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1.
Front Immunol ; 15: 1352330, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-38694513

RESUMEN

Introduction: COVID-19 patients can develop autoantibodies against a variety of secreted and membrane proteins, including some expressed on lymphocytes. However, it is unclear what proportion of patients might develop anti-lymphocyte antibodies (ALAb) and what functional relevance they might have. Methods: We evaluated the presence and lytic function of ALAb in the sera of a cohort of 85 COVID-19 patients (68 unvaccinated and 17 vaccinated) assigned to mild (N=63), or moderate/severe disease (N=22) groups. Thirty-seven patients were followed-up after recovery. We also analyzed in vivo complement deposition on COVID-19 patients' lymphocytes and examined its correlation with lymphocyte numbers during acute disease. Results: Compared with healthy donors (HD), patients had an increased prevalence of IgM ALAb, which was significantly higher in moderate/severe disease patients and persisted after recovery. Sera from IgM ALAb+ patients exhibited complement-dependent cytotoxicity (CDC) against HD lymphocytes. Complement protein C3b deposition on patients' CD4 T cells was inversely correlated with CD4 T cell numbers. This correlation was stronger in moderate/severe disease patients. Discussion: IgM ALAb and complement activation against lymphocytes may contribute to the acute lymphopenia observed in COVID-19 patients.


Asunto(s)
Autoanticuerpos , COVID-19 , Activación de Complemento , Inmunoglobulina M , SARS-CoV-2 , Humanos , COVID-19/inmunología , COVID-19/sangre , Inmunoglobulina M/sangre , Inmunoglobulina M/inmunología , Masculino , Femenino , Persona de Mediana Edad , Autoanticuerpos/sangre , Autoanticuerpos/inmunología , Activación de Complemento/inmunología , SARS-CoV-2/inmunología , Anciano , Adulto , Linfocitos/inmunología , Prevalencia , Linfocitos T CD4-Positivos/inmunología , Linfopenia/inmunología , Linfopenia/sangre , Complemento C3b/inmunología
2.
Clin Exp Immunol ; 216(3): 252-261, 2024 May 16.
Artículo en Inglés | MEDLINE | ID: mdl-38310540

RESUMEN

Psoriasis is a chronic inflammatory skin disease with a characteristic isomorphic reaction, i.e. the Köbner reaction, induced by slight epidermal trauma. In this study, the tape-stripping technique was used to induce the development of Köbner reaction in 18 subjects with psoriasis. Eight subjects developed a positive reaction. To study the early cellular changes, skin biopsies were taken at the baseline and subsequent time points of 2 h, 1 d, 3 d, and 7 d for the immunostaining of complement C3c, iC3b, and cells expressing complement receptor 3 (CD11b/CD18; a receptor of iC3b) or CD14. The results show that the positive Köbner reaction is associated with rapid (2 h-1 d) and sustained (3-7 d) increase in the expression of epidermal C3c and iC3b and dermal C3c. In addition, there was a positive correlation between CD11b+ and CD14+ cells in baseline and 2 h-1 d biopsies with a subsequent increase in CD11b+ and CD14+ cells in 3-7 d biopsies in the Köbner-positive group. In the Köbner-negative group, only a transient increase in epidermal iC3b at 2 h-1 d, as well as rapid (2 h-1 d) and sustained increase (3-7 d) in dermal iC3b and CD14+ cells, was observed. In experiments with cultured monolayer keratinocytes, a slight cell damage already at 30 mJ/cm2 ultraviolet B irradiation led to increased expression of C3c, but not iC3b. Therefore, there are marked differences between Köbner groups in respect to the expression of C3c, iC3b, and cells expressing CD11b or CD14. Of note is the rapid and sustained increase in epidermal C3c and iC3b in the positive Köbner reaction.


Asunto(s)
Antígeno CD11b , Complemento C3b , Receptores de Lipopolisacáridos , Psoriasis , Humanos , Receptores de Lipopolisacáridos/metabolismo , Masculino , Psoriasis/inmunología , Psoriasis/metabolismo , Femenino , Antígeno CD11b/metabolismo , Adulto , Persona de Mediana Edad , Complemento C3b/metabolismo , Complemento C3b/inmunología , Piel/patología , Piel/inmunología , Piel/metabolismo , Piel/efectos de la radiación , Biopsia , Epidermis/metabolismo , Epidermis/inmunología , Epidermis/patología
3.
Front Immunol ; 13: 946522, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-36091057

