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1.
Cells ; 10(5)2021 05 12.
Artigo em Inglês | MEDLINE | ID: mdl-34065953

RESUMO

Macrophages play a key role in induction of inflammatory responses. These inflammatory responses are mostly considered to be instigated by activation of pattern recognition receptors (PRRs) or cytokine receptors. However, recently it has become clear that also antibodies and pentraxins, which can both activate Fc receptors (FcRs), induce very powerful inflammatory responses by macrophages that can even be an order of magnitude greater than PRRs. While the physiological function of this antibody-dependent inflammation (ADI) is to counteract infections, undesired activation or over-activation of this mechanism will lead to pathology, as observed in a variety of disorders, including viral infections such as COVID-19, chronic inflammatory disorders such as Crohn's disease, and autoimmune diseases such as rheumatoid arthritis. In this review we discuss how physiological ADI provides host defense by inducing pathogen-specific immunity, and how erroneous activation of this mechanism leads to pathology. Moreover, we will provide an overview of the currently known signaling and metabolic pathways that underlie ADI, and how these can be targeted to counteract pathological inflammation.


Assuntos
Anticorpos/metabolismo , Proteína C-Reativa/metabolismo , Inflamação/imunologia , Componente Amiloide P Sérico/metabolismo , Anticorpos/imunologia , Proteína C-Reativa/imunologia , Interações Hospedeiro-Patógeno/imunologia , Humanos , Imunidade Inata , Inflamação/metabolismo , Inflamação/microbiologia , Macrófagos/imunologia , Macrófagos/metabolismo , Redes e Vias Metabólicas/imunologia , Receptores Fc/metabolismo , Componente Amiloide P Sérico/imunologia , Transdução de Sinais/imunologia
2.
Int J Mol Sci ; 22(7)2021 Mar 24.
Artigo em Inglês | MEDLINE | ID: mdl-33805142

RESUMO

Alzheimer's disease (AD) is a debilitating neurological disorder, and currently, there is no cure for it. Several pathologic alterations have been described in the brain of AD patients, but the ultimate causative mechanisms of AD are still elusive. The classic hallmarks of AD, including amyloid plaques (Aß) and tau tangles (tau), are the most studied features of AD. Unfortunately, all the efforts targeting these pathologies have failed to show the desired efficacy in AD patients so far. Neuroinflammation and impaired autophagy are two other main known pathologies in AD. It has been reported that these pathologies exist in AD brain long before the emergence of any clinical manifestation of AD. Microglia are the main inflammatory cells in the brain and are considered by many researchers as the next hope for finding a viable therapeutic target in AD. Interestingly, it appears that the autophagy and mitophagy are also changed in these cells in AD. Inside the cells, autophagy and inflammation interact in a bidirectional manner. In the current review, we briefly discussed an overview on autophagy and mitophagy in AD and then provided a comprehensive discussion on the role of these pathways in microglia and their involvement in AD pathogenesis.


Assuntos
Doença de Alzheimer/patologia , Autofagia , Microglia/metabolismo , Mitofagia , Peptídeos beta-Amiloides/metabolismo , Animais , Encéfalo/patologia , Proteínas do Sistema Complemento/metabolismo , Humanos , Inflamação , Camundongos , Camundongos Transgênicos , Mitocôndrias/metabolismo , Doenças Neurodegenerativas/patologia , Neurônios/metabolismo , Receptores Fc/metabolismo , Receptores Depuradores/metabolismo , Proteínas tau/metabolismo
3.
Int J Mol Sci ; 22(6)2021 Mar 17.
Artigo em Inglês | MEDLINE | ID: mdl-33802650

RESUMO

As an essential modulator of IgG disposition, the neonatal Fc receptor (FcRn) governs the pharmacokinetics and functions many therapeutic modalities. In this review, we thoroughly reexamine the hitherto elucidated biological and thermodynamic properties of FcRn to provide context for our assessment of more recent advances, which covers antigen-binding fragment (Fab) determinants of FcRn affinity, transgenic preclinical models, and FcRn targeting as an immune-complex (IC)-clearing strategy. We further comment on therapeutic antibodies authorized for treating SARS-CoV-2 (bamlanivimab, casirivimab, and imdevimab) and evaluate their potential to saturate FcRn-mediated recycling. Finally, we discuss modeling and simulation studies that probe the quantitative relationship between in vivo IgG persistence and in vitro FcRn binding, emphasizing the importance of endosomal transit parameters.


