Chabaudi AS Research Articles

Parasite‐specific IgM plays a significant role in the protective immune response to asexual erythrocytic stage Plasmodium chabaudi AS infection*

A comparison of Plasmodium chabaudi AS infection in BALB/c and BALB/c IgM-deficient mice demonstrated a protective role for IgM during infection. IgM-/- mice, unlike microMT mice, display competent B cell humoral immune responses. Increased susceptibility of IgM-/- mice was demonstrated by increased mortality, an advanced ascending infection and higher peak parasitaemia, as well as enhanced anaemia and weight loss compared with wild-type mice. The recrudescent parasitaemias were also higher in the IgM-/- mice. Early specific IgM production in P. chabaudi-infected wild-type mice was followed by IgG1 and IgG2a production, while IgG1 and IgG2a production in IgM-/- mice was preceded by specific IgD production. No protective role for natural IgM against P. chabaudi AS infection was detected as passive transfer of naïve WT serum into IgM-/- mice did not alter the disease outcome or reduce parasite numbers. Passive transfer of WT antiserum, containing predominantly specific IgM, into IgM-/- mice delayed the ascending parasitaemia and reduced mortality. Similarly, coating parasitized red blood cells with WT antiserum, but not IgM-/- antisera, prior to infection also slightly delayed the ascending acute parasitaemia. Specific IgM therefore plays an important role in the limitation of parasite replication during asexual erythrocytic P. chabaudi AS infection.

Role of CD28 in Polyclonal and Specific T and B Cell Responses Required for Protection against Blood Stage Malaria

The role of B7/CD28 costimulatory pathway in the polyclonal and specific lymphocyte activation induced by blood stages of Plasmodium chabaudi AS was investigated in CD28 gene knockout (CD28(-/-)) and C57BL/6 (CD28(+/+)) mice. Analysis of the spleen during the acute infection revealed a similar increase in T and B cell populations in both groups of mice. Moreover, CD28(-/-) mice were able to develop a polyclonal IgM response to P. chabaudi. On the contrary, the polyclonal IgG2a response was markedly reduced in the absence of CD28. Production of IFN-gamma; up-regulation of CD69, CD40L, CD95 (Fas), and CD95L (Fas ligand); and induction of apoptosis were also affected by the lack of CD28. Interestingly, the ability to control the first parasitemia peak was not compromised in acutely infected CD28(-/-) mice, but CD28(-/-) mice failed to eradicate the parasites that persisted in the blood for >3 mo after infection. In addition, drug-cured CD28(-/-) mice were unable to generate memory T cells, develop an anamnesic IgG response, or eliminate the parasites from a secondary challenge. The incapacity of CD28(-/-) mice to acquire a full protective immunity to P. chabaudi correlated with an impaired production of specific IgG2a. Moreover, reinfected CD28(-/-) mice were protected by the adoptive transfer of serum from reinfected CD28(+/+) mice containing specific IgG2a. Our results demonstrate that the polyclonal lymphocyte response is only partially affected by the absence of CD28, but this coreceptor is essential to generate specific T and B cell responses required for complete protection against P. chabaudi malaria.

Comparison of pathology in susceptible A/J and resistant C57BL/6J mice after infection with different sub-strains of Plasmodium chabaudi

Susceptible A/J and more resistant C57BL/6J mice were infected with Plasmodium chabaudi chabaudi 54X, P.c. chabaudi AS and Plasmodium chabaudi adami 408XZ. As expected, most C57BL/6J mice survived the infections with the different isolates. But in contrast to previous observations, not all A/J mice succumbed to infection: just over 50% of A/J mice survived infections with P.c. chabaudi 54X, while 80% survived P.c. chabaudi AS. The more virulent parasite, P.c. adami 408XZ, was able to kill all A/J mice and 20% of C57BL/6J mice after an intravenous infection with 10 5 pRBC. A detailed study of four parameters of pathology (body weight, body temperature, blood glucose and RBC counts) in both mouse strains after a P.c. adami 408XZ infection showed similar patterns to those previously reported after infection with P.c. chabaudi AS. These data suggest that environmental factors as well as parasite polymorphisms might influence the severity of malaria between susceptible and resistant mice.

