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1.
Mol Diagn Ther ; 28(2): 189-199, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38261250

RESUMO

The complement system plays a dual role in the body, either as a first-line defense barrier when balanced between activation and inhibition or as a potential driver of complement-associated injury or diseases when unbalanced or over-activated. C4b-binding protein (C4BP) was the first circulating complement regulatory protein identified and it functions as an important complement inhibitor. C4BP can suppress the over-activation of complement components and prevent the complement system from attacking the host cells through the binding of complement cleavage products C4b and C3b, working in concert as a cofactor for factor I in the degradation of C4b and C3b, and consequently preventing or reducing the assembly of C3 convertase and C5 convertase, respectively. C4BP, particularly C4BP α-chain (C4BPα), exerts its unique inhibitory effects on complement activation and opsonization, systemic inflammation, and platelet activation and aggregation. It has long been acknowledged that crosstalk or interplay exists between the complement system and platelets. Our unpublished preliminary data suggest that circulating C4BPα exerts its antiplatelet effects through inhibition of both complement activity levels and complement-induced platelet reactivity. Plasma C4BPα levels appear to be significantly higher in patients sensitive to, rather than resistant to, clopidogrel, and we suggest that a plasma C4BPα measurement could be used to predict clopidogrel resistance in the clinical settings.


Assuntos
Proteína de Ligação ao Complemento C4b , Proteínas do Sistema Complemento , Humanos , Biomarcadores , Clopidogrel , Convertases de Complemento C3-C5/metabolismo , Proteína de Ligação ao Complemento C4b/metabolismo
2.
Kidney Int ; 105(2): 328-337, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38008161

RESUMO

Renin, an aspartate protease, regulates the renin-angiotensin system by cleaving its only known substrate angiotensinogen to angiotensin. Recent studies have suggested that renin may also cleave complement component C3 to activate complement or contribute to its dysregulation. Typically, C3 is cleaved by C3 convertase, a serine protease that uses the hydroxyl group of a serine residue as a nucleophile. Here, we provide seven lines of evidence to show that renin does not cleave C3. First, there is no association between renin plasma levels and C3 levels in patients with C3 Glomerulopathies (C3G) and atypical Hemolytic Uremic Syndrome (aHUS), implying that serum C3 consumption is not increased in the presence of high renin. Second, in vitro tests of C3 conversion to C3b do not detect differences when sera from patients with high renin levels are compared to sera from patients with normal/low renin levels. Third, aliskiren, a renin inhibitor, does not block abnormal complement activity introduced by nephritic factors in the fluid phase. Fourth, aliskiren does not block dysregulated complement activity on cell surfaces. Fifth, recombinant renin from different sources does not cleave C3 even after 24 hours of incubation at 37 °C. Sixth, direct spiking of recombinant renin into sera samples of patients with C3G and aHUS does not enhance complement activity in either the fluid phase or on cell surfaces. And seventh, molecular modeling and docking place C3 in the active site of renin in a position that is not consistent with a productive ground state complex for catalytic hydrolysis. Thus, our study does not support a role for renin in the activation of complement.


Assuntos
Ativação do Complemento , Complemento C3 , Nefropatias , Renina , Humanos , Amidas , Síndrome Hemolítico-Urêmica Atípica , Complemento C3/metabolismo , Convertases de Complemento C3-C5/metabolismo , Via Alternativa do Complemento , Fumaratos , Renina/antagonistas & inibidores , Renina/sangue , Renina/metabolismo
4.
J Immunol ; 211(5): 862-873, 2023 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-37466368

RESUMO

Trypanosomes are known to activate the complement system on their surface, but they control the cascade in a manner such that the cascade does not progress into the terminal pathway. It was recently reported that the invariant surface glycoprotein ISG65 from Trypanosoma brucei interacts reversibly with complement C3 and its degradation products, but the molecular mechanism by which ISG65 interferes with complement activation remains unknown. In this study, we show that ISG65 does not interfere directly with the assembly or activity of the two C3 convertases. However, ISG65 acts as a potent inhibitor of C3 deposition through the alternative pathway in human and murine serum. Degradation assays demonstrate that ISG65 stimulates the C3b to iC3b converting activity of complement factor I in the presence of the cofactors factor H or complement receptor 1. A structure-based model suggests that ISG65 promotes a C3b conformation susceptible to degradation or directly bridges factor I and C3b without contact with the cofactor. In addition, ISG65 is observed to form a stable ternary complex with the ligand binding domain of complement receptor 3 and iC3b. Our data suggest that ISG65 supports trypanosome complement evasion by accelerating the conversion of C3b to iC3b through a unique mechanism.


