Retention of uninfected red blood cells causing congestive splenomegaly is the major mechanism of anemia in malaria.

Splenomegaly frequently occurs in patients with Plasmodium falciparum (Pf) or P. vivax (Pv) malarial anemia, but mechanisms underlying this co-occurrence are unclear. In malaria-endemic Papua, Indonesia, we prospectively analyzed red blood cell (RBC) concentrations in the spleen and spleen-mimetic retention in 37 subjects splenectomized for trauma or hyperreactive splenomegaly, most of whom were infected with Plasmodium. Splenomegaly (median 357 g [range: 80-1918 g]) was correlated positively with the proportion of red-pulp on histological sections (median 88.1% [range: 74%-99.4%]; r = .59, p = .0003) and correlated negatively with the proportion of white-pulp (median 8.3% [range: 0.4%-22.9%]; r = -.50, p = .002). The number of RBC per microscopic field (>95% uninfected) was correlated positively with spleen weight in both Pf-infected (r = .73; p = .017) and Pv-infected spleens (r = .94; p = .006). The median estimated proportion of total-body RBCs retained in Pf-infected spleens was 8.2% (range: 1.0%-33.6%), significantly higher than in Pv-infected (2.6% [range: 0.6%-23.8%]; p = .015) and PCR-negative subjects (2.5% [range: 1.0%-3.3%]; p = .006). Retained RBCs accounted for over half of circulating RBC loss seen in Pf infections. The proportion of total-body RBC retained in Pf- and Pv-infected spleens correlated negatively with hemoglobin concentrations (r = -.56, p = .0003), hematocrit (r = -.58, p = .0002), and circulating RBC counts (r = -.56, p = .0003). Splenic CD71-positive reticulocyte concentrations correlated with spleen weight in Pf (r = 1.0; p = .003). Retention rates of peripheral and splenic RBCs were correlated negatively with circulating RBC counts (r = -.69, p = .07 and r = -.83, p = .008, respectively). In conclusion, retention of mostly uninfected RBC in the spleen, leading to marked congestion of the red-pulp, was associated with splenomegaly and is the major mechanism of anemia in subjects infected with Plasmodium, particularly Pf.

Extracellular traps are evident in Romanowsky-stained smears of bronchoalveolar lavage from children with non-cystic fibrosis bronchiectasis.

BACKGROUND AND OBJECTIVE: The importance of extracellular traps (ETs) in chronic respiratory conditions is increasingly recognized but their role in paediatric bronchiectasis is poorly understood. The specialized techniques currently required to study ETs preclude routine clinical use. A simple and cost-effective ETs detection method is needed to support diagnostic applications. We aimed to determine whether ETs could be detected using light microscopy-based assessment of Romanowsky-stained bronchoalveolar lavage (BAL) slides from children with bronchiectasis, and whether the ETs cellular origin could be determined. METHODS: Archived Romanowsky-stained BAL slides from a cross-sectional study of children with bronchiectasis were examined for ETs using light microscopy. The cellular origin of individual ETs was determined based on morphology and physical contact with surrounding cell(s). RESULTS: ETs were observed in 78.7% (70/89) of BAL slides with neutrophil (NETs), macrophage (METs), eosinophil (EETs) and lymphocyte (LETs) ETs observed in 32.6%, 51.7%, 4.5% and 9%, respectively. ETs of indeterminate cellular origin were present in 59.6% of slides. Identifiable and indeterminate ETs were co-detected in 43.8% of slides. CONCLUSION: BAL from children with bronchiectasis commonly contains multiple ET types that are detectable using Romanowsky-stained slides. While specialist techniques remain necessary to determining the cellular origin of all ETs, screening of Romanowsky-stained slides presents a cost-effective method that is well-suited to diagnostic settings. Our findings support further research to determine whether ETs can be used to define respiratory endotypes and to understand whether ETs-specific therapies may be required to resolve airway inflammation among children with bronchiectasis.

Increased circulating myeloid-derived suppressor cells in vivax malaria and severe falciparum malaria.

