Involvement of Akt/NF-κB pathway in antitumor effects of parthenolide on glioblastoma cells in vitro and in vivo.

BACKGROUND: Glioblastoma is the most common and most aggressive form of malignant glioma and is very difficult to treat. Controlling tumour cell invasion and angiogenesis is essential to improve the prognosis of glioblastoma patients. Since constitutive activation of nuclear factor-κB (NF-κB) is necessary for tumour progression, NF-κB may be an important pharmacological target for this disease. Our study aimed to evaluate the antitumour effects of parthenolide, a NF-κB inhibitor, in two human glioblastoma cell lines (U87MG and U373) and in glioblastoma xenografts. Furthermore, we aimed to investigate the molecular mechanisms underlying these effects. METHODS: The anti-invasive and anti-angiogenic effects of parthenolide were analysed using in vitro invasion and angiogenesis assays. Parthenolide-induced growth inhibition of glioblastoma cells in vitro was determined using the MTT (methyl thiazolyl tetrazolium) assay. In addition, the effect of parthenolide on orthotropic implantation in vivo was evaluated using an intracerebral human glioblastoma xenograft model. RESULTS: We found that parthenolide suppresses proliferation, invasion, and tumour- induced angiogenesis of glioblastoma cells. Molecular studies demonstrated that parthenolide suppresses gene and protein expression of angiogenic factors. Furthermore, parthenolide reduced Akt phosphorylation and activated mitochondrial signalling, suggesting that the antitumour function of parthenolide may be mediated not only by the inhibition of NF-κB but also by the inhibition of Akt signalling and the activation of apoptotic proteins. Parthenolide suppressed neovascularity and tumour growth in glioblastoma xenografts. CONCLUSION: The present study identified parthenolide as a new therapeutic agent for glioblastomas.

Stereoscopic virtual realistic surgical simulation in intracranial aneurysms.

BACKGROUND: Three-dimensional (3D)-computed tomographic angiography (CTA) has been widely used for surgical simulation of intracranial aneurysms. Stereo imaging technology is progressing rapidly in recent years and stereo imaging may make more realistic surgical simulation possible. Therefore, we aimed at the establishment of a technique for stereoscopic viewing of minute volume rendering images while pursuing a low cost. MATERIALS AND METHODS: Between January 2009 and June 2011, 54 patients with ruptured intracranial aneurysms were enrolled in this study. CTA data was transferred to the workstation equipped with image-processing software, and multilayer fusion images were processed by neurosurgeons. Image data for stereoscopic viewing of multilayer fusion image from arbitrary directions were collected form rotational trajectories around an aneurysm and were output to MPEG file. Stereoscopic viewing using MPEG data was achieved by the freeware named Stereo Movie Maker. Stereo viewing method using QuickTime VR format was also tried. RESULTS: Multilayer fusion image created from CTA data displayed clearly the anatomical information about not only the aneurysm but also the surrounding structures, such as parent artery, venous system, brain tissue, skull bone, and scalp. The quality of the resulting multilayer fusion image was suitable for surgical simulation with virtual reality. Virtual realistic surgical simulation became possible by the combination of minute multilayer fusion image and stereoscopic viewing by our method. CONCLUSIONS: Our method for stereo viewing of multilayer fusion images resulted in an improvement in the capability of diagnostic imaging and the image-guided support for neurosurgical procedures in intracranial aneurysm.

HA1077, a Rho kinase inhibitor, suppresses glioma-induced angiogenesis by targeting the Rho-ROCK and the mitogen-activated protein kinase kinase/extracellular signal-regulated kinase (MEK/ERK) signal pathways.

Malignant gliomas are characterized by high invasive potential and strong angiogenic ability. Constitutive activation of Rho and its downstream target Rho-kinase are crucial for tumor progression. We investigated the effect of the Rho-kinase inhibitor HA1077 on tumor-induced angiogenesis of malignant glioma cells and explored the molecular mechanisms underlying the effect of HA1077. Here, we demonstrated that HA1077 suppressed tube formation of endothelial cells in human umbilical vein endothelial cell (HUVEC)-glioma cell co-culture assay. Western blot, RT-PCR, ELISA and zymography demonstrated that HA1077 suppressed gene and protein expressions of vascular endothelial growth factor (VEGF), matrix metalloproteinase (MMP)-2 and MMP-9 in glioma cells. Furthermore, HA1077 attenuated the phosphorylation of extracellular signal-regulated kinases 1 and 2 (ERK1/2) and the DNA binding activity of activator protein-1, a key downstream transcriptional factor of ERK1/2. HA1077 also suppressed the migration of HUVEC in vitro. Thus, it is suggested that the anti-angiogenic effect of HA1077 may be due to the combination of ROCK inhibition and mitogen-activated protein kinase kinase (MEK)/ERK pathway inhibition. Moreover, an in vivo intracerebral human glioma cell xenograft mouse model demonstrated that HA1077 suppressed neovascularity and tumor growth. The results of the present study suggest that HA1077 has a therapeutic potential as an anti-angiogenic agent against malignant gliomas.

