Clinical Prognostic Implications of Wnt Hub Genes Expression in Medulloblastoma.

Medulloblastoma is the most common type of pediatric malignant primary brain tumor, and about one-third of patients die due to disease recurrence and most survivors suffer from long-term side effects. MB is clinically, genetically, and epigenetically heterogeneous and subdivided into at least four molecular subgroups: WNT, SHH, Group 3, and Group 4. We evaluated common differentially expressed genes between a Brazilian RNA-seq GSE181293 dataset and microarray GSE85217 dataset cohort of pediatric MB samples using bioinformatics methodology in order to identify hub genes of the molecular subgroups based on PPI network construction, survival and functional analysis. The main finding was the identification of five hub genes from the WNT subgroup that are tumor suppressors, and whose lower expression is related to a worse prognosis for MB patients. Furthermore, the common genes correlated with the five tumor suppressors participate in important pathways and processes for tumor initiation and progression, as well as development and differentiation, and some of them control cell stemness and pluripotency. These genes have not yet been studied within the context of MB, representing new important elements for investigation in the search for therapeutic targets, prognostic markers or for understanding of MB biology.

Expression of microRNAs miR-21 and miR-326 associated with HIF-1α regulation in neurospheres of glioblastoma submitted to ionizing radiation treatment.

Background: Glioblastoma is an incurable neoplasm. Its hypoxia mechanism associated with cancer stem cells (CSCs) demonstrates hypoxia-inducible factor 1α (HIF-1α) expression regulation, which is directly related to tumor malignancy. The aim of this study was to identify a possible tumor malignancy signature associated with regulation of HIF-1α by microRNAs miR-21 and miR-326 in the subpopulation of tumor stem cells which were irradiated by ion in primary culture of patients diagnosed with glioblastoma. Materials and methods: We used cellular cultures from surgery biopsies of ten patients with glioblastoma. MicroRNA expressions were analyzed through real-time polymerase chain reaction (PCR ) and correlated with mortality and recurrence. The ROC curve displayed the cutoff point of the respective microRNAs in relation to the clinical prognosis, separating them by group. Results: The miR-21 addressed high level of expression in the irradiated neurosphere group (p = 0.0028). However, miR-21 was not associated with recurrence and mortality. miR-326 can be associated with tumoral recurrence (p = 0.032) in both groups; every 0.5 units of miR-326 increased the chances of recurrence by 1,024 (2.4%). Conclusion: The high expression of miR-21 in the irradiated group suggests its role in the regulation of HIF-1α and in the radioresistant neurospheres. miR-326 increased the chances of recurrence in both groups, also demonstrating that positive regulation from miR-326 does not depend on ionizing radiation treatment.

Expression of pluripotency-related genes in human glioblastoma.

Background: Cancer is a group of heterogeneous diseases characterized by several disruptions of the genetic and epigenetic components of cell biology. Some types of cancer have been shown to be constituted by a mosaic of cells with variable differentiation states, with more aggressive tumors being more undifferentiated. In most cases, undifferentiated tumor cells express associated embryonic markers such as the OCT4, NANOG, SOX2, and CARM1 genes. The ectopic or reminiscent expression of some master regulator genes of pluripotency has been indicated as the cause of the poorly differentiated state of tumors, and based on the evidence of some reports, can be used as a possible therapeutic target. Considering this information, a more detailed investigation of the expression of pluripotency-associated genes is necessary to evaluate the roles of these genes in the etiology of some tumors and their use targets of therapy. Methods: The expression of four pluripotency-related genes was investigated (OCT4, NANOG, SOX2, and CARM1) in the most malignant primary human brain tumor, glioblastoma (GBM). Results and Conclusion: The results demonstrated a signature of OCT4/SOX2/CARM1 genes and a significant increase of CARM1 expression in GBM cases.

Decreased hippocampal serotonin 5HT1A expression in mesial temporal lobe of epilepsy patients.

