RESUMEN
Thoracic aortic aneurysm/dissection (TAAD) is a lethal vascular disease, and several pathological factors participate in aortic medial degeneration. We previously discovered that the complement C3a-C3aR axis in smooth muscle cells promotes the development of thoracic aortic dissection (TAD) through regulation of matrix metalloproteinase 2. However, discerning the specific complement pathway that is activated and elucidating how inflammation of the aortic wall is initiated remain unknown. We ascertained that the plasma levels of C3a and C5a were significantly elevated in patients with TAD and that the levels of C3a, C4a, and C5a were higher in acute TAD than in chronic TAD. We also confirmed the activation of the complement in a TAD mouse model. Subsequently, knocking out Cfb (Cfb) or C4 in mice with TAD revealed that the alternative pathway and Cfb played a significant role in the TAD process. Activation of the alternative pathway led to generation of the anaphylatoxins C3a and C5a, and knocking out their receptors reduced the recruitment of inflammatory cells to the aortic wall. Moreover, we used serum from wild-type mice or recombinant mice Cfb as an exogenous source of Cfb to treat Cfb KO mice and observed that it exacerbated the onset and rupture of TAD. Finally, we knocked out Cfb in the FBN1C1041G/+ Marfan-syndrome mice and showed that the occurrence of TAA was reduced. In summary, the alternative complement pathway promoted the development of TAAD by recruiting infiltrating inflammatory cells. Targeting the alternative pathway may thus constitute a strategy for preventing the development of TAAD.NEW & NOTEWORTHY The alternative complement pathway promoted the development of TAAD by recruiting infiltrating inflammatory cells. Targeting the alternative pathway may thus constitute a strategy for preventing the development of TAAD.
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Aneurisma de la Aorta Torácica , Disección Aórtica , Azidas , Desoxiglucosa/análogos & derivados , Humanos , Ratones , Animales , Vía Alternativa del Complemento , Metaloproteinasa 2 de la Matriz , Aneurisma de la Aorta Torácica/genética , Aneurisma de la Aorta Torácica/metabolismo , Aneurisma de la Aorta Torácica/patología , Disección Aórtica/genética , InflamaciónRESUMEN
Complement constitutes a major part of the innate immune system. The study of complement in human health has historically focused on infection risks associated with complement protein deficiencies; however, recent interest in the field has focused on overactivation of complement as a cause of immune injury and the development of anticomplement therapies to treat human diseases. The kidneys are particularly sensitive to complement injury, and anticomplement therapies for several kidney diseases have been investigated. Overactivation of complement can result from loss-of-function mutations in complement regulators; gain-of-function mutations in key complement proteins such as C3 and factor B; or autoantibody production, infection, or tissue stresses, such as ischemia and reperfusion, that perturb the balance of complement activation and regulation. Here, we provide a high-level review of the status of anticomplement therapies, with an emphasis on the transition from rare diseases to more common kidney diseases.
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Enfermedades Renales , Enfermedades Raras , Humanos , Enfermedades Raras/tratamiento farmacológico , Enfermedades Raras/genética , Proteínas Inactivadoras de Complemento , Enfermedades Renales/tratamiento farmacológico , Enfermedades Renales/genética , MutaciónRESUMEN
Complement factor D (FD) is a rate-limiting enzyme of the alternative pathway (AP). Recent studies have suggested that it is synthesized as an inactive precursor and that its conversion to enzymatically active FD is catalyzed by mannan-binding lectin-associated serine protease 3 (MASP3). However, whether MASP3 is essential for AP complement activity remains uncertain. It has been shown that Masp1/3 gene knockout did not prevent AP complement overactivation in a factor H-knockout mouse, and a human patient lacking MASP3 still retained AP complement activity. In this study, we have assessed AP complement activity in a Masp3-knockout mouse generated by CRISPR/Cas9 editing of the Masp1/3 gene. We confirmed specific Masp3 gene inactivation by showing intact MASP1 protein expression and absence of mature FD in the mutant mice. Using several assays, including LPS- and zymosan-induced C3b deposition and rabbit RBC lysis tests, we detected plasma concentration-dependent AP complement activity in Masp3 gene-inactivated mice. Thus, although not measurable in 5% plasma, significant AP complement activity was detected in 20-50% plasma of Masp3 gene-inactivated mice. Furthermore, whereas FD gene deletion provided more than 90% protection of CD55/Crry-deficient RBCs from AP complement-mediated extravascular hemolysis, Masp3 gene deletion only provided 30% protection in the same study. We also found pro-FD to possess intrinsic catalytic activity, albeit at a much lower level than mature FD. Our data suggest that MASP3 deficiency reduces but does not abrogate AP complement activity and that this is explained by intrinsic pro-FD activity, which can be physiologically relevant in vivo.
