Effect of Leukotriene B4 on Enhancement of Superoxide Production Evoked by Formyl-Methionyl-Leucyl- Phenylalanine in Myeloid Differentiated HL-60 Cell$: Possible Involvement of Intracellular Calcium Influx and High Affinity Receptor for Leukotriene

Exposure of a human leukemic cell line HL-60 to 1 % dimethylsulfoxide (DMSO) for 4 days induced myeloid differentiation. DMSO-differentiated HL-60 cells displayed high and low-affinity binding sites for leukotriene B4 (LTB4). The pretreatment of myeloid differentiated HL-60 cells with 1-10 nM LTB4 enhanced superoxide production evoked by 100 nM formyl-methionyl-leucylphenylalanine (fMLP) to 127-137% of the controls stimulated by ±MLP alone. A concentration eliciting a half maximal increase (EC50) of LTB4 for the enhancing effect on superoxide production evoked by ±MLP was 0.32 nM. This was roughly similar to the dissociation constant (Kd) of high affinity receptors for LTB4 (0.23 nM). These results suggest that high affinity receptors transduce the enhancing effect of LTB4 on fMLP-induced superoxide production. Although it seems possible that enhancement of fMLP-induced superoxide production is associated with a substantial increase and/or an affinity alteration in receptors for fMLP, LTB4-pretreated cells failed to show significant changes in ±MLP binding compared to non-pretreated ones. It seems likely that Ca+ influx transduces enhancement of ±MLP-induced superoxide production, because extracellular Ca + is necessary for an enhancing effect of fMLP-induced superoxide production. Also, EC50 of LTB4 for Ca 2 + influx (0.78 nM) was similar to that of the enhancing effect of superoxide generation evoked by ±MLP. Although pretreatment of LTB4 failed to enhance the maximal level of ±MLP-induced intracellular Ca + rise, transient overshoot in intracellular Ca + evoked by ±MLP declined more rapidly after LTB4 pretreatment. Possible involvement of high affinity binding sites for LTB4 and Ca + influx was noted in the LTB4-enhancement of ±MLPinduced superoxide production in DMSO-differentiated HL-60 cells. However, the significance of the rapid attenuation of intracellular Ca + overshoot needs further evaluation.