INTRODUCTION: Mesial temporal lobe epilepsy related to hippocampal sclerosis (MTLE-HS) is a surgically remediable epilepsy with a relatively high prevalence and psychiatric comorbidities. Depressive disorders may occur in up to 25% of MTLE-HS patients suggesting a common molecular mechanism underlying both conditions. OBJECTIVE: To compare the gene expression comprising serotonin 5HT1A and 5HT2A, noradrenaline (NA) ADRA1A, and ADRA2A receptors in the hippocampus of MTLE-HS patients with and without major depression. METHODS: A cross-sectional study allocated 31 patients in three groups: MTLE-HS without psychiatric diagnosis (MTLE-HS group), MTLE-HS with major depression (MTLE-HS-D group) and a control group consisting of healthy volunteers without any neurological or psychiatric disorders. Demographic and clinical characteristics were compared among groups. Gene expression of receptors were analyzed using general linear mixed models (GLMM), with an unstructured matrix, normal link. RESULTS: The three groups showed a similar distribution regarding age, gender (p > 0.16), history of initial precipitating injury, family history of epilepsy, monthly frequency of seizures, side of hippocampal sclerosis, interictal spike distribution and anti-seizure medications did not differ between MTLE-HS and MTLE-HS-D groups (p > 0.05). We observed a greater expression of the 5HT1A receptor in the control group when compared to the MTLE-HS (P = .004) and MTLE-HS-D (P = .007). Nevertheless, we did not observe any difference when MTLE-HS and MTLE-HS-D groups were compared to the controls for the ADRA1A (P = .931; P = .931), ADRA2A (P = .120; P = .121) and 5HT2A (P = .638; P = .318, respectively) gene expression. CONCLUSION: Mesial temporal lobe epilepsy related to hippocampal sclerosis and MTLE-HS-D patients showed a lowered expression of the 5HT1A receptors when compared with the controls adjusted for age and schooling. Data suggest that temporal lobe epilepsy plasticity may affect serotonin receptors, which may lead to more frequent cases of major depression in this population. More studies comprising wider samples are necessary to confirm these results; they also should investigate serotonin reuptake drugs as an adjuvant therapeutic option for MTLE-HS disorder.

Modulation of Genes and MicroRNAs in the Neurospheres of Glioblastoma Cell Lines U343 and T98G Induced by Ionizing Radiation and Temozolomide Therapy.

INTRODUCTION: Glioblastoma is the most prevalent primary malignant neoplasm of the central nervous system. It has increased its incidence, while the overall survival remains over 14 months. PURPOSE: The purpose is to evaluate the expression of the genes EGFR, PTEN, MGMT, and IDH1/2, and microRNAs miR-181b, miR-145, miR-149, and miR-128a in adhered cells (AC) and neurospheres (NS) from cell lines (T98G and U343) submitted to temozolomide (TMZ) and ionizing radiation (IR). METHODS: T98G and U343 were treated with TMZ, IR, and TMZ+IR. The analysis of gene expression and miRNAs was performed using real-time PCR. RESULTS: This study demonstrated: a) an improvement in the expression of IDH1 after IR and TMZ + IR in the NS (T98G); b) an increase in the expression of MGMT in NS (T98G) in IR groups and TMZ + IR. The expression of miRNAs results as a) AC (U343) expressed more miR-181b after TMZ, IR, and TMZ + IR; and miR-128a improved after TMZ, IR, and TMZ + IR; b) NS (T98G) after TMZ + IR expressed: miR-181b; miR-149; miR-145 and miR-128a; c) NS (U343) after IR huge expressed miR-149 and miR-145. CONCLUSION: IR was an independent and determining radioresistance factor in NS. However, we observed no complementarity action of oncomiRs regulation.

The Role of MicroRNA 181d as a Possible Biomarker Associated With Tumor Progression in Meningiomas.

