The relation between the immune response and the clinical features of severe falciparum malaria was studied in Burundian adults with (n = 31) and without (n = 17) cerebral involvement. At the time of admission, mean values for age, temperature, and blood levels of hemoglobin, creatinine, bilirubin, and glucose were similar in the two groups. Plasma levels of tumor necrosis factor alpha, interferon gamma, interleukin 10 (IL-10), and soluble intercellular adhesion molecule 1 were similarly elevated in the two groups. Mean parasite counts and mean plasma levels of soluble E-selectin were higher in severe noncerebral malaria than in cerebral malaria and were correlated with each other. After adjustment for parasitemia, levels of soluble E-selectin remained higher in noncerebral malaria. All seven patients who died had cerebral disease. These patients had higher levels of creatinine, bilirubin, IL-10, and soluble E-selectin than did patients with nonfatal cerebral malaria. After adjustment for creatinine and bilirubin levels, IL-10 and soluble E-selectin concentrations were similar in fatal and nonfatal cases of cerebral infection. In these African adults, none of the immunologic variables investigated was specific to cerebral malaria or to a fatal outcome.

Adolescent↗

PubMed1994

High levels of circulating IL-10 in human malaria.

IL-10 is a monocyte/lymphocyte derived cytokine which has been shown to inhibit certain cellular immune responses such as delayed hypersensitivity. In particular, the production of tumour necrosis factor (TNF), IL-1 and IL-6, which are involved in malaria pathology, are strongly inhibited by IL-10. Accordingly, we examined whether IL-10 could be involved in a human acute parasitic infection such as Plasmodium falciparum malaria. Human IL-10 levels in plasma were determined by two-site ELISA method, taking care to avoid non-specific reactions due to autoantibodies. Fourteen cerebral, 11 severe, and 20 mild malaria cases had mean IL-10 levels of 2812, 2882 and 913 pg/ml, respectively, while 98% of healthy individuals had undetectable (less than 100 pg/ml) circulating IL-10. Thirteen of the 25 cerebral/severe cases had > 2000 pg/ml. In 11 hospitalized patients, circulating IL-10 levels were found to return to virtually normal levels 7 days after antimalarial chemotherapy when biological and clinical malaria features had disappeared (mean levels fell from 3880 to 333 pg/ml). Further studies are required to determine whether these elevated levels of IL-10 play a beneficial role by reducing the parasite-induced inflammatory response, or a detrimental one by decreasing the cellular immune responses.

Adolescent↗

PubMed1994

The multifactorial and multistage character of protective immunity to Plasmodium falciparum, naturally acquired by an indigenous population in Burkina Faso.

In malaria-endemic areas, protective immunity is acquired gradually. Some authors have proposed that different stages can be distinguished during development. To test this hypothesis, several in vitro assays of the host immune response to P. falciparum were performed in three groups of individuals: 'unprotected' children with clinical attacks, 'semi-immune' children, without clinical attacks but with transient high parasitaemias during the transmission period, and 'protected' adults with low residual parasitaemias. By comparison of immune responses in these groups and multifactorial analyses, discriminant factors and potential protective mechanisms were identified. Anti-RESA antibody levels were lower in 'unprotected' than in 'semi-immune' children, while specific cellular responses, TNF levels and percentage of activated T lymphocytes were higher. Low humoral immunity and high cellular activation in children were followed by high humoral immunity and low cellular activation in adults. Therefore, protective immunity seems to pass through different stages and to result from the association of different immune mechanisms according to the level and duration of the individual experience of malaria.

Adolescent↗

PubMed1994

Soluble intercellular adhesion molecule-1 and E-selectin levels in plasma of falciparum malaria patients and their lack of correlation with levels of tumor necrosis factor alpha, interleukin 1 alpha (IL-1 alpha), and IL-10.

Intercellular adhesion molecule-1 and E-selectin levels were increased in the plasma of 60 falciparum malaria patients and were not related to levels of tumor necrosis factor alpha, interleukin 10, or interleukin 1 alpha. Soluble E-selectin was correlated to disease; its level in plasma was related to levels of both tumor necrosis factor soluble receptors and biological markers of disease severity and returned to baseline after parasite clearance faster than that of soluble intercellular adhesion molecule-1.

