IMMUNOLOGY - Roitt I. - Mir 2000

Chapter 18. Immunity to Parasitic Infections

T LYMPHOCYTES PLAY A PIVOTAL ROLE IN THE DEVELOPMENT OF IMMUNITY

In most parasitic infections, protective Immunity can be experimentally transferred to healthy animals by administering Spleen Cells—primarily T lymphocytes—from immune animals. The critical role of T lymphocytes in immunity is further evidenced by the fact that T-Cell-depleted and nude (athymic) mice fail to control protozoan infections such as those caused by T. cruzi or P. yoelii, whereas T-cell-depleted rats cannot clear the intestinal helminth Nippostrongylus brasiliensis (Fig. 18.9). However, in some instances, transferring T cells from heavily infected animals can suppress the protective response and lead to the death of recipients. This occurs because these T cells secrete IL-4 and IL-10, cytokines that inhibit the production and activity of IFNγ, which is essential for macrophage activation and parasite destruction.

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Fig. 18.9. The first two graphs illustrate the post-infection increase in Blood parasite levels (parasitemia) of proliferating Protozoa.

1. In thymectomized and irradiation-depleted T-cell mice (Thym X), Trypanosoma cruzi cells multiply more rapidly than in normal animals, leading to mortality. In normal mice, trypanosomes are cleared from the blood by day 16 post-infection. Transferring T lymphocytes from immune animals (immune T cells) to T-cell-depleted mice restores their ability to clear the infection. In these experiments, thymectomized animals in both groups received fetal Liver cells to restore vital hematopoietic function. 2. In normal mice, Plasmodium yoelii causes a self-limiting infection, being cleared from the blood by day 20. In athymic (nude) mice, the plasmodia continue to proliferate, causing death approximately 30 days post-infection. 3. This graph illustrates the dynamics of the expulsion of the intestinal nematode Nippostrongylus brasiliensis in rats. In normal animals, the helminth is completely expelled by day 13, as determined by fecal egg counts. Conversely, athymic rats develop chronic intestinal infestation, demonstrating that T cells are required for helminth expulsion.

Previously, The Role of cytokines in parasitic infections could only be investigated by administering specific cytokines to infected animals or by depleting them using Monoclonal Antibodies. Today, a more powerful approach is available, utilizing cytokine-Transgenic Mice and Gene-knockout mice lacking specific cytokine genes. This methodology has revealed that many cytokines not only act on effector cells by enhancing their cytotoxic or cytostatic activity, but also function as growth factors, promoting the proliferation of specific cell lineages. For instance, the monocytosis and Splenomegaly observed in malaria—driven by massive cellular expansion—are dependent on T-lymphocyte products. Other Examples of T-dependent effects include macrophage accumulation in hepatic granulomas during Schistosomiasis, helminth-associated eosinophilia, and the migration of eosinophils and mast cells into the intestinal mucosa during gastrointestinal helminth infections. Mucosal mast cells and eosinophils, which are crucial for helminth clearance, proliferate in response to T-cell products, specifically IL-3 + GM-CSF and IL-5, respectively.

However, increased cell proliferation can also have adverse outcomes. For example, administering IL-3 to mice infected with Leishmania major can exacerbate local infection and enhance parasite dissemination, potentially by stimulating the proliferation of Bone Marrow precursors of the host cells.

Both CD4⁺ and CD8⁺ T Lymphocytes Are Required for Protection Against Certain Parasites

The subset of T lymphocytes involved in combating an infection varies depending on The Nature of the parasite and the stage of the disease, and is determined by the cytokine profile produced by these cells. For example, CD4⁺ and CD8⁺ T lymphocytes act against different stages of malaria parasites: CD4⁺ T cells mediate the Immune Response against the erythrocytic stage of P. yoelii, whereas CD8⁺ T cells target the hepatic stage of P. berghei. The action of CD8⁺ T lymphocytes is twofold: they secrete IFNγ, which inhibits parasite Replication within hepatocytes, and they lyse infected hepatocytes. Hepatocytes express MHC class I but not class II Antigens, meaning CD4⁺ T lymphocytes cannot recognize them or be activated to secrete IFNγ. Conversely, CD8⁺ T cells are ineffective against the erythrocytic stage of the parasite because erythrocytes do not express MHC class I antigens.

