RESUMEN
The development of agonists capable of activating the human complement system by binding to the C1 complex presents a novel approach for targeted cell killing. Bispecific nanobodies and Abs can successfully use C1 for this purpose; however, efficacy varies significantly between epitopes, Ab type, and bispecific design. To address this variability, we investigated monomeric agonists of C1 in the form of bispecific nanobodies, which lack Fc domains that lead to oligomerization in Abs. These therefore offer an ideal opportunity to explore the geometric parameters crucial for C1 activation. In this study, we explored the impact of linker length as a metric for Ag and epitope location. DNA nanotechnology and protein engineering allowed us to design linkers with controlled lengths and flexibilities, revealing a critical range of end-to-end distances for optimal complement activation. We discovered that differences in complement activation were not caused by differential C1 activation or subsequent cleavage of C4, but instead impacted C4b deposition and downstream membrane lysis. Considering the importance of Ab class and subclass, this study provides insights into the structural requirements of C1 binding and activation, highlighting linker and hinge engineering as a potential strategy to enhance potency over specific cellular targets. Additionally, using DNA nanotechnology to modify geometric parameters demonstrated the potential for synthetic biology in complement activation. Overall, this research offers valuable insights into the design and optimization of agonists for targeted cell killing through complement activation.
Asunto(s)
Anticuerpos Biespecíficos , Activación de Complemento , Ingeniería de Proteínas , Humanos , Activación de Complemento/inmunología , Anticuerpos Biespecíficos/inmunología , Anticuerpos Biespecíficos/farmacología , Complemento C1/inmunología , Anticuerpos de Dominio Único/inmunología , Epítopos/inmunología , Unión Proteica , Complemento C4b/inmunologíaRESUMEN
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.
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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-2RESUMEN
Human complement C4 is one of the most diverse but heritable effectors for humoral immunity. To help understand the roles of C4 in the defense and pathogenesis of autoimmune and inflammatory diseases, we determined the bases of polymorphisms including the frequent genetic deficiency of C4A and/or C4B isotypes. We demonstrated the diversities of C4A and C4B proteins and their gene copy number variations (CNVs) in healthy subjects and patients with autoimmune disease, such as type 1 diabetes, systemic lupus erythematosus (SLE) and encephalitis. We identified subjects with (a) the fastest migrating C4B allotype, B7, or (b) a deficiency of C4B protein caused by genetic mutation in addition to gene copy-number variation. Those variants and mutants were characterized, sequenced and specific techniques for detection developed. Novel findings were made in four case series. First, the amino acid sequence determinant for C4B7 was likely the R729Q variation at the anaphylatoxin-like region. Second, in healthy White subject MS630, a C-nucleotide deletion at codon-755 led to frameshift mutations in his single C4B gene, which was a private mutation. Third, in European family E94 with multiplex lupus-related mortality and low serum C4 levels, the culprit was a recurrent haplotype with HLA-A30, B18 and DR7 that segregated with two defective C4B genes and identical mutations at the donor splice site of intron-28. Fourth, in East-Asian subject E133P with anti-NMDA receptor encephalitis, the C4B gene had a mutation that changed tryptophan-660 to a stop-codon (W660x), which was present in a haplotype with HLA-DRB1*04:06 and B*15:27. The W660x mutation is recurrent among East-Asians with a frequency of 1.5% but not detectable among patients with SLE. A meticulous annotation of C4 sequences revealed clusters of variations proximal to sites for protein processing, activation and inactivation, and binding of interacting molecules.