RESUMEN

Numerous publications have underlined the link between complement C5a and the clinical course of COVID-19. We previously reported that levels of C5a remain high in the group of severely ill patients up to 90 days after hospital discharge. We have now evaluated which complement pathway fuels the elevated levels of C5a during hospitalization and follow-up. The alternative pathway (AP) activation marker C3bBbP and the soluble fraction of C4d, a footprint of the classical/lectin (CP/LP) pathway, were assessed by immunoenzymatic assay in a total of 188 serial samples from 49 patients infected with SARS-CoV-2. Unlike C5a, neither C3bBbP nor C4d readouts rose proportionally to the severity of the disease. Detailed correlation analyses in hospitalization and follow-up samples collected from patients of different disease severity showed significant positive correlations of AP and CP/LP markers with C5a in certain groups, except for the follow-up samples of the patients who suffered from highly severe COVID-19 and presented the highest C5a readouts. In conclusion, there is not a clear link between persistently high levels of C5a after hospital discharge and markers of upstream complement activation, suggesting the existence of a non-canonical source of C5a in patients with a severe course of COVID-19.


Asunto(s)
COVID-19 , Activación de Complemento , Complemento C3b , Complemento C4b , Complemento C5a , Factor B del Complemento , Fragmentos de Péptidos , Biomarcadores/sangre , COVID-19/sangre , COVID-19/inmunología , Activación de Complemento/inmunología , Complemento C3b/inmunología , Complemento C4b/inmunología , Complemento C5a/análisis , Complemento C5a/inmunología , Factor B del Complemento/inmunología , Proteínas del Sistema Complemento/inmunología , Humanos , Fragmentos de Péptidos/inmunología , SARS-CoV-2
4.
Front Immunol ; 13: 755694, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-35126388

RESUMEN

Atypical hemolytic uremic syndrome (aHUS) is a rare but severe type of thrombotic microangiopathy that is triggered by the abnormal activation of the alternative complement pathway. Previous studies have reported that three completely linked coding variants of CFHR1 form two haplotypes, namely, CFHR1*A (c.469C, c.475C, c.523G) and CFHR1*B (c.469T, c.475G, c.523C). CFHR1*B is associated with susceptibility to aHUS. To explore the genetic mechanism by which CFHR1 isoforms contribute to aHUS, we compared the structures of FHR1*A and FHR1*B by homology modeling and found differences in the angles between SCR3 and SCR4-SCR5, as FHR1*B had a larger angle than FHR1*A. Then, we expressed FHR1*A and FHR1*B recombinant proteins and compared their functions in complement system regulation and inflammation. We found that FHR1*B presented a significantly higher capacity for binding C3b and necrotic cells than FHR1*A. In a cofactor assay, the FHR-1*B showed stronger influence on FH mediated cofactor function than the FHR-1*A, resulted in fewer C3b cleavage products. In the C3 convertase assays, FHR1*B showed more powerful effect compared with FHR1*A regarding to de-regulate FH function of inhibition the assembling of C3bBb. Additionally, we also found that FHR1*B triggered monocytes to secrete higher levels of IL-1ß and IL-6 than FHR1*A. In the present study, we showed that variants of CFHR1 might differently affect complement activation and sterile inflammation. Our findings provide a possible mechanism underlying the predisposition to aHUS caused by CFHR1 isoform CFHR1*B.


Asunto(s)
Síndrome Hemolítico Urémico Atípico/etiología , Síndrome Hemolítico Urémico Atípico/metabolismo , Proteínas Sanguíneas/metabolismo , Activación de Complemento/inmunología , Inflamación/inmunología , Inflamación/metabolismo , Síndrome Hemolítico Urémico Atípico/diagnóstico , Unión Competitiva/inmunología , Biomarcadores , Proteínas Sanguíneas/química , Complemento C3b/inmunología , Complemento C3b/metabolismo , Susceptibilidad a Enfermedades , Células Endoteliales/metabolismo , Humanos , Inflamación/complicaciones , Modelos Moleculares , Necrosis/inmunología , Necrosis/metabolismo , Unión Proteica , Conformación Proteica , Isoformas de Proteínas , Relación Estructura-Actividad
5.
Front Immunol ; 13: 1020712, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-36591264

RESUMEN

Introduction: Platelets have essential functions as first responders in the immune response to pathogens. Activation and aggregation of platelets in bacterial infections can lead to life-threatening conditions such as arterial thromboembolism or sepsis-associated coagulopathy. Methods: In this study, we investigated the role of complement in Escherichia coli (E. coli)-induced platelet aggregation in human whole blood, using Multiplate® aggregometry, flow cytometry, and confocal microscopy. Results and Discussion: We found that compstatin, which inhibits the cleavage of complement component C3 to its components C3a and C3b, reduced the E. coli-induced platelet aggregation by 42%-76% (p = 0.0417). This C3-dependent aggregation was not C3a-mediated as neither inhibition of C3a using a blocking antibody or a C3a receptor antagonist, nor the addition of purified C3a had any effects. In contrast, a C3b-blocking antibody significantly reduced the E. coli-induced platelet aggregation by 67% (p = 0.0133). We could not detect opsonized C3b on platelets, indicating that the effect of C3 was not dependent on C3b-fragment deposition on platelets. Indeed, inhibition of glycoprotein IIb/IIIa (GPIIb/IIIa) and complement receptor 1 (CR1) showed that these receptors were involved in platelet aggregation. Furthermore, aggregation was more pronounced in hirudin whole blood than in hirudin platelet-rich plasma, indicating that E. coli-induced platelet aggregation involved other blood cells. In conclusion, the E. coli-induced platelet aggregation in human whole blood is partly C3b-dependent, and GPIIb/IIIa and CR1 are also involved in this process.


