Proliferation Of Human Glioma Cells Research Articles

The combination of novel low molecular weight inhibitors of RAF (LBT613) and target of rapamycin (RAD001) decreases glioma proliferation and invasion

Monotherapies have proven largely ineffective for the treatment of glioblastomas, suggesting that increased patient benefit may be achieved by combining therapies. Two protumorigenic pathways known to be active in glioblastoma include RAS/RAF/mitogen-activated protein kinase and phosphatidylinositol 3-kinase/AKT/target of rapamycin (TOR). We investigated the efficacy of a combination of novel low molecular weight inhibitors LBT613 and RAD001 (everolimus), which were designed to target RAF and TOR, respectively. LBT613 decreased phosphorylation of extracellular signal-regulated kinase 1 and 2, downstream effectors of RAF, in a human glioma cell line. RAD001 resulted in decreased phosphorylation of the TOR effector S6. To determine if targeting RAF and TOR activities could result in decreased protumorigenic glioma cellular behaviors, we evaluated the abilities of LBT613 and RAD001 to affect the proliferation, migration, and invasion of human glioma cells. Treatment with either LBT613 or RAD001 alone significantly decreased the proliferation of multiple human glioma cell lines. Furthermore, LBT613 and RAD001 in combination synergized to decrease glioma cell proliferation in association with G(1) cell cycle arrest. Glioma invasion is a critical contributor to tumor malignancy. The combination of LBT613 and RAD001 inhibited the invasion of human glioma cells through Matrigel to a greater degree than treatment with either drug alone. These data suggest that the combination of LBT613 and RAD001 reduces glioma cell proliferation and invasion and support examination of the combination of RAF and TOR inhibitors for the treatment of human glioblastoma patients.

Role of Wnt5a in the proliferation of human glioblastoma cells

Wnt5a operates as either a tumor suppressor or a tumor stimulator, according to tumor type. The functions of Wnt5a in human glioblastoma (GBM) have yet to be determined. We initially evaluated the expression of Wnt5a in human glioma. The results of immunohistochemical analyses have revealed that Wnt5a expression was higher in human GBM than in normal brain tissue and low-grade astrocytoma. In order to assess the role of Wnt5a on proliferation in human glioblastoma cells, we employed U87MG and GBM-05, a newly established GBM cell line. GBM-05 was established from a patient diagnosed with GBM. GBM-05 cells were shown to express Nestin, but did not express GFAP and Map2ab. GBM-05 cells formed infiltrating brain tumors after being intracerebrally transplanted into nude mice, and xenotransplanted GBM-05 cells were observed to differentiate into neuronal and astrocyte lineages. Wnt5a expression in the xenotransplanted tumors was higher than that detected in the surrounding brain tissues. The overexpression of Wnt5a increased the proliferation of GBM-05 and U87MG in vitro. By way of contrast, the downregulation of Wnt5a expression as the result of RNA interference reduced proliferation from GBM-05 and U87MG cells in vitro, and reduced tumorigenicity in vivo. Our data indicate that Wnt5a signaling is an important regulator in the proliferation of human glioma cells.

Thrombin-induced cell proliferation and platelet-derived growth factor-AB release from A172 human glioblastoma cells.

