Studies of factors involved in the survival of phorbol ester TPA-treated leukemic HL60 cells

Institute of Biochemistry, Vilnius, Lithuania The aim of this study was to determine the factors responsible for the survival of 12-O-tetradecanoyl-phorbol-13-acetate (TPA)-treated differentiating HL60 cells. After TPA treatment, HL60 cells in nonadherent cell fraction undergo apoptosis, whereas they survive and differentiate when attached to substrate. We have found that the amount of viable TPA-treated cells grown in a shaker decreased greatly in comparison to control cells grown under standard conditions, suggesting the importance of cell adhesion for their survival. Different approaches were used to evaluate the role of the transcription factor NF-źB in the TPA-induced process of cell survival/differentiation. We have found that NF-kB-DNA binding activity was much stronger in TPA-treated differentiating HL60 cells than in apoptotic ones, as was shown by an electrophoretic mobility shift assay (EMSA). The treatments directed to inactivate NF-kB by using the inhibitors of NF-kB signalling pathway, pyrrolidine dithiocarbamate (PDTC) and (E)-3-[(4-methylphenylsulfonyl]-2propenenitrile (BAY 11-7082), effectively promoted TPA-induced cell death. The possible activator of NF-kB, TNFα, was positively involved both in TPA-induced differentiation and apoptosis of HL60 cells, – monoclonal antibodies against TNFα attenuated TPA-induced HL60 cell differentiation (adhesion to substrate) and the death process in suspension cell fraction. It can be concluded that cell adhesion and NF-kB activity are essential for the survival of TPA-treated HL60 cells.