Serum biomarkers identification by iTRAQ and verification by MRM: S100A8/S100A9 levels predict tumor-stroma involvement and prognosis in Glioblastoma.

Despite advances in biology and treatment modalities, the prognosis of glioblastoma (GBM) remains poor. Serum reflects disease macroenvironment and thus provides a less invasive means to diagnose and monitor a diseased condition. By employing 4-plex iTRAQ methodology, we identified 40 proteins with differential abundance in GBM sera. The high abundance of serum S100A8/S100A9 was verified by multiple reaction monitoring (MRM). ELISA and MRM-based quantitation showed a significant positive correlation. Further, an integrated investigation using stromal, tumor purity and cell type scores demonstrated an enrichment of myeloid cell lineage in the GBM tumor microenvironment. Transcript levels of S100A8/S100A9 were found to be independent poor prognostic indicators in GBM. Medium levels of pre-operative and three-month post-operative follow-up serum S100A8 levels predicted poor prognosis in GBM patients who lived beyond median survival. In vitro experiments showed that recombinant S100A8/S100A9 proteins promoted integrin signalling dependent glioma cell migration and invasion up to a threshold level of concentrations. Thus, we have discovered GBM serum marker by iTRAQ and verified by MRM. We also demonstrate interplay between tumor micro and macroenvironment and identified S100A8 as a potential marker with diagnostic and prognostic value in GBM.

Rare manifestation of common disease with an unique method of minimally invasive spine stabilization: Cervical 2-3 facet lesion.

Cervical granulomatous infections of the posterior elements are very rare, it is often difficult to diagnose due to rarity and variable presentation of symptoms. Any cervical surgical procedure carries a certain morbid risk to the patient. We present a case of cervical 2-3 facet joint lesion which was managed by a minimally invasive technique with a favorable outcome.

Multichannel EEG based inter-ictal seizures detection using Teager energy with backpropagation neural network classifier.

A long-term multichannel electroencephalogram recording plays a crucial role in recognizing the epileptic seizure activities from the brain lobes. This research study investigates the automated detection of epileptic seizures from multichannel electroencephalogram recordings using Teager energy feature. A supervised back-propagation neural network model was implemented to classify the inter-ictal seizures. The study was conducted on multichannel electroencephalogram data that was obtained from Institute of Neuroscience, Ramaiah Memorial Hospital, Bengaluru, India, after ethical clearance from the from the Institutional Ethics Board. Initially, notch filter was applied to remove the 50 Hz power line noise from raw electroencephalogram followed by independent component analysis to remove eye blinks and muscular activities. A time domain feature called Teager energy was estimated which detects the rapid changes in the given electroencephalogram time series. A 1 s windowing was introduced to ensure stationarity for estimation of Teager energy. The descriptive and box plot analysis ensures the suitability of the Teager energy for the seizure detection. The performance of the multilayer perceptron neural network classifier was evaluated using sensitivity, specificity, and false detection rate. Simulation results showed the highest sensitivity, specificity and false detection rate of 96.66%, 99.15%, and 0.30 per hour respectively. It can be concluded that procedure can be applied for real-time seizure detection.

Automated epileptic seizures detection using multi-features and multilayer perceptron neural network.

Detection of epileptic seizure activities from long-term multi-channel electroencephalogram (EEG) signals plays a significant role in the timely treatment of the patients with epilepsy. Visual identification of epileptic seizure in long-term EEG is cumbersome and tedious for neurologists, which might also lead to human error. Therefore, an automated tool for accurate detection of seizures in a long-term multi-channel EEG is essential for the clinical diagnosis. This study proposes an algorithm using multi-features and multilayer perceptron neural network (MLPNN) classifier. After appropriate approval from the ethical committee, recordings of EEG data were collected from the Institute of Neurosciences, Ramaiah Memorial College and Hospital, Bengaluru. Initially, preprocessing was performed to remove the power-line noise and motion artifacts. Four features, namely power spectral density (Yule-Walker), entropy (Shannon and Renyi), and Teager energy, were extracted. The Wilcoxon rank-sum test and descriptive analysis ensure the suitability of the proposed features for pattern classification. Single and multi-features were fed to the MLPNN classifier to evaluate the performance of the study. The simulation results showed sensitivity, specificity, and false detection rate of 97.1%, 97.8%, and 1 h-1, respectively, using multi-features. Further, the results indicate the proposed study is suitable for real-time seizure recognition from multi-channel EEG recording. The graphical user interface was developed in MATLAB to provide an automated biomarker for normal and epileptic EEG signals.

