- Email: [email protected]
Phagocyte-specific S100 proteins: a novel group of proinflammatory molecules
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Phagocyte-specific S100 proteins: a novel group of proinflammatory molecules Johannes Roth1,2, Thomas Vogl1, Clemens Sorg1 and Cord Sunderko¨tter1,3 1
Institute of Experimental Dermatology, University of Muenster, Von-Esmarch-Str. 58, D-48149 Muenster, Germany Department of Paediatrics, University of Muenster, Von-Esmarch-Str. 58, D-48149 Muenster, Germany 3 Department of Dermatology and Allergology, University of Ulm, Maienweg 12, 89081 Ulm, Germany 2
Three members of the S100 family of calcium-binding proteins comprise a new group of proinflammatory molecules released by phagocytes. A novel inflammatory syndrome defined by extraordinarily high expression of S100A8 and S100A9 confirmed recent observations in vitro demonstrating a role of these proteins during recruitment of leukocytes. S100A12 directly activates endothelial cells, mononuclear phagocytes and lymphocytes through interaction with the receptor for advanced glycation end products. Thus, these S100-proteins are attractive targets to modulate inflammation. Even although the S100 protein family is the largest group of calcium-binding proteins, the main function of these molecules has not been finally defined [1]. Members of this family are characterized by a relatively low molecular weight and the presence of two calcium-binding sites of the EF-hand type (helix-loop-helix calcium-binding domains). Their tissue-specific expression pattern is of particular interest. Recent work points to prominent proinflammatory functions performed by at least three members of this protein family, S100A8, S100A9 and S100A12, which are mainly expressed by phagocytes. S100A8 and S100A9 and phagocyte activation The initial description of S100A8 and S100A9 already suggested that these proteins are involved in phagocyte biology during inflammation [2]. They attracted major interest in view of their high cytosolic concentration and their high intracellular calcium-binding capacity in phagocytes [3]. Targeted deletion of S100A8 in mice is incompatible with life owing to rapid resorption of homozygous null embryos by day 9.5. It is not clear whether abnormalities in migration of embryonic cells or infiltration by maternal leukocytes are responsible for resorption [4]. However, this model does not enable further analysis of S100A8 or S100A9 biology in phagocytes. By contrast, the recently generated S100A92/2 mouse is viable and fertile. These mice revealed that complexes of S100A8 and S100A9 are involved in migration of phagocytes [5]. This finding was not a complete surprise because calcium-induced complexes of S100A8 and S100A9 were known to modulate Corresponding author: Cord Sunderko¨tter ([email protected]). http://treimm.trends.com
cytoskeletal-membrane interactions during activation of human phagocytes [3]. A S100A8 – S100A9 heterodimer and a calcium-induced (S100A8 – S100A9)2 tetramer seem to be the functionally relevant structures of these proteins (Fig. 1). Monomers, homodimers and trimers are controversially discussed, however, because the existence of those complexes is not supported by current available structural data [6 – 9]. Interaction of S100A8 – S100A9 complexes with microtubules is involved in specific secretion of these proteins by phagocytes. This novel, so-called alternative, pathway of secretion bypasses the classical route, which involves the endoplasmatic reticulum and Golgi complex. It is dependent on the parallel induction of two independent signal pathways in phagocytes. One is the activation of protein kinase C, which is induced by different inflammatory stimuli, such as chemokines or bacterial products. The second pathway presents the elevation of intracellular calcium levels, which is induced in phagocytes by contact with tumor necrosis factor (TNF)-stimulated endothelial cells. By contrast, contact with resting endothelium inhibits secretion of S100A8 and S100A9. This explains why these S100 proteins are released especially at sites of inflammation and why serum concentrations of S100A8– S100A9 correlate specifically with disease activity in different inflammatory disorders, such as rheumatoid arthritis [10]. Subsequent studies have indicated a positive feedback mechanism, according to which these S100 proteins, released by primed phagocytes under inflammatory conditions, would promote further recruitment of leukocytes (Fig. 2). Extracellular complexes of S100A8– S100A9 interact with specific binding sites on endothelial cells. Binding to heparan sulfate, as found for several chemotactic factors, results in accumulation of S100A8 and S100A9 at the endothelial surface [11]. Subsequent interaction of S100A8– S100A9 with novel carboxylated glycans, exclusively expressed on inflammatory activated endothelial cells, promotes adhesion of phagocytes to the vascular endothelium [12]. Simultaneously, S100A9 increases the binding activity of the integrin receptor CD11b – CD18 on phagocytes, thus providing a synergistic action on leukocyte adherence [13]. The alternative pathway of release, as well as effects on endothelial cells, are thus novel molecular targets for modulation of inflammatory reactions. In addition, murine S100A8 exhibits chemotactic activity under some experimental conditions.