Asunto(s)
Enfermedades Autoinmunes/genética , Complemento C4b/genética , Variaciones en el Número de Copia de ADN , Dosificación de Gen , Inmunidad Humoral/genética , Mutación , Enfermedades Autoinmunes/diagnóstico , Enfermedades Autoinmunes/etnología , Enfermedades Autoinmunes/inmunología , Estudios de Casos y Controles , Complemento C4a/deficiencia , Complemento C4a/genética , Complemento C4a/inmunología , Complemento C4b/deficiencia , Complemento C4b/inmunología , Femenino , Frecuencia de los Genes , Predisposición Genética a la Enfermedad , Humanos , Masculino , FenotipoRESUMEN
Objectives: This study aimed to determine the prevalence and localization of complement factor C4d in renal biopsies from patients with lupus nephritis (LN), as well as its associations with the disease's clinico-pathological features. The correlation between arteriolar C4d deposition and renal microvascular lesions (RVLs) was further analyzed. Methods: A total of 325 biopsy-proven LN patients were enrolled, and their clinico-pathological data were collected. C4d staining of renal biopsies was performed by immunohistochemistry. The associations between C4d deposition and the clinico-pathological features were further analyzed. Results: C4d deposition was present in most (98.8%) renal specimens in our cohort. These deposits were localized in the glomeruli (98.2%), tubular basement membrane (TBM) (43.7%), arterioles (31.4%), and peritubular capillary (33.8%). Patients with TBM C4d staining had higher disease activity (measured with the Systemic Lupus Erythematous Disease Activity Index) and higher National Institutes of Health pathological activity and chronicity indices (all P < 0.01). Patients with arteriolar C4d deposition were more likely to develop RVLs (91.2%) compared to those with no arteriolar C4d deposition (78.0%; P = 0.004), especially with two or more types of RVLs (P < 0.001). During the mean follow-up of 55.8 months, arteriolar C4d was related to worse renal outcomes [hazard ration (HR): 2.074, 95% confidence interval (CI) 1.056-4.075, P = 0.034]. Multivariate Cox hazard analysis showed that co-deposition of arteriolar C4d and C3c was an independent risk factor (HR: 3.681, 95% CI 1.519-8.921, P = 0.004) for predicting renal outcomes. Conclusions: C4d deposition was common in renal tissues from LN patients. TBM C4d deposition was related to the disease activity, and arteriolar C4d deposition was associated with RVLs and worse renal outcomes.
Asunto(s)
Complemento C4b/inmunología , Complemento C4b/metabolismo , Susceptibilidad a Enfermedades , Nefritis Lúpica/etiología , Nefritis Lúpica/metabolismo , Fragmentos de Péptidos/inmunología , Fragmentos de Péptidos/metabolismo , Adulto , Biomarcadores , Biopsia , Complemento C1q/inmunología , Complemento C1q/metabolismo , Complemento C3c/inmunología , Complemento C3c/metabolismo , Femenino , Técnica del Anticuerpo Fluorescente , Humanos , Inmunohistoquímica , Glomérulos Renales/inmunología , Glomérulos Renales/metabolismo , Glomérulos Renales/patología , Nefritis Lúpica/diagnóstico , Nefritis Lúpica/mortalidad , Masculino , Persona de Mediana Edad , Pronóstico , Adulto JovenRESUMEN
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/virologíaRESUMEN
Immunoglobulin A nephropathy (IgAN) is considered as mesangiopathy since it initiates in the mesangium; however, other glomerular components are involved and the glomerular capillary wall offers the first contact to circulating macromolecular IgA1. Acute and active forms of IgAN are associated with endocapillary hypercellularity and vascular damage of various degrees, in severe cases with microangiopathy (MA) without or with thrombosis [thrombotic microangiopathy (TMA)]. Vascular damage activates complement and coagulation cascades. A defective complement regulation has recently been detected in active and progressive cases of IgAN. C4d deposits in renal biopsies have been found to be an early risk factor. These observations have raised interest in manifestation of MA and TMA in progressive cases of IgAN. MA-TMA lesions have been found in various percentages (2-53%) of patients with IgAN according to patients' selection and pathology definition of TMA. The association with hypertension (HTN) was so strong that it led to the hypothesis that MA/TMA in IgAN was a mere consequence of severe HTN. Old and new clinical and experimental data indicate that in IgAN the interaction of the glomerular capillary wall with immune reactants and complement uncontrolled activation leading to C4b deposits favours the development of MA-TMA, which plays a role in progression and renal function decline. The central role of complement activation is relevant also for the new therapeutic interventions offered by the pharma.
