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Frontiers in Immunology 2017Tunicates are the closest relatives of vertebrates, and their peculiar phylogenetic position explains the increasing interest toward tunicate immunobiology. They are... (Review)
Review
Tunicates are the closest relatives of vertebrates, and their peculiar phylogenetic position explains the increasing interest toward tunicate immunobiology. They are filter-feeding organisms, and this greatly influences their defense strategies. The majority of the studies on tunicate immunity were carried out in ascidians. The tunic acts as a first barrier against pathogens and parasites. In addition, the oral siphon and the pharynx represent two major, highly vascularized, immune organs, where circulating hemocytes can sense non-self material and trigger immune responses that, usually, lead to inflammation and phagocytosis. Inflammation involves the recruitment of circulating cytotoxic, phenoloxidase (PO)-containing cells in the infected area, where they degranulate as a consequence of non-self recognition and release cytokines, complement factors, and the enzyme PO. The latter, acting on polyphenol substrata, produces cytotoxic quinones, which polymerize to melanin, and reactive oxygen species, which induce oxidative stress. Both the alternative and the lectin pathways of complement activation converge to activate C3: C3a and C3b are involved in the recruitment of hemocytes and in the opsonization of foreign materials, respectively. The interaction of circulating professional phagocytes with potentially pathogenic foreign material can be direct or mediated by opsonins, either complement dependent or complement independent. Together with cytotoxic cells, phagocytes are active in the encapsulation of large materials. Cells involved in immune responses, collectively called immunocytes, represent a large fraction of hemocytes, and the presence of a cross talk between cytotoxic cells and phagocytes, mediated by secreted humoral factors, was reported. Lectins play a pivotal role as pattern-recognition receptors and opsonizing agents. In addition, variable region-containing chitin-binding proteins, identified in the solitary ascidian , control the settlement and colonization of bacteria in the gut.
PubMed: 28649250
DOI: 10.3389/fimmu.2017.00674 -
Proceedings of the National Academy of... Jan 2022Gram-positive organisms with their thick envelope cannot be lysed by complement alone. Nonetheless, antibody-binding on the surface can recruit complement and mark these...
Gram-positive organisms with their thick envelope cannot be lysed by complement alone. Nonetheless, antibody-binding on the surface can recruit complement and mark these invaders for uptake and killing by phagocytes, a process known as opsonophagocytosis. The crystallizable fragment of immunoglobulins (Fcγ) is key for complement recruitment. The cell surface of is coated with Staphylococcal protein A (SpA). SpA captures the Fcγ domain of IgG and interferes with opsonization by anti- antibodies. In principle, the Fcγ domain of therapeutic antibodies could be engineered to avoid the inhibitory activity of SpA. However, the SpA-binding site on Fcγ overlaps with that of the neonatal Fc receptor (FcRn), an interaction that is critical for prolonging the half-life of serum IgG. This evolutionary adaptation poses a challenge for the exploration of Fcγ mutants that can both weaken SpA-IgG interactions and retain stability. Here, we use both wild-type and transgenic human FcRn mice to identify antibodies with enhanced half-life and increased opsonophagocytic killing in models of infection and demonstrate that antibody-based immunotherapy can be improved by modifying Fcγ. Our experiments also show that by competing for FcRn-binding, staphylococci effectively reduce the half-life of antibodies during infection. These observations may have profound impact in treating cancer, autoimmune, and asthma patients colonized or infected with and undergoing monoclonal antibody treatment.
Topics: Amino Acid Sequence; Antibodies, Bacterial; Antibodies, Monoclonal; Antibody-Dependent Cell Cytotoxicity; Complement Activation; Dose-Response Relationship, Drug; Dose-Response Relationship, Immunologic; Humans; Opsonization; Phagocytosis; Protein Binding; Protein Engineering; Protein Interaction Domains and Motifs; Receptors, Fc; Staphylococcal Protein A; Staphylococcus aureus
PubMed: 35058363
DOI: 10.1073/pnas.2114478119 -
Nature Communications Aug 2023The opportunistic fungal pathogen Cryptococcus neoformans causes lethal infections in immunocompromised patients. Macrophages are central to the host response to...