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Fig. 1. Complex formation of S100A8 and S100A9. (a) A ribbon representation of a model of the heterodimer of S100A8 (red) and S100A9 (blue). The model was generated by an energy minimization calculation using the known structures of both subunits. The C-terminus of S100A9 is missing because no structural X-ray data are available owing to the high flexibility of this part of the molecule. The coordinates for S100A8 and S100A9 were obtained from Protein Data Bank (www.rcsb. org/pdb/). Asterisks indicate calcium-binding sites of the EF-hand type (helix-loophelix calcium-binding domain). This model fits well with published data obtained by nuclear magnetic resonance (NMR) studies [8]. (b) Summary of calcium (Ca2þ)dependent interactions of human S100A8 (red) and S100A9 (blue) as derived from data obtained by mass spectrometry [9]. S100A9 exists in two isoforms, the smaller one, S100A9* (dark blue), lacks the first five amino acids owing to an alternative translation start site at the second methionine at position five. Both S100A9 isoforms interact with S100A8 in a heterodimer in the absence of calcium. These S100A8– S100A9 heterodimers associate in the presence of calcium to form three (S100A8– S100A9)2 heterotetramers differing in their composition regarding S100A9 isoforms. There is no covalent binding involved in S100A8 –S100A9 interactions.
However, the physiological relevance of this action is not clear because mice overexpressing S100A8 and S100A9 in keratinocytes do not show any infiltration of leukocytes into their skin and also because human S100A8 did not show any chemotactic activity [14,15]. Complexes of human S100A8–S100A9 bind arachidonic acid in a calcium-dependent manner, a property that is not found for the murine homologues [7]. There are controversial reports regarding binding of these complexes of S100A8–S100A9 and arachidonic acid to the scavenger receptor CD36 [11,16] (Fig. 2). Overexpression of S100A8 and S100A9 defines a novel inflammatory disorder The particular relevance of S100A8 and S100A9 for inflammation has been underlined by identification of a new inflammatory disorder just described, whose hallmark is an extraordinarily high abundance of these two http://treimm.trends.com
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molecules. This novel disease is characterized by recurrent infections, hepatosplenomegaly, anaemia, cutaneous vasculitis and evidence of systemic inflammation [17]. As a result of the zinc-binding properties of S100A8 and S100A9, which might be mediated by a HEXXH motif also found in zinc-dependent metalloproteinases, these patients present with an impressive hyperzincaemia. Besides the proinflammatory actions described above, cytotoxic effects of S100A8 –S100A9 might be responsible for the clinical picture of these patients because this complex induces apoptosis and the inhibition of proliferation in several cell types when present in high concentrations [18]. Because there is simultaneous overexpression of both S100A8 and S100A9, the cause is more likely to be a mutation in a common regulatory element rather than mutations of the individual genes [17]. Little is known about the regulation of S100A8 and S100A9, and the regulatory elements of these genes have not been completely identified. Many of the S100 genes, including those for S100A8 and S100A9, are clustered at human chromosome 1q21 in a so-called differentiation complex, which might be controlled by a common regulatory mechanism, probably linked to distinct differentiation stages [1,15]. A mutation in a master regulatory element might thus be a possible cause for the high elevation of two independent gene products. Another possible explanation for the accumulation of S100A8 and S100A9 in these patients could be defective catabolism of the proteins. Because there are no published data about the physiological metabolism of S100A8 and S100A9 it is not clear whether reduced clearance could be due to changes in the proteins or in a specific cellular receptor. S100A12 –RAGE interaction: a novel inflammatory signal pathway S100A12 has extensive homology with S100A8 and S100A9 at protein and DNA level, but does not interact with either of these proteins, and thus has a distinct role during inflammation [19]. Similar to S100A8 and S100A9, S100A12 is secreted by activated human neutrophils [20]. The interaction of S100A12 with the receptor for advanced glycation end products (RAGE) revealed the first plausible receptor– ligand model for an S100 protein [21] (Fig. 2). RAGE is expressed on macrophages, lymphocytes and endothelial cells. Interaction of S100A12 with this receptor triggers cellular activation through the NF-kB pathway and results in synthesis and secretion of proinflammatory mediators. In addition, S100A12 exhibits potent chemotactic activity comparable to other chemoattractants. Consequently, blockade of this interaction succeeded in significant inhibition of delayed-type hypersensitivity and inflammatory colitis in mice [21,22]. In addition, activation of the RAGE pathway is important in wound healing, tumor growth and metastasis, as well as systemic amyloidosis [22– 24]. The relevance of these findings has now been confirmed by high expression of S100A12 in inflammatory bowel disease and systemic vasculitis in humans [25,26]. There are some highly conserved amino acids in the NH2-terminus of several S100 proteins shared with the RAGE ligand, amphoterin [27]. However, the
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Fig. 2. Extracellular effects of S100A8– S100A9 and S100A12 during inflammatory processes. Secretion of S100A8– S100A9 by phagocytes is a tightly controlled process. It depends on two independent signals: activation of protein kinase C (PKC) [signal 1, e.g. by lipopolysaccharide (LPS) or certain cytokines], and elevation of intracellular calcium (Ca2þ) concentrations induced by contact of phagocytes with pre-activated endothelial cells [signal 2, e.g. after endothelial activation with tumor necrosis factor (TNF)]. S100A12 is also released by activated phagocytes but the exact regulation of this process is currently not clear. Extracellular S100A8–S100A9 interacts with endothelial cells by binding to heparan sulfate and specifically carboxylated glycans. Interaction of S100A8– S100A9 with two other surface receptors on endothelial cells [i.e. CD36 and receptor for advanced glycation end products (RAGE)] is still a matter of debate. The intracellular signal pathways and effector mechanisms induced by binding of S100A8–S100A9 to endothelial cells are not defined so far. However, interaction of S100A8–S100A9 with phagocytes upregulates binding activity of the integrin receptor CD11b– CD18, which forms one of the major adhesion pathways of leukocytes to vascular endothelium. Specific receptors or signal pathways responsible for this effect are currently not clear. S100A12 is a specific ligand of the receptor for advanced glycation end products (RAGE) expressed by endothelial cells as well as by leukocytes. Interaction of S100A12 with RAGE activates NFkB-binding activity in these cells, which subsequently induces expression of many proinflammatory molecules, such as various cytokines or adhesion receptors. Thus, release and extracellular functions of these S100 proteins represent a positive feedback mechanism by which phagocytes promote further recruitment of leukocytes to sites of inflammatory processes.
