Search PubMed⌕ Search

Biomedical subjects

M Weller

Publications and source records attributed to M Weller.

At least 145 records · Page 8Linked to original sources
PubMed1999

p53 enhances BAK and CD95 expression in human malignant glioma cells but does not enhance CD95L-induced apoptosis.

The temperature-sensitive murine p53val135 mutant was introduced into 3 human malignant glioma cell lines to examine the effects of the p53 status on BCL-2 family protein expression, CD95 expression, and sensitivity to CD95 ligand (CD95L)-induced apoptosis. p53val135 behaves as a dominant negative mutant at 38.5 degrees C but assumes p53 wild-type properties. In order to dissect (i) specific effects of wild-type versus mutant p53, and (ii) transdominant-negative versus gain-of-function effects of mutant p53, we included glioma cell lines with functional wild-type (LN-229), mutant (LN-18) or deleted (LN-308) p53 genes. Wild-type, but not mutant, p53val135 promoted G2/M arrest and accumulation of BAK protein in all cell lines. The levels of other BCL-2 family members including BAX, BCL-2, BCL-X or MCL-1 were not consistently modulated by mutant or wild-type p53val135. Wild-type, but not mutant, p53val135 enhanced CD95 expression in all cell lines. CD95L-evoked caspase 3 activity was unaffected by wild-type p53 in all cell lines. Unexpectedly, mutant p53val135 differentially modulated caspase 3 activity in a gain-of-function fashion in that caspase 3 activity induced by CD95L was enhanced in LN-229 and LN-308 cells but reduced in LN-18 cells. Yet, mutant p53val135 enhanced the sensitivity to CD95L in LN-18 cells, had no effect in LN-229 cells, and decreased the sensitivity of LN-308 cells. Corresponding to the unaltered CD95L-evoked caspase 3 activity, wild-type p53val135 had no major effect on CD95L-induced apoptosis, except for a moderate sensitization of LN-229 cells but only when protein synthesis was inhibited. Thus, wild-type p53 induces BAK and CD95 expression in human glioma cells without enhancing their susceptibility to CD95-mediated apoptosis, and mutant p53 modulates CD95L-evoked apoptotic signalling in a gain-of-function fashion up-stream and down-stream of caspase 3 activation.

Animals↗
PubMed1999

Proteasome inhibitors induce p53/p21-independent apoptosis in human glioma cells.

The proteasome is a multiprotein complex involved in the degradation of ubiquitinated proteins. Three proteasome inhibitors, calpain inhibitor I, lactacystin and MG132, induced apoptosis in several human malignant glioma cell lines. Although proteasome inhibitors induced p53 accumulation in a cell line retaining wild-type p53 activity, p53 activity was dispensable for apoptosis since transdominant-negative p53 abrogated p53-dependent p21 induction but did not modulate apoptosis. Further, p21 was induced by higher concentrations of proteasome inhibitors in a p53-independent manner both in p53 wild-type and in p53 mutant cell lines. Although there was a strong G2/M arrest in response to proteasome inhibition in glioma cells, this G2/M arrest was also observed in p21(-/-) colon carcinoma cells, suggesting that p21 is dispensable for the G2/M arrest associated with proteasome inhibition. Interestingly, the p21(-/-) cells were more resistant to protease inhibitors than parental p21(+/+) cells. In summary, our data indicate that proteasome inhibition induces a p21-independent G2/M arrest and p53-independent apoptosis in human malignant glioma cells.

Acetylcysteine↗
PubMed1999

Characterization of Tu-2449, a glioma cell line derived from a spontaneous tumor in GFAP-v-src-transgenic mice: comparison with established murine glioma cell lines.

Glial fibrillary acidic protein (GFAP)-v-src transgenic mice develop spontaneous gliomas with a high incidence of malignant progression. We characterize the first glial cell line derived from v-src transgenic mice, Tu-2449 in comparison with a virally induced murine glioma, SRB-10, and a spontaneous murine glioma, P497. Doubling times were lowest, as clonogenicity in soft agar was highest for Tu-2449, and to a lesser degree, Tu-2449 cells formed spheroids and showed migratory behaviour and invaded fetal rat brain aggregates. BCL-2 and BAX expression were lower in Tu-2449 and P497 than in SRB-10 cells. Only Tu-2449 cells accumulated p53 protein in response to genotoxic stress. Tu-2449 and SRB-10 cells that showed low CD95 expression were resistant to CD95 ligand (CD95L)-induced apoptosis unless coexposed to CD95L and inhibitors of RNA or protein synthesis. A chemosensitivity profile revealed Tu-2449 to be rather chemoresistant. Tu-2449 thus displays growth characteristics and patterns of resistance to apoptosis similar to those of other mouse and human glioma cell lines and may therefore become a valuable tool to evaluate new therapies for malignant gliomas in vitro and in vivo.

