Stilbene disulfonic acids inhibit synexin-mediated membrane aggregation and fusion

Stilbene disulfonic acids inhibit surfactant secretion from lung epithelial type II cells by an undefined mechanism, and inhibit CD4 mediated cell-cell fusion. We have previously shown that lung synexin promotes in vitro fusion of lamellar bodies and plasma membranes, an obligatory process for surfactant secretion. This study investigates the effect of stilbene disulfonic acids, 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS), 4-acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic acid (SITS), and 4-acetamido-4'-maleimidylstilbene-2,2'-disulfonic acid (AMDS), on synexin-mediated liposome aggregation and fusion. Structurally, these three stilbene compounds differ in the number of isothiocyano groups present (DIDS = 2, SITS = 1, and AMDS = 0). At 10 micrograms synexin/ml, DIDS and SITS inhibited synexin-mediated liposome aggregation with an EC50 of 3.5 microM and 148 microM, respectively. In comparison, AMDS was least inhibitory (EC50 > 1 mM). Thus, the inhibitory potency (DIDS > SITS > AMDS) was partly dependent upon the number of isothiocyano groups. The EC50 was also dependent on synexin concentration. Stilbene disulfonic acids were also inhibitory for arachidonic acid-enhanced synexin-mediated liposome fusion. The EC50 for DIDS and SITS for fusion were similar to that for liposome aggregation. Ca(2+)-induced synexin polymerization, measured by 90 degrees light scattering, was increased by DIDS, suggesting binding of stilbene disulfonic acids to synexin. The binding of DIDS to synexin was dependent on the molar ratio of synexin to DIDS. These results indicate that stilbene disulfonic acids interact directly with synexin to inhibit membrane aggregation and fusion. Our results suggest that such inhibition of synexin activity may contribute towards inhibition of surfactant secretion by DIDS, and support a physiological role for synexin in lung surfactant secretion.