Asunto(s)
Activación de Complemento/inmunología , Complemento C4b/inmunología , Glomerulonefritis por IGA/patología , Glomérulos Renales/patología , Microangiopatías Trombóticas/patología , Enfermedades Vasculares/patología , Glomerulonefritis por IGA/inmunología , Humanos , Glomérulos Renales/inmunología , Microangiopatías Trombóticas/inmunología , Enfermedades Vasculares/inmunologíaRESUMEN
Solid tissue transplant is a growing medical need that is further complicated by a limited donor organ supply. Acute and chronic rejection occurs in nearly all transplants and reduces long-term graft survival, thus increasing the need for repeat transplantation. Viruses have evolved highly adapted responses designed to evade the host's immune defenses. Immunomodulatory proteins derived from viruses represent a novel class of potential therapeutics that are under investigation as biologics to attenuate immune-mediated rejection and damage. These immune-modulating proteins have the potential to reduce the need for traditional posttransplant immune suppressants and improve graft survival. The myxoma virus-derived protein M-T7 is a promising biologic that targets chemokine and glycosaminoglycan pathways central to kidney transplant rejection. Orthotopic transplantations in mice are prohibitively difficult and costly and require a highly trained microsurgeon to successfully perform the procedure. Here we describe a kidney-to-kidney subcapsular transplant model as a practical and simple method for studying transplant rejection, a model that requires fewer mice. One kidney can be used as a donor for transplants into six or more recipient mice. Using this model there is lower morbidity, pain, and mortality for the mice. Subcapsular kidney transplantation provides a first step approach to testing virus-derived proteins as new potential immune-modulating therapeutics to reduce transplant rejection and inflammation.
Asunto(s)
Antiinflamatorios/farmacología , Rechazo de Injerto/prevención & control , Supervivencia de Injerto , Factores Inmunológicos/farmacología , Trasplante de Riñón/métodos , Myxoma virus/química , Proteínas Virales/farmacología , Animales , Antiinflamatorios/inmunología , Antiinflamatorios/metabolismo , Biomarcadores/análisis , Quimiocinas/biosíntesis , Complemento C4b/genética , Complemento C4b/inmunología , Femenino , Expresión Génica , Rechazo de Injerto/genética , Rechazo de Injerto/inmunología , Rechazo de Injerto/patología , Factores Inmunológicos/biosíntesis , Factores Inmunológicos/inmunología , Riñón/inmunología , Riñón/cirugía , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Fragmentos de Péptidos/genética , Fragmentos de Péptidos/inmunología , Receptores de Interferón/biosíntesis , Receptores de Interferón/inmunología , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/inmunología , Proteínas Recombinantes/farmacología , Trasplante Homólogo , Proteínas Virales/biosíntesis , Proteínas Virales/inmunologíaRESUMEN
BACKGROUND: Glomerulonephritis (GN), including post-transplant IgAN (post-Tx IgAN) is an important contributor to decreased long-term allograft survival. The immunopathological detection of the complement degradation product C4d in glomeruli (C4dG) has been recently described as a risk factor in native kidney IgAN, however little is known about C4dG deposition in post-Tx IgAN. We hypothesized that glomerular C4d may indicate a more aggressive disease course and worse allograft survival in patients with post-Tx IgAN. METHODS: In this retrospective study we assessed the presence and clinical relevance of C4dG in patients with post-transplant IgAN. We analyzed 885 renal allograft recipients, including 84 patients with post-transplant GN. All patients were transplanted between January 1999 and April 2006 and underwent at least one biopsy for differnt causes. The primary endpoint was death-censored graft survival, with a median follow-up of 9.6 (IQR 3.8-13.2) years. RESULTS: The prevalence of post-Tx GN was 9.5%. Twenty-seven patients with post-Tx IgAN were included. C4dG positive patients (N = 18, 66.7%) had significantly worse allograft survival compared to C4dG negative post-Tx IgAN patients and patients without post-Tx IgAN [C4dG positive: 27.8% vs. 55.6% and 66.0%; log-rank: p = 0.01]. C4dG remained a significant risk factor (HR 2.22, 95% CI 1.27-3.87) for allograft loss even after adjustment for T cell mediated rejection (TCMR) and antibody mediated rejection. CONCLUSION: Glomerular C4d deposition is an independent risk factor for worse graft-survival in patients with post-Tx IgAN, even after adjusting for other risk factors such as antibody mediated rejection. Assessment of glomerular C4d deposition may provide a valuable prognostic risk assessment tool to identify high risk patients in post-Tx IgAN.