Assuntos
Trypanosoma brucei brucei , Camundongos , Animais , Humanos , Trypanosoma brucei brucei/metabolismo , Complemento C3b/metabolismo , Receptores de Complemento 3b , Ativação do Complemento , Fator H do Complemento/metabolismo , Fibrinogênio , Via Alternativa do Complemento , Convertases de Complemento C3-C5/metabolismo
5.
Nat Commun ; 14(1): 2403, 2023 04 27.
Artigo em Inglês | MEDLINE | ID: mdl-37105991

RESUMO

African Trypanosomes have developed elaborate mechanisms to escape the adaptive immune response, but little is known about complement evasion particularly at the early stage of infection. Here we show that ISG65 of the human-infective parasite Trypanosoma brucei gambiense is a receptor for human complement factor C3 and its activation fragments and that it takes over a role in selective inhibition of the alternative pathway C5 convertase and thus abrogation of the terminal pathway. No deposition of C4b, as part of the classical and lectin pathway convertases, was detected on trypanosomes. We present the cryo-electron microscopy (EM) structures of native C3 and C3b in complex with ISG65 which reveal a set of modes of complement interaction. Based on these findings, we propose a model for receptor-ligand interactions as they occur at the plasma membrane of blood-stage trypanosomes and may facilitate innate immune escape of the parasite.


Assuntos
Complemento C3 , Trypanosoma brucei gambiense , Humanos , Ativação do Complemento , Complemento C3/metabolismo , Convertases de Complemento C3-C5/metabolismo , Complemento C5/metabolismo , Via Alternativa do Complemento , Microscopia Crioeletrônica , Ligação Proteica , Trypanosoma brucei gambiense/metabolismo
6.
Blood Adv ; 7(16): 4258-4268, 2023 08 22.
Artigo em Inglês | MEDLINE | ID: mdl-36897252

RESUMO

Dysregulated activation of the complement system is implicated in the onset or progression of several diseases. Most clinical-stage complement inhibitors target the inactive complement proteins present at high concentrations in plasma, which increases target-mediated drug disposition and necessitates high drug levels to sustain therapeutic inhibition. Furthermore, many efforts are aimed at inhibiting only terminal pathway activity, which leaves opsonin-mediated effector functions intact. We describe the discovery of SAR443809, a specific inhibitor of the alternative pathway C3/C5 convertase (C3bBb). SAR443809 selectively binds to the activated form of factor B (factor Bb) and inhibits alternative pathway activity by blocking the cleavage of C3, leaving the initiation of classical and lectin complement pathways unaffected. Ex vivo experiments with patient-derived paroxysmal nocturnal hemoglobinuria erythrocytes show that, although terminal pathway inhibition via C5 blockade can effectively inhibit hemolysis, proximal complement inhibition with SAR443809 inhibits both hemolysis and C3b deposition, abrogating the propensity for extravascular hemolysis. Finally, intravenous and subcutaneous administration of the antibody in nonhuman primates demonstrated sustained inhibition of complement activity for several weeks after injection. Overall, SAR443809 shows strong potential for treatment of alternative pathway-mediated disorders.