In a previous study, we found that thrombin induced proliferation of TM-1 and T98G human glioma cells and that the mitogenic effect was abolished by hirudin. We investigated thrombin's effects on the proliferation of A172 human glioblastoma cells and the induction of growth factors. Furthermore, we examined whether or not the expression of heparin cofactor II (HCII) in A172 cells using adenovirus vector could suppress thrombin's effects. The effect of thrombin on cell proliferation was assessed using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide assay. The amount of growth factors in the conditioned medium was measured by enzyme-linked immunosorbent assay. The level of platelet-derived growth factor (PDGF)-B mRNA was assessed by reverse transcriptase-polymerase chain reaction analysis. Thrombin-induced proliferation of A172 cells primarily depended on the enhanced secretion of PDGF-AB by thrombin. The action of thrombin depended on its proteolytic activity. However, thrombin-induced PDGF-AB secretion was not abolished by anti-protease-activated receptor (PAR) antibody. The PAR-1 agonist peptide had no effect on cell growth and PDGF-AB levels. Thrombin did not increase PDGF-B gene expression. Expression of HCII effectively suppressed thrombin-induced PDGF-AB release. These results indicate that thrombin may play an important role in the proliferation of A172 cells by inducing PDGF-AB secretion and that thrombin's action is mediated by its proteolytic activity. Inhibition of thrombin's proteolytic activity may be a new therapeutic method for gliomas.

Differential expression and role of p21cip/waf1 and p27kip1 in TNF-α-induced inhibition of proliferation in human glioma cells

BackgroundThe role of TNF-α in affecting the fate of tumors is controversial, while some studies have reported apoptotic or necrotic effects of TNF-α, others provide evidence that endogenous TNF-α promotes growth and development of tumors. Understanding the mechanism(s) of TNF-α mediated growth arrest will be important in unraveling the contribution of tissue associated macrophages in tumor resistance. The aim of this study was to investigate the role of Cyclin Dependent Kinase Inhibitors (CDKI) – p21cip/waf1 and p27kip1 in TNF-α mediated responses in context with p53 and activation of NF-κB and Akt pathways. The study was done with human glioma cell lines -LN-18 and LN-229 cells, using monolayer cultures and Multicellular Spheroids (MCS) as in vitro models.ResultsTNF-α induced inhibition of proliferation and enhanced the expression of p21cip/waf1 and p27kip1 in LN-18 cells. p21 was induced on exposure to TNF-α, localized exclusively in the nucleus and functioned as an inhibitor of cell cycle but not as an antiapoptotic protein. In contrast, p27 was constitutively expressed, localized predominantly in the cytoplasm and was not involved in arrest of proliferation. Our data using IκBα mutant LN-18 cells and PI3K/Akt inhibitor-LY294002 revealed that the expression of p21 is regulated by NF-κB. Loss of IκBα function in LN-229 cells (p53 positive) did not influence TNF-α induced accumulation of pp53 (Ser-20 p53) suggesting that p53 was not down stream of NF-κB. Spheroidogenesis enhanced p27 expression and p21 induced by TNF-α was significantly increased in the MCS compared to monolayers.ConclusionThis study demarcates the functional roles for CDKIs-p21cip/waf1 and p27kip1 during TNF-α stimulated responses in LN-18 glioma cells. Our findings provide evidence that TNF-α-induced p21 might be regulated by NF-κB or p53 independently. p21 functions as an inhibitor of cell proliferation and does not have a direct role in rendering the cells resistant to TNF-α mediated cytotoxicity.

Evaluation of combination gene therapy with PTEN and antisense hTERT for malignant glioma in vitro and xenografts.

Telomerase activation is a critical event in cell immortalization, and an increase in human telomerase reverse transcriptase (hTERT) expression is the key step in activating telomerase. The phosphatase and tensin homolog (PTEN) gene encodes a double-specific phosphatase that induces cell cycle arrest, inhibits cell growth, and causes apoptotic cell death. Here, we evaluated a combined PTEN and antisense hTERT gene therapy for experimental glioma in vitro and in vivo. We demonstrated that infection with antisense-hTERT and wild-type-PTEN adenoviruses significantly inhibited human U251 glioma cell proliferation in vitro and glioma growth in a xenograft mouse model. The efficacy of therapy was obviously higher in the tumor xenografts infected with both PTEN and antisense hTERT than in the gliomas infected with either agent alone at the same total viral dose. Consistent with these results, we showed that telomerase activity and hTERT protein levels were markedly reduced in the glioma cells following adenovirus infection. In contrast, the levels of PTEN protein expression were dramatically increased in these cells. Our data indicate that combination treatment with antisense hTERT and wild-type PTEN effectively suppresses the malignant growth of human glioma cells in vitro and in tumor xenografts, suggesting a promising new approach in glioma gene therapy that warrants further investigation.