The immune response against T. cruzi depends not only on CD4⁺ and CD8⁺ T lymphocytes, but also on natural killer (NK) cells and antibody production; the same holds true for the immune response against T. gondii. Experimentally, CD8⁺ T cells confer protection in CD4⁺-T-cell-depleted mice, both through the release of IFNγ and via direct cytotoxic action on infected macrophages. Another source of IFNγ is NK cells activated by macrophage-derived IL-12. Chronic infections are associated with impaired IFNγ production. Collectively, these findings may explain the high incidence of Toxoplasmosis in AIDS patients, who typically suffer from CD4⁺ T-cell suppression.

CD4⁺ T Cells Function Differently in Various Infections

CD4⁺ helper T cells are divided into two principal subsets: Th1 and Th2 (see Chapter 10). In the Cytology/cytology/16.html">Early stages of infection, both cell types may be present in roughly equal numbers; however, their relative proportions can shift over the course of a prolonged disease. Because Th1 and Th2 cytokines are mutually antagonistic, the T-cell subset that ultimately becomes dominant plays a decisive role in parasite clearance (the Nature of the immune response is heavily influenced by the parasite species and can be unpredictable). Because the relative importance of Th1 and Th2 cells generally differs between protozoan and helminth infections, it is best to examine each disease individually. (The cytokines produced by these subsets and their roles in immunity are discussed in Chapters 10 and 11.)

Th1 and Th2 Cell Activity in Malaria. Th1 cells act against the hepatic stage of the malaria parasite: administering IFNγ (a cytokine produced by Th1 cells) to chimpanzees infected with P. vivax sporozoites reduces parasitemia. Furthermore, studies have shown that the resistance of immune mice to P. berghei sporozoite challenge can be abrogated by administering anti-IFNγ antibodies. Th2 lymphocytes typically promote antibody production, thereby enhancing the Specificity of the immune response. An example of an antibody-dependent reaction is the destruction of the erythrocytic stage of the malaria parasite in the spleen by activated effector cells (antibody-dependent cellular cytotoxicity). The function of an intact spleen is essential for maintaining this immunity.

The Th1 Subset Enhances the Elimination of Intracellular Protozoa. The cytokine IFNγ activates macrophages to destroy intracellular protozoan parasites such as L. major, T. cruzi, and T. gondii (Figs. 18.10 and 18.11), and also amplifies effector responses in Infections caused by other pathogens. The critical role of IFNγ is strongly supported by studies of Leishmania infection. In resistant mouse strains, Skin lesion development caused by L. major is prevented by the action of IFNγ (a cytokine secreted by Th1 cells). In susceptible animals with progressive disease, Th2 cells predominate; the IL-4 they secrete suppresses IFNγ production. Administering anti-IL-4 antibodies resolves the infection by dampening Th12 cell activity and expanding the Th1 population. Direct administration of recombinant IFNγ to susceptible mice is insufficient to achieve this; thus, susceptibility in this model stems from excessive Th2 activity rather than a deficiency in Th1 function. Meanwhile, IL-12 (produced by macrophages and B cells) stimulates the proliferation of activated Th1 and NK cells, as well as the synthesis of Leishmania-clearing cytokines, including IFNγ, while suppressing Th2 development and promoting recovery when administered to susceptible mice during early infection.

Fig. 18.10. Suppression of parasite replication in cytokine-treated macrophages. Peritoneal macrophages from BALB/c mice infected with Leishmania donovani amastigotes were treated 72 hours post-infection with an activated T-cell extract (containing cytokines) or control T cells. Upon subsequent culture: parasites were undetectable in the cytokine-treated cells (1), whereas untreated macrophages harbored large numbers of parasites (2). Subsequent experiments using recombinant IFNγ and monoclonal anti-IFNγ antibodies demonstrated that this cytokine mediates parasite suppression. (Micrographs kindly provided by Dr. N. Murray; reproduced with permission from J. Immunol., 1982; 129: 344-357, © American Association of Immunologists.)

Fig. 18.11. Effect of administering the T-cell cytokine IFNγ in an acute Trypanosoma cruzi infection. In this mouse strain, parasites multiply and cause mortality approximately 3 weeks post-infection. Administration of recombinant IFNγ suppresses trypanosome replication and drives parasite clearance.