Assuntos
Fator B do Complemento , Via Alternativa do Complemento , Animais , Fator B do Complemento/antagonistas & inibidores , Eritrócitos/efeitos dos fármacos , Hemólise/efeitos dos fármacos , Convertases de Complemento C3-C5/antagonistas & inibidores , Via Alternativa do Complemento/efeitos dos fármacos , Doenças do Sistema Imunitário/tratamento farmacológico , Doenças do Sistema Imunitário/enzimologia , Humanos , Macaca fascicularis , Anticorpos/administração & dosagem , Proteólise/efeitos dos fármacos
7.
Front Immunol ; 14: 1113015, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36891314

RESUMO

Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disorder affecting the joints. Many patients carry anti-citrullinated protein autoantibodies (ACPA). Overactivation of the complement system seems to be part of the pathogenesis of RA, and autoantibodies against the pathway initiators C1q and MBL, and the regulator of the complement alternative pathway, factor H (FH), were previously reported. Our aim was to analyze the presence and role of autoantibodies against complement proteins in a Hungarian RA cohort. To this end, serum samples of 97 ACPA-positive RA patients and 117 healthy controls were analyzed for autoantibodies against FH, factor B (FB), C3b, C3-convertase (C3bBbP), C1q, MBL and factor I. In this cohort, we did not detect any patient with FH autoantibodies but detected C1q autoantibodies in four patients, MBL autoantibodies in two patients and FB autoantibodies in five patients. Since the latter autoantibodies were previously reported in patients with kidney diseases but not in RA, we set out to further characterize such FB autoantibodies. The isotypes of the analyzed autoantibodies were IgG2, IgG3, IgGκ, IgGλ and their binding site was localized in the Bb part of FB. We detected in vivo formed FB-autoanti-FB complexes by Western blot. The effect of the autoantibodies on the formation, activity and FH-mediated decay of the C3 convertase in solid phase convertase assays was determined. In order to investigate the effect of the autoantibodies on complement functions, hemolysis assays and fluid phase complement activation assays were performed. The autoantibodies partially inhibited the complement-mediated hemolysis of rabbit red blood cells, inhibited the activity of the solid phase C3-convertase and C3 and C5b-9 deposition on complement activating surfaces. In summary, in ACPA-positive RA patients we identified FB autoantibodies. The characterized FB autoantibodies did not enhance complement activation, rather, they had inhibitory effect on complement. These results support the involvement of the complement system in the pathomechanism of RA and raise the possibility that protective autoantibodies may be generated in some patients against the alternative pathway C3 convertase. However, further analyses are needed to assess the exact role of such autoantibodies.


Assuntos
Artrite Reumatoide , Fator B do Complemento , Animais , Coelhos , Autoanticorpos , Hemólise , Complemento C1q , Convertases de Complemento C3-C5/metabolismo
8.
J Biol Chem ; 299(3): 102930, 2023 03.
Artigo em Inglês | MEDLINE | ID: mdl-36682494

RESUMO

Hyperactivation of the complement system, a major component of innate immunity, has been recognized as one of the core clinical features in severe covid-19 patients. However, how the virus escapes the targeted elimination by the network of activated complement pathways still remains an enigma. Here, we identified SARS-CoV-2-encoded ORF8 protein as one of the major binding partners of human complement C3/C3b components and their metabolites. Our results demonstrated that preincubation of ORF8 with C3/C3b in the fluid phase has two immediate functional consequences in the alternative pathway; this preincubation inhibits factor I-mediated proteolysis and blocks factor B zymogen activation into active Bb. ORF8 binding results in the occlusion of both factor H and factor B from C3b, rendering the complexes resistant to factor I-mediated proteolysis and inhibition of pro-C3-convertase (C3bB) formation, respectively. We also confirmed the complement inhibitory activity of ORF8 in our hemolysis-based assay, where ORF8 prevented human serum-induced lysis of rabbit erythrocytes with an IC50 value of about 2.3 µM. This inhibitory characteristic of ORF8 was also supported by in-silico protein-protein docking analysis, as it appeared to establish primary interactions with the ß-chain of C3b, orienting itself near the C3b CUB (C1r/C1s, Uegf, Bmp1) domain like a peptidomimetic compound, sterically hindering the binding of essential cofactors required for complement amplification. Thus, ORF8 has characteristics to act as an inhibitor of critical regulatory steps in the alternative pathway, converging to hasten the decay of C3-convertase and thereby, attenuating the complement amplification loop.