Asunto(s)
Plaquetas , Complemento C3b , Escherichia coli , Agregación Plaquetaria , Humanos , Plaquetas/efectos de los fármacos , Plaquetas/inmunología , Complemento C3b/inmunología , Hirudinas/farmacología , Agregación Plaquetaria/efectos de los fármacos , Agregación Plaquetaria/inmunología , Complejo GPIIb-IIIa de Glicoproteína Plaquetaria/inmunología , Técnicas In Vitro
6.
Nat Immunol ; 23(1): 62-74, 2022 01.
Artículo en Inglés | MEDLINE | ID: mdl-34764490

RESUMEN

The molecular mechanisms governing orderly shutdown and retraction of CD4+ type 1 helper T (TH1) cell responses remain poorly understood. Here we show that complement triggers contraction of TH1 responses by inducing intrinsic expression of the vitamin D (VitD) receptor and the VitD-activating enzyme CYP27B1, permitting T cells to both activate and respond to VitD. VitD then initiated the transition from pro-inflammatory interferon-γ+ TH1 cells to suppressive interleukin-10+ cells. This process was primed by dynamic changes in the epigenetic landscape of CD4+ T cells, generating super-enhancers and recruiting several transcription factors, notably c-JUN, STAT3 and BACH2, which together with VitD receptor shaped the transcriptional response to VitD. Accordingly, VitD did not induce interleukin-10 expression in cells with dysfunctional BACH2 or STAT3. Bronchoalveolar lavage fluid CD4+ T cells of patients with COVID-19 were TH1-skewed and showed de-repression of genes downregulated by VitD, from either lack of substrate (VitD deficiency) and/or abnormal regulation of this system.


Asunto(s)
Interferón gamma/inmunología , Interleucina-10/inmunología , SARS-CoV-2/inmunología , Células TH1/inmunología , Vitamina D/metabolismo , 25-Hidroxivitamina D3 1-alfa-Hidroxilasa/metabolismo , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/metabolismo , Líquido del Lavado Bronquioalveolar/citología , COVID-19/inmunología , COVID-19/patología , Complemento C3a/inmunología , Complemento C3b/inmunología , Humanos , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Activación de Linfocitos/inmunología , Receptores de Calcitriol/metabolismo , Síndrome de Dificultad Respiratoria/inmunología , Síndrome de Dificultad Respiratoria/patología , Síndrome de Dificultad Respiratoria/virología , Factor de Transcripción STAT3/metabolismo , Transducción de Señal/inmunología , Transcripción Genética/genética
7.
Mol Immunol ; 141: 273-279, 2022 01.
Artículo en Inglés | MEDLINE | ID: mdl-34906905

RESUMEN

BACKGROUND: Complement activation plays an important pathogenic role in numerous diseases. The ratio between an activation product and its parent protein is suggested to be more sensitive to detect complement activation than the activation product itself. In the present study we explored whether the ratio between the activation product and the parent protein for C3 (C3bc/C3) and for C5 (sC5b-9/C5) increased the sensitivity to detect complement activation in acute clinical settings compared to the activation product alone. MATERIALS AND METHODS: Samples from patients with acute heart failure following ST-elevated myocardial infarction (STEMI) and from patients with out-of-hospital cardiac arrest (OHCA) were used. C3, C3bc and C5, sC5b-9 were analysed in 629 and 672 patient samples, respectively. Healthy controls (n = 20) served to determine reference cut-off values for activation products and ratios, defined as two SD above the mean. RESULTS: Increased C3bc/C3- and sC5b-9/C5 ratios were vastly dependent on C3bc and sC5b-9. Thus, 99.5 % and 98.1 % of the increased C3bc/C3- and sC5b-9/C5 ratios were solely dependent on increased C3bc and sC5b-9, respectively. Significantly decreased C3 and C5 caused increased ratios in only 3/600 (0.5 %) and 4/319 (1.3 %) samples, respectively. Strong correlations between C3bc and C3bc/C3-ratio and between sC5b-9 and sC5b-9/C5-ratio were found in the STEMI- (r = 0.926 and r = 0.786, respectively) and the OHCA-population (r = 0.908 and r = 0.843, respectively; p < 0.0001 for all). Importantly, sC5b-9 identified worse outcome groups better than sC5b-9/C5-ratio. CONCLUSION: C3bc and sC5b-9 were sensitive markers of complement activation. The ratios of C3bc/C3 and sC5b-9/C5 did not improve detection of complement activation systemically.