BACKGROUND: Circulating myeloid-derived-suppressor-cells (MDSC) with immunosuppressive function are increased in human experimental Plasmodium falciparum infection, but have not been studied in clinical malaria. METHODS: Using flow-cytometry, circulating polymorphonuclear-MDSC were evaluated in cryopreserved samples from patients with uncomplicated Plasmodium vivax (n = 8) and uncomplicated (n = 4) and severe (n = 16) falciparum malaria from Papua, Indonesia. RESULTS: The absolute number of circulating polymorphonuclear-MDSC were significantly elevated in severe falciparum malaria patients compared to controls (n = 10). Polymorphonuclear-MDSC levels in uncomplicated vivax malaria were also elevated to levels comparable to that seen in severe falciparum malaria. CONCLUSION: Control of expansion of immunosuppressive MDSC may be important for development of effective immune responses in falciparum and vivax malaria.

Evaluation of splenic accumulation and colocalization of immature reticulocytes and Plasmodium vivax in asymptomatic malaria: A prospective human splenectomy study.

BACKGROUND: A very large biomass of intact asexual-stage malaria parasites accumulates in the spleen of asymptomatic human individuals infected with Plasmodium vivax. The mechanisms underlying this intense tropism are not clear. We hypothesised that immature reticulocytes, in which P. vivax develops, may display high densities in the spleen, thereby providing a niche for parasite survival. METHODS AND FINDINGS: We examined spleen tissue in 22 mostly untreated individuals naturally exposed to P. vivax and Plasmodium falciparum undergoing splenectomy for any clinical indication in malaria-endemic Papua, Indonesia (2015 to 2017). Infection, parasite and immature reticulocyte density, and splenic distribution were analysed by optical microscopy, flow cytometry, and molecular assays. Nine non-endemic control spleens from individuals undergoing spleno-pancreatectomy in France (2017 to 2020) were also examined for reticulocyte densities. There were no exclusion criteria or sample size considerations in both patient cohorts for this demanding approach. In Indonesia, 95.5% (21/22) of splenectomy patients had asymptomatic splenic Plasmodium infection (7 P. vivax, 13 P. falciparum, and 1 mixed infection). Significant splenic accumulation of immature CD71 intermediate- and high-expressing reticulocytes was seen, with concentrations 11 times greater than in peripheral blood. Accordingly, in France, reticulocyte concentrations in the splenic effluent were higher than in peripheral blood. Greater rigidity of reticulocytes in splenic than in peripheral blood, and their higher densities in splenic cords both suggest a mechanical retention process. Asexual-stage P. vivax-infected erythrocytes of all developmental stages accumulated in the spleen, with non-phagocytosed parasite densities 3,590 times (IQR: 2,600 to 4,130) higher than in circulating blood, and median total splenic parasite loads 81 (IQR: 14 to 205) times greater, accounting for 98.7% (IQR: 95.1% to 98.9%) of the estimated total-body P. vivax biomass. More reticulocytes were in contact with sinus lumen endothelial cells in P. vivax- than in P. falciparum-infected spleens. Histological analyses revealed 96% of P. vivax rings/trophozoites and 46% of schizonts colocalised with 92% of immature reticulocytes in the cords and sinus lumens of the red pulp. Larger splenic cohort studies and similar investigations in untreated symptomatic malaria are warranted. CONCLUSIONS: Immature CD71+ reticulocytes and splenic P. vivax-infected erythrocytes of all asexual stages accumulate in the same splenic compartments, suggesting the existence of a cryptic endosplenic lifecycle in chronic P. vivax infection. Findings provide insight into P. vivax-specific adaptions that have evolved to maximise survival and replication in the spleen.

Reduced circulating dendritic cells in acute Plasmodium knowlesi and Plasmodium falciparum malaria despite elevated plasma Flt3 ligand levels.