13-cis Retinoic Acid Induces Apoptosis and Cell Cycle Arrest in Human SEB-1 Sebocytes

Isotretinoin (13-cis retinoic acid (13-cis RA)) is the most potent inhibitor of sebum production, a key component in the pathophysiology of acne, yet its mechanism of action remains largely unknown. The effects of 13-cis RA, 9-cis retinoic acid (9-cis RA), and all-trans retinoic acid (ATRA) on cell proliferation, apoptosis, and cell cycle proteins were examined in SEB-1 sebocytes and keratinocytes. 13-cis RA causes significant dose-dependent and time-dependent decreases in viable SEB-1 sebocytes. A portion of this decrease can be attributed to cell cycle arrest as evidenced by decreased DNA synthesis, increased p21 protein expression, and decreased cyclin D1. Although not previously demonstrated in sebocytes, we report that 13-cis RA induces apoptosis in SEB-1 sebocytes as shown by increased Annexin V-FITC staining, increased TUNEL staining, and increased cleaved caspase 3 protein. Furthermore, the ability of 13-cis RA to induce apoptosis cannot be recapitulated by 9-cis RA or ATRA, and it is not inhibited by the presence of a retinoid acid receptor (RAR) pan-antagonist AGN 193109. Taken together these data indicate that 13-cis RA causes cell cycle arrest and induces apoptosis in SEB-1 sebocytes by a RAR-independent mechanism, which contributes to its sebosuppressive effect and the resolution of acne.

Coadministration of Sorafenib with Rottlerin Potently Inhibits Cell Proliferation and Migration in Human Malignant Glioma Cells

Mitogen-activated protein kinase (MAPK) and protein kinase C (PKC) are activated in the majority of gliomas and contribute to tumor cell growth and survival. Sorafenib (Bay43-9006; Nexavar) is a dual-action Raf and vascular endothelial growth factor receptor inhibitor that blocks receptor phosphorylation and MAPK-mediated signaling and inhibits growth in a number of tumor types. Because our initial studies of this agent in a series of glioma cell lines showed only partial growth inhibition at clinically achievable concentrations, we questioned whether inhibition of PKC signaling using the PKC-delta inhibitor rottlerin might potentiate therapeutic efficacy. Proliferation assays, apoptosis induction studies, and Western immunoblot analysis were conducted in cells treated with sorafenib and rottlerin as single agents or in combination. Sorafenib and rottlerin reduced proliferation in all cell lines when used as single agents, and the combination produced marked potentiation of growth inhibition. Flow-cytometric measurements of cells stained with Annexin V-propidium iodide and immunocytochemical assessment of cytochrome c and apoptosis-inducing factor release demonstrated that addition of rottlerin resulted in significantly higher levels of apoptosis than sorafenib alone. In addition, the combination of sorafenib and rottlerin reduced or completely inhibited the phosphorylation of extracellular signal-regulated kinase and Akt and down-regulated cell cycle regulatory proteins such as cyclin-D1, cyclin-D3, cyclin-dependent kinase (cdk)4, and cdk6 in a dose- and time-dependent manner. Our results clearly indicate that inhibition of PKC-delta signaling enhances the antiproliferative effect of sorafenib in malignant human glioma cell lines and support the examination of combinations of signaling inhibitors in these tumors.

