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1.
Blood ; 138(16): 1481-1489, 2021 10 21.
Article in English | MEDLINE | ID: covidwho-1484294

ABSTRACT

A subset of patients with coronavirus disease 2019 (COVID-19) become critically ill, suffering from severe respiratory problems and also increased rates of thrombosis. The causes of thrombosis in severely ill patients with COVID-19 are still emerging, but the coincidence of critical illness with the timing of the onset of adaptive immunity could implicate an excessive immune response. We hypothesized that platelets might be susceptible to activation by anti-severe acute respiratory syndrome coronavirus 2 (anti-SARS-CoV-2) antibodies and might contribute to thrombosis. We found that immune complexes containing recombinant SARS-CoV-2 spike protein and anti-spike immunoglobulin G enhanced platelet-mediated thrombosis on von Willebrand factor in vitro, but only when the glycosylation state of the Fc domain was modified to correspond with the aberrant glycosylation previously identified in patients with severe COVID-19. Furthermore, we found that activation was dependent on FcγRIIA, and we provide in vitro evidence that this pathogenic platelet activation can be counteracted by the therapeutic small molecules R406 (fostamatinib) and ibrutinib, which inhibit tyrosine kinases Syk and Btk, respectively, or by the P2Y12 antagonist cangrelor.


Subject(s)
Blood Platelets/pathology , COVID-19/complications , Immunoglobulin G/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/metabolism , Thrombosis/pathology , von Willebrand Factor/metabolism , Antibodies, Viral/blood , Antibodies, Viral/immunology , Antigen-Antibody Complex/immunology , Blood Platelets/immunology , Blood Platelets/metabolism , COVID-19/immunology , COVID-19/virology , Glycosylation , Humans , Platelet Activation/immunology , Thrombosis/immunology , Thrombosis/virology , von Willebrand Factor/genetics
2.
Blood ; 138(22): 2256-2268, 2021 12 02.
Article in English | MEDLINE | ID: covidwho-1443788

ABSTRACT

SARS-CoV-2 vaccine ChAdOx1 nCoV-19 (AstraZeneca) causes a thromboembolic complication termed vaccine-induced immune thrombotic thrombocytopenia (VITT). Using biophysical techniques, mouse models, and analysis of VITT patient samples, we identified determinants of this vaccine-induced adverse reaction. Super-resolution microscopy visualized vaccine components forming antigenic complexes with platelet factor 4 (PF4) on platelet surfaces to which anti-PF4 antibodies obtained from VITT patients bound. PF4/vaccine complex formation was charge-driven and increased by addition of DNA. Proteomics identified substantial amounts of virus production-derived T-REx HEK293 proteins in the ethylenediaminetetraacetic acid (EDTA)-containing vaccine. Injected vaccine increased vascular leakage in mice, leading to systemic dissemination of vaccine components known to stimulate immune responses. Together, PF4/vaccine complex formation and the vaccine-stimulated proinflammatory milieu trigger a pronounced B-cell response that results in the formation of high-avidity anti-PF4 antibodies in VITT patients. The resulting high-titer anti-PF4 antibodies potently activated platelets in the presence of PF4 or DNA and polyphosphate polyanions. Anti-PF4 VITT patient antibodies also stimulated neutrophils to release neutrophil extracellular traps (NETs) in a platelet PF4-dependent manner. Biomarkers of procoagulant NETs were elevated in VITT patient serum, and NETs were visualized in abundance by immunohistochemistry in cerebral vein thrombi obtained from VITT patients. Together, vaccine-induced PF4/adenovirus aggregates and proinflammatory reactions stimulate pathologic anti-PF4 antibody production that drives thrombosis in VITT. The data support a 2-step mechanism underlying VITT that resembles the pathogenesis of (autoimmune) heparin-induced thrombocytopenia.