Introduction Meningiomas are slow-growing intracranial neoplasms that originate from arachnoid meningothelial cells and represent 13-26% of intracranial tumors, thus being the most common. There are numerous technological advances available for a better understanding of the molecular pathways correlated with tumorigenesis and tumor progression of meningiomas. In this context, the role of microRNAs (miRNAs), which are non-coding RNAs (ncRNAs) consisting of 18 to 25 nucleotides whose function is the silencing of mRNA at the posttranscriptional level, has been highlighted. Recent studies suggest that miRNAs may act as possible biomarkers as well as therapeutic targets for various diseases, including brain tumors. Therefore, the objective of our study was to evaluate the tissue and plasma expression of the miRNAs miR-181d, miR-181c, and miR-130a. Methods The miRNAs miR-181d, miR-181c, and miR-130a were selected from our group's prior study by the large-scale microarray analysis technique. In this work, the expression of these miRNAs in the tumor tissue and plasma of patients with grade I (16 patients), II (16 patients), and III (eight patients) meningiomas was evaluated. Results MiR-181d was overexpressed in both tumor tissue and plasma in the studied groups. The level of expression was higher according to the progression of tumor grade. MiR-181c and miR-130a showed no significant difference in the studied groups in either tumor tissue or plasma. Conclusions MiR-181d has potential as a biomarker for meningiomas and is associated with the tumor progression of meningiomas.

Epidemiological features of meningiomas: a single Brazilian center's experience with 993 cases.

BACKGROUND: Meningiomas are the most frequent primary central nervous system (CNS) tumors. Their geographical and ethnic characteristics need to be known, in order to enable rational treatment. OBJECTIVE: To investigate clinical and epidemiological aspects in a series of patients with meningiomas. METHODS: Retrospective analysis on the demographic profile, location and histopathology of 993 patients with meningiomas (768 operated and 225 not operated). RESULTS: Meningiomas represented 43.8% of the primary CNS tumors; 6.8% were multiple tumors (14.7% with neurofibromatosis 2) and 0.6% were radiation-induced tumors. The mean ages were 53.0 and 63.9 years for operated and non-operated patients and the female/male ratios were 3.2:1 and 6.3:1. Diagnosis was made later among females. The peak incidences were in the 6th and 7th decades respectively for operated and non-operated patients. The incidence was low at early ages and higher among patients aged 70+ years. The meningiomas were intracranial in 96.5% and most were WHO grade I (88.9%) and transitional. In the spinal canal (3.5%), they occurred mainly in the dorsal region (all grade I; mostly transitional). The racial distribution was 1.0% in Asian-Brazilians, 87% in Caucasians and 12% in African-Brazilians. 83.4% and 51.6% of the patients were estimated to be recurrence-free at 10 and 20 years, and the mortality rate was 3%. CONCLUSIONS: Most of the demographic data were similar to what has been observed in other western centers. Differences were higher incidence of meningiomas, female and older predominance in non-operated patients, predominance in Caucasian, and higher association with neurofibromatosis 2.

High-throughput microRNA profile in adult and pediatric primary glioblastomas: the role of miR-10b-5p and miR-630 in the tumor aggressiveness.

Glioblastoma (GBM) is the most common primary malignant neoplasm of the central nervous system and, despite the standard therapy; the patients' prognoses remain dismal. The miRNA expression profiles have been associated with patient prognosis, suggesting that they may be helpful for tumor diagnosis and classification as well as predictive of tumor response to treatment. We described the microRNA expression profile of 29 primary GBM samples (9 pediatric GBMs) and 11 non-neoplastic white matter samples as controls (WM) by microarray analysis and we performed functional in vitro assays on these 2 most differentially expressed miRNAs. Hierarchical clustering analysis showed 3 distinct miRNA profiles, two of them in the GBM samples and a group consisting only of cerebral white matter. When adult and pediatric GBMs were compared to WM, 37 human miRNAs were found to be differentially expressed, with miR-10b-5p being the most overexpressed and miR-630 the most underexpressed. The overexpression of miR-630 was associated with reduced cell proliferation and invasion in the U87 GBM cell line, whereas the inhibition of miR-10b-5p reduced cell proliferation and colony formation in the U251 GBM cell line, suggesting that these miRNAs may act as tumor-suppressive and oncogenic miRNAs, respectively. The present study highlights the distinct epigenetic profiling of adult and pediatric GBMs and underscores the biological importance of mir-10b-5p and miR-630 for the pathobiology of these lethal tumors.

Expression of circulating microRNAs as predictors of diagnosis and surgical outcome in patients with mesial temporal lobe epilepsy with hippocampal sclerosis.

