Characterization of VAR2CSA-deficient Plasmodium falciparum-infected erythrocytes selected for adhesion to the BeWo placental cell line

Background. Malaria in pregnancy is characterized by accumulation of infected erythrocytes (IE) in the placenta. The key ligand identified as mediating this process is a Plasmodium falciparum erythrocyte membrane protein 1 family member, termed VAR2CSA. VAR2CSA appears to be the main ligand responsible for adhesion to chondroitin sulphate A (CSA). Whether other PfEMP1 molecules can also mediate placental adhesion, independent of CSA binding, is unclear. Methods. The parasite line CS2 carrying a disrupted var2csa gene (CS2KO) was selected for adhesion to the BeWo choriocarcinoma cell line, which has been proposed as a model for placental malaria. The selected and control IE were tested for adhesion to placental sections and flow cytometry was used to measure recognition of IE by three serum sets from malaria-exposed men and women. Results. Wild-type CS2 adhere to BeWo and placental tissue via CSA. CS2KO IE were successfully selected for adhesion to BeWo, and adhered by a CSA-independent mechanism. They bound to immobilized ICAM-1 and CD36. BeWo-selected CS2KO bound at moderate levels to placental sections, but most binding was to placental villi rather than to the syncytiotrophoblast to which IE adherence occurs in vivo. This binding was inhibited by a blocking antibody to CD36 but not to ICAM-1. As expected, sera from malaria-exposed adults recognized CS2 IE in a gender and parity dependent manner. In one serum set, there was a similar but less pronounced pattern of antibody binding to selected CS2KO IE, but this was not seen in two others. One var gene, It4var19, was particularly abundant in the selected line and was detected as full length transcripts in BeWo-selected IE, but not unselected CS2KO. Conclusion. This study suggests that IE with characteristics similar to the CS2KO have a limited role in the pathogenesis of placental malaria. VAR2CSA appear to be the major ligand for placental adhesion, and could be the basis for a vaccine against pregnancy malaria

The relationship of Plasmodium falciparum humeral immunity with HIV-1 immunosuppression and treatment efficacy in Zambia

<p>Abstract</p> <p>Background</p> <p>HIV-1 infection affects malaria humeral immunity during pregnancy, but data for non-pregnant adults are lacking. This study reports the impact of HIV-1 infection and other variables on the level of malaria humeral immunity in adults with clinical malaria and whether humeral immune suppression was a risk factor for treatment failure.</p> <p>Methods</p> <p>Sera of 224 HIV-1 infected and 115 uninfected adults were compared for IgG to merozoite antigens AMA-1 and MSP2 (3D7 and FC27 types) determined by ELISA, and for IgG to the Variant Surface Antigens (VSA) of three different parasite line E8B, A4 and HCD6 determined by flow cytometry.</p> <p>Results</p> <p>Compared to HIV-1 uninfected adults, AMA-1 IgG was lower in HIV-1 infected (<it>P </it>= 0.02) and associated with low CD4 count AMA-1 IgG (<it>P </it>= 0.003). Low IgG to all three merozoite antigens was associated with less anemia (<it>P </it>= 0.03). High parasite load was associated with low MSP2 IgG 3D7 and FC27 types (<it>P </it>= 0.02 and <it>P </it>= 0.08). Antibody levels to VSA did not differ between HIV-1 infected and uninfected adults. However, low VSA IgGs were associated with high parasite load (<it>P </it>≤ 0.002 for each parasite line) and with treatment failure (<it>P </it>≤ 0.04 for each parasite line).</p> <p>Conclusion</p> <p>HIV-1 affects humeral responses to AMA-1, but seems to marginally or not affect humeral responses to other merozoite antigens and VSAs. The latter were important for controlling parasite density and predict treatment outcome.</p

Antibodies to Variant Surface Antigens of Plasmodium falciparum –Infected Erythrocytes Are Associated with Protection from Treatment Failure and the Development of Anemia in Pregnancy

In pregnancy associated malaria (PAM), Plasmodium falciparum infected erythrocytes (IEs) express variant surface antigens (VSA-PAM) that evade existing immunity and mediate placental sequestration. Antibodies to VSA-PAM develop with gravidity and block placental adhesion or opsonise IEs for phagocytic clearance, protecting women from anemia and low birth weigh

Prevention of inflammatory activation of human gestational membranes in an ex vivo model using a pharmacological NF-κB inhibitor

