Enhancement of macrophage cytotoxicity against murine gliomas by interferon beta increase in nitric oxide production in response to glioma-derived soluble factors

2002 ◽  
Vol 97 (3) ◽  
pp. 619-626 ◽  
Author(s):  
Tomohiro Kito ◽  
Etsushi Kuroda ◽  
Akira Yokota ◽  
Uki Yamashita
Keyword(s):  
Nitric Oxide ◽  
Cytotoxic Activity ◽  
Glioma Cells ◽  
Cell Contact ◽  
No Production ◽  
Soluble Factors ◽  
Content Type ◽  
Culture Supernatants ◽  
Fine Print

Object. In previous studies interferon-β (IFNβ) has been shown to suppress tumor growth. In this report, the antitumor effect of macrophages stimulated with IFNβ is investigated in murine gliomas in vitro. Methods. The authors examined the cytotoxic activity of IFNβ-stimulated peritoneal macrophages in glioma cells labeled with [3H]thymidine. The addition of IFNβ enhanced cytotoxic activity in gliomas as well as the nitric oxide (NO) production of macrophages in cocultures. Addition of NG-monomethyl-l-arginine (l-NMMA) and l-N6-(1-iminoethyl)-lysine, but not d-NMMA (an inactive analog of l-NMMA), blocked this cytotoxic activity. The addition of IFNβ had no direct effect on the growth of glioma cells. Because NO was not produced from macrophages treated with IFNβ alone and IFNβ-induced cytotoxic activity did not need cell-to-cell contact, the authors suspected that gliomas produce some soluble factors that act as cofactors for IFNβ-induced cytotoxic activity. Macrophages stimulated with IFNβ in the presence of glioma culture supernatants showed higher cytotoxicity against glioma cells than macrophages stimulated with IFNβ alone. Furthermore, NO was markedly produced by IFNβ-stimulated macrophages in the presence of glial culture supernatants. Conclusions. These data indicate that the antiglioma activity of IFNβ through macrophages is due to NO produced by macrophages and that glioma-derived soluble factors play a role as an essential cofactor in this activity.

Cytotoxicity in glioma cells due to interleukin-12 and interleukin-18—stimulated macrophages mediated by interferon-γ regulated by nitric oxide

2003 ◽  
Vol 98 (2) ◽  
pp. 385-392 ◽  
Author(s):  
Tomohiro Kito ◽  
Etsushi Kuroda ◽  
Akira Yokota ◽  
Uki Yamashita
Keyword(s):  
Nitric Oxide ◽  
Cytotoxic Activity ◽  
Nk Cells ◽  
Gene Knockout ◽  
Glioma Cells ◽  
Interferon Γ ◽  
Interleukin 12 ◽  
No Production ◽  
Content Type ◽  
Fine Print

Object. Interleukin (IL)-12 and IL-18 synergistically mediate antitumor responses through the production of interferon-γ (IFNγ) by T and natural killer (NK) cells. Recently, it has been reported that macrophages stimulated with these cytokines also produce IFNγ, which led the authors to investigate the antiglioma activity of macrophages stimulated by the combination of these cytokines in vitro. Methods. Dish-adherent peritoneal exudate cells, which had been elicited in thioglycollate broth as a source of macrophages, were used in the experiment. The murine glioma cell lines VM-glioma and 203G were labeled with [3H]thymidine for a cytotoxicity assay of macrophages. In response to the combined stimulation by IL-12 and IL-18, macrophages expressed potent cytotoxic activity against glioma cells in association with increasing production of IFNγ and nitric oxide (NO). Inhibitors of NO abrogated the cytotoxic activity of the macrophages, which had been induced by IL-12 and IL-18, despite the increase in IFNγ production. Neutralization of IFNγ or use of macrophages obtained from IFNγ gene-knockout mice markedly reduced not only cytotoxic activity, but also NO production. Depletion of T and NK cells from the macrophage population, which was achieved using antibody plus complement treatment, slightly reduced macrophage activities, suggesting that these are the main effector cells, although T and NK cells may partially participate in this cytotoxicity. Conclusions. Macrophages stimulated with IL-12 and IL-18 produced IFNγ and NO, which in turn mediated the antiglioma response. Therefore, macrophages as well as T and NK cells play an important role in antitumor responses stimulated by IL-12 and IL-18.

