myelin-basic-protein and Glioblastoma

Glioblastoma multiforme (GBM), the most common type of central nervous system tumor in humans, is highly proliferative and resistant to apoptosis associated with genetic mutations that deregulate cell cycle. Signal transduction and activation of transcription 3 (Stat3) is a key signal transduction protein that mediated signaling by cytokines and contributed to oncogenesis. It is constitutively activated in numerous cancers including glioblastoma. To determine the effect on proliferation and differentiation of glioblastoma U251 cells after inhibiting STAT3 expression by RNAi, STAT3 gene was silenced with lentivirus vector in U251 cells. We demonstrate that a lentivirus-based shRNA vector had highly infecting efficiency to U251 cells and lentivirus vector-mediated RNAi significantly suppressed Stat3 expression and activation in U251 cells. Knockdown of STAT3 expression by RNAi suppressed the growth and induced apoptosis of U251 cells by down-regulating expression of Bcl-2. It was found that the cell proportion of G0/G1 phase significantly increased after silencing Stat3 by down-regulating expression of cyclin D1. Knockdown of Stat3 also induces morphological changes of U251 cell. It increases significantly expression of myelin basic protein (MBP) in U251 cells. This study demonstrates that STAT3 silencing with lentivirus effectively inhibits STAT3 gene expression and activation. Stat3 is associated with the survival, growth and differentiation of U251 cells. Lentivirus vector-mediated RNAi may be serve as a novel therapeutic strategy for treatment of GBM with expression constitutively and activation of STAT3 gene.

Topics: Analysis of Variance; Apoptosis; Cell Differentiation; Cell Line, Tumor; Cell Proliferation; Flow Cytometry; Gene Expression Regulation, Neoplastic; Glial Fibrillary Acidic Protein; Glioblastoma; Green Fluorescent Proteins; Humans; Lentivirus; Myelin Basic Protein; RNA Interference; STAT3 Transcription Factor; Time Factors; Transduction, Genetic

Many of the strategies developed in the last few years to treat cancer by gene therapy are based on putative killer-suicide genes whose products convert a prodrug into a toxic compound. When the therapy is applied to humans, a vector carrying the killer gene is first inoculated into the tumor of the patient, who 1 week later receives the corresponding prodrug that will selectively kill the cells able to process it to its toxic derivative. A strategy that obviates the need for a prodrug to destroy the cancer cells would be preferable because the patient would only need one treatment instead of two consecutive ones. In the following study, we describe the construction of retroviral vectors in which a reporter or a toxin gene (either the Pseudomonas exotoxin or the Ricinus communis toxin, ricin) is placed under the control of the thyroid hormone (T3) regulatable promoter of the rat myelin basic protein (MBPp). We demonstrate that the expression of these genes under the control of MBPp is regulated by T3 in vitro and in vivo. In vitro, the MBPp is switched off when T3 is removed from the serum of the culture medium, allowing the production of retroviruses carrying the toxic gene. In vivo, the toxin gene bearing retroviruses is capable of eradicating experimentally induced brain tumors in Wistar rats. The gene therapy strategy described here does not require the use of a prodrug to destroy the neoplastic cells.

Topics: Animals; Brain Neoplasms; Female; Fluorescent Antibody Technique; Genes, Reporter; Genetic Therapy; Genetic Vectors; Glioblastoma; Magnetic Resonance Imaging; Mice; Mice, Inbred BALB C; Mice, SCID; Myelin Basic Protein; Neoplasm Transplantation; Plasmids; Promoter Regions, Genetic; Rats; Rats, Wistar; Retroviridae; Ricin; Thyroid Hormones; Toxins, Biological; Transfection

Topics: Animals; Brain Neoplasms; Cell Differentiation; Cell Survival; Female; Fluorescent Antibody Technique, Indirect; Glial Fibrillary Acidic Protein; Glioblastoma; Humans; Male; Mice; Mice, Nude; Myelin Basic Protein; Neoplasm Transplantation; Neoplasms, Experimental; Transplantation, Heterologous; Tumor Cells, Cultured