Inappropriately low reticulocytosis in severe malarial anemia correlates with suppression in the development of late erythroid precursors

AbstractInappropriately low reticulocytosis may exacerbate malarial anemia, but the under-lying mechanism is not clear. In this study, naive and infected mice were treated with recombinant murine erythropoietin (EPO), and the upstream events of erythropoiesis affected by blood-stage Plasmodium chabaudi AS were investigated. Malaria infection, with or without EPO treatment, led to a suboptimal increase in TER119+ erythroblasts compared with EPO-treated naive mice. Furthermore, a lower percentage of TER119+ erythroblasts in infected mice were undergoing terminal differentiation to become mature hemoglobin-producing erythroblasts. The impaired maturation of erythroblasts during infection was associated with a shift in the transferrin receptor (CD71) expression from the TER119+ population to B220+ population. Moreover, the suboptimal increase in TER119+ erythroblasts during infection coincided with a blunted proliferative response by splenocytes to EPO stimulation in vitro, although a high frequency of these splenocytes expressed EPO receptor (EPOR). Taken together, these data suggest that during malaria, EPO-induced proliferation of early EPOR-positive erythroid progenitors is suppressed, which may lead to a suboptimal generation of TER119+ erythroblasts. The shift in CD71 expression may result in impaired terminal maturation of these erythroblasts. Thus, inadequate reticulocytosis during malaria is associated with suppressed proliferation, differentiation, and maturation of erythroid precursors.

Effect of anemia and renal cytokine production on erythropoietin production during blood-stage malaria

Renal dysfunction and severe anemia are clinical complications of blood-stage malaria. Erythropoietin (Epo) is a hormone produced by the kidney and plays an essential role in stimulating erythrocyte production. Renal dysfunction in malaria is associated with changes in renal cytokine levels, which may affect the production of Epo and the alleviation of anemia. Resistant C57BL/6 (B6) and susceptible A/J mice were infected with Plasmodium chabaudi AS. The levels of Epo and cytokines were measured by enzyme-linked immunosorbent assay (ELISA) and the degree of anemia was determined by hematocrit. Regression analyses were employed to estimate the influences of anemia and renal cytokines on the production of Epo during infection. A/J mice developed higher peak parasitemia, more severe anemia, and succumbed as compared to B6 mice, which survived the infection. B6 mice had higher levels of renal tumor necrosis factor-alpha (TNF-alpha) and interleukin (IL)-10, whereas A/J mice had higher levels of IL-12p70, granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-4, and Epo. Regression analyses revealed that kidney Epo levels were influenced most strongly by changes in hematocrit levels. In addition, albeit to a much weaker degree, kidney Epo levels correlated negatively with GM-CSF levels but positively with IL-10 levels. Blood-stage malaria infection modulates the production of renal pro- and anti-inflammatory cytokines in resistant versus susceptible strains of mice differentially. However, despite the fluctuations of renal cytokines, the degree of anemia is the main determinant for Epo production during blood-stage malaria while kidney cytokines may exert secondary influences.

Is the expression of genes encoding enzymes of glutathione (GSH) metabolism involved in chloroquine resistance in Plasmodium chabaudi parasites?

The genes encoding enzymes involved in glutathione (GSH) metabolism may modulate responses to antimalarial drugs, but the role of most of them in antimalarial drug resistance has scarcely been investigated. Using an in silico/PCR combined approach, we have isolated from Plasmodium chabaudi, full sequences of five Plasmodium falciparum gene orthologues involved in GSH metabolism: the gamma-glutamylcysteine synthetase (Pc-gammagcs), glutathione-synthetase (Pc-gs), glutathione peroxidase (Pc-gpx), glutathione reductase (Pc-gr) and glutathione-S-transferase (Pc-gst). DNA sequencing of these genes from drug sensitive parasites, P. chabaudi AS (0CQ), and ones isolated from parasite lines that show genetically stable resistance to chloroquine (CQ) at low, intermediate and high levels, AS (3CQ), AS (15CQ) and AS (30CQ), respectively, revealed no point mutations in the resistant parasites. We used these sequences to design internal oligonucleotide primers to compare relative mRNA amounts of these genes between all P. chabaudi clones, in untreated mice or following CQ treatment with sub-curative doses, by real-time PCR. Analysis of three independent experiments revealed that transcription levels of the Pc-gammagcs, Pc-gs, Pc-gpx, Pc-gr and Pc-gst genes were not changed between chloroquine sensitive and resistant parasite clones, and that treatment with chloroquine did not induce an alteration in the expression of these genes in sensitive or resistant parasites. We concluded that chloroquine resistance in this species is determined by a mechanism that is independent of these genes, and most likely, of GSH metabolism.