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Lectina de Unión a Manosa , Serina Proteasas Asociadas a la Proteína de Unión a la Manosa , Animales , Humanos , Ratones , Conejos , Factor D del Complemento/metabolismo , Vía Alternativa del Complemento/fisiología , Lectina de Unión a Manosa de la Vía del Complemento , Proteínas del Sistema Complemento , Ratones Noqueados , Serina Proteasas Asociadas a la Proteína de Unión a la Manosa/genéticaRESUMEN
Despite their cytotoxic capacity, neutrophils are often co-opted by cancers to promote immunosuppression, tumor growth, and metastasis. Consequently, these cells have received little attention as potential cancer immunotherapeutic agents. Here, we demonstrate in mouse models that neutrophils can be harnessed to induce eradication of tumors and reduce metastatic seeding through the combined actions of tumor necrosis factor, CD40 agonist, and tumor-binding antibody. The same combination activates human neutrophils in vitro, enabling their lysis of human tumor cells. Mechanistically, this therapy induces rapid mobilization and tumor infiltration of neutrophils along with complement activation in tumors. Complement component C5a activates neutrophils to produce leukotriene B4, which stimulates reactive oxygen species production via xanthine oxidase, resulting in oxidative damage and T cell-independent clearance of multiple tumor types. These data establish neutrophils as potent anti-tumor immune mediators and define an inflammatory pathway that can be harnessed to drive neutrophil-mediated eradication of cancer.
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Antineoplásicos , Neoplasias , Ratones , Animales , Humanos , Neutrófilos , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Leucotrieno B4/metabolismo , Leucotrieno B4/farmacología , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
The M-type phospholipase A2 receptor (PLA2R) is the major autoantigen of primary membranous nephropathy (MN). Despite many studies on B-cell epitopes recognized by antibodies, little is known about T-cell epitopes. Herein, we synthesized 123 linear peptides, each consisting of 15-22 amino acids with 8-12 amino acid overlaps, across ten domains of PLA2R. Their binding capacity to risk (DRB1∗1501, DRB1∗0301) and protective (DRB1∗0901, DRB1∗0701) HLA molecules was then assessed by flow cytometry. Proliferation of CD4+ T cells from patients with anti-PLA2R positive MN was analyzed after peptide stimulation. Cytokines produced by activated peripheral blood mononuclear cells were measured by cytometric bead arrays. We identified 17 PLA2R peptides that bound to both DRB1∗1501 and DRB1∗0301 molecules with high capacity. Some of these peptides showed decreased binding to heterozygous DRB1∗1501/0901 and DRB1∗0301/0701. Ten of the 17 peptides (CysR1, CysR10, CysR12, FnII-3, CTLD3-9, CTLD3-10, CTLD3-11, CTLD5-2-1, CTLD7-1 and CTLD7-2) induced significant proliferation of CD4+ T cells from patients with MN than cells from healthy individuals. Upon activation by these peptides, peripheral blood mononuclear cells from patients with MN produced higher levels of pro-inflammatory cytokines, predominantly IL-6, TNF-α, IL-10, IL-9 and IL-17. Thus, we mapped and identified ten peptides in the CysR, FnII, CTLD3, CTLD5, and CTLD7 domains of PLA2R as potential T-cell epitopes of MN. These findings are a first step towards developing peptide-specific immunotherapies.
