonstrated blood and cerebrospinal fluid antibodies to grey and white matter during the “infectious” onset of schizophrenia, but not in the chronic, demented phase.

scholarly journals Discrimination of prostate cancer cells and non-malignant cells using secondary ion mass spectrometry

The Analyst ◽  
2008 ◽  
Vol 133 (2) ◽  
pp. 175-179 ◽  
Author(s):  
Matthew J. Baker ◽  
Michael D. Brown ◽  
Ehsan Gazi ◽  
Noel W. Clarke ◽  
John C. Vickerman ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Mass Spectrometry ◽  
Cancer Cells ◽  
Secondary Ion Mass Spectrometry ◽  
Malignant Cells ◽  
Prostate Cancer Cells ◽  
Ion Mass Spectrometry ◽  
Secondary Ion

scholarly journals An immunohistochemical study of lymphatic elements in the human brain

2021 ◽  
Vol 118 (3) ◽  
pp. e2002574118
Author(s):  
Éva Mezey ◽  
Ildikó Szalayova ◽  
Christopher T. Hogden ◽  
Alexandra Brady ◽  
Ágnes Dósa ◽  
...  
Keyword(s):  
Human Brain ◽  
Dura Mater ◽  
Cranial Nerves ◽  
Lamina Cribrosa ◽  
Brain Parenchyma ◽  
Pia Mater ◽  
Postmortem Human Brain ◽  
Waste Products ◽  
T Cell Migration ◽  
The Brain

Almost 150 papers about brain lymphatics have been published in the last 150 years. Recently, the information in these papers has been synthesized into a picture of central nervous system (CNS) “glymphatics,” but the fine structure of lymphatic elements in the human brain based on imaging specific markers of lymphatic endothelium has not been described. We used LYVE1 and PDPN antibodies to visualize lymphatic marker-positive cells (LMPCs) in postmortem human brain samples, meninges, cavernous sinus (cavum trigeminale), and cranial nerves and bolstered our findings with a VEGFR3 antibody. LMPCs were present in the perivascular space, the walls of small and large arteries and veins, the media of large vessels along smooth muscle cell membranes, and the vascular adventitia. Lymphatic marker staining was detected in the pia mater, in the arachnoid, in venous sinuses, and among the layers of the dura mater. There were many LMPCs in the perineurium and endoneurium of cranial nerves. Soluble waste may move from the brain parenchyma via perivascular and paravascular routes to the closest subarachnoid space and then travel along the dura mater and/or cranial nerves. Particulate waste products travel along the laminae of the dura mater toward the jugular fossa, lamina cribrosa, and perineurium of the cranial nerves to enter the cervical lymphatics. CD3-positive T cells appear to be in close proximity to LMPCs in perivascular/perineural spaces throughout the brain. Both immunostaining and qPCR confirmed the presence of adhesion molecules in the CNS known to be involved in T cell migration.

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