Subject(s)
Antigen-Antibody Complex/immunology , Autoantibodies/immunology , COVID-19/prevention & control , Capsid Proteins/adverse effects , Drug Contamination , Genetic Vectors/adverse effects , HEK293 Cells/immunology , Immunoglobulin G/immunology , Platelet Factor 4/immunology , Purpura, Thrombocytopenic, Idiopathic/etiology , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/adverse effects , Adenoviridae/immunology , Animals , Antigen-Antibody Complex/ultrastructure , Autoantibodies/biosynthesis , Capillary Leak Syndrome/etiology , Capsid Proteins/immunology , Cell Line, Transformed , /immunology , Dynamic Light Scattering , Epitopes/chemistry , Epitopes/immunology , Extracellular Traps/immunology , Extravasation of Diagnostic and Therapeutic Materials/etiology , Genetic Vectors/immunology , HEK293 Cells/chemistry , Humans , Imaging, Three-Dimensional , Immunoglobulin G/biosynthesis , Inflammation , Mice , Microscopy/methods , Platelet Activation , Proteomics , Purpura, Thrombocytopenic, Idiopathic/blood , Purpura, Thrombocytopenic, Idiopathic/immunology , Sinus Thrombosis, Intracranial/diagnostic imaging , Sinus Thrombosis, Intracranial/immunology , Spike Glycoprotein, Coronavirus/immunology , Virus Cultivation
3.
J Infect Dis ; 224(4): 575-585, 2021 08 16.
Article in English | MEDLINE | ID: covidwho-1358459

ABSTRACT

Severe coronavirus disease 2019 (COVID-19) is associated with an overactive inflammatory response mediated by macrophages. Here, we analyzed the phenotype and function of neutrophils in patients with COVID-19. We found that neutrophils from patients with severe COVID-19 express high levels of CD11b and CD66b, spontaneously produce CXCL8 and CCL2, and show a strong association with platelets. Production of CXCL8 correlated with plasma concentrations of lactate dehydrogenase and D-dimer. Whole blood assays revealed that neutrophils from patients with severe COVID-19 show a clear association with immunoglobulin G (IgG) immune complexes. Moreover, we found that sera from patients with severe disease contain high levels of immune complexes and activate neutrophils through a mechanism partially dependent on FcγRII (CD32). Interestingly, when integrated in immune complexes, anti-severe acute respiratory syndrome coronavirus 2 IgG antibodies from patients with severe COVID-19 displayed a higher proinflammatory profile compared with antibodies from patients with mild disease. Our study suggests that IgG immune complexes might promote the acquisition of an inflammatory signature by neutrophils, worsening the course of COVID-19.


Subject(s)
Antibodies, Viral/immunology , Antigen-Antibody Complex/immunology , COVID-19/immunology , Immunoglobulin G/immunology , Neutrophil Activation/immunology , Adult , Aged , Antibodies, Viral/blood , Antigen-Antibody Complex/blood , Antigens, CD/immunology , CD11b Antigen/immunology , Cell Adhesion Molecules/immunology , Female , GPI-Linked Proteins/immunology , Humans , Immunoglobulin G/blood , Interleukin-8/immunology , Male , Middle Aged , Neutrophils/immunology , Receptors, IgG/immunology , SARS-CoV-2/immunology , Young Adult
4.
Int J Mol Sci ; 22(13)2021 Jun 22.
Article in English | MEDLINE | ID: covidwho-1304657

ABSTRACT

The innate immune system's natural killer (NK) cells exert their cytolytic function against a variety of pathological challenges, including tumors and virally infected cells. Their activation depends on net signaling mediated via inhibitory and activating receptors that interact with specific ligands displayed on the surfaces of target cells. The CD94/NKG2C heterodimer is one of the NK activating receptors and performs its function by interacting with the trimeric ligand comprised of the HLA-E/ß2m/nonameric peptide complex. Here, simulations of the all-atom multi-microsecond molecular dynamics in five immune complexes provide atomistic insights into the receptor-ligand molecular recognition, as well as the molecular events that facilitate the NK cell activation. We identify NKG2C, the HLA-Eα2 domain, and the nonameric peptide as the key elements involved in the molecular machinery of signal transduction via an intertwined hydrogen bond network. Overall, the study addresses the complex intricacies that are necessary to understand the mechanisms of the innate immune system.