In humans, cases of diffuse cutaneous leishmaniasis and progressive visceral leishmaniasis are characterized by deficient IFNγ synthesis and upregulated expression of IL-10—a Th2-associated cytokine that inhibits Th1 cell proliferation and function. In vitro, IFNγ-induced human monocyte activity against L. donovani is inhibited by IL-4 (Fig. 18.12). These findings suggest that therapeutic administration of IL-12, combined with the simultaneous neutralization of antagonistic cytokines such as IL-4, IL-10, and TGFβ, could prove effective in treating leishmaniasis.

Fig. 18.12. During the Immune Response to Leishmania infection, cytokines released by distinct T-cell subsets either promote infection resolution or disease progression. Note that IL-12, which is also produced by B cells, stimulates the proliferation of both NK cells and Th1 cells; these cells secrete IFNγ, a cytokine critical for parasite elimination.

Th1 and Th2 Responses Are Essential for Protection Against Helminths. The hallmark Features of the immune response to helminth infection—elevated IgE levels and eosinophilia—depend on cytokines secreted by Th2 lymphocytes. However, the relative contributions of the Th1 and Th2 subsets to immunity against these parasites remain incompletely understood. Investigating this question is complicated by the fact that responses differ among mice, rats, and humans, at least in the case of schistosomiasis: in humans, Th2-mediated effects appear crucial for immunity, as resistance to reinfection following drug Treatment correlates with IgE production. Conversely, in mice, vaccine-induced protection relies on Th1 cells and IFNγ, whereas Th2 cells are associated with immunopathology driven by parasite egg deposition (egg antigens initiate the switch toward Th2 cell generation).

Infected animals may differ from vaccinated animals in their profile of synthesized cytokines. For example, mice infected with S. mansoni predominantly harbor Th2 cells that produce IL-5. In immunized mice, IgE levels and eosinophil counts are relatively low, and Th1 lymphocytes predominate. IFNγ secretion by Th1 cells activates effector cells that destroy schistosomula during their pulmonary migratory phase via nitric oxide release. Once adult worms begin laying eggs, these eggs release soluble antigens whose effects manifest exclusively in susceptible animals. This egg antigen suppresses Th1 cell function and IFNγ levels while upregulating IL-5 production by Th2 cells.

In some parasitic infections, The Immune System fails to completely eliminate the parasites, but instead walls them off from the body's Tissues using cells localized in the inflammatory zone. In this process, the host Organism responds to locally released antigens, which stimulate the secretion of cytokines that attract cells to the affected site. Such reactions have been demonstrated in mice vaccinated with radiation-attenuated schistosome cercariae. The infiltrating cells, predominantly Th1 lymphocytes, surround

Immunity to schistosomula parasitic infections in lung tissue develops as early as 24 h after intravenous parasite administration (Fig. 18.13). This prevents the subsequent migration of schistosomula into Organs and tissues where they could otherwise develop into mature adults. Another example of a host response aimed at isolating parasites is The formation of granulomas in the liver at sites where schistosomes deposit their eggs. This reaction is a chronic cell-mediated response to soluble antigens from eggs trapped in the liver. Macrophages accumulate and release fibrogenic factors that stimulate the formation of granulomatous tissue and, ultimately, The Development of fibrosis. Although this reaction can benefit the host by shielding liver cells from toxins secreted by the parasite eggs, it is also a major cause of pathology, as it induces irreversible liver damage and loss of liver function. In the absence of T cells, granulomas do not form, and subsequent fibrous encapsulation does not occur.

Fig. 18.13. Micrographs of mouse lung tissue showing the formation of focal lesions around migrating Schistosoma mansoni schistosomula. Schistosomulae were administered intravenously. 1. Lungs of an unvaccinated mouse 24 h after parasite administration. In mice protected by vaccination with radiation-attenuated cercariae, infiltrating cells appear as early as 24 h (2). 3–6. Sequential development of lesions at 2, 4, 8, and 12 days post-infection. Immunocytochemical analysis of bronchoalveolar samples revealed that CD4⁺ T lymphocytes are the primary component of the pulmonary infiltrates. The major cytokine produced by these cells in culture is IFN-γ; mRNA synthesis for IFN-γ is induced throughout the lung tissue, implying that the protective response is mediated by Th1 cells. (Photographs kindly provided by Dr. L. Smythies; reproduced with permission from Parasite Immunol. 1996; 18: 359–69.)

