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Free labile hemin acts as a damage-associated molecular pattern during acute and chronic hemolysis and muscle injury, supporting platelet activation and thrombosis.

Transplacental delivery of maternal immunoglobulin G (IgG) provides humoral protection during the first months of life until the newborn’s immune system reaches maturity. The maternofetal interface has been exploited therapeutically to replace missing enzymes in the fetus, as shown in experimental mucopolysaccharidoses, or to shape adaptive immune repertoires during fetal development and induce tolerance to self-antigens or immunogenic therapeutic molecules.

Independent of etiology, hemolytic diseases are associated with thrombosis, inflammation and immune dysregulation, all together contributing to organ damage and poor outcome. Beyond anemia and the loss of the anti-inflammatory functions of red blood cells, hemolysis leads to the release of damage-associated molecular patterns including ADP, hemoglobin, and heme, which act through multiple receptors and signaling pathways fostering a hyperinflammatory and hypercoagulable state. Extracellular free heme is promiscuous alarmin capable of triggering oxido-inflammatory and thrombotic events by inducing the activation of platelets, endothelial and innate cells as well as the coagulation and complement cascades. In this review, we discuss the main mechanisms by which hemolysis and, in particular, heme, drive this thrombo-inflammatory milieu and discuss the consequences of hemolysis on the host response to secondary infections.

Antibodies are essential components of adaptive immunity. A typical antibody repertoire comprises an enormous diversity of antigen-binding specificities, which are generated by the genetic processes of recombination and mutation. Accumulating evidence suggests that the immune system can exploit additional strategies to diversify the repertoire of antigen specificities. These unconventional mechanisms exclusively target the antigen-binding sites of immunoglobulins and include the insertion of large amino acid sequences, post-translational modifications, conformational heterogeneity and use of nonprotein cofactor molecules. Here, we describe the different unconventional routes for diversification of antibody specificities. Furthermore, we highlight how the immune system has a much greater level of adaptability and plasticity than previously anticipated, which goes far beyond that encoded in the genome or generated by the acquisition of somatic mutations.

THE CIRCUIT IN Figure 1 is a proportional thermoregulator in which the switching of the triac occurs when the mains voltage crosses the zero level. As a result, the circuit [...]

We evaluated the potential clinical relevance of malondialdehyde (MDA) and autoantibodies to copper oxidized low-density lipoprotein (CuOx-LDL) in type 2 diabetes occurrence.

Human high-affinity antibodies to pathogens often recognize unrelated ligands. The molecular origin and the role of this polyreactivity are largely unknown. Here, we report that HIV-1 broadly neutralizing antibodies (bNAbs) are frequently polyreactive, cross-reacting with non-HIV-1 molecules, including self-antigens. Mutating bNAb genes to increase HIV-1 binding and neutralization also results in de novopolyreactivity. Unliganded paratopes of polyreactive bNAbs with improved HIV-1 neutralization exhibit a conformational flexibility, which contributes to enhanced affinity of bNAbs to various HIV-1 envelope glycoproteins and non-HIV antigens. Binding adaptation of polyreactive bNAbs to the divergent ligands mainly involves hydrophophic interactions. Plasticity of bNAbs’ paratopes may, therefore, facilitate accommodating divergent viral variants, but it simultaneously triggers promiscuous binding to non-HIV-1 antigens. Thus, a certain level of polyreactivity can be a mark of adaptable antibodies displaying optimal pathogens’ recognition.

Emicizumab is a bispecific, chimeric, humanized immunoglobulin G (IgG)4 that mimics the procoagulant activity of factor (F) VIII (FVIII). Its long half-life and subcutaneous route of administration have been life-changing in treating patients with hemophilia A (HA) with or without FVIII inhibitors. However, emicizumab only partially mimics FVIII activity; it prevents but does not treat acute bleeds. Emergency management is particularly complicated in patients with FVIII inhibitors receiving emicizumab prophylaxis in whom exogenous FVIII is inefficient. We have shown recently that Imlifidase (IdeS), a bacterial IgG-degrading enzyme, efficiently eliminates human anti-FVIII IgG in a mouse model of severe HA with inhibitors and opens a therapeutic window for the administration of exogenous FVIII.

Healthy immune repertoire contains a fraction of immunoglobulins that do not possess exquisite antigen specificity but are able to recognize numerous unrelated antigens with similar values of the binding affinity. These antibodies are referred to as polyreactive. Besides natural polyreactive antibodies immune repertoires contain antibodies that acquire polyreactivity post-translationally, upon structural changes in their variable regions. In this article we made an overview of the recent findings about antibody polyreactivity. After introduction of the concept, and description of the origin, functions, and molecular mechanisms of polyreactive antibodies, we discussed their role in autoimmunity, malignancy and infectious diseases. We made a parallel with similar data about antibodies with induced polyreactivity. This review highlights the importance of natural and acquired antibody polyreactivity in immune defense and surveillance and reveals their potential as a new type of therapeutics.

