Differentiation Of Human Leukemia HL-60 Cells Induced By A Sesquiterpene BS1 And H₂O₂-activated NADPH Oxidase

It has been made some progress in clinic to cure human leukemia through inducing cell differentiation, however, the differentiation therapy is restricted for wide application, due to that most differentiation inducers at present have severe toxic side-effects, and also due to that cells are easy to develop drug resistance. After a great deal of screening through in vitro experiments, it was explored that one bisabolane sesquiterpene could specially induce the differentiation of human leukemia HL-60 cells with low toxicity and high efficiency. On the other hand, the mechanism of HL-60 cell differentiation was discussed to some extent. The role of NADPH oxidase was examined in the process of differentiation induced by H₂O₂ in HL-60 cells, because reactive oxygen species (ROS) play important roles during cacinogenesis, and also because NADPH oxidase is the main enzyme in cells to generate ROS.Firstly, it showed that a bisabolane sesquiterpene could induce differntiation of HL-60 cells. This bisabolane sesquiterpene, rel-(1S,4R,5S,6R)-4,5-diacetoxy-6-[(R)-5-hydroxy-1,5-dimethylhex-3-enyl]-3-methylcyclohex-2-enyl (Z)-2-methylbut-2-enoate (BS1 for short in this paper), was newly isolated from the roots of Leontopodium longifolium. BS1 presented special anticancer activity against human leukemia HL-60 cells, but did not inhibit proliferation of human hepatoma SMMC-7721 cells and human normal hepatocytes L02 cells. Nitroblue tetrazolium (NBT) reduction, phagocytosis of latex beads, and cell electrophoresis all demonstrated that this bisabolane sesquiterpene presented its anticancer activity against human leukemia HL-60 cells in vitro via inducing cell differentiation. These results may have implication for treatment of human leukemia with BS1.Subsequently, we studied the effect of NADPH oxidase in the process of differentiation in HL-60 cells induced by H₂O₂. Different concentrations of H₂O₂ had different effects. Our present work showed that 200μM H₂O₂ induced the differentiation of HL-60 cells significantly, that is, promoted phagocytosis of latex beads and prolonged the electrophoresis time, which could be respectively blocked by Diphenylene iodoniurn (DPI) and Apocynin (APO), inhibitors of NADPH oxidase. At the same time, H₂O₂ stimulated the generation of O₂^-, while DPI and APO diminished this production. These results demonstrated that NADPH oxidase was probably involved in the process of differentiation induced by H₂O₂ in human leukemia HL-60 cells.In brief, this paper presented the special anticancer activity of BS1 in HL-60 cells, and implicated the important role of NADPH oxidase during the differentiation of HL-60 cells induced by H₂O₂, which might provide some new clues to cure human leukaemia.