question of whether these conserved structures enable direct binding of other S100 proteins, such as S100A8 and S100A9, to RAGE, is currently unanswered. Conclusion Recently published data provide growing evidence that S100A8, S100A9 and S100A12 comprise a new group of proinflammatory proteins expressed by phagocytes, which are attractive targets for novel diagnostic and therapeutic approaches to manipulate the innate immune system in inflammatory diseases. Acknowledgements The ribbon model of the S100A8 – S100A9 heterodimer was kindly provided by G. Fritz, University of Zu¨rich, Switzerland.
References 1 Heizmann, C.W. (2002) The multifunctional S100 protein family. Methods Mol. Biol. 172, 69 – 80 2 Odink, K. et al. (1987) Two calcium-binding proteins in infiltrate macrophages of rheumatoid arthritis. Nature 330, 80 – 82 3 Roth, J. et al. (1993) MRP8 and MRP14, S-100-like proteins associated with myeloid differentiation, are translocated to plasma membrane and intermediate filaments in a calcium-dependent manner. Blood 82, 1875 – 1883 4 Passey, R.J. et al. (1999) A null mutation in the inflammationassociated S100 protein S100A8 causes early resorption of the mouse embryo. J. Immunol. 163, 2209– 2216 5 Manitz, M.P. et al. (2003) Loss of S100A9 (MRP14) results in a reduced IL-8 induced CD11b surface expression, a polarized microfilament system and a diminished responsiveness upon chemoattractants in vitro. Mol. Cell. Biol. 23, 1034 – 1043 6 Berntzen, H.B. and Fagerhol, M.K. (1990) L1, a major granulocyte protein; isolation of high quantities of its subunits. Scand. J. Clin. Lab. Invest. 50, 769 – 774 http://treimm.trends.com
7 Kerkhoff, C. et al. (1998) Novel insights into structure and function of MRP8 (S100A8) and MRP14 (S100A9). Biochim. Biophys. Acta 1448, 200 – 211 8 Hunter, M.J. and Chazin, W.J. (1998) High level expression and dimer characterization of the S100 EF-hand proteins, migration inhibitory factor-related proteins 8 and 14. J. Biol. Chem. 273, 12427 – 12435 9 Vogl, T. et al. (1999) Calcium-induced noncovalently linked tetramers of MRP8 and MRP14 detected by ultraviolet matrix-assisted laser desorption/ionization mass spectrometry. J. Am. Soc. Mass Spectrom. 10, 1124 – 1130 10 Frosch, M. et al. (2000) Myeloid-related proteins 8 and 14 are specifically secreted during interaction of phagocytes and activated endothelium and are useful markers for monitoring disease activity in pauciarticular-onset juvenile rheumatoid arthritis. Arthritis Rheum. 43, 628 – 637 11 Robinson, M.J. et al. (2002) The S100 family heterodimer, MRP-8/14, binds with high affinity to heparin and heparan sulphate glycosaminoglycans on endothelial cells. J. Biol. Chem. 277, 3658– 3665 12 Srikrishna, G. et al. (2001) Two proteins modulating transendothelial migration of leukocytes recognize novel carboxylated glycans on endothelial cells. J. Immunol. 166, 4678– 4688 13 Newton, R.A. and Hogg, N. (1998) The human S100 protein MRP-14 is a novel activator of the b2 integrin Mac-1 on neutrophils. J. Immunol. 160, 1427– 1435 14 Geczy, C. (1996) Regulation and proinflammatory properties of the chemotactic protein, Cp 10. Biochim. Biophys. Acta 1313, 246– 252 15 Thorey, I.S. et al. (2001) The Ca2þ-binding proteins S100A8 and S100A9 are encoded by novel injury-regulated genes. J. Biol. Chem. 276, 35818 – 35825 16 Kerkhoff, C. et al. (2001) Interaction of S100A8 – S100A9– arachidonic acid complexes with the scavenger receptor CD36 may facilitate fatty acid uptake by endothelial cells. Biochemistry 40, 241– 248 17 Sampson, B. et al. (2002) Hyperzincaemia with hypercalprotectinaemia: a new disorder of zinc metabolism. Identification of calprotectin (S100A8/S100A9) as the zinc binding protein in patients with hyperzincaemia. Lancet 360, 1742 – 1745 18 Yui, S. et al. (1997) Growth inhibitory and apoptosis inducing activities
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of calprotectin derived from inflammatory exudate cells on normal fibroblasts: regulation by metal ions. J. Leukoc. Biol. 61, 50 – 57 Vogl, T. et al. (1999) S100A12 is expressed exclusively by granulocytes and acts independently from MRP8 and MRP14. J. Biol. Chem. 274, 25291 – 25296 Boussac, M. and Garin, J. (2000) Calcium-dependent secretion in human neutrophils: a proteomic approach. Electrophoresis 21, 665 – 672 Hofmann, M.A. et al. (1999) RAGE mediates a novel proinflammatoryaxis: a central cell surface receptor for S100/calgranulin polypeptides. Cell 97, 889– 901 Schmidt, A.M. et al. (2001) The multiligand receptor RAGE as a progression factor amplifying immune and inflammatory responses. J. Clin. Invest. 108, 949 – 955