BACKGROUND: Plasmodium falciparum malaria increases plasma levels of the cytokine Fms-like tyrosine kinase 3 ligand (Flt3L), a haematopoietic factor associated with dendritic cell (DC) expansion. It is unknown if the zoonotic parasite Plasmodium knowlesi impacts Flt3L or DC in human malaria. This study investigated circulating DC and Flt3L associations in adult malaria and in submicroscopic experimental infection. METHODS: Plasma Flt3L concentration and blood CD141+ DC, CD1c+ DC and plasmacytoid DC (pDC) numbers were assessed in (i) volunteers experimentally infected with P. falciparum and in Malaysian patients with uncomplicated (ii) P. falciparum or (iii) P. knowlesi malaria. RESULTS: Plasmodium knowlesi caused a decline in all circulating DC subsets in adults with malaria. Plasma Flt3L was elevated in acute P. falciparum and P. knowlesi malaria with no increase in a subclinical experimental infection. Circulating CD141+ DCs, CD1c+ DCs and pDCs declined in all adults tested, for the first time extending the finding of DC subset decline in acute malaria to the zoonotic parasite P. knowlesi. CONCLUSIONS: In adults, submicroscopic Plasmodium infection causes no change in plasma Flt3L but does reduce circulating DCs. Plasma Flt3L concentrations increase in acute malaria, yet this increase is insufficient to restore or expand circulating CD141+ DCs, CD1c+ DCs or pDCs. These data imply that haematopoietic factors, yet to be identified and not Flt3L, involved in the sensing/maintenance of circulating DC are impacted by malaria and a submicroscopic infection. The zoonotic P. knowlesi is similar to other Plasmodium spp in compromising DC in adult malaria.

Platelets kill circulating parasites of all major Plasmodium species in human malaria.

Platelets are understood to assist host innate immune responses against infection, although direct evidence of this function in any human disease, including malaria, is unknown. Here we characterized platelet-erythrocyte interactions by microscopy and flow cytometry in patients with malaria naturally infected with Plasmodium falciparum, Plasmodium vivax, Plasmodium malariae, or Plasmodium knowlesi Blood samples from 376 participants were collected from malaria-endemic areas of Papua, Indonesia, and Sabah, Malaysia. Platelets were observed binding directly with and killing intraerythrocytic parasites of each of the Plasmodium species studied, particularly mature stages, and was greatest in P vivax patients. Platelets preferentially bound to the infected more than to the uninfected erythrocytes in the bloodstream. Analysis of intraerythrocytic parasites indicated the frequent occurrence of platelet-associated parasite killing, characterized by the intraerythrocytic accumulation of platelet factor-4 and terminal deoxynucleotidyl transferase deoxyuridine triphosphate nick-end labeling of parasite nuclei (PF4+TUNEL+ parasites). These PF4+TUNEL+ parasites were not associated with measures of systemic platelet activation. Importantly, patient platelet counts, infected erythrocyte-platelet complexes, and platelet-associated parasite killing correlated inversely with patient parasite loads. These relationships, taken together with the frequency of platelet-associated parasite killing observed among the different patients and Plasmodium species, suggest that platelets may control the growth of between 5% and 60% of circulating parasites. Platelet-erythrocyte complexes made up a major proportion of the total platelet pool in patients with malaria and may therefore contribute considerably to malarial thrombocytopenia. Parasite killing was demonstrated to be platelet factor-4-mediated in P knowlesi culture. Collectively, our results indicate that platelets directly contribute to innate control of Plasmodium infection in human malaria.

Circulating Neutrophil Extracellular Traps and Neutrophil Activation Are Increased in Proportion to Disease Severity in Human Malaria.

BACKGROUND: Neutrophil activation results in Plasmodium parasite killing in vitro, but neutrophil products including neutrophil extracellular traps (NETs) mediate host organ damage and may contribute to severe malaria. The role of NETs in the pathogenesis of severe malaria has not been examined. METHODS: In Papua, Indonesia, we enrolled adults with symptomatic Plasmodium falciparum (n = 47 uncomplicated, n = 8 severe), Plasmodium vivax (n = 37), or Plasmodium malariae (n = 14) malaria; asymptomatic P falciparum (n = 19) or P vivax (n = 21) parasitemia; and healthy adults (n = 23) without parasitemia. Neutrophil activation and NETs were quantified by immunoassays and microscopy and correlated with parasite biomass and disease severity. RESULTS: In patients with symptomatic malaria, neutrophil activation and NET counts were increased in all 3 Plasmodium species. In falciparum malaria, neutrophil activation and NET counts positively correlated with parasite biomass (Spearman rho = 0.41, P = .005 and r2 = 0.26, P = .002, respectively) and were significantly increased in severe disease. In contrast, NETs were inversely associated with parasitemia in adults with asymptomatic P falciparum infection (r2 = 0.24, P = .031) but not asymptomatic P vivax infection. CONCLUSIONS: Although NETs may inhibit parasite growth in asymptomatic P falciparum infection, neutrophil activation and NET release may contribute to pathogenesis in severe falciparum malaria. Agents with potential to attenuate these processes should be evaluated.