MicroRNAs have been progressively investigated as post-transcriptional regulators playing important roles in epilepsy pathophysiology. Here we investigate three promising microRNAs (miR-27a-3p, miR-328-3p and miR-654-3p) previously described in the literature as possible peripheral biomarkers for epilepsy diagnose and surgical prognosis. Serum samples from 28 patients with mesial temporal lobe epilepsy with hippocampal sclerosis (MTLE-HS) were analyzed, 14 with good surgical prognosis (Engel I) and 14 with unfavorable surgical prognosis (Engel III-IV). Serum samples from 11 healthy volunteers were the control group. The microRNAs expression analysis was performed using real-time PCR. The present results did not endorse the role of miR-27a-3p as a peripheral biomarker for epilepsy diagnosis or surgical prognosis. MiR-328-3p, however, presented significant area under the curve (AUC) values when comparing controls to Engel I (90.3%), controls to Engel III-IV (96.8%) and controls to Engel I + Engel III-IV (i.e., epilepsy patients, AUC = 93.5%). Additionally, miR-654-3p displayed AUC = 74.7% when comparing controls to Engel I patients (p = 0.004), and AUC = 73.6% (p = 0.04) in the attempt to discriminate unfavorable from favorable surgical prognosis. In conclusion, the ANOVA and ROC analyzes with the respective AUC, specificity and sensitivity values allows us to conclude that miR-328-3p is the most important peripheral biomarker for the diagnosis of MTLE-HS. In terms of predicting the surgical prognosis of MTLE-HS patients, miR-654-3p proved to be the only microRNA evaluated to present statistical power to differentiate, as a peripheral biomarker, Engel I from Engel III-IV patients.

Drebrin expression patterns in patients with refractory temporal lobe epilepsy and hippocampal sclerosis.

OBJECTIVE: Drebrins are crucial for synaptic function and dendritic spine development, remodeling, and maintenance. In temporal lobe epilepsy (TLE) patients, a significant hippocampal synaptic reorganization occurs, and synaptic reorganization has been associated with hippocampal hyperexcitability. This study aimed to evaluate, in TLE patients, the hippocampal expression of drebrin using immunohistochemistry with DAS2 or M2F6 antibodies that recognize adult (drebrin A) or adult and embryonic (pan-drebrin) isoforms, respectively. METHODS: Hippocampal sections from drug-resistant TLE patients with hippocampal sclerosis (HS; TLE, n = 33), of whom 31 presented with type 1 HS and two with type 2 HS, and autopsy control cases (n = 20) were assayed by immunohistochemistry and evaluated for neuron density, and drebrin A and pan-drebrin expression. Double-labeling immunofluorescences were performed to localize drebrin A-positive spines in dendrites (MAP2), and to evaluate whether drebrin colocalizes with inhibitory (GAD65) and excitatory (VGlut1) presynaptic markers. RESULTS: Compared to controls, TLE patients had increased pan-drebrin in all hippocampal subfields and increased drebrin A-immunopositive area in all hippocampal subfields but CA1. Drebrin-positive spine density followed the same pattern as total drebrin quantification. Confocal microscopy indicated juxtaposition of drebrin-positive spines with VGlut1-positive puncta, but not with GAD65-positive puncta. Drebrin expression in the dentate gyrus of TLE cases was associated negatively with seizure frequency and positively with verbal memory. TLE patients with lower drebrin-immunopositive area in inner molecular layer (IML) than in outer molecular layer (OML) had a lower seizure frequency than those with higher or comparable drebrin-immunopositive area in IML compared with OML. SIGNIFICANCE: Our results suggest that changes in drebrin-positive spines and drebrin expression in the dentate gyrus of TLE patients are associated with lower seizure frequency, more preserved verbal memory, and a better postsurgical outcome.

Modulation of NMDA receptor by miR-219 in the amygdala and hippocampus of patients with mesial temporal lobe epilepsy.