Asunto(s)
Complemento C4b , Glomerulonefritis por IGA , Supervivencia de Injerto , Trasplante de Riñón , Adulto , Aloinjertos , Antagonistas de Receptores de Angiotensina , Inhibidores de la Enzima Convertidora de Angiotensina , Biopsia , Complemento C4b/inmunología , Femenino , Glomerulonefritis por IGA/inmunología , Rechazo de Injerto , Humanos , Masculino , Persona de Mediana Edad , Estudios RetrospectivosRESUMEN
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 terapéuticoRESUMEN
OBJECTIVES: To describe the histopathologic and immunophenotypic features of a temporary splenic allograft exposed to massive donor-specific antibody (DSA) insult. METHODS: A human cadaveric donor splenic allograft was temporarily transplanted in a highly sensitized patient with the intention of removing DSA before intestinal transplantation from the same donor. Before splenic transplant, the patient had several preformed cytotoxic DSAs that resulted in positive flow cytometric and complement-dependent cytotoxicity crossmatch. The splenic allograft was removed before intestinal transplantation and evaluated by H&E and immunohistochemical stains. RESULTS: Explanted donor splenic allograft showed several histopathologic changes: expanded red pulp secondary to congestion and marked neutrophilic and macrophage infiltration in the sinusoids, numerous neutrophilic microabscesses, and focal capillaritis. The C4d and IgG immunohistochemical stains were diffusely positive in the endothelial lining of the capillaries and sinusoidal lining, indicating diffuse IgG deposition and complement activation. CONCLUSIONS: We propose that the noted changes are features of splenic acute antibody-mediated rejection (AMR). Additional cases are required to determine all the features of splenic AMR. To our knowledge, this is the first report of histopathologic changes in donor spleen after exposure to DSA for a short duration.
Asunto(s)
Rechazo de Injerto/inmunología , Prueba de Histocompatibilidad , Bazo/inmunología , Bazo/patología , Donantes de Tejidos , Adulto , Biopsia , Complemento C4b/inmunología , Prueba de Histocompatibilidad/métodos , Humanos , Isoanticuerpos/inmunología , Trasplante de Riñón/métodos , Masculino , Fragmentos de Péptidos/inmunología , Bazo/trasplanteRESUMEN
In complement-driven thrombotic microangiopathies, failure to regulate complement activation leads to end-organ damage. The modified Ham (mHam) test measures complement-mediated killing of a nucleated cell in vitro but lacks a confirmatory assay and reliable positive controls. We demonstrate that C5b-9 accumulation on the surface of TF1 PIGAnull cells correlates with cell killing in the mHam. We also show that Sialidase treatment of cells or addition of Shiga toxin 1 to human serum serve as a more reliable positive control for the mHam than cobra venom factor or lipopolysaccharide. Simultaneously performing the mHam and measuring C5b-9 accumulation either in GVB++ or GVB0 MgEGTA buffer with the addition of complement pathway specific inhibitors (anti-C5 antibody or a factor D inhibitor, ACH-145951) can be used to localize defects in complement regulation. As more targeted complement inhibitors become available, these assays may aid in the selection of personalized treatments for patients with complement-mediated diseases.
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Síndrome Antifosfolípido/inmunología , Síndrome Hemolítico Urémico Atípico/inmunología , Activación de Complemento/efectos de los fármacos , Inactivadores del Complemento/farmacología , Adulto , Bioensayo , Línea Celular Tumoral , Complemento C3c/inmunología , Complemento C4b/inmunología , Complejo de Ataque a Membrana del Sistema Complemento/inmunología , Venenos Elapídicos/farmacología , Femenino , Humanos , Lipopolisacáridos/farmacología , Masculino , Persona de Mediana Edad , Neuraminidasa/farmacología , Fragmentos de Péptidos/inmunología , Toxina Shiga I/farmacologíaRESUMEN
Antibody-mediated rejection (ABMR) in organ transplantation has been recognized as the main cause of graft rejection. Binding of donor-specific HLA antibody (DSA) and A/B blood type antibody on graft endothelial cells causes complement-dependent tissue damage. C4d, a product of the complement cascade, has long been an indicator of graft tissue damage in graft endothelial cells. By contrast, recent evidences indicated histological findings of ABMR without C4d deposition in many cases and Banff classification criteria included a category of C4d-negative ABMR. Several mechanisms have been proposed for complement-independent tissue injury in the presence of DSA. It is well known that activated monocytes and macrophages infiltrate into graft tissues. The inflammatory environment triggered by the binding of DSA to endothelial cells alone can induce an allo-reaction of CD4 T-cells via graft endothelial cell HLA-class II. Accommodation is a condition that no rejections occur even in the presence of an antibody against donor organs and becomes attracting considerable attention as a therapeutic strategy to acquire long-term survival of the transplanted organs. Several recent publications have suggested some mechanistic insights about graft accommodation, including the upregulation of antioxidant, anti-apoptotic, and complement regulatory proteins genes via activation of PI3K/AKT survival signal or inactivation of extracellular signal-regulated protein kinase pro-inflammatory signals after DSA and anti-A/B antibody ligation on endothelial cells.