Antisense telomerase RNA inhibits the growth of human glioma cells in vitro and in vivo

Telomerase is implicated in the development of cellular immortality and oncogenesis. It has been shown that telomerase activity is considerably higher in the tissue of many different cancers than in normal tissue, and that the inhibition or downregulation of telomerase activity can prevent the malignant proliferation of tumor cells. Antisense oligonucleotides have been widely used in suppressing the expression of genes and, therefore, in the present research, we evaluated the effect of antisense human telomerase RNA (hTR) on glioma cell growth in vitro and in vivo. We showed that antisense hTR cDNA significantly inhibited TJ905 human glioma cell proliferation in vitro and tumor growth in vivo, as determined by MTT assay and by measuring the volume of glioma in nude mice. Consistent with these results, we found that telomerase activity and the mRNA levels of hTR and hTERT (human telomerase reverse transcriptase) expression were markedly decreased in tumor cells treated with antisense hTR cDNA, as assessed by TRAP (telomeric repeat amplification protocol) assay and RT-PCR (reverse transcription-polymerase chain reaction) analysis. Our study conclusively demonstrates that antisense hTR effectively inhibits the growth of human glioma cells in vitro and in vivo and, thus, may be potentially used for gene therapy of malignant gliomas and other cancers.

The effect of γ-tocopherol on proliferation, integrin expression, adhesion, and migration of human glioma cells

The effect of vitamin E on proliferation, integrin expression, adhesion, and migration in human glioma cells has been studied. γ-tocopherol at 50 μM concentration exerted more inhibitory effect than α-tocopherol at the same concentration on glioma cell proliferation. Integrin α5 and β1 protein levels were increased upon both α- and γ-tocopherol treatments. In parallel, an increase in the α5β1 heterodimer cell surface expression was observed. The tocopherols inhibited glioma cell-binding to fibronectin where γ-tocopherol treatment induced glioma cell migration. Taken together, the data reported here are consistent with the notion that the inhibition of glioma cell proliferation induced by tocopherols may be mediated, at least in part, by an increase in integrin α5 and β1 expression. Cell adhesion is also negatively affected by tocopherols, despite a small increase in the surface appearance of the α5β1 heterodimer. Cell migration is stimulated by γ-tocopherol. It is concluded that α5 and β1 integrin expression and surface appearance are not sufficient to explain all the observations and that other integrins or in general other factors may be associated with these events.

Nonsteroidal anti-inflammatory drugs inhibit the growth of C6 and U138-MG glioma cell lines

Nonsteroidal anti-inflammatory drugs (NSAIDs) are widely used drugs for the treatment of inflammatory disease and have a chemopreventive effect in a variety of tumors. Several studies have demonstrated unequivocally that certain NSAIDs cause antiproliferative effects independent of cyclooxygenase (COX) activity. In this study, we investigated the effect of chemically unrelated NSAIDs in the proliferation of glioma cell lines and the possible mechanisms involved in indomethacin-mediated inhibition of proliferation in glioma cells lines. The glioma cell lines were treated with NSAIDs and proliferation was measured by cell counting. Indomethacin, acetaminophen, sulindac sulfide and NS-398 ( N-[2-cyclohexyloxy)-4-nitrophenyl]methane-sulfonamide) induced a time- and concentration-dependent inhibition of C6 rat glioma cell proliferation. The inhibition of COX by chemically unrelated NSAIDs leads to inhibition of rat and human glioma cell proliferation. The tetrazolium reduction assay (MTT) indicated a reduction in cell viability induced by indomethacin. None of the NSAIDs tested induced caspase-3/7 activation, assayed with a fluorigenic substrate. The indomethacin-induced inhibition of C6 cells proliferation was abrogated by the use of the c-Src inhibitor, PP2 and the MEK inhibitor, PD 098059, suggesting COX-independent mechanisms. Indomethacin decreased the percentage of cells in the S phase, with relative increases in the G 0/G 1 and/or the G 2/M phase. NSAIDs may be clinically important for pharmacological intervention in gliomas.

