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Acute Phase Levels of C-Reactive Protein Enhance IL-lp and 11-lra Production by Human Blood Monocytes but Inhibit 11-10 and IL-1 raProduction by Alveolar Macrophages' Charles A. Pue,* Richard F. Mortensen,+ Clay B. Marsh,* Heidi A. Pope,* and Mark D. Wewers2* C-reactive protein (CRP), the major acute phase protein in humans, was purified free of endotoxin (LPS) (e10 pg of LPS/mg of purified CRP) and evaluated for its ability to modulate LPS-induced production of IL-lp and 11-1 receptor antagonist (11-lra) from human PBMC and lung macrophages. PBMC (5 X 106/ml) released low levels of 11-lp in response to either CRP (250 pg/ml) or LPS (100 ng/ml) for 18 h (0.3 f 0.1 and 1.5 f 0.7 ng/ml, respectively). However, when CRP (250 wg/ml) and LPS (100 ng/ml) were combined, PBMC released 9.7 f 2.9 ng/ml ( p < 0.001vsLPS alone). This synergy was removed by immunodepletion of CRP before stimulation. With respect to 11-1ra, although CRP induced 11-1ra production fromPBMC (0.8 2 0.3 ng/ml control, 2.6 f 1.3 ng/ml with CRP), CRP did not synergize with LPS for 11-1ra production (15.0 f 0.7 ng/ml LPS alone vs 15.4 k 1.4 ng/ml LPS and CRP). In contrast, lung macrophages respondedto CRP quite differently than PBMC. Macrophages (106/ml) were not stimulated to produce 11-1 or 11-1ra by CRP alone. When combined with LPS, CRP inhibited 11-1p and 11-lra release induced by LPS (for IL-10 release, LPS induced 3.0 +. 1.7 ng/ml vs 1.1 -C 0.4 for combined LPS and CRP; for IL-lra release, LPS induced 12.9 -t 2.3 ng/ml vs 7.6 k 2.3 ng/ml for combined LPS and CRP). These data suggest that acute phase levels of CRP may have divergent effects depending on the target population. CRP may belargely proinflammatory to blood monocytes responding to LPS since N-lP production is augmented over 11-1ra production. However, in tissue compartments the effects of The lournal of Immunology, CRP may be largely immunosuppressive to LPS-induced tissue macrophage 11-1/3 production. 1996,156: 1594-1600.
T
he systemic acute phase response involves a complex cascade of synthesis and secretion of both cytokines and hepatocyte-derived acute phase proteins. The focus of these events is to remove the inciting stimulus; however, this response may become overexuberant and result in tissue injury and damage to the host. For example, the damage manifest in the sepsis syndrome is shock and diffuse organ dysfunction (1). In human sepsis, there is an increase in production of the inflammatory cytokines IL-lp, TNF-a, IL-6, and IL-I receptor antagonist (IL-lra)3 (2-4). These cytokines modulate the degree of inflammation in a number of ways. For example, both IL-IP and TNF-a activate neutrophil oxidant production (5). They also signal mesenchymal cells to proliferate andto release collagenase and prostaglandin E, (6-9); furthermore, IL-16 and TNF-a augment neutrophil recruitment via IL-8 induction from parenchymal cells (10-12). In contrast, ILIra, which is also up-regulated with sepsis, modulates IL-1 effects by blocking IL-1 receptor activation (13).
Departments of 'Internal Medicine and 'Microbiology, the Ohio State University, Columbus, OH 43210. Received for publication February 6 , 1995. Accepted for publication December 6, 1995. The costs of publication of this article were defrayed in part by the payment of pagecharges.This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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This work was supported by National Institutes of Health Grant HL40871 and an American Lung Association Fellowship. Address correspondence and reprint requests to Dr.Mark D. Wewers, Division of Pulmonary and Critical Care Medicine, Ohio State University, 1654 Upham Drive, N325 Means Hall, Columbus, O H 43210-1228.
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Abbreviations used in this paper: IL-lra, IL-1 receptor antagonist; CRP, C-reactive protein; D-PBS, Dulbecco's PBS; PC, phosphorylcholine; LBP, LPS-binding protein. Copyright 0 1996 by The American Association of Immunologists
Hepatocytes respond to exogenous IL-1 and IL-6 in the blood by accelerating the new synthesis and production of acute phase proteins, especially the prototypical human acute phase reactant, Creactive protein (CRP) (14-18). The magnitude of this increase correlates with the extent of tissue involvement; therefore, the level of CRP in the blood is used extensively as a gauge of inflammation. In severe illness, plasma levels can increase from baseline levels of 200 pg/ml (19, 20). Patients with sepsis-induced adult respiratory distress syndrome have elevated levels of CRP in the alveolar lining fluid; therefore, elevated CRP concentrations are present at tissue macrophage sites (21). Although a precise or unique function has not been ascribed to CRP, its well documented activities such as complement activation, opsonization, and selective deposition at sites of tissue damage (reviewed in Ref. 22) are consistent with a proinflammatory designation. Nonetheless, the interaction of CRP with phagocytic leukocytes may modify this cellular component of inflammation. Specific receptors for CRP have been identified on human monocytes and mouse macrophages (23-27); however, little is known about the effect that CRP exerts on cytokine production by this cell lineage. CRP, or peptides derived from CRP, have been reported to induce IL-1 (28-33); however, the mechanism and significance of this induction and the potential interaction with LPS has not been examined. To evaluate the hypothesis that CRP actively modulates cytokine production by mononuclear phagocytes, we examined the influence of purified, LPS-free human CRP on the production of IL-I@and IL-lra by both blood mononuclear cells and alveolar macrophages in culture. The findings indicate that CRP alters production of the two cytokines in a manner that depends upon the cell source. 0022-1 767/96/$02.00
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Materials and Methods
Table I.
Lipopolysaccharide levels in CRP samples "
Purification of PBMC
"
~
~
~
LPS a Venous blood from normal, healthy adult volunteers was obtained in 15 Source of CRP (ngimg of CRP) U/ml sodium heparin (Elkins-Shinn, Inc., Cherry Hill, NJ). The volunteers Commercial source Ab 8.0 were free of nonsteroidal anti-inflammatory agents for at least 72 h. PBMC Commercial source B' 112.7 were purified by polysucroselsodium diatrizoate (Histopaque, Sigma Standard techniqued purification 5.8 Chemical Co.Diagnostics, St, Louis, MO) density gradient centrifugation, LPS free purificatione
