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Differential expression of novH and CTGF in human glioma cell lines.

ABSTRACT: Aims-(1) To investigate the expression in human derived glioblastoma cell lines of two structurally related genes, novH (nephroblastoma overexpressed gene) and CTGF (connective tissue growth factor), which encode putative insulin-like growth factor binding proteins of a novel type. (2) To investigate whether the same transcription factors regulate CTGF and novH expression.Methods-Expression of novH and CTGF was analysed in 24 glioblastoma derived cell lines by northern blotting. The CTGF promoter region was characterised by nucleotide sequencing, RNase protection experiments, by transient transfections, and CAT assays.Results-CTGF and novH mRNA levels differed in the glioma cell lines studied. NovH and CTGF genes were not co-expressed in all cell lines. The CTGF promoter region was highly conserved compared with the corresponding region in the mouse (FISP12) and exhibited in vitro transcriptional activity.Conclusions-Although the coding regions of novH and CTGF are highly homologous, their promoter regions are substantially different, suggesting that these two genes may be regulated by different mechanisms. Considering that novH and CTGF are likely to be, respectively, negative and positive regulators of growth and that some glioma cell lines expressing novH are not tumorigenic, expression of these two genes might represent a key element in determining the stage of differentiation or the malignant potential, or both, of some tumour cell lines.

SUBMITTER: Xin LW

PROVIDER: S-EPMC408028 | biostudies-other | 1996 Apr

REPOSITORIES: biostudies-other

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Differential expression of novH and CTGF in human glioma cell lines.

Xin L W LW Martinerie C C Zumkeller W W Westphal M M Perbal B B

Clinical molecular pathology 19960401 2

Aims-(1) To investigate the expression in human derived glioblastoma cell lines of two structurally related genes, novH (nephroblastoma overexpressed gene) and CTGF (connective tissue growth factor), which encode putative insulin-like growth factor binding proteins of a novel type. (2) To investigate whether the same transcription factors regulate CTGF and novH expression.Methods-Expression of novH and CTGF was analysed in 24 glioblastoma derived cell lines by northern blotting. The CTGF promote ...[more]

PMID: 16696057

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Project description:Zidovudine (3'-azido-3'-deoxythymidine; AZT) is the most widely used nucleoside reverse transcriptase inhibitor for the treatment of AIDS patients and prevention of mother-to-child transmission of HIV-1. Previously, we demonstrated that AZT had significantly greater growth inhibitory effects upon the human liver carcinoma cell line HepG2 as compared to the immortalized human liver cell line THLE2. We have now used gene expression profiling to determine the molecular pathways associated with toxicity in both cell lines. HepG2 cells were incubated with 0, 2, 20, or 100 ?M AZT for 2 weeks; THLE2 cells were treated with 0, 50, 500, or 2,500 ?M AZT, concentrations that were equi-toxic to those used in the HepG2 cells. After the treatment, total RNA was isolated and subjected to microarray analysis. Global analysis of gene expression, with a false discovery rate ?0.01 and a fold change ?1.5, indicated that 6- to 70-fold more genes were differentially expressed in a significant concentration-dependent manner in HepG2 cells when compared to THLE2 cells. Comparative analysis indicated that 7 % of these genes were common to both cell lines. Among the common differentially expressed genes, 70 % changed in the same direction, most of which were associated with cell death and survival, cell cycle, cell growth and proliferation, and DNA replication, recombination, and repair. As determined by the uptake of [methyl-(3)H]AZT, the intracellular levels of total AZT were approximately twofold higher in THLE2 cells than in HepG2 cells. The expression of thymidine kinase 1 (TK1) and UDP-glucuronosyltransferase 2B7 (UGT2B7) genes that regulate the metabolic activation and deactivation of AZT, respectively, was increased in HepG2 cells but decreased in THLE2 cells after treatment with AZT. This differential response in AZT metabolism was confirmed by real-time PCR, western blotting, and/or enzymatic assays. These data indicate that molecular pathways involved with cell death and survival, cell cycle, cell growth and proliferation, and DNA replication, recombination, and repair are involved in the toxicities associated with AZT in both human cell lines, and that the difference in expression of TK1 and UGT2B7 in response to AZT treatment in HepG2 cells and THLE2 cells might explain why HepG2 cells are more sensitive than THLE2 cells to the toxicity of AZT.

Project description:BackgroundGlioblastoma multiforme (GBM) is the most common primary intracranial tumor and despite recent advances in treatment regimens, prognosis for affected patients remains poor. Active cell migration and invasion of GBM cells ultimately lead to ubiquitous tumor recurrence and patient death. To further understand the genetic mechanisms underlying the ability of glioma cells to migrate, we compared the matched transcriptional profiles of migratory and stationary populations of human glioma cells. Using a monolayer radial migration assay, motile and stationary cell populations from seven human long term glioma cell lines and three primary GBM cultures were isolated and prepared for expression analysis.ResultsGene expression signatures of stationary and migratory populations across all cell lines were identified using a pattern recognition approach that integrates a priori knowledge with expression data. Principal component analysis (PCA) revealed two discriminating patterns between migrating and stationary glioma cells: i) global down-regulation and ii) global up-regulation profiles that were used in a proband-based rule function implemented in GABRIEL to find subsets of genes having similar expression patterns. Genes with up-regulation pattern in migrating glioma cells were found to be overexpressed in 75% of human GBM biopsy specimens compared to normal brain. A 22 gene signature capable of classifying glioma cultures based on their migration rate was developed. Fidelity of this discovery algorithm was assessed by validation of the invasion candidate gene, connective tissue growth factor (CTGF). siRNA mediated knockdown yielded reduced in vitro migration and ex vivo invasion; immunohistochemistry on glioma invasion tissue microarray confirmed up-regulation of CTGF in invasive glioma cells.ConclusionGene expression profiling of migratory glioma cells induced to disperse in vitro affords discovery of genomic signatures; selected candidates were validated clinically at the transcriptional and translational levels as well as through functional assays thereby underscoring the fidelity of the discovery algorithm.

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Differential expression of novH and CTGF in human glioma cell lines.

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Differential expression of novH and CTGF in human glioma cell lines.

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