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23 Yan, S.D. et al. (2000) Receptor-dependent cell stress and amyloid accumulation in systemic amyloidosis. Nat. Med. 6, 643– 651 24 Taguchi, A. et al. (2000) Blockade of RAGE-amphoterin signalling suppresses tumour growth and metastases. Nature 405, 354– 360 25 Foell, D. et al. Neutrophil derived human S100A12 (EN-RAGE) is strongly expressed during chronic active inflammatory bowel disease. Gut (in press) 26 Foell, D. et al. S100A12 (EN-RAGE): A novel serum marker in the monitoring of Kawasaki disease. Lancet (in press) 27 Huttunen, H.J. et al. (2002) Receptor for advanced glycation end products-binding COOH-terminal motif of amphoterin inhibits invasive migration and metastasis. Cancer Res. 62, 4805– 4811 1471-4906/03/$ - see front matter q 2003 Elsevier Science Ltd. All rights reserved. doi:10.1016/S1471-4906(03)00062-0
Tryptase, a novel link between allergic inflammation and fibrosis Francesca Levi-Schaffer and Adrian M. Piliponsky Department of Pharmacology, School of Pharmacy, Faculty of Medicine, The Hebrew University of Jerusalem, Jerusalem, Israel
Allergy is a complex inflammatory disease, the etiology of which is well defined. It has recently been proposed that eosinophil, mast-cell and fibroblast interactions contribute to allergy perpetuation. Moreover, mast-cellderived tryptase might act as a link among these cells. This hypothesis is supported by two recent papers that show that tryptase, seemingly through the proteaseactivated receptor-2, mediates eosinophil infiltration in the airways and fibroblast proliferation that depends on both cyclooxygenase and prostaglandin synthesis. Allergy is a complex disease characterized by a specific pattern of inflammation that is largely driven by immunoglobulin (Ig)E-dependent mechanisms triggered in mast cells. Mast-cell mediators contribute to the development of allergic inflammation by supporting the influx of inflammatory cells, such as eosinophils, macrophages, lymphocytes and basophils [1]. Eosinophils have the ability to persist in the inflamed tissue and to release membrane phospholipid metabolites, cytokines, reactive oxygen species and basic granule proteins; cytokines and reactive oxygen species are cytotoxic for airway epithelium. Altogether this has led to a consensus that eosinophils are major effector cells for tissue inflammation [2]. However, new evidence indicates that fibroblasts are also important effector cells of the inflammatory response because of their ability to respond to different stimuli and to release mediators that modulate mast-cell and eosinophil functionality. Therefore, it is apparent that no single inflammatory cell is exclusively responsible for the development of the allergic inflammation and its possible consequences. Yet, some cells more than others seem to contribute to the allergic inflammation process. Corresponding author: Francesca Levi-Schaffer ([email protected]). http://treimm.trends.com
…no single inflammatory cell is exclusively responsible for the development of the allergic inflammation and its possible consequences. In this Research Focus, we will discuss recent work, focusing on the interactions among what we consider the main cellular players of the allergic inflammatory response: mast cells, eosinophils and fibroblasts. Interactions between mast cells and eosinophils Recent studies suggest that an important cross-talk occurs between mast cells and eosinophils. We have demonstrated that eosinophil survival is enhanced by rat peritoneal mast-cell-derived tumor necrosis factor-a (TNF-a) that induces the autocrine production of granulocyte –monocyte-colony stimulating factor (GM-CSF) [3 – 4] (Fig. 1a). Moreover, we have recently shown that mastcell-derived tryptase is the primary mediator that induces interleukin-6 (IL-6) and IL-8 release in eosinophils by mitogen-activated protein kinase (MAPK) and activator protein-1 (AP-1) pathways [5] (Fig. 2b). Mast cells can also be influenced by eosinophil constituents. For example, eosinophil major basic protein (MBP) induces histamine and prostaglandin D2 (PGD2) from human lung and cordblood-derived mast cells through an IgE-independent mechanism [6] (Fig. 1b). Furthermore, MBP reactivates IgE-challenged rat peritoneal mast cells in an in vitro system that mimics the development of the late phase of the allergic process [7]. This evidence indicates that mast-cell and eosinophil products could prolong and even intensify allergic
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Phagocyte-specific S100 proteins: a novel group of proinflammatory molecules Johannes Roth1,2, Thomas Vogl1, Clemens Sorg1 and Cord Sunderko¨tter1,3 1
Institute of Experimental Dermatology, University of Muenster, Von-Esmarch-Str. 58, D-48149 Muenster, Germany Department of Paediatrics, University of Muenster, Von-Esmarch-Str. 58, D-48149 Muenster, Germany 3 Department of Dermatology and Allergology, University of Ulm, Maienweg 12, 89081 Ulm, Germany 2