Assuntos
COVID-19 , Fator B do Complemento , Animais , Humanos , Coelhos , Ativação do Complemento , Convertases de Complemento C3-C5/metabolismo , Complemento C3b/metabolismo , Fator B do Complemento/metabolismo , Fator H do Complemento/metabolismo , Via Alternativa do Complemento/fisiologia , SARS-CoV-2/metabolismo , Ligação Proteica , Simulação por Computador
9.
J Clin Periodontol ; 50(5): 657-670, 2023 05.
Artigo em Inglês | MEDLINE | ID: mdl-36632003

RESUMO

AIMS: To use experimental periodontitis models in rats to investigate the correlation between local expression of the complement components C3b and C4b in periodontal tissues and disease severity, and to assess the therapeutic effects of targeting C3b/C4b on inflammatory bone loss. MATERIALS AND METHODS: The gingival expression of C3, C3b, and C4b in animal experimental periodontitis models were analysed immunohistochemically. The therapeutic effects of the C3b/C4b inhibitor (SB002) on ligation-induced experimental periodontitis was examined using biochemical, histological, and immunohistochemical analyses. RESULTS: The gingival expression levels of C3, C3b, and C4b were positively correlated with the severity of periodontitis. Moreover, both single and multiple injections of the C3b/C4b inhibitor had preventive and therapeutic effects on alveolar bone loss in ligation-induced experimental periodontitis with no associated adverse consequences. CONCLUSIONS: The association between C3b/C4b and periodontitis may provide a basis for the development of novel therapeutic strategies for periodontitis and other inflammatory diseases.


Assuntos
Complemento C4b , Periodontite , Ratos , Animais , Complemento C4b/metabolismo , Complemento C3b/metabolismo , Convertases de Complemento C3-C5/metabolismo , Inflamação , Periodontite/complicações , Periodontite/tratamento farmacológico
10.
Curr Top Behav Neurosci ; 61: 243-264, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36059003

RESUMO

BACKGROUND: Herpesviruses alter cognitive functions in humans following acute infections; progressive cognitive decline and dementia have also been suggested. It is important to understand the pathogenic mechanisms of such infections. The complement system - comprising functionally related proteins integral for systemic innate and adaptive immunity - is an important component of host responses. The complement system has specialized functions in the brain. Still, the dynamics of the brain complement system are still poorly understood. Many complement proteins have limited access to the brain from plasma, necessitating synthesis and specific regulation of expression in the brain; thus, complement protein synthesis, activation, regulation, and signaling should be investigated in human brain-relevant cellular models. Cells derived from human-induced pluripotent stem cells (hiPSCs) could enable tractable models. METHODS: Human-induced pluripotent stem cells were differentiated into neuronal (hi-N) and microglial (hi-M) cells that were cultured with primary culture human astrocyte-like cells (ha-D). Gene expression analyses and complement protein levels were analyzed in mono- and co-cultures. RESULTS: Transcript levels of complement proteins differ by cell type and co-culture conditions, with evidence for cellular crosstalk in co-cultures. Hi-N and hi-M cells have distinct patterns of expression of complement receptors, soluble factors, and regulatory proteins. hi-N cells produce complement factor 4 (C4) and factor B (FB), whereas hi-M cells produce complement factor 2 (C2) and complement factor 3 (C3). Thus, neither hi-N nor hi-M cells can form either of the C3-convertases - C4bC2a and C3bBb. However, when hi-N and hi-M cells are combined in co-cultures, both types of functional C3 convertase are produced, indicated by elevated levels of the cleaved C3 protein, C3a. CONCLUSIONS: hiPSC-derived co-culture models can be used to study viral infection in the brain, particularly complement receptor and function in relation to cellular "crosstalk." The models could be refined to further investigate pathogenic mechanisms.