Intra-arterial ACNU therapy for malignant brain tumors

1983 ◽  
Vol 59 (3) ◽  
pp. 424-430 ◽  
Author(s):  
Junkoh Yamashita ◽  
Hajime Handa ◽  
Yasuhiko Tokuriki ◽  
Young Soo Ha ◽  
Shin-Ichi Otsuka ◽  
...  
Keyword(s):  
Survival Time ◽  
Postoperative Radiotherapy ◽  
Glioma Cells ◽  
Mantel Test ◽  
Survival Times ◽  
Content Type ◽  
The Difference ◽  
Intracarotid Administration ◽  
Fine Print

✓ The authors examined the growth rate of mouse 203 glioma cells in vitro and found it to be markedly inhibited after exposure to ACNU for 5 minutes at a drug concentration of 100 µg/ml. Rats that had undergone intracranial implantation of T1 neurogenic tumor were treated by 5 mg/kg of ACNU administered either intravenously or intra-arterially. The median survival times for the control animals and the animals undergoing intravenous or intracarotid administration of ACNU were 23, 29, and 46 days, respectively. The difference in survival time between the intravenous and intracarotid administration groups was statistically significant (p < 0.01) when examined by the Cox-Mantel test. In a clinical trial, 17 patients with glioblastoma were treated by ACNU, eight intravenously and nine by the intra-arterial route. The drug was given in doses of 2 to 3 mg/kg at least twice before and twice after a course of postoperative radiotherapy. Intra-arterial administration was performed over a period of 5 minutes under local anesthesia. The median postoperative survival time for the patients in the intra-arterial group was 12.5 months, compared with 9.0 months for those in the intravenous group. The survival rate for the intra-arterial group was slightly higher, although statistically not significant, probably because the number of cases was small. The degree of thrombocytopenia due to ACNU tended to be less marked in the intra-arterially treated patients. The theoretical advantages of the intra-arterial administration of ACNU are discussed.

In vitro photoradiation therapy of the rat 9L gliosarcoma

1986 ◽  
Vol 64 (5) ◽  
pp. 775-779 ◽  
Author(s):  
Douglas Hamilton ◽  
John D. S. McKean ◽  
John Tulip ◽  
Donald Boisvert ◽  
Judy Cummins
Keyword(s):  
Trypan Blue ◽  
Glioma Cells ◽  
Laser Source ◽  
Trypan Blue Exclusion ◽  
Content Type ◽  
9L Gliosarcoma ◽  
Trypan Blue Exclusion Test ◽  
9L Glioma ◽  
Fine Print

✓ The authors have investigated various factors involved in the photoradiation treatment of 9L glioma cells. The cells were grown in tissue culture and exposed to light from a laser source that allowed accurate quantitation of the light energy. Cell death was determined following treatment using the trypan blue exclusion test. It was shown that the treatment is very wavelength-dependent following the absorption spectrum of hematoporphyrin derivative (HPD). The absorption peaks in the lower part of the spectrum are more efficient than those of higher wavelengths. Photoradiation therapy is more effective the higher the concentration of HPD. Intensity of light is a very important factor in calculating the total dose of light necessary for this treatment.

Improvement of endothelial function in insulin-resistant carotid arteries treated with pravastatin

2001 ◽  
Vol 95 (3) ◽  
pp. 466-471 ◽  
Author(s):  
Aaron S. Dumont ◽  
M. Eric Hyndman ◽  
Randall J. Dumont ◽  
Paul M. Fedak ◽  
Neal F. Kassell ◽  
...  
Keyword(s):  
Risk Factors ◽  
Endothelial Function ◽  
Vascular Reactivity ◽  
No Production ◽  
Vascular Effects ◽  
Content Type ◽  
Insulin Resistant ◽  
Induced Hypertension ◽  
Fine Print