Antiphosphatidylserine Immunoglobulin M and Immunoglobulin G Antibodies Are Higher in Vivax Than Falciparum Malaria, and Associated With Early Anemia in Both Species.

BACKGROUND: Anemia is a major complication of vivax malaria. Antiphosphatidylserine (PS) antibodies generated during falciparum malaria mediate phagocytosis of uninfected red blood cells that expose PS and have been linked to late malarial anemia. However, their role in anemia from non-falciparum Plasmodium species is not known, nor their role in early anemia from falciparum malaria. METHODS: We measured PS immunoglobulin G (IgG) and immunoglobulin M (IgM) antibodies in Malaysian patients with vivax, falciparum, knowlesi, and malariae malaria, and in healthy controls, and correlated antibody titres with hemoglobin. PS antibodies were also measured in volunteers experimentally infected with Plasmodium vivax and Plasmodium falciparum. RESULTS: PS IgM and IgG antibodies were elevated in patients with vivax, falciparum, knowlesi, and malariae malaria (P < .0001 for all comparisons with controls) and were highest in vivax malaria. In vivax and falciparum malaria, PS IgM and IgG on admission correlated inversely with admission and nadir hemoglobin, controlling for parasitemia and fever duration. PS IgM and IgG were also increased in volunteers infected with blood-stage P. vivax and P. falciparum, and were higher in P. vivax infection. CONCLUSIONS: PS antibodies are higher in vivax than falciparum malaria, correlate inversely with hemoglobin, and may contribute to the early loss of uninfected red blood cells found in malarial anemia from both species.

Plasmodium falciparum Activates CD16+ Dendritic Cells to Produce Tumor Necrosis Factor and Interleukin-10 in Subpatent Malaria.

Background: The malaria causing parasite Plasmodium subverts host immune responses by several strategies including the modulation of dendritic cells (DCs). Methods: In this study, we show that Plasmodium falciparum skewed CD16+ DC cytokine responses towards interleukin (IL)-10 production in vitro, distinct to the cytokine profile induced by Toll-like receptor ligation. To determine CD16+ DC responsiveness in vivo, we assessed their function after induced P falciparum infection in malaria-naive volunteers. Results: CD16+ DCs underwent distinctive activation, with increased expression of maturation markers human leukocyte antigen (HLA)-DR and CD86, enhanced tumor necrosis factor (TNF) production, and coproduction of TNF/IL-10. In vitro restimulation with P falciparum further increased IL-10 production. In contrast, during naturally acquired malaria episode, CD16+ DCs showed diminished maturation, suggesting increased parasite burden and previous exposure influence DC subset function. Conclusions: These findings identify CD16+ DCs as the only DC subset activated during primary blood-stage human Plasmodium infection. As dual cytokine producers, CD16+ DCs contribute to inflammatory as well as regulatory innate immune processes.

Dysregulated IL-1ß-GM-CSF Axis in Acute Rheumatic Fever That Is Limited by Hydroxychloroquine.

