Impediment to calcium influx and reactive oxygen production accounts for the inhibition of neutrophil mac-1 up-regulation and adhesion by tetrandrine

We studied the mechanisms by which the plant alkaloid tetrandrine (TTD) inh ibits Mac-1-dependent neutrophil adhesion to fibrinogen. TTD (0.1-10 mu M) significantly inhibited Mac-1 upregulation and neutrophil adhesion, as indu ced by N-formylmethionyl-leucyl-phenylalanine (fMLP) or phorbol-myristate a cetate (PMA). Treatment of neutrophils with fMLP or PMA caused a rapid infl ux of Ca++ and accumulation of reactive oxygen species (ROS), both of which have been shown to enhance neutrophil adhesion via Mac-1 up-regulation. Be cause TTD antagonizes Ca++ influx and abrogates ROS, we examined the relati onship between Ca++ influx, ROS formation, and Mac-1 expression in TTD-inhi bited neutrophil adhesion. TID alone caused a slight but statistically sign ificant increase in [Ca++](i) with no effect on adhesion. In contrast, TTD as well as two Ca++ channel antagonists, verapamil and nifedipine, markedly diminished fMLP- and PMA-induced Ca++ influx, Mac-1 up-regulation, and adh esion. TTD also inhibited increases in [Ca++](i) and adhesion induced by th e ionophore A23187 but failed to inhibit those induced by thapsigargin, an agent mobilizing Ca++ from intracellular stores. Thus, TTD impeded Ca++ inf lux from outward to avert: neutrophil adhesion. Similarly, TTD and two ROS scavengers, superoxide dismutase and catalase, abolished ROS production, Ma c-1 up-regulation, and neutrophil adhesion. Ca++ and ROS, therefore, repres ent two essential signals for Mac-1 up-regulation upon fMLP or PMA stimulat ion. Our data suggest that the antiadherent effect of TTD is mediated, in p art, by the inhibition of Ca++ influx and ROS formation, resulting in suppr essed up-regulation of Mac-1 and, in turn, neutrophil adhesion to fibrinoge n.