Animals↗

PubMed1993

An improved assay for the detection of Toxoplasma gondii antibodies in human serum by flow cytometry.

The diagnosis of toxoplasmosis relies primarily on the demonstration of specific antibodies to Toxoplasma gondii. We describe a flow cytometry method for the determination of antibodies to whole fixed tachyzoites by indirect immunofluorescence. Fixed tachyzoites in suspension have characteristic light scattering properties. The amount of IgM, IgG, and IgA antibodies from patients' sera bound to the tachyzoites can be estimated from the mean fluorescence intensities observed using class-specific conjugates for different Ig heavy chains. Appropriate serum dilutions for the estimation of specific IgG titres and for the discrimination between sera positive and negative for IgA and IgM antibodies were established using 40 random sera from pregnant women. The method proved to be quantitative and highly sensitive as compared with currently used assays. Coefficients of variation between series ranged from 6.4% to 12.2% and could be controlled by the inclusion of positive and negative standard sera in the test series. The method might represent a major improvement in cost effectiveness as compared with routine immunofluorescence assay by fluorescent microscopy when the volume of testing is sufficient. It can be fully automated, and its large scale application is currently under investigation.

Animals↗

PubMed1993

Neopterin levels in plasma during a longitudinal study in an area endemic for malaria.

Neopterin levels in plasma were measured during a longitudinal study of 75 patients living in an area endemic for malaria. The maximum levels were observed in August/September, the period of peak transmission of the disease. During the dry season, levels remain elevated. No relationship was found between the level of neopterin and the individual's immune status against malaria. Tumor necrosis factor (TNF), activated T lymphocytes subpopulations, soluble interleukin 2 receptors, and anti-circumsporozoite protein antibodies were not correlated with neopterin levels during the period of peak transmission. In contrast, neopterin levels were correlated with anti-Plasmodium falciparum antibodies detected by immunofluorescence. It was concluded that, in a population of chronic parasite carriers subjected to repeated infestations, neopterin was a good indicator of slowly acquired immunity in stable transmission area, but it poorly reflected acute immune responses.

Adolescent↗

PubMed1993

Soluble interleukin-2 receptor is involved during systemic mycosis.

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Adult↗

PubMed1993

Ring-infected erythrocyte surface antigen (Pf/155RESA) induces tumour necrosis factor-alpha production.

Cerebral malaria is probably related to an overstimulation of the immune system and the cytokine network. We have previously demonstrated that tumour necrosis factor (TNF) secretion by human macrophages can be induced by soluble and heat-stable malarial antigens. Indirect evidence from epidemiological and in vitro studies suggests that Pf155/RESA can be considered as a candidate for triggering TNF secretion. Thus we conducted experiments to investigate the relationship between Pf155/RESA and TNF production. The SGE1 strain of Plasmodium falciparum was compared with the P. falciparum FCR3 strain, which does not express Pf155/RESA protein, for ability to induce TNF secretion by normal human macrophages in vitro. Synthetic peptides from the Pf155/RESA antigen ((EENV)4, (EENVEHDA)4, (DDEHVEEPTVA)3), were used in some experiments. TNF levels were measured by an immunoradiometric assay. We observed that the RESA-defective strain induces lower levels of TNF after schizont rupture than the SGE1 strain. Moreover, substantial TNF secretion was detected when macrophages were incubated with all three peptides, maximum levels being obtained with the (EENV)4 peptide. Although previous reports have described TNF-inducing activity of phospholipid from P. falciparum, these findings strengthen the evidence for Pf155/RESA antigens also being involved in TNF production during malaria.

Amino Acid Sequence↗

PubMed1993

Parasite virulence factors during falciparum malaria: rosetting, cytoadherence, and modulation of cytoadherence by cytokines.