Animals↗
PubMed1999

Betulinic acid-induced apoptosis in glioma cells: A sequential requirement for new protein synthesis, formation of reactive oxygen species, and caspase processing.

Betulinic acid (BA), a pentacyclic triterpene, is an experimental cytotoxic agent for malignant melanoma. Here, we show that BA triggers apoptosis in five human glioma cell lines. BA-induced apoptosis requires new protein, but not RNA, synthesis, is independent of p53, and results in p21 protein accumulation in the absence of a cell cycle arrest. BA-induced apoptosis involves the activation of caspases that cleave poly(ADP ribose)polymerase. Interactions of death ligand/receptor pairs of the CD95/CD95 ligand family do not mediate BA-induced caspase activation. BA enhances the levels of BAX and BCL-2 proteins but does not alter the levels of BCL-xS or BCL-xL. Ectopic expression of BCL-2 prevents BA-induced caspase activation, DNA fragmentation, and cell death. Furthermore, BA induces the formation of reactive oxygen species that are essential for BA-triggered cell death. The generation of reactive oxygen species is blocked by BCL-2 and requires new protein synthesis but is unaffected by caspase inhibitors, suggesting that BA toxicity sequentially involves new protein synthesis, formation of reactive oxygen species, and activation of crm-A-insensitive caspases.

Antineoplastic Agents, Phytogenic↗
PubMed1998

Predicting chemoresistance in human malignant glioma cells: the role of molecular genetic analyses.

Less than 30% of malignant gliomas respond to adjuvant chemotherapy. Here, we asked whether alterations in the p53 and RB pathways and the expression of six BCL-2 family proteins predicted acute cytotoxicity and clonogenic cell death induced by BCNU, vincristine, cytarabine, teniposide, doxorubicin, camptothecin or beta-lapachone in 12 human malignant glioma cell lines. Neither wild-type p53 status, nor p53 protein accumulation, nor p21 or MDM-2 levels, nor differential expression of BCL-2 family proteins predicted drug sensitivity, except for an association of BAX with higher beta-lapachone sensitivity in acute cytotoxicity assays. p16 protein expression was associated with high doubling time and chemoresistance. We conclude that some important molecular changes, which are involved in the development of gliomas and attributed a role in regulating vulnerability to apoptosis, may not determine the response to chemotherapy in these tumors.

Antineoplastic Agents↗
PubMed1998

BCL-2 promotes migration and invasiveness of human glioma cells.

Malignant progression in gliomas is correlated with increased migratory capacity which involves metalloproteolytic activity. Here, we report that ectopic expression of BCL-2 in two malignant glioma sublines markedly promoted glioma cell migration from spheroids and invasion into Matrigel-coated membranes. Invasion of fetal rat-brain aggregates was enhanced by BCL-2. Zymography revealed activation of matrix metalloproteinase-2 (MMP-2) in BCL-2-expressing cells. BCL-2 expressing cells showed an increase in MMP-2/-3/-12 (LN-18), and MMP-9/-12 and cell surface urokinase-type plasminogen activator (u-PA) (LN-229) mRNA and a reduction in tissue inhibitors of metalloproteinases (TIMP)-2 mRNA (LN-229). Taken together, we propose a novel function for BCL-2 in the malignant phenotype of glioma cells, that is, to enhance migration and invasion by altering the expression of a set of metalloproteinases and their inhibitors.

Animals↗
PubMed1998

Synthesis and biological effects of NO in malignant glioma cells: modulation by cytokines including CD95L and TGF-beta, dexamethasone, and p53 gene transfer.

