Signalling through the IL-2 receptor γ(c) peptide (CD132) is essential for the expression of immunity to Plasmodium chabaudi adami blood-stage malaria.

A genetic dissection approach was employed to determine whether the IL-2 receptor complex (IL-2R) comprised of α, ß and γ chains is required for the suppression of Plasmodium chabaudi adami parasitemia. Blood-stage infections in IL-2Rγ(c)(-/y) mice failed to cure with parasitemia remaining elevated for > 50 days indicating the IL-2Rγ(c) through which all members of the γ(c) family of cytokines signal has an essential role in protective immunity against blood-stage malarial parasites. In contrast, the curing of parasitemia in IL-2/15Rß⁻/⁻ mice, deficient in both IL-2 and IL-15 signalling was significantly delayed but did occur, indicating that neither cytokine plays an essential role in parasite clearance. Moreover, the observation that the time course of parasitemia in IL-15⁻/⁻ mice was nearly identical to that seen in controls suggests that the parasitemia-suppressing role of stimulating through the IL-2/15Rß chain is owing to IL-2 signalling and not a redundant function of IL-15.

Expansion of the gammadelta T cell subset in vivo during bloodstage malaria in B cell-deficient mice.

Mice rendered B cell-deficient either by chronic anti-mu treatment initiated at birth or by gene knockout (JHD and mu-MT mice) suppressed acute Plasmodium chabaudi infections with a time course similar to intact control mice. Moreover, both kinds of B cell-deficient mice showed a 50- to 100-fold increase in splenic gammadelta T cell number after suppression of parasitemia compared with uninfected B cell-deficient controls; the magnitude of this increase resulted in significantly (P< 0.05) greater numbers of splenic gammadelta T cells in the B cell-deficient mice than in infected B cell-intact controls (about 10-fold). In contrast, the number of splenic CD4+ alphabeta T cells was only slightly elevated (< 2-fold) in both kinds of B cell-deficient mice compared with their intact controls. The number of splenic gammadelta T cells following suppression of P. vinckei parasitemia was approximately ninefold greater in JHD mice than in C57BL/6 controls, whereas similar numbers of splenic CD4+ alphabeta T cells were detected. Maximal numbers of gammadelta T cells were in cell-cycle in both JHD and C57BL/6 mice during descending P. chabaudi parasitemia, but the number of gammadelta T cells in cell-cycle was greater in B cell-deficient mice than in intact controls. Interleukin-10 (IL-10), a potent TH1 cell-suppressive molecule, does not appear to down-regulate the gammadelta T cell response during malaria in B cell-intact mice because the magnitude of the gammadelta T cell response was not significantly greater in IL-10 knockout mice compared with heterozygote controls. These findings collectively indicate that a markedly enhanced expansion of the gamma delta T cell population occurs in the absence of B cells, and this expansion occurs predominantly during acute malaria when parasite burdens are similar in B cell-deficient animals and intact controls.

Cell-mediated immunity to the asexual blood stages of malarial parasites: animal models.

Studies using experimental models of malaria in immunodeficient mice and chickens have shown that resistance to blood-stage infection is mediated by protective antibodies and T cell-dependent cell-mediated mechanisms of immunity. Depending on the infecting species of Plasmodium and prior experience of the host, either humoral or cell-mediated immune mechanisms predominate. Cell-mediated immunity has been adoptively transferred with CD4+ splenic T cells, and with antigen-specific T cell lines and clones. Since ascending parasitemia occurs in all instances, the transferred cells do not kill plasmodia directly but appear to activate effector mechanisms capable of destroying the invading parasites. Both CD4+ and CD8+ T cells were found to increase in the spleens of malarious mice. Depletion of CD4+ T cells prevented nude mice adoptively transferred with immune splenic T cells from clearing parasitemia. In contrast, late treatment with anti-CD4 antibody had little if any effect. The converse was true when anti-CD8 antibody was utilized, i.e., a significant number of mice treated with anti-CD8 antibody after parasitemia became patent and had difficulty clearing blood parasites. These data suggest that during infection CD8+ T cells may become activated by CD4+ T cells responding to malarial antigens. These CD8+ T cells may be directly cytotoxic or secrete additional cytokines thereby amplifying the role of CD4+ T cells in the activation of anti-parasite effector mechanisms. Finally, a hypothesis is presented to explain how the parasite in natural infections may activate T cell-dependent effector mechanisms in order to control its numbers in host tissues thereby ensuring the survival of both parasite and host.

The effects of interleukin-15 on human gammadelta T cell responses to Plasmodium falciparum in vitro.