To determine virulence factors of isolates of Plasmodium falciparum and the potential role of cytokines in cerebral malaria, 46 Malagasy patients presenting with cerebral (n = 10), severe (n = 10), and uncomplicated (n = 26) malaria were enrolled in a study. The capacity of 21 of 46 P. falciparum isolates to form rosettes in vitro and to adhere to human umbilical vein endothelial cells (HUVECs) that express intercellular adhesion molecule-1 receptors and to C32 amelanotic melanoma cells that express mainly CD36 receptors was investigated together with the effects of tumor necrosis factor alpha (TNF-alpha), granulocyte macrophage-colony-stimulating factor (GM-CSF), interleukin-3 (IL-3), and IL-6 alone and in two-by-two combinations on the cytoadherence of infected erythrocytes to HUVECs. Plasma levels of these cytokines were also measured in the patients at admission. The percentage of rosette formation was higher for the isolates from patients with cerebral (n = 6; 19.5%) and severe (n = 6; 30.5%) malaria than for those from patients with uncomplicated malaria (n = 9; 5%) (P < 0.002). The cytoadherence properties of the isolates did not differ among the three groups whatever the target cell used, but adherence to melanoma cells was systematically higher than that to HUVECs. Adhesion to HUVECs was increased more after TNF-alpha stimulation than after GM-CSF, IL-3, or IL-6 stimulation (P < 0.01). Only the combination of TNF-alpha and IL-3 enhanced cytoadherence more than TNF-alpha used alone (P < 0.02). No difference in the modulation of cytoadherence by cytokines was found in relation to the severity of the disease. TNF-alpha and IL-6 levels in peripheral blood were higher in the patients with cerebral and severe malaria than in the patients with uncomplicated malaria (P < 0.005). Most of the patients' sera contained little or no IL-3 or GM-CSF. Our results challenge the role of intercellular adhesion molecule-1 as the principal receptor mediating the cytoadherence of P. falciparum-infected erythrocytes and contrast with data obtained in the murine model.

Adolescent↗

PubMed1993

Possible role of specific immunoglobulin M antibodies to Plasmodium falciparum antigens in immunoprotection of humans living in a hyperendemic area, Burkina Faso.

Two seroepidemiological studies were performed in an area of Burkina Faso hyperendemic for malaria to estimate the protective role of immunoglobulin M (IgM) antibodies. Six cross-sectional surveys were carried out on children (ages, < 16 years) in the village of Karankasso. The evolution of antibodies to crude extracts of Plasmodium falciparum (IgG or IgM antisomatic and IgG antiexoantigens) were tested by IFI or enzyme-linked immunosorbent assay and were followed up according to the fluctuations of the parasite densities. Specific IgG antibodies had the same evolution as parasite densities. By contrast, specific IgM antibodies increased when IgG and parasite densities began to decrease (despite a high inoculation rate). A longitudinal survey of 77 children and adults was conducted in another village (Dafinso). In that study, clinical follow-up of the selected individuals allowed us to define three groups in the population. Children in group 1 were considered nonimmune (children with one or more malaria attacks). Group 2 was composed of semiimmune children who did not present with any malarial attack during the survey but who had high levels of parasitemia during the transmission period. Group 3 was composed of immunoprotected adults. Specific IgM and IgG antibodies to crude extracts or a recombinant antigen (glutamate-rich protein) of P. falciparum were tested. Specific IgM antibodies were lower in group 1 (nonimmune) than in groups 2 (semiimmune) and 3 (immunoprotected). Furthermore, there was a negative correlation between parasite densities and the levels of specific IgM antibodies. We discuss the possible role of IgM antibodies in the acquisition of immunity to malaria.

Adolescent↗

PubMed1993

Chloroquine-induced inhibition of the production of TNF, but not of IL-6, is affected by disruption of iron metabolism.

There is now considerable evidence that cerebral malaria may be related to the over-production of tumour necrosis factor (TNF). Nevertheless, our knowledge is very poor concerning the biological events which lead up to this TNF over-production. Furthermore, interleukin-6 (IL-6) is produced in large amounts during malaria infection and seems to have inhibitory action on TNF production. Anti-malarial drugs were investigated for their ability to interfere with TNF and IL-6 secretion by human non-immune macrophages stimulated by lipopolysaccharides (LPS) or Plasmodium falciparum culture supernatant. Macrophages were pretreated with chloroquine, quinine, proguanil, mefloquine or halofantrine before stimulation. TNF and IL-6 production were suppressed in a dose-dependent manner when macrophages were treated with chloroquine, but not with other anti-malarial drugs. Considering that chloroquine probably acts via lysosomotropic mechanisms, and that iron metabolism may interfere with the non-specific immune response, we focused our attention on these biochemical events in order to investigate the mechanisms by which chloroquine inhibits cytokine production. Our results demonstrated that chloroquine-induced inhibition of TNF and IL-6 production is not mediated through a lysosomotropic mechanism, and that chloroquine probably acts on TNF secretion by disrupting iron homeostasis. Inhibition of IL-6 production seems not to be mediated through these pathways. These observations suggest that chloroquine may help to prevent cerebral malaria whatever the drug sensitivity of the parasite strain, and may provide new tools for an anti-disease therapy regardless of the emergence of parasite multi-drug resistance.