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Glomerulonefritis Membranosa , Humanos , Epítopos de Linfocito T , Receptores de Fosfolipasa A2 , Leucocitos Mononucleares , Aminoácidos , Fosfolipasas A2 , Citocinas , AutoanticuerposRESUMEN
A complement effect on homeostasis during infection is determined by both cytotoxic (activate complement component 5 (C5a) terminal cytotoxic complex (TCC)), and cytoprotective elements (complement factor H (FH), as well as apolipoprotein E (ApoE)). Here, we investigated the gap in knowledge in their blood milieu during SARS-CoV-2 infection with respect to the viral burden, level of tissue necrosis, and immunological response. 101 patients hospitalized with a PCR-confirmed diagnosis of COVID-19 had blood collected at H1 (48 h), H2 (3-4 Days), H3 (5-7 days), H4 (more than 7 days up to 93 days). Pre-existing conditions, treatment, the incidence of cerebrovascular events (CVA), a history of deep venous thrombosis (DVT) and pulmonary embolism (PE), and mortality was collected using electronic medical records. Plasma C5a, TCC, FH, and ApoE were considered as a complement milieu. Tissue necrosis (HMGB1, RAGE), non-specific inflammatory responses (IL-6, C-reactive protein), overall viral burden (SARS-CoV-2 spike protein), and specific immune responses (IgG, IgA, IgM directed αS- & N-proteins) were assessed simultaneously. C5a remained elevated across all time points, with the peak at 5-7 days. Studied elements of complement coalesced around three clusters: #0 (↑↑↑C5a, ↑↑TCC, ↓↓ApoE), #1 ↑C5a, ↑TCC, ↑↑↑FH); #2 (↑C5a, ↑TCC, ↑FH, ↑↑↑ApoE). The decline in FH and ApoE was a predictor of death, while TCC and C5a correlated with patient length of stay, APACHE, and CRP. Increased levels of C5a (Δ = 122.64; p = 0.0294; data not shown) and diminished levels of FH (Δ = 836,969; p = 0.0285; data not shown) co-existed with CVA incidence. C5a correlated storngly with blood RAGE and HMGB1, but not with viral load and immunological responsiveness. Remdesivir positively affected FH preservation, while convalescent plasma treatment elevated C5a levels. Three clusters of complement activation demonstrated a various milieu of ApoE & FH vs C5a & TCC in COVID-19 patients. Complement activation is linked to increased necrosis markers but not to viral burden or immune system response.
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COVID-19 , Proteína HMGB1 , Apolipoproteínas E/genética , Proteína C-Reactiva , COVID-19/terapia , Activación de Complemento , Complemento C5a , Factor H de Complemento , Humanos , Inmunización Pasiva , Necrosis , Factores Protectores , SARS-CoV-2 , Glicoproteína de la Espiga del Coronavirus , Sueroterapia para COVID-19RESUMEN
A safe and effective vaccine against multidrug-resistant gonorrhea is urgently needed. An experimental peptide vaccine called TMCP2 that mimics an oligosaccharide epitope in gonococcal lipooligosaccharide, when adjuvanted with glucopyranosyl lipid adjuvant-stable emulsion, elicits bactericidal immunoglobulin G and hastens clearance of gonococci in the mouse vaginal colonization model. In this study, we show that efficacy of TMCP2 requires an intact terminal complement pathway, evidenced by loss of activity in C9-/- mice or when C7 function was blocked. In conclusion, TMCP2 vaccine efficacy in the mouse vagina requires membrane attack complex. Serum bactericidal activity may serve as a correlate of protection for TMCP2.
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Gonorrea , Neisseria gonorrhoeae , Animales , Vacunas Bacterianas , Proteínas del Sistema Complemento , Modelos Animales de Enfermedad , Femenino , Gonorrea/prevención & control , Lipopolisacáridos , RatonesRESUMEN
Complement opsonization is among the biggest challenges facing nanomedicine. Nearly instantly after injection into blood, nanoparticles are opsonized by the complement protein C3, leading to clearance by phagocytes, fouling of targeting moieties, and release of anaphylatoxins. While surface polymers such as poly(ethylene glycol) (PEG) partially decrease complement opsonization, most nanoparticles still suffer from extensive complement opsonization, especially when linked to targeting moieties. To ameliorate the deleterious effects of complement, two of mammals' natural regulators of complement activation (RCAs), Factors H and I, are here conjugated to the surface of nanoparticles. In vitro, Factor H or I conjugation to PEG-coated nanoparticles decrease their C3 opsonization, and markedly reduce nanoparticle uptake by phagocytes. In an in vivo mouse model of sepsis-induced lung injury, Factor I conjugation abrogates nanoparticle uptake by intravascular phagocytes in the lungs, allowing the blood concentration of the nanoparticle to remain elevated much longer. For nanoparticles targeted to the lung's endothelium by conjugation to anti-ICAM antibodies, Factor I conjugation shifts the cell-type distribution away from phagocytes and toward endothelial cells. Finally, Factor I conjugation abrogates the severe anaphylactoid responses common to many nanoparticles, preventing systemic capillary leak and preserving blood flow to visceral organs and the brain. Thus, conjugation of RCAs, like Factor I, to nanoparticles is likely to help in nanomedicine's long battle against complement, improving several key parameters critical for clinical success.
