Peripheral Blood Cell Signatures of Plasmodium falciparum Infection during Pregnancy

Abstract

Sequestration of Plasmodium falciparum-infected erythrocytes in placental intervillous spaces causes inflammation and pathology. Knowledge of the profiles of immune cells associated with the physiopathology of pregnancy-associated malaria (PAM) is scarce. We conducted a longitudinal, prospective study, both in Benin and Tanzania, including ∼1000 pregnant women in each site with systematic follow-up at scheduled antenatal visits until delivery. We used ex vivo flow cytometry to identify peripheral blood mononuclear cell (PBMC) profiles that are associated with PAM and anaemia, determining the phenotypic composition and activation status of PBMC in selected sub-groups with and without PAM both at inclusion and at delivery in a total of 302 women. Both at inclusion and at delivery PAM was associated with significantly increased frequencies both of B cells overall and of activated B cells. Infection-related profiles were otherwise quite distinct at the two different time-points. At inclusion, PAM was associated with anaemia, with an increased frequency of immature monocytes and with a decreased frequency of regulatory T cells (Treg). At delivery, infected women presented with significantly fewer plasmacytoid dendritic cells (DC), more myeloid DC expressing low levels of HLA-DR, and more effector T cells (Teff) compared to uninfected women. Independent associations with an increased risk of anaemia were found for altered antigen-presenting cell frequencies at inclusion, but for an increased frequency of Teff at delivery. Our findings emphasize the prominent role played by B cells during PAM whenever it arises during pregnancy, whilst also revealing signature changes in other circulating cell types that, we conclude, primarily reflect the relative duration of the infections. Thus, the acute, recently-acquired infections present at delivery were marked by changes in DC and Teff frequencies, contrasting with infections at inclusion, considered chronic in nature, that were characterized by an abundance of immature monocytes and a paucity of Treg in PBMC.

Submicroscopic Infections with Plasmodium falciparum during Pregnancy and Their Association with Circulating Cytokine, Chemokine, and Cellular Profiles

“…After giving written informed consent, ϳ1,000 pregnant women at gestational ages of Յ24 weeks were included in the district of Comé, located in the Mono province 70 km west of Cotonou, the economic capital of Benin. The STOPPAM study design in Benin has been described in detail elsewhere (25), as has the procedure used to select women for the immunological substudy described here (18). First, at inclusion in the study, we selected a subgroup of 131 women who were identified as being infected with P. falciparum (for methods, see below) and an uninfected control group appropriately matched for age, gravidity, and gestational age.…”

“…Venous blood samples, collected in Vacutainer tubes with citrate phosphate dextrose adenine (CPDA) anticoagulant, were transported to the research laboratories within 4 h. Plasma samples were separated and stored at Ϫ80°C. Peripheral blood mononuclear cells (PBMCs) were isolated using Leucosep tubes (Greiner-Bio), according to the manufacturer's instructions, and were subsequently used for immunophenotyping as described previously (18).…”

“…PBMCs were washed and resuspended at a concentration of 10 million cells/ml. Cells were then labeled using a panel of surface and intracellular markerspecific monoclonal antibodies, as described elsewhere (18). Human antiblood dendritic cell antigen 1 (BDCA-1)-phycoerythrin (PE) was used for myeloid dendritic cell (mDC) detection and anti-BDCA-2-PE for plasmacytoid DC (pDC) detection (all from Miltenyi Biotec).…”

“…We recently showed that P. falciparum infection during pregnancy is associated with modified peripheral blood mononuclear cell (PBMC) profiles, consistently characterized by enhanced B cell activity (18), a finding consistent with the prominent B cell component of the inflammatory response to PfiEs in the placenta reported by others (19). B cells are recognized as having multiple functions, not only as antigen-presenting cells and antibody producers but also as regulators of dendritic cell (DC) function and maturation through their production of the cytokine interleukin 10 (IL-10) (20,21).…”

Pregnancy and Malaria Exposure Are Associated with Changes in the B Cell Pool and in Plasma Eotaxin Levels

“…Although multiple studies have analyzed Ab responses to malaria parasites during pregnancy, only two recent studies have analyzed B cells in malaria during pregnancy. The first one reported an increase in B cells and activated B cells in this condition (15); the second one showed altered frequencies of B cell subsets (16). However, the phenotypic resolution of these studies was very limited.…”