Nitric oxide (NO) is thought to play an important role in neurotransmission, inflammation, and regulation of cell death in the mammalian brain. Here, we examined the synthesis and biological effects of NO in human malignant glioma cells. Exposure to cytokines such as interferon (IFN)-gamma, tumor necrosis factor (TNF)-alpha or interleukin (IL)-1beta and lipopolysaccharide (LPS) induced NO synthesis in rat C6 and A172 human glioma cells, but not in LN-229, T98G or LN-18 human malignant glioma cells. Induced release of NO involved enhanced expression of inducible NO synthase (iNOS). Failure to detect NO release in the latter cell lines was not overcome by neutralization of endogenous TGF-beta or by coexposure to cytokines, LPS, and antioxidants. Apoptosis induced by CD95 ligand (CD95L) did not involve NO formation. Neither NOS inhibitors nor NO donators modulated CD95L-induced apoptosis. Dexamethasone (DEX)-mediated protection of glioma cells from CD95L-induced apoptosis was also independent of DEX effects on NO metabolism. DEX inhibited not only cytokine/LPS-evoked NO release but also attenuated the toxicity of NO in three of five cell lines. Forced expression of temperature-sensitive p53 val135 in C6 cells in either mutant or wild-type conformation inhibited cytokine/LPS-induced NO synthesis. Further, accumulation of p53 in both mutant or wild-type conformation protected glioma cells from the toxicity of exogenous NO, consistent with a gain of p53 function associated with p53 accumulation. We conclude that resistance to NO-dependent immune defense mechanisms may contribute to the malignant progression of human cancers with p53 alterations, notably those associated with the accumulation of mutant p53 protein.

Animals↗
PubMed1998

Dexamethasone-mediated protection from drug cytotoxicity: association with p21WAF1/CIP1 protein accumulation?

Dexamethasone (DEX)-mediated inhibition of drug-induced, but not CD95 ligand-induced, apoptosis in malignant glioma cells correlates with wild-type p53 status. Here, we examined mechanisms underlying DEX-mediated protection from apoptosis. DEX did not induce p53 expression in two p53 wild-type cell lines (U87MG, LN-229) and did not alter drug-induced p53 accumulation. Forced expression of temperature-sensitive p53val135 in mutant conformation failed to prevent accumulation of endogenous wild-type p53 but acted in a transdominant negative manner to inhibit p53-mediated, camptothecin-induced p21WAF1/CIP1 expression. p53val135-transfected cells retained responsiveness to DEX at restrictive temperature, suggesting that p53 activity is not required for cytoprotection. Forced expression of wild-type p53val135 abrogated the protective effect of DEX, suggesting redundant cytoprotective effects of DEX and p53. Indeed, DEX induced moderate accumulation of p21WAF1/CIP1 in U87MG, LN-229 and p53 mutant LN-18 cells, but not in p53 mutant LN-308 or T98G cells. LN-18 is also the p53 mutant cell line with the best cytoprotective response to DEX. p21WAF1/CIP1 accumulation occurred in the absence of changes in p21WAF1/CIP1 mRNA expression. Wild-type p53 was not required for this DEX effect since DEX induced p21WAF1/CIP1 accumulation in p53val135-transfected LN-229 cells, too. DEX failed to induce p21WAF1/CIP1 expression or cytoprotection in untransformed rat astrocytes. The same lack of modulation of p21WAF1/CIP1 expression and drug toxicity was observed in p21(+/+), p21(+/-) and p21(-/-) human colon carcinoma cells. Paradoxically, while only p21(+/+) and p21(+/-) mouse embryonic fibroblasts showed enhance p21WAF1/CIP1 levels after exposure to DEX, only p21(-/-) fibroblasts were protected from drug toxicity by DEX. The present study links DEX-mediated protection from cancer chemotherapy to a p53-independent pathway of regulating p21WAF1/CIP1 expression in glioma cells but this effect appears to cell type-specific.

Animals↗
PubMed1998

CD95-mediated apoptosis: no variation in cellular sensitivity during cell cycle progression.