Essential role of METTL3-mediated m6A modification in glioma stem-like cells maintenance and radioresistance.

Despite advances in biology and therapeutic modalities, existence of highly tumorigenic glioma stem-like cells (GSCs) makes glioblastomas (GBMs) invincible. N6-methyl adenosine (m6A), one of the abundant mRNA modifications catalyzed by methyltransferase-like 3 and 14 (METTL3/14), influences various events in RNA metabolism. Here, we report the crucial role of METTL3-mediated m6A modification in GSC (neurosphere) maintenance and dedifferentiation of glioma cells. METTL3 expression is elevated in GSC and attenuated during differentiation. RNA immunoprecipitation studies identified SOX2 as a bonafide m6A target of METTL3 and the m6A modification of SOX2 mRNA by METTL3 enhanced its stability. The exogenous overexpression of 3'UTR-less SOX2 significantly alleviated the inhibition of neurosphere formation observed in METTL3 silenced GSCs. METTL3 binding and m6A modification in vivo required intact three METTL3/m6A sites present in the SOX2-3'UTR. Further, we found that the recruitment of Human antigen R (HuR) to m6A-modified RNA is essential for SOX2 mRNA stabilization by METTL3. In addition, we found a preferential binding by HuR to the m6A-modified transcripts globally. METTL3 silenced GSCs showed enhanced sensitivity to γ-irradiation and reduced DNA repair as evidenced from the accumulation of γ-H2AX. Exogenous overexpression of 3'UTR-less SOX2 in METTL3 silenced GSCs showed efficient DNA repair and also resulted in the significant rescue of neurosphere formation from METTL3 silencing induced radiosensitivity. Silencing METTL3 inhibited RasV12 mediated transformation of mouse immortalized astrocytes. GBM tumors have elevated levels of METTL3 transcripts and silencing METTL3 in U87/TIC inhibited tumor growth in an intracranial orthotopic mouse model with prolonged mice survival. METTL3 transcript levels predicted poor survival in GBMs which are enriched for GSC-specific signature. Thus our study reports the importance of m6A modification in GSCs and uncovers METTL3 as a potential molecular target in GBM therapy.

Clinical and radiological assessment of cerebral hemodynamics after cranioplasty for decompressive craniectomy - A Clinical study.

OBJECTIVES: To find the correlation between radiologically proven improvement in cerebral hemodynamics with clinical improvement in patients undergoing cranioplasty. MATERIAL AND METHODS: The study is a prospective observational study of 10 cases, in M S Ramaiah Institute of Neurosciences, involving patients treated by a decompressive craniectomy for intractable intra cranial hypertension either due to trauma or stroke and afterwards underwent cranioplasty. RESULTS: Of the 10 patients, 70% patients showing significant improvement in motor functions on Barthel index scale, 60% patients showed improvement in speech, mean duration from date of decompressive craniectomy to cranioplasty being 122.4days. Cerebral perfusion was remarkably better after cranioplasty, as demonstrated decrease in the Pulsatility index on the ipsilateral side of decompression on Trans cranial Doppler (<0.73 mean). This data also favored improved cerebral blood flow and permeability on the CT perfusion with increase in cerebral blood flow (CBF), Cerebral Blood Volume (CBV) and decrease in Time to Peak (TTP) and a positive outcome when correlated with Barthel index with P-values of 0.093, 0.017 and 0.001 respectively. CONCLUSION: Cranioplasty influences the cerebral hemodynamics after cranioplasty and has a positive correlation on the functional outcome and cerebral blood flow in the MCA territory.

Integrative functional genomic analysis identifies epigenetically regulated fibromodulin as an essential gene for glioma cell migration.