Assuntos
Antígenos de Histocompatibilidade Classe I/química , Antígenos de Histocompatibilidade Classe I/metabolismo , Receptores Fc/química , Receptores Fc/metabolismo , Animais , Anticorpos Monoclonais/metabolismo , Anticorpos Monoclonais/farmacocinética , COVID-19/tratamento farmacológico , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Imunoglobulina G/imunologia , Imunoglobulina G/metabolismo , Receptores Fc/imunologia , Distribuição Tecidual/imunologia
4.
Front Immunol ; 12: 640093, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33717193

RESUMO

COVID-19 (SARS-CoV-2) disease severity and stages varies from asymptomatic, mild flu-like symptoms, moderate, severe, critical, and chronic disease. COVID-19 disease progression include lymphopenia, elevated proinflammatory cytokines and chemokines, accumulation of macrophages and neutrophils in lungs, immune dysregulation, cytokine storms, acute respiratory distress syndrome (ARDS), etc. Development of vaccines to severe acute respiratory syndrome (SARS), Middle East Respiratory Syndrome coronavirus (MERS-CoV), and other coronavirus has been difficult to create due to vaccine induced enhanced disease responses in animal models. Multiple betacoronaviruses including SARS-CoV-2 and SARS-CoV-1 expand cellular tropism by infecting some phagocytic cells (immature macrophages and dendritic cells) via antibody bound Fc receptor uptake of virus. Antibody-dependent enhancement (ADE) may be involved in the clinical observation of increased severity of symptoms associated with early high levels of SARS-CoV-2 antibodies in patients. Infants with multisystem inflammatory syndrome in children (MIS-C) associated with COVID-19 may also have ADE caused by maternally acquired SARS-CoV-2 antibodies bound to mast cells. ADE risks associated with SARS-CoV-2 has implications for COVID-19 and MIS-C treatments, B-cell vaccines, SARS-CoV-2 antibody therapy, and convalescent plasma therapy for patients. SARS-CoV-2 antibodies bound to mast cells may be involved in MIS-C and multisystem inflammatory syndrome in adults (MIS-A) following initial COVID-19 infection. SARS-CoV-2 antibodies bound to Fc receptors on macrophages and mast cells may represent two different mechanisms for ADE in patients. These two different ADE risks have possible implications for SARS-CoV-2 B-cell vaccines for subsets of populations based on age, cross-reactive antibodies, variabilities in antibody levels over time, and pregnancy. These models place increased emphasis on the importance of developing safe SARS-CoV-2 T cell vaccines that are not dependent upon antibodies.


Assuntos
Anticorpos Facilitadores , COVID-19/imunologia , Mastócitos/imunologia , Coronavírus da Síndrome Respiratória do Oriente Médio/fisiologia , Fagócitos/imunologia , SARS-CoV-2/fisiologia , Síndrome de Resposta Inflamatória Sistêmica/imunologia , Animais , Anticorpos Antivirais/metabolismo , COVID-19/transmissão , Criança , Reações Cruzadas , Feminino , Humanos , Recém-Nascido , Transmissão Vertical de Doenças Infecciosas , Modelos Imunológicos , Gravidez , Receptores Fc/metabolismo , Risco , Linfócitos T/imunologia
5.
Methods Mol Biol ; 2224: 123-132, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33606211

RESUMO

Proteinuria is a widely used marker of renal disease and is strongly associated with renal and cardiovascular outcomes. The molecular mechanisms underlying filtration of serum proteins through the glomerular filtration barrier (GFB) remain to be determined. Since the GFB is a complex structure, studies of albumin or IgG trafficking in cultured cells in vitro may not fully recapitulate these processes in vivo. In other epithelial cells including renal proximal tubular cells, the neonatal Fc receptor (FcRn) is required to divert albumin and IgG from the degradative pathway which allows these proteins to be recycled or transcytosed. To examine the role of podocyte FcRn in albumin and IgG trafficking in vivo, we detail the creation of a podocyte-specific FcRn knockout mouse and describe methods for examining intraglomerular detection of albumin and IgG in these mice.


Assuntos
Antígenos de Histocompatibilidade Classe I/metabolismo , Podócitos/metabolismo , Receptores Fc/metabolismo , Albuminas/metabolismo , Animais , Células Epiteliais/metabolismo , Feminino , Imunoglobulina G/metabolismo , Túbulos Renais Proximais/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transporte Proteico/fisiologia , Proteinúria/metabolismo , Transcitose/fisiologia
6.
PLoS Pathog ; 17(1): e1009252, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33513208

RESUMO

Neonatal echovirus infections are characterized by severe hepatitis and neurological complications that can be fatal. Here, we show that expression of the human homologue of the neonatal Fc receptor (hFcRn), the primary receptor for echoviruses, and ablation of type I interferon (IFN) signaling are key host determinants involved in echovirus pathogenesis. We show that expression of hFcRn alone is insufficient to confer susceptibility to echovirus infections in mice. However, expression of hFcRn in mice deficient in type I interferon (IFN) signaling, hFcRn-IFNAR-/-, recapitulate the echovirus pathogenesis observed in humans. Luminex-based multianalyte profiling from E11 infected hFcRn-IFNAR-/- mice revealed a robust systemic immune response to infection, including the induction of type I IFNs. Furthermore, similar to the severe hepatitis observed in humans, E11 infection in hFcRn-IFNAR-/- mice caused profound liver damage. Our findings define the host factors involved in echovirus pathogenesis and establish in vivo models that recapitulate echovirus disease in humans.


