Intrace|lular protozoa, Leishmania spp, Trypanosoma cruzi, and Toxoplasma gondii, have as a pathogenic characteristic a particular tropism for host mononuclear phagocytes. Successful invasion of host macrophages (M4~) 1 by Leishmania promastigotes requires an initial interaction at the cell surface that has not been defined. In the presence of serum, enhanced binding ofLeishmania promastigotes to murine resident peritoneal M~b (RPM) is mediated by complement (1). In the absence of serum, the lectin-like Mq~-specific mannosyl/fucosyl receptor (MFR) may play a role (2), consistent with studies (3, 4) demonstrating mannose-rich glycoconjugates on the surface of L. donovani promastigotes. We report an additional role for the type three complement receptor (CR3) in serum-independent binding ofL. donovani to RPM. Modulation experiments in which either CR3 or MFR are rendered inaccessible demonstrate that both receptors must be present on the segment of M4~ membrane to which the parasite binds. The combined function of these two distinct M4~ receptors may represent a general mechanism for the entry of pathogenic protozoa into mononuclear phagocytes. Materials and MethodsMice. C57BL/10ScSn mice were from OLAC(1976) Ltd., Bicester, Oxon, or bred at the London School of Hygiene and Tropical Medicine and males or females used between 6 and 12 wk.Reagents. The rat anti-mouse CR3 monoclonal antibody (mAb) Ml/70 (anti-Mac-l) (5) was used at saturation (6). Fab fragments of M1/70 and rat anti-mouse mAb 2.4G2, which binds to Mq~ IgG2b/IgG 1 Fc receptors (7), were kind gifts from Dr. T. Springer, Harvard Medical School, Boston and Dr. J. Unkeless, The Rockefeller University, New York. F4/80, a rat antibody (Ab) specific for mature mouse M4~ (8), was used as concentrated hybridoma supernatant. Anti-C3 Fab, prepared from a rabbit anti-human C3 that crossreacts with mouse C3, was used at a concentration (200 t~g/ml) known to

The concentrations of IL-8, IL-6, G-CSF and MCP-1 are similar to the levels found in reproductive tract fluids of patients with infection. The constitutive secretion and compartmentalization of large quantities of bioactive chemokines and cytokines provide additional evidence for the role of epithelial cells as gatekeepers of innate immune protection in the female reproductive tract.

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When tachyzoites were incubated with human peripheral blood leukocytes in vitro, more monocytes and dendritic cells than neutrophils or lymphocytes were infected. Although tachyzoites were able to divide in each of these cell types, monocytes and dendritic cells were more permissive to rapid tachyzoite division than neutrophils or lymphocytes.Toxoplasma gondii is an obligate intracellular parasite that can infect any nucleated host cell by a process of active penetration (7). Acute toxoplasmosis is characterized by dissemination and intracellular growth of the tachyzoite in a variety of host organs. In the innate phase of the immune response, nonspecific inflammatory cells, including neutrophils, monocytes, and dendritic cells, are elicited to the site of infection. The ensuing response of these cells allows for the establishment of the specific cell-mediated immune response that provides for long-term protection in the host against recurrent infection (13). Previous studies have reported that human neutrophils and monocytes become infected by tachyzoites. However, for each cell type there are conflicting reports of the subsequent fate of these intracellular parasites; both parasite stasis (14,16,30) and parasite division (15, 19) are described. In a recent study it was shown that there was markedly less uptake and division of tachyzoites in adherent monocytes than in nonadherent monocytes (9). In the present study the response of human peripheral blood leukocytes to tachyzoites was evaluated.Infection of human leukocytes is parasite dose dependent. Dendritic cells, monocytes, lymphocytes, and neutrophils were isolated from peripheral blood from toxoplasma-seronegative donors under endotoxin-free conditions. To obtain dendritic cells, mononuclear cells were isolated from blood by density gradient centrifugation (2:1 [vol/vol] ratio of blood to 1.07 g of Ficoll-Hypaque per ml; 500 ϫ g for 20 min). Washed cells were plated for 2 h at 37°C in tissue culture flasks, and then nonadherent cells were removed. The adherent cells were cultured in medium supplemented with 2,000 U of recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF; generously supplied by Immunex, Seattle, Wash.) and 20 ng of recombinant human interleukin-4 (rhIL-4; PeproTech Inc., Rocky Hill, N.J.) per ml. Every 3 days the medium in these flasks was supplemented with rhGM-CSF and rhIL-4. At day 8, the cells demonstrating dendritic cell morphology (nonadherent with projections) were harvested, and their phenotype was characterized by flow cytometry as described previously (27).Monocytes and lymphocytes were isolated from peripheral blood mononuclear cells obtained by cytophoresis carried out with volunteers as described previously (2). Briefly, monocytes were enriched to 85 to 95% by aggregation of washed cytophoresed cells at 4°C. Supernatants containing enriched lymphocytes were removed and purified further (see below). Platelets were removed from monocytes by washing cells twice in Versene buffer (0.2 g of EDTA per ml in phos...

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