Uncertainty remains about the cellular origins of the earliest phase of the proinflammatory cytokine response to malaria. Here we show by fluorescence-activated cell sorter analysis that ␥␦ T cells and CD14 ؉ cells from nonimmune donors produce tumor necrosis factor and that ␥␦ T cells also produce gamma interferon within 18 h of contact with mycoplasma-free Plasmodium falciparum-infected erythrocytes in vitro. This early cytokine response is more effectively induced by intact than by lysed parasitized erythrocytes. However, the IFN-␥ response to lysed parasites is considerably enhanced several days after peripheral blood mononuclear cells are primed with low numbers of intact parasitized erythrocytes, and in this case it derives from both ␣␤ and ␥␦ T cells. These data show that naïve ␥␦ T cells can respond very rapidly to malaria infection but that malaria fever may involve a multistage process in which the priming of both ␥␦ and ␣␤ T-cell populations boosts the cytokine response to lysed parasite products released at schizont rupture.Malaria-infected individuals produce large amounts of proinflammatory cytokines, such as tumor necrosis factor (TNF) and gamma interferon (IFN-␥). This innate cytokine response is responsible for the high levels of fever that occur within a few days of the onset of blood stage infection in nonimmune individuals (15,18,20,21,33). Since TNF and IFN-␥ have important antiparasitic actions but are also believed to play a major role in the pathogenesis of severe complications, such as cerebral malaria and severe malarial anemia, understanding the cellular basis of the early innate cytokine response may be of considerable importance in relation to both malaria immunity and pathogenesis.Until recently, it has been widely assumed that the early host response to malaria is broadly similar to that evoked by bacterial endotoxin, whereby parasite factors (toxins) stimulate monocytes and macrophages to release TNF and related cytokines (5,19). This model was supported by evidence that some isolates of Plasmodium falciparum strongly stimulate TNF production by human peripheral blood mononuclear cells (PBMC) (1) and by a significant literature documenting TNF production by monocytes or macrophages within a few hours of exposure to malaria parasite preparations in vitro (examples are given in references 2, 25, and 34). However, it has recently become apparent that many malaria culture preparations are contaminated with mycoplasma species which have potent macrophage-stimulatory factors (22,30,36). Mycoplasma species have been found in parasite lines obtained from different laboratories around the world, and even after infection has been eradicated, continuous cultures are susceptible to reinfection within a few months unless their mycoplasma status is continually monitored. These observations have made it necessary to reevaluate the cellular basis of the early cytokine response to malaria.We have recently observed that the pattern of early cytokine production by nonimmune human PBMC following stim...

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-alpha (TNF-alpha), and interferon-gamma (IFN-gamma). 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-alpha, and IL-6, thus enhancing the Th1 response of naïve T cells. IFN-gamma and TNF-alpha 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-beta. 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.

The N-terminal src-homology 2 domain of the p85a subunit of phosphatidylinositol 3' kinase (SH2-N) binds specifically to phosphotyrosine-containing sequences. Notably, it recognizes phosphorylated Tyr 75 1 within the kinase insert of the cytoplasmic domain of the activated OPDGF receptor. A titration of a synthetic 12-residue phosphopeptide (ESVDY*VPMLDMK) into a solution of the SH2-N domain was monitored using heteronuclear 2D and 3D NMR spectroscopy. 2D-( "N-IH) heteronuclear single-quantum correlation (HSQC) experiments were performed at each point of the titration to follow changes in both I5N and ' H chemical shifts in NH groups. When mapped onto the solution structure of the SH2-N domain, these changes indicate a peptide-binding surface on the protein. Line shape analysis of ID profiles of individual ( "N-'H )-HSQC peaks at each point of the titration suggests a kinetic exchange model involving at least 2 steps. To characterize changes in the internal dynamics of the domain, the magnitude of the ( "N-IH J heteronuclear NOE for the backbone amide of each residue was determined for the SH2-N domain with and without bound peptide. These data indicate that, on a nanosecond timescale, there is no significant change in the mobility of either loops or regions of secondary structure. A mode of peptide binding that involves little conformational change except in the residues directly involved in the 2 binding pockets of the p85a SH2-N domain is suggested by this study.

We have investigated the widely held view that malaria parasites induce pro-inflammatory cytokines primarily through an endotoxin-like stimulatory effect on macrophages. We report that the pattern of cytokine production by non-immune human peripheral blood mononuclear cells following stimulation by Plasmodium falciparum-infected erythrocytes (Pfe) in vitro differs considerably from that induced by bacterial endotoxin. The Pfe-induced TNF response at day 1 is associated with a much higher level of IFN-gamma production and a much lower level of IL-12 p40 and IL-10 expression than a comparable endotoxin-induced TNF response. Both CD3(+) and CD14(+) populations are required for this early TNF response to Pfe, whereas the endotoxin-induced response is unaffected by depletion of the CD3(+) population. Pfe fails to stimulate the monocyte-like cell line MonoMac6 to express pro-inflammatory cytokines. These findings suggest that the early inflammatory response to malaria is critically dependent on lymphocyte subpopulations that play a lesser role in the response to bacterial endotoxin.

The 19 kDa C‐terminal fragment of the malaria parasite merozoite surface protein 1 (MSP119) is a leading malaria vaccine candidate. In rodents, high antibody levels to this protein confer protective immunity, and can be generated by immunization with the antigen in adjuvants. In natural human infections, however, MSP119‐specific antibody responses can be short‐lived andcomparatively low, despite repeated exposure to infection. The tightly folded structure of MSP119 is stabilized by five or six disulfide bonds. These bonds impede antigen processing and, thereby, may affect the generation of CD4+ T cells providing help for B cells. Asparagine endopeptidase could digest unfolded, but not native MSP119 in vitro. Immunization with unfolded MSP119 resulted in a faster antibody response, and a combination of unfolded and native MSP119 increased antibody responses to the native form. Immunization with either form of the antigen activated similar numbers of CD4+ T cells, but, unlike the antibody response, CD4+ T cells immunized with one form of MSP119 were able to respond in vitro to the other form of the protein. Although the reduced form of MSP119 does not induce protective antibodies, our data suggest that inclusion of unfolded protein may improve the efficacy of MSP119 as a vaccine.See correction http://dx.doi.org/10.1002/eji.200490004