[Show abstract][Hide abstract] ABSTRACT: In this study, 293T cells were genetically engineered to secrete tissue inhibitor of metalloproteinase-2 (TIMP2) and encapsulated into alginate microcapsules to continuously release TIMP2 protein.
The anti-invasive potential of the microcapsules was studied in vitro using brain tumor cells. The TIMP2 gene was transfected to 293T cells, and genetically engineered 293TIMP2 cells were encapsulated into alginate microcapsules. Release of TIMP2 protein was detected with Western blot analysis and the anti-invasive potential against U87MG cells was tested using gelatin zymography and a Matrigel assay.
Cell viability within the alginate microcapsules was maintained at a cell density of 5 × 10(6). Because polycationic polymers are helpful for maintaining the mechanical strength of microcapsules with good cell viability, the alginate microcapsules were reinforced with chitosan (0.1% w/v). Expression of TIMP2 protein in cell lysates and secretion of TIMP2 into the conditioned medium was confirmed by Western blot analysis. Alginate microcapsules encapsulating 293TIMP2 cells released TIMP2 protein into the medium efficiently, where the TIMP2 protein participated in degradation of the matrix metalloproteinase-2 enzyme and inhibited invasion of U87MG cells.
Alginate microcapsules encapsulating 293TIMP2 cells are promising candidates for anti-invasive treatment of glioma.
International Journal of Nanomedicine 01/2013; 8:4351-4359. · 4.20 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Metallothionein 1E (MT1E) has been found to be highly expressed in motile cell lines. We investigated whether MT1E actually modulates the migration and invasion of human glioma cell lines and the types of factors that have an effect on MT1E. RNA differential display was performed using Genefishing™ technology in the human glioma cell lines U343MG-A, U87MG and U87MG-10'; the results were validated by RT-PCR and northern blot analysis, in order to detect possible genetic changes as the determining factors for migration ability in malignant glioma. MT1E was identified in U87MG, a highly motile cell line. The migration and invasion abilities of human glioma cell lines, and MT1E transfectants were investigated using simple scratch testing and Matrigel invasion assays. Morphological and cytoskeletal (actin, vimentin) changes were documented by light and confocal microscopy. The expression of MT1E in four glioma cell lines was assessed by RT-PCR and western blotting. In addition, the effects of MT1E on the activity of the NF-κB p50/p65 transcription factor, MMP-2 and -9 were examined by western blotting and zymography. The endogenous MT1E expression in the human glioma cell lines was statistically correlated with their migratory abilities and invasion. The U87-MT-AS cells became more round and had decreased stress fibers, compared with the U87MG cells. Endogenous MT1E expression in the four human glioma cell lines was directly correlated with migration. Two antisense MT1E-transfected cell lines showed decreased NF-κB p50 translocation into the nucleus, which led to decreased activity of MMP-9 in conditioned media. It may be postulated that MT1E can enhance the migration and invasion of human glioma cells by inducing MMP-9 inactivation via the upregulation of NF-κB p50.
International Journal of Oncology 07/2012; · 2.66 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Although bone invasion and hyperostosis are common phenomena in patients with intracranial meningiomas, the basic pathomechanism is not fully understood. Based on an immunohistochemical study of surgically resected samples with hyperostosis, we postulate a possible mechanism of hyperostosis in patients with intracranial meningiomas.
Forty-six meningiomas were evaluated in this study. Twenty-six meningiomas associated with hyperostosis specimens served as the study group, and 20 meningiomas without any bony changes served as controls. An immunohistochemical staining technique was used to detect the expression of matrix metalloproteinase (MMP)-2, -9, and -13, membrane type (MT)1-MMP, estrogen receptor (ER), and progesterone receptor (PR) in the main tumor and hyperostotic portions of the studied samples.
In the non-hyperostosis group, expression of MMP-13, MT1-MMP, and ER was significantly less than in the main tumor portion of hyperostotic meningiomas, while there was no difference in the expression of MMP-2 and -9 and PR in the main tumor between the two groups. In the hyperostosis group, the immunoreactivity of MMP-2 in the hyperostotic portion revealed a higher pattern of expression than the main tumor (p < 0.002). The expression of MMP-9, MT1-MMP, ER, and PR had relatively positive immunoreactivity in the main tumor portion (P < 0.05).
