Redox reagents and staurosporine inhibit stimulation of the transcription regulator NF-kappa B following tumour necrosis factor treatment of chronic B-leukaemia cells.

B-chronic lymphocytic leukaemia (B-CLL) and hairy cell leukaemia cells (HCL) are refractory to stimulation by several cytokines which activate normal B-cells. However, tumour necrosis factor (TNF) promotes the proliferation of these cells. TNF regulates some of its cellular responses via the transcription factor NF-kappa B. Using an electrophoretic mobility shift assay, we demonstrate that TNF treatment of B-CLL and HCL cells in vitro resulted in the augmentation of NF-kappa B levels. In haemopoietic cell lines, TNF induction of NF-kappa B is mediated via the generation of reactive oxygen intermediates and by the activation of protein kinase C (PKC). We have used activators and inhibitors of these pathways to unravel TNF signalling in the cells of ten patients with B-CLL and two with HCL, using the increase in NF-kappa B levels following TNF treatment as an end point. Raising glutathione levels with N-acetyl cysteine substantially reduced NF-kappa B induction by TNF in two of four samples tested. These data suggest that redox mechanisms are involved in TNF signalling in these cells. Treatment with the PKC activator phorbol myristate acetate failed to activate NF-kappa B suggesting that this enzyme does not mediate the induction of NF-kappa B in these cells. However, the protein kinase inhibitor staurosporine inhibited TNF induction of NF-kappa B in four of five samples, suggesting that staurosporine-sensitive protein kinases (other than PKC) are involved in the signalling pathway. Our results suggest that PKC-independent pathways, including pathways sensitive to redox reagents, mediate the induction of NF-kappa B by TNF in chronic B-leukaemia cells. Additionally, these data suggest that defects in PKC-mediated pathways may contribute to the general reluctance of B-CLL and HCL cells to respond to mitogenic signals.