Effects of nitrogen mustard on potassium transport systems and membrane structure of Ehrlich ascites tumor cells.
By Ehrlich ascites tumor cells 86Rb+ has been shown to be a suitable tracer for K+-transport. Sixty percent of the total 86Rb-uptake into these cells is ouabain-inhibitable, 30% is sensitive to furosemide and 10% enters the cells by ouabain and furosemide-insensitive systems. N-Mustard inhibits both the ouabain-sensitive and the furosemide-inhibitable systems. The uptake which is resistant to both inhibitors is not affected by the alkylating drug. At N-mustard concentrations below 10 microM, the reduction of the Rb-uptake is predominantly due to the inhibition of the furosemide-sensitive transport. Higher concentrations are required before a significant inhibition of the ouabain-sensitive transport can be observed. The dose response curve of the furosemide-sensitive transport--not, however, of the ouabain inhibitable pump--corresponds to the dose response curve for the antiproliferative activity of N-mustard. The recovery of the furosemide-sensitive transport after a single exposure to N-mustard is relatively slow and--in contrast to the repair of DNA cross-links--is characterized by an initial 4-hr lag period. Furosemide alone does not interfere with cell multiplication. The inhibition of the transport system alone does, therefore, not explain the antitumor activity of N-mustard. The effect is discussed as a marker for membrane lesions after exposure to alkylating agents. In order to investigate the influence of N-mustard on membrane structure, membranes were labelled with diiodofluoresceiniodoacetamide. Anisotropy curves obtained from time-dependent depolarization of delayed fluorescence indicated a mustard induced immobilization of membrane constituents. Lateral diffusion of lipophilic probes was determined by following the quenching of fluorescence of pyrene by cetylpyridinium. The latter studies yielded no evidence for a change in membrane lipid fluidity. The data are interpreted as the results of cross-links of membrane proteins by the bifunctional alkylating agent.
D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002286:Carcinoma, Ehrlich Tumor / Q000378:metabolism* / Q000473:pathology; D002455:Cell Division / Q000187:drug effects; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D004247:DNA / Q000378:metabolism; D005665:Furosemide / Q000494:pharmacology; D008466:Mechlorethamine / Q000494:pharmacology*; D010042:Ouabain / Q000494:pharmacology; D011188:Potassium / Q000378:metabolism*; D012413:Rubidium / Q000378:metabolism
D008466:Mechlorethamine; D010042:Ouabain; D005665:Furosemide; D004247:DNA; D012413:Rubidium; D011188:Potassium