Increased expression of MMP-13 and MT1-MMP in the tumor portion of hyperostosis of meningiomas might contribute to the initiation of osteolysis. Activated MMP-2 in hyperostotic lesions may change the physiological metabolism of the skull bone, thus playing an important role in hyperostosis formation.
[Show abstract][Hide abstract] ABSTRACT: Nogo-A belongs to the reticulon protein family and is expressed in the inner and outer loops of myelin sheaths of oligodendrocytes. We analyzed the patterns of Nogo-A expression in human gliomas in an effort to identify a useful marker for the characterization of oligodendroglial tumors. We determined the expression of Nogo-A in a panel of 58 astrocytic and oligodendroglial tumors using immunohistochemistry and compared the expression of Nogo-A with Olig-2, a recently identified marker for oligodendrogliomas. To localize Nogo-A expression, immunofluorescent staining was performed using other glial markers (MAP-2 and GFAP). We also confirmed the overexpression of the Nogo-A protein in 53 astrocytic and oligodendroglial tumors using Western blot analysis. Based on immunohistochemical analysis, Nogo-A and Olig-2 had specificity in the detection of oligodendroglial tumors from astrocytic tumors (P=0.001). The level of Nogo-A staining was highly correlated with Olig-2 (P=0.001). The sensitivity and specificity of Nogo-A for oligodendroglial tumors was 86.9% and 57.1%, respectively. Nogo-A expression overlapped that of other oligodendroglial markers, but with different patterns of expression. Western blot analysis revealed that Nogo-A is predominantly expressed in 85.7% of oligodendroglioma cells and 93.7% of anaplastic oligodendroglioma cells. Like other oligodendroglial markers, Nogo-A is highly expressed in oligodendroglial tumors; however, it does not serve as a definite marker specific for oligodendroglial tumors.
[Show abstract][Hide abstract] ABSTRACT: Celecoxib, a cyclo-oxygenase (COX)-2 inhibitor, has been reported to mediate growth inhibitory effects and to induce apoptosis in various cancer cell lines. In this study, we examined the potential effects of celecoxib on glioma cell proliferation, migration, and inhibition of COX-2 expression in vitro.
Celecoxib was incorporated into poly DL-lactide-co-glycolide (PLGA) nanoparticles for antitumor drug delivery.
PLGA nanoparticles incorporating celecoxib had spherical shapes and their particle sizes were in the range of 50-200 nm. Drug-loading efficiency was not significantly changed according to the solvent used, except for acetone. Celecoxib was released from the PLGA nanoparticles for more than 2 days, and the higher the drug content, the longer the duration of drug release. PLGA nanoparticles incorporating celecoxib showed cytotoxicity against U87MG tumor cells similar to that of celecoxib administered alone. Furthermore, celecoxib did not affect the degree of migration of U87MG cells. PLGA nanoparticles incorporating celecoxib showed dose-dependent cytotoxicity similar to that of celecoxib alone in C6 rat glioma cells. Western blot assay of the C6 cells showed that neither celecoxib alone nor PLGA nanoparticles incorporating celecoxib affected COX-2 expression.
PLGA nanoparticles incorporating celecoxib had antitumor activity similar to that of celecoxib alone, even though these particles did not affect the degree of migration or COX-2 expression in the tumor cells.
