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Demographics of participants in case-control studies.
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Severe malaria-associated anemia and cerebral malaria are life-threatening complications of Plasmodium falciparum infection. Red blood cell (RBC) complement regulatory proteins (CRPs) have been implicated in the pathogenesis of both. We
sought to determine whether there are age-related changes in the expression of CRPs that could explain the suscep...
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Variant surface antigens (VSAs) play a critical role in severe malaria pathogenesis. Defining gaps, or "lacunae", in immunity to these Plasmodium falciparum antigens in children with severe malaria would improve our understanding of vulnerability to severe malaria and how protective immunity develops. Using a protein microarray with 179 antigen var...
Citations
... It can be hypothesized that this should make aged erythrocytes less prone to necroptosis, simultaneously promoting eryptosis and hence immunologically silent clearance of old RBCs. It is worth mentioning that CD59 levels are higher in severe malaria-associated anemia [143,144]. Notably, non-infected erythrocytes don't have elevated CD59 [145]. CD59 overexpression might make erythrocytes more susceptible to necroptosis fueling the immune response via erythroid DAMPs release. ...
Necroptosis is considered a programmed necrosis that requires receptor-interacting protein kinase 1 (RIPK1), receptor-interacting protein kinase 3 (RIPK3), and pore-forming mixed lineage kinase domain-like protein (MLKL) to trigger a regulated cell membrane lysis. Membrane rupture in necroptosis has been shown to fuel innate immune response due to release of damage-associated molecular patterns (DAMPs). Recently published studies indicate that mature erythrocytes can undergo necroptosis as well. In this review, we provide an outline of multiple cell death modes occurring in erythrocytes, discuss possible immunological aspects of diverse erythrocyte cell deaths, summarize available evidence related to the ability of erythrocytes to undergo necroptosis, outline key involved molecular mechanisms, and discuss the potential implication of erythrocyte necroptosis in the physiology and pathophysiology. Furthermore, we aim to highlight the interplay between necroptosis and eryptosis signaling in erythrocytes, emphasizing specific characteristics of these pathways distinct from their counterparts in nucleated cells. Thus, our review provides a comprehensive summary of the current knowledge of necroptosis in erythrocytes. To reflect critical differences between necroptosis of nucleated cells and necroptosis of erythrocytes, we suggest a term erythronecroptosis for necroptosis of enucleated cells.
... Age of the subjects studied was probably important. Indeed, Waitumbi et al. (33) observed a lower RBC complement regulatory protein in young children and increased into adulthood. In contrast, we included only adult patients in our study. ...
Background
During sepsis, red blood cell (RBC) deformability is altered. Persistence of these alterations is associated with poor outcome. Activation of the complement system is enhanced during sepsis and RBCs are protected by membrane surface proteins like CD35, CD55 and CD59. In malaria characterized by severe anemia, a study reported links between the modifications of the expression of these RBCs membrane proteins and erythrophagocytosis. We studied the evolution of RBCs deformability and the expression of RBC membrane surface IgG and regulatory proteins in septic patients.
Methods
By flow cytometry technics, we measured at ICU admission and at day 3–5, the RBC membrane expression of IgG and complement proteins (CD35, 55, 59) in septic patients compared to RBCs from healthy volunteers. Results were expressed in percentage of RBCs positive for the protein. RBC shape was assessed using Pearson's second coefficient of dissymmetry (PCD) on the histogram obtained with a flow cytometer technique. A null value represents a perfect spherical shape. RBC deformability was determined using ektacytometry by the elongation index in relation to the shear stress (0.3–50 Pa) applied to the RBC membrane. A higher elongation index indicates greater RBC deformability.
Results
RBCs from 11 septic patients were compared to RBCs from 21 volunteers. At ICU admission, RBCs from septic patients were significantly more spherical and RBC deformability was significantly lower in septic patients for all shear stress ≥1.93 Pa. These alterations of shape and deformability persists at day 3–5. We observed a significant decrease at ICU admission only in CD35 expression on RBCs from septic patients. This low expression remained at day 3–5.
Conclusions
We observed in RBCs from septic patients a rapid decrease expression of CD35 membrane protein protecting against complement activation. These modifications associated with altered RBC deformability and shape could facilitate erythrophagocytosis, contributing to anemia observed in sepsis. Other studies with a large number of patients and assessment of erythrophagocytosis were needed to confirm these preliminary data.
... Other than erythrophagocytosis, erythrocytes with C3bcontaining ICs are taken up by splenic reticuloendothelial cells. This may lead to stripping off of the CR1 from the erythrocyte surface (76,79,80). CR1-deficient erythrocytes are recirculated and are susceptible to complement attack, implicating complement deposition as a driver of severe malarial anaemia. ...
