Anne-Marit Sponaas

Immunity to malaria: more questions than answers

Malaria is one of the main health problems facing developing countries today. At present, preventative and treatment strategies are continuously hampered by the issues of the ever-emerging parasite resistance to newly introduced drugs, considerable costs and logistical problems. The main hope for changing this situation would be the development of effective malaria vaccines. An important part of this process is understanding the mechanisms of naturally acquired immunity to malaria. This review will highlight key aspects of immunity to malaria, about which surprisingly little is known and which will prove critical in the search for effective malaria vaccines.

Malaria infection changes the ability of splenic dendritic cell populations to stimulate antigen-specific T cells

The capacity of splenic CD11c+ dendritic cell (DC) populations to present antigen (Ag) to T cells differs during malarial infection with Plasmodium chabaudi in mice. Both CD11c+CD8+ and CD8− DCs presented malarial peptides on their surface during infection. However, although both DC subsets expressing malaria peptides could induce interferon-γ production by CD4 T cells, only CD8− DCs isolated at the acute phase of infection stimulated Ag-specific T cell proliferation and interleukin (IL)-4 and -10 production from MSP1-specific T cell receptor for Ag transgenic T cells coincidental with our reported Th1 to Th2 switch at this stage in response to the pathogen. The timing of these distinct DC responses coincided with increased levels of apoptosis in the CD8+ population and an increase in the numbers of CD8− DCs in the spleen. Our data suggest that the switch in CD4 T cell responses observed in P. chabaudi–infected mice may be the result of the presentation by different DC populations modified by the malaria infection.

Dendritic cells, pro-inflammatory responses, and antigen presentation in a rodent malaria infection.

Summary:  An infection of mice with Plasmodium chabaudi is characterized by a rapid and marked inflammatory response with a rapid but regulated production of interleukin‐12 (IL‐12), tumor necrosis factor‐α (TNF‐α), and interferon‐γ (IFN‐γ). Recent studies have shown that dendritic cells (DCs) are activated in vivo in the spleen, are able to process and present malaria antigens during infection, and may provide a source of cytokines that contribute to polarization of the CD4 T‐cell response. P. chabaudi‐infected erythrocytes are phagocytosed by DCs, and peptides of malaria proteins are presented on major histocompatibility complex (MHC) class II. The complex disulfide‐bonded structure of some malaria proteins can impede their processing in DCs, which may affect the magnitude of the CD4 T‐cell response and influence T‐helper 1 (Th1) or Th2 polarization. DCs exhibit a wide range of responses to parasite‐infected erythrocytes depending on their source, their maturational state, and the Plasmodium species or strain. P. chabaudi‐infected erythrocytes stimulate an increase in the expression of costimulatory molecules and MHC class II on mouse bone marrow‐derived DCs, and they are able to induce the production of pro‐inflammatory cytokines such as IL‐12, TNF‐α, and IL‐6, thus enhancing the Th1 response of naïve T cells. IFN‐γ and TNF‐α play a role in both protective immunity and the pathology of the infection, and the inflammatory disease may be regulated by IL‐10 and transforming growth factor‐β. It will therefore be important to elucidate the host and parasite molecules that are involved in activation or suppression of the DCs and to understand the interplay between these opposing forces on the host response in vivo during a malaria infection.

Migrating monocytes recruited to the spleen play an important role in control of blood stage malaria

Host responses controlling blood-stage malaria include both innate and acquired immune effector mechanisms. During Plasmodium chabaudi infection in mice, a population of CD11b(high)Ly6C(+) monocytes are generated in bone marrow, most of which depend on the chemokine receptor CCR2 for migration from bone marrow to the spleen. In the absence of this receptor mice harbor higher parasitemias. Most importantly, splenic CD11b(high)Ly6C(+) cells from P chabaudi-infected wild-type mice significantly reduce acute-stage parasitemia in CCR2(-/-) mice. The CD11b(high)Ly6C(+) cells in this malaria infection display effector functions such as production of inducible nitric oxide synthase and reactive oxygen intermediates, and phagocytose P chabaudi parasites in vitro, and in a proportion of the cells, in vivo in the spleen, suggesting possible mechanisms of parasite killing. In contrast to monocyte-derived dendritic cells, CD11b(high)Ly6C(+) cells isolated from malaria-infected mice express low levels of major histocompatibility complex II and have limited ability to present the P chabaudi antigen, merozoite surface protein-1, to specific T-cell receptor transgenic CD4 T cells and fail to activate these T cells. We propose that these monocytes, which are rapidly produced in the bone marrow as part of the early defense mechanism against invading pathogens, are important for controlling blood-stage malaria parasites.

gp96-Peptide Vaccination of Mice against Intracellular Bacteria

ABSTRACT This work demonstrates that gp96 preparations isolated from cells infected with intracellular bacteria induce cytotoxic T-lymphocyte responses and confer protection. Our findings extend previous reports on the immunogenicity of gp96-associated peptides to antigens derived from intracellular bacteria. Immunization with gp96 may therefore represent a promising vaccination strategy against bacterial pathogens.

Classical CD11c+ dendritic cells, not plasmacytoid dendritic cells, induce T cell responses to Plasmodium chabaudi malaria.