Asunto(s)
Activación de Complemento/inmunología , Complemento C3/inmunología , Complemento C3b/inmunología , Complemento C5/inmunología , Complejo de Ataque a Membrana del Sistema Complemento/inmunología , Fragmentos de Péptidos/inmunología , Adulto , Estudios de Casos y Controles , Femenino , Humanos , Masculino , Persona de Mediana Edad
8.
Nat Commun ; 12(1): 7172, 2021 12 09.
Artículo en Inglés | MEDLINE | ID: mdl-34887405

RESUMEN

Complement receptor of immunoglobulin superfamily (CRIg) is expressed on liver macrophages and directly binds complement component C3b or Gram-positive bacteria to mediate phagocytosis. CRIg plays important roles in several immune-mediated diseases, but it is not clear how its pathogen recognition and phagocytic functions maintain homeostasis and prevent disease. We previously associated cytolysin-positive Enterococcus faecalis with severity of alcohol-related liver disease. Here, we demonstrate that CRIg is reduced in liver tissues from patients with alcohol-related liver disease. CRIg-deficient mice developed more severe ethanol-induced liver disease than wild-type mice; disease severity was reduced with loss of toll-like receptor 2. CRIg-deficient mice were less efficient than wild-type mice at clearing Gram-positive bacteria such as Enterococcus faecalis that had translocated from gut to liver. Administration of the soluble extracellular domain CRIg-Ig protein protected mice from ethanol-induced steatohepatitis. Our findings indicate that ethanol impairs hepatic clearance of translocated pathobionts, via decreased hepatic CRIg, which facilitates progression of liver disease.


Asunto(s)
Enterococcus faecalis/inmunología , Infecciones por Bacterias Grampositivas/inmunología , Hepatopatías Alcohólicas/inmunología , Macrófagos/inmunología , Receptores de Complemento 3b/inmunología , Receptores de Complemento/inmunología , Animales , Traslocación Bacteriana , Complemento C3b/inmunología , Enterococcus faecalis/fisiología , Etanol/efectos adversos , Femenino , Tracto Gastrointestinal/microbiología , Infecciones por Bacterias Grampositivas/genética , Infecciones por Bacterias Grampositivas/microbiología , Humanos , Hígado/efectos de los fármacos , Hígado/inmunología , Hígado/microbiología , Hepatopatías Alcohólicas/etiología , Hepatopatías Alcohólicas/genética , Hepatopatías Alcohólicas/microbiología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptores de Complemento/deficiencia , Receptores de Complemento/genética , Receptores de Complemento 3b/genética
9.
Biol Futur ; 72(1): 7-13, 2021 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-34554501

RESUMEN

Integrins are essential membrane proteins that provide a tightly regulated link between the extracellular matrix and the intracellular cytoskeletal network. These cell surface proteins are composed of a non-covalently bound α chain and ß chain. The leukocyte-specific complement receptor 3 (CR3, αMß2, CD11b/CD18) and complement receptor 4 (CR4, αXß2, CD11c/CD18) belong to the family of ß2-integrins. These receptors bind multiple ligands like iC3b, ICAMs, fibrinogen or LPS, thus allowing them to partake in phagocytosis, cellular adhesion, extracellular matrix rearrangement and migration. CR3 and CR4 were generally expected to mediate identical functions due to their structural homology, overlapping ligand specificity and parallel expression on human phagocytes. Despite their similarities, the expression level and function of these receptors differ in a cell-type-specific manner, both under physiological and inflammatory conditions.We investigated comprehensively the individual role of CR3 and CR4 in various functions of human phagocytes, and we proved that there is a "division of labour" between these two receptors. In this review, I will summarize our current knowledge about this area.


Asunto(s)
Antígenos CD18/inmunología , Integrina alfaXbeta2/inmunología , Antígeno de Macrófago-1/inmunología , Fagocitos/inmunología , Fagocitosis/inmunología , Antígeno CD11b/inmunología , Antígeno CD11b/metabolismo , Antígeno CD11c/inmunología , Antígeno CD11c/metabolismo , Antígenos CD18/metabolismo , Complemento C3b/inmunología , Complemento C3b/metabolismo , Humanos , Integrina alfaXbeta2/metabolismo , Antígeno de Macrófago-1/metabolismo , Macrófagos/inmunología , Macrófagos/metabolismo , Fagocitos/metabolismo
10.
Clin J Am Soc Nephrol ; 16(11): 1639-1651, 2021 11.
Artículo en Inglés | MEDLINE | ID: mdl-34551983