An integrative functional genomics study of multiple forms of data are vital for discovering molecular drivers of cancer development and progression. Here, we present an integrated genomic strategy utilizing DNA methylation and transcriptome profile data to discover epigenetically regulated genes implicated in cancer development and invasive progression. More specifically, this analysis identified fibromodulin (FMOD) as a glioblastoma (GBM) upregulated gene because of the loss of promoter methylation. Secreted FMOD promotes glioma cell migration through its ability to induce filamentous actin stress fiber formation. Treatment with cytochalasin D, an actin polymerization inhibitor, significantly reduced the FMOD-induced glioma cell migration. Small interfering RNA and small molecule inhibitor-based studies identified that FMOD-induced glioma cell migration is dependent on integrin-FAK-Src-Rho-ROCK signaling pathway. FMOD lacking C-terminus LRR11 domain (ΔFMOD), which does not bind collagen type I, failed to induce integrin and promote glioma cell migration. Further, FMOD-induced integrin activation and migration was abrogated by a 9-mer wild-type peptide from the FMOD C-terminus. However, the same peptide with mutation in two residues essential for FMOD interaction with collagen type I failed to compete with FMOD, thus signifying the importance of collagen type I-FMOD interaction in integrin activation. Chromatin immunoprecipitation-PCR experiments revealed that transforming growth factor beta-1 (TGF-ß1) regulates FMOD expression through epigenetic remodeling of FMOD promoter that involved demethylation and gain of active histone marks with a simultaneous loss of DNMT3A and EZH2 occupancy, but enrichment of Sma- and Mad-related protein-2 (SMAD2) and CBP. FMOD silencing inhibited the TGF-ß1-mediated glioma cell migration significantly. In univariate and multivariate Cox regression analysis, both FMOD promoter methylation and transcript levels predicted prognosis in GBM. Thus, this study identified several epigenetically regulated alterations responsible for cancer development and progression. Specifically, we found that secreted FMOD as an important regulator of glioma cell migration downstream of TGF-ß1 pathway and forms a potential basis for therapeutic intervention in GBM.

Artery of Percheron infarction after endoscopic pituitary surgery.

Infarction in the artery of Percheron territory is a rare phenomenon in which occlusion of an unpaired perforating artery arising from the P1 segment on one side results in infarcts in the bilateral paramedian thalami with or without midbrain infarcts. We describe the case of a 40-year-old male who developed this complication following re-exploratory trans-sphenoidal surgery for a pituitary adenoma. In this first report of its kind in endoscopic pituitary surgery, the pathogenesis and clinico-radiological features of this rare vascular event are discussed.

Anaplastic hemangiopericytoma in the frontal horn of the lateral ventricle.

Intracranial hemangiopericytomas are uncommon tumors, and their intraventricular occurrence is even rarer. We report a 40-year-old man who presented with raised intracranial pressure. His MRI showed a 3.3 × 3.2 × 3.2 cm heterogeneously enhancing lesion in the left frontal horn obstructing the foramen of Monro and causing hydrocephalus. The tumor was excised through an anterior interhemispheric, transcallosal approach, and histopathology revealed an anaplastic hemangiopericytoma (World Health Organization grade III). To our knowledge this is the first report of this rare pathology being located within the frontal horn of the lateral ventricle.

Infantile holocord cellular ependymoma with communicating hydrocephalus: unusual presentation of a rare case.

We present a case of infantile holocord ependymoma in a 4-month-old boy who presented with infection of ventriculoperitoneal shunt done elsewhere for a communicating hydrocephalus. On magnetic resonance imaging, a diffuse holocord T2-hyperintense, T1-hypointense intramedullary bulky lesion with syringomyelia in the cervical level was seen. To the best of our knowledge, this is the first case of infantile holocord ependymoma. As the extent of morbidity associated with a spinal cord tumor is high, an increased level of suspicion and the need for a complete spinal cord screening in a case of infantile hydrocephalus without obvious clinical and radiological evidence of intracranial abnormality is emphasized.

An interesting case of post traumatic tibialis anterior muscle herniation.