Three members of the S100 family of calcium-binding proteins comprise a new group of proinflammatory molecules released by phagocytes. A novel inflammatory syndrome defined by extraordinarily high expression of S100A8 and S100A9 confirmed recent observations in vitro demonstrating a role of these proteins during recruitment of leukocytes. S100A12 directly activates endothelial cells, mononuclear phagocytes and lymphocytes through interaction with the receptor for advanced glycation end products. Thus, these S100-proteins are attractive targets to modulate inflammation. Even although the S100 protein family is the largest group of calcium-binding proteins, the main function of these molecules has not been finally defined [1]. Members of this family are characterized by a relatively low molecular weight and the presence of two calcium-binding sites of the EF-hand type (helix-loop-helix calcium-binding domains). Their tissue-specific expression pattern is of particular interest. Recent work points to prominent proinflammatory functions performed by at least three members of this protein family, S100A8, S100A9 and S100A12, which are mainly expressed by phagocytes. S100A8 and S100A9 and phagocyte activation The initial description of S100A8 and S100A9 already suggested that these proteins are involved in phagocyte biology during inflammation [2]. They attracted major interest in view of their high cytosolic concentration and their high intracellular calcium-binding capacity in phagocytes [3]. Targeted deletion of S100A8 in mice is incompatible with life owing to rapid resorption of homozygous null embryos by day 9.5. It is not clear whether abnormalities in migration of embryonic cells or infiltration by maternal leukocytes are responsible for resorption [4]. However, this model does not enable further analysis of S100A8 or S100A9 biology in phagocytes. By contrast, the recently generated S100A92/2 mouse is viable and fertile. These mice revealed that complexes of S100A8 and S100A9 are involved in migration of phagocytes [5]. This finding was not a complete surprise because calcium-induced complexes of S100A8 and S100A9 were known to modulate Corresponding author: Cord Sunderko¨tter ([email protected]). http://treimm.trends.com
cytoskeletal-membrane interactions during activation of human phagocytes [3]. A S100A8 – S100A9 heterodimer and a calcium-induced (S100A8 – S100A9)2 tetramer seem to be the functionally relevant structures of these proteins (Fig. 1). Monomers, homodimers and trimers are controversially discussed, however, because the existence of those complexes is not supported by current available structural data [6 – 9]. Interaction of S100A8 – S100A9 complexes with microtubules is involved in specific secretion of these proteins by phagocytes. This novel, so-called alternative, pathway of secretion bypasses the classical route, which involves the endoplasmatic reticulum and Golgi complex. It is dependent on the parallel induction of two independent signal pathways in phagocytes. One is the activation of protein kinase C, which is induced by different inflammatory stimuli, such as chemokines or bacterial products. The second pathway presents the elevation of intracellular calcium levels, which is induced in phagocytes by contact with tumor necrosis factor (TNF)-stimulated endothelial cells. By contrast, contact with resting endothelium inhibits secretion of S100A8 and S100A9. This explains why these S100 proteins are released especially at sites of inflammation and why serum concentrations of S100A8– S100A9 correlate specifically with disease activity in different inflammatory disorders, such as rheumatoid arthritis [10]. Subsequent studies have indicated a positive feedback mechanism, according to which these S100 proteins, released by primed phagocytes under inflammatory conditions, would promote further recruitment of leukocytes (Fig. 2). Extracellular complexes of S100A8– S100A9 interact with specific binding sites on endothelial cells. Binding to heparan sulfate, as found for several chemotactic factors, results in accumulation of S100A8 and S100A9 at the endothelial surface [11]. Subsequent interaction of S100A8– S100A9 with novel carboxylated glycans, exclusively expressed on inflammatory activated endothelial cells, promotes adhesion of phagocytes to the vascular endothelium [12]. Simultaneously, S100A9 increases the binding activity of the integrin receptor CD11b – CD18 on phagocytes, thus providing a synergistic action on leukocyte adherence [13]. The alternative pathway of release, as well as effects on endothelial cells, are thus novel molecular targets for modulation of inflammatory reactions. In addition, murine S100A8 exhibits chemotactic activity under some experimental conditions.