Phenotypic expression of pyruvate kinase deficiency and protection against malaria in a mouse model.

The recombinant congenic mouse strains AcB55 and AcB61 are extremely resistant to malaria (Plasmodium chabaudi AS) despite the presence of susceptibility alleles at the known Char1/Char2 resistance loci. Resistance in AcB55 and AcB61 is controlled by a locus on chromosome 3 (Char4) shown to be allelic with or tightly linked to a loss-of-function mutation in pyruvate kinase (Pklr). AcB55 and AcB61 show important splenomegaly prior to infection caused by the expansion of the red pulp, and display histological signs of extramedullary erythropoiesis in the liver. Examination of splenic cell populations by flow cytometry demonstrates elevated numbers of TER119-positive erythroid precursor cells (>30% of total spleen cells), while RNA expression studies show elevated expression of erythrocyte-specific transcripts such as globin, transferrin receptor, and Nramp2/Slc11a2 in the spleen of both strains. Hematological profiling in both strains is consistent with the presence of anemia as evidenced by low total erythrocyte counts, decreased hemoglobin, as well as abnormally high numbers of circulating reticulocytes (15-20%). These results strongly suggest that the mutant Pklr allele (Pklr(269A)) of AcB55/61 strains causes hemolytic anemia compensated by constitutive erythropoiesis, which in turn protects the mice against P. chabaudi infection. The possible molecular basis of the Pklr protective effect is discussed and is under current investigation in these two strains.

Early treatment during a primary malaria infection modifies the development of cross immunity.

We have used a murine model to study the kinetics of cross-protection when a primary infection is halted at different times. We analysed how parasitaemia is modified during a second infection with the homologous parasite, a heterologous parasite, or a mixture of the two. In addition, possible mechanisms involved in cross-protection were analysed. Results show that treatment with pyrimethamine on day 5 during a primary infection with P. chabaudi AS (non-lethal), prevents the generation of cross-protection to a new challenge with lethal P. yoelii 17XL. In contrast, when treatment is on day 7, mice survive a P. yoelii infection. Differences between both groups suggest that in order for 'preimmune' mice to survive a lethal challenge, a predominantly TH2-type response is required, with a higher mRNA expression level of IL-4 and IL-10, and a lower mRNA expression of IFN-gamma. This work shows that an early treatment of a malaria infection produced by a non-lethal parasite drives the immune response towards a loss of cross-protection to further infections, in particular with more virulent parasites. This finding should be taken into account for the development of effective malaria vaccines.

Different responses of three rodent Plasmodia species, Plasmodium yoelii 17XL, P. berghei NK65 and P. chabaudi AS on treatment with febrifugine and isofebrifugine mixture from Hydrangea macrophylla var. Otaksa leaf in ICR mice.

The antimalarial activity of Hydrangea macrophylla var. Otaksa alkaloids was evaluated against Plasmodium yoelii 17XL, P. berghei NK65 and P. chabaudi AS in ICR mice. For trials in P. yoelii 17XL or P. chabaudi AS infections, mice were infected intraperitoneally with 10(5), 10(6) and 10(7) parasitized erythrocytes, respectively, and in P. berghei NK65 infections, mice were infected intraperitoneally with 10(3), 10(4) and 10(5) parasitized erythrocytes, respectively. Three days after injection, mice were orally given febrifugine and isofebrifugine mixture at 1 mg/kg in the treated group and 0.5% cremophor EL solution in the untreated, infected one, respectively, twice a day for 5 consecutive days. In P. yoelii 17XL infections, mice in all the non-treated controls died from 5 to 9 dpi with a gradual body weight loss and increasing parasitemias. In the treated groups, the mouse body weight gradually decreased after the end of administration but turned to increase in several days, and except one mouse in the group given 10(6) parasitized erythrocytes, other mice survived during the experiment. Mice given orally the mixture showed low parasitemia levels during administration. Following a transient recrudescence of malaria parasites in the bloodstream of treated mice, no parasites could be detected by a microscopic examination. In P. berghei NK65 infections, mice in all the non-treated controls died from 7 to 12 dpi with a gradual body weight loss and increasing parasitemias. In the treated groups, the body weight gradually decreased from 11 dpi and all mice died from 12 to 30 dpi. During a mixture administration all mice showed slight suppression of multiplication of malaria parasites. After the end of administration, however, malaria parasites increased in the bloodstream of the treated mice and all mice died. In P. chabaudi AS infections, there were two different patterns in the course of infection; lethal infection or recovery in both the non-treated control and treated groups. In the non-treated and treated groups, mice showed a gradual body weight loss. But the body weights of survivals in both groups turned to increase in several days. Mice in control and treated groups showed as the same profile in the changes of parasitemia. In the non-treated controls, after a transient peak parasitemia malaria parasites in the bloodstream of survivals could not be detected by a microscopic examination. During a mixture administration, all mice showed suppression of multiplication of malaria parasites. After the end of medication, some mice died with increasing parasitemia. After a transient recrudescence, however, malaria parasites in the bloodstream of survivals could not be detected by a microscopic examination.

