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This report describes further attempts to define the nature of the parasite product(s) responsible for the extensive changes in lymphoid tissue in mice during infection with Trypanosoma brucei. As previously described, potent mitogenic and immunosuppressive effects are induced by a trypanosome-derived crude membrane fraction in vivo. There was no enrichment in these activities when purified parasite surface membranes were used. Mitogenic activity can be recovered from soluble trypanosome material only when it is incubated with peritoneal macrophages before transfer into syngeneic recipients. Thus, by encouraging association with a critical target cell, soluble parasite products can be studied, and their active components can be separated by conventional methods. Preliminary fractionation of high-spin trypanosome supernatant over Sepharose 4B confined the mitogenic activity to the high-molecular-weight fraction, which is a macromolecular complex of proteins, glycoproteins, and lipid. Extracted lipid from this material was able to significantly suppress a primary immunoglobulin G anti-sheep erythrocyte response. The activity was periodate sensitive and pronase resistant. The use of macrophages in vitro may be a general method whereby important biological activities lost as a result of fractionation procedures can be recovered and the active components studied in greater detail.

African trypanosomiasis is accompanied by profound general immunosuppression. The experiments described here were designed to characterize the contribution of macrophages to the immune pathology of this disease. We used peptone-stimulated, uninfected mice and injected them intraperitoneally with lethally irradiated and 35S-labeled Trypanosoma brucei and parasite-specific antisera. Peritoneal macrophages were thus induced to take up in vivo a defined number of trypanosomes. After the phagocytosis of parasites, macrophages were transferred into uninfected syngeneic mice, where they mimicked some of the important immunological changes normally associated with active trypanosome infection: (i) splenic background plaque-forming cells increased nonspecifically and (ii) the specific immune response to sheep erythrocytes was either enhanced or suppressed, depending on the timing of the antigen challenge: priming simultaneously with the transfer of trypanosome-containing macrophages enhanced immune responsiveness; in contrast, if parasite-containing macrophages were transferred and recipient mice were primed 4 days later, the immune response was suppressed. A contribution of suppressor T cells was ruled out by the treatment of peritoneal exudate cells with anti-Thy 1.2 and complement before transfer into recipient mice. The results indicate that macrophages are key cells in the mediation of parasite-induced immune dysfunction.

Previous reports have described profound effects on the function of the lymphoid system, especially the spleen, in mice infected with Trypanosoma brucei. This study provides further evidence of major change in the cell populations of the blood, peritoneum, and bone marrow, but shows that at least some of the stem cells of the bone marrow survive the damage caused by trypanosomes and retain their ability to repopulate the animal. In these infected mice the initial parasitemia was terminated by day 11 and was followed by a subpatent period of approximately 7 days before a final, lethal parasitemia occurred. Lymphopenia preceded the initial and final waves of parasites in the blood, and there was a marked increase in circulating neutrophils and large mononuclear cells for 1 week after the termination of the first wave of bloodstream parasitemia and during the final lethal parasitemia. Dividing macrophages were detected in the peritoneum only briefly during week 1 of infection, but the total number of peritoneal cells was increased from day 8 until the mice died. The bone marrow is severely stressed by the parasite infection. Total cell numbers and spleen colony-forming cells in the bone marrow were profoundly depleted during the resolution of the first parasitemia, but both these parameters largely recovered during the subpatent period before the mice were killed by the disease. Immune function was restored gradually after treatment with Berenil late in infection. We conclude that the progenitors of lymphocytes as well as the mature cells are affected by trypanosomes, but that some of the early bone marrow stem cells escape and rapidly repopulate the peripheral organs upon removal of the parasites.

African trypanosomiasis is accompanied by a profound general immunosuppression in which both suppressive T cells and macrophages (M phi) have been implicated. The present studies define changes in the M phi surface, endocytic and secretory properties, during the infection of mice by Trypanosoma brucei. Peritoneal M phi obtained after the control of the first wave of parasitemia displayed characteristics similar to those activated by intracellular pathogens, such as Mycobacterium bovis bacillus Calmette-Guérin, e.g., the enhanced expression of Ia antigen, decreased M phi-specific antigens, receptors mediating the pinocytosis of mannose-terminal glycoproteins, and an increased ability to secrete plasminogen activator, superoxide anion, and H2O2. Some markers of macrophage activation persisted during the subpatent period before the recurrence of parasitemia, whereas others reverted to normal. Mature T cell function appears not to be essential for M phi activation by T. brucei since the infection of athymic nude mice also induced Ia antigens and plasminogen activator. These studies show that M phi activated by different pathways express common features which may contribute to immune dysfunction observed in trypanosomiasis, as well as in other infections.

Trypanosoma brucei infection results in hypertrophy of the reticuloendothelial system. Peritoneal macrophages from infected mice were larger, spread out more on glass, and had increased receptor-mediated phagocytic activity both for C3b-and immunoglobulin G-coated erythrocytes. These observations suggest that macrophages were activated as a result of this infection.