Antibody immunity against malaria is effective but non-sterile. In addition to antibody-mediated inhibition, neutralisation or opsonisation of malaria parasites, antibody-mediated complement activation is also important in defense against infection. Antibodies form immune complexes with parasite-derived antigens that can activate the classical complement pathway. The complement system provides efficient surveillance for infection, and its activation leads to parasite lysis or parasite opsonisation for phagocytosis. The induction of complement-fixing antibodies contributes significantly to the development of protective immunity against clinical malaria. These complement-fixing antibodies can form immune complexes that are recognised by complement receptors on innate cells of the immune system. The efficient clearance of immune complexes is accompanied by complement receptor internalisation, abrogating the detrimental consequences of excess complement activation. Here, we review the mechanisms of activation of complement by alternative, classical, and lectin pathways in human malaria at different stages of the Plasmodium life cycle with special emphasis on how complement-fixing antibodies contribute to protective immunity. We briefly touch upon the action of anaphylatoxins, the assembly of membrane attack complex, and the possible reasons underlying the resistance of infected erythrocytes towards antibody-mediated complement lysis, relevant to their prolonged survival in the blood of the human host. We make suggestions for further research on effector functions of antibody-mediated complement activation that would guide future researchers in deploying complement-fixing antibodies in preventive or therapeutic strategies against malaria.
... The complement cascade is under tight regulation to avoid consumption of complement proteins and activated C1s would usually be inhibited by C1 esterase inhibitor. During malaria infection, complement is activated (49,50) and complement regulatory proteins are depleted (49,51,52). Therefore, PfEMP1 proteins at the IE surface may be exposed to activated serine proteases, such as complement C1s and thrombin, which then cleave PfEMP1 and interfere with cytoadhesion in vivo. ...
Significance
Mature asexual stages of Plasmodium falciparum -infected erythrocytes (IEs) bind to endothelium to avoid splenic clearance. The PfEMP1 family is the major cytoadhesion ligand at the IE surface and is essential for parasite sequestration. In this study, we show that complement component 1s (C1s), found in serum, cleaves PfEMP1 on the IEs to prevent binding to endothelial cells. We find that the C1s cleavage sites are maintained at semiconserved arginine motifs located at interdomain regions between PfEMP1 receptor-binding domains and upstream of the transmembrane region. This suggests that parasites have taken advantage of a human serum protease, either to escape antibody recognition of sequestered parasites or to dampen sequestration and pathogenesis from uncontrolled inflammatory responses threatening host survival.
... CR1/E decreases by one third during the 120-day life of E in normal individuals (Cohen et al., 1992), but a larger proportion of CR1/E can be lost in acute situations, making CR1/E measurement a cumulative assessment of what happened during the E lifespan, like HbA1c, but in the opposite direction. An acquired decrease of CR1/E has been described in AIDS , SLE (Cohen et al., 1992), Alzheimer's disease (Mahmoudi et al, 2015(Mahmoudi et al, , 2018, and malaria (Waitumbi et al., 2004). CR1 allele structural polymorphisms mainly located in the exon 29 mutation hot spot have been found to be associated with resistance or susceptibility to malaria in Africa (Stoute, 2011). ...
In order to study the mechanisms of COVID-19 damage following the complement activation phase occurring during the innate immune response to SARS-CoV-2, CR1 (the regulating complement activation factor, CD35, the C3b/C4b receptor), C4d deposits on Erythrocytes (E), and the products of complement activation C3b/C3bi, were assessed in 52 COVID-19 patients undergoing O2 therapy or assisted ventilation in ICU units in Rheims France. An acquired decrease of CR1 density on E from COVID-19 patients was observed (Mean 418, SD 162, N=52) versus healthy individuals (Mean = 592, SD = 287, N= 400), Student’s t-test p<10⁻⁶, particularly among fatal cases, and in parallel with several parameters of clinical severity. Large deposits of C4d on E in patients were well above values observed in normal individuals, mostly without concomitant C3 deposits, in more than 80% of the patients. This finding is reminiscent of the increased C4d deposits on E previously observed to correlate with sub endothelial pericapillary deposits in organ transplant rejection, and with clinical SLE flares. Conversely, significant C3 deposits on E were only observed among ¼ of the patients. The decrease of CR1/E density, deposits of C4 fragments on E and previously reported detection of virus spikes or C3 on E among COVID-19 patients, suggest that the handling and clearance of immune complex or complement fragment coated cell debris may play an important role in the pathophysiology of SARS-CoV-2. Measurement of C4d deposits on E might represent a surrogate marker for assessing inflammation and complement activation occurring in organ capillaries and CR1/E decrease might represent a cumulative index of complement activation in COVID-19 patients.