Intrauterine inflammation plays a major role in the etiology of preterm labor and birth. We established an ex vivo model employing perfused full-thickness term gestational membranes to study membrane transport, function, and inflammatory responses. Exposure of the maternal (decidual) face of the membranes to LPS (5 μg/ml) resulted in increased accumulation of proinflammatory cytokines in the maternal compartment within 4 h, followed by a response in the fetal (amniotic) compartment. Using cytokine arrays, exposure to LPS was found to result in increased secretion of a large number of cytokines and chemokines in both compartments, most notably IL-5, IL-6, IL-7, MDC (macrophage-derived chemokine), MIG (monokine induced by IFN-γ), TARC (thymus and activation-regulated chemokine), TGF-β, and TNF-α. PGE2 accumulation also increased in response to LPS, particularly in the fetal compartment. Cotreatment with sulfasalazine, which inhibited nuclear translocation of NF-κB p65, had a rapid and marked inhibitory effect on the rate of cytokine accumulation in the maternal compartment, with lesser but significant effects observed in the fetal compartment. While membrane integrity was not discernibly impaired with LPS or sulfasalazine exposure, rates of chorionic apoptosis after 20 h were doubled in sulfasalazine-treated tissues. We conclude that the system described provides a means of accurately modeling human gestational membrane functions and inflammatory activation ex vivo. Decidual LPS exposure was shown to elicit a robust inflammatory response in both the maternal and fetal compartments. Sulfasalazine was an effective antiinflammatory agent in this model, but also exerted proapoptotic effects that raise concerns regarding its placental effects when administered in pregnancy

Serum Lipoproteins Promote Efficient Presentation of the Malaria Virulence Protein PfEMP1 at the Erythrocyte Surface▿

The virulence of the malaria parasite Plasmodium falciparum is related to its ability to express a family of adhesive proteins known as P. falciparum erythrocyte membrane protein 1 (PfEMP1) at the infected red blood cell surface. The mechanism for the transport and delivery of these adhesins to the erythrocyte membrane is only poorly understood. In this work, we have used specific immune reagents in a flow cytometric assay to monitor the effects of serum components on the surface presentation of PfEMP1. We show that efficient presentation of the A4 and VAR2CSA variants of PfEMP1 is dependent on the presence of serum in the bathing medium during parasite maturation. Lipid-loaded albumin supports parasite growth but allows much less efficient presentation of PfEMP1 at the red blood cell surface. Analysis of the serum components reveals that lipoproteins, especially those of the low-density lipoprotein fraction, promote PfEMP1 presentation. Cytoadhesion of infected erythrocytes to the host cell receptors CD36 and ICAM-1 is also decreased in infected erythrocytes cultured in the absence of serum. The defect appears to be in the transfer of PfEMP1 from parasite-derived structures known as the Maurer's clefts to the erythrocyte membrane or in surface conformation rather than a down-regulation or switching of particular PfEMP1 variants

Relationship between Human Immunodeficiency Virus Type 1 Coinfection, Anemia, and Levels and Function of Antibodies to Variant Surface Antigens in Pregnancy-Associated Malaria ▿

Human immunodeficiency virus type 1 (HIV-1) coinfection decreases antibodies to variant surface antigens implicated in pregnancy-associated malaria (VSA-PAM) caused by Plasmodium falciparum. The effect of HIV-1 on antibody functions that may protect mothers from pregnancy-associated malaria is unknown. Sera from multigravid pregnant women with malaria and HIV-1 coinfection (n = 58) or malaria alone (n = 29) and from HIV-1-infected (n = 102) or -uninfected (n = 54) multigravidae without malaria were analyzed for anti-VSA-PAM antibodies by flow cytometry, the ability to inhibit adhesion to chondroitin sulfate A, or to opsonize CS2-infected erythrocytes for phagocytosis by THP-1 cells. In women with malaria, anti-VSA-PAM levels correlated better with opsonic activity (r = 0.60) than with adhesion-blocking activity (r = 0.33). In univariate analysis, HIV-1 coinfection was associated with lower opsonic activity but not adhesion-blocking activity or anti-VSA-PAM levels. Malaria-infected women with anemia (hemoglobin levels of <11.0 g/dl) had lower opsonic activity than nonanemic women (P = 0.007) independent of HIV-1 status. By multivariate analysis, in malaria-infected women, anemia (but not HIV status) was associated with opsonic activity. In women without malaria, opsonic activity was not associated with either anemia or HIV-1 status. In multigravid pregnant women with malaria, impaired serum opsonic activity may contribute to anemia and possibly to the decreased immunity to pregnancy-associated malaria associated with HIV-1

Adhesion of CS2KO IE to BeWo cells following repeated cycles of selection

Adhesion is expressed as IE bound per 100 BeWo nuclei, in the presence (black bars) or absence (white bars) of CSA 10 μg/ml. Data shown are the mean + SEM of at least two independent experiments, performed in triplicate.<p><b>Copyright information:</b></p><p>Taken from "Characterization of VAR2CSA-deficient -infected erythrocytes selected for adhesion to the BeWo placental cell line"</p><p>http://www.malariajournal.com/content/7/1/51</p><p>Malaria Journal 2008;7():51-51.</p><p>Published online 26 Mar 2008</p><p>PMCID:PMC2329659.</p><p></p