Assuntos
Enterovirus Humano B/patogenicidade , Infecções por Enterovirus/virologia , Genoma Viral/genética , Hepatite/virologia , Antígenos de Histocompatibilidade Classe I/metabolismo , Interferon Tipo I/metabolismo , Receptores Fc/metabolismo , Transdução de Sinais , Animais , Enterovirus Humano B/genética , Infecções por Enterovirus/imunologia , Feminino , Expressão Gênica , Hepatite/imunologia , Hepatócitos/imunologia , Hepatócitos/virologia , Antígenos de Histocompatibilidade Classe I/genética , Humanos , Imunidade , Fígado/imunologia , Fígado/virologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Receptores Fc/genética
7.
Biochem Biophys Res Commun ; 536: 32-37, 2021 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-33360096

RESUMO

The neonatal Fc receptor (FcRn) interacts with IgG and albumin at acidic pH within endosomes, thus protecting these plasma proteins from degradation. Recently, we proposed fibrinogen as a new binding partner of FcRn. This work was aimed at providing a direct demonstration of FcRn-fibrinogen binding at acidic pH by Fluorescence Correlation Spectroscopy. The increase in diffusion time between free and fibrinogen-bound FITC-labelled FcRn was assumed as the binding indicator. We observed that, at acidic pH (pH = 5.3), FcRn diffusion time shifted from ≈730 µs (FITC-labelled FcRn alone) to >1200 µs (FITC-labelled FcRn added with fibrinogen). A similar trend was exhibited by albumin, a known FcRn interactor, while no significant variations in diffusion time were observed upon incubation with catalase as negative control. Our results demonstrate a binding interaction between fibrinogen, one of the most abundant plasma proteins, and FcRn, a receptor involved in the regulation of the levels of IgG and albumin. This interaction is likely responsible for fibrinogen protection from intracellular degradation and recycling in plasma. Fibrinogen is crucial not only in haemostasis but also in acute inflammatory response and in some pathological conditions. The interaction with FcRn can influence not only the levels of fibrinogen in plasma and other tissues, but also the levels of other FcRn binding partners, among which are some plasma proteins of clinical relevance.


Assuntos
Fibrinogênio/metabolismo , Antígenos de Histocompatibilidade Classe I/metabolismo , Receptores Fc/metabolismo , Espectrometria de Fluorescência , Catalase/metabolismo , Difusão , Células HEK293 , Humanos , Concentração de Íons de Hidrogênio , Ligação Proteica , Albumina Sérica Humana/metabolismo
8.
PLoS One ; 15(12): e0230401, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33370294

RESUMO

Podocytes have been proposed to be antigen presenting cells (APCs). In traditional APCs, the neonatal Fc receptor (FcRn) is required for antigen presentation and global knockout of FcRn protects against glomerulonephritis. Since podocytes express FcRn, we sought to determine whether the absence of podocyte FcRn ameliorates immune-mediated disease. We examined MHCII and costimulatory markers expression in cultured wild type (WT) and FcRn knockout (KO) podocytes. Interferon gamma (IFNγ) induced MHCII expression in both WT and KO podocytes but did not change CD80 expression. Neither WT nor KO expressed CD86 or inducible costimulatory ligand (ICOSL) at baseline or with IFNγ. Using an antigen presentation assay, WT podocytes but not KO treated with immune complexes induced a modest increase in IL-2. Induction of the anti-glomerular basement membrane (anti-GBM) model resulted in a significant decrease in glomerular crescents in podocyte-specific FcRn knockout mouse (podFcRn KO) versus controls but the overall percentage of crescents was low. To examine the effects of the podocyte-specific FcRn knockout in a model with a longer autologous phase, we used the nephrotoxic serum nephritis (NTS) model. We found that the podFcRn KO mice had significantly reduced crescent formation and glomerulosclerosis compared to control mice. This study demonstrates that lack of podocyte FcRn is protective in immune mediated kidney disease that is dependent on an autologous phase. This study also highlights the difference between the anti-GBM model and NTS model of disease.


Assuntos
Glomerulonefrite/metabolismo , Antígenos de Histocompatibilidade Classe I/metabolismo , Podócitos/metabolismo , Receptores Fc/metabolismo , Animais , Células Cultivadas , Modelos Animais de Doenças , Citometria de Fluxo , Membrana Basal Glomerular/metabolismo , Glomerulonefrite/genética , Antígenos de Histocompatibilidade Classe I/genética , Camundongos , Camundongos Knockout , Receptores Fc/genética
9.
J Vis Exp ; (162)2020 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-32831311