Prospect for research on deregulation of cell proliferation and apoptosis in human gliomas

胶质瘤占原发性脑肿瘤的60%～70%,且多数呈浸润性生长,手术不易全切.化疗和放疗既不能高度特异地杀伤胶质瘤细胞,又可能产生中枢神经系统,乃至全身不能耐受的毒副作用,治疗效果差.在全身肿瘤中,恶性胶质瘤5年病死率仅次于胰腺癌和肺癌居第3位,5年生存率不足5%.近20年中,胶质瘤的疗效和患者预后没有明显改善.故寻求有效的治疗措施势在必行,但阐明胶质瘤的发病机制是实现该目标必须解决的关键问题.已知无限增殖和凋亡抑制是胶质瘤发生、发展的根本原因[1],而对其细胞增殖及凋亡调控紊乱认识的不断深入是近10年来胶质瘤研究的重要进展之一,为进一步深入探讨胶质瘤发病的分子机制提供了重要启示。

Human U251 Glioma Cell Proliferation Is Suppressed by HET0016 [N-Hydroxy-N′-(4-butyl-2-methylphenyl)formamidine], a Selective Inhibitor of CYP4A

We have previously reported that HET0016 [N-hydroxy-N'-(4-butyl-2 methylphenyl)formamidine], a selective inhibitor of CYP4A and thus 20-HETE (20-hydroxyeicosatetraenoic acid) synthesis, inhibits endothelial cell proliferation and decreases angiogenesis induced by human glioma cell U251. A stable 20-HETE agonist, WIT003 [20-hydroxyeicosa-5(Z),14(Z)-dienoic acid (1 microM)], increased U251 cell proliferation from 3.9- to 4.8-folds from T(0) (time of the treatment). We examined the effects of HET0016 on the growth of U251. HET0016 inhibited U251 basal cell proliferation in a dose-dependent manner. 10 microM HET0016 suppressed 56% of U251 proliferation and significantly increased the proportions of the cells arrested in the G(0)/G(1) phase of the cell cycle. Exposure to HET0016 (as early as 4 h) reduced protein tyrosine and p42/p44 MAPK (mitogen-activated protein kinase) phosphorylation. Furthermore, HET0016 significantly inhibited the U251 proliferation and phosphorylation of both the epidermal growth factor (EGF) receptor and p42/p44 MAPK induced by EGF. CYP4A mRNA and proteins were both present in U251. This suggests that HET0016 inhibited U251 proliferation by inhibiting 20-HETE synthesis. However, U251 did not synthesize 20-HETE in the presence of arachidonic acid. This implies that HET0016 suppresses U251 proliferation by mechanisms that are not yet clear but may involve activities other than inhibition of 20-HETE synthesis. We concluded that HET0016 may be the prototype of novel compounds that suppress human glioma cell proliferation.

SDF-1 and CXCR4 are up-regulated by VEGF and contribute to glioma cell invasion

Glioma cells produce vascular endothelial growth factor (VEGF) to induce vascularization and thereby supply the malignant tissue with oxygen and nutrients. However, little is known about the direct effects of VEGF on tumor cells. In this study, we investigate the ability of VEGF to promote proliferation and invasion of human glioma cells (U251n). Since the chemokine and its receptor, SDF-1/CXCR4, promote glioma cell proliferation and are up-regulated in human glioblastomas, we also tested the effects of VEGF on SDF-1 and CXCR4 mRNA expression. Using cell culture, the effect of VEGF on proliferation of U251n cells was measured using ELISA to detect incorporated BrdU as a marker of DNA syntheses. The effects of VEGF and SDF-1 on U251n cell invasion and proliferation were measured using inhibitors to VEGF receptor1 and receptor2, DC101 and MF1, respectively, and a CXCR4 antagonist (AMD3100). SDF-1 and CXCR4 mRNA expression in U251n and U87MG cells were measured using quantitative PCR. VEGF antisense phosphorothioate oligodeoxynucleotide (AS-VEGF) was also used to down-regulate VEGF expression in U251n cells. VEGF significantly increased U251n cell proliferation and invasion in a dose-dependent manner. These effects were blocked by the VEGF receptor inhibitors, DC101/MF1. The CXCR4 antagonist AMD3100 blocked U251n increased invasion, but not proliferation. CXCR4 and SDF-1 mRNA were up-regulated when U251n and U87MG cells were treated with VEGF. Eight micrometer VEGF antisense phosphorothioate oligodeoxynucleotide (AS-VEGF) down-regulated CXCR4 and SDF-1 mRNA levels in U251n cells. VEGF has a direct effect on U251n glioma cell proliferation and invasion. VEGF up-regulates SDF-1 and CXCR4 mRNA expression, and contributes to U251n cell invasion.

