Long intervening non-coding RNA 00320 is human brain-specific and highly expressed in the cortical white matter.

Pervasive transcription of the genome produces a diverse array of functional non-coding RNAs (ncRNAs). One particular class of ncRNAs, long intervening non-coding RNAs (lincRNAs) are thought to play a role in regulating gene expression and may be a major contributor to organism and tissue complexity. The human brain with its heterogeneous cellular make-up is a rich source of lincRNAs; however, the functions of the majority of lincRNAs are unknown. Recently, by completing RNA sequencing (RNA-Seq) of the human frontal cortex, we identified linc00320 as being highly expressed in the white matter compared to grey matter in multiple system atrophy (MSA) brain. Here, we further investigate the expression patterns of linc00320 and conclude that it is involved in specific brain regions rather than having involvement in the MSA disease process. We also show that the full-length linc00320 is only expressed in human brain tissue and not in other primates, suggesting that it may be involved in improved functional connectivity for higher human brain cognition.

Landscape of chromosomal copy number aberrations in gangliogliomas and dysembryoplastic neuroepithelial tumours.

AIM: Gangliogliomas (GGs) and dysembryoplastic neuroepithelial tumours (DNTs) represent the most common histological entities within the spectrum of glioneuronal tumours (GNTs). The wide variability of morphological features complicates histological classification, including discrimination from prognostically distinct diffuse low-grade astrocytomas (AIIs). This study was performed to increase our understanding of these tumours. METHODS: We studied chromosomal copy number aberrations (CNAs) by genome-wide sequencing in a large cohort of GNTs and linked these to comprehensive histological analysis and clinical characteristics. One hundred fourteen GNTs were studied: 50 GGs and 64 DNTs. Also, a data set of CNAs from 38 diffuse AIIs was included. RESULTS: The most frequent CNAs in both GGs and DNTs were gains at chromosomes 5 and 7, often concurrent, and gain at chromosome 6. None of the CNAs was linked to histological subtype, immunohistochemical features or to clinical characteristics. Comparison of AIIs and diffuse GNTs revealed that gain at whole chromosome 5 is only observed in GNTs. CNA patterns indicative of chromothripsis were detected in three GNTs. CONCLUSION: We conclude that GNTs with diverse morphologies share molecular features, and our findings support the need to improve classification and differential diagnosis of tumour entities within the spectrum of GNTs, as well as their distinction from other gliomas.

Differential expression of major histocompatibility complex class I in developmental glioneuronal lesions.

PURPOSE: The expression of the major histocompatibility complex class I (MHC-I) in the brain has received considerable interest not only because of its fundamental role in the immune system, but also for its non-immune functions in the context of activity-dependent brain development and plasticity. METHODS: In the present study we evaluated the expression and cellular pattern of MHC-I in focal glioneuronal lesions associated with intractable epilepsy. MHC-I expression was studied in epilepsy surgery cases with focal cortical dysplasia (FCD I, n = 6; FCD IIa, n = 6 and FCD IIb, n = 15), tuberous sclerosis complex (TSC, cortical tubers; n = 6) or ganglioglioma (GG; n = 15) using immunocytochemistry. Evaluation of T lymphocytes with granzyme-B+ granules and albumin immunoreactivity was also performed. RESULTS: All lesions were characterized by MHC-I expression in blood vessels. Expression in both endothelial and microglial cells as well as in neurons (dysmorphic/dysplastic neurons) was observed in FCD II, TSC and GG cases. We observed perivascular and parenchymal T lymphocytes (CD8+, T-cytotoxic) with granzyme-B+ granules in FCD IIb and TSC specimens. Albumin extravasation, with uptake in astrocytes, was observed in FCD IIb and GG cases. CONCLUSIONS: Our findings indicate a prominent upregulation of MHC-I as part of the immune response occurring in epileptogenic glioneuronal lesions. In particular, the induction of MHC-I in neuronal cells appears to be a feature of type II FCD, TSC and GG and may represent an important accompanying event of the immune response, associated with blood-brain barrier dysfunction, in these developmental lesions.