Assuntos
Infecções por Herpesviridae , Células-Tronco Pluripotentes Induzidas , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Complemento C3/metabolismo , Neurônios/metabolismo , Convertases de Complemento C3-C5/metabolismo , Encéfalo/metabolismo , Infecções por Herpesviridae/metabolismo
11.
Front Immunol ; 13: 981375, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36189215

RESUMO

The complement system is part of the innate immune system. The crucial step in activating the complement system is the generation and regulation of C3 convertase complexes, which are needed to generate opsonins that promote phagocytosis, to generate C3a that regulates inflammation, and to initiate the lytic terminal pathway through the generation and activity of C5 convertases. A growing body of evidence has highlighted the interplay between the complement system, coagulation system, platelets, neutrophils, and endothelial cells. The kidneys are highly susceptible to complement-mediated injury in several genetic, infectious, and autoimmune diseases. Atypical hemolytic uremic syndrome (aHUS) and lupus nephritis (LN) are both characterized by thrombosis in the glomerular capillaries of the kidneys. In aHUS, congenital or acquired defects in complement regulators may trigger platelet aggregation and activation, resulting in the formation of platelet-rich thrombi in the kidneys. Because glomerular vasculopathy is usually noted with immunoglobulin and complement accumulation in LN, complement-mediated activation of tissue factors could partly explain the autoimmune mechanism of thrombosis. Thus, kidney glomerular capillary thrombosis is mediated by complement dysregulation and may also be associated with complement overactivation. Further investigation is required to clarify the interaction between these vascular components and develop specific therapeutic approaches.


Assuntos
Síndrome Hemolítico-Urêmica Atípica , Trombose , Síndrome Hemolítico-Urêmica Atípica/genética , Capilares/metabolismo , Convertases de Complemento C3-C5/metabolismo , Proteínas do Sistema Complemento/metabolismo , Células Endoteliais/metabolismo , Humanos , Rim/metabolismo , Proteínas Opsonizantes/metabolismo , Trombose/metabolismo
12.
Front Immunol ; 13: 1006761, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36172347

RESUMO

During organ transplantation, ischemia/reperfusion injury and pre-formed anti-HLA antibodies are the main cause of delayed graft function and recovery through the activation of the complement system. By supplying oxygen during transplantation, M101 is suspected to avoid complement activation, however, a direct effect exerted by M101 on this pathway is unknown. This was tested by using functional assays (lymphocytotoxic crossmatch test, C3d Luminex-based assay, 50% complement hemolysis [CH50], and 50% alternative complement pathway [AP50/AH50]), and quantitative assays (C3, C3a, C4, C5, C5a, C6, C7, C8, C9 and sC5b-9). M101 interferes with the anti-HLA lymphocytotoxic crossmatch assay, and this effect is complement-dependent as M101 inhibits the classical complement pathway (CH50) in a dose-dependent and stable manner. Such inhibition was independent from a proteolytic effect (fractions C3 to C9) but related to a dose-dependent inhibition of the C3 convertase as demonstrated by exploring downstream the release of the anaphylatoxins (C3a and C5a), C3d, and sC5b-9. The C3 convertase inhibition in the presence of M101 was further demonstrated in the AP50/AH50 assay. In conclusion, the use of M101 avoids the activation of the complement pathway, which constitutes an additional advantage for this extracellular hemoglobin to preserve grafts from ischemia/reperfusion injury and preformed anti-HLA antibodies.


Assuntos
Preservação de Órgãos , Traumatismo por Reperfusão , Anafilatoxinas , Ativação do Complemento , Complemento C3 , Convertases de Complemento C3-C5 , Hemoglobinas , Humanos , Isquemia , Oxigênio , Traumatismo por Reperfusão/prevenção & controle
13.
Protein Sci ; 31(10): e4432, 2022 10.
Artigo em Inglês | MEDLINE | ID: mdl-36173177