The opportunistic fungal pathogen Cryptococcus neoformans causes lethal infections in immunocompromised patients. Macrophages are central to the host response to cryptococci; however, it is unclear how C. neoformans is recognised and phagocytosed by macrophages. Here we investigate the role of TLR4 in the non-opsonic phagocytosis of C. neoformans. We find that loss of TLR4 function unexpectedly increases phagocytosis of non-opsonised cryptococci by murine and human macrophages. The increased phagocytosis observed in Tlr4 cells was dampened by pre-treatment of macrophages with oxidised-LDL, a known ligand of scavenger receptors. The scavenger receptor, macrophage scavenger receptor 1 (MSR1) (also known as SR-A1 or CD204) was upregulated in Tlr4 macrophages. Genetic ablation of MSR1 resulted in a 75% decrease in phagocytosis of non-opsonised cryptococci, strongly suggesting that it is a key non-opsonic receptor for this pathogen. We go on to show that MSR1-mediated uptake likely involves the formation of a multimolecular signalling complex involving FcγR leading to SYK, PI3K, p38 and ERK1/2 activation to drive actin remodelling and phagocytosis. Altogether, our data indicate a hitherto unidentified role for TLR4/MSR1 crosstalk in the non-opsonic phagocytosis of C. neoformans.
Topics: Animals; Humans; Mice; Cryptococcosis; Cryptococcus neoformans; Macrophages; Phagocytosis; Toll-Like Receptor 4; Scavenger Receptors, Class A
PubMed: 37580395
DOI: 10.1038/s41467-023-40635-w -
Journal of Infection and Chemotherapy :... Jun 2013Streptococcus pneumoniae is a major human pathogen responsible for the majority of bacterial pneumonia cases as well as invasive pneumococcal diseases with high... (Review)
Review
Streptococcus pneumoniae is a major human pathogen responsible for the majority of bacterial pneumonia cases as well as invasive pneumococcal diseases with high mortality and morbidity. Use of conjugate vaccines targeting the pneumococcal capsule has dramatically reduced the incidence of invasive diseases, and there are active efforts to further improve the conjugate vaccines. However, in children new pneumococcal vaccines can no longer be tested with placebo-based clinical trials because effective vaccines are currently available. Thus, vaccine studies must depend on surrogate markers of vaccine efficacy. Although traditional antibody levels (e.g., ELISA) are useful as a surrogate marker of protection, they have limitations, and a bioassay measuring the capacity of antibodies to opsonize pneumococci has been developed. This opsonophagocytosis assay (OPA) replicates the in vivo mechanism of antibody protection and should therefore better reflect protection by vaccine-induced antibodies. Technical improvements of OPA have made this bioassay rapid, multiplexed, and practical for analyzing small samples including those from children. Strong correlations between ELISA and OPA have been observed in many studies of young children. However, poor correlations have been found in some important clinical situations (such as determination of protection by cross-reactive antibodies) and populations (such as elderly adults and immunodeficient patients). In these settings, OPA has become a useful supplementary measure of pneumococcal vaccine immunogenicity. Current efforts to standardize OPA will further expand its uses.
Topics: Antibodies, Bacterial; Biomarkers; Cytological Techniques; Enzyme-Linked Immunosorbent Assay; Humans; Opsonin Proteins; Phagocytosis; Pneumococcal Vaccines; Streptococcus pneumoniae; Vaccines, Conjugate
PubMed: 23657429
DOI: 10.1007/s10156-013-0601-1 -
Traffic (Copenhagen, Denmark) Aug 2012Phagocytosis is a key cellular process, both during homeostasis and upon infection or tissue damage. Receptors on the surface of professional phagocytic cells bind to... (Review)
Review
Phagocytosis is a key cellular process, both during homeostasis and upon infection or tissue damage. Receptors on the surface of professional phagocytic cells bind to target particles either directly or through opsonizing ligands, and trigger actin-mediated ingestion of the particles. The process must be carefully controlled to ensure that phagocytosis is triggered efficiently and specifically, and that the antimicrobial cytotoxic responses that often accompany it are initiated only when required. In this review, we will describe and compare the molecular mechanisms that regulate phagocytosis triggered by Fcγ receptors, which mediate the uptake of immunoglobulin G-opsonized targets, and Dectin-1, which is responsible for internalization of fungi with exposed cell wall β-glucan. We will examine how these receptors detect their ligands, how signal transduction is initiated and regulated, and how internalization is instructed to achieve rapid and yet controlled uptake of their targets.