International Journal of Nanomedicine 01/2011; 6:2621-31. · 4.20 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In this study, methoxy poly(ethylene glycol)-grafted carboxymethyl chitosan (CMCPEG) was synthesized to make nanoparticles with doxorubicin (DOX) by ion complex formation. Since DOX has positive amine groups, it can interact with the carboxymethyl group of CMCPEG. The particle size of DOX-incorporated nanoparticles of CMCPEG was < 300 nm and nanoparticles had spherical shapes at morphological observation, indicating that DOX/CMCPEG mixtures can form spherical nanoparticles. In a drug release study, higher drug content induced an extended release of drug. Drug release was significantly changed by the release media pH. DOX release was faster at an acidic pH than a neutral or basic pH. The antitumor activity of DOX-incorporated nanoparticles in vitro was tested with DOX-resistant C6 glioma cells. Nanoparticles showed increased cytotoxicity compared to DOX alone. These results suggest that DOX was unable to penetrate into cells and did not effectively inhibit cell proliferation. In contrast, nanoparticles can penetrate into cells and effectively inhibit cell proliferation. Observation of cells under red fluorescence confirmed these results, i.e., nanoparticle-treated C6 cells, unlike DOX-treated cells, had strong red fluorescence. Since DOX has strong red fluorescence, DOX-incorporated nanoparticles entered into the tumor cells more than DOX alone. As a result, we suggest that DOX-incorporated nanoparticles of CMCPEG are superior candidates for antitumor drug delivery.
[Show abstract][Hide abstract] ABSTRACT: We determined whether the expression of GRIM-19 is correlated with pathologic types and malignant grades in gliomas, and determined the function of GRIM-19 in human gliomas.
Tumor tissues were isolated and frozen at -80 just after surgery. The tissues consisted of normal brain tissue (4), astrocytomas (2), anaplastic astrocytomas (2), oligodendrogliomas (13), anaplastic oligodendrogliomas (11), and glioblastomas (16). To profile tumor-related genes, we applied RNA differential display using a Genefishing DEG kit, and validated the tumor-related genes by reverse transcription polymerase chain reaction (RT-PCR). A human glioblastoma cell line (U343MG-A) was used for the GRIM-19 functional studies. The morphologic and cytoskeletal changes were examined via light and confocal microscopy. The migratory and invasive abilities were investigated by the simple scratch technique and Matrigel assay. The antiproliferative activity was determined by thiazolyl blue Tetrazolium bromide (MTT) assay and FACS analysis.
Based on RT-PCR analysis, the expression of GRIM-19 was higher in astrocytic tumors than oligodendroglial tumors. The expression of GRIM-19 was higher in high-grade tumors than low-grade tumors or normal brain tissue; glioblastomas showed the highest expression. After transfection of GRIM-19 into U343MG-A, the morphology of the sense-transfection cells became larger and more spindly. The antisense-transfection cells became smaller and rounder compared with wild type U343MG-A. The MTT assay showed that the sense-transfection cells were more sensitive to the combination of interferon-beta and retinoic acid than U343MG-A cells or antisense-transfection cells; the anti-proliferative activity was related to apoptosis.
GRIM-19 may be one of the gene profiles which regulate cell death via apoptosis in human gliomas.
Journal of Korean Neurosurgical Society 07/2010; 48(1):20-30. · 0.60 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The aim of this study was to evaluate the correlation and prognostic significance of MGMT promoter methylation and protein expression in patients with glioblastoma.
Eighty-three patients with glioblastoma underwent surgery followed by radiotherapy and temozolomide chemotherapy between October 2000 and June 2008. To investigate the correlation between MGMT methylation and MGMT expression, methylation-specific polymerase chain reaction (MSP) and immunohistochemical staining was performed. To analyze the correlation between MGMT methylation and MGMT expression according to location, biopsies were obtained from 37 different sites within the tumors in 12 patients. Age, sex, Karnofsky Performance Scale status, extent of removal, chemotherapeutic methods, and MGMT promoter methylation and protein expression were analyzed as prognostic factors.
The total median survival was 15.8 months (range, 12.6-19.1 months). The results of MSP were the same at various sites in 12 patients. A correlation between MSP and immunohistochemical staining was observed in 50% of the patients. In 73 patients, negative MGMT expression was detected in 70.5% of 44 patients with MGMT promoter methylation, and positive expression was observed in 55.2% of the 29 patients with unmethylated promoters. Multivariate analysis revealed that the extent of removal (P = 0.001) and the combination of MGMT promoter methylation and negative MGMT expression (median survival, 20.06 months; P = 0.006) were significantly associated with longer survival.