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Rechazo de Injerto/etiología , Antígenos HLA/inmunología , Isoanticuerpos/inmunología , Trasplante de Órganos/efectos adversos , Complemento C4b/inmunología , Células Endoteliales/inmunología , Rechazo de Injerto/inmunología , Humanos , Fragmentos de Péptidos/inmunología , Donantes de TejidosAsunto(s)
Betacoronavirus/inmunología , Complemento C4b/inmunología , Infecciones por Coronavirus/complicaciones , Livedo Reticularis/diagnóstico , Fragmentos de Péptidos/inmunología , Neumonía Viral/complicaciones , Adolescente , Betacoronavirus/aislamiento & purificación , Biopsia , COVID-19 , Niño , Complemento C4b/análisis , Infecciones por Coronavirus/inmunología , Infecciones por Coronavirus/virología , Femenino , Humanos , Livedo Reticularis/inmunología , Masculino , Pandemias , Fragmentos de Péptidos/análisis , Neumonía Viral/inmunología , Neumonía Viral/virología , SARS-CoV-2 , Hermanos , Piel/inmunología , Piel/patología , Factores de TiempoRESUMEN
Objectives: As accumulating data supporting the potential role of the complement system in the pathogenesis of diabetic kidney disease (DKD), the present study aimed to explore the association of glomerular complement C4c deposition with the baseline clinicopathological characteristics and the prognosis of DKD in type 2 diabetes (T2DM) patients. Methods: A total of 79 T2DM patients with biopsy-proven DKD were enrolled. Clinicopathological features and renal outcomes were compared between groups divided by the glomerular C4c deposition patterns and median values of serum C4. Renal outcomes were defined by doubling of serum creatinine level or progression to end-stage renal disease (ESRD). A Cox proportional hazards model was employed to identify the risk factors associated with renal events. Results: Patients with glomerular C4c deposition had worse renal insufficiency than those without C4c deposits, along with higher 24-h urinary protein, triglyceride, but lower serum albumin and higher interstitial inflammation score. Besides, serum C4 levels positively correlated with urinary protein and serum C3 levels. During 21.85 ± 16.32 months of follow-up, Kaplan-Meier curve analysis showed significantly faster deterioration of renal function for patients with positive glomerular C4c deposition as well as higher levels of serum C4. More specifically, more than 50% of the patients with glomerular C4c had co-deposition of C3c or C1q, and patients with glomerular complement complex of C4c and one or two of C3/C1q deposition had more severe proteinuria and a higher rate of DKD progression than those with negative C4c deposits. The univariate Cox regression indicated that factors of combined serum and glomerular C4, urinary protein, serum creatinine, serum C3, combined glomerular C4c and IgM and interstitial inflammation were associated with an increased risk of DKD, but only glomerular C4c intensity (HR 1.584, 95% CI [1.001, 2.508], p = 0.0497), as well as baseline age and diabetic neuropathy, were independent risk factors for renal survival by the multivariate Cox analysis. Conclusions: Glomerular C4c deposition was associated with deteriorated renal function and outcomes in patients with T2DKD. Glomerular C4c deposition was an independent risk factor for DKD progression.