RESUMEN

BACKGROUND AND OBJECTIVES: Membranoproliferative GN and C3 glomerulopathy are rare and overlapping disorders associated with dysregulation of the alternative complement pathway. Specific etiologic data for pediatric membranoproliferative GN/C3 glomerulopathy are lacking, and outcome data are based on retrospective studies without etiologic data. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: A total of 80 prevalent pediatric patients with membranoproliferative GN/C3 glomerulopathy underwent detailed phenotyping and long-term follow-up within the National Registry of Rare Kidney Diseases (RaDaR). Risk factors for kidney survival were determined using a Cox proportional hazards model. Kidney and transplant graft survival was determined using the Kaplan-Meier method. RESULTS: Central histology review determined 39 patients with C3 glomerulopathy, 31 with immune-complex membranoproliferative GN, and ten with immune-complex GN. Patients were aged 2-15 (median, 9; interquartile range, 7-11) years. Median complement C3 and C4 levels were 0.31 g/L and 0.14 g/L, respectively; acquired (anticomplement autoantibodies) or genetic alternative pathway abnormalities were detected in 46% and 9% of patients, respectively, across all groups, including those with immune-complex GN. Median follow-up was 5.18 (interquartile range, 2.13-8.08) years. Eleven patients (14%) progressed to kidney failure, with nine transplants performed in eight patients, two of which failed due to recurrent disease. Presence of >50% crescents on the initial biopsy specimen was the sole variable associated with kidney failure in multivariable analysis (hazard ratio, 6.2; 95% confidence interval, 1.05 to 36.6; P<0.05). Three distinct C3 glomerulopathy prognostic groups were identified according to presenting eGFR and >50% crescents on the initial biopsy specimen. CONCLUSIONS: Crescentic disease was a key risk factor associated with kidney failure in a national cohort of pediatric patients with membranoproliferative GN/C3 glomerulopathy and immune-complex GN. Presenting eGFR and crescentic disease help define prognostic groups in pediatric C3 glomerulopathy. Acquired abnormalities of the alternative pathway were commonly identified but not a risk factor for kidney failure.


Asunto(s)
Autoanticuerpos/sangre , Complemento C3/metabolismo , Glomerulonefritis Membranoproliferativa/sangre , Glomerulonefritis Membranoproliferativa/etiología , Fenotipo , Adolescente , Niño , Preescolar , Complemento C3/genética , Complemento C3b/inmunología , Complemento C4/metabolismo , Factor B del Complemento/inmunología , Factor H de Complemento/inmunología , Progresión de la Enfermedad , Femenino , Estudios de Seguimiento , Tasa de Filtración Glomerular , Glomerulonefritis Membranoproliferativa/patología , Glomerulonefritis Membranoproliferativa/terapia , Supervivencia de Injerto , Humanos , Estimación de Kaplan-Meier , Fallo Renal Crónico/etiología , Fallo Renal Crónico/cirugía , Trasplante de Riñón , Masculino , Pronóstico , Modelos de Riesgos Proporcionales , Estudios Prospectivos , Recurrencia , Sistema de Registros , Factores de Riesgo
11.
Am J Physiol Lung Cell Mol Physiol ; 321(2): L485-L489, 2021 08 01.
Artículo en Inglés | MEDLINE | ID: mdl-34231390

RESUMEN

COVID-19, the disease caused by the SARS-CoV-2 virus, can progress to multisystem organ failure and viral sepsis characterized by respiratory failure, arrhythmias, thromboembolic complications, and shock with high mortality. Autopsy and preclinical evidence implicate aberrant complement activation in endothelial injury and organ failure. Erythrocytes express complement receptors and are capable of binding immune complexes; therefore, we investigated complement activation in patients with COVID-19 using erythrocytes as a tool to diagnose complement activation. We discovered enhanced C3b and C4d deposition on erythrocytes in COVID-19 sepsis patients and non-COVID sepsis patients compared with healthy controls, supporting the role of complement in sepsis-associated organ injury. Our data suggest that erythrocytes may contribute to a precision medicine approach to sepsis and have diagnostic value in monitoring complement dysregulation in COVID-19-sepsis and non-COVID sepsis and identifying patients who may benefit from complement targeted therapies.


