IL-17 stimulates the production and expression of proinflammatory cytokines, IL-beta and TNF-alpha, by human macrophages

Author(s): Jovanovic DV, Di Battista JA, Martel-Pelletier J, Jolicoeur FC, He Y, et al.


IL-17 is a newly described, T cell-derived cytokine with ill-defined physiologic properties. As such, we examined the release of proinflammatory mediators by human macrophages in response to recombinant human (rh) IL-17. IL-1β and TNF-α expression and synthesis were up-regulated by rhIL-17 in a dose (ED50 was 50 ± 9 ng/ml)- and time-dependent fashion, with cytokine accumulation reaching a zenith after 9 h. Release of IL-6, PGE2, IL-10, IL-12, IL-1R antagonist, and stromelysin was also stimulated by rhIL-17. IL-1β and TNF-α mRNA expression levels were controlled by rhIL-17 in a complex manner with an initial 30-min inhibitory phase, and then up-regulation beginning at 1 h and reaching a plateau at about 3 h. The latter expression pattern closely mirrored the nuclear accumulation of the transcription factor nuclear factor-κB. cAMP mimetics isobutyl-1-methylxanthine (IBMX), forskolin, PGE2, and cholera toxin reversed rhIL-17-induced release of TNF-α, but had no consistent effect on induced IL-1β synthesis. Induced release of TNF-α was also inhibited by serine/threonine protein kinase inhibitors KT-5720 (protein kinase A) and Calphostin C (protein kinase C), mitogen-activated protein kinase kinase inhibitor PD098059, and a nonspecific tyrosine kinase inhibitor, genistein. Calphostin C alone abrogated the rhIL-17-induced release of IL-1β. The antiinflammatory cytokines IL-4 (p < 0.01) and IL-10 (p < 0.02) completely reversed rhIL-17-stimulated IL-1β release, while IL-13 and TGF-β2 were partially effective (59 and 43% diminution, respectively). IL-10 exerted a significant suppressive effect on IL-17-induced TNF-α release (99%, p < 0.02), while the inhibitory effects of IL-4, IL-13, and TGF-β2 on TNF-α secretion were partial (48, 10, and 23%, respectively). The data suggest a pivotal role for IL-17 in initiating and/or sustaining an inflammatory response. Macrophages are migratory phagocytic cells that play a central role in the host defense system. They are essential APCs that process Ags and present them to T cells, initiating an immune response. Macrophages are also an important source of cytokines, as well as of costimulatory molecules that play a crucial role in the efficient activation of T and B cells (1). Cytokines mediate so-called cross-talk between macrophages and Th cells, and are now widely accepted as the major factor involved in the communication between T cells, macrophages, and other immune cells in the course of an immune response (2). Monocyte/macrophage-produced cytokines, including TNF-α, IL-1, IL-6, IL-8, IL-10, IL-12, granulocyte-macrophage CSF, and IFN-γ, may affect differentiation toward Th1 and Th2 pathways in their own specific ways (2). Activated Th1 and Th2 CD4+ lymphocytes also produce cytokines that, with cellular hormones of other cellular sources, alter macrophage function by causing selective changes in macrophage gene expression (3). The response of macrophages to any given stimulus is dependent upon the balance of exogenous ambient cytokines. Th1-associated cytokines, such as IL-2 and IFN-γ, are considered macrophage activators (4), whereas cytokines secreted by Th2 lymphocytes, such as IL-4, IL-10, IL-13, and TGF-β, exhibit suppressive activities on macrophage functions and antagonize the effect of Th1-secreted cytokines (5, 6). All of this may reflect the complexity of the feedback regulatory mechanisms that occur through the cytokine network in macrophages. Human IL-17 is a 20- to 30-kDa homodimeric variably glycosylated polypeptide secreted by CD4+ activated memory (CD45+RO+) T cells (7). There appears to be a ubiquitous nature to the tissue distribution of the IL-17R (8), although it is not yet known whether all cells expressing the IL-17R respond to IL-17. Murine rIL-17 exhibits biologic activity with a variety of cell types, including induction of IL-6 (8) and IL-8 (7) secretion from stromal cell elements such as fibroblasts, endothelial, and epithelial cells. IL-17 costimulates T cell proliferation induced by suboptimal amounts of PHA (9), increases surface expression of intracellular adhesion molecule-1 (ICAM-1)2 by human fibroblasts (7), and stimulates the secretion of PGE2. IL-17 up-regulates a number of gene products involved in cell activation, growth, and proliferation (10, 11), including the proinflammatory cytokines IL-1β, TNF-α, and IL-6. To date, there have been no reports alluding to a role for IL-17 in terms of macrophage activation. Fossiez et al. (9) found that hIL-17 did not alter the secretion of proinflammatory cytokines by blood monocytes, suggesting a limited proinflammatory role for IL-17 on T cell-driven inflammatory processes. Cytokines have no effect on nonactivated monocytes in vitro, and the latter observation may explain why circulating monocytes are not activated routinely by circulating cytokines (12). Freshly isolated monocytes and freshly seeded monocytes produce considerably less TNF-α and IL-6 than macrophages derived by monocyte adherence (13). It is known that adherence activates macrophages, inducing high steady state levels of mRNA for TNF-α, c-fos, and monocyte/macrophage CSF-1 (14), thereby enhancing their response to stimuli (15). The attachment occurs in vivo when the monocytes adhere to the vascular endothelium and migrate into the extravascular space (16), providing an activation signal that leads to biologic activation of the macrophages. Knowing that macrophages and T cells are indissolubly linked in the activation of the immune system as a whole (1), and that all cells express the IL-17R (8), we have tested the effect of rhIL-17, the newly described T cell-derived cytokines, on macrophage function. In this work, we report for the first time the stimulation by rhIL-17 of IL-1β and TNF-α mRNA expression and synthesis, using activated human monocytes in primary culture. In addition, we show that the cytokine up-regulates IL-6, IL-10, IL-1Ra, PGE2, and stromelysin (MMP-3) release, but does not induce nitric oxide (NO) synthesis. Using cell-permeable chemical inhibitors of protein kinases, we explored signaling pathways that may possibly mediate IL-17 action. We used the human peripheral blood monocytes isolated and activated by adherence, to examine the effect of the second activating signal on these cells provided by IL-17.

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