Asunto(s)
Complemento C4b/inmunología , Diabetes Mellitus Tipo 2/complicaciones , Nefropatías Diabéticas/etiología , Nefropatías Diabéticas/metabolismo , Glomérulos Renales/inmunología , Glomérulos Renales/metabolismo , Fragmentos de Péptidos/inmunología , Adulto , Biomarcadores , Biopsia , Complemento C3/inmunología , Complemento C3/metabolismo , Nefropatías Diabéticas/mortalidad , Nefropatías Diabéticas/patología , Progresión de la Enfermedad , Susceptibilidad a Enfermedades , Femenino , Técnica del Anticuerpo Fluorescente , Humanos , Estimación de Kaplan-Meier , Pruebas de Función Renal , Glomérulos Renales/patología , Masculino , Persona de Mediana Edad , Pronóstico , Curva ROCRESUMEN
The classical and lectin pathways of the complement system are important for the elimination of pathogens and apoptotic cells and stimulation of the adaptive immune system. Upon activation of these pathways, complement component C4 is proteolytically cleaved, and the major product C4b is deposited on the activator, enabling assembly of a C3 convertase and downstream alternative pathway amplification. Although excessive activation of the lectin and classical pathways contributes to multiple autoimmune and inflammatory diseases and overexpression of a C4 isoform has recently been linked to schizophrenia, a C4 inhibitor and structural characterization of the convertase formed by C4b is lacking. In this study, we present the nanobody hC4Nb8 that binds with picomolar affinity to human C4b and potently inhibits in vitro complement C3 deposition through the classical and lectin pathways in human serum and in mouse serum. The crystal structure of the C4b:hC4Nb8 complex and a three-dimensional reconstruction of the C4bC2 proconvertase obtained by electron microscopy together rationalize how hC4Nb8 prevents proconvertase assembly through recognition of a neoepitope exposed in C4b and reveals a unique C2 conformation compared with the alternative pathway proconvertase. On human induced pluripotent stem cell-derived neurons, the nanobody prevents C3 deposition through the classical pathway. Furthermore, hC4Nb8 inhibits the classical pathway-mediated immune complex delivery to follicular dendritic cells in vivo. The hC4Nb8 represents a novel ultrahigh-affinity inhibitor of the classical and lectin pathways of the complement cascade under both in vitro and in vivo conditions.
Asunto(s)
Convertasas de Complemento C3-C5 de la Vía Clásica/metabolismo , Complemento C3/metabolismo , Complemento C4b/metabolismo , Células Madre Pluripotentes Inducidas/fisiología , Neuronas/fisiología , Esquizofrenia/metabolismo , Anticuerpos de Dominio Único/metabolismo , Animales , Afinidad de Anticuerpos , Complejo Antígeno-Anticuerpo/metabolismo , Diferenciación Celular , Células Cultivadas , Activación de Complemento , Complemento C4b/genética , Complemento C4b/inmunología , Humanos , Ratones , Ratones Noqueados , Multimerización de Proteína , Regulación hacia ArribaRESUMEN
OBJECTIVE: To evaluate the frequency of cell-bound complement activation products (CB-CAPs) as a marker of complement activation in patients with suspected systemic lupus erythematosus (SLE) and the usefulness of this biomarker as a predictor of the evolution of probable SLE into SLE as classified by the American College of Rheumatology (ACR) criteria. METHODS: Patients in whom SLE was suspected by lupus experts and who fulfilled 3 ACR classification criteria for SLE (probable SLE) were enrolled, along with patients with established SLE as classified by both the ACR and the Systemic Lupus International Collaborating Clinics (SLICC) criteria, patients with primary Sjögren's syndrome (SS), and patients with other rheumatic diseases. Individual CB-CAPs were measured by flow cytometry, and positivity rates were compared to those of commonly assessed biomarkers, including serum complement proteins (C3 and C4) and autoantibodies. The frequency of a positive multianalyte assay panel (MAP), which includes CB-CAPs, was also evaluated. Probable SLE cases were followed up prospectively. RESULTS: The 92 patients with probable SLE were diagnosed more recently than the 53 patients with established SLE, and their use of antirheumatic medications was lower. At the enrollment visit, more patients with probable SLE were positive for CB-CAPs (28%) or MAP (40%) than had low complement levels (9%) (P = 0.0001 for each). In probable SLE, MAP scores of >0.8 at enrollment predicted fulfillment of a fourth ACR criterion within 18 months (hazard ratio 3.11, P < 0.01). CONCLUSION: Complement activation occurs in some patients with probable SLE and can be detected with higher frequency by evaluating CB-CAPs and MAP than by assessing traditional serum complement protein levels. A MAP score above 0.8 predicts transition to classifiable SLE according to ACR criteria.