Mesial temporal lobe epilepsy with hippocampal sclerosis is the most frequent form of focal epilepsy in adults, and it is often refractory to drug treatment. Regardless of the efforts on developing new antiepileptic drugs for refractory cases, studies suggest a need for better understanding the molecular bases of epilepsy. The microRNAs have been progressively investigated as potential targets for both epilepsy mechanisms elucidation and treatment. Therefore, the goal of this study was to evaluate the differential expression of miR-219, miR-181b, and miR-195, previously described as regulators of the excitatory neurotransmitter receptors NMDA-R1 and AMPA-GluR2 and inhibitory neurotransmitter GABAA (α2, ß3, and γ2 subunits) in the amygdala and hippocampus of patients with mesial temporal lobe epilepsy. Based on genes and miRNAs' quantitative Polymerase Chain Reaction (qPCR) from 18 patients with epilepsy, our results showed an inverse relationship between miR-219 and NMDA-NR1 expression in both the amygdala and hippocampus in comparison to their expression in controls. NR1 and GluR2 were upregulated in the amygdala of epileptic patients. Low miR-195 expression was observed in the amygdala of patients with epilepsy. Our findings indicate that miR-219 has a possible regulatory role in excitatory neurotransmission in patients with epilepsy, contributing to the new avenue of miRNA biology in drug-resistant epilepsy, reserving huge potential for future applications and clinical interventions in conjunction with existing therapies.

Histological correlates of hippocampal magnetization transfer images in drug-resistant temporal lobe epilepsy patients.

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 < 0.05. RESULTS: Compared to the respective controls, TLE patients had reduced hippocampal MTR, increased reactive astrocytes and activated microglia, increased extracellular chondroitin sulfate, and reduced neuron density, compares to controls. MTR correlated positively with neuron density in CA3 and with chondroitin sulfate in CA3 and CA1. Multiple linear regressions reinforced the correlations between chondroitin sulfate and MTR. SIGNIFICANCE: Our data indicate that extracellular matrix molecules are the most significant histological correlates of magnetization transfer ratio in the hippocampus of TLE patients.

Analysis of Caspase-9 protein and microRNAs miR-21, miR-126 and miR-155 related to the apoptosis mechanism in the cerebellum of rats submitted to focal cerebral ischemia associated with an alcoholism model.

OBJECTIVE: This study aimed to analyze the cerebellum of rats submitted to an experimental focal cerebral ischemia, by middle cerebral artery occlusion for 90 minutes, followed by reperfusion for 48 hours, associated with an alcoholism model. METHODS: Fifty adult Wistar rats were used, subdivided into five experimental groups: control group (C): animals submitted to anesthesia only; sham group (S): animals submitted to complete simulation of the surgical procedure; ischemic group (I): animals submitted to focal cerebral ischemia for 90 minutes followed by reperfusion for 48 hours; alcoholic group (A): animals that received daily absolute ethanol diluted 20% in water for four weeks; and, ischemic and alcoholic group (I + A): animals receiving the same treatment as group A and, after four weeks, submitted to focal cerebral ischemia for 90 minutes, followed by reperfusion for 48 hours. The cerebellum samples were collected and immunohistochemical analysis of Caspase-9 protein and serum analysis by RT-PCR of microRNAs miR-21, miR-126 and miR155 were performed. RESULTS: The expression of Caspase-9 was higher in groups I, A and I + A. In the microRNAs analyses, miR-126 was higher in groups A and I + A, miR-155 was higher in groups I and I + A. CONCLUSIONS: We conclude that apoptosis occurs in the cerebellar cortex, even if it is distant from the ischemic focus, and that microRNAs 126 and 155 show a correlation with cellular apoptosis in ischemic rats and those submitted to the chronic alcohol model.

Expression of MicroRNAs miR-145, miR-181c, miR-199a and miR-1183 in the Blood and Hippocampus of Patients with Mesial Temporal Lobe Epilepsy.