Intracranial nervous-system tumours were diagnosed in three of 1092 bovine necropsy specimens submitted to the Department of Veterinary Pathology, Obihiro University between April 1983 and March 1996. A fourth case was a referral from the Department of Veterinary Pathology, Rakuno Gakuen University. Histopathological examination revealed four types of tumour: intracranial malignant peripheral nerve sheath tumour (MPNST), choroid plexus papilloma, differentiated fibrillary astrocytoma and anaplastic (malignant) astrocytoma. Immunohistochemically, the intracranial MPNST was strongly positive for S-100 protein and vimentin, and in places weakly positive for glial fibrillary acid protein (GFAP). The choroid plexus papilloma was strongly positive for epithelial membrane antigen (EMA), keratin, S-100 protein and vimentin, and positive for GFAP in places. The cytoplasm and fibrous component in the differentiated fibrillary astrocytoma were strongly positive for S-100 protein and GFAP. The anaplastic (malignant) astrocytoma was strongly positive for vimentin, S-100 protein and keratin in the cytoplasm and fibrous processes, and weakly positive for GFAP and EMA in places. Myelin basic protein (MBP) and synaptophysin showed a weak positive reaction in the marginal areas of the tumour.

Topics: Animals; Astrocytoma; Brain Neoplasms; Cattle; Cattle Diseases; Glial Fibrillary Acidic Protein; Glioblastoma; Glioma; Immunohistochemistry; Keratins; Mucin-1; Myelin Basic Protein; Nerve Sheath Neoplasms; S100 Proteins; Vimentin

Pathological differentiation of oligodendroglioma and mixed oligoastrocytoma from astrocytoma is difficult, relying on morphological characteristics due to the lack of reliable immunohistochemical stains. Oligodendrocytes, the presumed cell of origin of oligodendrogliomas, highly express the genes encoding myelin basic protein (MBP) and proteolipid protein (PLP). We analyzed the expression of these genes to determine whether they might be useful molecular markers of oligodendrocytic tumors. MBP and PLP were highly expressed in all oligodendrogliomas and minimally expressed in glioblastomas multiforme. MBP was highly expressed in mixed oligoastrocytomas, whereas PLP expression was minimal. The association between tumor classification and expression of the MBP and PLP genes was statistically significant. Expression of these genes may serve as a useful molecular marker for some subtypes of human gliomas.

Topics: Adult; Aged; Biomarkers, Tumor; Brain Neoplasms; Female; Gene Expression Regulation, Neoplastic; Glioblastoma; Glioma; Humans; Male; Middle Aged; Myelin Basic Protein; Myelin Proteolipid Protein; Oligodendroglia; Oligodendroglioma; Reverse Transcriptase Polymerase Chain Reaction; Survival Analysis; Time Factors

Human JC virus (JCV) is a neurotropic human polyomavirus that was found in the plaques and oligodendroglial cells of the brains of patients with the fatal demyelinating disease, progressive multifocal leukoencephalopathy (PML). Transgenic mice expressing JCV large tumor (T)-antigen from integrated DNA showed dysmyelination in the central nervous system. However, the role of T-antigen from episomal DNA in the demyelination in PML remains unclear. In this report, we examined the effect of episomally expressed JCV T-antigen on the expression of myelin basic protein (MBP) in U-87 MG human glioblastoma cells to study the mechanism of demyelination. Expression assays of the MBP promoter in U-87 MG detected a 2.5-fold reduction in cells expressing intact T-antigen. Next, U-87 MG expressing T-antigen were examined by RNase protection assays for mRNA accumulation from the endogenous MBP promoter. Also, the expression of the MBP promoter plasmid was determined using in vitro transcription assays with extracts from T-antigen expressing cells. Both assays found a similar down-regulation of the MBP promoter by T-antigen, confirming that negative regulation occurred at the transcriptional level for the endogenous and exogenous MBP promoters. Furthermore, in situ immunofluorescence assays and quantitative Western blot analysis provided convincing evidence of a similar reduction in the level of MBP produced from the functional endogenous gene in U-87 MG glioblastoma cells expressing T-antigen. Thus, we provide evidence for the role of T-antigen in a transcriptional control mechanism for the demyelination that is caused by JCV in PML patients.

