Of the 119 articles that met the search criteria, 113 were excluded on the basis of title and abstract (Fig. 3). An additional 47 papers, identified from the references of the 6 original papers, contributed to this review.
Leukocytes and neutrophils variations during pregnancy
The total white blood cell (WBC) count was found to be higher in pregnant women than in non-pregnant women due to an increase in neutrophils [13]. The number of neutrophils increases at the time of the oestrogen peak of a normal menstrual cycle and, if fertilization has occurred, neutrophils continue to increase [14]. Subsequently, there is an increase in neutrophils from day 45 of pregnancy, reaching a plateau during the second and third trimesters [13]. Breastfeeding may prolong the rise in neutrophil counts (reviewed by [15]). Increases in neutrophil count and metabolic activity are apparently the result of oestrogen stimulation [16–18].
Neutrophils and cytokine variations in PAM
In response to stress, such as inflammation or infection, neutrophils are released into the blood circulation. Normally, neutrophils are released in large numbers, more than any other leukocytes. In case of PAM, recently, Okezie Caleb Okamgba et al showed that the number and type of WBC lineages that may be preferentially increased against the pathogenic malaria parasite in peripheral and placental blood [19]. Comparison of the mean of the neutrophil counts between malaria-infected peripheral and placental blood, showed that the number of neutrophils is higher in placental blood [19].
Bostrom S et al. showed that the number of neutrophils in the peripheral blood stream was reduced in pregnant women with P. falciparum infection compared to pregnant women without malaria infection (Fig. 4A) [20]. In contrast, other studies reported an increase in neutrophil counts in African children with uncomplicated malaria [21, 22]. In contrast to African children, a significant reduction of neutrophil counts in Thailand adults with P. falciparum malaria was described [23]. This difference in immune response may reflect different inflammatory responses and differences in neutrophils functions. Age difference and hormonal status could also play a role in immune response.
Although undetectable in the peripheral blood, the malaria parasite can be present in the placenta and induce the immune response in the peripheral blood. It has been investigated that the number of maternal peripheral granulocytes decreases with placental malaria (PM) in the presence and absence of pre-existing human immunodeficiency virus (HIV) infection [24]. Even though the study did not perform a differential analysis targeting the number of neutrophils among granulocytes, these results are consistent with previous publications that have reported reduced numbers of neutrophils in malaria-infected pregnant women compared with uninfected women [20, 25]. This reduction in the number of neutrophils in the peripheral blood may be explained by their accumulation in the placenta infected with malaria parasites. Other publications confirmed an important accumulation of neutrophils in the placenta during PM [19, 26].
The decreased neutrophil counts observed in peripheral blood might partially reflect the accumulation of neutrophils in the infected placenta as part of immune response. Neutrophils, eosinophils, basophils and mast cells have in fact often been observed in histological studies of placental biopsies from pregnant women with malaria infection [27, 28].
Moreover, cytokine levels such as interferon gamma (IFN-γ), tumor necrosis factor alpha (TNF-α), interleukin-4 (IL-4), IL-6, and IL-10 were elevated in pregnant women with malaria compared to those uninfected by malaria parasites [15]. Other studies reported that IFN-γ and IL-6 were significantly higher in malaria-infected pregnant women than in non-pregnant uninfected women [16, 17]. Additional finding (Torre et al., 2002) [31], showed that a significantly higher levels of IFN-γ, TNF-α, and IL-10 were found in malaria-infected pregnant women than in their control counterparts. Data from East Africa, published by Bayoumi et al [32] and Boström et al [33], show a significant increase in IL-10 in malaria infected pregnant women compared to uninfected pregnant women. However, data from Nigeria [19] described that IFN-γ, IL-4 and IL-10 were higher in peripheral blood not infected with malaria parasites than in infected peripheral blood. The authors believe that this may be due to regional differences, as Nigeria is a region where malaria is very prevalent. From these findings, it can be speculated that the intensity of malaria prevalence can modulate cytokine levels.
In addition to in vivo studies, in vitro models have also been used to observe the activation of immunological parameters including neutrophils. In vitro studies showed that stimulation of human placental cells line that originates from a choriocarcinoma (BeWo cells) [34] with malaria infected red blood cells (iRBCs) resulted in a pronounced release of interleukin 8 (IL-8) and strongly induced neutrophil migration in a transwell assay. This is in the same line with other co-culture studies where iRBCs incubated with whole blood [35] or brain endothelial [36] cells also led to IL-8 production. In addition to this, IL-8 as well as macrophage inflammatory protein-1 alpha (MIP-1α) were elevated in placenta plasma of pregnant women with malaria [12]. This suggests that the PAM associated with choriocarcinoma increases further neutrophil activities.