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Activación de Complemento/inmunología , Complemento C3/inmunología , Complemento C4b/inmunología , Lupus Eritematoso Sistémico/inmunología , Fragmentos de Péptidos/inmunología , Adulto , Anciano , Anciano de 80 o más Años , Anticuerpos Antiproteína Citrulinada/inmunología , Anticuerpos Antinucleares/inmunología , Autoanticuerpos/inmunología , Linfocitos B/metabolismo , Estudios de Casos y Controles , Complemento C4/inmunología , Complemento C4b/metabolismo , Progresión de la Enfermedad , Eritrocitos/metabolismo , Femenino , Citometría de Flujo , Humanos , Masculino , Persona de Mediana Edad , Fragmentos de Péptidos/metabolismo , Estudios Prospectivos , Enfermedades Reumáticas/inmunología , Factor Reumatoide/inmunología , Síndrome de Sjögren/inmunología , Adulto JovenRESUMEN
Urticarial vasculitis (UV) is a clinicopathologic entity characterized by persistent urticarial lesions with biopsy features of vasculitis. Currently, only certain clinical features such as arthralgia and serum complement concentrations are used to identify UV patients at risk for an underlying systemic disease. Hypocomplementemic urticarial vasculitis (HUV) is in contrast to normocomplementemic urticarial vasculitis (NUV), strongly associated with underlying systemic disease, especially systemic lupus erythematosus (SLE). The aim of this study was to find specific histopathological features associated with HUV and underlying systemic disease in UV. In addition, the use of complement C4d deposition in skin biopsies was evaluated as a diagnostic adjunct for HUV- and UV-associated systemic disease. In this retrospective study, the clinical, histopathological, and immunohistological (C4d) features of 43 patients with UV were compared between HUV and NUV and analyzed for association with UV-associated systemic disease. Eight of 43 patients with UV (19%) had hypocomplementemia. Patients with HUV showed a significantly higher number of perivascular neutrophils and lower number of eosinophils compared to NUV. Of all histopathological features, alignment of neutrophils along the dermal-epidermal junction (DEJ) and dermal granular C4d deposition were found to be strongly associated with HUV and underlying SLE. This study shows that both the alignment of neutrophils along the DEJ and dermal C4d deposition are strongly associated with HUV and SLE. Therefore, these (immuno)histopathological features can be used as an easy diagnostic adjunct for early detection of underlying systemic disease in UV.
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Complemento C4b/metabolismo , Lupus Eritematoso Sistémico/complicaciones , Neutrófilos/patología , Fragmentos de Péptidos/metabolismo , Vasculitis Leucocitoclástica Cutánea/inmunología , Vasculitis Leucocitoclástica Cutánea/patología , Adulto , Autoanticuerpos/inmunología , Autoantígenos/inmunología , Complemento C1q/inmunología , Complemento C4b/inmunología , Femenino , Humanos , Lupus Eritematoso Sistémico/inmunología , Masculino , Persona de Mediana Edad , Neutrófilos/inmunología , Fragmentos de Péptidos/inmunología , Estudios RetrospectivosRESUMEN
BACKGROUND: Linear C4d staining in the peritubular capillaries is considered a sensitive and useful marker of active or chronic active antibody-mediated rejection (ABMR) in transplanted kidneys. However, the diagnostic significance of glomerular C4d deposits (gC4d) is still undetermined. The aim of this study is to evaluate the association of gC4d with clinicopathologic features and to assess its diagnostic value. METHODS: From 2013 to 2018, a total of 158 cases of allograft kidney biopsy specimens were obtained from the Korea University Anam Hospital. The histologic features were evaluated according to the Banff classification. The gC4d were determined through immunohistochemical analyses and classified based on scores of 0 to 3 according to the extent of gC4d. RESULTS: A total of 73 cases (46.2%) showed gC4d, and 37 cases (23.4%), 23 cases (14.6%), and 13 cases (8.2%) were classified with a score of 1+, 2+, and 3+, respectively. The gC4d showed a significant correlation with antibody-associated histologic lesions, including peritubular capillaritis, glomerulitis, and transplant glomerulopathy (P < .001). However, gC4d showed no significant association with cell-mediated injuries such as tubulitis, interstitial inflammation, acute tubular necrosis, and thrombotic microangiopathy. Although positive gC4d alone was associated with nonspecific findings without ABMR, most cases of gC4d combined with glomerulitis or transplant glomerulopathy showed typical histologic features of ABMR, clinically with higher antibody titers and severe functional deterioration. CONCLUSIONS: Glomerular C4d deposits may be an alternate useful marker in the diagnosis of active or chronic active ABMR when combined with histologic features of glomerular lesions.