We report the feasibility of using MSP combined with immunohistochemical staining as a prognostic factor. The results of the present study suggest that MGMT promoter methylation in combination with negative MGMT expression might be a good prognostic factor in patients with glioblastoma.
[Show abstract][Hide abstract] ABSTRACT: Hyaluronidase (HAse), a degrading enzyme of hyaluronic acid (HA), is highly expressed in patients with malignant glioma. The purpose of this study was to verify whether HAse is related to the invasion of glioma cells. We also investigated if glioma cells with higher mobility in 2-dimensioal (2-D) method have also higher mobility at 3-dimensional (3-D) environment.
Malignant glioma cell lines (U87MG, U251MG, U343MG-A, and U373MG) were used, and their HAse expressions were evaluated by HA zymography. The migration ability was evaluated by simple scratch technique. The invasiveness of each cell lines was evaluated by Matrigel invasion assay and HA hydrogel invasion assay. In HA hydrogel invasion assay, colonies larger than 150 microm were regarded as positive ones and counted. Statistical analysis of migration ability and invasion properties of each cell lines was performed using t-test.
In scratch test to examine migration ability of each cell lines, U87MG cells were most motile than others, and U343MG-A least motile. The HAse was expressed in U251MG and U343MG-A cell lines. However, U87MG and U373MG cell lines did not express HAse activity. In Matrigel invasion assay, the cell lines expressing HAse (U251MG and U343MG-A) were more invasive in the presence of HA than HAse deficient cell lines (U87MG and U373MG). In HA hydrogel invasion assay, the HAse-expressing cell lines formed colonies more invasively than HAse-deficient ones.
Malignant Glioma cells expressing HAse were more invasive than HAse-deficient ones in 3-dimensional environment. Therefore, it might be suggested that invasion of malignant gliomas is suppressed by inhibition of HAse expression or HA secretion. Additionally, the ability of 2-D migration and 3-D invasion might not be always coincident to each other in malignant glioma cells.
Journal of Korean Neurosurgical Society 11/2009; 46(5):472-8. · 0.60 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Although matrix metalloproteinases (MMPs) play a crucial role in the invasion and growth of malignant gliomas, their increased activity in tumor environment can be used as a specific target for chemotherapy. We investigated whether polymer-drug conjugates formed via MMP-cleavable peptide linkages could provide MMP-responsive tumor targeting and cytotoxicity for malignant glioma cells. One end of an MMP-cleavable peptide was attached to the end of methoxy polyethylene glycol (MPEG) while the other end was attached to adriamycin (ADR). The release of drugs in the presence of conditioned media of U87MG cells was investigated. The cytotoxicities of the MMP-cleavable MPEG-peptide-ADR (PPA) conjugates and non-cleavable MPEG-ADR (PA) conjugates were investigated using U87MG cells. The (1)H nuclear magnetic resonance (NMR) spectra confirmed the conjugation of the two ends of the peptide to the ends of MPEG and ADR, respectively. Gelatin zymography showed that MMP-2 was strongly expressed in the media of U87MG cells. The PA conjugate did not release ADR either in the phosphate buffered saline (PBS) or conditioned media of U87MG cells. The PPA conjugate released ADR in the presence of the conditioned media of U87MG cells, but not in PBS only. In the cytotoxicity test using U87MG cells, ADR and PPA conjugate showed similar anti-proliferative activities, while the cytotoxicity of PA conjugate was lower than that of ADR. Considering that the cytotoxicity of the PPA conjugate was similar to that of ADR, MMP-cleavable polymer-drug conjugates can be used as targeting carriers for the purpose of inhibiting the proliferation of malignant glioma cells.
International Journal of Pharmaceutics 11/2009; 387(1-2):209-14. · 3.99 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Galectin-1 is highly expressed in motile cell lines. The authors investigated whether galectin-1 actually modulates the migration and invasion of human glioblastoma multiforme (GBM) cell lines, and whether its expression with respect to invasion and prognosis is attributable to certain glioma subgroups.
