Neonatal and maternal immunological responses to conserved epitopes within the DBL-gamma3 chondroitin sulfate A-binding domain of Plasmodium falciparum erythrocyte membrane protein 1

Abstract

Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) mediates the adherence of P. falciparum-infected erythrocytes to placental syncytiotrophoblasts via interactions with chondroitin sulfate A (CSA), a characteristic of pregnancy-associated malaria. Pregnancy-associated malaria predicts increased susceptibility of newborns to malaria, and it is postulated that transplacental passage of parasite antigen induces immune regulatory activity in the neonate. We wished to examine the immune responsiveness to a CSA-binding domain of PfEMP1, the DBL-gamma3 domain, in cord and maternal venous blood obtained from pregnancies with various histories of P. falciparum infection. We assessed in vitro T-cell cytokine and plasma immunoglobulin G (IgG) and IgM responses to four peptides corresponding to highly conserved regions of a DBL-gamma3 domain common to central African parasite isolates. The presence of placental P. falciparum infection at delivery was associated with elevated frequencies of DBL-gamma3 peptide-specific CD3+ interleukin-10-positive T cells in cord blood, while treatment and clearance of infection prior to delivery was associated with elevated frequencies of CD3+ gamma interferon-positive T cells. DBL-gamma3 peptide-specific IgM antibodies were detected in 12 of 60 (20%) cord plasma samples from those born to mothers with P. falciparum infection during pregnancy. Consistent with polyclonal anti-PfEMP1 antibody responses that are associated with protection against pregnancy-associated malaria, the presence of maternal IgG antibodies with specificity for one of the DBL-gamma3 peptides showed a parity-dependent profile. These data demonstrate that peptides corresponding to conserved regions of the DBL-gamma3 domain of PfEMP1 are immunogenic in P. falciparum-infected mothers and their offspring.

FIG. 1.

A) Amino acid sequences of peptides corresponding to conserved regions of DBL-γ3, and B) their SYFPEITHI score for HLA-DR phenotypes.

FIG. 2.

Intracellular cytokine activity of cord and maternal T cells following stimulation of cord blood mononuclear cells and peripheral blood mononuclear cells with DBL-γ3 peptides. A) Percentage of samples containing CD3⁺ cells with a cytokine response to DBL-γ3 peptides in cord and maternal blood. The mean net percent gated CD3⁺ cells in cord and maternal blood that produce IFN- γ, IL-13, or IL-10 in response to B) DBL-γ3 peptides or C) phytohemagglutinin (PHA) in groups segregated by maternal P. falciparum infection history during pregnancy. Bar graphs illustrate mean percentages with standard deviations. Sample sizes: European control, 7; Cords: negative, 19; placenta positive, 14; treated, 17; Mothers: negative, 8; placenta positive, 7; treated, 7. *, P < 0.05 by either Fisher exact test or the nonparametric Mann-Whitney test.

FIG. 3.

IgM and IgG antibodies with specificity for DBL-γ3 peptides are present in cord blood. A) The proportions of cord blood samples containing IgM antibodies with specificity for DBL-γ3 peptides, glutamate-rich protein (GLURP), or purified protein derivative (PPD) in groups segregated by maternal P. falciparum infection history during pregnancy. B) The proportions of cord plasma samples containing IgM with specificity for at least one DBL-γ3 peptide. C) Dot plot of DBL120 IgM antibody concentrations in plasma from malaria-unexposed Europeans (positivity threshold: mean + 3 standard deviations, shown as a dotted line), Gabonese mothers' peripheral blood and cord blood segregated by maternal P. falciparum infection history during pregnancy. D) Correlation between cord and maternal venous blood IgG antibody levels for all DBL-γ3 peptides. Sample sizes: negative, 25; placenta positive, 32; treated, 28; European controls, 12. *, P < 0.05 compared to negative group by Fisher exact test.

FIG. 4.

A) IgG and B) IgM antibodies with specificity for the DBL-γ3 peptides, glutamate-rich protein (GLURP), and purified protein derivative (PPD) in peripheral blood plasma from mothers at the time of delivery. Samples are segregated by P. falciparum infection history during pregnancy. Box-whisker plots show medians with 25th and 75th and whiskers for 10th and 90th percentiles. Sample sizes: negative, 25; placenta positive, 32; treated, 28; European controls, 12. *, P < 0.05 compared to negative group, Mann-Whitney test.

FIG. 5.

Relationship between parity and maternal A) IgG and B) IgM antibody levels with specificity for DBL-γ peptides, glutamate-rich protein (GLURP), or purified protein derivative (PPD). *, P < 0.05. Spearman rank correlation.