The aim of this study was to analyze the expression profiles of the microRNAs (miRNAs) miR-145, miR-181c, miR-199a and miR-1183 in the hippocampus and blood of patients with mesial temporal lobe epilepsy with hippocampal sclerosis (MTLE-HS) and to investigate whether these can be used as diagnosis and prognosis biomarkers for epilepsy. Hippocampus and blood samples were collected from 20 patients with MTLE-HS, ten of whom had a favorable surgical outcome (Engel I) and ten with an unfavorable surgical outcome (Engel III-IV). Hippocampus samples from autopsied individuals with no neurological or psychiatric medical history (necropsy samples) and blood samples from healthy individuals were used as controls. Real-time quantitative PCR (RQ-PCR) was used to analyze miRNA expression. The results showed that the expressions of these miRNAs differed quantitatively in the hippocampus and blood of patients with MTLE-HS in comparison to the respective control. This difference was most pronounced for miR-145, which was hypo-expressed in the hippocampus and hyper-expressed in the blood of MTLE-HS patients. MiRNAs miR-145, miR-181c, miR-199a and miR-1183 were hyper-expressed in the blood of patients with MTLE-HS. No statistical differences in the levels of these miRNAs in the blood or hippocampus were found between Engel I patients and Engel III-IV patients. These results suggest that the analyzed microRNAs are potential circulating biomarkers for epilepsy diagnosis.

microRNA-181d associated with the methylation status of the MGMT gene in Glioblastoma multiforme cancer stem cells submitted to treatments with ionizing radiation and temozolomide.

Despite the increased understanding of the oncological mechanisms underlying Glioblastoma multiforme (GBM) pathophysiology, and recent advances in therapeutic strategies such as maximal surgical resection and post-operative radiotherapy with concomitant and adjuvant temozolomide chemotherapy, the prognosis for patients with brain tumors remains limited. Evidences indicate that the assessment of DNA methylation status in cancer stem cells would allow identifying molecules expressed in these cells, to lead to targeted elimination of this critical population from brain tumors, making the glioblastoma treatment more effective. This study aimed to analyze the role of microRNA-181d associated with the methylation status of the O6-methylguanine methyl transferase (MGMT) gene in Glioblastoma multiforme cancer stem cells subjected to treatment with temozolomide and ionizing radiation. Such responses were analyzed in terms of cell survival, evaluation of the MGMT gene methylation status by MS-HRM (Methylation-Sensitive High Resolution Melting), and analysis of miRNA-181d and MGMT gene expression by relative quantification of mRNA levels in cancer stem cells subjected to treatment with temozolomide and ionizing radiation, isolated or combined. We showed that ionizing radiation and temozolomide reduced the viability of cancer stem cells from GBM patients, as well as modified MGMT gene and miRNA-181d expression in cancer stem cells, suggesting that miRNA-181d interferes in the glioblastoma cancer stem cell response to treatment with temozolomide and ionizing radiation.

Spontaneous subdural hematoma associated with microcystic meningioma: first case report in the literature.

Background: Non traumatic subdural hematomas are rare, especially those associated with intracranial meningiomas. Among the most common meningiomas associated with spontaneous bleeding are angioblastic and malignant meningioma variants. The pathophysiological mechanisms of this association are not yet fully understood. The association of chronic subdural hematoma with microcystic meningioma histological subtype has not yet been described in the literature. Case report: The authors present a case report of a patient with a spontaneous non traumatic chronic subdural hematoma associated with a microcystic subtype grade I meningioma of the parietal convexity. Epidemiological, etiology, natural history, pathophysiology, risk factors of bleeding and treatment options are reviewed. Conclusion: Spontaneous subdural hematomas associated with meningiomas are rare, specially related to the microcystic variant of meningioma. Careful pre-operative consideration of specific anatomy and pathophysiological features are paramount to their full treatment.

Manual Hippocampal Subfield Segmentation Using High-Field MRI: Impact of Different Subfields in Hippocampal Volume Loss of Temporal Lobe Epilepsy Patients.

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.

Morphological and immunohistochemical analysis of proteins CASPASE 3 and XIAP in rats subjected to cerebral ischemia and chronic alcoholism.

