Stimulation by tumor necrosis factor of HL-60 thymidine salvage pathway metabolism dissociated from proliferation.
The effect of human tumor necrosis factor (TNF) on early-passage HL-60 cells was studied. A transient phase of increased [3H]thymidine (TdR) incorporation was noted at 20-24 hr of exposure to TNF. This increase was disproportionate to the much slighter stimulation of the percentage of S-phase cells, which was measured by flow cytometry. Evidence for increased metabolic trapping of [3H]TdR following TNF treatment was apparent from whole cell uptake experiments. The salvage pathway enzyme TdR kinase was therefore measured and was found to be elevated comparably to [3H]TdR uptake. The mechanism of TNF regulation of TdR kinase was further investigated by a series of combination treatment experiments using other biologic factors and pharmacologic inhibitors of various intracellular steps. The response to TNF was not potentiated or reproduced by IL-1, IL-2, IL-3, IL-4, G-CSF, M-CSF, GM-CSF or alpha- or gamma-interferon. Blockers of early signal transduction steps, including H7, W7, sphingosine, and pertussis toxin, failed to inhibit TNF stimulation of [3H]TdR incorporation. mRNA synthesis inhibition with alpha-amanitin blocked this TNF effect, as did cAMP but not cGMP analogues. A sensitizing effect was noted with amiloride or cytochalasin B, characterized by greater relative increases of [3H]TdR incorporation and TdR kinase activity in response to TNF. In the presence of cytochalasin B, TNF treatment resulted in no change or slight decreases in the percentage of S-phase cells. Regulation of TdR kinase could thereby be dissociated from the usual cell cycle control. This study thus documents a unique example of stimulation of thymidine salvage pathway metabolism by a biologic factor, dissociable from overall cell cycle regulation.
Elias
L
L
Cancer Center, University of New Mexico School of Medicine, Albuquerque 87131.
D000584:Amiloride / Q000494:pharmacology; D002453:Cell Cycle; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D003571:Cytochalasin B / Q000378:metabolism; D006801:Humans; D015470:Leukemia, Myeloid, Acute / Q000378:metabolism*; D013936:Thymidine / Q000378:metabolism* / Q000493:pharmacokinetics; D013937:Thymidine Kinase / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D014409:Tumor Necrosis Factor-alpha; D003571:Cytochalasin B; D000584:Amiloride; D013937:Thymidine Kinase; D013936:Thymidine