TCR signal strength controls thymic differentiation of discrete proinflammatory gamma delta T cell subsets

The mouse thymus produces discrete gd T cell subsets that make either interferon-g (IFN-g) or interleukin 17 (IL-17), but the role of the T cell antigen receptor (TCR) in this developmental process remains controversial. Here we show that Cd3g+/− Cd3d+/− (CD3 double-haploinsufficient (CD3DH)) mice have reduced TCR expression and signaling strength on gd T cells. CD3DH mice had normal numbers and phenotypes of ab thymocyte subsets, but impaired differentiation of fetal Vg6+ (but not Vg4+) IL-17- producing gd T cells and a marked depletion of IFN-g-producing CD122+ NK1.1+ gd T cells throughout ontogeny. Adult CD3DH mice showed reduced peripheral IFN-g+ gd T cells and were resistant to experimental cerebral malaria. Thus, TCR signal strength within specific thymic developmental windows is a major determinant of the generation of proinflammatory gd T cell subsets and their impact on pathophysiology

Longevity and Composition of Cellular Immune Responses Following Experimental Plasmodium falciparum Malaria Infection in Humans

Cellular responses to Plasmodium falciparum parasites, in particular interferon-gamma (IFNγ) production, play an important role in anti-malarial immunity. However, clinical immunity to malaria develops slowly amongst naturally exposed populations, the dynamics of cellular responses in relation to exposure are difficult to study and data about the persistence of such responses are controversial. Here we assess the longevity and composition of cellular immune responses following experimental malaria infection in human volunteers. We conducted a longitudinal study of cellular immunological responses to sporozoites (PfSpz) and asexual blood-stage (PfRBC) malaria parasites in naïve human volunteers undergoing single (n = 5) or multiple (n = 10) experimental P. falciparum infections under highly controlled conditions. IFNγ and interleukin-2 (IL-2) responses following in vitro re-stimulation were measured by flow-cytometry prior to, during and more than one year post infection. We show that cellular responses to both PfSpz and PfRBC are induced and remain almost undiminished up to 14 months after even a single malaria episode. Remarkably, not only ‘adaptive’ but also ‘innate’ lymphocyte subsets contribute to the increased IFNγ response, including αβT cells, γδT cells and NK cells. Furthermore, results from depletion and autologous recombination experiments of lymphocyte subsets suggest that immunological memory for PfRBC is carried within both the αβT cells and γδT compartments. Indeed, the majority of cytokine producing T lymphocytes express an CD45RO+ CD62L- effector memory (EM) phenotype both early and late post infection. Finally, we demonstrate that malaria infection induces and maintains polyfunctional (IFNγ+IL-2+) EM responses against both PfRBC and PfSpz, previously found to be associated with protection. These data demonstrate that cellular responses can be readily induced and are long-lived following infection with P. falciparum, with a persisting contribution by not only adaptive but also (semi-)innate lymphocyte subsets. The implications hereof are positive for malaria vaccine development, but focus attention on those factors potentially inhibiting such responses in the field

Opsonising antibodies to P. falciparum Merozoites associated with immunity to clinical malaria

Naturally acquired humoral immunity to the malarial parasite Plasmodium falciparum can protect against disease, although the precise mechanisms remain unclear. Although antibody levels can be measured by ELISA, few studies have investigated functional antibody assays in relation to clinical outcomes. In this study we applied a recently developed functional assay of antibody-mediated opsonisation of merozoites, to plasma samples from a longitudinal cohort study conducted in a malaria endemic region of Papua New Guinea (PNG). Phagocytic activity was quantified by flow cytometry using a standardized and high-throughput protocol, and was subsequently evaluated for association with protection from clinical malaria and high-density parasitemia. Opsonising antibody responses were found to: i) increase with age, ii) be enhanced by concurrent infection, and iii) correlate with protection from clinical episodes and high-density parasitemia. Stronger protective associations were observed in individuals with no detectable parasitemia at baseline. This study presents the first evidence for merozoite phagocytosis as a correlate of acquired immunity and clinical protection against P. falciparum malaria

Peripheral blood mononuclear cells derived from grand multigravidae display a distinct cytokine profile in response to P. falciparum infected erythrocytes

Immunopathology of placental malaria is most significant in women in their first pregnancy especially in endemic areas, due to a lack of protective immunity to Plasmodium falciparum, which is acquired in successive pregnancies. In some studies (but not all), grand multigravidae (defined as 5 or more pregnancies, G5–7) are more susceptible to poor birth outcomes associated with malaria compared to earlier gravidities. By comparing peripheral cellular responses in primigravidae (G1), women in their second to fourth pregnancy (G2–4) and grand multigravidae we sought to identify key components of the dysregulated immune response. PBMC were exposed to CS2-infected erythrocytes (IE) opsonised with autologous plasma or unopsonised IE, and cytokine and chemokine secretion was measured. Higher levels of opsonising antibody were present in plasma derived from multigravid compared to primigravid women. Significant differences in the levels of cytokines and chemokines secreted in response to IE were observed. Less IL-10, IL-1β, IL-6 and TNF but more CXCL8, CCL8, IFNγ and CXCL10 were detected in G5–7 compared to G2–4 women. Our study provides fresh insight into the modulation of peripheral blood cell function and effects on the balance between host protection and immunopathology during placental malaria infection