Object. Insulin resistance and hypertension are independent risk factors for stroke. Endothelial dysfunction in response to risk factors and carotid artery (CA) disease are important in the pathogenesis of stroke. Pravastatin may have cholesterol-independent pleiotropic effects. In the present study the authors examined the effects of short-course pravastatin treatment on endothelial function in CAs obtained in control and insulin-resistant rats with fructose-induced hypertension. Methods. Thirty rats were divided into two experimental groups, in which 14 were fed a regular diet and 16 were fed a fructose-enriched diet for 3 weeks. The rats were then divided into four groups: control, pravastatin-treated control, fructose-fed, and pravastatin-treated fructose-fed. Pravastatin was administered (20 mg/kg/day) for 2 weeks. Excretion of the urinary nitric oxide (NO) metabolite nitrite (NO2−) was also assayed. The CAs from all rats were subsequently removed and assessed for endothelium-dependent and -independent vascular reactivity in vitro. The rats in the fructose-fed group were insulin resistant, hyperinsulinemic, and hypertensive relative to the rats in the control and pravastatin-treated control groups and exhibited diminished endothelium-dependent vasomotion and urinary NO2− excretion (p < 0.05), with preserved endothelium-independent vasomotion. Strikingly, pravastatin treatment restored endothelium-dependent vasomotion and urinary NO2− excretion in rats in the fructose-fed pravastatin-treated relative to the fructose-fed group (p < 0.05). Conclusions. The authors report, for the first time, that pravastatin restores endothelial function in CAs from insulin-resistant rats with fructose-induced hypertension. These beneficial effects were ascribed to direct, cholesterol-independent vascular effects of pravastatin and are likely the result of augmentation of NO production. These data provide impetus for further investigation of nonlipid-lowering indications for pravastatin therapy in the prevention and treatment of CA disease.

Nitric oxide mediation of chemoregulation but not autoregulation of cerebral blood flow in primates

1996 ◽  
Vol 84 (1) ◽  
pp. 71-78 ◽  
Author(s):  
B. Gregory Thompson ◽  
Ryszard M. Pluta ◽  
Mary E. Girton ◽  
Edward H. Oldfield
Keyword(s):  
Nitric Oxide ◽  
Blood Flow ◽  
Cerebral Blood Flow ◽  
No Production ◽  
Direct Role ◽  
Content Type ◽  
Continuous Release ◽  
Fine Print

✓ The authors sought to develop a model for assessing in vivo regulation of cerebral vasoregulation by nitric oxide (NO), originally described as endothelial-derived relaxing factor, and to use this model to establish the role of NO in the regulation of cerebral blood flow (CBF) in primates. By using regional intraarterial perfusion, the function of NO in cerebral vasoregulation was examined without producing confounding systemic physiological effects. Issues examined were: whether resting vasomotor tone requires NO; whether NO mediates vasodilation during chemoregulation and autoregulation of CBF; and whether there is a relationship between the degree of hypercapnia and hypotension and NO production. Twelve anesthetized (0.5% isoflurane) cynomolgus monkeys were monitored continuously for cortical CBF, PaCO2, and mean arterial pressure (MAP), which were systematically altered to provide control and experimental curves of chemoregulation (CBF vs. PaCO2) and autoregulation (CBF vs. MAP) during continuous intracarotid infusion of 1) saline and 2) an NO synthase inhibitor (NOSI), either l-n-monomethyl arginine or nitro l-arginine. During basal conditions (PaCO2 of 38–42 mm Hg) NOSI infusion of internal carotid artery (ICA) reduced cortical CBF from 62 (saline) to 53 ml/100 g/per minute (p < 0.01), although there was no effect on MAP. Increased CBF in response to hypercapnia was completely blocked by ICA NOSI. The difference in regional (r)CBF between ICA saline and NOSI infusion increased linearly with PaCO2 when PaCO2 was greater than 40 mm Hg, indicating a graded relationship of NO production, increasing PaCO2, and increasing CBF. Diminution of CBF with NOSI infusion was reversed by simultaneous ICA infusion of l-arginine, indicating a direct role of NO synthesis in the chemoregulation of CBF. Hypotension and hypertension were induced with trimethaphan camsylate (Arfonad) and phenylephrine at constant PaCO2 (40 ± 1 mm Hg). Autoregulation in response to changes in MAP from 50 to 140 mm Hg was unaffected by ICA infusion of NOSI. In primates, cerebral vascular tone is modulated in vivo by NO; continuous release of NO is necessary to maintain homeostatic cerebral vasodilation; vasodilation during chemoregulation of CBF is mediated directly by NO production; autoregulatory vasodilation with hypotension is not mediated by NO; and increasing PaCO2 induces increased NO production.