MicroRNA519d and microRNA4758 can identify gangliogliomas from dysembryoplastic neuroepithelial tumours and astrocytomas.

Glioneuronal tumours, including gangliogliomas and dysembryoplastic neuroepithelial tumours, represent the most common low-grade epilepsy-associated brain tumours and are a well-recognized cause of intractable focal epilepsy in children and young adults. Classification is predominantly based on histological features, which is difficult due to the broad histological spectrum of these tumours. The aim of the present study was to find molecular markers that can be used to identify entities within the histopathology spectrum of glioneuronal tumours. The focus of this study was on microRNAs (miRNAs). miRNAs are important post-transcriptional regulators of gene expression and are involved in the pathogenesis of different neurological diseases and oncogenesis. Using a miRNA array, miR-519d and miR-4758 were found to be upregulated in gangliogliomas (n=26) compared to control cortex (n=17), peritumoural tissue (n=7), dysembryoplastic neuroepithelial tumours (n=9) and astrocytomas (grade I-IV; subependymal giant cell astrocytomas, n=10; pilocytic astrocytoma, n=15; diffuse astrocytoma grade II, n=10; grade III, n=14 and glioblastoma n=15). Furthermore, the PI3K/AKT3/P21 pathway, which is predicated to be targeted by miR-519d and miR-4758, was deregulated in gangliogliomas. Functionally, overexpression of miR-519d in an astrocytic cell line resulted in a downregulation of CDKN1A (P21) and an increase in cell proliferation, whereas co-transfection with miR-4758 counteracted this effect. These results suggest that miR-519d and miR-4758 might work in concert as regulators of the cell cycle in low grade gliomas. Furthermore, these miRNAs could be used to distinguish gangliogliomas from dysembryoplastic neuroepithelial tumours and other low and high grade gliomas and may lead to more targeted therapy.

Impaired oligodendroglial turnover is associated with myelin pathology in focal cortical dysplasia and tuberous sclerosis complex.

Conventional antiepileptic drugs suppress the excessive firing of neurons during seizures. In drug-resistant patients, treatment failure indicates an alternative important epileptogenic trigger. Two epilepsy-associated pathologies show myelin deficiencies in seizure-related brain regions: Focal Cortical Dysplasia IIB (FCD) and cortical tubers in Tuberous Sclerosis Complex (TSC). Studies uncovering white matter-pathology mechanisms are therefore urgently needed to gain more insight into epileptogenesis, the propensity to maintain seizures, and their associated comorbidities such as cognitive defects. We analyzed epilepsy surgery specimens of FCD IIB (n = 22), TSC (n = 8), and other malformations of cortical development MCD (n = 12), and compared them to autopsy and biopsy cases (n = 15). The entire lesional pathology was assessed using digital immunohistochemistry, immunofluorescence and western blotting for oligodendroglial lineage, myelin and mTOR markers, and findings were correlated to clinical parameters. White matter pathology with depleted myelin and oligodendroglia were found in 50% of FCD IIB and 62% of TSC cases. Other MCDs had either a normal content or even showed reactive oligodendrolial hyperplasia. Furthermore, myelin deficiency was associated with increased mTOR expression and the lower amount of oligodendroglia was linked with their precursor cells (PDGFRa). The relative duration of epilepsy (normalized to age) also correlated positively to mTOR activation and negatively to myelination. Decreased content of oligodendroglia and missing precursor cells indicated insufficient oligodendroglial development, probably mediated by mTOR, which may ultimately lead to severe myelin loss. In terms of disease management, an early and targeted treatment could restore normal myelin development and, therefore, alter seizure threshold and improve cognitive outcome.

BRAF V600E mutation is associated with mTOR signaling activation in glioneuronal tumors.