Plasmodium chabaudi chabaudi AS: modification of acute infection in CBA/Ca mice as a result of pre-treatment with erythrocyte band 3 in adjuvant

In this paper, in vivo data are presented that suggest a role for host recognition of erythrocyte band 3 in the control of malaria parasitaemia. The course of Plasmodium chabaudi chabaudi AS acute infection in CBA/Ca mice was suppressed or enhanced as a result of treatment on two occasions with enriched preparations of normal erythrocyte band 3 in adjuvant. Co-treatment with band 3 and a recombinant polypeptide encoding the C-terminal region of the P. c. chabaudi AS merozoite surface protein 1, which on its own had no clear effect on parasitaemia, appeared to modulate band 3-induced inhibition. Despite several-fold reductions in ascending parasitaemias in some band 3-immunized groups, there was a lack of obvious or unexpected anaemia prior to, or during infection, indicating a degree of specificity in the parasitaemia modifying response for infected rather than uninfected erythrocytes. These findings support a role for modified host recognition of erythrocyte band 3 in the partial immunity that transcends phenotypic and genotypic antigenic variation by malaria parasites. Index Descriptors and Abbreviations: malaria; Plasmodium chabaudi; band 3; MSP-1, merozoite surface protein 1; MBP, maltose binding protein; E, erythrocyte; PE, parasitized erythrocyte; PBS, phosphate-buffered saline.

Comparison of murine Epo ELISA and Epo bioassays in detecting serum Epo levels during anemia associated with malaria infection

A highly sensitive sandwich ELISA specific for murine erythropoietin (mEpo) was developed using commercially available monoclonal anti-mouse Epo antibody and polyclonal anti-human Epo antibody. This newly developed Epo ELISA protocol and the traditional Epo bioassay method were used to analyze Epo production in response to anemia induced during blood-stage Plasmodium chabaudi AS ( P. chabaudi AS) malaria infection in C57BL/6 mice. Both methods revealed an inverse correlation between the serum Epo concentration and hematocrit level, but Epo values estimated by the Epo bioassay were between 5- and 20-fold higher than those estimated by the ELISA. Further study demonstrated that the estimated Epo level in bioassay was strongly influenced by other cytokines present in the samples. Therefore, the Epo bioassay detects the net erythropoiesis promoting activities, whereas the ELISA method specifically measures the level of Epo in the samples. Combined with the Epo bioassay, the murine Epo ELISA will be an extremely useful tool in specifically measuring the Epo response and facilitating the understanding of mechanisms involved in the development of anemia-associated diseases using mouse models.

Mouse splenic CD4+ and CD8+ T cells undergo extensive apoptosis during a Plasmodium chabaudi chabaudi AS infection.

The presence and phenotype of apoptotic lymphocytes was studied in spleen cell suspensions taken from CB6F1 mice infected with Plasmodium chabaudi chabaudi AS. High levels of apoptotic cells were found, associated with high parasitaemias and splenomegaly. This was also accompanied by expansion and disarray of spleen white pulp. Apoptosis levels lowered when parasitaemia was cleared, but were still higher than in normal mice. At this time, the spleen was diminishing in size and the white pulp was contracting and rearranging. When parasitaemia was patent, the cells most affected by apoptosis were CD4+ T cells followed by CD8+ T cells, and to a lesser extent B220+ B cells. When parasitaemia was cleared, CD8+ T cells and B220+ B cells returned to basal levels of apoptosis, while CD4+ T cells still had higher apoptosis levels than normal mice. A similar pattern of lymphocyte subpopulation apoptosis was found in infected BALB/c mice, despite the fact that, for this mouse model, it has been reported that B cells are the cells that are most affected by apoptosis. We consider that the high levels of apoptosis in CD4+ T cells when parasitaemias are still high are not easily explained by a normal mechanism of down regulation of the immune response.

