A Praziquantel Treatment Study of Immune and Transcriptome Profiles in Schistosoma haematobium-Infected Gabonese Schoolchildren.

BACKGROUND: Although Schistosoma haematobium infection has been reported to be associated with alterations in immune function, in particular immune hyporesponsiveness, there have been only few studies that have used the approach of removing infection by drug treatment to establish this and to understand the underlying molecular mechanisms. METHODS: Schistosoma haematobium-infected schoolchildren were studied before and after praziquantel treatment and compared with uninfected controls. Cellular responses were characterized by cytokine production and flow cytometry, and in a subset of children RNA sequencing (RNA-Seq) transcriptome profiling was performed. RESULTS: Removal of S haematobium infection resulted in increased schistosome-specific cytokine responses that were negatively associated with CD4+CD25+FOXP3+ T-cells and accompanied by increased frequency of effector memory T-cells. Innate responses to Toll like receptor (TLR) ligation decreased with treatment and showed positive association with CD4+CD25+FOXP3+ T-cells. At the transcriptome level, schistosome infection was associated with enrichment in cell adhesion, whereas parasite removal was associated with a more quiescent profile. Further analysis indicated that alteration in cellular energy metabolism was associated with S haematobium infection and that the early growth response genes 2 and 3 (EGR 2 and EGR3), transcription factors that negatively regulate T-cell activation, may play a role in adaptive immune hyporesponsiveness. CONCLUSIONS: Using a longitudinal study design, we found contrasting effects of schistosome infection on innate and adaptive immune responses. Whereas the innate immune system appears more activated, the adaptive immunity is in a hyporesponsive state reflected in alterations in CD4+CD25+FOXP3+ T-cells, cellular metabolism, and transcription factors involved in anergy

Cytokine Responses to <i>Schistosoma mansoni</i> and <i>Schistosoma haematobium</i> in Relation to Infection in a Co-endemic Focus in Northern Senegal

<div><p>Background</p><p>In Africa, many areas are co-endemic for the two major <i>Schistosoma</i> species, <i>S. mansoni</i> and <i>S. haematobium</i>. Epidemiological studies have suggested that host immunological factors may play an important role in co-endemic areas. As yet, little is known about differences in host immune responses and possible immunological interactions between <i>S. mansoni</i> and <i>S. haematobium</i> in humans. The aim of this study was to analyze host cytokine responses to antigens from either species in a population from a co-endemic focus, and relate these to <i>S. mansoni</i> and <i>S. haematobium</i> infection.</p><p>Methodology</p><p>Whole blood cytokine responses were investigated in a population in the north of Senegal (n = 200). Blood was stimulated for 72 h with schistosomal egg and adult worm antigens of either <i>Schistosoma</i> species. IL-10, IL-5, IFN-γ, TNF-α, and IL-2 production was determined in culture supernatants. A multivariate (i.e. multi-response) approach was used to allow a joint analysis of all cytokines in relation to <i>Schistosoma</i> infection.</p><p>Principal Findings</p><p><i>Schistosoma haematobium</i> egg and worm antigens induced higher cytokine production, suggesting that <i>S. haematobium</i> may be more immunogenic than <i>S. mansoni</i>. However, both infections were strongly associated with similar, modified Th2 cytokine profiles.</p><p>Conclusions/Significance</p><p>This study is the first to compare <i>S. mansoni</i> and <i>S. haematobium</i> cytokine responses in one population residing in a co-endemic area. These findings are in line with previous epidemiological studies that also suggested <i>S. haematobium</i> egg and worm stages to be more immunogenic than those of <i>S. mansoni</i>.</p></div

CD4⁺CD25^hiFOXP3⁺ Regulatory T Cells and Cytokine Responses in Human Schistosomiasis before and after Treatment with Praziquantel