RESUMO

The protocol describes how to set up and run a flow cytometry-based phagocytosis assay of Plasmodium falciparum-infected erythrocytes (IEs) opsonized by naturally acquired IgG antibodies specific for VAR2CSA. VAR2CSA is the parasite antigen that mediates the selective sequestration of IEs in the placenta that can cause a severe form of malaria in pregnant women, called placental malaria (PM). Protection from PM is mediated by VAR2CSA-specific antibodies that are believed to function by inhibiting placental sequestration and/or by opsonizing IEs for phagocytosis. The assay employs late-stage-synchronized IEs that have been selected in vitro to express VAR2CSA, plasma/serum-antibodies from women with naturally acquired PM-specific immunity, and the phagocytic cell line THP-1. However, the protocol can easily be modified to assay the functionality of antibodies to any parasite antigen present on the IE surface, whether induced by natural exposure or by vaccination. The assay offers simple and high-throughput evaluation, with good reproducibility, of an important functional aspect of antibody-mediated immunity in malaria. It is, therefore, useful when evaluating clinical immunity to P. falciparum malaria, a major cause of morbidity and mortality in the tropics, particularly in sub-Saharan Africa.


Assuntos
Anticorpos Antiprotozoários/análise , Bioensaio/métodos , Citometria de Fluxo/métodos , Parasitos/imunologia , Fagocitose , Plasmodium falciparum/imunologia , Animais , Antígenos de Protozoários/imunologia , Eritrócitos/parasitologia , Feminino , Humanos , Malária Falciparum/sangue , Malária Falciparum/parasitologia , Proteínas Opsonizantes/metabolismo , Gravidez , Receptores Fc/metabolismo , Reprodutibilidade dos Testes , Células THP-1
10.
Appl Environ Microbiol ; 86(17)2020 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-32591386

RESUMO

Staphylococcus aureus and other coagulase-positive Staphylococcus spp. bind the Fc region of IgG antibodies through expression of protein A (SpA). These species have consequently been a source of false-positive signals in antibody-based assays designed to detect other target bacteria. Here, flow cytometry was used to study the influence of a number of factors on the SpA-mediated binding of single cells to an anti-human IgG antibody, including strain, heat killing, overnight storage, growth phase, cell physiology, surface adhesion, and growth in model food systems. Through the costaining of antibody-stained cells with the permeability dye propidium iodide and calcein violet AM, the cell physiological status was related to SpA-mediated antibody binding. Generally, permeabilized cells lacking esterase activity did not strongly bind antibody. The binding of a number of commercially available polyclonal IgG antibodies to non-Staphylococcus spp. was also characterized. Not all SpA-expressing species showed strong binding of mouse IgG, and one species not known to express SpA showed strong binding. Most SpA-expressing strains bound rabbit IgG antibodies to some extent, whereas only one strain bound goat IgG. To reduce or eliminate SpA-mediated IgG binding, the following products were evaluated as blocking reagents and applied prior to staining with primary or secondary antibody: normal rabbit serum, mouse IgG isotype control, goat IgG, and a commercial FcR blocking reagent. Only the FcR blocking reagent consistently reduced SpA-mediated binding of Staphylococcus spp. to antibodies against other species and could be recommended as a blocking reagent in immunoassays designed to detect non-Staphylococcus species.IMPORTANCE This study characterizes a widespread but little-studied problem associated with the antibody-based detection of microbes-the Staphylococcus protein A (SpA)-mediated binding of IgG antibodies-and offers a solution: the use of commercial FcR blocking reagent. A common source of false-positive signals in the detection of microbes in clinical, food, or environmental samples can be eliminated by applying this study's findings. Using flow cytometry, the authors demonstrate the extent of heterogeneity in a culture's SpA-mediated binding of antibodies and that the degree of SpA-mediated antibody binding is strain, growth phase, and food matrix dependent and influenced by simulated food processing treatments and cell adherence. In addition, our studies of SpA-mediated binding of Staphylococcus spp. to antibodies against other bacterial species produced a very nuanced picture, leading us to recommend testing against multiple strains of S. aureus and S. hyicus of all antibodies to be incorporated into any immunoassay designed to detect a non-Staphylococcus spp.


Assuntos
Anticorpos Antibacterianos/imunologia , Receptores Fc/metabolismo , Proteína Estafilocócica A/imunologia , Staphylococcus/metabolismo , Anticorpos Antibacterianos/metabolismo , Citometria de Fluxo , Ligação Proteica , Proteína Estafilocócica A/metabolismo
11.
Am J Respir Crit Care Med ; 202(7): 1013-1023, 2020 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-32501729