Haemopedia: an expression atlas of murine hematopoietic cells

Hematopoiesis is a multistage process involving the differentiation of stem and progenitor cells into distinct mature cell lineages. Here we present Haemopedia, an atlas of murine gene-expression data containing 54 hematopoietic cell types, covering all the mature lineages in hematopoiesis. We include rare cell populations such as eosinophils, mast cells, basophils, and megakaryocytes, and a broad collection of progenitor and stem cells. We show that lineage branching and maturation during hematopoiesis can be reconstructed using the expression patterns of small sets of genes. We also have identified genes with enriched expression in each of the mature blood cell lineages, many of which show conserved lineage-enriched expression in human hematopoiesis. We have created an online web portal called Haemosphere to make analyses of Haemopedia and other blood cell transcriptional datasets easier. This resource provides simple tools to interrogate gene-expression-based relationships between hematopoietic cell types and genes of interest

The contribution of Natural Killer Complex loci to the development of experimental cerebral malaria

Background: The Natural Killer Complex (NKC) is a genetic region of highly linked genes encoding several receptors involved in the control of NK cell function. The NKC is highly polymorphic and allelic variability of various NKC loci has been demonstrated in inbred mice, providing evidence for NKC haplotypes. Using BALB.B6-Cmv1(r) congenic mice, in which NKC genes from C57BL/6 mice were introduced into the BALB/c background, we have previously shown that the NKC is a genetic determinant of malarial pathogenesis. C57BL/6 alleles are associated with increased disease-susceptibility as BALB.B6-Cmv1(r) congenic mice had increased cerebral pathology and death rates during P. berghei ANKA infection than cerebral malaria-resistant BALB/c controls.\ud \ud Methods: To investigate which regions of the NKC are involved in susceptibility to experimental cerebral malaria (ECM), intra-NKC congenic mice generated by backcrossing recombinant F2 progeny from a (BALB/c x BALB.B6-Cmv1(r)) F1 intercross to BALB/c mice were infected with P. berghei ANKA.\ud \ud Results: Our results revealed that C57BL/6 alleles at two locations in the NKC contribute to the development of ECM. The increased severity to severe disease in intra-NKC congenic mice was not associated with higher parasite burdens but correlated with a significantly enhanced systemic IFN-gamma response to infection and an increased recruitment of CD8(+) T cells to the brain of infected animals.\ud \ud Conclusions: Polymorphisms within the NKC modulate malarial pathogenesis and acquired immune responses to infection

CD27 is a thymic determinant of the balance between interferon-γ- and interleukin 17–producing γδ T cell subsets

The production of cytokines such as interferon-gamma and interleukin 17 by alphabeta and gammadelta T cells influences the outcome of immune responses. Here we show that most gammadelta T lymphocytes expressed the tumor necrosis factor receptor family member CD27 and secreted interferon-gamma, whereas interleukin 17 production was restricted to CD27(-) gammadelta T cells. In contrast to the apparent plasticity of alphabeta T cells, the cytokine profiles of these distinct gammadelta T cell subsets were essentially stable, even during infection. These phenotypes were established during thymic development, when CD27 functions as a regulator of the differentiation of gammadelta T cells at least in part by inducing expression of the lymphotoxin-beta receptor and genes associated with trans-conditioning and interferon-gamma production. Thus, the cytokine profiles of peripheral gammadelta T cells are predetermined mainly by a mechanism involving CD2

Significant Association of KIR2DL3-HLA-C1 Combination with Cerebral Malaria and Implications for Co-evolution of KIR and HLA

Cerebral malaria is a major, life-threatening complication of Plasmodium falciparum malaria, and has very high mortality rate. In murine malaria models, natural killer (NK) cell responses have been shown to play a crucial role in the pathogenesis of cerebral malaria. To investigate the role of NK cells in the developmental process of human cerebral malaria, we conducted a case-control study examining genotypes for killer immunoglobulin-like receptors (KIR) and their human leukocyte antigen (HLA) class I ligands in 477 malaria patients. We found that the combination of KIR2DL3 and its cognate HLA-C1 ligand was significantly associated with the development of cerebral malaria when compared with non-cerebral malaria (odds ratio 3.14, 95% confidence interval 1.52–6.48, P = 0.00079, corrected P = 0.02). In contrast, no other KIR-HLA pairs showed a significant association with cerebral malaria, suggesting that the NK cell repertoire shaped by the KIR2DL3-HLA-C1 interaction shows certain functional responses that facilitate development of cerebral malaria. Furthermore, the frequency of the KIR2DL3-HLA-C1 combination was found to be significantly lower in malaria high-endemic populations. These results suggest that natural selection has reduced the frequency of the KIR2DL3-HLA-C1 combination in malaria high-endemic populations because of the propensity of interaction between KIR2DL3 and C1 to favor development of cerebral malaria. Our findings provide one possible explanation for KIR-HLA co-evolution driven by a microbial pathogen, and its effect on the global distribution of malaria, KIR and HLA