GMP-140 (P-selectin), a 140-kDa granular membrane glycoprotein localized to the alpha granules of platelets and the Weibel-Palade bodies of endothelial cells, is thought to play an important role in adhesive interactions predominantly between granulocytes, platelets and vascular endothelial cells during inflammation. Although GMP-140 binds to granulocytes, its binding to lymphocytes has not been demonstrated. Using genetically engineered IgG C gamma 1 fusion protein of the extracellular domains of GMP-140, we demonstrate that GMP-140 binds to chronically antigen (Ag)-stimulated CD4+ T cells. Freshly isolated CD4+ T cells did not bind GMP-140, but priming and subsequent stimulation with alloantigen induced and gradually increased expression of GMP-140-reactive structures on their surface. T cells isolated from rheumatoid synovial fluids also exhibited strong binding to GMP-140. The binding of GMP-140 to primed T cells is not influenced by preactivation with phorbol 12-myristate 13-acetate, is almost completely abolished by pretreatment of T cells with neuraminidase or trypsin, and is also strongly inhibited by EDTA, the soluble sulfated glycans dextran sulfate, fucoidan, and heparin, but not by chondroitin sulfates. In spite of its strong binding to Ag-primed T cells, GMP-140 did not modulate the proliferative responses of these cells to various stimuli. However, GMP-140 in conjunction with anti-T cell receptor alpha beta monoclonal antibodies augmented the production of granulocyte-macrophage colony-stimulating factor GM-CSF and inhibited the production of interleukin-8 by Ag-primed T cells without influencing their tumor necrosis factor-alpha production. These results suggest that GMP-140 binds to chronically stimulated CD4+ T cells and differentially modulates their production of proinflammatory cytokines. The ability of Ag-primed T cells to bind GMP-140 may facilitate interactions with activated platelets and endothelial cells affecting the course of inflammation.

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The mechanism by which mycobacteria elicit class I-restricted T-cell responses remains undefined because these organisms have been shown to reside exclusively within membrane-bound vesicles in macrophages (M), their primary host cells. We studied the interaction of M. avium with dendritic cells (DC) because they are the most potent antigen-presenting cells and are abundant at M. avium infection sites. We observed that both DC and M, generated from human peripheral blood monocytes by short-term culture, internalized M. avium. The onset of programmed cell death and the percentage of apoptotic cells in infected DC and M were comparable. However, following infection, DC secreted significantly larger amounts of interleukin-12, but not interleukin-1␤, than infected autologous M. Further analysis of infected cells showed that while phagosomes failed to acidify in both M. avium-infected DC and M, bacilli grew more slowly in DC. Electron microscopy studies revealed that M. avium resided within endocytic vacuoles in both cell types. The vacuolar membrane surrounding some bacilli in approximately 10% of the vacuoles in DC possessed several breaks. The importance of this finding will have to be addressed in future studies.Organisms of the Mycobacterium avium complex are rarely pathogenic for healthy individuals (17) but cause disseminated disease in patients with AIDS (24, 40) and localized pulmonary infection in non-AIDS patients with underlying chronic lung disease (26). Infection with M. avium poses staggering public health problems because of the limited susceptibility of this organism to available antibiotics (12) and the ability of these bacilli to become resistant to commonly used antituberculosis agents (19). To develop new strategies for treating M. avium infection, we need to better understand the interactions of this organism with the host's immune system.As is the case with other mycobacteria, the importance of CD8 ϩ T cells in resistance to M. avium is controversial (6, 27) because the mechanism by which M. avium-derived molecules gain access to the cytoplasmic presentation pathway to elicit major histocompatibility complex class I-restricted T cells remains undefined. It is widely accepted that M. avium, a facultative intracellular bacillus, impedes macrophages' (M) processing and presentation of antigen by restricting vacuole maturation (10,36,39). In view of the paradox concerning the involvement of mycobacterium-specific major histocompatibility complex class I-restricted T cells in the control of infection and the finding that M. avium remains primarily within membrane-bound vesicles in M, we analyzed human dendritic cells (DC) infected in vitro with M. avium for uptake and intracellular growth of bacilli, apoptotic death, production of interleukin-12 (IL-12), fusigenicity of bacilli containing vacuoles with lysosomes, and the intracellular localization of the bacteria. The belief that M. avium may display behavior in DC different from its behavior in M was inspired by studies showing that in DC there is a...

Bisphosphonates are potent inhibitors of bone resorption in vivo and are emerging as important and widely used drugs for the treatment of a variety of abnormal bone resorptive processes. In the current study we investigated the in vitro effects of 4-amino-1-hydroxybutylidene-1,1-bisphosphonate (alendronate), a recently developed, extremely potent bisphosphonate, on the immune functions of human peripheral blood mononuclear cells (PBMCs). PBMC proliferation induced by lectins, alloantigens, and a nominal antigen (tetanus toxoid) was inhibited in a dose-dependent manner by alendronate. Pretreatment of monocytes, but not T cells, with the compound at concentrations ranging from 10(-4) to 10(-8) M was inhibitory, indicating that alendronate acts selectively on antigen-presenting cells (APCs). Alendronate did not affect the viability of monocytes or T cells or the expression of cell surface molecules known to play critical roles in antigen presentation. Alendronate exhibited dose-dependent inhibition of the production of interleukin-1 beta (IL-1 beta), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF-alpha) by activated monocytes. The inhibitory effect of 10(-6) M alendronate on PBMC proliferation was reversed by 10 U/ml recombinant rIL-1 beta, whereas other cytokines such as IL-6, TNF-alpha, and granulocyte-macrophage colony-stimulating factor (GM-CSF) had no effect. Thus, alendronate acts on monocytes to inhibit their antigen-presenting/accessory cell functions through a mechanism that can be overcome by exogenous IL-1. The inhibitory effect of this agent on cytokine production may contribute to its inhibitory effect on bone resorption.