<div><p>Background</p><p>Chronic schistosomiasis is associated with T cell hypo-responsiveness and immunoregulatory mechanisms, including induction of regulatory T cells (Tregs). However, little is known about Treg functional capacity during human <i>Schistosoma haematobium</i> infection.</p><p>Methodology</p><p>CD4⁺CD25^hiFOXP3⁺ cells were characterized by flow cytometry and their function assessed by analysing total and Treg-depleted PBMC responses to schistosomal adult worm antigen (AWA), soluable egg antigen (SEA) and Bacillus Calmette-Guérin (BCG) in <i>S</i>. <i>haematobium</i>-infected Gabonese children before and 6 weeks after anthelmintic treatment. Cytokines responses (IFN-γ, IL-5, IL-10, IL-13, IL-17 and TNF) were integrated using Principal Component Analysis (PCA). Proliferation was measured by CFSE.</p><p>Principal Findings</p><p><i>S</i>. <i>haematobium</i> infection was associated with increased Treg frequencies, which decreased post-treatment. Cytokine responses clustered into two principal components reflecting regulatory and Th2-polarized (PC1) and pro-inflammatory and Th1-polarized (PC2) cytokine responses; both components increased post-treatment. Treg depletion resulted in increased PC1 and PC2 at both time-points. Proliferation on the other hand, showed no significant difference from pre- to post-treatment. Treg depletion resulted mostly in increased proliferative responses at the pre-treatment time-point only.</p><p>Conclusions</p><p><i>Schistosoma</i>-associated CD4⁺CD25^hiFOXP3⁺Tregs exert a suppressive effect on both proliferation and cytokine production. Although Treg frequency decreases after praziquantel treatment, their suppressive capacity remains unaltered when considering cytokine production whereas their influence on proliferation weakens with treatment.</p></div

Levels of <i>Schistosoma</i>-induced cytokine responses in 72 h whole blood cultures (n = 200).

<p>Blood samples from one individual were divided into five and stimulated with <i>Schistosoma</i> antigens (SEAm, SEAh, AWAm, or AWAh), and with medium only (negative control; see <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0003080#s2" target="_blank">Materials and Methods</a>).</p>a<p>Crude cytokine levels are reported. IQR: Interquartile range (Tukey's hinges).</p>b<p>Wilcoxon Signed Rank test comparing <i>S. mansoni</i>- and <i>S. haematobium</i>-induced cytokine levels within individuals (either for SEA or AWA).</p>c<p><i>Schistosoma</i> egg antigen.</p>d<p>Adult worm antigen.</p

Variation in <i>Schistosoma</i> antigen-induced cytokine responses in relation to <i>Schistosoma</i> infection intensity.

<p>Each three-dimensional (3D) nMDS ordination is represented in two 2D planes (<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0003080#pntd.0003080.s001" target="_blank">Supporting Information S1</a>). Left and right panels represent the 1^st and 2^nd, and 2^nd and 3^rd dimensions, respectively. <b>Panels A</b> and <b>B</b> show the <i>S. mansoni</i> egg antigen (SEAm)-induced cytokine profile, <b>Panels C</b> and <b>D</b> that of <i>S. haematobium</i> SEA(h), <b>Panels E</b> and <b>F</b> that of <i>S. mansoni</i> adult worm antigens (AWAm), and <b>Panels G</b> and <b>H</b> show <i>S. haematobium</i> AWA(h)-induced cytokine profiles. Green dots represent individuals. Distances between dots approximate the rank order of dissimilarities in cytokine profiles between the respective individuals with stress values (i.e. discrepancies) of 0.051 for SEAm, 0.041 for SEAh, 0.058 for AWAm, and 0.061 for AWAh. Red arrows indicate linear gradients of normalized net cytokine responses on which the nMDS is based. Green dot sizes are proportional to individual values of normalized infection intensity of <i>S. mansoni</i> (for simplicity dots were only labelled with <i>S. mansoni</i> (not <i>S. haematobium</i>) infection intensity). Black arrows indicate linear gradients of post hoc fitted normalized infection intensity of <i>S. mansoni</i> (‘Sm’) and <i>S. haematobium</i> (‘Sh’). The length of the arrows is proportional to the goodness of fit onto the cytokine profile within one 2D plane, but lengths cannot be compared between cytokine and infection intensity arrows. Arrows are only depicted if their fit was significant at the level of <i>p</i> = 0.05 in 3D ordinations (see <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0003080#pntd-0003080-t004" target="_blank">Table 4</a>), as well as in the respective 2D planes. In Panel H, the arrows of IL-5 response and <i>S. mansoni</i> infection intensity are overlapping and their labels are therefore illegible. ^aThe biological a posteriori interpretation of nMDS1 (left x-axis) and nMDS2 (y-axis) were added between brackets on the axis labels, but nMDS3 (right x-axis) could not be interpreted.</p