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Fig. 1. Complex formation of S100A8 and S100A9. (a) A ribbon representation of a model of the heterodimer of S100A8 (red) and S100A9 (blue). The model was generated by an energy minimization calculation using the known structures of both subunits. The C-terminus of S100A9 is missing because no structural X-ray data are available owing to the high flexibility of this part of the molecule. The coordinates for S100A8 and S100A9 were obtained from Protein Data Bank (www.rcsb. org/pdb/). Asterisks indicate calcium-binding sites of the EF-hand type (helix-loophelix calcium-binding domain). This model fits well with published data obtained by nuclear magnetic resonance (NMR) studies [8]. (b) Summary of calcium (Ca2þ)dependent interactions of human S100A8 (red) and S100A9 (blue) as derived from data obtained by mass spectrometry [9]. S100A9 exists in two isoforms, the smaller one, S100A9* (dark blue), lacks the first five amino acids owing to an alternative translation start site at the second methionine at position five. Both S100A9 isoforms interact with S100A8 in a heterodimer in the absence of calcium. These S100A8– S100A9 heterodimers associate in the presence of calcium to form three (S100A8– S100A9)2 heterotetramers differing in their composition regarding S100A9 isoforms. There is no covalent binding involved in S100A8 –S100A9 interactions.
However, the physiological relevance of this action is not clear because mice overexpressing S100A8 and S100A9 in keratinocytes do not show any infiltration of leukocytes into their skin and also because human S100A8 did not show any chemotactic activity [14,15]. Complexes of human S100A8–S100A9 bind arachidonic acid in a calcium-dependent manner, a property that is not found for the murine homologues [7]. There are controversial reports regarding binding of these complexes of S100A8–S100A9 and arachidonic acid to the scavenger receptor CD36 [11,16] (Fig. 2). Overexpression of S100A8 and S100A9 defines a novel inflammatory disorder The particular relevance of S100A8 and S100A9 for inflammation has been underlined by identification of a new inflammatory disorder just described, whose hallmark is an extraordinarily high abundance of these two http://treimm.trends.com
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molecules. This novel disease is characterized by recurrent infections, hepatosplenomegaly, anaemia, cutaneous vasculitis and evidence of systemic inflammation [17]. As a result of the zinc-binding properties of S100A8 and S100A9, which might be mediated by a HEXXH motif also found in zinc-dependent metalloproteinases, these patients present with an impressive hyperzincaemia. Besides the proinflammatory actions described above, cytotoxic effects of S100A8 –S100A9 might be responsible for the clinical picture of these patients because this complex induces apoptosis and the inhibition of proliferation in several cell types when present in high concentrations [18]. Because there is simultaneous overexpression of both S100A8 and S100A9, the cause is more likely to be a mutation in a common regulatory element rather than mutations of the individual genes [17]. Little is known about the regulation of S100A8 and S100A9, and the regulatory elements of these genes have not been completely identified. Many of the S100 genes, including those for S100A8 and S100A9, are clustered at human chromosome 1q21 in a so-called differentiation complex, which might be controlled by a common regulatory mechanism, probably linked to distinct differentiation stages [1,15]. A mutation in a master regulatory element might thus be a possible cause for the high elevation of two independent gene products. Another possible explanation for the accumulation of S100A8 and S100A9 in these patients could be defective catabolism of the proteins. Because there are no published data about the physiological metabolism of S100A8 and S100A9 it is not clear whether reduced clearance could be due to changes in the proteins or in a specific cellular receptor. S100A12 –RAGE interaction: a novel inflammatory signal pathway S100A12 has extensive homology with S100A8 and S100A9 at protein and DNA level, but does not interact with either of these proteins, and thus has a distinct role during inflammation [19]. Similar to S100A8 and S100A9, S100A12 is secreted by activated human neutrophils [20]. The interaction of S100A12 with the receptor for advanced glycation end products (RAGE) revealed the first plausible receptor– ligand model for an S100 protein [21] (Fig. 2). RAGE is expressed on macrophages, lymphocytes and endothelial cells. Interaction of S100A12 with this receptor triggers cellular activation through the NF-kB pathway and results in synthesis and secretion of proinflammatory mediators. In addition, S100A12 exhibits potent chemotactic activity comparable to other chemoattractants. Consequently, blockade of this interaction succeeded in significant inhibition of delayed-type hypersensitivity and inflammatory colitis in mice [21,22]. In addition, activation of the RAGE pathway is important in wound healing, tumor growth and metastasis, as well as systemic amyloidosis [22– 24]. The relevance of these findings has now been confirmed by high expression of S100A12 in inflammatory bowel disease and systemic vasculitis in humans [25,26]. There are some highly conserved amino acids in the NH2-terminus of several S100 proteins shared with the RAGE ligand, amphoterin [27]. However, the
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PKC Heperan sulfate
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Expression of pro-inflammatory cytokines and adhesion receptors TRENDS in Immunology
Fig. 2. Extracellular effects of S100A8– S100A9 and S100A12 during inflammatory processes. Secretion of S100A8– S100A9 by phagocytes is a tightly controlled process. It depends on two independent signals: activation of protein kinase C (PKC) [signal 1, e.g. by lipopolysaccharide (LPS) or certain cytokines], and elevation of intracellular calcium (Ca2þ) concentrations induced by contact of phagocytes with pre-activated endothelial cells [signal 2, e.g. after endothelial activation with tumor necrosis factor (TNF)]. S100A12 is also released by activated phagocytes but the exact regulation of this process is currently not clear. Extracellular S100A8–S100A9 interacts with endothelial cells by binding to heparan sulfate and specifically carboxylated glycans. Interaction of S100A8– S100A9 with two other surface receptors on endothelial cells [i.e. CD36 and receptor for advanced glycation end products (RAGE)] is still a matter of debate. The intracellular signal pathways and effector mechanisms induced by binding of S100A8–S100A9 to endothelial cells are not defined so far. However, interaction of S100A8–S100A9 with phagocytes upregulates binding activity of the integrin receptor CD11b– CD18, which forms one of the major adhesion pathways of leukocytes to vascular endothelium. Specific receptors or signal pathways responsible for this effect are currently not clear. S100A12 is a specific ligand of the receptor for advanced glycation end products (RAGE) expressed by endothelial cells as well as by leukocytes. Interaction of S100A12 with RAGE activates NFkB-binding activity in these cells, which subsequently induces expression of many proinflammatory molecules, such as various cytokines or adhesion receptors. Thus, release and extracellular functions of these S100 proteins represent a positive feedback mechanism by which phagocytes promote further recruitment of leukocytes to sites of inflammatory processes.