BACKGROUND: Acute rheumatic fever (ARF) and rheumatic heart disease are autoimmune consequences of group A streptococcus infection and remain major causes of cardiovascular morbidity and mortality around the world. Improved treatment has been stymied by gaps in understanding key steps in the immunopathogenesis of ARF and rheumatic heart disease. This study aimed to identify (1) effector T cell cytokine(s) that might be dysregulated in the autoimmune response of patients with ARF by group A streptococcus, and (2) an immunomodulatory agent that suppresses this response and could be clinically translatable to high-risk patients with ARF. METHODS: The immune response to group A streptococcus was analyzed in peripheral blood mononuclear cells from an Australian Aboriginal ARF cohort by a combination of multiplex cytokine array, flow cytometric analysis, and global gene expression analysis by RNA sequencing. The immunomodulatory drug hydroxychloroquine was tested for effects on this response. RESULTS: We found a dysregulated interleukin-1ß-granulocyte-macrophage colony-stimulating factor (GM-CSF) cytokine axis in ARF peripheral blood mononuclear cells exposed to group A streptococcus in vitro, whereby persistent interleukin-1ß production is coupled to overproduction of GM-CSF and selective expansion of CXCR3+CCR4-CCR6- CD4 T cells. CXCR3+CCR4-CCR6- CD4 T cells are the major source of GM-CSF in human CD4 T cells and CXCL10, a CXCR3 ligand and potent T helper 1 chemoattractant, was elevated in sera from patients with ARF. GM-CSF has recently emerged as a key T cell-derived effector cytokine in numerous autoimmune diseases, including myocarditis, and the production of CXCL10 may explain selective trafficking of these cells to the heart. We provide evidence that interleukin-1ß amplifies the expansion of GM-CSF-expressing CD4 T cells, which is effectively suppressed by hydroxychloroquine. RNA sequencing showed shifts in gene expression profiles and differentially expressed genes in peripheral blood mononuclear cells derived from patients at different clinical stages of ARF. CONCLUSIONS: Given the safety profile of hydroxychloroquine and its clinical pedigree in treating autoimmune diseases such as rheumatoid arthritis, where GM-CSF plays a pivotal role, we propose that hydroxychloroquine could be repurposed to reduce the risk of rheumatic heart disease after ARF.

Early Immune Regulatory Changes in a Primary Controlled Human Plasmodium vivax Infection: CD1c+ Myeloid Dendritic Cell Maturation Arrest, Induction of the Kynurenine Pathway, and Regulatory T Cell Activation.

Plasmodium vivax malaria remains a major public health problem. The requirements for acquisition of protective immunity to the species are not clear. Dendritic cells (DC) are essential for immune cell priming but also perform immune regulatory functions, along with regulatory T cells (Treg). An important function of DC involves activation of the kynurenine pathway via indoleamine 2,3-dioxygenase (IDO). Using a controlled human experimental infection study with blood-stage P. vivax, we characterized plasmacytoid DC (pDC) and myeloid DC (mDC) subset maturation, CD4+ CD25+ CD127lo Treg activation, and IDO activity. Blood samples were collected from six healthy adults preinoculation, at peak parasitemia (day 14; ∼31,400 parasites/ml), and 24 and 48 h after antimalarial treatment. CD1c+ and CD141+ mDC and pDC numbers markedly declined at peak parasitemia, while CD16+ mDC numbers appeared less affected. HLA-DR expression was selectively reduced on CD1c+ mDC, increased on CD16+ mDC, and was unaltered on pDC. Plasma IFN-γ increased significantly and was correlated with an increased kynurenine/tryptophan (KT) ratio, a measure of IDO activity. At peak parasitemia, Treg presented an activated CD4+ CD25+ CD127lo CD45RA- phenotype and upregulated TNFR2 expression. In a mixed-effects model, the KT ratio was positively associated with an increase in activated Treg. Our data demonstrate that a primary P. vivax infection exerts immune modulatory effects by impairing HLA-DR expression on CD1c+ mDC while activating CD16+ mDC. Induction of the kynurenine pathway and increased Treg activation, together with skewed mDC maturation, suggest P. vivax promotes an immunosuppressive environment, likely impairing the development of a protective host immune response.

Profoundly Reduced CD1c+ Myeloid Dendritic Cell HLA-DR and CD86 Expression and Increased Tumor Necrosis Factor Production in Experimental Human Blood-Stage Malaria Infection.

