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We characterize the mouse gene imap38 and its inducibility by Plasmodium chabaudi malaria among different lymphoid tissues and mouse strains of different H-2 complex and non-H-2 background. Imap38 is a single copy gene assigned to chromosome 6B. It consists of only one exon of 1900 base pairs encoding a highly basic 25.8-kDa protein. Confocal laser scanning microscopy localizes differently tagged IMAP38 proteins in nuclei of transfected cells. Reporter gene assays reveal that the 1730-base pair 5'-flanking region, containing an RSINE1 repeat immediately adjacent to initiation site +1, exhibits promoter activity in nonmurine cells, while it is largely repressed in diverse mouse cell lines, which corresponds to the situation in mouse tissues. P. chabaudi malaria induces imap38 expression almost exclusively in the spleen but not in other lymphoid organs. Parasite lysates are able to induce imap38 in the spleen, but not in spleen cells ex vivo. Activation of spleen cells by LPS and other stimuli is not sufficient to induce imap38. Inducibility of imap38 requires signals from both parasites and the intact spleen, and it is controlled by genes of that non-H-2 background, which also controls development of protective immunity against P. chabaudi malaria.

We report the identification of a novel mouse cDNA encoding IAP38, a putative plasma membrane protein of 38 kDa in splenic macrophages, B cells and T cells. The expression of iap38 is induced by blood-stage infections of Plasmodium chabaudi malaria and is testosterone-sensitive in non-immune mice. However, when mice have acquired testosterone-resistant immunity to P. chabaudi, there is an about 40-fold increase in the expression of iap38, which has then largely lost its responsiveness to infection and testosterone. The gene iap38 is suggested to be involved in imparting spleen cells the ability to mediate testosterone-resistant immunity to P. chabaudi malaria.

The immunogenicity of a 93-kDa Plasmodium chabaudi protein that contains glutamate-rich tandem repeats was investigated in this study. Immunoblotting with various monoclonal antibodies indicates that this 93-kDa protein is equivalent to a potential P. chabaudi RESA analogue. However, the sequence of the P. chabaudi protein does not exhibit any significant homology to Pf155/RESA. Antibodies against the 93-kDa protein appear early during P. chabaudi infection and reach high titers. The highest antibody titers are found when the parasitemia is descending, suggesting that this protein may play some role in immunity. Immunization of mice with the recombinant protein also results in high antibody titers, indicating that the protein is quite immunogenic. However, mice immunized with recombinant protein and challenged with P. chabaudi do not exhibit a delayed appearance of parasitemia, a reduced parasitemia, or a shortened duration of parasitemia. Glutamate-rich P. falciparum proteins such as Pf155/RESA, are being considered as vaccine candidates. The studies with P. chabaudi suggest that interpretation of serological data using glutamate-rich proteins should proceed with caution. The glutamate-rich repeats, although highly immunogenic, may not be important in host immunity against malaria. However, antibodies that appear late in the P. chabaudi infection do appear to play a role in anti-malarial immunity.

Vaccination with surface membranes isolated from Plasmodium chabaudi-infected erythrocytes can protect B10.A mice from the lethal outcome of P. chabaudi malaria. However, the efficacy depends on gender and testosterone levels. Thus, vaccination protects over 90% of female mice, but only about 55% of male mice and only about 34% of female mice when pretreated with testosterone for 4 weeks. The suppressive testosterone effect remains imprinted in females even at 10 weeks after the testosterone treatment. These data indicate that not only genetic but also environmental factors restrict the host's immune response to a malaria vaccine.

Testosterone (Te) is known to suppress immunity and to increase host susceptibility to many parasites. This study investigates the action of Te on immunity acquired against blood-stages of the malaria parasite Plasmodium chabaudi in female mice of the inbred strain C57BL/10. Our data show: (i) About 90% of mice infected with 10(6) P. chabaudi-infected erythrocytes are able to develop protective immune mechanisms which become evident in self-healing the infection. The capability of self-healing is lost when mice are pretreated with Te for 3 weeks. (ii) Mice which have self-healed infections acquire immunity to homologous rechallenge. Concomitantly, mice become Te-unresponsive in that their acquired immunity is not suppressible by Te-treatment. (iii) Flow cytometry reveals that Te-pretreatment entails an increase of CD8+ cells and a decrease of Ig+ cells by about 4% in spleens of non-immune mice. In immune mice, however, there is a Te-unresponsiveness of the percental distribution of splenic cell populations. (iv) Adoptive transfer experiments indicate that immunity is conferred by spleen cells, presumably non-T-cells. These cells are Te-unresponsive, since they exert their effect in Te-pretreated mice in the presence of Te. (v) Te-unresponsive immunity can be also transferred by serum, especially the IgG-fraction, obtained from immune mice. Our data demonstrate that Te prevents the development of protective immunity against P. chabaudi infections. However, when once established, protective immunity becomes unresponsive to Te. Our data suggest that the effector mechanisms of protective immunity involve Te-unresponsive B cells secreting protective IgG-antibodies.

The effect of circulating concentrations of testosterone (Te) on resistance to Plasmodium chabaudi malaria was investigated in the H-2 congenic mouse strains C57BL/10, B10.A, B10.A(3R), B10.A(4R), and B10.D2. Te-levels were determined by radioimmunoassay and resistance was expressed in terms of percent self-healers after challenge with 10(6) P. chabaudi-infected erythrocytes. Our data indicate: (i) Females and castrated males reveal very similar interstrain variations of resistance. These do not correlate with the interstrain variations of the Te-levels. This is consistent with the view that resistance to P. chaubaudi is controlled by genes of the H-2 complex and genes of the non-H-2 B10-background, (ii) The polygenic control of resistance is inefficacious at high Te-levels. This is evident as high susceptibilities of males, Te-treated females and Te-treated castrated males. Moreover, high Te-levels correlate with susceptibilities to P. chabaudi within mice of the same sex of a given strain, (iii) B10-males chemically castrated using buserelin display the same low Te-level as those surgically castrated. The latter become resistant, while the former remain as highly susceptible to P. chabaudi as untreated B10-males. Obviously, other gonadal factor(s), besides Te, impose restrictions on genes controlling resistance to P. chabaudi malaria.

This study investigates the effects of the male sex hormone, testosterone (Te), on self-healing of Plasmodium chabaudi malaria as well as on protein expression and functional properties of total spleen cells and splenic T cells in females of the mouse strain C57BL/10. About 90% of the B10 females survive a challenge with 10(6) P. chabaudi-infected erythrocytes. The percentage of self-healers, however, is reduced to about 60%, 40%, and 0% after pretreatment with Te for 1, 2, and 3 weeks, respectively. The progressive loss of the capability of self-healing is correlated with an increasing expression of five proteins in splenic non-T cells as revealed by two-dimensional fluorography after metabolic labelling of total spleen cells and T cells with [35S]methionine. These have molecular masses (isoelectric points) of about 10 kDa (pI 5.7), 14 kDa (pI 6.3), 14 kDa (pI 6.4), 38 kDa (pI 6.5), and 46 kDa (pI 5.5), respectively. Splenic non-T cells from mice treated with Te for 3 weeks have gained an increased capability to stimulate the concanavalin A-induced proliferative response of T cells. Te induces the changes in functional properties and protein expression of splenic non-T cells only in vivo and not in vitro. This suggests that the changes in splenic non-T cells as well as the prevention of self-healing P. chabaudi malaria are not directly induced by Te but rather indirectly, i.e. by a Te metabolite and/or Te-induced factor(s).