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Effect of car☐ylic ionophores, membrane active and lysosomotropic agents on adriamycin uptake and histamine release in rat peritoneal mast cells
|706 Thus the usage of the developed pharmaceutical form (MCD) of anticancer drugs gives us with the help of magnetic field, control over drug distribution on in organism and localisation of its cytotoxic action mainly in the tumor tissue.
Effect of carboxylic ionophores, membrane active and lysosomotropic agents on adrimnydn uptake and histamine release in rat peritoneal mast cells Bartoli K l u g m a n n , F., Ek'corti, G., Crivellato *, E., M a l l a r d i * *, F., Candussio, L. a n d Baldini, L. l.~titutc of Pharmacology, Via Valerio 32, 1-34100 Trieste, * Institute of Anatomy, Via Manzoni 16, 1-34100 Trieste and ** Chair of Anutomy, Univt'rsity of Udine, 1-33100 Udine, Italy
It has been recently shown that the antineoplastic drug adriamycin exhibits a high affinity for mast cells in ¢cmp~-ison vci~ other normal and neoplastic cells, and that the uptake of the anthracycline is related to a non cytotoxic histamine release (Decorti, 1989). Adiiamycinis acct,,nulated in mast cells by an active transport system~ which closely resembles the outward transport system of multi drug resistant (MDR) cells. Recently it has been proposed (Beck, 1987) that in MDR cells, drugs such as anthracyclines and vinca alkaloids, which are weak bases, are entrapped in the endosomal or lysosomal vescicles, which have an acidic pH, and are then extruded to the outside by "fusion of the.~e drug-containing vescicles with plasma membrane. Many orgamc cationic compounds, which interfere with membrane traffic processes such as recycling, endocytosis, vescicle fusion and exocytosis, have been succesfully used to reverse resistance in the M D R cells. The present study was undertaken in order to test the effect of substances that are known to limit or reverse resistance in MDR cells on adriamycin uptake and histamine reease in rat peritoneal mast cells. The substances used were the carboxylic ;_onophores monensin and nigericin, the membrane active substances Triton WR-1339, Tween 80, deoxycholate and amphotericin B, and the lysosomotropic amines chloroquine, nicotine, propranolol, atropine, amanl~dine, methylamine, ammonium chloride, and quinacrine. Purified rat peritoneal mast 6ells were incubated at 37°C with various concentrations of the inhibitors, and then adriamycin (50 ug/ml) was added and the incubation continued for 10 additional rain. The amount of histamine released and adriamycin uptake were measured by f'aorimetric methods. Cells treated as detailed above were also processed for epifluorescence microscopy The carboxylic ionophores were extremely efficacious in preventing adriamycin uptake and histamine release in rat peritoneal cells; in particular, monensin effect was dependent on the time that cells were exposed to the inhibitor prior to the addition of the secretagogue. Removal of monensin before stimulation with adriamycin did not abolish the inhibitory effect. Among the tested membrane active substances, Tween 80 and Triton WR-1339 were active; on the contrary, the lysosomotropic amines were almost completely ineffective at non cytotoxic concentrations. The "~icroscopical examination revealed that in mast cells treated with adriamycin an intense fluorescence was present in cytoplasmic g r a ~ e s ; on the contrary, mast cells preincubated with monensin showed hardly any fluorescence. These data suggest that in mast cells, the plasma menzbrane and membrane trafficking are particularly important in adriamycin uptake; on the contrary, the relative inactivity of substances that alter the pH of lysosomes indicate that the transport of the drug in these cells is partially different from that of MDR cells.
References Decor,A, G., Bartoli Klugmann, F., Candussio, L., Furlani, A., Scarcia, V., Baldini, L., 1989, Cancer Res. 49, 1921. Beck, W.T., 1987, Biochem. Pharmacol. 36, 2879.
Effect of carboxylic ionophores, membrane active and lysosomotropic agents on adrimnydn uptake and histamine release in rat peritoneal mast cells Bartoli K l u g m a n n , F., Ek'corti, G., Crivellato *, E., M a l l a r d i * *, F., Candussio, L. a n d Baldini, L. l.~titutc of Pharmacology, Via Valerio 32, 1-34100 Trieste, * Institute of Anatomy, Via Manzoni 16, 1-34100 Trieste and ** Chair of Anutomy, Univt'rsity of Udine, 1-33100 Udine, Italy
It has been recently shown that the antineoplastic drug adriamycin exhibits a high affinity for mast cells in ¢cmp~-ison vci~ other normal and neoplastic cells, and that the uptake of the anthracycline is related to a non cytotoxic histamine release (Decorti, 1989). Adiiamycinis acct,,nulated in mast cells by an active transport system~ which closely resembles the outward transport system of multi drug resistant (MDR) cells. Recently it has been proposed (Beck, 1987) that in MDR cells, drugs such as anthracyclines and vinca alkaloids, which are weak bases, are entrapped in the endosomal or lysosomal vescicles, which have an acidic pH, and are then extruded to the outside by "fusion of the.~e drug-containing vescicles with plasma membrane. Many orgamc cationic compounds, which interfere with membrane traffic processes such as recycling, endocytosis, vescicle fusion and exocytosis, have been succesfully used to reverse resistance in the M D R cells. The present study was undertaken in order to test the effect of substances that are known to limit or reverse resistance in MDR cells on adriamycin uptake and histamine reease in rat peritoneal mast cells. The substances used were the carboxylic ;_onophores monensin and nigericin, the membrane active substances Triton WR-1339, Tween 80, deoxycholate and amphotericin B, and the lysosomotropic amines chloroquine, nicotine, propranolol, atropine, amanl~dine, methylamine, ammonium chloride, and quinacrine. Purified rat peritoneal mast 6ells were incubated at 37°C with various concentrations of the inhibitors, and then adriamycin (50 ug/ml) was added and the incubation continued for 10 additional rain. The amount of histamine released and adriamycin uptake were measured by f'aorimetric methods. Cells treated as detailed above were also processed for epifluorescence microscopy The carboxylic ionophores were extremely efficacious in preventing adriamycin uptake and histamine release in rat peritoneal cells; in particular, monensin effect was dependent on the time that cells were exposed to the inhibitor prior to the addition of the secretagogue. Removal of monensin before stimulation with adriamycin did not abolish the inhibitory effect. Among the tested membrane active substances, Tween 80 and Triton WR-1339 were active; on the contrary, the lysosomotropic amines were almost completely ineffective at non cytotoxic concentrations. The "~icroscopical examination revealed that in mast cells treated with adriamycin an intense fluorescence was present in cytoplasmic g r a ~ e s ; on the contrary, mast cells preincubated with monensin showed hardly any fluorescence. These data suggest that in mast cells, the plasma menzbrane and membrane trafficking are particularly important in adriamycin uptake; on the contrary, the relative inactivity of substances that alter the pH of lysosomes indicate that the transport of the drug in these cells is partially different from that of MDR cells.
References Decor,A, G., Bartoli Klugmann, F., Candussio, L., Furlani, A., Scarcia, V., Baldini, L., 1989, Cancer Res. 49, 1921. Beck, W.T., 1987, Biochem. Pharmacol. 36, 2879.