In the human GBM cell lines U343MG-A, U87MG, and U87MG-10', the RNA differential display was evaluated using Genefishing technology. The results were validated by reverse transcription polymerase chain reaction and Northern blot analysis to detect possible genetic changes as the determining factors for the motility of the malignant glioma. The migration and invasion abilities were investigated in human GBM cell lines and galectin-1 transfectant using an in vitro brain slice invasion model and a simple scratch technique. The morphological and cytoskeletal (such as the development of actin and vimentin) changes were examined under light and confocal microscopy. Galectin-1 expression was assessed on immunohistochemical tests and Western blot analysis.
Endogenous galectin-1 expression in the human GBM cell lines was statistically correlated with migratory abilities and invasiveness. The U87-G-AS cells became more round than the U87MG cells and lacked lamellipodia. On immunohistochemical staining, galectin-1 expression was increased in higher-grade glioma subgroups (p = 0.027).
Diffuse gliomas demonstrated higher expression levels than pilocytic astrocytoma in the Western blot. Galectin-1 appears to modulate migration and invasion in human glioma cell lines and may play a role in tumor progression and invasiveness in human gliomas.
Journal of Neurosurgery 09/2008; 109(2):273-84. · 3.15 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The aim of this study is to prepare cisplatin-incorporated nanoparticles based on ion complex formation between hyaluronic acid (HA) and cisplatin for antitumor drug delivery. To prepare nanoparticles using HA, bulk HA was degraded by hyaluronidases (HAses). Cisplatin-incorporated HA nanoparticles were prepared by mixing cisplatin with an aqueous solution of HA and then the nanoparticle solution was dialyzed to remove trace elements. Since glioma tumor cell lines are able to secrete HAse, extracts from U343MG and U87MG cell lines were used to test the release of cisplatin from the nanoparticles. The morphological observation of the cisplatin-incorporated nanoparticles showed that they had spherical shapes with a particle size around 100-200 nm. The loading efficiency of cisplatin in the nanoparticles was about 67-81% (w/w) and cisplatin was continuously released from the nanoparticles for 4 days. Especially, the release rate of cisplatin from the nanoparticles increased when HAse was added to the release medium. In the results of the HA zymography, the U343MG cell line secreted HAse, while the U87MG cell line did not. When the extracts from U343MG were added to the release medium, the release rate of cisplatin was slightly increased, while the extracts from U87MG did not significantly affect the release rate of cisplatin. In conclusion, cisplatin-incorporated nanoparticles have sufficiently small particle sizes to use as a drug targeting system. The release of cisplatin from the nanoparticles was responsive to the secretion of HAse. These nanoparticles are suitable vehicles for an antitumor drug targeting system.
Journal of Pharmaceutical Sciences 04/2008; 97(3):1268-76. · 3.13 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Subdural fluid collections appear in about 39% of patients after the removal of intra- and paraventricular tumors. This extracerebral fluid collection requires surgical intervention when progressive fluid accumulation takes place. The authors retrospectively and prospectively studied the efficacy of gelfoam and fibrin adhesive in closing cortical and ependymal defects after intraventricular and/or paraventricular lesion resection to prevent the development of SFCs.
From 1999 to 2004, we used gelfoam and fibrin adhesive on the cortical and ependymal defects of 28 patients who underwent the resection of intraventricular and/or paraventricular lesions via the transcortical approach associated with the communicated ventricle. We investigated the percentage of symptomatic and asymptomatic SFC.
The patients median age was 59.5 years (range, 30-76 years), and the male/female ratio was 16:12. A frontal approach was performed in 18 patients, an occipital approach in 2, a parietal approach in 4, and a temporal approach in 4. The incidence of SFCs was 7% (2 patients). Of the 2 patients with SFCs, 1 required temporary drainage. The other patient was asymptomatic, and the SFCs were spontaneously absorbed 2 months later.
The use of gelfoam and fibrin adhesive to seal cortical and ependymal defects after a transcortical procedure might be a viable method of preventing the development of SFC.