Subject(s)
Antigen-Antibody Complex/chemistry , Histocompatibility Antigens Class I/chemistry , Models, Molecular , NK Cell Lectin-Like Receptor Subfamily C/chemistry , NK Cell Lectin-Like Receptor Subfamily D/chemistry , Peptides/chemistry , Amino Acid Sequence , Antigen-Antibody Complex/immunology , Antigen-Antibody Complex/metabolism , Binding Sites , Histocompatibility Antigens Class I/immunology , Histocompatibility Antigens Class I/metabolism , Humans , Hydrogen Bonding , Ligands , NK Cell Lectin-Like Receptor Subfamily C/metabolism , NK Cell Lectin-Like Receptor Subfamily D/metabolism , Peptides/metabolism , Protein Binding , Protein Conformation , Protein Interaction Domains and Motifs , Signal Transduction , Structure-Activity Relationship
5.
Proc Natl Acad Sci U S A ; 118(27)2021 07 06.
Article in English | MEDLINE | ID: covidwho-1285962

ABSTRACT

IgA is the second most abundant antibody present in circulation and is enriched at mucosal surfaces. As such, IgA plays a key role in protection against a variety of mucosal pathogens including viruses. In addition to neutralizing viruses directly, IgA can also stimulate Fc-dependent effector functions via engagement of Fc alpha receptors (Fc-αRI) expressed on the surface of certain immune effector cells. Neutrophils are the most abundant leukocyte, express Fc-αRI, and are often the first to respond to sites of injury and infection. Here, we describe a function for IgA-virus immune complexes (ICs) during viral infections. We show that IgA-virus ICs potentiate NETosis-the programmed cell-death pathway through which neutrophils release neutrophil extracellular traps (NETs). Mechanistically, IgA-virus ICs potentiated a suicidal NETosis pathway via engagement of Fc-αRI on neutrophils through a toll-like receptor-independent, NADPH oxidase complex-dependent pathway. NETs also were capable of trapping and inactivating viruses, consistent with an antiviral function.


Subject(s)
Extracellular Traps/immunology , Immunoglobulin A/immunology , Neutrophils/immunology , Virus Diseases/immunology , Antigen-Antibody Complex/immunology , Antigens, CD/metabolism , Extracellular Traps/virology , Humans , Influenzavirus A/immunology , NADPH Oxidases/metabolism , Neutrophils/pathology , Neutrophils/virology , Receptors, Fc/metabolism , SARS-CoV-2/immunology , Signal Transduction , Virion
6.
J Infect Dis ; 224(4): 575-585, 2021 08 16.
Article in English | MEDLINE | ID: covidwho-1169670

ABSTRACT

Severe coronavirus disease 2019 (COVID-19) is associated with an overactive inflammatory response mediated by macrophages. Here, we analyzed the phenotype and function of neutrophils in patients with COVID-19. We found that neutrophils from patients with severe COVID-19 express high levels of CD11b and CD66b, spontaneously produce CXCL8 and CCL2, and show a strong association with platelets. Production of CXCL8 correlated with plasma concentrations of lactate dehydrogenase and D-dimer. Whole blood assays revealed that neutrophils from patients with severe COVID-19 show a clear association with immunoglobulin G (IgG) immune complexes. Moreover, we found that sera from patients with severe disease contain high levels of immune complexes and activate neutrophils through a mechanism partially dependent on FcγRII (CD32). Interestingly, when integrated in immune complexes, anti-severe acute respiratory syndrome coronavirus 2 IgG antibodies from patients with severe COVID-19 displayed a higher proinflammatory profile compared with antibodies from patients with mild disease. Our study suggests that IgG immune complexes might promote the acquisition of an inflammatory signature by neutrophils, worsening the course of COVID-19.