RESUMO

Rationale: A subpopulation of B cells (age-associated B cells [ABCs]) is increased in mice and humans with infections or autoimmune diseases. Because depletion of these cells might be valuable in patients with certain lung diseases, the goal was to find out if ABC-like cells were at elevated levels in such patients.Objectives: To measure ABC-like cell percentages in patients with lung granulomatous diseases.Methods: Peripheral blood and BAL cells from patients with sarcoidosis, beryllium sensitivity, or hypersensitivity pneumonitis and healthy subjects were analyzed for the percentage of B cells that were ABC-like, defined by expression of CD11c, low levels of CD21, FcRL 1-5 (Fc receptor-like protein 1-5) expression, and, in some cases, T-bet.Measurements and Main Results: ABC-like cells in blood were at low percentages in healthy subjects and higher percentages in patients with sarcoidosis as well as at high percentages among BAL cells of patients with sarcoidosis, beryllium disease, and hypersensitivity pneumonitis. Treatment of patients with sarcoidosis led to reduced percentages of ABC-like cells in blood.Conclusions: Increased levels of ABC-like cells in patients with sarcoidosis may be useful in diagnosis. The increase in percentage of ABC-like cells in patients with lung granulomatous diseases and decrease in treated patients suggests that depletion of these cells may be valuable.


Assuntos
Alveolite Alérgica Extrínseca/sangue , Subpopulações de Linfócitos B/metabolismo , Beriliose/sangue , Líquido da Lavagem Broncoalveolar/citologia , Sarcoidose Pulmonar/sangue , Adulto , Idoso , Idoso de 80 Anos ou mais , Alveolite Alérgica Extrínseca/imunologia , Subpopulações de Linfócitos B/imunologia , Beriliose/imunologia , Antígeno CD11c/metabolismo , Estudos de Casos e Controles , Feminino , Humanos , Masculino , Proteínas de Membrana/metabolismo , Pessoa de Meia-Idade , Receptores de Superfície Celular/metabolismo , Receptores de Complemento 3d/metabolismo , Receptores Fc/metabolismo , Receptores Imunológicos/metabolismo , Sarcoidose Pulmonar/imunologia , Proteínas com Domínio T/metabolismo , Adulto Jovem
12.
Proc Natl Acad Sci U S A ; 117(23): 12943-12951, 2020 06 09.
Artigo em Inglês | MEDLINE | ID: mdl-32461366

RESUMO

The IgG Fc domain has the capacity to interact with diverse types of receptors, including the neonatal Fc receptor (FcRn) and Fcγ receptors (FcγRs), which confer pleiotropic biological activities. Whereas FcRn regulates IgG epithelial transport and recycling, Fc effector activities, such as antibody-dependent cellular cytotoxicity (ADCC) and phagocytosis, are mediated by FcγRs, which upon cross-linking transduce signals that modulate the function of effector leukocytes. Despite the well-defined and nonoverlapping functional properties of FcRn and FcγRs, recent studies have suggested that FcγRs mediate transplacental IgG transport, as certain Fc glycoforms were reported to be enriched in fetal circulation. To determine the contribution of FcγRs and FcRn to the maternal-fetal transport of IgG, we characterized the IgG Fc glycosylation in paired maternal-fetal samples from patient cohorts from Uganda and Nicaragua. No differences in IgG1 Fc glycan profiles and minimal differences in IgG2 Fc glycans were noted, whereas the presence or absence of galactose on the Fc glycan of IgG1 did not alter FcγRIIIa or FcRn binding, half-life, or their ability to deplete target cells in FcγR/FcRn humanized mice. Modeling maternal-fetal transport in FcγR/FcRn humanized mice confirmed that only FcRn contributed to transplacental transport of IgG; IgG selectively enhanced for FcRn binding resulted in enhanced accumulation of maternal antibody in the fetus. In contrast, enhancing FcγRIIIa binding did not result in enhanced maternal-fetal transport. These results argue against a role for FcγRs in IgG transplacental transport, suggesting Fc engineering of maternally administered antibody to enhance only FcRn binding as a means to improve maternal-fetal transport of IgG.


Assuntos
Sangue Fetal/imunologia , Antígenos de Histocompatibilidade Classe I/metabolismo , Imunoglobulina G/metabolismo , Troca Materno-Fetal/imunologia , Circulação Placentária/imunologia , Receptores Fc/metabolismo , Animais , Feminino , Antígenos de Histocompatibilidade Classe I/genética , Humanos , Imunoglobulina G/imunologia , Camundongos , Camundongos Transgênicos , Gravidez , Ensaios Clínicos Controlados Aleatórios como Assunto , Receptores Fc/genética , Receptores de IgG/genética , Receptores de IgG/metabolismo
13.
Curr Top Microbiol Immunol ; 429: 103-115, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32300915

RESUMO

Mycobacteria have unique lipids on their cell walls, and the structures and physiological activities of these lipid components have been the subject of many studies. Although the host receptors for mycobacterial lipid have long been elusive, in recent years C-type lectin receptors (CLRs) have been reported to recognize these components. The dendritic cell immunoactivating receptor (DCAR), a CLR member, is encoded by Clec4b1. DCAR, which was identified in 2003, is reported to be associated with the immunoreceptor tyrosine-based activation motif (ITAM)-containing adaptor protein, the Fc receptor γ chain (FcRγ). However, its physiological ligand and biological function were unknown. We recently identified DCAR as an activating receptor for mycobacteria. DCAR recognizes acylated phosphatidyl-inositol mannosides (PIMs) in mycobacteria to promote Th1 responses during mycobacterial infection. This review summarizes recent discoveries about the ligands and immunological roles of DCAR.