Neutrophil levels in the peripheral bloodstream and poor birth outcome during PAM
It has been reported based on data collected in Papua New Guinea that women in the second or third trimester of pregnancy with increased numbers in pigmented neutrophils gave birth to babies with lower weights (Fig. 4B) [37]. Furthermore, the prevalence of children with low birth weight (LBW) was higher in pregnant women with high number of pigmented peripheral neutrophils than in PAM women with low numbers [37]. When the women were grouped into primigravidae, secundigravidae and multigravidae, the study found no significant difference in either birth weight or pigmented neutrophil count among the different groups [37]. The role of neutrophils in the pathogenesis of PAM needs to be further explored (Fig. 1 and Fig. 2), but the available data are consistent with the role of neutrophil activation in the pathogenesis of idiopathic intrauterine growth restriction and preeclampsia [38, 39]. Evidence suggests that abnormally pigmented neutrophils clearance in malaria-infected women may be delayed in the peripheral circulation, leading to chronic inflammation and thus impacting fetal growth [40]. Furthermore, pigmented neutrophil counts were positively correlated with levels of TNF-α [41], a pro-inflammatory cytokine steady associated with LBW [4, 42]. Similarly, sequestration of maternal monocytes in placental intervillous blood has been linked to LBW in placental malaria [5, 43]. It could be that their excessive cytokine production impairs placental nutrient transport [44] and placental trophoblast invasion [45], thereby contributing to poor fetal growth.
Circulating pigmented neutrophils counts can be used to predict malaria fatality with more accuracy compared to peripheral parasitemia [46] but remains speculative regarding poor delivery outcome.
Neutrophil activation markers and malaria or malaria and HIV coinfection in pregnant women
Neutrophil activation results in massive production of its markers such as myeloperoxidase (MPO), matrix metalloproteinase (MMP9), neutrophil elastase (NE) and proteinase 3 (PRTN3). MPO is involved in the oxidative burst of neutrophils and macrophages. Interestingly, the placental blood MPO and PRTN3 levels are increased with PM in both HIV negative and HIV positive pauci-gravidae women relative to PM-HIV negative women [24]. In addition, MPO was found in placental tissue sections, suggesting that this factor may be produced by both neutrophils and intervillous macrophages. However, most MPO-expressing cells observed by immunofluorescence also stain for NE, identifying them as neutrophils. Furthermore, it has been found that placental levels of MPO, MMP9, and PRTN3 correlated positively with placental malaria parasite density [24]. This further confirms that MPO is derived from neutrophils whose activation is proportional to the parasite density. MPO levels in placental blood were significantly increased in chronic, inflammatory PM in primigravida. MPO is associated with vascular dysfunction [47], which underlies the pathophysiology of many vascular inflammatory diseases, including arteriosclerosis and coronary artery disease [48, 49]. High neutrophil activation, proved by higher plasma concentrations of not only MPO but also PRTN3 and NE, is correlated with severe pediatric malaria [50]. Relatedly, MMP9, an endopeptidase discharged by neutrophils and monocytes, is incriminated in the pathogenesis of severe malaria [51–53]. A polymorphism in MMP9 protects against PM, implying an important role for this enzyme in P. falciparum infection [54]. High concentrations of neutrophil-derived antimicrobial compounds can render these cells harmful to the host [55–57]. Neutrophils can rapidly be recruited to sites of infection and tissue injury [8], generate ROS which can exacerbate preeclampsia [58–61].
These findings suggest that malaria during pregnancy associated with HIV infection exacerbates neutrophil activation.
Neutrophil and Plasmodium vivax exflagellation in pregnant woman first description
Soo In Choi et al [62] reported a case of exflagellated microgametes in neutrophils in Pakistani pregnant woman. She was presenting a fever > 40°C lasting for 7 days with P. vivax mono-specific infection detected by rapid diagnostic tests and confirmed by nested PCR. In her blood, filamentous microgametes of 15–20 µm surrounding round to oval chromatin structures were detected. Microgametes were spread outside the RBCs, whereas others looked to be exflagellated from the gametocyte. Some microgametes were observed in the cytoplasm of neutrophils. Neutrophils containing microgametes have phagosomes in the cytoplasm and often nuclear condensation and ruffled plasma membranes (Fig. 5). The patient ended up having an abortion four days later and a P. vivax malaria relapse one month after the initial diagnosis.
It has been previously reported by MacCallum in 1897 [2] that the presence of exflagellated microgametes in blood is rare and involve only RBC. According to Soo In Choi et al [62], they made the first report of the involvement of Plasmodium exflagellates neutrophils and that further studies are needed to determine if neutrophils can be infected by Plasmodium and its drive from one site to another during its migration.