Nitric oxide and reactive nitrogen intermediates during lethal and nonlethal strains of murine malaria.

The virulence of Plasmodia depends partly on the strain of parasite and partly on the host. In this study, Plasmodium berghei N/13/1A/4/203 caused the death of mice, whereas Plasmodium chabaudi chabaudi AS was not lethal. Current opinion is that nitric oxide (NO) and other reactive nitrogen intermediates (RNI) are produced in several host organs during malaria to resist infection or produce tissue damage. NO and RNI production in blood or plasma, brain, liver and spleen in MF1 mice was investigated during P. berghei and P. c. chabaudi infection, in order to help determine whether changes in NO production are beneficial or detrimental to the host in vivo. NO production was measured both directly and indirectly as nitrites and nitrates, to represent RNI. No changes in blood NO were detected in P. berghei infected mice, but increases were observed in brain, liver and spleen. In P. c. chabaudi infected mice, rises in NO concentration were observed in blood and spleen, whereas a decline in liver NO was seen, but there were no changes in brain. Liver contained the highest concentration of RNI, but increasing concentrations were seen in both plasma and spleen in both P. berghei and P. c. chabaudi infected mice. These results show that NO and RNI production alters during murine malaria. The changes depend upon the tissue, the day of infection, the degree of parasitaemia, the strain of Plasmodia and the method of measuring NO biosynthesis. Lethal P. berghei induced NO production in the mid and late stages of infection in mice when parasitaemia was high, whereas in nonlethal P. c. chabaudi infection, NO production was increased in the early and late stages when parasitaemia was low. These data are consistent with a role for NO in the protection of the MF1 mouse against Plasmodia. Failure to clear the parasite is associated with evidence of increased NO production in brain and liver, which may contribute to the pathology of malaria, but this hypothesis requires confirmation from other experimental approaches.

Possible Modulation by Male Sex Hormone of Th1/Th2 Function in Protection against Plasmodium chabaudi chabaudi AS Infection in Mice

Zhang, Z.-H., Chen, L., Saito, S., Kanagawa, O., and Sendo, F. 2000. Possible modulation by male sex hormone of Th1/Th2 function in protection against Plasmodium chabaudi chabaudi AS infection in mice. Experimental Parasitology96, 121–129. We examined the mortality, survival time, and parasitemia in interferon γ receptor (IFN-γR)-deficient (IFN-γR−/−) and IL-4-deficient (IL-4−/−) mice infected with Plasmodium chabaudi AS and compared them with the wild type counterparts (IFN-γR+/+ and IL-4+/+, respectively). (1) Mortality was higher and survival time was shorter in males of both IFN-γR−/− and IL-4−/− mice infected with P. chabaudi AS, compared with their wild type counterparts, whereas such a difference was not observed in female mice. (2) These differences between males and females were not observed when male mice were castrated; however, female castration had no effect on the data. (3) The rate of parasitemia in both male and female IFN-γR−/− and IL-4−/− mice was higher at some points during the observation than in the wild type counterparts. (4) These results on susceptibility vs resistance to P. chabaudi AS infection can be explained partially by the levels of expression of Th1/Th2 cytokine and chemokine mRNAs in the spleen cells of the infected mice. These results suggest that male sex hormones modulate the function of Th1/Th2 cells and that these T cells counteract the activity of these hormones in protection against P. chabaudi AS infection in mice.

Protective immunity to erythrocytic Plasmodium chabaudi AS infection involves IFNgamma-mediated responses and a cellular infiltrate to the liver.

