Tenidap and other anion transport inhibitors disrupt cytolytic T lymphocyte-mediated IL-1 beta post-translational processing

Abstract

LPS-activated murine peritoneal macrophages produce IL-1 beta but externalize little mature cytokine in the absence of a secondary stimulus, and CTLs previously were reported to serve this capacity. The release of 17-kDa IL-1 beta from LPS-activated BALB/c macrophages occurred rapidly after the addition of C57/B1-derived allogeneic CTLs; within 30 min of coculture, mature IL-1 beta was observed in the medium, and maximum release was achieved within 4 h. CTL-induced post-translational processing was efficient, and >80% of newly synthesized pro-IL-1 beta was released into the medium as the 17-kDa species. Externalization of IL-1 beta required active recognition of the macrophage target by the CTL preparation; C57/B1 CTLs promoted the release of mature IL-1 beta from allogeneic BALB/c macrophages, but not from syngeneic C57/B1 macrophages. In contrast, extracellular ATP promoted mature IL-1 beta release from both macrophage populations. CTL-induced cytokine post-translational processing was blocked by anion transport inhibitors, including 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid, UK5099, and the anti-inflammatory agent tenidap. An analogue of tenidap, CP-100,829, was more effective as an inhibitor of both IL-1 beta post-translational processing and anion transport. In contrast, the close structural analogue CP-236,492 inhibited neither process. Tenidap's activity was reversible and was not mimicked by cyclooxygenase inhibitors or by cycloheximide. Therefore, tenidap disrupted CTL-induced IL-1 beta post-translational processing by a mechanism dependent on anion transport inhibition. Multiple stimuli are likely to operate in vivo to promote IL-1 beta post-translational processing, and anion transport inhibitors such as tenidap that suppress cytokine processing independently of the initiating stimulus thus represent attractive candidates as therapeutic regulators of IL-1 production.