BRAF V600E mutations have been recently reported in glioneuronal tumors (GNTs). To evaluate the expression of the BRAF V600E mutated protein and its association with activation of the mammalian target of rapamycin (mTOR) pathway, immunophenotype and clinical characteristics in GNTs, we investigated a cohort of 174 GNTs. The presence of BRAF V600E mutations was detected by direct DNA sequencing and BRAF V600E immunohistochemical detection. Expression of BRAF-mutated protein was detected in 38/93 (40.8%) gangliogliomas (GGs), 2/4 (50%) desmoplastic infantile gangliogliomas (DIGs) and 23/77 (29.8%) dysembryoplastic neuroepithelial tumors (DNTs) by immunohistochemistry. In both GGs and DNTs, the presence of BRAF V600E mutation was significantly associated with the expression of CD34, phosphorylated ribosomal S6 protein (pS6; marker of mTOR pathway activation) in dysplastic neurons and synaptophysin (P < 0.05). In GGs, the presence of lymphocytic cuffs was more frequent in BRAF-mutated cases (31 vs. 15.8%; P=0.001). The expression of both BRAF V600E and pS6 was associated with a worse postoperative seizure outcome in GNT (P < 0.001). Immunohistochemical detection of BRAF V600E-mutated protein may be valuable in the diagnostic evaluation of these glioneuronal lesions and the observed association with mTOR activation may aid in the development of targeted treatment involving specific pathogenic pathways.

Fetal brain lesions in tuberous sclerosis complex: TORC1 activation and inflammation.

Tuberous sclerosis complex (TSC) is an autosomal dominant disorder caused by mutations in either the TSC1 or TSC2 genes and characterized by developmental brain abnormalities. We defined the spectrum of brain abnormalities in fetal TSC brain ranging from 23 to 38 gestational weeks. We hypothesized (i) prenatal activation of the target-of-rapamycin complex 1 (TORC1) signaling pathway; and (ii) activation of inflammatory pathways in fetal brain lesions. Immunocytochemical analysis of cortical tubers, as well as subependymal lesions in all cases confirmed the cell-associated activation of the TORC1 signaling pathway in both the cortical tubers and subependymal lesions (including a congenital subependymal giant cell astrocytoma) with expression of pS6, p4EBP1 and c-myc proteins, as well as of p70 S6 kinase 1. The lesions contained macrophages and T-lymphocytes; giant cells within the lesions expressed inflammatory response markers including major histocompatibility complex class I and II, Toll-like receptors (TLR) 2 and 4 and receptor for advanced glycation end products (RAGE). These observations indicate that brain malformations in TSC are likely a consequence of increased TORC1 activation during embryonic brain development. We also provide evidence supporting the possible immunogenicity of giant cells and the early activation of inflammatory pathways in TSC brain.

Role of p53 in irinotecan-induced intestinal cell death and mucosal damage.

Irinotecan treatment of colorectal cancers results in high-grade intestinal mucositis in a large proportion of patients. The mechanisms behind irinotecan-induced mucosal injury, however, have yet to be fully explained. The aim of this study was to investigate the role of the p53 protein in the onset of intestinal damage following irinotecan treatment in two different settings. IEC-6 and FHs 74 intestinal cell lines were treated with irinotecan with and without a temporary p53 inhibitor, pifithrin-alpha, and examined for changes in proliferation and survival along with expression of p53 and related proteins. Forty tumour-bearing rats also underwent irinotecan treatment with and without pifithrin-alpha, and the effects on intestinal morphology, gene expression, apoptosis and other toxicities were assessed. Irinotecan caused a dose-dependent reduction in cell viability that was not prevented by pifithrin-alpha in either cell line. Rats responded to irinotecan with diarrhoea, weight loss, histopathological changes to the small and large intestine, increased crypt apoptosis, and a mild inflammatory response. Pifithrin-alpha reduced severity and duration of intestinal apoptosis; however, it did not significantly affect other parameters including p53 expression. Temporary inhibition of p53 activation does not markedly prevent intestinal cell death or mucositis following irinotecan treatment. Irinotecan may act through upregulation of proapoptotic proteins Bax and Bak to induce cell death.