Involvement of Na+,K(+)-ATPase inhibition in K562 cell differentiation induced by bufalin.
Human leukemia K562 cell differentiation induction by naturally occurring bufadienolides purified from the Chinese drug Senso and synthetic bufalin derivatives was examined by a nitro blue tetrazolium reduction assay. Bufalin showed the strongest activity among all the bufadienolides tested in this study. The degree of the induction of nitro blue diformazan positive cells by the bufadienolides correlated well with their inhibitory activities against Na+,K(+)-ATPase prepared from K562 cells in vitro. N+,K(+)-ATPases from a variant K562 clone (ouabain resistant, OuaR) and murine leukemia cell line M1-T22, which were insensitive to the bufadienolides in terms of growth inhibition and cell differentiation, appeared to be refractory to bufalin in vitro. A binding study of 3H-bufalin and 3H-ouabain revealed that saturated levels of both ligands associated with K562 cells were virtually similar; however, affinity of 3H-bufalin was considerably higher than 3H-ouabain. The saturated level of 3H-bufalin observed in the OuaR cells was approximately half of that observed in K562 cells without a change in its affinity. Association of 3H-bufalin with K562 cells was completely blocked by pretreatment of the cells with cold ouabain at concentrations saturating the binding sites. These results suggest that bufalin acts on the cells by binding to sites on the cell membrane which also bind ouabain. It is thus proposed that N+,K(+)-ATPase inhibition is closely related to the initiation process in the induction of K562 cell differentiation induced by bufalin.
Department of Biochemical Toxicology, School of Pharmaceutical Sciences, Showa University, Tokyo, Japan.
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S
Numazawa
M A
MA
Shinoki
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H
Ito
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T
Yoshida
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Kuroiwa
D000818:Animals; D002018:Bufanolides / Q000494:pharmacology*; D002454:Cell Differentiation / Q000187:drug effects; D002471:Cell Transformation, Neoplastic / Q000473:pathology*; D004351:Drug Resistance; D006801:Humans; D007942:Leukemia, Experimental / Q000378:metabolism / Q000473:pathology*; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000378:metabolism / Q000473:pathology*; D051379:Mice; D010042:Ouabain / Q000494:pharmacology; D000254:Sodium-Potassium-Exchanging ATPase / Q000037:antagonists & inhibitors* / Q000502:physiology*; D014316:Tritium; D014407:Tumor Cells, Cultured
D002018:Bufanolides; D014316:Tritium; D010042:Ouabain; D000254:Sodium-Potassium-Exchanging ATPase; C022777:bufalin