Dual role of erythrocyte complement receptor type 1 in immune complex-mediated macrophage stimulation: implications for the pathogenesis of Plasmodium falciparum malaria.

Given the ability of erythrocytes to bind immune complexes (ICs), we postulated that they can serve a dual role during inflammatory or infectious processes. Erythrocytes could restrict stimulation of macrophages by free ICs by binding C3b-opsonized ICs via their complement receptor 1 (CR1). Conversely, IC-loaded erythrocytes could stimulate macrophages to produce proinflammatory cytokines such as tumour necrosis factor (TNF)-α. To test our hypothesis we selected 72 individuals with low, medium or high red cell CR1 expression and determined their IC binding capacity. We tested the in vitro ability of red cells to inhibit IC-mediated stimulation of TNF-α production by macrophages or to stimulate TNF-α production when loaded with ICs. Plain erythrocytes inhibited IC-induced TNF-α production by macrophages and low CR1 expressors showed the lowest inhibitory capacity. IC-loaded erythrocytes stimulated macrophages to release TNF-α, but the effect was not proportional to the CR1 level. These data support our hypothesis that erythrocytes can serve a dual role in regulation of cytokine responses in a setting of IC formation. Our findings suggest that individuals with low CR1 expression are ill-equipped to clear ICs and prevent IC-mediated stimulation of macrophages. In addition, IC-loaded red cells in areas of sluggish circulation such as in the spleen or in brain capillaries blocked by sequestered malaria-infected red cells may induce inflammation by stimulating monocytes and macrophages, the latter leading to the development of cerebral malaria.

Distinct pattern of class and subclass antibodies in immune complexes of children with cerebral malaria and severe malarial anaemia.

Plasmodium falciparum infection can lead to deadly complications such as severe malaria-associated anaemia (SMA) and cerebral malaria (CM). Children with severe malaria have elevated levels of circulating immune complexes (ICs). To further investigate the quantitative differences in antibody class/subclass components of ICs in SMA and CM, we enrolled 75 children with SMA and 32 children with CM from hospitals in western Kenya and matched them to 74 and 52 control children, respectively, with uncomplicated symptomatic malaria. Total IgG IC levels were always elevated in children with malaria upon enrollment, but children with CM had the highest levels of any group. Conditional logistic regression showed a borderline association between IgG4-containing IC levels and increased risk of SMA (OR = 3.11, 95% CI 1.01-9.56, P = 0.05). Total IgG ICs (OR = 2.84, 95% CI 1.08-7.46, P = 0.03) and IgE-containing ICs (OR = 6.82, OR 1.88-24.73, P < or = 0.01) were associated with increased risk of CM. These results point to differences in the contribution of the different antibody class and subclass components of ICs to the pathogenesis of SMA and CM and give insight into potential mechanisms of disease.

Red cell surface changes and erythrophagocytosis in children with severe plasmodium falciparum anemia.

Severe anemia is one of the most lethal complications in children infected with Plasmodium falciparum. The pathogenesis of this anemia is not completely understood. Experimental data from malaria-infected humans and animal models suggest that uninfected red cells have a shortened life span. This study looked for changes in the red cell surfaces of children with severe malarial anemia that could explain this accelerated destruction. A prospective case-control study was conducted of children with severe P falciparum anemia (hemoglobin of 5 g/dL or lower) admitted to a large general hospital in western Kenya. Children with severe anemia were compared with children who had symptoms of uncomplicated malaria and with asymptomatic children. Cytofluorometry was used to quantify in vitro erythrophagocytosis and to measure red cell surface immunoglobulin G (IgG) and the complement regulatory proteins CR1, CD55, and CD59. Red cells from patients with severe anemia were more susceptible to phagocytosis and also showed increased surface IgG and deficiencies in CR1 and CD55 compared with controls. Red cell surface CD59 was elevated in cases of severe anemia compared with asymptomatic controls but not as compared with symptomatic controls. The surface of red cells of children with severe P falciparum anemia is modified by the deposition of IgG and alterations in the levels of complement regulatory proteins. These changes could contribute to the accelerated destruction of red cells in these patients by mechanisms such as phagocytosis or complement-mediated lysis. (Blood. 2000;95:1481-1486)

Malaria vaccines: triumphs or tribulations?

A safe and effective malaria vaccine will greatly facilitate efforts to control the global spread of malaria. This paper discusses the conceptual framework for developing malaria vaccines and some of the difficulties that the various approaches face. It emphasizes the role of pre-erythrocytic malaria vaccines, which are designed to protect against malaria infection, rather than simply prevent clinical disease. It describes recent encouraging results in human subjects with the RTS,S vaccine, a promising pre-erythrocytic malaria vaccine candidate.

Long-term efficacy and immune responses following immunization with the RTS,S malaria vaccine.

The malaria sporozoite vaccine candidate RTS,S, formulated with an oil-in-water emulsion plus the immunostimulants monophosphoryl lipid A and the saponin derivative QS21 (vaccine 3), recently showed superior efficacy over two other experimental formulations. Immunized volunteers were followed to determine the duration of protective immune responses. Antibody levels decreased to between one-third and one-half of peak values 6 months after the last dose of vaccine. T cell proliferation and interferon-gamma production in vitro were observed in response to RTS,S or hepatitis B surface antigen. Seven previously protected volunteers received sporozoite challenge, and 2 remained protected (1/1 for vaccine 1, 0/1 for vaccine 2, and 1/5 for vaccine 3). The prepatent period was 10.8 days for the control group and 13.2 days for the vaccinees (P < .01). Immune responses did not correlate with protection. Further optimization in vaccine composition and/or immunization schedule will be required to induce longer-lasting protective immunity.