Antimalarials↗

PubMed1993

Pharmacokinetic and pharmacodynamic study of amodiaquine and its two metabolites after a single oral dose in human volunteers.

The pharmacokinetics of amodiaquine (AQ, Flavoquine, CAS 6398-98-7) and its metabolites, mono (AQml) and bis-desethyl amodiaquine (AQm2) were investigated in 8 healthy volunteers after an oral dose of 306.2 mg of AQ. Metabolic clearance was the main AQ elimination pathway. AQ disappeared rapidly, from the plasma and blood, whereas AQml appeared rapidly in keeping with a hepatic first-pass effect. By contrast, AQ was little excreted in urine and AQm2 formation from AQm1 was low. Blood AQm1 concentrations were higher than plasma levels, with an AQm1/AQ concentration ratio of 5 to 10. This result was related to strong uptake of AQm1 by white blood cells, as shown by an in vitro study. On the basis of plasma concentrations, there was no preferential uptake by red blood cells, the pharmacological target cells; effective AQ concentrations should thus be analyzed in plasma rather than in whole blood. The inhibitory activity of patients' sera on Plasmodium falciparum growth in vitro appears to be directly related to the AQm1 concentration.

Adult↗

PubMed1992

Monocyte tissue factor expression induced by Plasmodium falciparum-infected erythrocytes.

Monocytes are active elements of the host response against Plasmodium falciparum. They are able to express tissue factor and trigger the extrinsic pathway of blood coagulation the activation of which remained unclear in malaria. Our aim was to assess the tissue factor expression of purified blood monocytes stimulated by cultured Plasmodium falciparum-infected erythrocytes. Malaria parasite induced an early generation of tissue factor with a peak between 8 and 12 h of stimulation. Maximum expression was observed for parasitemia ranging from 1 to 2%. Plasmodium falciparum culture supernatants had the same effect showing the existence of a soluble factor able to induce the tissue factor expression. These data, demonstrating an activation of the tissue factor pathway by the malaria parasite, emphasize thrombin generation. Therefore, thrombin could participate in malaria pathology either in the microcirculatory blockade via platelet and fibrinogen activation or as a mitotic.

Animals↗

PubMed1992

Relationships between circulating S-antigens, naturally acquired antibodies to Plasmodium falciparum exoantigens and malaria attack in a mesoendemic area.

A survey involving 77 individuals living in two savannah villages near Bobo Dioulasso (Burkina Faso, West Africa), was performed in June 1987 (before), August-September (during) and January 1988 after the seasonal transmission. The clinical longitudinal study during the seasonal period permitted us to define three different groups in terms of both age and occurrence of malaria attack (MA; greater than or equal to 5000 parasites/mm3 of blood and axillary fever greater than or equal to 37.8 degrees C). The presence of circulating stable antigen (S-Ag) and the antibody responses against exoantigens (E-Ag) of Plasmodium falciparum were also evaluated at three observations periods: beginning, during and after the transmission season. The adult group (III) had the highest rates of IgG Ab to E-Ag although, IgM prevalence to E-Ag was maximal in the group II (individuals with no malaria attack and age less than or equal to 15 years old). Group I (persons with less than or equal to 15 years old and who contracted at least one MA) did not have any S-Ag at the first observation period and showed the lowest rate of antibodies to E-Ag. The probability of occurrence of an MA calculated from these parameters at the beginning of the transmission period were correct in 78.9% of the cases in children (Groups I & II) and in 71.8% of adults during the subsequent transmission period. Therefore these values could be used for evaluating the probability of occurrence of a clinical MA during the transmission period in a mesoendemic area. S-Ag and antibodies to E-Ag could participate positively in the mechanisms involved in the development of the immune status.

Adolescent↗