Topics: Antigens, Polyomavirus Transforming; Demyelinating Diseases; Down-Regulation; Gene Expression Regulation, Viral; Glioblastoma; Humans; JC Virus; Myelin Basic Protein; Promoter Regions, Genetic; RNA, Messenger; Transcription, Genetic; Tumor Cells, Cultured

We measured the level of myelin basic protein (MBP) in the cerebrospinal fluid (CSF) of patients with various kinds of tumors, including malignant tumors, using radioimmunoassay. The CSF had been obtained by lumbar puncture through an Ommaya reservoir or a shunt device placed in the lateral ventricle. The level of MBP was high (> 4 ng/ml) in the patients with meningeal dissemination of malignant tumors, but in those who showed a good response to chemotherapy and/or radiation, it decreased or returned to the normal level, with improvement on the computed tomography and magnetic resonance imaging, cytological, general CSF, and neurological findings. Of seven malignant gliomas without CSF dissemination, six showed an elevated level of MBP before selective intra-arterial chemotherapy with a combination of etoposide and cisplatin administered via a microcatheter placed at A1, M1, P1-P2, and the basilar top. All CSF specimens obtained during the period of the intra-arterial chemotherapy showed an abnormally high (> 4 ng/ml) level of MBP that exceeded the prechemotherapy level. The MBP level decreased or returned to normal in the patients with a good response to chemotherapy after intra-arterial chemotherapy. In some patients with multiple metastatic brain tumors, the MBP level was elevated before treatment and returned to normal after treatment (surgical removal, chemotherapy, and/or irradiation) in all except one. Thus, there was a clear correlation between the timing of treatment and changes in imaging studies and MBP levels.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adult; Aged; Astrocytoma; Biomarkers, Tumor; Brain Damage, Chronic; Brain Neoplasms; Chemotherapy, Adjuvant; Cisplatin; Combined Modality Therapy; Cranial Irradiation; Etoposide; Female; Follow-Up Studies; Glioblastoma; Humans; Infusions, Intra-Arterial; Lymphoma, B-Cell; Lymphoma, Large B-Cell, Diffuse; Male; Meningeal Neoplasms; Meningioma; Middle Aged; Myelin Basic Protein; Treatment Outcome

We have taken a transgenic approach in an effort to specifically transform oligodendrocytes, the myelinating glial cells of the central nervous system (CNS). Transgenic mice were generated with a DNA construct that contained the activated neu oncogene under the transcriptional control of the myelin basic protein (MBP) gene. The MBP/c-neu transgenic animals have experienced a low incidence of brain tumors that express molecular markers specific to oligodendrocytes, providing a mouse model to study the formation and progression of oligodendrocyte tumors. A tumor from a transgenic animal has been dispersed in culture, and transformed cells that express properties of oligodendrocytes and astrocytes have been maintained. The degree to which these cells express phenotypic characteristic of oligodendrocytes or astrocytes is influenced by culture conditions. These transformed cells should serve as a valuable resource with which to study various molecular and biochemical aspects of the myelination process, as well as the lineage interrelationship of CNS glial cells.

Topics: Animals; Astrocytes; Base Sequence; Biomarkers; Biomarkers, Tumor; Brain Neoplasms; Cell Differentiation; Cell Transformation, Neoplastic; Gene Expression Regulation; Genes, Synthetic; Glioblastoma; Mice; Mice, Inbred C57BL; Mice, Inbred DBA; Mice, Transgenic; Molecular Sequence Data; Myelin Basic Protein; Neoplasm Proteins; Nerve Tissue Proteins; Oligodendroglia; Oncogenes; Organ Specificity; Phenotype; Promoter Regions, Genetic; Proto-Oncogene Proteins; Receptor, ErbB-2; Recombinant Fusion Proteins; Tumor Cells, Cultured

Topics: Animals; Brain; Brain Neoplasms; Cell Line; Encephalomyelitis, Autoimmune, Experimental; Female; Freund's Adjuvant; Glioblastoma; Guinea Pigs; Humans; Immunization; Macaca fascicularis; Male; Myelin Basic Protein; Neoplasm Proteins