Antiproliferative effect of trapidil, a platelet-derived growth factor antagonist, on a glioma cell line in vitro

1990 ◽  
Vol 73 (3) ◽  
pp. 436-440 ◽  
Author(s):  
Jun-ichi Kuratsu ◽  
Yukitaka Ushio
Keyword(s):  
Growth Factor ◽  
Cell Lines ◽  
Glioma Cell ◽  
Glioma Cells ◽  
Inhibitory Effect ◽  
Platelet Derived Growth Factor ◽  
Content Type ◽  
Glioma Cell Lines ◽  
Fine Print

✓ Platelet-derived growth factor (PDGF) is produced by glioma cells. However, there is heterogeneity among glioma cell lines in the production of PDGF. It has been demonstrated that U251MG cells produce a PDGF-like molecule while U105MG cells do not. Trapidil, a specific antagonist of PDGF, competes for receptor binding with PDGF. Therefore, the inhibitory effect of trapidil on the proliferation of glioma cells was investigated in vitro using two glioma cell lines. At 100 µg/ml, trapidil significantly inhibited the proliferation of U251MG cells (which produce the PDGF-like molecule). At the same trapidil concentration, the proliferation of U105MG cells (which do not produce the PDGF-like molecule) was not inhibited. The inhibitory effect of trapidil was remarkable on Days 3 and 4 of culture. After 4 days of incubation, the proliferation of U251MG cells was 46% of the control preparation. Trapidil enhanced the antitumor effect of 3-((4-amino-2-methyl-5-pyrimidinyl)ethyl)-1-(2-chloroethyl)-1-nitro-sourea (ACNU) against U251MG cells. The enhancing effect was highest on Days 4 and 6 of culture. After 6 days of incubation in the presence of 100 µg/ml trapidil and 1 µg/ml ACNU, the proliferation of U251MG cells was 18% of the control preparation. These findings suggest that trapidil interrupts the autocrine loop at the PDGF and PDGF-receptor level and that combination therapy with trapidil and ACNU may be useful in the treatment of glioma.

Modulation of phorbol ester—induced regulation of matrix metalloproteinases and tissue inhibitors of metalloproteinases by SB203580, a specific inhibitor of p38 mitogen-activated protein kinase

2002 ◽  
Vol 97 (1) ◽  
pp. 112-118 ◽  
Author(s):  
Myung-Jin Park ◽  
In-Chul Park ◽  
Jin-Heang Hur ◽  
Mi-Suk Kim ◽  
Hyung-Chan Lee ◽  
...  
Keyword(s):  
Protein Kinase ◽  
P38 Mapk ◽  
Specific Inhibitor ◽  
Glioma Cells ◽  
Tissue Inhibitors Of Metalloproteinases ◽  
Boyden Chamber ◽  
Content Type ◽  
In Vitro Invasion ◽  
Fine Print