Anti-invasive and antiangiogenic effects of MMI-166 on malignant glioma cells.

BACKGROUND: The constitutive overexpression of matrix metalloproteinases (MMPs) is frequently observed in malignant tumours. In particular, MMP-2 and MMP-9 have been reported to be closely associated with invasion and angiogenesis in malignant gliomas. Our study aimed to evaluate the antitumour effects of MMI-166 (Nalpha-[4-(2-Phenyl-2H- tetrazole-5-yl) phenyl sulfonyl]-D-tryptophan), a third generation MMP inhibitor, on three human glioma cell lines (T98G, U87MG, and ONS12) in vitro and in vivo. METHODS: The effects of MMI-166 on the gelatinolytic activity was analysed by gelatine zymography. The anti-invasive effect of MMI-166 was analysed by an in vitro invasion assay. An in vitro angiogenesis assay was also performed. In vitro growth inhibition of glioma cells by MMI-166 was determined by the MTT assay. The effect of MMI-166 on an orthotropic implantation model using athymic mice was also evaluated. RESULTS: Gelatine zymography revealed that MMP-2 and MMP-9 activities were suppressed by MMI-166. The invasion of glioma cells was suppressed by MMI-166. The angiogenesis assay showed that MMI-166 had a suppressive effect on glioma cell-induced angiogenesis. However, MMI-166 did not suppress glioma cell proliferation in the MTT assay. In vivo, MMI-166 suppressed tumour growth in athymic mice implanted orthotropically with T98G cells and showed an inhibitory effect on tumour-induced angiogenesis and tumour growth. This is the first report of the effect of a third generation MMP inhibitor on malignant glioma cells. CONCLUSIONS: These results suggest that MMI-166 may have potentially suppressive effects on the invasion and angiogenesis of malignant gliomas.

Enhanced expression of cancer testis antigen genes in glioma stem cells.

Cancer stem cells are an important target for effective therapy, since they show tumorigenicity, chemoresistance, and radioresistance. We isolated cancer stem cells from glioma cell lines and tissues and examined the expression of cancer testis antigen (CTA) genes as potential target molecules for cancer vaccine therapy. CTA genes were highly and frequently expressed in cancer stem cells compared with differentiated cells. In addition, histone acetylation levels in the promoter regions of CTA genes were high in cancer stem cells and low in differentiated cells, while DNA methylation analysis of the promoter regions revealed hypomethylation in cancer stem cells. This epigenetic difference between cells leads to heterogeneous expression of CTA genes in the tumor mass, which consists of cells at various levels of differentiation. Moreover, the expression level of HLA class I antigens was not affected by the differentiation status, suggesting that CTA genes may present as surface antigens in cancer stem cells. Taken together, these findings suggest that CTA genes may be attractive candidates for targeted vaccine therapy against cancer stem cells in glioma patients.

Enhanced MDR1 expression and chemoresistance of cancer stem cells derived from glioblastoma.

We established a cancer stem (CS) cell line, U87CS, by means of spheroid culture of U87MG cells derived from glioblastoma (GBM) in neuronal stem cell medium. U87CS cells presented positive immunohistochemical staining for multidrug resistance (MDR)1 and CD133, a marker for a subset of leukemia and GBM CS cells. The gene expression of MDR1 and CD133 on U87CS cells increased by an average of 8.51 and 47.18 times, respectively, compared to the levels on U87MG cells by real-time quantitative RT-PCR. U87CS cells possessed stronger drug-resistance to conventional anti-cancer drugs, such as doxorubicin (Dox), etoposide (VP-16), carboplastin, and BCNU than U87MG cells. Double immunofluoresence staining showed co-expression of MDR1 and CD133 on U87CS cells transplanted into nude mice brains. In addition, we identified the crossreactivity of CD133 and MDR1 in a surgical specimen of GBM. Our results suggest that CS cells may be resistant to current chemotherapy and represent a novel target for GBM therapeutics.