The effect of cyclin D expression on cell proliferation in human gliomas

The expression of three cyclin D subtypes was defined immunohistochemically with cyclin D1, D2 and D3 monoclonal antibodies in 52 human glioma biopsies and eight control samples of normal brain tissue. PCNA labeling indices (LI) were used to evaluate proliferation in the glioma biopsies. LI of cyclin D1, D2 and D3 were compared with histological grade and the proliferating cell nuclear antigen (PCNA) LI. Cyclin D1 expression only was observed in normal brain tissue, but marked overexpression of cyclin D1 and cyclin D3 was observed in glioma. Cyclin D1 LI increased with malignancy, in parallel with an increase in PCNA LI. Lower expression of cyclin D2 was found in a small fraction of the gliomas, but its LI did not vary significantly with grade. Cyclin D3 was mainly expressed by malignant gliomas and was rarely observed in low-grade glioma. Cyclin D2 and D3 expression correlated with PCNA LI, but not as strongly as for cyclin D1. Expression of cyclin D1 is closely related to both the oncogenesis and progression of glioma, while cyclin D3 is associated with transformation to a malignant phenotype. Cyclin D2 is weakly expressed and shows no marked relationship with any aspect of tumorigenesis. The exact contribution of cyclin D subtypes to cell cycle progression in neoplastic and reactive cells remains to be defined.

Suppression of proliferation of human U-87MG glioma cells by overexpression of the AMPA receptor subunit GluR2

Suppression of proliferation of human U-87MG glioma cells by overexpression of the AMPA receptor subunit GluR2

Mda-7 (IL-24) Inhibits Growth and Enhances Radiosensitivity of Glioma Cells In Vitro via JNK Signaling

Despite therapeutic interventions including surgery, chemotherapy and radiotherapy, glioblastoma multiforme (GBM) has a very poor prognosis and novel therapies are required. MDA-7 (IL-24), when expressed via a recombinant replication defective adenovirus, Ad.mda-7, has profound anti-proliferative and cytotoxic effects in a variety of tumor cells, but not in non-transformed cells. The present studies examined the combined impact of Ad.mda-7 and ionizing radiation on the proliferation and survival of GBM cells. Ad.mda-7 reduced the proliferation of rodent and human glioma cells in MTT assays and in colony formation assays. The anti-proliferative effects of Admda-7 were enhanced by radiation in a greater than additive fashion. In vitro, this cellular change correlated with enhanced cell numbers in G1/G0 and G2/M phases of the cell cycle, implying Ad.mda-7 radiosensitizes tumor cells in a cell cycle-independent manner. The radiosensitizing effects were not observed in cultures of non-transformed primary astrocytes. The enhanced reduction in growth correlated with increased necrosis and DNA degradation. Ad.mda-7 enhanced p38 and ERK1/2 activity but did not alter JNK or Akt activity. Irradiation of cells expressing MDA-7 suppressed ERK1/2 activity and dramatically enhanced JNK1/2 activity without altering either Akt or p38 activity. Inhibition of JNK1/2, but not p38, signaling abolished the radiosensitizing properties of MDA-7. Inhibition of neither ERK1/2 nor PI3K signaling enhanced the anti-proliferative effects of Ad.mda-7, whereas combined inhibition of both pathways enhanced cell killing, suggesting that ERK and PI3K signaling can be protective against MDA-7 lethality.