Topics: Animals; Antigens, Fungal; Humans; Immunoglobulin G; Lectins, C-Type; Phagocytosis; Receptor Aggregation; Receptors, Fc; Signal Transduction; beta-Glucans
PubMed: 22624959
DOI: 10.1111/j.1600-0854.2012.01382.x -
Frontiers in Immunology 2022Crustins are the most abundant class of antimicrobial peptides in crustaceans and are essential for protecting animals from infection. Among them, type II crustins...
Crustins are the most abundant class of antimicrobial peptides in crustaceans and are essential for protecting animals from infection. Among them, type II crustins usually exhibit potent antimicrobial activity. Interestingly, in this study, a newly identified type II crustin gene homolog (named SpCrus8) from mud crab , the recombinant proteins of which (rSpCrus8 and rTrx-SpCrus8) showed no obvious antibacterial effects, but could significantly reduce the bacterial load in crab hemolymph and improve the survival rate of crabs infected with . The immune-related function of SpCrus8 and the underlying mechanism deserve further investigation. It was found that the SpCrus8 gene was widely distributed in various tissues of adult crabs. In the hepatopancreas of crabs infected with or , transcripts of the SpCrus8 gene were remarkably induced, indicating that the SpCrus8 gene was involved in the immune response to bacterial infection . In addition, rSpCrus8 and rTrx-SpCrus8 had strong binding activity not only to microbial surface components (lipopolysaccharide, lipoteichoic acid, peptidoglycan, and glucan), but also to the tested bacteria (, and ). Notably, rSpCrus8 and rTrx-SpCrus8 could significantly promote hemocyte phagocytosis. After rSpCrus8 and rTrx-SpCrus8 treatment, a large number of fluorescent microspheres were observed to aggregate into clusters and be phagocytosed by multiple hemocytes, while hemocytes in the control group phagocytosed only individual microspheres, indicating that SpCrus8 played an important role in opsonization. When the SpCrus8 gene was knocked down, the expression levels of the key phagocytosis-related genes SpRab5 and SpRab7 were significantly downregulated, as well as the IMD signaling pathway genes SpIKKβ and SpRelish, and another crustin gene SpCrus5. Correspondingly, all the SpIKKβ, SpRelish and SpCrus5 genes were significantly upregulated after rSpCrus8 treatment, suggesting that SpCrus8 might be involved in the immunomodulation of . Taken together, this study revealed the immune-related functions of the SpCrus8 gene in opsonization and regulation, which will help us further understand the role of the crustin gene family in the immune system of mud crabs and provide new insights into the function of type II crutins.
Topics: Animals; Antimicrobial Cationic Peptides; Arthropod Proteins; Brachyura; Gene Expression Regulation; Immunity, Innate; Immunomodulation; Opsonization; Staphylococcal Infections; Staphylococcus aureus
PubMed: 35874773
DOI: 10.3389/fimmu.2022.946227 -
Microbiology Spectrum Dec 2022The human pathogen Streptococcus pyogenes causes substantial morbidity and mortality. It is unclear if antibodies developed after infections with this pathogen are...