Asunto(s)
COVID-19/complicaciones , Activación de Complemento/inmunología , Complemento C3b/inmunología , Complemento C4b/inmunología , Eritrocitos/inmunología , Fragmentos de Péptidos/inmunología , Insuficiencia Respiratoria/diagnóstico , Sepsis/diagnóstico , COVID-19/inmunología , COVID-19/virología , Complemento C3b/metabolismo , Complemento C4b/metabolismo , Eritrocitos/metabolismo , Eritrocitos/virología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Fragmentos de Péptidos/metabolismo , Insuficiencia Respiratoria/inmunología , Insuficiencia Respiratoria/metabolismo , Insuficiencia Respiratoria/virología , SARS-CoV-2/aislamiento & purificación , Sepsis/inmunología , Sepsis/metabolismo , Sepsis/virología
12.
Immunobiology ; 226(4): 152106, 2021 07.
Artículo en Inglés | MEDLINE | ID: mdl-34147816

RESUMEN

BACKGROUND: The complement system is a key player in innate immunity and a modulator of the adaptive immune system. Among the three pathways of complement, the alternative pathway (AP) accounts for most of the complement activation. Factor B (FB) is a major protease of the AP, making it a promising target to inhibit the AP activity in conditions of uncontrolled complement activation. METHODS: Based on the data obtained from sequence analysis and conformational changes associated with FB, we expressed and purified a recombinant FB fragment (FBfr). We tested the inhibitory activity of the protein against the AP by in vitro assays. RESULTS: FBfr protein was proven to inhibit the complement AP activity when tested by C3b deposition assay and rabbit erythrocyte hemolytic assay. CONCLUSION: Our recombinant FBfr was able to compete with the native human FB, which allowed it to inhibit the AP activity. This novel compound is a good candidate for further characterization and testing to be used in complement diagnostic tests and as a drug lead in the field of complement therapeutics.


Asunto(s)
Complemento C3b/inmunología , Factor B del Complemento/inmunología , Vía Alternativa del Complemento , Animales , Factor B del Complemento/genética , Eritrocitos , Escherichia coli/genética , Hemólisis , Humanos , Hígado/inmunología , Conejos , Proteínas Recombinantes/inmunología
13.
Front Immunol ; 12: 690821, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-34177949

RESUMEN

Complement factor B (FB) mutant variants are associated with excessive complement activation in kidney diseases such as atypical hemolytic uremic syndrome (aHUS), C3 glomerulopathy and membranoproliferative glomerulonephritis (MPGN). Patients with aHUS are currently treated with eculizumab while there is no specific treatment for other complement-mediated renal diseases. In this study the phenotype of three FB missense variants, detected in patients with aHUS (D371G and E601K) and MPGN (I242L), was investigated. Patient sera with the D371G and I242L mutations induced hemolysis of sheep erythrocytes. Mutagenesis was performed to study the effect of factor D (FD) inhibition on C3 convertase-induced FB cleavage, complement-mediated hemolysis, and the release of soluble C5b-9 from glomerular endothelial cells. The FD inhibitor danicopan abrogated C3 convertase-associated FB cleavage to the Bb fragment in patient serum, and of the FB constructs, D371G, E601K, I242L, the gain-of-function mutation D279G, and the wild-type construct, in FB-depleted serum. Furthermore, the FD-inhibitor blocked hemolysis induced by the D371G and D279G gain-of-function mutants. In FB-depleted serum the D371G and D279G mutants induced release of C5b-9 from glomerular endothelial cells that was reduced by the FD-inhibitor. These results suggest that FD inhibition can effectively block complement overactivation induced by FB gain-of-function mutations.


Asunto(s)
Síndrome Hemolítico Urémico Atípico/inmunología , Activación de Complemento , Factor B del Complemento/genética , Factor D del Complemento/antagonistas & inhibidores , Glomerulonefritis Membranoproliferativa/inmunología , Animales , Síndrome Hemolítico Urémico Atípico/genética , Niño , Convertasas de Complemento C3-C5/inmunología , Complemento C3b/inmunología , Factor B del Complemento/inmunología , Factor D del Complemento/inmunología , Células Endoteliales/inmunología , Eritrocitos , Femenino , Glomerulonefritis Membranoproliferativa/genética , Hemólisis , Humanos , Lactante , Glomérulos Renales/citología , Masculino , Persona de Mediana Edad , Mutación , Fenotipo , Conejos , Ovinos
14.
J Immunol ; 206(12): 3032-3042, 2021 06 15.
Artículo en Inglés | MEDLINE | ID: mdl-34117107

RESUMEN

Complement receptor 3 (CR3, also known as Mac-1, integrin αMß2, or CD11b/CD18) is expressed on a subset of myeloid and certain activated lymphoid cells. CR3 is essential for the phagocytosis of complement-opsonized particles such as pathogens and apoptotic or necrotic cells opsonized with the complement fragment iC3b and, to a lesser extent, C3dg. Although the interaction between the iC3b thioester domain and the ligand binding CR3 αM I-domain is structurally and functionally well characterized, the nature of additional CR3-iC3b interactions required for phagocytosis of complement-opsonized objects remains obscure. In this study, we analyzed the interaction between iC3b and the 150-kDa headpiece fragment of the CR3 ectodomain. Surface plasmon resonance experiments demonstrated a 30 nM affinity of the CR3 headpiece for iC3b compared with 515 nM for the iC3b thioester domain, whereas experiments monitoring binding of iC3b to CR3-expressing cells suggested an affinity of 50 nM for the CR3-iC3b interaction. Small angle x-ray scattering analysis revealed that iC3b adopts an extended but preferred conformation in solution. Upon interaction with CR3, iC3b rearranges to form a compact receptor-ligand complex. Overall, the data suggest that the iC3b-CR3 interaction is of high affinity and relies on minor contacts formed between CR3 and regions outside the iC3b thioester domain. Our results rationalize the more efficient phagocytosis elicited by iC3b than by C3dg and pave the way for the development of specific therapeutics for the treatment of inflammatory and neurodegenerative diseases that do not interfere with the recognition of noncomplement CR3 ligands.