Memory B-cell and antibody responses to the SARS-CoV-2 spike protein contribute to long-term immune protection against severe COVID-19, which can also be prevented by antibody-based interventions. Here, wide SARS-CoV-2 immunoprofiling in Wuhan COVID-19 convalescents combining serological, cellular, and monoclonal antibody explorations revealed humoral immunity coordination. Detailed characterization of a hundred SARS-CoV-2 spike memory B-cell monoclonal antibodies uncovered diversity in their repertoire and antiviral functions. The latter were influenced by the targeted spike region with strong Fc-dependent effectors to the S2 subunit and potent neutralizers to the receptor-binding domain. Amongst those, Cv2.1169 and Cv2.3194 antibodies cross-neutralized SARS-CoV-2 variants of concern, including Omicron BA.1 and BA.2. Cv2.1169, isolated from a mucosa-derived IgA memory B cell demonstrated potency boost as IgA dimers and therapeutic efficacy as IgG antibodies in animal models. Structural data provided mechanistic clues to Cv2.1169 potency and breadth. Thus, potent broadly neutralizing IgA antibodies elicited in mucosal tissues can stem SARS-CoV-2 infection, and Cv2.1169 and Cv2.3194 are prime candidates for COVID-19 prevention and treatment.

Vascular endothelial cells (ECs) link hemostasis, thrombosis, and complement. ECs synthesize both the clotting initiator von Willebrand factor (VWF) and the complement regulator factor H (FH). VWF is stored in EC Weibel-Palade bodies (WPBs), but the intracellular location of FH is not well defined. We found that FH colocalizes with VWF in WPBs of human umbilical vein ECs. Moreover, FH bound to VWF with an apparent nanomolar affinity and the complex was present in normal plasma. The binding of VWF to FH enhanced FH cofactor activity toward factor I–mediated downregulation of complement activation. Besides, this interaction inhibited ADAMTS13-mediated proteolysis of VWF and promoted platelet aggregation. Here, we describe a novel interaction between complement and hemostasis. The simultaneous secretion of VWF and FH by activated ECs may promote adhesion of platelets to endothelial injury sites to assure wound healing, simultaneously dampening the proinflammatory effect of complement to limit bystander tissue damage.

Decrypting the B cell ontogeny of HIV-1 broadly neutralizing antibodies (bNAbs) is paramount for vaccine design. Here, we characterized IgA and IgG bNAbs of three distinct B cell lineages in a viremic controller, two of which comprised only IgG+ or IgA+ blood memory B cells; the third combined both IgG and IgA clonal variants. 7-269 bNAb in the IgA-only lineage displayed the highest neutralizing capacity despite limited somatic mutation, and delayed viral rebound in humanized mice. bNAbs in all three lineages targeted the N332 glycan supersite. The 2.8-Å resolution cryo-EM structure of 7-269-BG505 SOSIP.664 complex showed a similar pose as 2G12, on an epitope mainly composed of sugar residues comprising the N332 and N295 glycans. Binding and cryo-EM structural analyses showed that antibodies from the two other lineages interact mostly with glycans N332 and N386. Hence, multiple B cell lineages of IgG and IgA bNAbs focused on a unique HIV-1 site of vulnerability can codevelop in HIV-1 viremic controllers.

Atypical hemolytic uremic syndrome (aHUS) is characterized by genetic and acquired abnormalities of the complement system leading to alternative pathway (AP) overactivation and by glomerular endothelial damage, thrombosis, and mechanical hemolysis. Mutations per se are not sufficient to induce aHUS, and nonspecific primary triggers are required for disease manifestation. We investigated whether hemolysis-derived heme contributes to aHUS pathogenesis. We confirmed that heme activates complement AP in normal human serum, releasing C3a, C5a, and sC5b9. We demonstrated that heme-exposed endothelial cells also activate the AP, resulting in cell-bound C3 and C5b9. This was exacerbated in aHUS by genetic abnormalities associated with AP overactivation. Heme interacted with C3 close to the thioester bond, induced homophilic C3 complexes, and promoted formation of an overactive C3/C5 convertase. Heme induced decreased membrane cofactor protein (MCP) and decay-accelerating factor (DAF) expression on endothelial cells, giving Factor H (FH) a major role in complement regulation. Finally, heme promoted a rapid exocytosis of Weibel-Palade bodies, with membrane expression of P-selectin known to bind C3b and trigger the AP, and the release of the prothrombotic von Willebrand factor. These results strongly suggest that hemolysis-derived heme represents a common secondary hit amplifying endothelial damage and thrombosis in aHUS.