Subject(s)
Antibodies, Viral/immunology , Antigen-Antibody Complex/immunology , COVID-19/immunology , Immunoglobulin G/immunology , Neutrophil Activation/immunology , Adult , Aged , Antibodies, Viral/blood , Antigen-Antibody Complex/blood , Antigens, CD/immunology , CD11b Antigen/immunology , Cell Adhesion Molecules/immunology , Female , GPI-Linked Proteins/immunology , Humans , Immunoglobulin G/blood , Interleukin-8/immunology , Male , Middle Aged , Neutrophils/immunology , Receptors, IgG/immunology , SARS-CoV-2/immunology , Young Adult
7.
Mol Med Rep ; 22(6): 4485-4491, 2020 Dec.
Article in English | MEDLINE | ID: covidwho-979150

ABSTRACT

In December 2019, an emergence of pneumonia was detected in patients infected with a novel coronavirus (CoV) in Wuhan (Hubei, China). The International Committee on Taxonomy of Viruses named the virus severe acute respiratory syndrome­CoV­2 and the disease CoV disease­19 (COVID­19). Patients with COVID­19 present with symptoms associated with respiratory system dysfunction and hematological changes, including lymphopenia, thrombocytopenia and coagulation disorders. However, to the best of our knowledge, the pathogenesis of COVID­19 remains unclear. Therefore, understanding the mechanisms underlying the hematological changes that manifest during COVID­19 may aid in the development of treatments and may improve patient prognosis.


Subject(s)
Betacoronavirus , Coronavirus Infections/blood , Pneumonia, Viral/blood , Antibodies, Viral/immunology , Antigen-Antibody Complex/immunology , Antiviral Agents/pharmacology , Antiviral Agents/therapeutic use , Betacoronavirus/immunology , COVID-19 , Cellular Microenvironment , Coronavirus Infections/complications , Coronavirus Infections/drug therapy , Coronavirus Infections/therapy , Cytokine Release Syndrome/blood , Cytokine Release Syndrome/etiology , Cytokine Release Syndrome/prevention & control , Cytokines/blood , Diagnostic Tests, Routine , Endothelium, Vascular/pathology , Hematologic Tests , Hematopoiesis/drug effects , Hematopoietic Stem Cells/pathology , Humans , Hypoalbuminemia/etiology , Liver/physiopathology , Lung/physiopathology , Lymphopenia/etiology , Lymphopenia/physiopathology , Pandemics , Pneumonia, Viral/complications , Pneumonia, Viral/drug therapy , Pneumonia, Viral/therapy , Reperfusion Injury/etiology , SARS-CoV-2 , Thrombocytopenia/etiology , Thrombocytopenia/physiopathology , Thrombophilia/etiology
8.
Front Immunol ; 11: 566000, 2020.
Article in English | MEDLINE | ID: covidwho-922794

ABSTRACT

Because of particular properties of SARS-Cov-2, such as an high infection speed, its antigenic nature, evolutionarily unknown to the human immune system, and/or a viral interference on the immune response mechanisms, this virus would determine in the subjects a delayed anomalous (slow and/or low) immune response, ineffective and, finally, self-damaging. The hypothetical pathogenetic process for covid-19 could occur in three phases: a) Viral phase, asymptomatic or weakly symptomatic, with an a-specific innate immune response; b) Immunological phase, intermediately symptomatic, with an anomalous specific immune response (delayed, slow and/or low synthesis of IgM and IgG) in antigen excess conditions, immune complex formation and complement activation with tissue damages; c) Hemo-vascular phase, severely symptomatic, where complement-mediated tissue damages would induce vascular inflammation and systemic alteration of the coagulation homeostasis. This hypothesis is well supported by the immune-histochemical and microscopic demonstration in severe patient lungs of co-localized spike viral proteins, terminal components of the activated complement system (C5b-9 membrane attack complex) and microvascular deposits of small fibrin thrombi. This picture could be aggravated by the involvement of neutrophils and macrophages, releasing additional lytic and inflammatory factors. Thus, covid-19 would arise as a simple viral infection, develop as a diffuse immune complex hypersensitivity and explode as a systemic hemo-vascular pathology. If this hypothesized process would be real, suitable therapeutic interventions might be carried out, able to interfere with or block the critical factors in the various phases.


