Objective. Although monosodium urate monohydrate (MSU) crystals have been recognized since the 18th century as the etiologic agent of gout, it is still unknown why certain hyperuricemic individuals remain asymptomatic, and how an acute attack of gout spontaneously resolves. We hypothesized that mononuclear phagocytes hold the key to these questions, and that the state of monocyte/macrophage differentiation is critical.Methods. Human peripheral blood monocytes were differentiated for 1-7 days in vitro and examined with respect to 1) uptake of MSU crystals, 2) expression of macrophage, dendritic cell, and activation markers, 3) secretion of tumor necrosis factor ␣ (TNF␣), interleukin 1␤ (IL-1␤), IL-6, and IL-10, 4) activation of endothelial E-selectin expression, and 5) enhancement of secondary neutrophil recruitment by endothelial cells.Results. MSU crystals induced TNF␣, IL-1␤, and IL-6 (but not IL-10) secretion in undifferentiated monocytes, which in turn promoted endothelial cell E-selectin expression and secondary neutrophil capture under shear flow. In contrast, differentiation over 3-5 days led to development of a noninflammatory phenotype characterized by a lack of proinflammatory cytokine secretion, lack of endothelial cell activation, and lack of secondary neutrophil recruitment. Acquisition of the noninflammatory phenotype correlated with expression of macrophage antigen but not with expression of dendritic cell marker or activation marker. Monocytes and macrophages were similarly phagocytic, and a control particle, zymosan, elicited secretion of the full panel of cytokines in both cell types. However, coincubation with MSU led to a significant suppression of zymosaninduced TNF␣ secretion (P ‫؍‬ 0.009) from macrophages but not monocytes.Conclusion. These findings imply that differentiated macrophages provide a safe-disposal mechanism for the removal of inflammatory urate crystals. This may be of clinical relevance to the maintenance of asymptomatic hyperuricemia and the resolution of acute gout.

Objective. We have hypothesized that the process of monocyte to macrophage differentiation may alter the inflammatory response of mononuclear phagocytes to the uptake of monosodium urate monohydrate (MSU) crystals.Methods. Eight mouse monocyte/macrophage cell lines were arranged in increasing order of differentiation, as judged by expression of the macrophage markers F4/80 and BM 8 and by phagocytic capacity. Secretion of tumor necrosis factor ␣ (TNF␣) in response to MSU was measured by enzyme-linked immunosorbent assay.Results. The panel of monocyte/macrophage cell lines revealed a close linkage between the state of differentiation and the capacity of the cells to ingest MSU crystals. TNF␣ production, however, was not linked to phagocytic ability. Peak TNF␣ levels were synthesized by cells at an intermediate state of differentiation (3.2-14.1 ng/ml), whereas mature macrophages, which efficiently phagocytosed crystals, did not secrete TNF␣. Mature cell lines produced TNF␣ when stimulated with zymosan (5.9-6.2 ng/ml), but this was abolished by coincubation with MSU crystals. Suppression of the zymosan response was not due to apoptosis or steric hindrance by MSU crystals. Culture supernatants from mature macrophages did not stimulate endothelial cell activation, in contrast to MSU-treated cells at an earlier stage of differentiation, which stimulated intercellular adhesion molecule 1 expression on sEND endothelioma cells through the release of TNF␣ (inhibited 80.6% by anti-TNF␣).Conclusion. We demonstrated that phagocytosis and TNF␣ production are distinct events in the response of mononuclear phagocytes to urate crystals, and these events can be distinguished at the level of macrophage differentiation. The noninflammatory removal of urate crystals by mature macrophages defines a new pathway that may be important in controlling the development of acute gout in patients with hyperuricemia.Acute gouty arthritis is a self-limiting inflammatory response to the intraarticular deposition of monosodium urate monohydrate (MSU) microcrystals. Previous studies investigating the effect of MSU crystals in mononuclear phagocytes have focused primarily on the ability of peripheral blood monocytes or monocytic cell lines to secrete proinflammatory cytokines such as tumor necrosis factor ␣ (TNF␣), interleukin-1 (IL-1), . Investigators in our group have further shown that TNF␣ and IL-1 released by peripheral blood monocytes can activate vascular endothelial cell expression of E-selectin, intercellular adhesion molecule 1 (ICAM-1), and vascular cell adhesion molecule 1, thereby stimulating both the recruitment of leukocytes to the site of crystal deposition and the amplification of the inflammatory response (6-8).Hyperuricemia and precipitation of inflammatory microcrystals in synovial fluid per se are not always predictive of acute synovitis, as the severity of disease Supported by grants from the British Heart Foundation and Glaxo Wellcome PLC.

No abstract

P- and E-selectin are surface glycoproteins that mediate leukocyte rolling on the surface of endothelium in inflammation. We have cloned porcine P-selectin cDNA and generated a mAb, 12C5, with which to examine P-selectin expression by porcine aortic endothelial cells (PAEC) in comparison with that of E-selectin. Basal expression by PAEC of P-selectin was greater than that of E-selectin, whereas E-selectin expression was more prominently enhanced than that of P-selectin by stimulation with TNF-α or IL-1α. Both human or porcine IL-4 led to an increase in P-selectin expression, with kinetics that were delayed compared with those seen following stimulation with TNF-α or IL-1α, but IL-4 did not stimulate expression of E-selectin. When cells were stimulated with TNF-α in the presence of IL-4, we observed enhanced P-selectin expression with a parallel reduction in E-selectin expression. Finally, the increase in P-selectin expression due to human IL-4 was reduced in the presence of porcine but not human IFN-γ. These observations show that E-selectin and P-selectin expression are differentially regulated in PAEC, and that IL-4 leads to a shift in the relative surface density of the two molecules toward P-selectin. The ability of porcine IFN-γ to inhibit IL-4-induced P-selectin expression suggests that the balance between Th1 and Th2 cytokine production may determine the relative densities of the two selectins in chronic immune-mediated inflammation. Because the increased expression of P-selectin induced by human IL-4 was not inhibited by human IFN-γ, this balance may be shifted toward P-selectin expression in porcine xenografts infiltrated by human lymphocytes.

No abstract