IFNgamma receptor (IFNgammaR) deficient mice and IL-4 deficient mice were infected with blood-stage Plasmodium chabaudi AS in order to analyse the role of Th1 (IFNgamma) and Th2 (IL-4)-associated cytokines in the development of protective immunity to the parasite. A high mortality rate and failure to reduce the primary parasitaemia to subpatent levels was observed in the IFNgammaR deficient mice. IL-4 deficient mice controlled a primary P. chabaudi AS infection in a similar manner to control mice and no mortality was observed. IFNyR deficient mice had a reduction in parasite-specific IgG and a significantly increased level of total IgE compared to control mice. There was no reduction in the level of parasite-specific IgG in IL-4 deficient mice. Cytological analysis of the cells present in the spleen and liver during the primary parasitaemia revealed a reduction in the numbers of lymphocytes, monocytes and polymorphonuclear (PMN) cells in the liver at the peak of parasitaemia in both IFNgammaR deficient mice and IL-4 deficient mice compared to control mice. Adoptive transfer studies demonstrated that cells isolated from the liver at day 11 post-infection could confer some protective immunity to P. chabaudi AS infection.

MMCP-8, the first lineage-specific differentiation marker for mouse basophils. Elevated numbers of potent IL-4-producing and MMCP-8-positive cells in spleens of malaria-infected mice

In mice infected with the non-lethal malaria parasite Plasmodium chabaudi chabaudi AS, a prominent switch from a Th1 to a Th2 type of response occurs in CD4+ T cells at the time of peak parasitemia or shortly thereafter (9-15 days after infection). This is accompanied by a major increase in IL-4, and a similar decrease in IFN-gamma-producing cells. Non-B-non-T cells have been shown to be the main source of the IL-4 in these mice. The IL-4-producing cells are hyperresponsive to IL-3, indicating mast cell or basophil origin. To further characterize this cell population we have studied various organs at different time points of malarial infection by Northern blot analysis. No significant increase in the expression of any of the classical mouse mast cell serine proteases (MMCP)-1 to 7 or carboxypeptidase A was detected in the spleen during the entire infection. However, a marked increase in the expression of MMCP-8 was observed in the spleen at around day 15 post infection. Isolation of IgE receptor-positive cells from spleen shortly after peak parasitemia led to a prominent enrichment of MMCP-8-expressing cells. Fifty thousand of these cells were, after IL-3 stimulation, found to produce IL-4 to levels comparable with more than one million fully activated T cells. Our results show that basophil-like cells are very potent producers of IL-4 and that IL-4 produced by these cells may be of major importance for the initiation of a Th2 response. In addition, the detection of MMCP-8 in these cells has led to the identification of the first basophil-specific differentiation marker in the mouse.

Central role of endogenous gamma interferon in protective immunity against blood-stage Plasmodium chabaudi AS infection.

The role of endogenous gamma interferon (IFN-gamma) in protective immunity against blood-stage Plasmodium chabaudi AS malaria was studied using IFN-gamma gene knockout (GKO) and wild-type (WT) C57BL/6 mice. Following infection with 10(6) parasitized erythrocytes, GKO mice developed significantly higher parasitemia during acute infection than WT mice and had severe mortality. In infected GKO mice, production of interleukin 12 (IL-12) p70 and tumor necrosis factor alpha in vivo and IL-12 p70 in vitro by splenic macrophages was significantly reduced compared to that in WT mice and the enhanced nitric oxide (NO) production observed in infected WT mice was completely absent. WT and GKO mice had comparable numbers of total nucleated spleen cells and B220(+) and Mac-1(+) spleen cells both before and after infection. Infected WT mice, however, had significantly more F4/80(+), NK1.1(+), and F4/80(+)Ia(+) spleen cells than infected GKO mice; male WT had more CD3(+) cells than male GKO mice. In comparison with those from WT mice, splenocytes from infected GKO mice had significantly higher proliferation in vitro in response to parasite antigen or concanavalin A stimulation and produced significantly higher levels of IL-10 in response to parasite antigen. Infected WT mice produced more parasite-specific immunoglobulin M (IgM), IgG2a, and IgG3 and less IgG1 than GKO mice. Significant gender differences in both GKO and WT mice in peak parasitemia levels, mortality, phenotypes of spleen cells, and proliferation of and cytokine production by splenocytes in vitro were apparent during infection. These results thus provide unequivocal evidence for the central role of endogenous IFN-gamma in the development of protective immunity against blood-stage P. chabaudi AS.