Assuntos
Mycobacterium , Proteínas Adaptadoras de Transdução de Sinal , Células Dendríticas/metabolismo , Lectinas Tipo C/genética , Lectinas Tipo C/metabolismo , Mycobacterium/metabolismo , Receptores Fc/metabolismo
14.
Biochem Biophys Res Commun ; 526(4): 941-946, 2020 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-32284170

RESUMO

Human serum albumin (HSA) has been used to extend the serum half-lives of various protein therapeutics through genetic fusion because HSA exhibits an exceptionally long circulation time as a result of neonatal Fc receptor (FcRn)-mediated recycling. As another serum half-life extender, the human antibody Fab SL335 that strongly binds HSA was developed. When SL335 was fused to a protein therapeutic, SL335 was shown to prolong the half-life of the drug. Despite the significance of SL335-HSA binding in the extension of drug circulation time, it remains unclear how SL335 interacts with HSA at a molecular structural level. To reveal the structural basis of HSA recognition by SL335, we determined the crystal structure of the SL335-HSA complex at a resolution of 2.95 Å. SL335 binds HSA at a 1:1 stoichiometry. SL335 uses the exposed loops of its heavy and light chains to specifically recognize the IIa and IIb subdomains of HSA. The SL335 epitope is located on the opposite side of the FcRn-binding site and does not overlap with it, suggesting that SL335 extends the serum half-lives of itself and its fusion partner through an FcRn-dependent recycling mechanism.


Assuntos
Anticorpos/imunologia , Fragmentos Fab das Imunoglobulinas/imunologia , Proteínas Recombinantes/sangue , Proteínas Recombinantes/uso terapêutico , Albumina Sérica Humana/química , Albumina Sérica Humana/imunologia , Anticorpos/química , Reações Cruzadas , Meia-Vida , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Fragmentos Fab das Imunoglobulinas/química , Modelos Moleculares , Ligação Proteica , Receptores Fc/metabolismo
15.
J Immunol Methods ; 480: 112767, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32119889

RESUMO

IgG antibodies have been used to treat many diseases including cancer. IgG antibody-drug conjugates (ADCs) deliver cytotoxic drugs to target cells for cell elimination, but they have dose limiting toxicity due to target-independent uptake, including pinocytotic uptake. Neonatal Fc receptor (FcRn) recycles pinocytosed IgG in a pH-dependent manner and is the receptor responsible for the long half-life of IgG. Use of IgG variants with stronger FcRn binding at pH 6.0 for ADCs might improve recycling efficiency and reduce toxicity. However, these variants have residual FcRn binding at pH 7.4, which could lead to FcRn-mediated uptake and higher toxicity. Thus, the uptake of such variants at pH 7.4 needs to be evaluated. Here we report a reproducible and quantitative assay using an inducible HM7 colorectal cancer cell line to measure IgG uptake at endogenous and overexpressed FcRn levels. Our assay had comparable reproducibility at pH 6.0, 6.8 and 7.4. The wild type (WT) IgG had similar uptake at endogenous and overexpressed FcRn levels, as expected for pinocytotic uptake. We found similar uptake of a WT IgG and a stronger FcRn binding T307Q/N434A variant (QA variant) at endogenous FcRn levels at pH 7.4, although the QA variant had higher uptake at overexpressed FcRn levels. The QA variant also had higher uptake than the WT IgG at overexpressed FcRn levels at pH 6.8. Our assay can be used to characterize the stronger FcRn binding variants to aid in selection of suitable variants with low uptake at pH 7.4 for use as ADCs.


Assuntos
Neoplasias Colorretais/metabolismo , Antígenos de Histocompatibilidade Classe I/metabolismo , Imunoglobulina G/metabolismo , Pinocitose , Receptores Fc/metabolismo , Linhagem Celular Tumoral , Neoplasias Colorretais/genética , Antígenos de Histocompatibilidade Classe I/genética , Humanos , Concentração de Íons de Hidrogênio , Cinética , Ligação Proteica , Receptores Fc/genética , Regulação para Cima
16.
Sci Rep ; 10(1): 3685, 2020 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-32111886

RESUMO

The blood-brain barrier (BBB) hinders the brain delivery of therapeutic immunoglobulin γ (IgG) antibodies. Evidence suggests that IgG-specific processing occurs within the endothelium of the BBB, but any influence on transcytosis remains unclear. Here, involvement of the neonatal Fc receptor (FcRn), which mediates IgG recycling and transcytosis in peripheral endothelium, was investigated by evaluating the transcytosis of IgGs with native or reduced FcRn engagement across human induced pluripotent stem cell-derived brain endothelial-like cells. Despite differential trafficking, the permeability of all tested IgGs were comparable and remained constant irrespective of concentration or competition with excess IgG, suggesting IgG transcytosis occurs nonspecifically and originates from fluid-phase endocytosis. Comparison with the receptor-enhanced permeability of transferrin indicates that the phenomena observed for IgG is ubiquitous for most macromolecules. However, increased permeability was observed for macromolecules with biophysical properties known to engage alternative endocytosis mechanisms, highlighting the importance of biophysical characterizations in assessing transcytosis mechanisms.