Phase I/IIa safety, immunogenicity, and efficacy trial of NYVAC-Pf7, a pox-vectored, multiantigen, multistage vaccine candidate for Plasmodium falciparum malaria.

Candidate malaria vaccines have failed to elicit consistently protective immune responses against challenge with Plasmodium falciparum. NYVAC-Pf7, a highly attenuated vaccinia virus with 7 P. falciparum genes inserted into its genome, was tested in a phase I/IIa safety, immunogenicity, and efficacy vaccine trial in human volunteers. Malaria genes inserted into the NYVAC genome encoded proteins from all stages of the parasite's life cycle. Volunteers received three immunizations of two different dosages of NYVAC-Pf7. The vaccine was safe and well tolerated but variably immunogenic. While antibody responses were generally poor, cellular immune responses were detected in >90% of the volunteers. Of the 35 volunteers challenged with the bite of 5 P. falciparum-infected Anopheles mosquitoes, 1 was completely protected, and there was a significant delay in time to parasite patency in the groups of volunteers who received either the low or high dose of vaccine compared with control volunteers.

The current status of malaria vaccines.

A vaccine against Plasmodium falciparum malaria is needed now more than ever due the resurgence of the parasite and the increase in drug resistance. However, success in developing an effective malaria vaccine has been elusive. Among pre-erythrocytic antigens, the major antigen coating the surface of the sporozoite, the circumsporozoite protein (CS), has been, and continues to be, the major target for vaccine development. Despite initial limited success with CS-based vaccines, the use of new adjuvant formulations has led to the development of a promising candidate (the RTS,S vaccine) which has shown significant efficacy in a preliminary trial. In addition to CS, many other malaria antigens have been identified that play an important role in the parasite life cycle which are being considered for, or are currently undergoing, clinical trials. Among the blood stage antigens, the merozoite surface protein 1 (MSP-1) is the most promising vaccine candidate. New approaches to immunisation against malaria being considered include the use of multistage, multicomponent vaccines in attenuated viral vectors (NYVAC-Pf7), or in a combination DNA vaccine. While there is reason to be optimistic about the prospects for an effective vaccine, many challenges lie ahead that still have to be overcome. Among these are the antigenic polymorphism exhibited by wild parasite strains and the genetic restriction of immune responses.

A preliminary evaluation of a recombinant circumsporozoite protein vaccine against Plasmodium falciparum malaria. RTS,S Malaria Vaccine Evaluation Group.

BACKGROUND: The candidate vaccines against malaria are poorly immunogenic and thus have been ineffective in preventing infection. We developed a vaccine based on the circumsporozoite protein of Plasmodium falciparum that incorporates adjuvants selected to enhance the immune response. METHODS: The antigen consists of a hybrid in which the circumsporozoite protein fused to hepatitis B surface antigen (HBsAg) is expressed together with unfused HBsAg. We evaluated three formulations of this antigen in an unblinded trial in 46 subjects who had never been exposed to malaria. RESULTS: Two of the vaccine formulations were highly immunogenic. Four subjects had adverse systemic reactions that may have resulted from the intensity of the immune response after the second dose, which led us to reduce the third dose. Twenty-two vaccinated subjects and six unimmunized controls underwent a challenge consisting of bites from mosquitoes infected with P. falciparum. Malaria developed in all six control subjects, seven of eight subjects who received vaccine 1, and five of seven subjects who received vaccine 2. In contrast, only one of seven subjects who received vaccine 3 became infected (relative risk of infection, 0.14; 95 percent confidence interval, 0.02 to 0.88; P<0.005). CONCLUSIONS: A recombinant vaccine based on fusion of the circumsporozoite protein and HBsAg plus a potent adjuvant can protect against experimental challenge with P. falciparum sporozoites. After additional studies of protective immunity and the vaccination schedule, field trials are indicated for this new vaccine against P. falciparum malaria.

Induction of humoral immune response against Plasmodium falciparum sporozoites by immunization with a synthetic peptide mimotope whose sequence was derived from screening a filamentous phage epitope library.

The mouse monoclonal antibody 2A10 (immunoglobulin G), which recognizes the (NANP)n repeat of Plasmodium falciparum circumsporozoite surface protein, was used to screen a filamentous phage epitope library expressing random amino acid hexamers. The sequences obtained were TNRNPQ, SNRNPQ, NND-NPQ, SNYNPQ, and QNDNPQ (single-letter amino acid designation). These peptides showed 50% homology with the native epitope (PNANPN) and therefore were considered to mimic its structure (mimotopes). Two of these mimotopes (TNRNPQ and NNDNPQ) inhibited the binding of monoclonal antibody 2A10 to the recombinant protein R32LR, which contains the amino acid sequence [(NANP)15NVDP]2. Immunization of mice and rabbits using the peptide (TNRNPQ)4 induced a humoral response that recognized R32LR by an enzyme-linked immunosorbent assay and P. falciparum sporozoites by an immunofluorescence assay. These results suggest that phage epitope libraries can be exploited to screen for mimotopes in the design of subunit vaccines against infectious agents.

The battalion medical platoon: a flawed scalpel.

This article discusses the current organization and doctrine for a medical platoon in an armour-heavy task force in light of experience gained in Operations Desert Shield and Desert Storm. The organization of and doctrine for the use of the platoon is covered first. This is followed by a discussion of how doctrine was modified based on field exercises, and projected, and actual, combat operations. It concludes with a presentation of lessons learned and recommendations for changes.