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Complemento C3 , Nanomedicina , Nanopartículas , Animales , Activación de Complemento , Complemento C3/metabolismo , Complemento C3/farmacología , Factor H de Complemento/uso terapéutico , Células Endoteliales/metabolismo , Fibrinógeno/uso terapéutico , Mamíferos/metabolismo , Ratones , Nanomedicina/métodos , Nanopartículas/efectos adversos , Nanopartículas/uso terapéutico , OpsonizaciónRESUMEN
Background: Heart surgery results in complement activation with the potential for collateral end-organ damage, especially if the protective elements (complement factor H, Apolipoprotein J) are inadequate. Here, we have investigated if peri-operative stress results in an imbalance between complement activation and its protective mechanisms up to 3 months after heart surgery. Methods: 101 patients scheduled for non-emergent cardiac surgery donated blood before the procedure (tbaseline), and 24 h (t24h ), 7 days (t7d ) and 3 months (t3m ) after. Complement activation was measured as a serum level of soluble activated component 5 (sC5a) and soluble terminal complement complex (sTCC). Simultaneously, protective complement factor H (CfH), and apolipoprotein J (ApoJ) were measured. Inflammatory responses were quantified using C-reactive protein (CRP) and interleukin-6 (IL-6). Details regarding anesthesia, intensive care unit (ICU) stay, pre-existing conditions, the incidence of postoperative complications, and mortality were collected from medical records. Results: C5a declined at t24h to rebound at t7d and t3m . sTCC was significantly depressed at t24h and returned to baseline at later time points. In contrast, CfH and ApoJ were depressed at t3m . Milieu of complement factors aligned along two longitudinal patterns:cluster#1 (C5a/sTTC continuously increasing and CfH/ApoJ preserved at tbaseline) and cluster#2 (transient sC5a/sTTC increase and progressive decline of CfH). Most patients belonged to cluster #1 at t24h (68%), t7d (74%) and t3m (72%). sTCC correlated with APACHE1h (r 2 =-0.25; p < 0.031) and APACHE24h (r 2 = 0.27; p < 0.049). IL-6 correlated with C5a (r 2 =-0.28; p < 0.042) and sTTC (r 2 =-0.28; p < 0.015). Peri-operative administration of acetaminophen and aspirin altered the complement elements. Prolonged hospital stay correlated with elevated C5a [t (78) = 2.03; p = 0.048] and sTTC serum levels [U (73) = 2.07; p = 0.037]. Patients with stroke had a decreased serum level of C5a at t7d and t3m. Conclusion: There is a significant decrease in complement protective factors 3 months after cardiac surgery, while C5a seems to be slightly elevated, suggesting that cardiac surgery affects complement milieu long into recovery.
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.
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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
C3 glomerulopathy (C3G) is a rare renal disease characterized by predominant glomerular C3 staining. Complement alternative pathway dysregulation due to inherited complement defects is associated with C3G. To identify novel C3G-related genes, we screened 86 genes in the complement, coagulation and endothelial systems in 35 C3G patients by targeted genomic enrichment and massively parallel sequencing. Surprisingly, the most frequently mutated gene was VWF. Patients with VWF variants had significantly higher proteinuria levels, higher crescent formation and lower factor H (FH) levels. We further selected two VWF variants to transiently express the von Willebrand factor (vWF) protein, we found that vWF expression from the c.1519A > G variant was significantly reduced. In vitro results further indicated that vWF could regulate complement activation, as it could bind to FH and C3b, act as a cofactor for factor I-mediated cleavage of C3b. Thus, we speculated that vWF might be involved in the pathogenesis of C3G.