Assuntos
Barreira Hematoencefálica/metabolismo , Células Endoteliais/metabolismo , Antígenos de Histocompatibilidade Classe I/metabolismo , Imunoglobulina G/metabolismo , Receptores Fc/metabolismo , Transcitose , Animais , Barreira Hematoencefálica/citologia , Linhagem Celular , Células Endoteliais/citologia , Humanos , Ratos
17.
Immunol Cell Biol ; 98(4): 305-317, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32142167

RESUMO

Antibody-dependent complement activity is associated not only with autoimmune morbidity, but also with antitumor efficacy. In infectious disease, both recombinant monoclonal antibodies and polyclonal antibodies generated in natural adaptive responses can mediate complement activity to protective, therapeutic or disease-enhancing effect. Recent advances have contributed to the structural resolution of molecular complexes involved in antibody-mediated complement activation, defining the avid nature of participating interactions and pointing to how antibody isotype, subclass, hinge flexibility, glycosylation state, amino acid sequence and the contextual nature of the cognate antigen/epitope are all factors that can determine complement activity through impact on antibody multimerization and subsequent recruitment of complement component 1q. Beyond the efficiency of activation, complement activation products interact with various cell types that mediate immune adherence, trafficking, immune education and innate functions. Similarly, depending on the anatomical location and extent of activation, complement can support homeostatic restoration or be leveraged by pathogens or neoplasms to enhance infection or promote tumorigenic microenvironments, respectively. Advances in means to suppress complement activation by intravenous immunoglobulin (IVIG), IVIG mimetics and complement-intervening antibodies represent proven and promising exploratory therapeutic strategies, while antibody engineering has likewise offered frameworks to enhance, eliminate or isolate complement activation to interrogate in vivo mechanisms of action. Such strategies promise to support the optimization of antibody-based drugs that are able to tackle emerging and difficult-to-treat diseases by improving our understanding of the synergistic and antagonistic relationships between antibody mechanisms mediated by Fc receptors, direct binding and the products of complement activation.


Assuntos
Anticorpos Monoclonais/imunologia , Doenças Transmissíveis/imunologia , Ativação do Complemento/imunologia , Proteínas do Sistema Complemento/imunologia , Neoplasias/imunologia , Animais , Anticorpos Monoclonais/efeitos adversos , Anticorpos Monoclonais/metabolismo , Antígenos/imunologia , Antígenos/metabolismo , Autoanticorpos/efeitos adversos , Autoanticorpos/imunologia , Engenharia Biomédica , Doenças Transmissíveis/microbiologia , Doenças Transmissíveis/patologia , Doenças Transmissíveis/virologia , Complemento C1q/química , Complemento C1q/imunologia , Complemento C1q/metabolismo , Proteínas do Sistema Complemento/química , Proteínas do Sistema Complemento/metabolismo , Humanos , Fragmentos Fc das Imunoglobulinas/imunologia , Imunoglobulina G/química , Imunoglobulina G/imunologia , Imunoglobulina G/ultraestrutura , Neoplasias/patologia , Receptores Fc/imunologia , Receptores Fc/metabolismo
18.
Mol Immunol ; 121: 1-6, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32135400

RESUMO

The transglutaminase 2 (TG2) is one of the enigmatic enzymes with important functional diversity. It plays an important role in several pathologies such as celiac disease (CD). In patients with active CD, the abnormal retrotranscytosis of IgA/gliadin complexes is mediated by Transferrin Receptor 1 (TfR1). This triad association takes also place in IgA nephropathy (IgA-N). IgA-N is characterized by the formation of nephrotoxic complexes of IgA1 and soluble CD89 (sCD89). These complexes are abnormally deposited in the kidney. Using a humanized mouse model of IgA-N (α1KI-CD89Tg), we showed that IgA1-sCD89 complexes engender mesangial cell activation and proliferation with TfR1 and TG2 up-regulation, associated with IgA-N features. This TG2-TfR1 interaction enhances mesangial IgA1 deposition promoting inflammation. Humanized α1KI-CD89Tg mice deficient for TG2 show a decrease in TfR1 expression in kidney leading to reduced IgA1-sCD89 deposits and an improvement in IgA-N features. Moreover, TG2 is active and overexpressed in the intestine of IgA-N mice and gliadin participates to this renal pathology. In kidney as in intestine, the TG2 has a crucial role in the cooperation between TfR1-IgA and a central role in the pathogenic amplification.