Asunto(s)
Complemento C3b/inmunología , Antígeno de Macrófago-1/inmunología , Humanos
15.
Front Immunol ; 12: 607641, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33936031

RESUMEN

The Sez6 family consists of Sez6, Sez6L, and Sez6L2. Its members are expressed throughout the brain and have been shown to influence synapse numbers and dendritic morphology. They are also linked to various neurological and psychiatric disorders. All Sez6 family members contain 2-3 CUB domains and 5 complement control protein (CCP) domains, suggesting that they may be involved in complement regulation. We show that Sez6 family members inhibit C3b/iC3b opsonization by the classical and alternative pathways with varying degrees of efficacy. For the classical pathway, Sez6 is a strong inhibitor, Sez6L2 is a moderate inhibitor, and Sez6L is a weak inhibitor. For the alternative pathway, the complement inhibitory activity of Sez6, Sez6L, and Sez6L2 all equaled or exceeded the activity of the known complement regulator MCP. Using Sez6L2 as the representative family member, we show that it specifically accelerates the dissociation of C3 convertases. Sez6L2 also functions as a cofactor for Factor I to facilitate the cleavage of C3b; however, Sez6L2 has no cofactor activity toward C4b. In summary, the Sez6 family are novel complement regulators that inhibit C3 convertases and promote C3b degradation.


Asunto(s)
Convertasas de Complemento C3-C5/metabolismo , Complemento C3b/inmunología , Fibrinógeno/metabolismo , Proteínas de la Membrana/metabolismo , Complemento C3b/metabolismo , Proteínas Inactivadoras de Complemento/genética , Proteínas Inactivadoras de Complemento/metabolismo , Vía Alternativa del Complemento/efectos de los fármacos , Vía Clásica del Complemento/efectos de los fármacos , Expresión Génica , Humanos , Inmunohistoquímica , Proteínas de la Membrana/genética , Proteínas de la Membrana/farmacología , Proteolisis , Proteínas Recombinantes de Fusión
17.
Methods Mol Biol ; 2227: 133-139, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33847938

RESUMEN

Autoantibodies against complement proteins are involved in the pathological process of many diseases, including lupus nephritis, C3 glomerulopathies, and atypical hemolytic uremic syndrome. This method describes the detection of autoantibodies targeting the central complement component C3 by ELISA. These autoantibodies (IgG) are detected in up to 30% of the patients with lupus nephritis and more rarely in cases with C3 glomerulopathies. These autoantibodies recognize the active fragment C3b and have overt functional consequences. They enhance the formation of the C3 convertase and prevent the inactivation of C3b by Factor H and complement receptor 1. Moreover, they enhance the deposition of complement activation fragments on activator surfaces, such as apoptotic cells. The data currently available on the relations of anti-C3 autoantibodies with clinical, laboratory, and histological markers for activity of lupus nephritis, as well as the relations of anti-C3 with classical immunological markers for activity of autoimmune process in patients with lupus nephritis, such as hypocomplementemia and high levels of anti-dsDNA, could identify these autoantibodies as a potential marker for evaluation the activity of lupus nephritis. These autoantibodies correlate with the disease severity and can be used to identify patients with lupus nephritis who were prone to flare. Therefore, the detection of such autoantibodies could guide the clinicians to evaluate and predict the severity and to manage the therapy of lupus nephritis.


Asunto(s)
Autoanticuerpos/análisis , Complemento C3b/inmunología , Autoanticuerpos/sangre , Activación de Complemento , Complemento C3/inmunología , Complemento C3/metabolismo , Ensayo de Inmunoadsorción Enzimática/métodos , Glomerulonefritis/sangre , Glomerulonefritis/diagnóstico , Glomerulonefritis/inmunología , Humanos , Nefritis Lúpica/sangre , Nefritis Lúpica/diagnóstico , Nefritis Lúpica/inmunología
18.
Front Immunol ; 12: 789897, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-35069568