Taken together, these original findings highlight the participation of complement regulatory proteins and indicate that E are important in immune pathophysiology of COVID-19 patients. Besides a potential role for monitoring the course of disease, these observations suggest that novel therapies such as the use of CR1, or CR1-like molecules, in order to down regulate complement activation and inflammation, should be considered.
... Despite that low density, the number of E makes CR1/E the major source of CR1 available in the blood. An acquired decrease of CR1E has been described in AIDS , SLE (Cohen et al., 1992), Malaria (Waitumbi et al., 2004) and Alzheimer disease (Mahmoudi et al, 2015;Mahmoudi et al, 2018). Primates do express CR1 on E (instead of platelets in most mammals) and use it as the main mechanism of immune complex (IC) capture and transportation, as well for the removal of cellular debris, either recognized by natural IgM auto-antibodies or directly activating the alternate pathway of the complement (C) system (Cornacoff et al., 1983;Waxman et al., 1984;Waxman et al.,1986). ...
We determined CR1, CD35 the C3b, C4b receptor density, C3b/C3bi and C4d deposits densities on Erythrocytes (E) in 51 COVID-19 patients undergoing O2 therapy or assisted ventilation in ICU units in Rheims France. A clear acquired decrease of CR1 density of E from COVID-19 patients was observed, particularly among fatal cases, and paralleling several severity parameters.
Deposits of C4d largely above values observed in normal individuals, mostly without C3 deposits, have been observed in more than 80% of the patients, reminiscent of the sub endothelial pericapil-lary deposits in organ transplant rejection, already observed on E in parallel, as well as also observed on E in clinical SLE flares.
Conversely, significant C3 deposits were only observed among 1/4 of the patients. The decrease of CR1/E density, and the detection of virus spike, C3 or C4 fragment on E, among COVID-19 pa-tients, are likely to be two aspects of the same phenomenon of immune complexes or complement fragment coated cell debris handling and clearance.
Measurement of C4d deposit on E might represent a way for assessing inflammation and comple-ment activation occurring in organ capillaries. CR1/E decrease might represent a cumulative index of complement activation in COVID-19 patients.
Taken together, these original findings stress on the participation of the complement regulatory pro-teins in that disease and evidence that E matter in immune mechanisms in COVID-19 patients.
The use of CR1, or CR1-like molecules with the aim of down regulating complement activation and inflammation for therapy should also be considered.
... The second advantage of this method is that it is adaptable to the quantification of other cellular receptors whose density is low by simply replacing the primary anti-CR1 antibody with an antibody specifically directed against an epitope of the receptor to be studied. It is also adaptable to using 96 well plates instead of tube racks, which requires lower blood and reagent volumes 25,38 . The third advantage of this method is that it is flexible. ...
CR1 (CD35, Complement Receptor type 1 for C3b/C4b) is a high molecular weight membrane glycoprotein of about 200 kDa that controls complement activation, transports immune complexes, and participates in humoral and cellular immune responses. CR1 is present on the surface of many cell types, including erythrocytes, and exhibits polymorphisms in length, structure (Knops, or KN, blood group), and density. The average density of CR1 per erythrocyte (CR1/E) is 500 molecules per erythrocyte. This density varies from one individual to another (100–1,200 CR1/E) and from one erythrocyte to another in the same individual. We present here a robust flow cytometry method to measure the density of CR1/E, including in subjects expressing a low density, with the help of an amplifying immunostaining system. This method has enabled us to show the lowering of CR1 erythrocyte expression in diseases such as Alzheimer's disease (AD), systemic lupus erythematosus (SLE), AIDS, or malaria.
... Haemolysis and dyserythropoiesis are key features of SMA [4][5][6][7][8][9] , but the causal haematological and immune mediators of this fatal condition are not well defined. Inflammatory cytokine levels in plasma as well as complement regulatory protein levels on red blood cells (RBCs) differ between children presenting with SMA versus those with mild malaria [reviewed in [10][11][12][13] ], but these associations do not confirm causality. ...
... This can leave the RBC vulnerable to complement attack and haemolysis. Levels of CD35, and other complement regulatory proteins, such as CD55 and CD59, are lower on the surface of RBCs in patients with SMA [11][12][13] . RBCs from these patients also have increased susceptibility to phagocytosis versus those from patients with uncomplicated malaria 13 . ...
... As in children, increased EPO expression had no clear impact on reticulocyte levels in the blood of rhesus macaques. Increased pro-inflammatory cytokine responses [reviewed in 10 ] and low levels of CD35 on RBCs [11][12][13] have been seen in children at the time of SMA. Our data showing increased pro-inflammatory responses during early infection and low levels of CD35 on RBCs at baseline are consistent with the idea that these have causal roles in the pathogenesis of SMA. ...