Dendritic cells (DCs) are sentinels of the immune system that uniquely prime naive cells and initiate adaptive immune responses. CD1c (BDCA-1) myeloid DCs (CD1c(+) mDCs) highly express HLA-DR, have a broad Toll-like receptor (TLR) repertoire, and secrete immune modulatory cytokines. To better understand immune responses to malaria, CD1c(+) mDC maturation and cytokine production were examined in healthy volunteers before and after experimental intravenous Plasmodium falciparum infection with 150- or 1,800-parasite-infected red blood cells (pRBCs). After either dose, CD1c(+) mDCs significantly reduced HLA-DR expression in prepatent infections. Circulating CD1c(+) mDCs did not upregulate HLA-DR after pRBC or TLR ligand stimulation and exhibited reduced CD86 expression. At peak parasitemia, CD1c(+) mDCs produced significantly more tumor necrosis factor (TNF), whereas interleukin-12 (IL-12) production was unchanged. Interestingly, only the 1,800-pRBC dose caused a reduction in the circulating CD1c(+) mDC count with evidence of apoptosis. The 1,800-pRBC dose produced no change in T cell IFN-γ or IL-2 production at peak parasitemia or at 3 weeks posttreatment. Overall, CD1c(+) mDCs are compromised by P. falciparum exposure, with impaired HLA-DR and CD86 expression, and have an increased capacity for TNF but not IL-12 production. A first prepatent P. falciparum infection is sufficient to modulate CD1c(+) mDC responsiveness, likely contributing to hampered effector T cell cytokine responses and assisting parasite immune evasion.

Characterization of blood dendritic and regulatory T cells in asymptomatic adults with sub-microscopic Plasmodium falciparum or Plasmodium vivax infection.

BACKGROUND: Plasmodium falciparum and Plasmodium vivax infections compromise dendritic cell (DC) function and expand regulatory T (Treg) cells in both clinical disease (malaria) and experimental human sub-microscopic infection. Conversely, in asymptomatic microscopy-positive (patent) P. falciparum or P. vivax infection in endemic areas, blood DC increase or retain HLA-DR expression and Treg cells exhibit reduced activation, suggesting that DC and Treg cells contribute to the control of patent asymptomatic infection. The effect of sub-microscopic (sub-patent) asymptomatic Plasmodium infection on DC and Treg cells in malaria-endemic area residents remains unclear. METHODS: In a cross-sectional household survey conducted in Papua, Indonesia, 162 asymptomatic adults were prospectively evaluated for DC and Treg cells using field-based flow cytometry. Of these, 161 individuals (99 %) were assessed retrospectively by polymerase chain reaction (PCR), 19 of whom had sub-microscopic infection with P. falciparum and 15 with sub-microscopic P. vivax infection. Flow cytometric data were re-analysed after re-grouping asymptomatic individuals according to PCR results into negative controls, sub-microscopic and microscopic parasitaemia to examine DC and Treg cell phenotype in sub-microscopic infection. RESULTS: Asymptomatic adults with sub-microscopic P. falciparum or P. vivax infection had DC HLA-DR expression and Treg cell activation comparable to PCR-negative controls. Sub-microscopic P. falciparum infection was associated with lower peripheral CD4(+) T cells and lymphocytes, however sub-microscopic Plasmodium infection had no apparent effect on DC sub-set number or Treg cell frequency. CONCLUSIONS: In contrast to the impairment of DC maturation/function and the activation of Treg cells seen with sub-microscopic parasitaemia in primary experimental human Plasmodium infection, no phenotypic evidence of dysregulation of DC and Treg cells was observed in asymptomatic sub-microscopic Plasmodium infection in Indonesian adults. This is consistent with DC and Treg cells retaining their functional capacity in sub-microscopic asymptomatic infection with P. falciparum or P. vivax in malaria-endemic areas.

Preserved dendritic cell HLA-DR expression and reduced regulatory T cell activation in asymptomatic Plasmodium falciparum and P. vivax infection.