Rathke's cleft cyst with short-term size changes in response to glucocorticoid replacement.

An 81-year-old man was admitted to our hospital because of general fatigue. Hormonal examination showed that he had panhypopituitarism and central diabetes insipidus. MRI imaging revealed the presence of large cystic mass with suprasellar extension in his hypothalamo-pituitary region. Interestingly, the cystic mass shrank following the start of glucocorticoid replacement, and since then relatively high doses of cortisol administration were needed to prevent the re-enlargement of cystic size. Because of the concern over possible side effects of supraphysiological doses of glucocorticoid replacement, surgical treatment was eventually carried out, confirming the pathological feature of Rathke's cleft cyst. The present case suggests that the inflammatory nature of Rathke's cleft cyst may explain the observed short-term size changes in response to glucocorticoid administration.

Clinico-pathological significance of RCAS1 expression in gliomas: a potential mechanism of tumor immune escape.

RCAS1, one of the tumor cell surface antigens, is strongly expressed in aggressive tumors. RCAS1 suppresses the in vitro growth of immune effector cells. We investigated the expression of RCAS1 in 57 gliomas using immunohistochemistry. Furthermore, we examined the association of the RCAS1 expression with the infiltration of tumor infiltrating lymphocyte (TIL). RCAS1 overexpression was significantly correlated with high histological grade and poor prognosis. Reduced infiltration and increased apoptosis of TILs was observed in RCAS1-positive regions. Apoptotsis of TILs appeared to be induced by RCAS1. RCAS1 expression in gliomas may play roles in tumor progression and tumor immune escape.

Primary malignant B-cell-type dural lymphoma: Case report.

BACKGROUND: The primary malignant dural lymphoma of B-cell type is rare. Our review of the literature (24 cases) indicated that patients with this tumor had female predominance, immunocompetency, and longer survival compared with those with primary malignant intracerebral lymphomas. Based on its clinicopathological features, this dural lymphoma may be classified differently from other types of malignant lymphomas in the central nervous system. CASE DESCRIPTION: The authors report an example of a patient who had a favorable course of malignant dural lymphoma. A 59-year-old woman presented with primary malignant dural lymphoma in the frontal area. Histological diagnosis was a diffuse large B-cell-type lymphoma of intermediate malignancy, but MIB-1 index was extremely high. Subtotal tumor removal was followed by 4 courses of chemotherapy. Thirty months after surgery, the patient is asymptomatic with no MRI evidence of recurrence. A combination of subtotal resection and chemotherapy for a primary malignant dural lymphoma is an effective means to attempt cure of this tumor. CONCLUSION: These rare tumors may be managed successfully by subtotal resection and postoperative treatment. We believe that chemotherapy is the treatment of choice after subtotal resection of malignant dural lymphomas, as experienced in our case.

A rapidly enlarging nocardial brain abscess mimicking malignant glioma.

Nocardial brain abscesses are uncommon and are not preceded by clear infectious symptoms in most cases. Delayed identification of the bacteria is responsible for a high mortality rate. A 58-year-old afebrile woman was admitted to our hospital because of progressive right hemiparesis and aphasia. Magnetic resonance imaging (MRI) showed a single ring-enhanced lesion in the left frontal lobe. It was extremely difficult to establish the diagnosis of brain abscess, because the laboratory data provided little evidence of bacterial infection, (201)TlCl-scintigraphy revealed definite accumulation of thallium in the lesion, and follow-up MRI demonstrated rapid enlargement of the lesion. Total resection was performed because of the possibility of a malignant brain tumor, but brain abscess was finally diagnosed with histological examination. A nocardial species was detected through microscopic examination of the pus obtained at surgery, and this precise diagnosis of nocardial brain abscess in the early stage enabled the administration of appropriate antibiotics and the patient's quick recovery. Nocardial brain abscesses are often misdiagnosed as malignant brain tumors, and a definitive diagnosis may not be possible without detecting bacteria from the lesion. Total excision of the abscess can produce good results when the abscess is large and located superficially, but incomplete aspiration and drainage of a lesion is associated with a high chance of relapse.

Clinicopathologic significance of cystatin C expression in gliomas.