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Complemento C3/metabolismo , Glomerulonefritis Membranoproliferativa/genética , Glomerulonefritis/genética , Factor de von Willebrand/genética , Adolescente , Adulto , Estudios de Casos y Controles , China , Estudios de Cohortes , Complemento C3b/metabolismo , Factor H de Complemento/metabolismo , Vía Alternativa del Complemento , Femenino , Variación Genética , Glomerulonefritis/inmunología , Glomerulonefritis/patología , Glomerulonefritis Membranoproliferativa/inmunología , Glomerulonefritis Membranoproliferativa/patología , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Técnicas In Vitro , Glomérulos Renales/inmunología , Glomérulos Renales/patología , Masculino , Persona de Mediana Edad , Modelos Inmunológicos , Simulación de Dinámica Molecular , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Análisis de Secuencia de ADN , Adulto Joven , Factor de von Willebrand/química , Factor de von Willebrand/metabolismoRESUMEN
The statistical model (log-normal model), niche models (Zipf model, broken stick mo-del, niche preemption model), and neutral model were used to fit the species-abundance distribution patterns based on the measurements of environmental factors and inventory data of trees with DBH≥1 cm in a 1.5 hm2 plot in the primary forest (PF) and a 1.5 hm2 plot in the secondary forest (SF). The results showed that species-abundance distribution was affected by habitat heterogeneity in Q. aliena var. acutiserrata forest. Topography had a predominant impact on the species-abundance distribution in PF. Species distribution was affected by both neutral and niche processes, with neutral process having a less prominent effect in large convexity habitats. While the neutral model was rejected by the K-S and Chi-square test in low convexity habitats, the species-abundance distribution satisfied the assumption of niche theory. Niche process and neutral process were equally important in the community in areas with steep slopes, while niche differentiation was the dominant in flat areas. In SF, the main factors affecting species distribution were soil nutrients. The niche process was the mainly ecological process affected species-abundance distribution in habitats with high soil available phosphorus, while the niche and neutral processes existed simultaneously in habitats with low soil phosphorus availability. There was a significant scale effect on the species-abundance distribution pattern of Q. aliena var. acutiserrata forests in Taibai Mountain. The niche and neutral processes could protect the species-abundance distribution at the 20 m×20 m scale in PF, while the niche process could explain the species-abundance distribution at the 40 m×40 m and 70 m×70 m scales. The niche and neutral processes combined acted on the species abundance distribution at the 20 m×20 m, 40 m×40 m and 70 m×70 m scales in SF, with niche process being more important than neutral process. Moreover, besides the scale and habitat heterogeneity, the species-abundance distribution patterns of Q. aliena var. acutiserrata forests differed significantly between primary forest and secondary forest under anthropogenic disturbance.
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Quercus , China , Ecosistema , Bosques , ÁrbolesRESUMEN
Novel therapeutics against the global threat of multidrug-resistant Neisseria gonorrhoeae are urgently needed. Gonococci possess several mechanisms to evade killing by human complement, including binding of factor H (FH), a key inhibitor of the alternative pathway. FH comprises 20 short consensus repeat (SCR) domains organized in a head-to-tail manner as a single chain. N. gonorrhoeae binds two regions in FH; domains 6 and 7 and domains 18 through 20. We designed a novel anti-infective immunotherapeutic molecule that fuses domains 18-20 of FH containing a D-to-G mutation in domain 19 at position 1119 (called FH*) with human IgG1 Fc. FH*/Fc retained binding to gonococci but did not lyse human erythrocytes. Expression of FH*/Fc in tobacco plants was undertaken as an alternative, economical production platform. FH*/Fc was expressed in high yields in tobacco plants (300-600 mg/kg biomass). The activities of plant- and CHO-cell produced FH*/Fc against gonococci were similar in vitro and in the mouse vaginal colonization model of gonorrhea. The addition of flexible linkers [e.g., (GGGGS)2 or (GGGGS)3] between FH* and Fc improved the bactericidal efficacy of FH*/Fc 2.7-fold. The linkers also improved PMN-mediated opsonophagocytosis about 11-fold. FH*/Fc with linker also effectively reduced the duration and burden of colonization of two gonococcal strains tested in mice. FH*/Fc lost efficacy: i) in C6-/- mice (no terminal complement) and ii) when Fc was mutated to abrogate complement activation, suggesting that an intact complement was necessary for FH*/Fc function in vivo. In summary, plant-produced FH*/Fc represent promising prophylactic or adjunctive immunotherapeutics against multidrug-resistant gonococci.