RESUMO

Structure determination of macromolecular complexes is challenging if subunits can dissociate during crystallization or preparation of electron microscopy grids. We present an approach where a labile complex is stabilized by linking subunits though introduction of a peptide tag in one subunit that is recognized by a nanobody tethered to a second subunit. This allowed crystal structure determination at 3.9 Å resolution of the highly non-globular 320 kDa proconvertase formed by complement components C3b, factor B, and properdin. Whereas the binding mode of properdin to C3b is preserved, an internal rearrangement occurs in the zymogen factor B von Willebrand domain type A domain compared to the proconvertase not bound to properdin. The structure emphasizes the role of two noncanonical loops in thrombospondin repeats 5 and 6 of properdin in augmenting the activity of the C3 convertase. We suggest that linking of subunits through peptide specific tethered nanobodies represents a simple alternative to approaches like affinity maturation and chemical cross-linking for the stabilization of large macromolecular complexes. Besides applications for structural biology, nanobody bridging may become a new tool for biochemical analysis of unstable macromolecular complexes and in vitro selection of highly specific binders for such complexes.


Assuntos
Properdina , Anticorpos de Domínio Único , Convertases de Complemento C3-C5/química , Convertases de Complemento C3-C5/metabolismo , Fator B do Complemento/química , Fator B do Complemento/metabolismo , Precursores Enzimáticos , Substâncias Macromoleculares , Properdina/química , Properdina/metabolismo , Trombospondinas
14.
Nat Commun ; 13(1): 5409, 2022 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-36109509

RESUMO

Failure of the right ventricle plays a critical role in any type of heart failure. However, the mechanism remains unclear, and there is no specific therapy. Here, we show that the right ventricle predominantly expresses alternative complement pathway-related genes, including Cfd and C3aR1. Complement 3 (C3)-knockout attenuates right ventricular dysfunction and fibrosis in a mouse model of right ventricular failure. C3a is produced from C3 by the C3 convertase complex, which includes the essential component complement factor D (Cfd). Cfd-knockout mice also show attenuation of right ventricular failure. Moreover, the plasma concentration of CFD correlates with the severity of right ventricular failure in patients with chronic right ventricular failure. A C3a receptor (C3aR) antagonist dramatically improves right ventricular dysfunction in mice. In summary, we demonstrate the crucial role of the C3-Cfd-C3aR axis in right ventricular failure and highlight potential therapeutic targets for right ventricular failure.


Assuntos
Insuficiência Cardíaca , Disfunção Ventricular Direita , Animais , Complemento C3/genética , Convertases de Complemento C3-C5 , Fator D do Complemento , Insuficiência Cardíaca/genética , Camundongos , Camundongos Knockout , Remodelação Ventricular
15.
Front Immunol ; 13: 872536, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35935935

RESUMO

The complement system is a part of the innate immune system, where it labels intruding pathogens as well as dying host cells for clearance. If complement regulation is compromised, the system may contribute to pathogenesis. The proteolytic fragment C3b of complement component C3, is the pivot point of the complement system and provides a scaffold for the assembly of the alternative pathway C3 convertase that greatly amplifies the initial complement activation. This makes C3b an attractive therapeutic target. We previously described a nanobody, hC3Nb1 binding to C3 and its degradation products. Here we show, that extending the N-terminus of hC3Nb1 by a Glu-Trp-Glu motif renders the resulting EWE-hC3Nb1 (EWE) nanobody specific for C3 degradation products. By fusing EWE to N-terminal CCP domains from complement Factor H (FH), we generated the fusion proteins EWEnH and EWEµH. In contrast to EWE, these fusion proteins supported Factor I (FI)-mediated cleavage of human and rat C3b. The EWE, EWEµH, and EWEnH proteins bound C3b and iC3b with low nanomolar dissociation constants and exerted strong inhibition of alternative pathway-mediated deposition of complement. Interestingly, EWEnH remained soluble above 20 mg/mL. Combined with the observed reactivity with both human and rat C3b as well as the ability to support FI-mediated cleavage of C3b, this features EWEnH as a promising candidate for in vivo studies in rodent models of complement driven pathogenesis.


