TLR7 and TLR8 ligands and antiphospholipid antibodies show synergistic effects on the induction of IL-1β and caspase-1 in monocytes and dendritic cells

TLRs represent the first line of defense against invading pathogens in the innate immune system. Certain cytokines are important mediators and essentially necessary to assure an appropriately regulated immune response. Recent data gave initial evidence that IL-1 beta is one of the most relevant members of these regulating cytokines. We investigated the induction of IL-1 beta production in monocytes and pDCs stimulated with ligands for TLR7 and TLR8 and with antiphospholipid antibodies (aPL). Using human monocytes and pDCs for stimulation with specific TLR7 and TLR8 ligands such as resiquimod (R848) and single stranded RNA (RNA42) as well as with a human monoclonal aPL HL5B resulted in a specific upregulation of IL-1 beta mRNA and protein in these cells. Determination of expression-levels using real-time RT-PCR showed significantly augmented TLR-dependent IL-1 beta and caspase-1 expression. This increase could be substantially enhanced by adding the monoclonal aPL HL5B. To demonstrate the direct dependency between TLR stimulation and IL-1 beta production, specific TLR inhibitors were applied and the IL-1 beta and caspase-1 secretion could be explicitly decreased. The respective protein levels were determined using Western Blot, FACS analysis or ELISA assays. In conclusion we demonstrated that the downstream signaling pathway of TLR7 and TLR8 in Monocytes and pDCs after stimulation with specific ligands included not only the secretion of cytokines such as TNF alpha and IL-1 beta but as well the activation of necessary regulating proteins like caspase-1. APL seem to enforce this process hinting that endogenous stimulation of TLRs in the Anti phospholipid Syndrome (APS) patients resulted ill enhanced secretion of proinflammatory cytokines. (C) 2009 Elsevier GmbH. All rights reserved.