Altered extracellular ATP, ADP and AMP catabolism in glioma cell lines.

In order to characterize the enzymes involved in the purine nucleotide catabolism as indicators of invasiveness and aggressiveness of malignant gliomas, the degradation of extracellular nucleotides by five different glioma cell lines was investigated and compared with primary astrocytes. Rapid hydrolysis of extracellular ATP and ADP by astrocytes was observed, whereas all glioma cell lines examined presented low rates of ATP hydrolysis. In contrast, ecto-5'-nucleotidase activity was increased in glioma cell lines when compared to astrocytes. Considering that ATP is recognized as a mitogenic factor that induces proliferation in human glioma cells, the substantial decrease in ATP and ADP hydrolysis observed in gliomas leads us to suggest that alterations in the ecto-nucleotidases pathway may represent an important mechanism associated with malignant transformation of glioma cell lines.

Migration, proliferation, and invasion of human glioma cells following treatment with simvastatin.

This study describes the migration, proliferation, and invasion behaviour of two human glioma cell lines, GaMg and U-87 Mg, grown as multicellular tumour spheroids after 3-hydroxy-3-methylglutaryl coenzyme A (HMGCoA) reductase inhibitor (simvastatin) therapy. Migration and proliferation studies were performed using simvastatin in concentrations of 0.2-30 microg/ml(-1). A coculture system in which tumour spheroids were confronted with foetal rat brain aggregates was used for invasion studies. A dose-dependent growth and migratory inhibitory response to simvastatin treatment was observed. Marked invasion of the glioma spheroids into the brain aggregates could be seen in both treated and nontreated groups. Simvastatin therapy inhibits tumour cell growth and migration, but the invasiveness of the remaining tumour cells seems to be unaffected.

Proliferation, migration, and invasion of human glioma cells exposed to fractionated radiotherapy in vitro.

Radiotherapy is a well established treatment for malignant gliomas. This study describes the migration, proliferation, and invasion behaviour of two human glioma cell lines (GaMg and U-87 Mg) grown as multicellular tumour spheroids after radiotherapy. Migration and proliferation studies were performed using conventional and accelerated fractionation up to 60 Gy and 59.4 Gy, respectively. A dose-dependent growth and migratory response to irradiation independent of the type of fractionation was observed. A coculture system in which tumour spheroids were confronted with foetal rat brain aggregates was used for invasion studies. Marked invasion of the glioma spheroids into the brain aggregates occurred with or without radiotherapy. For the GaMg cells, flow cytometric DNA histograms after treatment with 10 Gy and 40 Gy showed an accumulation of cells in the G2/M phase of the cell cycle. Radiotherapy inhibits tumour cell growth and migration, but the invasiveness of the remaining tumour cells seems to be unaffected.

Inhibition of autocrine fibroblast growth factor signaling by the adenovirus-mediated expression of an antisense transgene or a dominant negative receptor in human glioma cells in vitro

Autocrine fibroblast growth factor (FGF) signaling was genetically manipulated in human gliomas by an adenovirus-mediated strategy. Antisense inhibition of endogenous basic FGF (bFGF) expression showed inhibition of the proliferation of human glioma cells (U251MG, NMC-G1) without promoting apoptosis. The reduction in proliferation in response to antisense inhibition was reversed with an additional supplement of exogenous bFGF. On the other hand, the overexpression of a kinase defective mutant of the type I FGF receptor (dominant negative FGF receptor) resulted in not only growth inhibition but also marked apoptosis in U251MG cells. In the cells expressing a dominant negative FGF receptor, the reduction in cell number was not reversed by the exogenous bFGF supplement. Our data further implicated the significance of an autocrine FGF signaling loop in human gliomas. Cell survival activity may largely depend on the receptor-mediated pathway, which should be considered in the development of the molecular based therapeutics.