PURPOSE: To evaluate histopathological and ultrastructural changes and expression of proteins related to apoptosis CASPASE 3 and XIAP after experimental induction of temporary focal cerebral ischemia (90 minutes) due to obstruction of the middle cerebral artery in alcoholism model. METHODS: Forty adult Wistar rats were used, subdivided into 5 experimental groups: control group (C); Sham group (S); Ischemic group (I); Alcoholic group (A); and Ischemic and Alcoholized group (I+A): animals submitted to the same treatment of group A and after four weeks were submitted to focal cerebral ischemia during 90 minutes, followed by reperfusion of 48 hours. Were processed for histopathological analysis and immunohistochemistry (for the protein expression of CASPASE -3 and XIAP). RESULTS: Greater histopathological changes were observed in the animals of groups I and I+A in the three areas analyzed. The neuronal loss was higher in the medial striatum region of the animals of groups I and I + A. The protein expression of CASPASE -3 was higher than that of XIAP in the groups I and I + A for both proteins. CONCLUSION: The expression of XIAP was slightly higher where the histopathological changes and expression of CASPASE -3 was less evident.

HIF1A is Overexpressed in Medulloblastoma and its Inhibition Reduces Proliferation and Increases EPAS1 and ATG16L1 Methylation.

BACKGROUND: Genetic and epigenetic modifications are closely related to tumor initiation and progression and can provide guidance for understanding tumor functioning, potentially leading to the discovery of new therapies. Studies have associated hypoxia-related genes to tumor progression and chemo/radioresistance in brain tumors. Information on the expression profile of hypoxiarelated genes in pediatric medulloblastoma, although scarce, may reveal relevant information that could support treatment decisions. OBJECTIVE: Our study focused on evaluation the of CA9, CA12, HIF1A, EPAS1, SCL2A1 and VEGF genes in 41 pediatric fresh-frozen medulloblastoma sample. Additionally, we analyzed the effect of hypoxia and normoxia in the pediatric medulloblastoma cell-line UW402. Furthermore, we assessed the effects of HIF1A knockdown in cell-proliferation and methylation levels of genes related to hypoxia, apoptosis and autophagy. METHOD: qPCR was performed to evaluate mRNA levels, and Western blot to confirm HIF1A silencing in both patient samples and cell line. Pyrosequencing was performed to asses the methylation levels after HIF1A knockdown in the UW402 cell line. RESULTS: A higher HIF1A mRNA level was observed in MB patients when compared to the cerebellum (non-tumor match). In UW402 MB cell-line, chemically induced hypoxic resulted in an increase of mRNA levels of HIF1A, VEGF, SCL2A1 and CA9 genes. Additionally, HIF1A knockdown induced a decrease in the expression of hypoxia related genes and a decrease of 30% in cell proliferation was also observed. Also, a significant increase in the methylation of ATG16L1 promoter and decrease in the methylation of EPAS1 promoter were observed after HIF1A knockdown. CONCLUSION: HIF1A knockdown in medulloblastoma cells lead to decreased cellular proliferation, suggesting that HIF1A can be a potential therapeutic target to be explored in the medulloblastoma. However, the mechanisms behind HIF1A protein stabilization and function are very complex and more data need to be generated to potentially use HIF1A as a therapeutical target.

High expression of XIAP and Bcl-2 may inhibit programmed cell death in glioblastomas.

Glioblastoma (GBM) is the most malignant glioma and represents 29% of all brain tumors. Tumorigenesis is intimately connected with characteristics acquired in the physiologic pathway of cellular death. OBJECTIVE: In the present study, the expression of anti-apoptotic (XIAP and Bcl-2) and apoptotic (cytochrome C, caspase 9, APAF-1), caspase 3 and the Smac/DIABLO genes related to the apoptosis pathway were evaluated in 30 samples of glioblastoma. METHODS: The gene expression was evaluated in 30 glioblastomas (WHO grade IV) and compared to 10 white matter control samples with real-time PCR. RESULTS AND CONCLUSION: There were higher expressions of XIAP (p = 0.0032) and Bcl-2 (p = 0.0351) in the glioblastoma samples compared to the control samples of normal brain. These results raise the question of whether Bcl-2 and XIAP genes can be responsible for the inhibition of programmed cell death in glioblastomas. Moreover, they provide additional information capable of allowing the development of new target therapy strategies.