Acquired hemophilia is a rare bleeding disorder characterized by the spontaneous occurrence of inhibitory antibodies against endogenous factor VIII (FVIII). IgG from some patients with acquired hemophilia hydrolyze FVIII. Because of the complex etiology of the disease, no clinical parameter, including the presence of FVIII-hydrolyzing IgG, has been associated with patient's survival or death. Here, we demonstrate the presence of anti-FIX antibodies in acquired hemophilia patients. IgG from some patients were found to hydrolyze FIX. In most cases, IgG-mediated FIX-hydrolysis resulted in FIX activation. IgG-mediated hydrolysis of FIX thus led to the significant generation of activated FIX in 25 of 65 patients. Based on the estimated kinetic parameters, patients' IgG activated up to 0.3nM FIX in 24 hours, an amount that restored thrombin generation in vitro provided the presence of more than or equal to 3% residual FVIII activity in plasma. This work identifies proteolytic IgG as novel molecules able to activate FIX under pathologic conditions. IgG-mediated FIX activation is a prevalent phenomenon among acquired hemophilia patients. The presence of FIX-activating IgG may partly compensate for the antibody-mediated inhibition of endogenous FVIII in restoring thrombin generation. This clinical trial was registered at www.clinicaltrials.gov as #NCT00213473.

Replacement therapy with exogenous factor VIII (FVIII) to treat hemorrhages induces anti-FVIII inhibitory immunoglobulin G in up to 30% of patients with hemophilia A. Chronic inflammation associated with recurrent bleedings is a proposed risk factor for FVIII inhibitor development. Heme oxygenase-1 (HO-1) is a stress-inducible enzyme with potent anti-inflammatory activity. Here, we demonstrate that induction of HO-1 before FVIII administration drastically reduces the onset of the anti-FVIII humoral immune response. The protective effect was specific for HO-1 because it was reproduced on administration of the end products of HO-1 activity, carbon monoxide, and bilirubin, and prevented by the pharmacologic inhibition of HO-1 using tin mesoporphyrin IX. HO-1 induction was associated with decreased major histocompatibility complex class II expression by splenic antigen-presenting cells and reduced T-cell proliferation. Triggering the endogenous anti-inflammatory machinery before FVIII administration may represent a novel therapeutic option for preventing the development of FVIII inhibitors in hemophilia A patients.

Complement is a major innate immune defense against pathogens, tightly regulated to prevent host tissue damage. Atypical hemolytic uremic syndrome (aHUS) is characterized by endothelial damage leading to renal failure and is highly associated with abnormal alternative pathway regulation. We characterized the functional consequences of 2 aHUS-associated mutations (D254G and K325N) in factor B, a key participant in the alternative C3 convertase. Mutant proteins formed high-affinity C3-binding site, leading to a hyperfunctional C3 convertase, resistant to decay by factor H. This led to enhanced complement deposition on the surface of alternative pathway activator cells. In contrast to native factor B, the 2 mutants bound to inactivated C3 and induced formation of functional C3-convertase on iC3b-coated surface. We demonstrated for the first time that factor B mutations lead to enhanced C3-fragment deposition on quiescent and adherent human glomerular cells (GEnCs) and human umbilical vein endothelial cells (HUVECs), together with the formation of sC5b-9 complexes. These results could explain the occurrence of the disease, since excessive complement deposition on endothelial cells is a central event in the pathogenesis of aHUS. Therefore, risk factors for aHUS are not only mutations leading to loss of regulation, but also mutations, resulting in hyperactive C3 convertase.

Rare individuals can naturally clear chronic hepatitis B virus (HBV) infection and acquire protection from reinfection as conferred by vaccination. To examine the protective humoral response against HBV, we cloned and characterized human antibodies specific to the viral surface glycoproteins (HBsAg) from memory B cells of HBV vaccinees and controllers. We found that human HBV antibodies are encoded by a diverse set of immunoglobulin genes and recognize various conformational HBsAg epitopes. Strikingly, HBsAg-specific memory B cells from natural controllers mainly produced neutralizing antibodies able to cross-react with several viral genotypes. Furthermore, monotherapy with the potent broadly neutralizing antibody Bc1.187 suppressed viremia in vivo in HBV mouse models and led to post-therapy control of the infection in a fraction of animals. Thus, human neutralizing HBsAg antibodies appear to play a key role in the spontaneous control of HBV and represent promising immunotherapeutic tools for achieving HBV functional cure in chronically infected humans.