RESUMEN

Complement factor I (FI) is a central inhibitor of the complement system, and impaired FI function increases complement activation, contributing to diseases such as age-related macular degeneration (AMD) and atypical hemolytic uremic syndrome (aHUS). Genetic variation in complement factor I (CFI) has been identified in both AMD and aHUS, with more than half of these variants leading to reduced FI secretion levels. For many of the variants with normal FI secretion, however, functional implications are not yet known. Here we studied 11 rare missense variants, with FI secretion levels comparable to wildtype, but a predicted damaging effects based on the Combined Annotation Dependent Depletion (CADD) score. Three variants (p.Pro50Ala, p.Arg339Gln, and p.Ser570Thr) were analyzed in plasma and serum samples of carriers affected by AMD. All 11 variants (nine for the first time in this study) were recombinantly expressed and the ability to degrade C3b was studied with the C3b degradation assay. The amount of degradation was determined by measuring the degradation product iC3b with ELISA. Eight of 11 (73%) mutant proteins (p.Pro50Ala, p.Arg339Gln, p.Ile340Thr, p.Gly342Glu, p.Gly349Arg, p.Arg474Gln, p.Gly487Cys, and p.Gly512Ser) showed significantly impaired C3b degradation, and were therefore classified as likely pathogenic. Our data indicate that genetic variants in CFI with a CADD score >20 are likely to affect FI function, and that monitoring iC3b in a degradation assay is a useful tool to establish the pathogenicity of CFI variants in functional studies.


Asunto(s)
Síndrome Hemolítico Urémico Atípico , Factor I de Complemento , Degeneración Macular , Mutación Missense , Sustitución de Aminoácidos , Síndrome Hemolítico Urémico Atípico/sangre , Síndrome Hemolítico Urémico Atípico/genética , Síndrome Hemolítico Urémico Atípico/inmunología , Complemento C3b/inmunología , Complemento C3b/metabolismo , Factor I de Complemento/genética , Factor I de Complemento/inmunología , Factor I de Complemento/metabolismo , Femenino , Humanos , Degeneración Macular/sangre , Degeneración Macular/genética , Degeneración Macular/inmunología , Masculino
19.
Eur J Immunol ; 51(2): 490-493, 2021 02.
Artículo en Inglés | MEDLINE | ID: mdl-33022775

RESUMEN

We show that the intraerythrocytic stages of the malaria parasite Plasmodium falciparum bind plasminogen and mediate its conversion into plasmin to inactivate parasite-bound C3b. This complement evasion mechanism counteracts terminal complex formation and hence promotes parasite survival in human blood.


Asunto(s)
Complemento C3b/inmunología , Evasión Inmune/inmunología , Malaria Falciparum/inmunología , Plasmodium falciparum/inmunología , Interacciones Huésped-Parásitos/inmunología , Humanos , Malaria Falciparum/parasitología
20.
J Biol Chem ; 296: 100200, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33334893

RESUMEN

Human complement receptor 1 (HuCR1) is a pivotal regulator of complement activity, acting on all three complement pathways as a membrane-bound receptor of C3b/C4b, C3/C5 convertase decay accelerator, and cofactor for factor I-mediated cleavage of C3b and C4b. In this study, we sought to identify a minimal soluble fragment of HuCR1, which retains the complement regulatory activity of the wildtype protein. To this end, we generated recombinant, soluble, and truncated versions of HuCR1 and compared their ability to inhibit complement activation in vitro using multiple assays. A soluble form of HuCR1, truncated at amino acid 1392 and designated CSL040, was found to be a more potent inhibitor than all other truncation variants tested. CSL040 retained its affinity to both C3b and C4b as well as its cleavage and decay acceleration activity and was found to be stable under a range of buffer conditions. Pharmacokinetic studies in mice demonstrated that the level of sialylation is a major determinant of CSL040 clearance in vivo. CSL040 also showed an improved pharmacokinetic profile compared with the full extracellular domain of HuCR1. The in vivo effects of CSL040 on acute complement-mediated kidney damage were tested in an attenuated passive antiglomerular basement membrane antibody-induced glomerulonephritis model. In this model, CSL040 at 20 and 60 mg/kg significantly attenuated kidney damage at 24 h, with significant reductions in cellular infiltrates and urine albumin, consistent with protection from kidney damage. CSL040 thus represents a potential therapeutic candidate for the treatment of complement-mediated disorders.


Asunto(s)
Activación de Complemento , Receptores de Complemento 3b/inmunología , Animales , Línea Celular , Complemento C3b/inmunología , Complemento C4b/inmunología , Femenino , Glomerulonefritis/inmunología , Glomerulonefritis/terapia , Humanos , Ratones , Ratones Endogámicos C57BL , Receptores de Complemento 3b/química , Receptores de Complemento 3b/uso terapéutico , Proteínas Recombinantes/química , Proteínas Recombinantes/inmunología , Proteínas Recombinantes/uso terapéutico
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