Dendritic cells play an important role in the development of immune responses in malaria, but the contribution of plasmacytoid dendritic cells (pDC) to CD4 T cell activation and immunopathology is unknown. We have investigated pDC in a Plasmodium chabaudi infection in mice. During infection, pDC increased in number and transiently up-regulated expression of Major Histocompatibility Complex class II and co-stimulatory molecules. However, in contrast to classical CD11c(high) DC, pDC could not phagocytose parasites or process parasite proteins, to activate CD4 T cells. Activation of naïve pDC, but not CD11c(high) DC, by infected red blood cells induced IFN alpha in vitro, which was dependent on the Toll-like receptor, TLR9. However, inactivation of TLR9 in knock-out mice had no effect on a P. chabaudi infection suggesting that TLR9 was not crucial for parasite elimination or pathology. Neither pDC nor IFN alpha beta were essential for parasite clearance as mice depleted of pDC or IFN alpha beta Receptor-knock-out mice could control infection. However, these mice lost significantly more weight than untreated or wild-type mice. We conclude that classical DC are the major antigen-presenting cells for CD4 T cells in this infection, but that pDC and IFN alpha beta may play minor roles in controlling the magnitude of acute stage pathology.

IL-22 Protects Against Liver Pathology and Lethality of an Experimental Blood-Stage Malaria Infection

The host response following malaria infection depends on a fine balance between levels of pro-inflammatory and anti-inflammatory mediators resulting in the resolution of the infection or immune-mediated pathology. Whilst other components of the innate immune system contribute to the pro-inflammatory milieu, T cells play a major role. For blood-stage malaria, CD4+ and γδ T cells are major producers of the IFN-γ that controls parasitemia, however, a role for TH17 cells secreting IL-17A and other cytokines, including IL-17F and IL-22 has not yet been investigated in malaria. TH17 cells have been shown to play a role in some protozoan infections, but they also are a source of pro-inflammatory cytokines known to be involved in protection or pathogenicity of infections. In the present study, we have investigated whether IL-17A and IL-22 are induced during a Plasmodium chabaudi infection in mice, and whether these cytokines contribute to either protection or to pathology induced during the infection. Although small numbers of IL-17- and IL-22-producing CD4 T cells are induced in the spleens of infected mice, a more pronounced induction is observed in the liver, where increases in mRNA for IL-17A and, to a lesser extent, IL-22 were observed and CD8+ T cells, rather than CD4 T cells, are a major source of these cytokines in this organ. Although the lack of IL-17 did not affect the outcome of infection or pathology, lack of IL-22 resulted in 50% mortality within 12 days after infection with significantly greater weight loss at the peak of infection and significant increase in alanine transaminase in the plasma in the acute infection. As parasitemias and temperature were similar in IL-22 KO and wild-type control mice, our observations support the idea that IL-22 but not IL-17 provides protection from the potentially lethal effects of liver damage during a primary P. chabaudi infection.

Alterations of Splenic Architecture in Malaria Are Induced Independently of Toll-Like Receptors 2, 4, and 9 or MyD88 and May Affect Antibody Affinity

ABSTRACT Splenic microarchitecture is substantially altered during acute malaria infections, which may affect the development and regulation of immune responses. Here we investigated whether engagement of host Toll-like receptor 2 (TLR2), TLR4, TLR9, and the adaptor protein MyD88 is required for induction of the changes and whether antibody responses are modified when immunization takes place during the period of splenic disruption. The alterations in splenic microarchitecture were maximal shortly after the peak of parasitemia and were not dependent on engagement of TLR2, TLR4, or TLR9, and they were only minimally affected by the absence of the MyD88 adaptor molecule. Although germinal centers were formed in infected mice, they did not contain the usual light and dark zones. Immunization of mice with chicken gamma globulin 2 weeks prior to acute Plasmodium chabaudi infection did not affect the quantity or avidity of the immunoglobulin G antibody response to this antigen. However, immunization at the same time as the primary P. chabaudi infection resulted in a clear transient reduction in antibody avidity in the month following immunization. These data suggest that the alterations in splenic structure, particularly the germinal centers, may affect the quality of an antibody response during a malaria infection and could impact the development of immunity to malaria or to other infections or immunizations given during a malaria infection.

Immunization with gp96 from Listeria monocytogenes-Infected Mice Is Due to N-Formylated Listerial Peptides

N-Formylated (N-f-met) peptides derived from proteins of the intracellular bacterium Listeria monocytogenes generate a protective, H2-M3-restricted CD8 T cell response in C57BL/6 mice. N-f-met peptide-specific CTL were generated in vitro when mice previously immunized with gp96 isolated from donor mice infected with L. monocytogenes were stimulated with these peptides. No significant peptide-specific CTL activity was observed in mice immunized with gp96 from uninfected animals. Masses corresponding to one N-f-met peptide were found by matrix-assisted laser desorption/ionization-mass spectrometry on gp96 isolated from C57BL/6 mice infected with L. monocytogenes, but not on gp96 from noninfected mice. Therefore, bacterial N-f-met peptides from intracellular bacteria can bind to gp96 in the infected host, and gp96 loaded with these peptides can generate N-f-met-peptide-specific CTL. We assume a unique role of gp96 in Ag processing through the H2-M3 pathway.

Scant activation of CD8 T cells by antigen loaded on heat shock protein

Heat shock proteins (HSP) not only function as chaperones for denatured proteins but also for antigenic peptides, thus inducing protective T cell responses. Here we show that vaccination with peptide‐loaded HSP70 causes initial interferon‐γ production by murine CD8 T cells but no T cell expansion. These CD8 T cells lacked cytotoxic activity in vitro and in vivo, which was not due to apoptosis. Restimulation with peptide‐pulsed dendritic cells both bypassed the proliferative block and suspended the non‐protective state of CD8 T lymphocytes in an infection model with the bacterial pathogen, Listeria monocytogenes. Cotransfer of antigen‐specific CD4 T cells circumvented the proliferative arrest of CD8 T cells. Our data suggest that HSP vaccines induce CD8 T cell unresponsiveness unless proficient help is provided. Assuming that this model reflects the antigenically experienced human condition where immunological space is restricted and any T cell response possibly leads to suppression of heterologous reactions, our findings bear implications for rational vaccination protocols including those for immunocompromised patients.