Sensitivity of CD95-mediated apoptosis has been reported to vary during cell cycle progression (FEBS Lett. (1997) 412, 91-93). Here, we report that three human glioma cell lines with different p53 status (i) undergo growth arrest and synchronous cell cycle re-entry after prolonged serum deprivation, (ii) do not exhibit cell cycle-related changes in CD95 expression at the cell surface, and (iii) do not exhibit cell cycle-related changes in susceptibility to DC95 ligand-induced apoptosis. In contrast, cell cycle-specific activity was demonstrated for various cancer chemotherapy drugs. Further, CD95 expression and susceptibility to CD95 ligand-induced apoptosis does not vary during cell cycle progression of Jurkat T cells, HeLa cervical carcinoma and HepG2 hepatocellular carcinoma cells. These results do not support a role for the cell cycle phase as an important predictor of vulnerability to CD95-mediated apoptosis.

Antibiotics, Antineoplastic↗
PubMed1998

Chemotherapy primes malignant glioma cells for CD95 ligand-induced apoptosis up-stream of caspase 3 activation.

The cytotoxic cytokine, CD95 ligand, is an experimental anti-cancer agent. Here, we describe a novel pathway of drug-mediated augmentation of CD95 ligand-induced apoptosis. We report that prolonged pre-exposure of human malignant glioma cells to different cytotoxic agents, VM26, cytarabine and cisplatin, induces strong sensitization to CD95 ligand-induced apoptosis. CD95 gene transfer does not prevent sensitization, suggesting that sensitization is not mediated by drug-induced CD95 expression. Priming with cytotoxic drugs greatly increases CD95 ligand-induced caspase 3 activity, indicating that the cytotoxic drugs positively modulate the CD95-dependent signalling cascade up-stream of caspase 3 activation.

Antineoplastic Agents↗
PubMed1998

Interferon-alpha enhances CD95L-induced apoptosis of human malignant glioma cells.

CD95 ligand (CD95L)-induced apoptosis is a novel immunotherapeutic approach to malignant glioma. Here, we report that interferon-alpha (IFN-alpha) sensitizes LN-229 and T98G human malignant glioma cells to CD95L-induced apoptosis. In contrast to the effects of IFN-gamma and TNF-alpha which sensitize glioma cells to CD95 antibody-induced apoptosis in part by enhancing CD95 expression, IFN-alpha has no effect on CD95 expression at the cell surface of LN-229 and T98G cells. To confirm that changes in CD95 expression are not required for the effects of IFN-alpha, we show that IFN-alpha enhances CD95L-induced apoptosis even in CD95-transfected LN-308 glioma cells. These LN-308 cells have little endogenous CD95 expression but express high levels of CD95 from a stably integrated CD95 expression plasmid. The sensitizing effects of IFN-alpha appear to be independent of cell cycle effects of IFN-alpha and are unaffected by ectopic expression of the bcl-2 proto-oncogene. IFN-alpha enhances CD95L-induced activation of caspase-3, a critical mediator of CD95L-induced cell death. IFN-alpha also increases the cytotoxic effects of BCNU, teniposide and cytarabine in both cell lines, and of vincristine in LN-229 cells. Doxorubicin and 5-fluorouracil toxicity are unaffected by IFN-alpha. IFN-alpha may be a useful adjunct to novel strategies of immunochemotherapy for malignant gliomas that target CD95-mediated apoptosis.

Antineoplastic Agents, Alkylating↗
PubMed1998

CD95 ligand-induced apoptosis of human medulloblastoma cells.

CD95 ligand (CD95L) is a cytotoxic cytokine that induces apoptosis in susceptible target cells. Medulloblastoma is the most common non-glial intrinsic malignancy of the brain. In this study, we have studied CD95-mediated apoptosis of human medulloblastoma cell lines. We found that DAOY, MED-1 and D-283 cells are susceptible to CD95L-induced apoptosis when RNA and protein synthesis are inhibited. Preexposure of D-283, but not DAOY or MED-1 cells, to interferon-gamma or tumor necrosis factor-alpha enhances CD95 expression and primes these cells for CD95-mediated apoptosis. Inhibitors of interleukin 1-converting enzyme (ICE)-like protease (caspase) activity block CD95L-induced cytotoxicity, suggesting that caspases mediate the death signal induced by CD95L in human medulloblastoma cells. Interestingly, medulloblastoma cells belong to an increasing number of tumor cell types that coexpress CD95 and CD95L. We conclude that CD95 may be a promising target of immunochemotherapy for human medulloblastoma.

Animals↗
PubMed1998

Serum CD95 of relapsing remitting multiple sclerosis patients protects from CD95-mediated apoptosis.