Assuntos
Complemento C3 , Complemento C3b , Animais , Ativação do Complemento , C3 Convertase da Via Alternativa do Complemento , Convertases de Complemento C3-C5/metabolismo , Complemento C3b/metabolismo , Fibrinogênio/metabolismo , Humanos , Ratos
16.
Front Immunol ; 13: 942482, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35958553

RESUMO

Multidrug-resistant Acinetobacter baumannii is known to be one of the leading pathogens that cause severe nosocomial infections. To overcome eradication by the innate immune system during infection, A. baumannii developed a number of immune evasion strategies. Previously, we identified CipA as a plasminogen-binding and complement-inhibitory protein. Here we show that CipA inhibits all three complement activation pathways and interacts with key complement components C3, C3b, C4b, C5, Factor B, Factor D, and in particular Factor I. CipA also targets function of the C5 convertase as cleavage of C5 was impaired. Systematic screening of CipA variants identified two separate binding sites for C3b and a Factor I-interacting domain located at the C-terminus. Structure predictions using AlphaFold2 and binding analyses employing CipA variants lacking Factor I-binding capability confirmed that the orientation of the C-terminal domain is essential for the interaction with Factor I. Hence, our analyses point to a novel Factor I-dependent mechanisms of complement inactivation mediated by CipA of A. baumannii. Recruitment of Factor I by CipA initiates the assembly of a quadripartite complex following binding of either Factor H or C4b-binding protein to degrade C3b and C4b, respectively. Loss of Factor I binding in a CipA-deficient strain, or a strain producing a CipA variant lacking Factor I-binding capability, correlated with a higher susceptibility to human serum, indicating that recruitment of Factor I enables A. baumannii to resist complement-mediated killing.


Assuntos
Acinetobacter baumannii , Ativação do Complemento , Convertases de Complemento C3-C5/metabolismo , Fator B do Complemento/metabolismo , Fibrinogênio/metabolismo , Humanos
17.
Dev Comp Immunol ; 137: 104520, 2022 12.
Artigo em Inglês | MEDLINE | ID: mdl-36041641

RESUMO

Complement plays an important role in the innate immune system, and it comprises about 35 individual proteins. In mammals, complement is activated via three different pathways, the classical pathway, the alternative pathway, and the lectin pathway. All three activation pathways produce C3-convertase in different forms. C3-convertase cleaves C3 to C3a and C3b and initiates a cascade of cleavage and activation, eventually resulting in the formation of the membrane attack complex. Complement activation results in the generation of activated fragments that are involved in microbial killing, phagocytosis, inflammatory reactions, immune complex clearance, and antibody production. Although the complement system has been studied extensively in mammals, complement is less well understood in teleosts. This review summarizes the current knowledge of the teleost complement components involved in phagocytosis, chemotaxis, and cell lysis. We report the characterized complement components in various teleost species. In addition, we provide a comprehensive compilation of complement regulators, and this information is used to analyze the role of complement regulators in pathogen infection. The influence of complement receptors on the immune responses of teleosts is reviewed. Finally, we propose directions for future study of the molecular evolution, structure, and function of complement components in teleosts. This review provides new insights into the complement system of recognition and defense, and such knowledge is essential for the development of new immune strategies in aquaculture.


Assuntos
Complemento C3 , Complexo de Ataque à Membrana do Sistema Complemento , Animais , Complexo Antígeno-Anticorpo , Ativação do Complemento , Complemento C3/metabolismo , Convertases de Complemento C3-C5/metabolismo , Complemento C3b , Lectinas , Mamíferos , Receptores de Complemento/metabolismo
18.
Semin Immunol ; 60: 101634, 2022 03.
Artigo em Inglês | MEDLINE | ID: mdl-35817659

RESUMO

C3 glomerulopathy (C3G) is a rare and complex kidney disease that primarily affects young adults. Renal outcomes remain poor in the absence of specific treatment. C3G is driven by uncontrolled overactivation of the alternative complement pathway, which is mainly of acquired origin. Functional characterization of complement abnormalities (i.e., autoantibodies targeting complement components and variants in complement genes) identified in patients and experimental models of the disease improved the understanding of the disease, making C3G a prototype of complement-mediated diseases. The contribution of C3 convertase, as well as C5 convertase, in disease occurrence, phenotype, and severity is now well established, offering various potential therapeutic interventions. However, the lack of sufficient efficiency in anti-C5 therapy highlights the extreme complexity of the disease and the need for new therapeutic approaches based on C3 and C3 convertase axis inhibition. Here, we provide an overview of the complement activation mechanism involved in C3G and discuss therapeutic options based on complement inhibitors, with a specific focus on C3 inhibition.