[Show abstract][Hide abstract] ABSTRACT: We report a patient with a intracranial subdural osteoma with a large cortical vein passing through the subdural calcified mass. A 60-year-old man presented with an approximately 3-year history of persistent headache. Computerized tomography (CT) scanning showed a homogeneous high-density nodule attached to the inner surface of the right frontal skull. Intraoperatively, the hard mass was found to be located in the intradural subarachnoid space. A large cortical vein passed through the subdural mass and was anastomosed in an end-to-end fashion after the excision of the segment involved by the tumor. The histopathologic examination showed lamellated bony trabeculae lined by osteoblasts and the underlying dura was uninvolved by the tumor cells.
Journal of Clinical Neuroscience 06/2007; 14(5):468-70. · 1.25 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The authors evaluated the clinical manifestations and surgical results in patients with cystic vestibular schwannoma (VS), and investigated the matrix metalloproteinase (MMP) expression of the cyst fluid and wall in an attempt to elucidate the pathogenesis and characteristics of this disease.
The clinical and neuroimaging features, perioperative findings, and surgical outcomes in 24 cases of cystic VS and 82 cases of solid VS, all of which were treated using the suboccipital approach, were retrospectively compared. To evaluate the role of MMP in cystic VS, gelatin zymography and immunohistochemical studies of the cyst fluid, wall, and solid portion were performed in nine cases of this disease. The mean duration of symptoms was shorter (14.0 months compared with 26.1 months; p = 0.04) and the mean size of the tumor was larger (43.8 mm compared with 34.2 mm; p = 0.048) in the cystic than the solid VS group. Although gross-total resection was easier to accomplish in this group (100% compared with 84.1%), adhesion to the facial nerve was more frequent (62.5% compared with 48.8%; p = 0.042). On gelatin zymography studies, MMP-2 expression was ubiquitously observed in all cyst fluids. Immunohistochemical analysis of the cyst wall showed that MMP-2 was apparently localized to the tumor cells on the luminal inner surface, adjacent to the cyst cavity.
Resection of cystic VS is complicated by severe adhesion of the tumor capsule to the facial nerve and the large size of the lesion. The authors believe that MMP-2 may be involved in the pathogenesis of cyst formation or in its enlargement and may aggravate adhesion to the facial nerve, either by promoting the enlargement of the tumor or engendering the degradation of the tumor-nerve barrier proteolytically.
Journal of Neurosurgery 06/2007; 106(5):866-71. · 3.15 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The most characteristic feature of a malignant astrocytoma is its early and extensive infiltration into adjacent parenchymal structures. We focused on detecting the possible expression changes as the determining factors for malignant astrocytoma's motile ability. We confirmed that four of 39 genes showed different expression on DD-PCR by RT-PCR and Northern blot analysis. These findings suggest that the genes identified may be important for determining high motility in astrocytoma cell lines. These findings may help us understand the molecular invasion mechanism in astrocytomas.
Journal of Neuro-Oncology 05/2007; 82(2):125-31. · 3.12 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Surgery for meningiomas involving the cavernous sinus remains controversial. Interdural cavernous sinus is called the lateral dural wall in the cavernous sinus, which is composed of two layers, the outer dural layer and the inner membranous layer. We encountered two cases of dumbbell-shaped middle cranial fossa meningioma with interdural cavernous sinus extension, which were successfully removed by surgical means.
A 57-year-old woman presented with headache and decreased visual acuity. Neurological assessment was normal. Computed tomography and magnetic resonance imaging showed the presence of a dumbbell-shaped, smooth-contoured, well-enhanced mass in the right mesial temporal area. The lateral wall of the cavernous sinus was exposed via frontotemporal craniotomy and the tumor originating in the lateral wall was totally removed. A 41-year-old man presented with seizure attacks and drowsy mental status. Magnetic resonance imaging showed the presence of a multilobulated, well-enhanced mass in the left parasellar area. The tumor was totally resected via a transsylvian temporopolar approach. The mass originated from tentorial edge and extended into the cavernous sinus by dural penetration.
Middle cranial fossa meningioma with interdural cavernous sinus extension can be removed more easily than other tumors with intracavernous sinus extension and, consequently, can be safely resected without any resulting cranial nerve deficit.