A muscle hernia is defined as a protrusion of the muscle belly through an acquired or congenital fascial defect. Muscle herniation through fascia is a relatively rare entity. Though predominantly asymptomatic, rarely they can be cause of vague pain in the leg, aggravated by exercises. Various conservative measures have been described for asymptomatic hernias, but treatment of symptomatic cases remains controversial. Here we present a case of symptomatic post traumatic tibialis anterior muscle herniation which was treated successfully with autologous fascia lata graft in Yenepoya Medical College, Mangalore in the month of January 2013. Muscle hernias should be kept in mind as a rare differential diagnosis whenever patients present with persisting vague leg pain with or without swelling. If conservative treatment fails, we recommend closure with autologous graft or fasciotomy to relieve the symptoms.

Chordoid glioma in suprasellar location with extension into the third ventricle: smear preparation morphology of a rare tumor.

We report a rare case of recurrent chordoid glioma in suprasellar region with extension into the third ventricle diagnosed peroperatively. On MRI, a brilliantly contrast enhancing lesion in the suprasellar region extending superiorly into the third ventricle and measuring 2.5 × 3 × 3.5 cm was identified. Patient had a previous history of subtotal removal of the tumor 8 months back and had received 56 Gy of postoperative radiotherapy. On smear preparation the tumor was composed of cells arranged in chords and rows in a myxoid background with sprinkling of lymphocytes. The individual cells were polygonal with moderate amount of faintly eosinophilic cytoplasm with cytoplasmic vacoules in few cells. The nuclei showed fine chromatin with small prominent nucleoli. Paraffin sections confirmed the diagnosis and on immunohistochemistry the tumor cells showed strong positivity for GFAP and focal weak positivity for EMA with low MIB-1 labeling index (<1%). To the best of our knowledge this is the first documented case of chordoid glioma described on crush cytology.

Higher topoisomerase 2 alpha gene transcript levels predict better prognosis in GBM patients receiving temozolomide chemotherapy: identification of temozolomide as a TOP2A inhibitor.

The search for molecular markers which predict response to chemotherapy is an important aspect of current neuro-oncology research. MGMT promoter methylation is the only proved marker of glioblastoma. The purpose of this study was to assess the effect of topoisomerase expression on glioblastoma survival and study the mechanisms involved. The transcript levels of all isoforms of the topoisomerase family in all grades of diffuse astrocytoma were assessed. A prospective study of patients with glioblastoma treated by a uniform treatment procedure was performed with the objective of correlating outcome with gene expression. The ability of TOP2A enzyme to relax the super coiled plasmid DNA in the presence of temozolomide was evaluated to assess its effect on TOP2A. The temozolomide cyctotoxicity of TOP2A-silenced U251 cells was assessed. The transcript levels of TOP2A, TOP2B, and TOP3A are upregulated significantly in GBM in comparison with lower grades of astrocytoma and normal brain samples. mRNA levels of TOP2A correlated significantly with survival of the patients. Higher TOP2A transcript levels in GBM patients predicted better prognosis (P = 0.043; HR = 0.889). Interestingly, we noted that temozolomide inhibited TOP2A activity in in-vitro enzyme assays. We also noted that siRNA knock down of TOP2A rendered a glioma cell line resistant to temozolomide chemotherapy. We demonstrated for the first time that temozolomide is also a TOP2A inhibitor and established that TOP2A transcript levels determine the chemosensitivity of glioblastoma to temozolomide therapy. Very high levels of TOP2A are a good prognostic indicator in GBM patients receiving temozolomide chemotherapy.

A clinicopathological study of diagnostically challenging meningioma mimics.

The occurrence of neoplastic and nonneoplastic dural based masses that mimic meningiomas is infrequent and may not be considered during radiological and intraoperative analysis. We describe single institute study of 20 such rare cases. This study included total of 20 cases of meningioma mimics. The clinical, radiological and histopathological findings were evaluated. Tissue fixed in 10% formalin was routinely processed and 5 µ thick sections were cut and stained with hematoxylin & eosin. Histochemistry and immunohistochemistry using avidin-biotin complex immunoperoxidase method was done wherever indicated. In the present study group, 15 were male and 5 female with a male: female ratio of 3:1. The age ranged from 14 to 78 years. Radiologically all these lesions were extra-axial in location, predominantly hypointense on T2W, isointense on T1W images and showed intense homogenous enhancement on contrast administration. Four cases were in pediatric age group with histopathological diagnosis of Rosai Dorfman disease, medulloblastoma, hemangiopericytoma and malignant melanoma. In the adult population, the histopathological diagnoses were hemangiopericytoma, undifferentiated sarcoma, extraskeletal osteosarcoma, Rosai Dorfman disease, medulloblastoma, and metastases from systemic malignancies. Of the total 6 cases of metastases 1 was nonseminomatous germ cell tumor from a primary in testis, 1 was adenocarcinoma from an unknown primary, and 4 were adenocarcinoma from lung. There was a single case of dural based frontal lobe malignant melanoma with congenital hairy nevi on anterior chest wall, scalp, anterior abdominal wall and inguinal region. As the management and biologic behaviour of many of the MM are different, it is essential to familiarize ourselves to them.