The human pathogen Streptococcus pyogenes causes substantial morbidity and mortality. It is unclear if antibodies developed after infections with this pathogen are opsonic and if they are strain specific or more broadly protective. Here, we quantified the opsonic-antibody response following invasive S. pyogenes infection. Four patients with S. pyogenes bacteremia between 2018 and 2020 at Skåne University Hospital in Lund, Sweden, were prospectively enrolled. Acute- and convalescent-phase sera were obtained, and the S. pyogenes isolates were genome sequenced (118, 85, and two 1 isolates). Quantitative antibody binding and phagocytosis assays were used to evaluate isolate-dependent opsonic antibody function in response to infection. Antibody binding increased modestly against the infecting isolate and across types in convalescent- compared to acute-phase sera for all patients. For two patients, phagocytosis increased in convalescent-phase serum both for the infecting isolate and across types. The increase was only across types for one patient, and one had no improvement. No correlation to the clinical outcomes was observed. Invasive S. pyogenes infections result in a modestly increased antibody binding with differential opsonic capacity, both nonfunctional binding and broadly opsonic binding across types. These findings question the dogma that an invasive infection should lead to a strong type-specific antibody increase rather than a more modest but broadly reactive response, as seen in these patients. Furthermore, our results indicate that an increase in antibody titers might not be indicative of an opsonic response and highlight the importance of evaluating antibody function in S. pyogenes infections. The bacterium Streptococcus pyogenes is a common cause of both mild and severe human diseases resulting in substantial morbidity and mortality each year. No vaccines are available, and our understanding of the antibody response to this human pathogen is still incomplete. Here, we carefully analyzed the opsonic antibody response following invasive infection in four patients. Unexpectedly, the patients did not always generate opsonic antibodies against the specific infecting strain. Instead, we found that some patients could generate cross-opsonic antibodies, leading to phagocytosis of bacteria across strains. The emergence of cross-opsonic antibodies is likely important for long-term immunity against S. pyogenes. Our findings question the dogma that mostly strain-specific immunity is developed after infection and add to our overall understanding of how immunity to S. pyogenes can evolve.
Topics: Humans; Streptococcal Infections; Phagocytosis; Streptococcus pyogenes; Antibodies, Bacterial; Bacteremia; Antigens, Bacterial
PubMed: 36314947
DOI: 10.1128/spectrum.02486-22 -
Frontiers in Immunology 2017Balancing surface functionalization and low immune recognition of nanomedicines is a major challenge. Opsonization with the third component of the complement protein...
Balancing surface functionalization and low immune recognition of nanomedicines is a major challenge. Opsonization with the third component of the complement protein (C3) plays a major role in immune cell recognition of nanomedicines. We used dextran-coated superparamagnetic iron oxide nanoworms (SPIO NWs) to study the effect of surface functionalization on C3 opsonization in mouse serum and subsequent macrophage/leukocyte recognition as well as on intravenous injection into mice. Previously, we found that in mouse serum, SPIO NWs became opsonized with C3 complement lectin pathway. Crosslinking the dextran shell with epichlorohydrin significantly decreased C3 opsonization and uptake by mouse peritoneal macrophages. Crosslinked nanoworms (NWs) further functionalized with polyethylene glycol (PEG) or with PEG-antibody (Ab) (~160 IgG molecules/particle) did not show an increase in C3 opsonization and peritoneal macrophage uptake . Following tail vein injection into mice, plain crosslinked NWs and PEGylated crosslinked NWs showed very low C3 opsonization and mouse leukocyte uptake. However, Ab-decorated crosslinked NWs showed significant C3 opsonization and high level of complement-dependent uptake by leukocytes in mice. Decreasing the number of conjugated Abs to 46 IgG molecules/particle significantly reduced C3 opsonization and leukocyte uptake. Using fresh mouse lepirudin plasma rather than serum showed better correlation with C3 opsonization . The reason for this difference could be related to the known instability of complement classical pathway in mouse sera. Our data illustrate that fine-tuning in nanoparticle surface functionalization with Abs is required to avoid excessive complement activation and complement-mediated immune uptake in mice, and raise issues with immunological assays of nanomedicines intended to mimic conditions.