Cathepsin B, one of the lysosomal cysteine proteases, has been related to tumor invasiveness. Cystatin C is the strongest inhibitor of cathepsin B. Knowledge of its participation in the progression of gliomas is limited. We investigated the expression of cystatin C and its association with the clinicopathologic features of 57 gliomas. Cystatin C and cathepsin B expressions were evaluated by immunohistochemical methods and by semiquantitative real-time polymerase chain reaction analysis for the corresponding messenger RNA. Disease-free survival was analyzed by the Kaplan-Meier method. Tumors with low cystatin C protein expression and high cathepsin B protein expression were significantly more likely to be of high grade, and this pattern was significantly correlated with high Ki-67 LI and tumor recurrence. Depressed expression of cystatin C messenger RNA in glioblastomas compared with low-grade astrocytomas was demonstrated. Multivariate analysis demonstrated high tumor grade, high Ki-67 labeling index, high cathepsin B expression, and low cystatin C expression correlated significantly with shorter disease-free survival. These results suggest that gliomas in patients with an unfavorable clinical outcome are characterized by depressed expression of cystatin C. Evaluation of cystatin C expression in gliomas provides useful clinical information, especially as a prognostic indicator.

Characterization of microglia induced from mouse embryonic stem cells and their migration into the brain parenchyma.

We derived microglia from mouse embryonic stem cells (ES cells) at very high density. Using the markers Mac1(+)/CD45(low) and Mac1(+)/CD45(high) to define microglia and macrophages, respectively, we show that Mac1(+) cells are induced by GM-CSF stimulation following neuronal differentiation of mouse ES cells using a five-step method. CD45(low) expression was high and CD45(high) expression was low on induced cells. We used a density gradient method to obtain a large amount of microglia-like cells, approximately 90% of Mac1(+) cells. Microglia-like cells expressed MHC class I, class II, CD40, CD80, CD86, and IFN-gammaR. The expression level of these molecules on microglia-like cells was barely enhanced by IFN-gamma. Intravenously transferred GFP(+) microglia derived from GFP(+) ES cells selectively accumulated in brain but not in peripheral tissues such as spleen and lymph node. GFP(+) cells were detected mainly in corpus callosum and hippocampus but were rarely seen in cerebral cortex, where Iba1, another marker of microglia, is primarily expressed. Furthermore, both GFP(+) and Iba1(+) cells exhibited a ramified morphology characteristic of mature microglia. These studies suggest that ES cell-derived microglia-like cells obtained using our protocol are functional and migrate selectively into the brain but not into peripheral tissues after intravenous transplantation.

Primitive neuroectodermal tumor (PNET) treated with interferon-beta after surgical removal and irradiation: case report.

This case report describes an 11-year-old boy with primitive neuroectodermal tumor (PNET), which remains the pediatric brain tumor with the worst prognosis despite combination treatment with surgery, irradiation and anti-cancer drugs. The boy was successfully treated with intratumoral administration of interferon-beta (IFN-beta) following surgical resection and irradiation. Two million units of IFN-beta were locally administered into the post-operative cavity through the Ommaya's reservoir every two weeks after discharge. He was managed as an outpatient without serious side-effects to IFN. The patient has shown no tumor recurrence, mental retardation, or neuroendocrine impairment for over three years and has lived a normal school life with a full Karnofsky Performance Status score. The local administration of IFN-beta may be warranted for maintenance therapy in children with PNET.

Prognostic significance of cyclin a expression in meningiomas.

Unconstrained cell proliferation is characteristic of tumors. It is caused by the functional disorders of proteins that constitute the cell cycle mechanism. The cell cycle is controlled by cyclins, cyclin-dependent kinases, and cyclin-dependent kinase inhibitors. Many reports have proved, in cancers, that cyclins, cyclin-dependent kinases, and cyclin-dependent kinase inhibitors are out of control. Cyclin A is a protein that regulates critical transition of the cell cycle. The expression of cyclin A in meningiomas by immunohistochemical method was investigated. Furthermore, the correlation among cyclin A expression, clinical course, and proliferative potential were also evaluated. Seventy-seven meningiomas were studied. The mean cyclin A labeling indices were as follows: benign meningiomas, 1.01% +/- 0.62%; atypical meningiomas, 4.23% +/- 1.82%; and anaplastic meningiomas, 7.72% +/- 0.88%. Analyses of variance showed that significant differences existed between tumor grades for cyclin A labeling indices. A linear positive correlation between the cyclin A labeling index and bromodeoxyuridine labeling index was observed. The multivariate analysis using Cox's hazards model showed a high cyclin A labeling index (>3%) was a significant risk factor for recurrence. A high Ki-67 labeling index (>5%) and high tumor grade (World Health Organization grade II, III) were also significant risk factors for recurrence. These results suggested that the evaluation of cyclin A expression in meningiomas provides significant clinical information, especially as an independent prognostic indicator.