question of whether these conserved structures enable direct binding of other S100 proteins, such as S100A8 and S100A9, to RAGE, is currently unanswered. Conclusion Recently published data provide growing evidence that S100A8, S100A9 and S100A12 comprise a new group of proinflammatory proteins expressed by phagocytes, which are attractive targets for novel diagnostic and therapeutic approaches to manipulate the innate immune system in inflammatory diseases. Acknowledgements The ribbon model of the S100A8 – S100A9 heterodimer was kindly provided by G. Fritz, University of Zu¨rich, Switzerland.
References 1 Heizmann, C.W. (2002) The multifunctional S100 protein family. Methods Mol. Biol. 172, 69 – 80 2 Odink, K. et al. (1987) Two calcium-binding proteins in infiltrate macrophages of rheumatoid arthritis. Nature 330, 80 – 82 3 Roth, J. et al. (1993) MRP8 and MRP14, S-100-like proteins associated with myeloid differentiation, are translocated to plasma membrane and intermediate filaments in a calcium-dependent manner. Blood 82, 1875 – 1883 4 Passey, R.J. et al. (1999) A null mutation in the inflammationassociated S100 protein S100A8 causes early resorption of the mouse embryo. J. Immunol. 163, 2209– 2216 5 Manitz, M.P. et al. (2003) Loss of S100A9 (MRP14) results in a reduced IL-8 induced CD11b surface expression, a polarized microfilament system and a diminished responsiveness upon chemoattractants in vitro. Mol. Cell. Biol. 23, 1034 – 1043 6 Berntzen, H.B. and Fagerhol, M.K. (1990) L1, a major granulocyte protein; isolation of high quantities of its subunits. Scand. J. Clin. Lab. Invest. 50, 769 – 774 http://treimm.trends.com
7 Kerkhoff, C. et al. (1998) Novel insights into structure and function of MRP8 (S100A8) and MRP14 (S100A9). Biochim. Biophys. Acta 1448, 200 – 211 8 Hunter, M.J. and Chazin, W.J. (1998) High level expression and dimer characterization of the S100 EF-hand proteins, migration inhibitory factor-related proteins 8 and 14. J. Biol. Chem. 273, 12427 – 12435 9 Vogl, T. et al. (1999) Calcium-induced noncovalently linked tetramers of MRP8 and MRP14 detected by ultraviolet matrix-assisted laser desorption/ionization mass spectrometry. J. Am. Soc. Mass Spectrom. 10, 1124 – 1130 10 Frosch, M. et al. (2000) Myeloid-related proteins 8 and 14 are specifically secreted during interaction of phagocytes and activated endothelium and are useful markers for monitoring disease activity in pauciarticular-onset juvenile rheumatoid arthritis. Arthritis Rheum. 43, 628 – 637 11 Robinson, M.J. et al. (2002) The S100 family heterodimer, MRP-8/14, binds with high affinity to heparin and heparan sulphate glycosaminoglycans on endothelial cells. J. Biol. Chem. 277, 3658– 3665 12 Srikrishna, G. et al. (2001) Two proteins modulating transendothelial migration of leukocytes recognize novel carboxylated glycans on endothelial cells. J. Immunol. 166, 4678– 4688 13 Newton, R.A. and Hogg, N. (1998) The human S100 protein MRP-14 is a novel activator of the b2 integrin Mac-1 on neutrophils. J. Immunol. 160, 1427– 1435 14 Geczy, C. (1996) Regulation and proinflammatory properties of the chemotactic protein, Cp 10. Biochim. Biophys. Acta 1313, 246– 252 15 Thorey, I.S. et al. (2001) The Ca2þ-binding proteins S100A8 and S100A9 are encoded by novel injury-regulated genes. J. Biol. Chem. 276, 35818 – 35825 16 Kerkhoff, C. et al. (2001) Interaction of S100A8 – S100A9– arachidonic acid complexes with the scavenger receptor CD36 may facilitate fatty acid uptake by endothelial cells. Biochemistry 40, 241– 248 17 Sampson, B. et al. (2002) Hyperzincaemia with hypercalprotectinaemia: a new disorder of zinc metabolism. Identification of calprotectin (S100A8/S100A9) as the zinc binding protein in patients with hyperzincaemia. Lancet 360, 1742 – 1745 18 Yui, S. et al. (1997) Growth inhibitory and apoptosis inducing activities
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of calprotectin derived from inflammatory exudate cells on normal fibroblasts: regulation by metal ions. J. Leukoc. Biol. 61, 50 – 57 Vogl, T. et al. (1999) S100A12 is expressed exclusively by granulocytes and acts independently from MRP8 and MRP14. J. Biol. Chem. 274, 25291 – 25296 Boussac, M. and Garin, J. (2000) Calcium-dependent secretion in human neutrophils: a proteomic approach. Electrophoresis 21, 665 – 672 Hofmann, M.A. et al. (1999) RAGE mediates a novel proinflammatoryaxis: a central cell surface receptor for S100/calgranulin polypeptides. Cell 97, 889– 901 Schmidt, A.M. et al. (2001) The multiligand receptor RAGE as a progression factor amplifying immune and inflammatory responses. J. Clin. Invest. 108, 949 – 955