Smear preparation of intracranial lesions: a retrospective study of 306 cases.

A cytohistological correlation with determination of accuracy rate of smear preparation result was done in a retrospective study of 306 cases of intracranial tumors. Cytomorphology of few new entities of CNS tumors are described. The cytological features and WHO grading of the tumors were described on smear preparation. The cases with discrepancy in cytological and histological diagnosis were reviewed again and a final possible diagnosis on smear preparation which should have been given is discussed. The clinical details like the age, sex, and site of the tumors were analyzed. The age range of the patients was from 3 years to 63 years with male:female ratio of 1.5:1. Of the total 306 cases, a cytohistological correlation was seen in 93% cases. Twenty-two (7.3%) cases showed discrepancy between the crush preparation diagnosis and final histopathological diagnosis. Majority of the tumors were located in the cerebral hemisphere (56%) and the most frequently diagnosed tumor was astrocytoma, in particular, pilocytic astrocytoma (18.5%) followed by meningioma (11.9%), medulloblastoma (7.3%), anaplastic oligodendroglioma (5%), ependymoma (4.3%), pituitary adenoma (3.3%), schwannoma (3.3%), etc. A few rarer tumors, in central nervous system like differentiating neuroblastoma, pineocytoma, atypical choroid plexus papilloma, piloxmyxoid astrocytoma, rosette forming glioneuronal tumor, etc. are also described, Smear/crush preparation is a very effective, simple, rapid and reliable technique for the diagnosis and WHO grading of central nervous system tumors. Diagnostic accuracy of cytology with final histopathological report is established with accuracy rate of 93%.

Angiolipoma in sellar, suprasellar and parasellar region: report on two new cases and review of literature.

Angiolipomas of CNS are very rare; more than 90% involve the spinal canal. There are only a few case reports documenting the tumor in the sellar/suprasellar region. We report 2 cases of angiolipoma involving sellar/suprasellar and right parasellar region in two female patients. On histopathology the lesion was composed of vascular component which was of cavernous type in one patient and of capillary caliber in the other with admixture of mature adipocytes. The MRI findings were distinctive as the lesion was predominantly iso-to-hypointense on T1W sequence, hyperintense on T2WI and FLAIR sequence. In addition, a few small hyperintense areas were noted on plain T1W images which may represent intralesional fat. One of the cases also showed lipid lactate peak on MR spectroscopy. To the best of our knowledge only 8 other cases of sellar, parasellar angiolipomas have been reported in indexed literature. MRI with fat suppression sequences should be routinely performed in lesions in such locations with hyperintense areas on T1WI as the incidence of intraoperative blood loss is very high. A preoperative suspicion of the diagnosis might help in reducing the patient morbidity.

A supratentorial primary parenchymal paraganglioma.

Primary intracerebral paraganglioma is an unusual entity in the diagnostic armamentarium of the neurosurgeon. Only two patients have been described with a supratentorial intra-parenchymal subtype. We report a 40-year-old man who presented with features of raised intracranial pressure and secondary visual deterioration of short duration. His MRI showed a 2 cm by 2 cm by 2.5 cm, brilliantly enhancing lesion in the left frontal region with flow voids and disproportionate perilesional edema. Magnetic resonance angiography demonstrated a "sun-burst" appearance. The lesion was totally excised, with subsequent resolution of symptoms. Histopathology was reported as a paraganglioma. The systemic workup confirmed the primary intracerebral nature of the lesion. To our knowledge, this report documents a hitherto undescribed, exclusive intraparenchymal location of a rare benign lesion.