A variety of mechanisms may be deployed to combat helminths affecting different sites of the body—such as the intestine (e.g., Trichuris trichiura) or tissues (e.g., Onchocerca volvulus)—and targeting different Stages of the parasite's life cycle (e.g., schistosomulae in the lungs versus adult schistosomes in the Veins).

Th2 cells are required for the expulsion of intestinal helminths. Experiments have established that Th2 cells play a crucial role in fighting intestinal helminths. For example, mice normally resistant to whipworm infection develop a chronic infestation if IL-4, a cytokine produced by Th2 cells, is neutralized. Conversely, susceptible mice clear the parasites if IL-4 activity is enhanced by administering neutralizing antibodies against IFN-γ. Similarly, administering IL-12 to rats shortly after infection with the intestinal helminth N. brasiliensis promotes IFN-γ production and delays worm expulsion. The action of IL-12 involves suppressing the synthesis of Th2 cytokines, specifically IL-4 and IL-5; these effects prevent IgE production and intestinal mast cell hypertrophy (which are mediated by IL-4), as well as the development of eosinophilia (mediated by IL-5).

The Mechanism of helminth expulsion involves the degranulation of mucosal mast cells triggered by products of antigen-activated T cells. Mast cell products enhance other effector processes, interact with eosinophils, and accelerate parasite expulsion (Fig. 18.14). Mast cell cytoplasmic granules contain preformed mediators (see Chapter 23), various cytokines, and proteases. Granule components alter intestinal permeability and cause epithelial shedding, which also aids in the clearance of certain parasitic protozoa. During intestinal nematode infections, goblet cells secrete mucus that traps the helminths and facilitates their expulsion. This is a specific response observed only in immune animals. By altering mucosal permeability, mast cell mediators allow Complement and serum antibodies to enter the intestinal lumen and can act on smooth Muscle to drive parasite expulsion via enhanced peristalsis.

Fig. 18.14. Expulsion of certain intestinal nematodes occurs spontaneously several weeks post-infection. Parasite clearance appears to be a two-stage process driven by the combined action of T-dependent and T-independent mechanisms. 1. T cells (predominantly Th2) respond to parasite antigens and induce (a) antibody production by B cells proliferating in response to IL-4 and IL-5, (b) proliferation of mucosal mast cells in response to IL-3, IL-4, IL-9, and IL-10, and (c) hyperplasia of mucus-secreting goblet cells in the intestinal epithelium. Damage to the helminths is caused by antibodies and products from IgE-sensitized mast cells that degranulate upon contact with antigen, releasing histamine, which increases intestinal epithelial permeability. However, these effects alone are insufficient for worm expulsion. 2. Macrophage-secreted non-specific inflammatory molecules, including TNF and IL-1, promote goblet cell proliferation and stimulate increased mucus secretion. By enveloping the worms, the mucus aids in their expulsion. The number of goblet cells in the jejunal epithelium and the volume of mucus secreted increase in proportion to the parasite burden. Antigen-specific effector T cells are generated early in the infection; the rate-limiting step for resolving the infection is the onset of worm damage mediated by antibodies. The relative contribution of these responses to parasite expulsion depends on the nematode species.

Parasites induce both non-specific and specific antibody production

Many parasitic infections feature non-specific hypergammaglobulinemia, likely driven largely by parasite-released compounds that exert a B-cell mitogenic effect. Total immunoglobulin levels rise: IgM in trypanosomiasis and malaria, and IgG in malaria and visceral leishmaniasis. The relative importance of antibody-dependent and antibody-independent responses varies depending on the type of infection (Fig. 18.15). The Mechanisms of action of specific antibodies in parasitic infections and their effects are summarized in Fig. 18.16 and listed below.

✵ Antibodies can act on protozoan cells directly or via complement activation (Fig. 18.17).

✵ Antibodies can directly neutralize parasites by blocking their attachment to new host cells; for instance, during infections caused by various malaria species, merozoites invade erythrocytes using a specific receptor—a process inhibited by specific antibodies (Fig. 18.18). Antibodies can also prevent the spread of infection, as seen in the acute phase of T. cruzi infection.