Object. Expression of matrix metalloproteinases (MMPs) has been postulated to play a central role in brain tumor invasion; however, its underlying mechanism is not yet fully understood. In the present study, by assessing the effect of a specific p38 mitogen-activated protein kinase (MAPK) inhibitor, SB203580, on the secretion of MMPs and in vitro invasion of various glioma cells, the authors attempt to define the role of the p38 MAPK pathway in the regulation of MMPs and tissue inhibitors of metalloproteinases (TIMPs) activated by phorbol ester (phorbol-12-myristate-13-acetate [PMA]) in the D54 human glioblastoma cell line. Methods. The activation of MAPKs was determined using Western blot analysis after addition of phospho-specific antibodies against these kinases, the status of MMPs and TIMPs was analyzed using gelatin zymography and Western blot analysis, and the invasion rate of D54 cells and other glioma cells was analyzed using a modified Boyden chamber assay. Treatment of D54 cells with PMA activated two distinct MAPKs, extracellular signal-regulated kinase (ERK) 1/2 and p38 MAPK, but not c-Jun N-terminal kinase/stress-activated protein kinase. Induction of MMP-9 production and MMP-2 activation by PMA were blocked by SB203580, a specific inhibitor of p38 MAPK, but not by PD98059, a specific inhibitor of ERK 1/2. In addition, PMA-induced downregulation of TIMP-1 and TIMP-2 secretion and upregulation of the membrane type 1 MMP, a major activator of MMP-2 on the cell surface, were reversed by SB203580 in these cells; the PMA-induced increase of invasion in vitro decreased when SB203580 was added to the top compartment of a modified Boyden chamber; and the inhibitor also reduced the MMP secretion and PMA-induced in vitro invasion in various glioma cell lines. Conclusions. These results indicate that activation of p38 MAPK by PMA plays a central role in the regulation of MMPs and TIMPs in D54 cells, which has a major influence in tumor invasion and metastasis. Furthermore, inhibition of p38 MAPK by SB203580 blocked the secretion of MMPs and in vitro invasion of various glioma cells, underscoring a possible role of p38 MAPK inhibitors as antiinvasive and/or antimetastatic agents of malignant gliomas.

Inhibitory effect of antisense epidermal growth factor receptor RNA on the proliferation of rat C6 glioma cells in vitro and in vivo

2000 ◽  
Vol 92 (1) ◽  
pp. 132-139 ◽  
Author(s):  
Peiyu Pu ◽  
Xuwen Liu ◽  
Aixue Liu ◽  
Jianling Cui ◽  
Yunting Zhang
Keyword(s):  
Survival Time ◽  
Glioma Cells ◽  
C6 Glioma ◽  
C6 Glioma Cells ◽  
C6 Cells ◽  
Content Type ◽  
Proliferation Activity ◽  
Fine Print

Object. The goal of this study was to evaluate the effect of antisense epidermal growth factor receptor (EGFR) RNA on the growth of rat glioma cells in vitro and in vivo and to determine the feasibility of targeting the EGFR gene for gene therapy in gliomas.Methods. Antisense EGFR complementary (c)DNA was transfected into C6 glioma cells by using lipofectamine. In vitro studies, Southern and Northern blot analyses, in situ hybridization, and immunohistochemical staining were designed to examine the integration and expression of antisense EGFR constructs. The 3′(4,5-dimethylthiazol-2-yl)2,5-diphenyl tetrazolium bromide (MTT) assay and the average number of argyrophilic nuclear organizer regions (Ag-NORs) were used to evaluate cell proliferation, whereas the terminal deoxynucleotidyl transferase—mediated deoxyuridine triphosphate nick-end labeling (TUNEL) method and microscopy were used to observe cell apoptosis. As part of the in vivo studies, parental C6 cells and C6 cells transfected with EGFR antisense cDNA were implanted stereotactically into the right caudate nucleus of Wistar rats (C6-injected animals and transfected C6-injected animals). Rats with well-established cerebral C6 glioma foci were treated intratumorally with either antisense EGFR cDNA or empty-vector DNA by using lipofectamine (treated-C6 and control treated group). The general behavior and survival of the rats, findings on magnetic resonance images of their brains, histopathological changes, proliferation activity, and apoptosis of the cerebral gliomas in each group of rats were examined.Exogenous antisense EGFR cDNA was integrated into the genome of C6 cells and expressed. In clones with a high expression of the antisense construct, there was a dramatic decrease in endogenous EGFR messenger RNA and protein levels, reduced proliferation activity, and induction of apoptosis in vitro. The mean survival time of rats injected with C6 cells was 17.3 days. The mean survival time of rats injected with C6 cells followed by treatment with empty vector in lipofectamine was 15.4 days. Survival time was significantly prolonged in 100% of the rats injected with antisense-transfected C6 cells and in two thirds of the rats injected with C6 cells followed by antisense EGFR cDNA. Magnetic resonance imaging revealed distinct cerebral tumor foci in C6-injected rats and in control rats of the treated group, but none were found in the rats injected with transfected C6 cells. Furthermore, tumor foci disappeared completely in C6-injected rats treated with antisense EGFR cDNA. The cerebral gliomas of the rats treated by injection of antisense EGFR RNA were characterized by reduced proliferation activity and the induction of apoptosis.Conclusions. The results of this study indicate that EGFR plays an important role in the genesis of malignant gliomas. It may, therefore, be an effective target of antisense gene therapy in patients with gliomas.