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Resistencia a Múltiples Medicamentos/inmunología , Fragmentos Fc de Inmunoglobulinas/inmunología , Neisseria gonorrhoeae/inmunología , Nicotiana/genética , Plantas Modificadas Genéticamente , Animales , Antibacterianos/farmacología , Factor H de Complemento/genética , Factor H de Complemento/inmunología , Gonorrea , Humanos , Inmunoglobulina G , Inmunoterapia , Ratones , Plantas Modificadas Genéticamente/genética , Proteínas Recombinantes de Fusión/inmunologíaRESUMEN
COVID-19, the disease caused by the SARS-CoV-2 virus, can progress to multi-organ failure characterized by respiratory insufficiency, arrhythmias, thromboembolic complications and shock. The mortality of patients hospitalized with COVID-19 is unacceptably high and new strategies are urgently needed to rapidly identify and treat patients at risk for organ failure. Clinical epidemiologic studies demonstrate that vulnerability to organ failure is greatest after viral clearance from the upper airway, which suggests that dysregulation of the host immune response is a critical mediator of clinical deterioration and death. Autopsy and pre-clinical evidence implicate aberrant complement activation in endothelial injury and organ failure. A potential therapeutic strategy warranting investigation is to inhibit complement, with case reports of successful treatment of COVID-19 with inhibitors of complement. However, this approach requires careful balance between the host protective and potential injurious effects of complement activation, and biomarkers to identify the optimal timing and candidates for therapy are lacking. Here we report the presence of complement activation products on circulating erythrocytes from hospitalized COVID-19 patients using flow cytometry. These findings suggest that novel erythrocyte-based diagnostics provide a method to identify patients with dysregulated complement activation.
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Proteínas del Sistema Complemento/inmunología , Infecciones por Coronavirus/complicaciones , Neumonía Viral/complicaciones , Microangiopatías Trombóticas/complicaciones , Betacoronavirus , COVID-19 , Activación de Complemento , Infecciones por Coronavirus/inmunología , Hemostasis , Humanos , Necrosis , Pandemias , Fenotipo , Neumonía Viral/inmunología , Síndrome de Dificultad Respiratoria , SARS-CoV-2 , TrombosisRESUMEN
Hemorrhagic shock (HS) is a potential life-threatening condition that may lead to injury to multiple organs, including the lung. The estrogen sulfotransferase (EST, or SULT1E1) is a conjugating enzyme that sulfonates and deactivates estrogens. In this report, we showed that the expression of Est was markedly induced in the liver but not in the lung of female mice subject to HS and resuscitation. Genetic ablation or pharmacological inhibition of Est effectively protected female mice from HS-induced acute lung injury (ALI), including interstitial edema, neutrophil mobilization and infiltration, and inflammation. The pulmonoprotective effect of Est ablation or inhibition was sex-specific, because the HS-induced ALI was not affected in male Est-/- mice. Mechanistically, the pulmonoprotective phenotype in female Est-/- mice was accompanied by increased lung and circulating levels of estrogens, attenuated pulmonary inflammation, and inhibition of neutrophil mobilization from the bone marrow and neutrophil infiltration to the lung, whereas the pulmonoprotective effect was abolished upon ovariectomy, suggesting that the protection was estrogen dependent. The pulmonoprotective effect of Est ablation was also tissue specific, as loss of Est had little effect on HS-induced liver injury. Moreover, transgenic reconstitution of human EST in the liver of global Est-/- mice abolished the pulmonoprotective effect, suggesting that it is the EST in the liver that sensitizes mice to HS-induced ALI. Taken together, our results revealed a sex- and tissue-specific role of EST in HS-induced ALI. Pharmacological inhibition of EST may represent an effective approach to manage HS-induced ALI.
