Abstract-Basic calcium phosphate (BCP) crystal deposition underlies the development of arterial calcification.Inflammatory macrophages colocalize with BCP deposits in developing atherosclerotic lesions and in vitro can promote calcification through the release of TNF alpha. Here we have investigated whether BCP crystals can elicit a proinflammatory response from monocyte-macrophages. BCP microcrystals were internalized into vacuoles of human monocyte-derived macrophages in vitro. This was associated with secretion of proinflammatory cytokines (TNF␣, IL-1␤ and IL-8) capable of activating cultured endothelial cells and promoting capture of flowing leukocytes under shear flow. Critical roles for PKC, ERK1/2, JNK, but not p38 intracellular signaling pathways were identified in the secretion of TNF alpha, with activation of ERK1/2 but not JNK being dependent on upstream activation of PKC. Using confocal microscopy and adenoviral transfection approaches, we determined a specific role for the PKC-alpha isozyme. The response of macrophages to BCP crystals suggests that pathological calcification is not merely a passive consequence of chronic inflammatory disease but may lead to a positive feed-back loop of calcification and inflammation driving disease progression. Key Words: atherosclerosis pathophysiology Ⅲ cell biology Ⅲ calcification Ⅲ inflammation Ⅲ macrophage C oronary artery calcification occurs as part of the atherosclerotic process, and is due to the deposition in the arterial intima of basic calcium phosphate (BCP) crystals, consisting mainly of calcium hydroyapatite (Ca 10 (PO 4 ) 6 (OH) 2 ) and similar to those that mineralize bone. [1][2][3][4][5][6] Using electron beam computed tomography, the amount of vessel calcification has been shown to closely correlate with the extent of plaque burden and may predict future coronary events. [7][8][9][10] However, although arterial calcification is now accepted to be an active and highly regulated process similar to that of bone ossification, [11][12][13][14] it has tended to be seen merely as a surrogate marker for the burden of atherosclerotic disease rather than a contributor to disease progression.Growing evidence from the study of degenerative arthritis, in which BCP crystals can be found in the majority of affected joints and closely correlate with the extent of joint destruction, suggests a pathogenic role in driving disease. 15,16 Thus, BCP crystals have been shown to activate synovial fibroblasts, inducing cellular proliferation and matrix metalloproteinase (MMP) secretion through a variety of intracellular signaling pathways, including protein kinase C (PKC), ERK1/2 MAP kinase, NFB, and AP-1. [17][18][19] Limited studies using murine cells have also demonstrated the ability of macrophages to interact with BCP crystals in vitro, resulting in DNA synthesis and cytokine production. 20,21 Cells of the monocyte/macrophage lineage form an important part of the innate immune system and play a key role in atherogenesis. 22 Inflammatory mediators derived from monocy...

Abstract-The recently described hemoglobin scavenger receptor CD163 mediates the endocytosis of hemoglobin:haptoglobin (Hb:Hp) complexes and thereby counters Hb-induced oxidative tissue damage after hemolysis. Although CD163 has been indirectly associated with antiinflammatory and atheroprotective activity, no ligand-receptor-effector pathway has yet been described for this receptor. To understand the significance of CD163 and more clearly define downstream pathways linked to inflammatory resolution, we studied the expression and function of CD163 in human monocytes/macrophages using both in vitro and in vivo models. Differentiation of human blood monocytes into macrophages either by in vitro culture or in resolving cantharidin-induced skin blisters led to an equivalent increase (Ͼ15ϫ) in CD163 expression. Elevated CD163 levels were also noted on circulating monocytes in cardiac surgical patients during the resolution phase of the systemic inflammatory response to cardiopulmonary bypass surgery. In each case, binding of Hb:Hp to CD163-bearing cells elicited potent interleukin-10 secretion, and this was inhibited by the anti-CD163 antibody RM3/1. Release of interleukin-10, in turn, induced heme oxygenase-1 stress protein synthesis via an autocrine mechanism. Such induction of heme oxygenase-1 was observed in vivo 24 to 48 hours after the onset of cardiopulmonary bypass surgery. These results identify novel antiinflammatory and cytoprotective effector pathways in human monocytes/macrophages related to Hb scavenging and metabolism, which may have relevance in atheroprotection, wound healing, and patient recovery postoperatively.

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.