Subject(s)
Antigen-Antibody Complex/immunology , COVID-19/therapy , SARS-CoV-2/physiology , COVID-19/immunology , Complement Activation , Humans , Immunity, Innate , Pandemics , SARS-CoV-2/genetics
9.
Sci Rep ; 10(1): 16219, 2020 10 01.
Article in English | MEDLINE | ID: covidwho-811544

ABSTRACT

COVID-19 pandemic has resulted in 16,114,449 cases with 646,641 deaths from the 217 countries, or territories as on July 27th 2020. Due to multifaceted issues and challenges in the implementation of the safety and preventive measures, inconsistent coordination between societies-governments and most importantly lack of specific vaccine to SARS-CoV-2, the spread of the virus that initially emerged at Wuhan is still uprising after taking a heavy toll on human life. In the present study, we mapped immunogenic epitopes present on the four structural proteins of SARS-CoV-2 and we designed a multi-epitope peptide based vaccine that, demonstrated a high immunogenic response with a vast application on world's human population. On codon optimization and in-silico cloning, we found that candidate vaccine showed high expression in E. coli and immune simulation resulted in inducing a high level of both B-cell and T-cell mediated immunity. The results predicted that exposure of vaccine by administrating three injections significantly subsidized the antigen growth in the system. The proposed candidate vaccine found promising by yielding desired results and hence, should be validated by practical experimentations for its functioning and efficacy to neutralize SARS-CoV-2.


Subject(s)
Epitopes/immunology , Molecular Docking Simulation , Vaccines, Subunit/immunology , Viral Vaccines/immunology , Antigen-Antibody Complex/chemistry , Antigen-Antibody Complex/immunology , Antigens, Viral/immunology , B-Lymphocytes/immunology , COVID-19 Vaccines , Coronavirus Infections/immunology , Coronavirus Infections/prevention & control , Coronavirus Nucleocapsid Proteins , Epitopes/chemistry , HLA Antigens/chemistry , HLA Antigens/immunology , Humans , Immunogenicity, Vaccine , Nucleocapsid Proteins/chemistry , Nucleocapsid Proteins/immunology , Phosphoproteins , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/immunology , T-Lymphocytes/immunology , Toll-Like Receptors/immunology , Vaccines, Subunit/chemistry , Viral Matrix Proteins/chemistry , Viral Matrix Proteins/immunology , Viral Vaccines/chemistry
10.
Paediatr Respir Rev ; 35: 81-87, 2020 Sep.
Article in English | MEDLINE | ID: covidwho-680602

ABSTRACT

The rapid spread of SARS-CoV-2 infection globally coupled with the relatively high case-fatality rate has led to immediate need for therapeutic intervention to prevent and treat COVID-19 disease. There is accumulating evidence that morbidity and mortality in COVID-19 may be exacerbated by a dysregulated host immune response resulting in significant hyperinflammation and cytokine release. The aim of this review is to describe the basis for the immune dysregulation caused by SARS-CoV-2 infection and to examine current investigations into immunomodulatory therapies aimed at targeting the excessive host immune response.


Subject(s)
Coronavirus Infections/immunology , Cytokine Release Syndrome/immunology , Immunologic Factors/therapeutic use , Pneumonia, Viral/immunology , Systemic Inflammatory Response Syndrome/immunology , Adrenal Cortex Hormones/therapeutic use , Antibodies, Monoclonal, Humanized/therapeutic use , Antigen-Antibody Complex/immunology , Betacoronavirus , COVID-19 , Child , Coronavirus Infections/drug therapy , Coronavirus Infections/physiopathology , Coronavirus Infections/therapy , Cytokine Release Syndrome/physiopathology , Cytokine Release Syndrome/therapy , Humans , Immunization, Passive/methods , Immunoglobulins, Intravenous/therapeutic use , Inflammation/immunology , Inflammation/physiopathology , Inflammation/therapy , Interleukin 1 Receptor Antagonist Protein/therapeutic use , Janus Kinase Inhibitors/therapeutic use , Pandemics , Pneumonia, Viral/physiopathology , Pneumonia, Viral/therapy , Receptors, Interleukin-6/antagonists & inhibitors , SARS-CoV-2 , Systemic Inflammatory Response Syndrome/physiopathology , Systemic Inflammatory Response Syndrome/therapy
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