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Lesión Pulmonar Aguda/patología , Choque Hemorrágico/complicaciones , Sulfotransferasas/metabolismo , Lesión Pulmonar Aguda/metabolismo , Lesión Pulmonar Aguda/prevención & control , Animales , Estrógenos/metabolismo , Femenino , Hígado/enzimología , Masculino , Ratones , Ratones Noqueados , Ratones Transgénicos , Resucitación , Factores Sexuales , Choque Hemorrágico/terapiaRESUMEN
Severe deficiency of plasma ADAMTS13 activity is the primary cause of thrombotic thrombocytopenic purpura (TTP) whereas overwhelming activation of complement via an alternative pathway results in atypical hemolytic uremic syndrome (aHUS), the prototypes of thrombotic microangiopathy (TMA). However, clinical and pathogenic distinctions between TTP and aHUS are often quite challenging. Clinical reports have suggested that complement activation may play a role in the development of TTP, which is caused by severe deficiency of plasma ADAMTS13 activity. However, the experimental evidence to support this hypothesis is still lacking. Here, we show that mice with either Adamts13 -/- or a heterozygous mutation of complement factor H (cfh) at amino acid residue of 1206 (ie, cfh W/R ) alone remain asymptomatic despite the presence of occasional microvascular thrombi in various organ tissues. However, mice carrying both Adamts13 -/- and cfh W/R exhibit thrombocytopenia, low haptoglobin, increased fragmentation of erythrocytes in peripheral blood smear, increased plasma levels of lactate dehydrogenase activity, blood urea nitrogen, and creatinine, as well as an increased mortality rate, consistent with the development of TMA. Moreover, mice with a homozygous mutation of cfh (ie, cfh R/R ) with or without Adamts13 -/- developed severe TMA. The mortality rate in mice with Adamts13 -/- cfh R/R was significantly higher than that in mice with cfh R/R alone. Histological and immunohistochemical analyses demonstrated the presence of disseminated platelet-rich thrombi in terminal arterioles and capillaries of major organ tissues in these mice that were either euthanized or died. Together, our results support a synergistic effect of severe ADAMTS13 deficiency and complement activation in pathogenesis of TMA in mice.
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Proteína ADAMTS13/genética , Activación de Complemento , Microangiopatías Trombóticas/genética , Proteína ADAMTS13/inmunología , Animales , Factor H de Complemento/genética , Factor H de Complemento/inmunología , Eliminación de Gen , Ratones Endogámicos C57BL , Microangiopatías Trombóticas/inmunología , Microangiopatías Trombóticas/patologíaRESUMEN
Multidrug-resistant Neisseria gonorrhoeae is a global health problem. Monoclonal antibody (mAb) 2C7 recognizes a gonococcal lipooligosaccharide epitope that is expressed by >95% of clinical isolates and hastens gonococcal vaginal clearance in mice. Chimeric mAb 2C7 (human immunoglobulin G1 [IgG1]) with an E430G Fc modification that enhances Fc:Fc interactions and hexamerization following surface-target binding and increases complement activation (HexaBody technology) showed significantly greater C1q engagement and C4 and C3 deposition compared to mAb 2C7 with wild-type Fc. Greater complement activation by 2C7-E430G Fc translated to increased bactericidal activity in vitro and, consequently, enhanced efficacy in mice, compared with "Fc-unmodified" chimeric 2C7. Gonococci bind the complement inhibitors factor H (FH) and C4b-binding protein (C4BP) in a human-specific manner, which dampens antibody (Ab)-mediated complement-dependent killing. The variant 2C7-E430G Fc overcame the barrier posed by these inhibitors in human FH/C4BP transgenic mice, for which a single 1 µg intravenous dose cleared established infection. Chlamydia frequently coexists with and exacerbates gonorrhea; 2C7-E430G Fc also proved effective against gonorrhea in gonorrhea/chlamydia-coinfected mice. Complement activation alone was necessary and sufficient for 2C7 function, evidenced by the fact that (1) "complement-inactive" Fc modifications that engaged Fc gamma receptor (FcγR) rendered 2C7 ineffective, nonetheless; (2) 2C7 was nonfunctional in C1q-/- mice, when C5 function was blocked, or in C9-/- mice; and (3) 2C7 remained effective in neutrophil-depleted mice and in mice treated with PMX205, a C5a receptor (C5aR1) inhibitor. We highlight the importance of complement activation for antigonococcal Ab function in the genital tract. Elucidating the correlates of protection against gonorrhea will inform the development of Ab-based gonococcal vaccines and immunotherapeutics.