The Effect of Chloroquine on the Production of Interferon-g, Interleukin (IL)-4, IL-6, and IL-10 in Plasmodium chabaudi chabaudi in Infected C57BL6 Mice

The effect of chloroquine (CQ) on the production pattern of interferon (IFN)-gamma, interleukin (IL)-4, IL-6, and IL-10 in female C57BL6 mice infected with Plasmodium chabaudi chabaudi AS was evaluated during a period of 35 days. Our data confirm that there is a switch from a T helper cell (Th)1 to a Th2 response during malaria infection in this model. Proliferation assays showed a decreased stimulation index in infected mice that was further reduced in infected mice treated with CQ. Noninfected control mice treated with CQ showed an increase production of IFN-gamma. However, no detectable changes in IL-4, IL-6, and IL-10 production were observed in this group. CQ treatment of infected mice resulted in parasite clearance that was associated with an earlier production of IL-4, IL-6, and IL-10 when compared with nontreated infected mice. We suggest that this earlier switch to a Th2 response is a consequence of parasite killing rather than CQ interference with cytokine production.

In Vivo IL-12 Production and IL-12 Receptors β1 and β2 mRNA Expression in the Spleen Are Differentially Up-Regulated in Resistant B6 and Susceptible A/J Mice During Early Blood-StagePlasmodium chabaudiAS Malaria

AbstractAs previously reported, blood-stage Plasmodium chabaudi AS malaria is lethal by days 10–12 postinfection in susceptible A/J mice that mount an early, predominantly Th2 response. In contrast, resistant C57BL/6 (B6) mice clear the infection by 4 wk with an early Th1 response. In this study, we analyzed in vivo production of IL-12, a potent Th1-inducing cytokine, during the first 5 days after P. chabaudi AS infection in these mice. By day 2, serum IL-12 p70 levels were significantly increased in B6 mice over basal levels and were also significantly higher compared with A/J mice that showed no significant changes in serum p70 levels after infection. Splenectomy of resistant B6 mice before infection demonstrated that the spleen is the major source of systemic IL-12 in these hosts. Splenic mRNA levels of both p40 and p35 were significantly higher in A/J mice; however, the ratios of p40/p35 mRNA levels were similarly up-regulated in both strains. Furthermore, B6 but not A/J mice showed significant up-regulation of splenic IL-12R β2 mRNA over basal levels by days 3 and 4, coincident with sustained up-regulation of splenic IFN-γ mRNA levels on days 3–5. However, IL-12R β1 mRNA levels in the spleen were similarly up-regulated in both mouse strains by day 3. Taken together, these data suggest that high systemic IL-12 production, accompanied by an early and sustained up-regulation of both IL-12R β1 and β2 mRNA levels in the spleen, as occurs in resistant B6 mice, appears to preferentially induce protective Th1 responses against blood-stage malaria.

Altered immune responses in mice with concomitant Schistosoma mansoni and Plasmodium chabaudi infections.

Mixed parasitic infections are common in many parts of the world. However, little is known about how concurrent infections affect the immunity to and/or pathogenesis of each other. Protection and elimination of blood-stage Plasmodium chabaudi chabaudi AS in resistant mice are characterized by a sequential activation of CD4(+) Th1 and Th2 cells. The patent egg-laying stage of the murine model of Schistosoma mansoni is associated with a strong Th2 response to both Schistosoma and unrelated antigens. In this study, we investigated how infection of mice with S. mansoni would affect the immune response to and pathogenesis of a P. chabaudi infection. C57BL/6 mice infected with S. mansoni for 8 weeks were infected with blood-stage P. chabaudi. Malaria parasitemias were significantly higher in these mice than in mice infected with P. chabaudi only. In doubly infected mice, both spleen cell proliferative and Th2 responses to S. mansoni soluble egg antigen (SEA) or anti-CD3 were suppressed up to 1 month after the malaria infection. Findings for SEA-specific immunoglobulin M (IgM) and IgG serum antibody levels were similar. No significant effects were seen on P. chabaudi-induced gamma interferon responses. However, tumor necrosis factor alpha (TNF-alpha) production was significantly lower in double-infected mice. Thus, a defect in TNF-alpha production might contribute to the increased malaria parasitemias seen in S. mansoni-P. chabaudi-infected mice. Taken together, our data show that schistosoma and malaria infections profoundly affect each other, findings which might have implications for the development of vaccines.