Assuntos
Complemento C3 , Nefropatias , Humanos , Complemento C3/metabolismo , Convertases de Complemento C3-C5/metabolismo , Via Alternativa do Complemento/genética , Nefropatias/tratamento farmacológico , Rim/metabolismo
19.
J Immunol Methods ; 507: 113295, 2022 08.
Artigo em Inglês | MEDLINE | ID: mdl-35679953

RESUMO

Factor D (also known as adipsin) is a serine protease and part of the complement system, involved in innate immune responses and effector functions of antibodies. Factor D cleaves factor B complexed with C3b, leading to the C3 convertase C3bBb. This C3 convertase is central in the alternative activation pathway and the amplification loop, which amplifies the two other complement activation pathways: the classical pathway and the lectin pathway. Adipocytes synthesize factor D as a pro-form comprising 6 additional residues that must be cleaved off to generate a mature form. The MBL-associated serine protease 3 (MASP-3), found in complex with the pattern recognition molecules of lectin activation pathway, converts the pro-form to mature factor D, which reportedly is the most abundant form found in the circulation at concentrations of 1-2 µg/ml among healthy individuals. The mature factor D is rate-limiting for complement activation, but little is known about the distribution of pro vs. mature factor D in the circulation, the regulation hereof and the potential activation stimuli of the lectin pathway, responsible for activation of MASP-3 and subsequent conversion of pro-form of factor D. In this light we established and validated an ELISA specific for measuring the pro-form of complement factor D. With a working range of 0.82-25 ng/ml, acceptable intra and inter assay CVs, and a relative recovery rate above 90%, we found that the median plasma concentration in Danish blood donors was 134 ng/ml; corresponding to that 8-15% factor D circulates as pro-form. We also found that blood sampling procedures affect conversion and hence the levels measured in serum and plasma.


Assuntos
Fator D do Complemento , Serina Proteases Associadas a Proteína de Ligação a Manose , Ativação do Complemento , Convertases de Complemento C3-C5 , Fator D do Complemento/metabolismo , Lectina de Ligação a Manose da Via do Complemento , Humanos , Lectinas/metabolismo , Serina Proteases Associadas a Proteína de Ligação a Manose/metabolismo
20.
Immunobiology ; 227(3): 152225, 2022 05.
Artigo em Inglês | MEDLINE | ID: mdl-35567980

RESUMO

Several disorders associated with the total or partial absence of components of the human complement system are known. Deficiencies of classical pathway (CP) components are generally linked to systemic lupus erythematosus (SLE) or SLE-like syndromes. However, only approximately one-third of patients who lack C2 show mild symptoms of SLE. The relatively high frequency of homozygous C2 deficiency without or with minor disease manifestation suggests that there might be a compensatory mechanism which allows the activation of the CP of complement without the absolute requirement of C2. In this study we show that factor B (FB), the C2 homologue of the alternative pathway (AP) of complement, can substitute for C2. This was confirmed by using C4b as immobilised ligand and FB as analyte in Surface Plasmon Resonance (BIACORE). C2 binding to the immobilised C3b-like molecule C3(CH3NH2) was not seen. The estimated binding constant for C4bB complex formation was 2.00 * 10-5 [M]. We were further able to demonstrate that C4b supports the cleavage of Factor B by Factor D. Finally, cleavage of 125I-C3 by C4bBb was evaluated and gave strong evidence that the "hybrid" convertase C4bBb can cleave and activate C3 in vitro. Cleavage activity is very low, but consistent with some of the "C2-bypass" observations of others.


Assuntos
Complemento C4 , Lúpus Eritematoso Sistêmico , Ativação do Complemento , Complemento C2/metabolismo , Complemento C3/metabolismo , Convertases de Complemento C3-C5/metabolismo , Complemento C3b , Fator B do Complemento , Via Clássica do Complemento , Humanos
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