Clinical illness with Plasmodium falciparum or Plasmodium vivax compromises the function of dendritic cells (DC) and expands regulatory T (Treg) cells. Individuals with asymptomatic parasitemia have clinical immunity, restricting parasite expansion and preventing clinical disease. The role of DC and Treg cells during asymptomatic Plasmodium infection is unclear. During a cross-sectional household survey in Papua, Indonesia, we examined the number and activation of blood plasmacytoid DC (pDC), CD141(+), and CD1c(+) myeloid DC (mDC) subsets and Treg cells using flow cytometry in 168 afebrile children (of whom 15 had P. falciparum and 36 had P. vivax infections) and 162 afebrile adults (of whom 20 had P. falciparum and 20 had P. vivax infections), alongside samples from 16 patients hospitalized with uncomplicated malaria. Unlike DC from malaria patients, DC from children and adults with asymptomatic, microscopy-positive P. vivax or P. falciparum infection increased or retained HLA-DR expression. Treg cells in asymptomatic adults and children exhibited reduced activation, suggesting increased immune responsiveness. The pDC and mDC subsets varied according to clinical immunity (asymptomatic or symptomatic Plasmodium infection) and, in asymptomatic infection, according to host age and parasite species. In conclusion, active control of asymptomatic infection was associated with and likely contingent upon functional DC and reduced Treg cell activation.

Neutrophils with myeloid derived suppressor function deplete arginine and constrain T cell function in septic shock patients.

INTRODUCTION: Impaired T cell function in sepsis is associated with poor outcome, but the mechanisms are unclear. In cancer, arginase-expressing myeloid derived suppressor cells (MDSCs) deplete arginine, impair T cell receptor CD3 zeta-chain expression and T cell function and are linked to poor clinical outcome, but their role during acute human infectious disease and in particular sepsis remains unknown. Hypoarginemia is prevalent in sepsis. This study aimed to determine whether neutrophils that co-purify with PBMC express arginase, and if arginine depletion constrains T cell CD3 zeta-chain expression and function in human sepsis. METHODS: Using flow cytometry, cell culture, HPLC, arginase activity and mRNA detection, our study examined whether neutrophils, with reduced buoyant density isolated in the Ficoll interface, metabolise L-arginine and suppress T cell proliferation in sepsis. A total of 35 sepsis patients (23 with septic shock) and 12 hospital controls in a tertiary referral hospital in tropical Australia were evaluated. RESULTS: Only sepsis patients had interphase neutrophils, neutrophils co-purifying with mononuclear cells (≤1.077 specific gravity). The percentage of interphase neutrophils in sepsis was proportional to sepsis severity and correlated with plasma IL-6 concentrations. Ex vivo, sepsis-derived interphase neutrophils expressed arginase, metabolised culture L-arginine and suppressed T cell proliferation and CD3 zeta-chain expression. In vivo, in septic shock there was a longitudinal inverse association between interphase neutrophil number and CD3 zeta-chain expression. Depletion or inhibition of interphase neutrophils in vitro restored zeta-chain expression and T cell function. CONCLUSIONS: For the first time during an acute human infection, interphase neutrophils that express arginase were found to circulate in sepsis, in proportion to disease severity. These neutrophil-MDSCs impair T cell CD3 zeta-chain expression and T cell function via L-arginine metabolism, and likely contribute to the T cell dysfunction seen in sepsis. Modulation of neutrophil-MDSC or their downstream effects warrant consideration as targets for novel adjunctive therapies in sepsis.

Photoperiod and metabolic health: evidence, mechanism, and implications.

Circadian rhythms are evolutionarily programmed biological rhythms that are primarily entrained by the light cycle. Disruption of circadian rhythms is an important risk factor for several metabolic disorders. Photoperiod is defined as total duration of light exposure in a day. With the extended use of indoor/outdoor light, smartphones, television, computers, and social jetlag people are exposed to excessive artificial light at night increasing their photoperiod. Importantly long photoperiod is not limited to any geographical region, season, age, or socioeconomic group, it is pervasive. Long photoperiod is an established disrupter of the circadian rhythm and can induce a range of chronic health conditions including adiposity, altered hormonal signaling and metabolism, premature ageing, and poor psychological health. This review discusses the impact of exposure to long photoperiod on circadian rhythms, metabolic and mental health, hormonal signaling, and ageing and provides a perspective on possible preventive and therapeutic approaches for this pervasive challenge.

Low-level Plasmodium falciparum blood-stage infection causes dendritic cell apoptosis and dysfunction in healthy volunteers.

