Vol. 55, Issue 1, 186-193, January 1999

Impediment to Calcium Influx and Reactive Oxygen Production Accounts for the Inhibition of Neutrophil Mac-1 Up-Regulation and Adhesion by Tetrandrine

Yuh-Chiang Shen, Chieh-Fu Chen, Sheng-Yuan Wang, and Yen-Jen Sung

Institutes of Pharmacology (Y.C.S.) and Anatomy and Cell Biology (Y.J.S.), School of Life Science, National Yang-Ming University, National Research Institute of Chinese Medicine (C.F.C.), and Department of Medical Research and Education, Veterans General Hospital (S.Y.W.), Taipei, Taiwan, Republic of China

We studied the mechanisms by which the plant alkaloid tetrandrine (TTD) inhibits Mac-1-dependent neutrophil adhesion to fibrinogen. TTD (0.1-10 µM) significantly inhibited Mac-1 up-regulation and neutrophil adhesion, as induced by N-formyl-methionyl-leucyl-phenylalanine (fMLP) or phorbol-myristate-acetate (PMA). Treatment of neutrophils with fMLP or PMA caused a rapid influx of Ca⁺⁺ and accumulation of reactive oxygen species (ROS), both of which have been shown to enhance neutrophil adhesion via Mac-1 up-regulation. Because TTD antagonizes Ca⁺⁺ influx and abrogates ROS, we examined the relationship between Ca⁺⁺ influx, ROS formation, and Mac-1 expression in TTD-inhibited neutrophil adhesion. TTD alone caused a slight but statistically significant 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 adhesion. TTD also inhibited increases in [Ca⁺⁺]_i and adhesion induced by the ionophore A23187 but failed to inhibit those induced by thapsigargin, an agent mobilizing Ca⁺⁺ from intracellular stores. Thus, TTD impeded Ca⁺⁺ influx from outward to avert neutrophil adhesion. Similarly, TTD and two ROS scavengers, superoxide dismutase and catalase, abolished ROS production, Mac-1 up-regulation, and neutrophil adhesion. Ca⁺⁺ and ROS, therefore, represent two essential signals for Mac-1 up-regulation upon fMLP or PMA stimulation. Our data suggest that the antiadherent effect of TTD is mediated, in part, by the inhibition of Ca⁺⁺ influx and ROS formation, resulting in suppressed up-regulation of Mac-1 and, in turn, neutrophil adhesion to fibrinogen.