✵ Macrophage-mediated phagocytosis is enhanced by antibodies, most effectively in the presence of complement. These effects are mediated via Fc and C3 receptors, the expression of which can be upregulated upon macrophage activation.

✵ Antibodies participate in antibody-dependent cell-mediated cytotoxicity (ADCC) reactions, such as in infections caused by T. cruzi, T. spiralis, S. mansoni, and filarial worms. Cytotoxic cells—such as macrophages, neutrophils, and eosinophils—attach to antibody-coated protozoa and helminths via Fc and C3 receptors and execute parasite exocytosis.

Fig. 18.15. Relative importance of two TYPES OF IMMUNE response, their mechanisms, and parasite evasion strategies against antibody-mediated damage in protozoan infections. Antibodies are critical for defense against blood-borne protozoa, such as African trypanosomes and malaria parasites, whereas cellular immunity is effective against tissue-dwelling parasites like Leishmania. Antibodies can directly damage parasite cells, stimulate phagocytosis, activate complement, or block parasite entry into host cells, thereby limiting the spread of infection. Once inside host cells, parasites evade all of these antibody effects. Trypanosoma cruzi and Leishmania cells are susceptible to oxygen metabolites generated during the macrophage respiratory burst and to nitric oxide. Cytokine treatment of macrophages enhances The production of these molecules, blocks cell entry, and reduces parasite survival. Intraerythrocytic malaria parasites can be destroyed by activated macrophage products, including hydrogen peroxide and other cytotoxic agents.

Fig. 18.16.

1. Direct antibody-mediated damage to parasites. Antibodies trigger classical pathway complement activation, leading to protozoan cell membrane disruption and increased vulnerability to other mediators.

2. Neutralization. Certain parasites, such as Plasmodium spp., invade cells using specific host cell receptors; antibodies block these binding sites, thereby preventing erythrocyte attachment and subsequent merozoite replication.

3. Enhanced phagocytosis. Deposition of the complement fragment C3b on the parasite surface opsonizes it for phagocytosis by cells bearing C3b receptors (e.g., macrophages). Macrophages also possess Fc receptors.

4. Eosinophils, neutrophils, platelets, and macrophages can exert cytotoxic effects (ADCC) on certain parasites by recognizing them via specific antibodies. This reaction is amplified by complement.

Fig. 18.17. Direct action of specific antibodies on malaria parasite sporozoites. These scanning electron micrographs show a sporozoite of Plasmodium berghei, the rodent malaria agent, before (1) and after (2) incubation with immune serum. Antibodies damage the sporozoite surface by acting on the outer membrane, causing fluid loss. Stage-specific antibodies confer protection against Plasmodium spp. infection at specific extracellular stages of The life cycle. Antibodies are stage-specific. (Photographs kindly provided by Dr. R. Nussenzweig.)

Fig. 18.18.

1. Administration of gamma globulin from immune adults to a child infected with Plasmodium falciparum causes a sharp decrease in parasitemia. Specific antibodies target merozoites—one of the stages in the parasite's life cycle—and prevent their further multiplication in the blood. However, the development of gametocytes from already existing intracellular forms is not suppressed.

2. Immune serum blocks the multiplication of P. knowlesi (the CAUSATIVE AGENT OF simian malaria) in culture, as indicated by ³H-leucine incorporation data. Plasmodium multiplication is inhibited at the stage following schizont rupture; the released merozoites fail to invade new erythrocytes. The activity of the immune serum may be reduced As a result of prior absorption of specific antibodies by free schizonts.

Different antibody isotypes can act through various mechanisms. As noted above, in schistosomiasis, specific IgE antibodies are associated with resistance to infection, showing an inverse correlation between blood IgE levels and susceptibility to reinfection. IgG4 antibodies appear to block the action of IgE; reinfection is most likely in childhood, which is characterized by high IgG4 levels. The development of immunity likely depends on a class switch from IgG4 production to IgE synthesis that occurs with age, as The rate of infection peaks at 10–14 years of age when IgG4 levels are particularly high.

For many parasitic infections, it is difficult to distinguish between cellular and humoral responses because they develop simultaneously. This is illustrated in Fig. 18.19, which summarizes data on the immune mechanisms acting against schistosomes.