Effects of dexamethasone and methylprednisolone on cell cultures of human glioblastomas

1971 ◽  
Vol 34 (3) ◽  
pp. 324-334 ◽  
Author(s):  
John Mealey ◽  
Tsu T. Chen ◽  
George P. Schanz
Keyword(s):  
Cell Cultures ◽  
Glioma Cells ◽  
Clinical Applications ◽  
Natural Rate ◽  
Content Type ◽  
Culture Growth ◽  
Different Origins ◽  
Dose Dependent ◽  
Fine Print

✓ Eight biopsied glioblastomas were propagated in vitro through multiple, serial cell cultures which were exposed to dexamethasone and methylprednisolone in concentrations ranging from 0.25 to 400 µg/ml. The higher concentrations of both steroids produced inhibition of culture growth and cytotoxic damage which appeared relatively nonspecific. Although the responses observed were dose-dependent, the sensitivity among glioma cultures of different origins varied. Of the two steroids, Methylprednisolone was more injurious to glioma cells in vitro and was cytolytic in doses of 400 µg/ml. Growth inhibition was demonstrated after a 1-day exposure to this corticoid, but this effect was usually transient at lower doses. Thereafter, surviving resistant cells resumed their natural rate of growth in the continued presence of the steroid as well as under standard conditions of culture. Potential clinical applications of the antineoplastic properties of corticosteroids against the gliomas are discussed, emphasizing the need to investigate additional, possibly more efficacious compounds.

Production of reactive oxygen species after reperfusion in vitro and in vivo: protective effect of nitric oxide

2000 ◽  
Vol 93 (1) ◽  
pp. 99-107 ◽  
Author(s):  
R. Bryan Mason ◽  
Ryszard M. Pluta ◽  
Stuart Walbridge ◽  
David A. Wink ◽  
Edward H. Oldfield ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Free Radical ◽  
Reperfusion Injury ◽  
Oxygen Free Radical ◽  
Content Type ◽  
Free Radical Production ◽  
Radical Production ◽  
Fine Print

Object. Thrombolytic treatments for ischemic stroke can restore circulation, but reperfusion injury, mediated by oxygen free radicals, can limit their utility. The authors hypothesized that, during reperfusion, nitric oxide (NO) provides cytoprotection against oxygen free radical species.Methods. Levels of NO and oxygen free radicals were determined in both reoxygenation in vitro and reperfusion in vivo models using an NO electrochemical probe and high-performance liquid chromatography with the 2,3- and 2,5-dihydroxybenzoic acid trapping method, before and after addition of the NO donor diethanolamine nitric oxide (DEA/NO).Reoxygenation after anoxia produced a twofold increase in NO release by human fetal astrocytes and cerebral endothelial cells (p < 0.005). In both cell lines, there was also a two- to threefold increase in oxygen free radical production (p < 0.005). In human fetal astrocytes and cerebral endothelial cells given a single dose of DEA/NO, free radical production dropped fivefold compared with peak ischemic levels (p < 0.001). In a study in which a rat global cerebral ischemia model was used, NO production in a vehicle-treated group increased 48 ± 16% above baseline levels after reperfusion. After intravenous DEA/NO infusion, NO reached 1.6 times the concentration of the postischemic peak in vehicle-treated animals. In vehicle-treated animals during reperfusion, free radical production increased 4.5-fold over basal levels (p < 0.01). After intravenous DEA/NO infusion, free radical production dropped nearly 10-fold compared with peak levels in vehicle-treated animals (p < 0.006). The infarct volume in the vehicle-treated animals was 111 ± 16.9 mm3; after DEA/NO infusion it was 64.8 ± 23.4 mm3 (p < 0.01).Conclusions. The beneficial effect of early restoration of cerebral circulation after cerebral ischemia is limited by reperfusion injury. These results indicate that NO release and oxygen free radical production increase during reperfusion, and suggest a possible early treatment of reperfusion injury using NO donors.

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