We observed that the gammadelta T cell subset expands when human peripheral blood mononuclear cells (PBMC) from malaria-naive donors are cultured with Plasmodium falciparum lysate in the presence of IL-2 or IL-15, cytokines that utilize two common IL-2 receptor subunits. IL-15 induced the expansion of the gammadelta T cell subset at all levels tested, whereas IL-2 was not stimulatory at high levels. Flow cytometric analysis of apoptosis using the TUNEL assay indicated that the percentage and absolute number of gammadelta T cells undergoing apoptosis were greater in cultures stimulated with antigen and IL-2 than in cultures stimulated with either antigen and IL-15 or control erythrocyte lysate and IL-2. The ability of IL-15 to enhance gammadelta T cell function was also assessed; the results suggest that IL-15 can function with IL-2 to enhance the capacity of gammadelta T cells to inhibit parasite replication. Together these data indicate that IL-2 and IL-15, which both bind to IL-2Rbeta and IL-2R(gamma)c, enhance gammadelta T cell function, but they appear to have different effects on proliferation and survival.

Increased gamma delta T cells in acute Plasmodium falciparum malaria.

The T cell receptor of gamma delta is normally expressed on a small percentage of peripheral lymphocytes. Although the role of gamma delta T cells in the physiologic immune response is still unknown, there is accumulating evidence that gamma delta T cells may participate in the immune response to mycobacterial and other infectious organisms. In this study, we have quantitated the number of circulating gamma delta T cells during acute Plasmodium falciparum malaria. The results indicate that gamma delta T cells are elevated during the acute infection and remain elevated for at least 4 weeks during convalescence. T cells may participate in the immune response against P. falciparum by functioning as non-MHC restricted cytotoxic cells against intraerythrocytic parasites. Alternatively, lymphokines may be produced on antigen stimulation which may have antiparasitic activity.

T-cell immunity to malaria in the B-cell deficient mouse.

Some B-cell deficient mice drug-rescued with clindamycin HCl from otherwise lethal infections with Plasmodium yoelii resisted subsequent challenge with the same parasite despite the fact that they lacked detectable antibody to plasmodia. Parasitemias remained patent but at low levels (less than or equal to 5%) in these mice for prolonged periods of time, suggesting that some T-cell function independent of antibody formation can in part mediate immunity to malaria.

Cultivation of Plasmodium falciparum in commercially available sera depleted of natural antibodies reactive with human erythrocytes.

The efficacy of commercially available sera for the continuous in vitro cultivation of Plasmodium falciparum is controversial. In this study, various commercial animal and human sera are shown to support the in vitro growth of P. falciparum after their adsorption with erythrocytes to remove "natural" anti-human erythrocyte antibodies. A simple and inexpensive method to accomplish cultivation is presented.

In vitro antibody response to the tetrapeptide repeats of Plasmodium falciparum circumsporozoite protein by splenocytes from mice infected with P. yoelii yoelii.

The antibody response to the recombinant protein, R32tet32, which contained the repetitive sequence (NANP)n of Plasmodium falciparum CSP was determined in C57BL/6 mice during the course of nonlethal infection with Plasmodium yoelii 17X. Marked suppression of the IgG antibody response to R32tet32 occurred when mice were immunized at peak parasitemia (on day 16). In vitro antibody responses of spleen cells from acutely infected mice to R32tet32 were similarly suppressed. Stimulation of normal spleen cells cultured for 5 days with 100 ng/ml of R32tet32 gave an optimal IgG antibody response, but spleen cells from infected mice obtained at peak parasitemia failed to respond to a broad range of antigen concentrations. Cocultivation studies employing enriched lymphocyte populations from infected and uninfected C57BL/6 mice indicated that both T and B cells from infected mice were defective in their response to R32tet32. The response to the repetitive region was restored by the addition of recombinant mouse interleukin-2 (IL-2) at a dose of 50 U/ml to cultures of spleen cells from infected mice.

Activation of antigen-specific suppressor T cells by the intravenous injection of soluble blood-stage malarial antigen.

The regulation of delayed-type hypersensitivity (DTH) to soluble antigens derived from blood-stage parasites was investigated. DTH responses to soluble blood-stage malarial antigen were induced by subcutaneous (sc) sensitization in the flanks and elicited by ear challenge with the same antigen 6 days later. Adoptive transfer studies revealed that T cells of the L3T4+ phenotype were mediating this response. When a high dose of malarial antigen was injected intravenously (iv) prior to sc sensitization, immunosuppression of DTH resulted. The degree of immunosuppression was dependent on the dose of antigen injected iv and the time at which it was administered prior to sc sensitization. Immunosuppression was antigen-specific and mediated by Lyt-2+ splenic T cells.

Malarial immunodepression in vitro: adherent spleen cells are functionally defective as accessory cells in the response to horse erythrocytes.