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Complemento C4b/análisis , Glomerulonefritis/inmunología , Rechazo de Injerto/diagnóstico , Trasplante de Riñón/efectos adversos , Fragmentos de Péptidos/análisis , Complicaciones Posoperatorias/inmunología , Enfermedad Aguda , Adulto , Anticuerpos/inmunología , Biomarcadores/análisis , Capilares/patología , Enfermedad Crónica , Complemento C4b/inmunología , Femenino , Glomerulonefritis/patología , Rechazo de Injerto/inmunología , Humanos , Riñón/inmunología , Glomérulos Renales/inmunología , Masculino , Persona de Mediana Edad , Fragmentos de Péptidos/inmunología , Complicaciones Posoperatorias/patología , República de Corea , Microangiopatías Trombóticas/inmunología , Microangiopatías Trombóticas/patología , Trasplante HomólogoRESUMEN
BACKGROUND: Enterococcus faecalis is considered to be the most important species of enterococci responsible for blood stream infections in critically ill patients. In blood, the complement system is activated via the classical pathway (CP), the lectin pathway (LP), or the alternative pathway (AP), and it plays a critical role in opsonophagocytosis of bacteria including E faecalis. METHODS: In a mouse model of enterococcus peritonitis, BALB-C mice were challenged with a high dose of E faecalis 12 hours after intraperitoneal administration of anti-Factor H (FH) antibodies or isotype control. Four hours later, control mice developed higher bacterial burden in blood and organs compared with mice treated with anti-FH antibodies. RESULTS: We demonstrate that complement recognition molecules C1q, CL-11, and murine ficolin-A bind the enterococcus and drive the CP and the LP in human and mouse. We further describe that E faecalis evades the AP by recruitment of FH on its surface. Our results show a strong C3b deposition on E faecalis via both the CP and the LP but not through the AP. CONCLUSIONS: These findings indicate that E faecalis avoids the complement phagocytosis by the AP via sequestering complement FH from the host blood.
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Factor H de Complemento/inmunología , Proteínas del Sistema Complemento/inmunología , Enterococcus faecalis/inmunología , Infecciones por Bacterias Grampositivas/inmunología , Peritonitis/inmunología , Animales , Anticuerpos Antibacterianos/sangre , Complemento C3b/inmunología , Complemento C4b/inmunología , Lectina de Unión a Manosa de la Vía del Complemento/inmunología , Modelos Animales de Enfermedad , Humanos , Lectinas , Ratones , Ratones Endogámicos BALB C , Peritonitis/microbiología , Peritonitis/patología , Fagocitosis/inmunología , FicolinasRESUMEN
C3 Glomerulopathy (C3G) is a renal disease mediated primarily by dysregulation of the alternative pathway of complement. Complement is the cornerstone of innate immunity. It targets infectious microbes for destruction, clears immune complexes, and apoptotic cells from the circulation, and augments the humoral response. In C3G, this process becomes dysregulated, which leads to the deposition of complement proteins-including complement component C3-in the glomerular basement membrane of the kidney. Events that trigger complement are typically environmental insults like infections. Once triggered, in patients who develop C3G, complement activity is sustained by a variety of factors, including rare or novel genetic variants in complement genes and autoantibodies that alter normal complement protein function and/or regulation. Herein, we review two such autoantibodies, one to Factor B and the other to C4b2a, the C3 convertase of the classical, and lectin pathways. These two types of autoantibodies are identified in a small fraction of C3G patients and contribute marginally to the C3G phenotype.