leukotriene-c4 and Glioma

Bradykinin and leukotriene C4 (LTC4) have been shown to increase molecular transport across the blood-tumor barrier (BTB). The aim of this study was to quantitatively assess whether an increase in vesicular transport or opening of tight junctions was responsible for this increase in permeability. Wistar rats bearing RG2 or C6 gliomas were infused with bradykinin or LTC4 through the right carotid artery for 15 min and then perfused to achieve fixation. Prepared specimens were observed using transmission electron microscopy. Infusion of either bradykinin or LTC4 resulted in significantly increased vesicular density in capillary endothelial cells of the BTB but not in normal brain capillaries. The opening of tight junctions, assessed by determining a cleft index, was found to be greater in tumor capillaries compared to normal controls. However, neither bradykinin nor LTC4 produce variations in the cleft index. A significant accumulation of horseradish peroxidase was seen in the intercellular peri-capillary spaces and in endothelial transport vesicles after infusion of bradykinin, demonstrating that the formation of vesicles was associated with macromolecular transcytosis. These findings suggest that pinocytotic vesicular transport is the primary means by which luminal to abluminal transport occurs in response to vasomodulation with bradykinin or LTC4.

Topics: Animals; Biological Transport; Bradykinin; Brain Neoplasms; Capillaries; Capillary Permeability; Endothelium, Vascular; Female; Glioma; Horseradish Peroxidase; Leukotriene C4; Microscopy, Electron; Rats; Rats, Wistar; Tight Junctions

Previous work in our laboratory has shown a correspondence between the chemosensitivity of C6 rat glioma and that of human glioblastoma (GBM) to a panel of chemotherapeutic agents in vitro, as determined by the MTT [3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H tetrazolium bromide] colorimetric assay. In the present study, an in vivo model of intracerebral C6 glioma in Sprague-Dawley rats was used to determine if a correlation exists between in vitro chemosensitivity and in vivo survival of the animals, and post-mortem histopathological changes in the tumor. Cisplatin (CDDP) and methotrexate (MTX), agents previously shown to demonstrate high and low in vitro cytotoxicity, respectively, against C6, were administered by intra-carotid infusion over the course of two days. In a separate series of animals, LTC4 was administered prior to infusion of CDDP or MTX; LTC4 was used in view of its known, selective, vasogenic effect on the permeability of brain tumor capillaries. It was found that survival of animals treated with CDDP alone was increased, although this did not reach statistical significance; histopathologically, CDDP-treated animals showed significant tumor necrosis. However, in CDDP-treated animals, pre-treatment with LTC4 increased survival to a statistically significant degree. When administered alone, LTC4 (not followed by CDDP) had no effect on either survival or histology. The survival-enhancing effect of CDDP, when combined with LTC4, was probably not due to any cytotoxic effect of LTC4; this is based on our finding that, on the in vitro MTT colorimetric assay, LTC4 showed low cytotoxicity for C6 glioma cells. By contrast with CDDP, MTX -- with or without pretreatment with LTC4 -- affected neither survival nor tumor histology. With respect to the question of correspondence between the MTT colorimetric in vitro assay and in vivo effect, MTX showed a clear correlation: low cytotoxicity in vitro and poor in vivo response. In the case of CDDP, the correspondence was not clear-cut: there was a high level of in vitro chemosensitivity of the C6 cell line to CDDP as well as post-mortem tumor necrosis, but in vivo testing showed no significant prolongation of survival. However, pre-treatment with LTC4 did significantly extend survival in animals treated with CDDP.

Topics: Animals; Brain Neoplasms; Cell Survival; Cisplatin; Colorimetry; Female; Glioma; Leukotriene C4; Male; Methotrexate; Premedication; Rats; Rats, Sprague-Dawley; Survival Analysis; Tetrazolium Salts; Thiazoles

Treatment of human glioma A172 cells with 1-(4-amino-2-methyl-5-pyrimidinyl)methyl-3-(2-chloroethyl)-3-nitrosourea (ACNU), an alkylating antitumor agent the primary target of which has been thought to be DNA, resulted in elevated expression of mRNA for multidrug resistance-associated protein (MRP) within the first 2 h and then a decrease in expression 24 h after the treatment. Western blot analyses revealed that levels of MRP in these ACNU-treated cells paralleled mRNA levels. Membrane vesicles prepared from ACNU-treated cells also displayed elevated transport activities for leukotriene C4, a known substrate for MRP. Gamma-glutamylcysteine synthetase (gamma-GCS) mRNA expression was coinduced with MRP by ACNU. Because gamma-GCS is the rate-limiting enzyme involved in the de novo biosynthesis of glutathione, increases in glutathione were also transiently induced by ACNU. These results demonstrate for the first time that the expression of functional MRP and gamma-GCS can be transiently coinduced by ACNU. Multiple short exposures (1 h) of ACNU following a long duration (1 week) of drug-free conditions resulted in the development of an ACNU-resistant population (designated A172R) that overexpressed MRP/gamma-GCS mRNA and had elevated transport activities for leukotriene C4. A172R exhibited cross-resistance to the antitumor drug doxorubicin and heavy metal sodium arsenate but not to cisplatin. Our results also demonstrate that intermittent treatments of human glioma cells with ACNU can lead to the development of MRP-related multidrug resistance. These results, taken together, reveal a possible new mechanism of the development of drug resistance for the antitumor nitrosoureas.

Topics: Amino Acid Sequence; Animals; Antibodies; Antineoplastic Agents; Arsenates; ATP-Binding Cassette Transporters; Biological Transport; Carrier Proteins; Cisplatin; Doxorubicin; Drug Resistance, Multiple; Enzyme Induction; Gene Expression Regulation, Neoplastic; Glioma; Glutamate-Cysteine Ligase; Glutathione; Humans; Leukotriene C4; Membrane Transport Proteins; Mice; Mice, Inbred BALB C; Models, Biological; Molecular Sequence Data; Multidrug Resistance-Associated Proteins; Nimustine; Peptide Fragments; RNA, Messenger; Transcription, Genetic; Tumor Cells, Cultured