PubMed: 28239384
DOI: 10.3389/fimmu.2017.00151 -
Journal of Translational Autoimmunity 2021Bactericidal/permeability-increasing protein (BPI) is an anti-microbial protein predominantly expressed in azurophilic granules of neutrophils. BPI has been shown to... (Review)
Review
Bactericidal/permeability-increasing protein (BPI) is an anti-microbial protein predominantly expressed in azurophilic granules of neutrophils. BPI has been shown to mediate cytocidal and opsonic activity against Gram-negative bacteria, while also blunting inflammatory activity of lipopolysaccharide (LPS). Despite awareness of these functions the magnitude of the contribution of BPI to innate immunity remains unclear, and the nature of the functional role of BPI has been submitted to limited investigation. Understanding this role takes on particular interest with the recognition that autoimmunity to BPI is tightly linked to a specific infectious trigger like in chronic lung infection. This has led to the notion that anti-BPI autoantibodies compromise the activity of BPI in innate immunity against , which is primarily mediated by neutrophils. In this review, we explore the three main mechanisms in bactericidal, opsonic, and anti-inflammatory of BPI. We address the etiology and the effects of BPI autoreactivity on BPI function. We explore BPI polymorphism and its link to multiple diseases. We summarize BPI therapeutic potential in both animal models and human studies, as well as offer therapeutic approaches to designing a sustainable and promising BPI molecule.
PubMed: 34142075
DOI: 10.1016/j.jtauto.2021.100105 -
Frontiers in Immunology 2023Syphilis, a sexually transmitted infection caused by the spirochete (), is resurging globally. 's repertoire of outer membrane proteins (OMPs) includes BamA (β-barrel...
INTRODUCTION
Syphilis, a sexually transmitted infection caused by the spirochete (), is resurging globally. 's repertoire of outer membrane proteins (OMPs) includes BamA (β-barrel assembly machinery subunit A/TP0326), a bipartite protein consisting of a 16-stranded β-barrel with nine extracellular loops (ECLs) and five periplasmic POTRA (polypeptide transport-associated) domains. BamA ECL4 antisera promotes internalization of by rabbit peritoneal macrophages.
METHODS
Three overlapping BamA ECL4 peptides and a two-stage, phage display strategy, termed "Epivolve" (for epitope evolution) were employed to generate single-chain variable fragments (scFvs). Additionally, antisera generated by immunizing mice and rabbits with BamA ECL4 displayed by a thioredoxin scaffold (Trx). MAbs and antisera reactivities were evaluated by immunoblotting and ELISA. A comparison of murine and rabbit opsonophagocytosis assays was conducted to evaluate the functional ability of the Abs (, opsonization) and validate the mouse assay. Sera from -infected mice (MSS) and rabbits (IRS) were evaluated for ECL4-specific Abs using Trx and overlapping ECL4 peptides in immunoblotting and ELISA assays.
RESULTS
Each of the five mAbs demonstrated reactivity by immunoblotting and ELISA to nanogram amounts of Trx. One mAb, containing a unique amino acid sequence in both the light and heavy chains, showed activity in the murine opsonophagocytosis assay. Mice and rabbits hyperimmunized with Trx produced opsonic antisera that strongly recognized the ECL presented in a heterologous scaffold and overlapping ECL4 peptides, including S2. In contrast, Abs generated during infection of mice and rabbits poorly recognized the peptides, indicating that S2 contains a subdominant epitope.
DISCUSSION
Epivolve produced mAbs target subdominant opsonic epitopes in BamA ECL4, a top syphilis vaccine candidate. The murine opsonophagocytosis assay can serve as an alternative model to investigate the opsonic potential of vaccinogens. Detailed characterization of BamA ECL4-specific Abs provided a means to dissect Ab responses elicited by infection.
Topics: Mice; Animals; Rabbits; Treponema pallidum; Antibodies, Monoclonal; Syphilis; Immune Sera; Bacteriophages; Epitopes
PubMed: 37675118
DOI: 10.3389/fimmu.2023.1222267