Assuntos
Antígenos CD/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Gliadina/imunologia , Glomerulonefrite por IGA/imunologia , Imunoglobulina A/metabolismo , Receptores da Transferrina/metabolismo , Transglutaminases/metabolismo , Animais , Antígenos CD/genética , Antígenos CD/imunologia , Doença Celíaca/imunologia , Doença Celíaca/patologia , Modelos Animais de Doenças , Proteínas de Ligação ao GTP/genética , Proteínas de Ligação ao GTP/imunologia , Gliadina/metabolismo , Humanos , Imunoglobulina A/imunologia , Mucosa Intestinal/imunologia , Mucosa Intestinal/patologia , Células Mesangiais/imunologia , Células Mesangiais/patologia , Camundongos , Camundongos Transgênicos , Receptores Fc/genética , Receptores Fc/imunologia , Receptores Fc/metabolismo , Receptores da Transferrina/imunologia , Transglutaminases/genética , Transglutaminases/imunologia , Regulação para Cima/imunologia
19.
J Immunol ; 204(4): 954-966, 2020 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-31915259

RESUMO

Neutrophils are critical to the generation of effective immune responses and for killing invading microbes. Paired immune receptors provide important mechanisms to modulate neutrophil activation thresholds and effector functions. Expression of the leukocyte Ig-like receptor (LILR)A6 (ILT8/CD85b) and LILRB3 (ILT5/CD85a) paired-receptor system on human neutrophils has remained unclear because of the lack of specific molecular tools. Additionally, there is little known of their possible functions in neutrophil biology. The objective of this study was to characterize expression of LILRA6/LILRB3 receptors during human neutrophil differentiation and activation, and to assess their roles in modulating Fc receptor-mediated effector functions. LILRB3, but not LILRA6, was detected in human neutrophil lysates following immunoprecipitation by mass spectrometry. We demonstrate high LILRB3 expression on the surface of resting neutrophils and release from the surface following neutrophil activation. Surface expression was recapitulated in a human PLB-985 cell model of neutrophil-like differentiation. Continuous ligation of LILRB3 inhibited key IgA-mediated effector functions, including production of reactive oxygen species, phagocytic uptake, and microbial killing. This suggests that LILRB3 provides an important checkpoint to control human neutrophil activation and their antimicrobial effector functions during resting and early-activation stages of the neutrophil life cycle.


Assuntos
Antígenos CD/metabolismo , Neutrófilos/imunologia , Receptores Fc/metabolismo , Receptores Imunológicos/metabolismo , Infecções Estafilocócicas/imunologia , Antígenos CD/genética , Antígenos CD/isolamento & purificação , Diferenciação Celular/imunologia , Linhagem Celular , Regulação para Baixo/imunologia , Humanos , Ativação de Neutrófilo , Neutrófilos/metabolismo , Fagocitose , Cultura Primária de Células , Espécies Reativas de Oxigênio/metabolismo , Receptores Imunológicos/genética , Receptores Imunológicos/isolamento & purificação , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Infecções Estafilocócicas/microbiologia , Staphylococcus capitis/imunologia
20.
Nature ; 577(7791): 543-548, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31915378

RESUMO

Although maternal antibodies protect newborn babies from infection1,2, little is known about how protective antibodies are induced without prior pathogen exposure. Here we show that neonatal mice that lack the capacity to produce IgG are protected from infection with the enteric pathogen enterotoxigenic Escherichia coli by maternal natural IgG antibodies against the maternal microbiota when antibodies are delivered either across the placenta or through breast milk. By challenging pups that were fostered by either maternal antibody-sufficient or antibody-deficient dams, we found that IgG derived from breast milk was crucial for protection against mucosal disease induced by enterotoxigenic E. coli. IgG also provides protection against systemic infection by E. coli. Pups used the neonatal Fc receptor to transfer IgG from milk into serum. The maternal commensal microbiota can induce antibodies that recognize antigens expressed by enterotoxigenic E. coli and other Enterobacteriaceae species. Induction of maternal antibodies against a commensal Pantoea species confers protection against enterotoxigenic E. coli in pups. This role of the microbiota in eliciting protective antibodies to a specific neonatal pathogen represents an important host defence mechanism against infection in neonates.


Assuntos
Anticorpos/imunologia , Escherichia coli Enterotoxigênica/imunologia , Infecções por Escherichia coli/imunologia , Infecções por Escherichia coli/prevenção & controle , Imunidade Materno-Adquirida/imunologia , Recém-Nascido/imunologia , Microbiota/imunologia , Leite Humano/imunologia , Animais , Anticorpos/sangue , Anticorpos/metabolismo , Aleitamento Materno , Reações Cruzadas/imunologia , Infecções por Escherichia coli/microbiologia , Feminino , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Imunoglobulina G/metabolismo , Masculino , Camundongos , Mães , Pantoea/imunologia , Receptores Fc/imunologia , Receptores Fc/metabolismo , Simbiose/imunologia
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