Severe malarial anaemia (SMA) is the most common life-threatening complication of Plasmodium falciparum infection in African children. SMA is characterised by haemolysis and inadequate erythropoiesis, and is associated with dysregulated inflammatory responses and reduced complement regulatory protein levels (including CD35). However, a deeper mechanistic understanding of the pathogenesis requires improved animal models. In this comparative study of two closely related macaque species, we interrogated potential causal factors for their differential and temporal relationships to onset of SMA. We found that rhesus macaques inoculated with blood-stage Plasmodium coatneyi developed SMA within 2 weeks, with no other severe outcomes, whereas infected cynomolgus macaques experienced only mild/ moderate anaemia. The abrupt drop in haematocrit in rhesus was accompanied by consumption of haptoglobin (haemolysis) and poor reticulocyte production. Rhesus developed a greater inflammatory response than cynomolgus macaques, and had lower baseline levels of CD35 on red blood cells (RBCs) leading to a significant reduction in the proportion of CD35+ RBCs during infection. Overall, severe anaemia in rhesus macaques infected with P. coatneyi has similar features to SMA in children. Our comparisons are consistent with an association of low baseline CD35 levels on RBCs and of early inflammatory responses with the pathogenesis of SMA.
... Consistent with previous studies demonstrating that CRP loss is associated with severe malarial anaemia [11,19], the current results show that the expression of CRPs on RBC is also reduced in individuals with mild malarial anaemia associated with either falciparum or vivax malaria. Loss of CRPs is associated with haemoglobin [11,19,24,25], suggesting that CRP loss may also be an important contributor to chronic malarial anaemia in low transmission settings. The present study also shows that CRPs are reduced on RBCs following experimental IBSM with P. falciparum where parasitaemia is very low. ...
Background:
Anaemia is a major consequence of malaria, caused by the removal of both infected and uninfected red blood cells (RBCs) from the circulation. Complement activation and reduced expression of complement regulatory proteins (CRPs) on RBCs are an important pathogenic mechanism in severe malarial anaemia in both Plasmodium falciparum and Plasmodium vivax infection. However, little is known about loss of CRPs on RBCs during mild malarial anaemia and in low-density infection.
Methods:
The expression of CRP CR1, CD55, CD59, and the phagocytic regulator CD47, on uninfected normocytes and reticulocytes were assessed in individuals from two study populations: (1) P. falciparum and P. vivax-infected patients from a low transmission setting in Sabah, Malaysia; and, (2) malaria-naïve volunteers undergoing P. falciparum induced blood-stage malaria (IBSM). For clinical infections, individuals were categorized into anaemia severity categories based on haemoglobin levels. For IBSM, associations between CRPs and haemoglobin level were investigated.
Results:
CRP expression on RBC was lower in Malaysian individuals with P. falciparum and P. vivax mild malarial anaemia compared to healthy controls. CRP expression was also reduced on RBCs from volunteers during IBSM. Reduction occurred on normocytes and reticulocytes. However, there was no significant association between reduced CRPs and haemoglobin during IBSM.
Conclusions:
Removal of CRPs occurs on both RBCs and reticulocytes during Plasmodium infection even in mild malarial anaemia and at low levels of parasitaemia.
... SERPING1 or C1inh and CR1 or CD35). Similarly, a previous study indicated diminished CD35 and CD55 levels expressed on erythrocyte membranes and enhanced erythrophagocytosis in Kenyan children with severe malarial anemia, compared to children with uncomplicated malaria [28]. Several markers, including low haptoglobin (Hp) levels due to higher Hp consumption, indicate increased intravascular hemolysis in these children [29]. ...
Background
Transcriptomic research of blood cell lineages supports the understanding of distinct features of the immunopathology in human malaria.
Methods
We used microarray hybridization, validated by real-time RT-PCR to analyze whole blood gene expression in healthy Gabonese children and children with various conditions of Plasmodium falciparum infection, including i) asymptomatic infection, ii) uncomplicated malaria, iii) malaria associated with severe anemia and iv) cerebral malaria.
Findings
Our data indicate that the expression profile of 22 genes significantly differed among the investigated groups. Immunoglobulin production, complement regulation and IFN beta signaling, in particular IRF7 and ISRE binding signatures in the corresponding genes, were most conspicuous. Down-regulation in cerebral malaria seems to rely on AhRF, GABP and HIF1 hypoxia transcription factors. ARG1, BPI, CD163, IFI27, HP and TNFAIP6 transcript levels correlated positively with lactatemia, and negatively with hemoglobin concentrations.
Interpretation
Differences in gene expression profile reflect distinct immunopathological mechanisms of P. falciparum infection. They emerge as potential prognostic markers for early therapeutic measures and need to be validated further.
Fund
This work was supported by a grant of the NGFN (Nationales Genomforschungsnetz 01GS0114) and by a CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico, Brazil) PhD scholarship for A. B. W. Boldt. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