The basis for the depressed response of malarial infected mice to horse red blood cells (HRBC) has been studied in vitro. Results presented show that the adherent spleen cells from infected mice (a) are defective in their ability to allow nonadherent spleen cells of both normal and infected mice to respond to HRBC whereas a response does occur with adherent spleen cells from normal mice (b) do not suppress the response of unfractionated spleen cells from normal mice to HRBC (c) contain phagocytic cells as measured by the uptake of neutral red in numbers which are of the same order of magnitude as in adherent spleen cells from normal mice, but which are unable to take up HRBC. We conclude that a splenic adherent cell, probably the macrophage is functionally defective as an accessory cell in the response to HRBC of mice infected with Plasmodium berghei yoelii.

Immunosuppressive effects of experimental infection with Plasmodium gallinaceum.

Experimental infection of chickens with P. gallinaceum mardedly suppressed the splenic PFC response to SRBC. Suppression was most pronounced when birds were immunized at the time of peak parasitemia. The PFC response to E. coli LPS was of low magnitiude in both normal and infected chickens; however, it, too, was suppressed in infected birds, but not to the same degree as observed in response to SRBC. Cellular immunity as evidenced by allograft rejection was not influenced by infection.

Immunity to Plasmodium chabaudi adami in the B-cell-deficient mouse.

Immunity to malaria has a multicomponent basis which requires the participation of both T- and B-lymphocyte systems. Previous studies have suggested that the T-lymphocyte system has an essential role in 're-infection immunity' to malaria, but that B cells and/or their products are necessary for the host to survive acute infection and to clear the blood of parasites during chronic malaria. Thus, B-cell-deficient mice and chickens died of fulminant malaria when infected with Plasmodium yoelii and Plasmodium gallinaceum, respectively, but when their acute infections were controlled with subcurative chemotherapy, B-cell-deficient host developed chronic low-grade infections and resisted challenge with homologous parasites. In contrast, athymic nude mice failed to control their endogenous P. yoelii infection after the termination of drug therapy unless they had been thymus grafted before initiation of acute infection. We now report that Plasmodium chabaudi adami (556KA) infection in B-cell-deficient mice results in an activation of a T-cell-dependent immune mechanism which terminates acute malaria in a similar way to that seen in immunologically intact mice. Furthermore, these immunized B-cell-deficient mice were resistant to homologous challenge infection as well as infections initiated with Plasmodium vinckei, but not with P. yoelii and Plasmodium berghei.

Antibody-independent immunity to reinfection malaria in B-cell-deficient mice.

Immunity to "reinfection malaria" or "premunition" was studied in B-cell-deficient mice which had previously experienced acute malaria caused by the avirulent plasmodia Plasmodium yoelii or P. chabaudi or by the lethal P. vinckei. Such mice resisted challenge infection with large numbers of homologous parasites but differed in their capacity to resist challenge with heterologous species. Mice immune to P. yoelii resisted infection with P. chabaudi but developed acute-type, albeit nonlethal, infections when challenged with P. vinckei. Whereas mice immune to P. chabaudi resisted challenge with P. vinckei and vice versa, they developed fulminating malaria and died when infected with P. yoelii. The data suggest that immunity to reinfection malaria in B-cell-deficient mice, although antibody independent, is mediated by different mechanisms of resistance depending upon the plasmodial species used to initiate acute infection. Additional evidence supporting this concept was gained from preliminary experiments in which immunity to reinfection was measured by the ability of chronically infected mice to control endogenous parasites at low levels. B-cell-deficient mouse strains showed genotypic differences in their ability to develop immunity to reinfection with P. yoelii. In contrast, the same mouse strains uniformly developed immunity to reinfection with P. chabaudi. These findings suggest that different genetic loci control resistance to reinfection malaria caused by different species of plasmodia. Finally, B-cell-deficient mice acutely infected with lethal plasmodia, P. vinckei or P. berghei, died at the same time or earlier than similarly infected immunologically intact mice, indicating that "early death" in virulent malarial infections is an antibody-independent phenomenon.

Splenomegaly, enhanced phagocytosis, and anemia are thymus-dependent responses to malaria.

Immunologically competent mice and mice with defined immunological deficiencies were infected with Plasmodium yoelii. Splenomegaly, enhanced phagocytosis, and anemia were most marked in infected mice having intact thymic tissue. Whereas the spleens of infected nude mice increased minimally in size, the relative blood hemoglobin levels and the rates of carbon clearance in these mice were similar to those of noninfected, immunologically intact mice. Thymus-reconstituted nude mice and B-cell-deficient mice responded to infection in a manner similar to that of infected immunocompetent mice. These data demonstrate that the hallmarks of malaria, i.e., splenomegaly, enhanced phagocytosis, and anemia, are thymus-dependent responses to infection.