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Activación de Complemento/inmunología , Gonorrea/inmunología , Neisseria gonorrhoeae/inmunología , Animales , Anticuerpos Antibacterianos/inmunología , Anticuerpos Monoclonales/metabolismo , Antígenos Bacterianos , Proteína de Unión al Complemento C4b/inmunología , Factor H de Complemento/inmunología , Proteínas del Sistema Complemento/inmunología , Proteínas del Sistema Complemento/metabolismo , Epítopos/inmunología , Femenino , Voluntarios Sanos , Humanos , Inmunoglobulina G/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Transgénicos , Neisseria gonorrhoeae/patogenicidadRESUMEN
Atypical hemolytic uremic syndrome (aHUS) is a form of thrombotic microangiopathy (TMA) caused by dysregulated complement activation. Clinically, aHUS is effectively treated by an anti-C5 monoclonal antibody (mAb) but whether the disease is mediated by the C5a receptor (C5aR) or C5b-9 pathway, or both, is unknown. Here we address this in a factor H mutant mouse (FHR/R) which developed complement-mediated TMA as well as macrovascular thrombosis caused by an aHUS-related factor H point mutation (mouse W1206R, corresponding to human W1183R). C5 deficiency and anti-C5 mAb treatment blocked all disease manifestations in FHR/R mice. C5aR1 gene deficiency prevented macrovascular thrombosis in various organs but did not improve survival or reduce renal TMA. Conversely, C6 or C9 deficiency significantly improved survival and markedly diminished renal TMA but did not prevent macrovascular thrombosis. Interestingly, as they aged both FHR/R C6-/- and FHR/R C9-/- mice developed glomerular disease reminiscent of C3 glomerulonephritis. Thus, C5aR and C5b-9 pathways drove different aspects of disease in FHR/R mice with the C5aR pathway being responsible for macrovascular thrombosis and chronic inflammatory injury while the C5b-9 pathway caused renal TMA. Our data provide new understanding of the pathogenesis of complement-mediated TMA and macrovascular thrombosis in FHR/R mice and suggest that C5 blockade is more effective for the treatment of aHUS than selectively targeting the C5aR or C5b-9 pathway alone.
Asunto(s)
Síndrome Hemolítico Urémico Atípico/inmunología , Factor H de Complemento/genética , Complejo de Ataque a Membrana del Sistema Complemento/inmunología , Glomérulos Renales/patología , Receptor de Anafilatoxina C5a/inmunología , Animales , Anticuerpos Monoclonales/farmacología , Anticuerpos Monoclonales/uso terapéutico , Síndrome Hemolítico Urémico Atípico/tratamiento farmacológico , Síndrome Hemolítico Urémico Atípico/genética , Síndrome Hemolítico Urémico Atípico/patología , Activación de Complemento/efectos de los fármacos , Activación de Complemento/genética , Activación de Complemento/inmunología , Complemento C6/genética , Complemento C6/inmunología , Complemento C6/metabolismo , Factor H de Complemento/inmunología , Complejo de Ataque a Membrana del Sistema Complemento/genética , Complejo de Ataque a Membrana del Sistema Complemento/metabolismo , Modelos Animales de Enfermedad , Femenino , Humanos , Glomérulos Renales/irrigación sanguínea , Glomérulos Renales/ultraestructura , Masculino , Ratones , Ratones Transgénicos , Microscopía Electrónica , Mutación Puntual , Receptor de Anafilatoxina C5a/antagonistas & inhibidores , Receptor de Anafilatoxina C5a/genética , Receptor de Anafilatoxina C5a/metabolismoRESUMEN
Single-nucleotide polymorphisms and rare mutations in factor H (FH; official name, CFH) are associated with age-related macular degeneration and atypical hemolytic uremic syndrome, a form of thrombotic microangiopathy. Mice with the FH W1206R mutation (FHR/R) share features with human atypical hemolytic uremic syndrome. Herein, we report that FHR/R mice exhibited retinal vascular occlusion and ischemia. Retinal fluorescein angiography demonstrated delayed perfusion and vascular leakage in FHR/R mice. Optical coherence tomography imaging of FHR/R mice showed retinal degeneration, edema, and detachment. Histologic analysis of FHR/R mice revealed retinal thinning, vessel occlusion, as well as degeneration of photoreceptors and retinal pigment epithelium. Immunofluorescence showed albumin leakage from blood vessels into the neural retina, and electron microscopy demonstrated vascular endothelial cell irregularity with narrowing of retinal and choroidal vessels. Knockout of C6, a component of the membrane attack complex, prevented the aforementioned retinal phenotype in FHR/R mice, consistent with membrane attack complex-mediated pathogenesis. Pharmacologic blockade of C5 also rescued retinas of FHR/R mice. This FHR/R mouse strain represents a model for retinal vascular occlusive disorders and ischemic retinopathy. The results suggest complement dysregulation can contribute to retinal vascular occlusion and that an anti-C5 antibody might be helpful for C5-mediated thrombotic retinal diseases.