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23 Yan, S.D. et al. (2000) Receptor-dependent cell stress and amyloid accumulation in systemic amyloidosis. Nat. Med. 6, 643– 651 24 Taguchi, A. et al. (2000) Blockade of RAGE-amphoterin signalling suppresses tumour growth and metastases. Nature 405, 354– 360 25 Foell, D. et al. Neutrophil derived human S100A12 (EN-RAGE) is strongly expressed during chronic active inflammatory bowel disease. Gut (in press) 26 Foell, D. et al. S100A12 (EN-RAGE): A novel serum marker in the monitoring of Kawasaki disease. Lancet (in press) 27 Huttunen, H.J. et al. (2002) Receptor for advanced glycation end products-binding COOH-terminal motif of amphoterin inhibits invasive migration and metastasis. Cancer Res. 62, 4805– 4811 1471-4906/03/$ - see front matter q 2003 Elsevier Science Ltd. All rights reserved. doi:10.1016/S1471-4906(03)00062-0
Tryptase, a novel link between allergic inflammation and fibrosis Francesca Levi-Schaffer and Adrian M. Piliponsky Department of Pharmacology, School of Pharmacy, Faculty of Medicine, The Hebrew University of Jerusalem, Jerusalem, Israel
Allergy is a complex inflammatory disease, the etiology of which is well defined. It has recently been proposed that eosinophil, mast-cell and fibroblast interactions contribute to allergy perpetuation. Moreover, mast-cellderived tryptase might act as a link among these cells. This hypothesis is supported by two recent papers that show that tryptase, seemingly through the proteaseactivated receptor-2, mediates eosinophil infiltration in the airways and fibroblast proliferation that depends on both cyclooxygenase and prostaglandin synthesis. Allergy is a complex disease characterized by a specific pattern of inflammation that is largely driven by immunoglobulin (Ig)E-dependent mechanisms triggered in mast cells. Mast-cell mediators contribute to the development of allergic inflammation by supporting the influx of inflammatory cells, such as eosinophils, macrophages, lymphocytes and basophils [1]. Eosinophils have the ability to persist in the inflamed tissue and to release membrane phospholipid metabolites, cytokines, reactive oxygen species and basic granule proteins; cytokines and reactive oxygen species are cytotoxic for airway epithelium. Altogether this has led to a consensus that eosinophils are major effector cells for tissue inflammation [2]. However, new evidence indicates that fibroblasts are also important effector cells of the inflammatory response because of their ability to respond to different stimuli and to release mediators that modulate mast-cell and eosinophil functionality. Therefore, it is apparent that no single inflammatory cell is exclusively responsible for the development of the allergic inflammation and its possible consequences. Yet, some cells more than others seem to contribute to the allergic inflammation process. Corresponding author: Francesca Levi-Schaffer ([email protected]). http://treimm.trends.com
…no single inflammatory cell is exclusively responsible for the development of the allergic inflammation and its possible consequences. In this Research Focus, we will discuss recent work, focusing on the interactions among what we consider the main cellular players of the allergic inflammatory response: mast cells, eosinophils and fibroblasts. Interactions between mast cells and eosinophils Recent studies suggest that an important cross-talk occurs between mast cells and eosinophils. We have demonstrated that eosinophil survival is enhanced by rat peritoneal mast-cell-derived tumor necrosis factor-a (TNF-a) that induces the autocrine production of granulocyte –monocyte-colony stimulating factor (GM-CSF) [3 – 4] (Fig. 1a). Moreover, we have recently shown that mastcell-derived tryptase is the primary mediator that induces interleukin-6 (IL-6) and IL-8 release in eosinophils by mitogen-activated protein kinase (MAPK) and activator protein-1 (AP-1) pathways [5] (Fig. 2b). Mast cells can also be influenced by eosinophil constituents. For example, eosinophil major basic protein (MBP) induces histamine and prostaglandin D2 (PGD2) from human lung and cordblood-derived mast cells through an IgE-independent mechanism [6] (Fig. 1b). Furthermore, MBP reactivates IgE-challenged rat peritoneal mast cells in an in vitro system that mimics the development of the late phase of the allergic process [7]. This evidence indicates that mast-cell and eosinophil products could prolong and even intensify allergic