BACKGROUND: Dendritic cells (DCs) are highly specialized antigen-presenting cells that are crucial for initiation of immune responses. During naturally acquired malaria, DC number and function is reduced. METHODS: The timing of, parasitemia threshold of, and contribution of apoptosis to DC loss were prospectively evaluated in 10 men after experimental challenge with approximately 1800 Plasmodium falciparum-parasitized red blood cells (pRBCs) and after drug cure initiated at a parasite level of ≥ 1000 parasites/mL. RESULTS: The nadir levels of total, myeloid, and plasmacytoid DCs occurred 8 days after infection. DC loss was partially attributable to apoptosis, which was first detected on day 5 (median parasite level, 238 parasites/mL) and maximal at day 7. Remaining DCs exhibited a reduced ability to uptake particulate antigen. DC numbers recovered approximately 60 hours after antimalarial drug administration. There was no loss of DC number or function before or after drug cure in 5 men inoculated with <180 pRBCs and treated on day 6, when their parasite level was approximately 200 parasites/mL. CONCLUSIONS: Plasmodium causes DC loss in vivo, which is at least partially explained by apoptosis in response to blood-stage parasites. In primary infection, loss of DC number and function occurs early during the prepatent period and before or with onset of clinical symptoms. These findings may explain in part the inadequate development of immunity to blood-stage malaria infection.

A complementary role for the tetraspanins CD37 and Tssc6 in cellular immunity.

The cooperative nature of tetraspanin-tetraspanin interactions in membrane organization suggests functional overlap is likely to be important in tetraspanin biology. Previous functional studies of the tetraspanins CD37 and Tssc6 in the immune system found that both CD37 and Tssc6 regulate T cell proliferative responses in vitro. CD37(-/-) mice also displayed a hyper-stimulatory dendritic cell phenotype and dysregulated humoral responses. In this study, we characterize "double knockout" mice (CD37(-/-)Tssc6(-/-)) generated to investigate functional overlap between these tetraspanins. Strong evidence for a cooperative role for these two proteins was identified in cellular immunity, where both in vitro T cell proliferative responses and dendritic cell stimulation capacity are significantly exaggerated in CD37(-/-)Tssc6(-/-) mice when compared with single knockout counterparts. Despite these exaggerated cellular responses in vitro, CD37(-/-)Tssc6(-/-) mice are not more susceptible to autoimmune induction. However, in vivo responses to pathogens appear poor in CD37(-/-)Tssc6(-/-) mice, which showed a reduced ability to produce influenza-specific T cells and displayed a rapid onset hyper-parasitemia when infected with Plasmodium yoelii. Therefore, in the absence of both CD37 and Tssc6, immune function is further altered when compared with CD37(-/-) or Tssc6(-/-) mice, demonstrating a complementary role for these two molecules in cellular immunity.

Differential cellular recognition of antigens during acute Plasmodium falciparum and Plasmodium vivax malaria.

BACKGROUND: Plasmodium falciparum and Plasmodium vivax are co-endemic in the Asia-Pacific region. Their capacity to induce and sustain diverse T-cell responses underpins protective immunity. We compared T-cell responses to the largely conserved merozoite surface protein-5 (PfMSP5) during acute and convalescent falciparum and vivax malaria. METHODS: Lymphoproliferation and IFN--γ secretion to PfMSP5 and purified protein derivate were quantified in adults with falciparum (n=34), and vivax malaria (n=12) or asymptomatic residents (n=10) of Papua, Indonesia. Responses were reassessed 7-28 days following treatment. RESULTS: The frequency of IFN-γ responders to PfMSP5 was similar in acute falciparum (63%) or vivax (67%) malaria. However, significantly more IFN-γ-secreting cells were detectable during vivax compared with falciparum infection. Purified protein derivative responses showed a similarly enhanced pattern. While rapidly lost in vivax patients, PfMSP5-specific responses in falciparum malaria remained to day 28. By contrast, frequency and magnitude of lymphoproliferation to PfMSP5 were similar for falciparum and vivax infections. CONCLUSION: Cellular PfMSP5-specific responses are most frequent during either acute falciparum or vivax malaria, indicating functional T-cell responses to conserved antigens. Both effector and central memory T-cell functions are increased. Greater IFN-γ responses in acute P. vivax, suggest enhancement of pre-existing effector T-cells during acute vivax infection.