Failure of CD95-mediated apoptosis as a potential negative regulatory mechanism of T cell expansion may be involved in T cell-mediated autoimmune diseases such as multiple sclerosis (MS). Recently, soluble CD95 has been shown to be elevated in MS patients with active disease. Here, we report that the sera of MS patients inhibit CD95 ligand-induced apoptosis of susceptible target cells in a concentration-dependent manner and dependent on the amount of serum CD95 levels. Thus, MS sera contain biologically active inhibitors of T cell apoptosis that may allow for prolonged abnormal immune responses.

Apoptosis↗
PubMed1998

APO2 ligand: a novel lethal weapon against malignant glioma?

APO2L (TRAIL) is a novel CD95L (Fas/APO-1-L) homologous cytotoxic cytokine that interacts with various receptors which transmit (DR4, DR5) or inhibit (DcR1, DcR2) an apoptotic signal. Here, we report that human glioma cell lines preferentially express mRNAs for agonistic death receptors DR4 (8/12) and DR5 (11/12) rather than the death-inhibitory decoy receptors DcR1 (4/12) and DcR2 (2/12). Ten of 12 cell lines are susceptible to APO2L-induced apoptosis. The resistant cell lines, U138MG and U373MG, are cross-resistant to CD95L-induced apoptosis. Similar to CD95L-induced apoptosis, APO2L-induced apoptosis is inhibited by ectopic expression of the caspase inhibitor, crm-A, or of bcl-2, or by coexposure to the corticosteroid, dexamethasone, or the lipoxygenase inhibitor, nordihydroguaretic acid. There is no correlation between p53 genetic status of the cell lines and their susceptibility to APO2L-induced apoptosis, but the latter is moderately enhanced by ectopic expression of wild-type p53. APO2L targeting may be a promising approach for selectively targeting apoptosis to human malignant glioma cells.

Apoptosis↗
PubMed1998

Endonucleolytic DNA fragmentation is not required for apoptosis of cultured rat cerebellar granule neurons.

Depolarizing concentrations of potassium (K+) promote maturation and survival of cerebellar granule neurons in vitro. Withdrawal of potassium from differentiated neurons induces morphological and biochemical features of apoptosis, including membrane blebbing, nuclear condensation, activation of caspases, and internucleosomal DNA fragmentation. Significant DNA fragmentation is detectable at 6 h after K+ withdrawal and slowly increases thereafter. Two observations indicate that endonucleolytic DNA degradation is neither required nor sufficient for K+ withdrawal-induced apoptosis in cerebellar granule neurons: (i) neurons are rescued from apoptosis by readdition of K+ up to 8 h after K+ withdrawal, when DNA fragmentation has already occurred. (ii) The endonuclease inhibitor, aurintricarboxylic acid, inhibits DNA fragmentation as assessed by quantitative DNA fluorometry, TUNEL staining, and DNA gel electrophoresis, but not cell death or chromatin condensation induced by K+ withdrawal.

Animals↗
PubMed1998

BCL-2 family protein expression in human malignant glioma: a clinical-pathological correlative study.

Malignant gliomas are rather refractory to current therapeutic approaches including surgery, radiotherapy, chemotherapy and immunotherapy. Acquired alterations in the pathways required for apoptotic cell death are thought to be responsible to the failure of glioma to respond to therapy. Here we have examined the expression of several proteins involved in the susceptibility to apoptosis in 20 human gliomas, including the BCL-2 family proteins BCL-2, BCL-X, BAX and MCL-1, as well as p53 and RB. Most gliomas expressed several BCL-2 family proteins. There was good correlation between expression of the functional antagonists, BCL-2/BCL-X and BAX, suggesting that changes in the BCL-2+BCL-X/BAX ratio are not responsible for the differential response of glioma patients to chemotherapy. The immunochemistry data were also analysed in regard to response to therapy and clinical outcome. All patients had cytoreductive surgery and received radiotherapy and nitrosourea-based adjuvant chemotherapy. There was no prominent association of outcome with the expression patterns of p53, RB, BCL-2, BCL-X or BAX. We find, however, that expression of the MCL-1 protein is associated with early tumour recurrence and shorter survival in this group of glioma patients. This preliminary observation will have to be confirmed in a larger independent sample of glioma patients.

Adolescent↗