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1.
BMJ Case Rep ; 14(2)2021 Feb 09.
Artigo em Inglês | MEDLINE | ID: mdl-33563691

RESUMO

While sporadic inclusion body myositis (sIBM) is the most commonly acquired inflammatory myopathy above 50 years of age, its refractory response to conventional immunosuppressive treatments raises questions about its perplexing pathogenesis. Muscle biopsy typically reveals major histocompatibility complex I antigens and CD8+ T cell endomysial infiltrates invading non-necrotic muscle fibres early in the disease course with rimmed vacuoles, protein aggregates and amyloid inclusions later in the disease. Transactive response DNA-binding protein-43 (TDP-43), a protein implicated in transcriptional repression in neurodegenerative diseases, is also found in sIBM. C5b-9 membrane attack complex, an effector protein involved in the complement cascade of the immune response, is commonly found in dermatomyositis, but has rarely been reported in IBM. We describe a novel case of IBM with simultaneous C5b-9 and TDP-43 staining on quadriceps biopsy, raising the question of a possibility of concurrent immune-mediated inflammatory and myodegenerative pathogenesis.


Assuntos
Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Proteínas de Ligação a DNA/metabolismo , Miosite de Corpos de Inclusão/diagnóstico , Miosite de Corpos de Inclusão/metabolismo , Idoso , Biomarcadores/sangue , Biópsia , Diagnóstico Diferencial , Humanos , Masculino , Coloração e Rotulagem
2.
Muscle Nerve ; 63(4): 506-515, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33346931

RESUMO

INTRODUCTION: Identification and treatment of immune-mediated polyneuropathies may lead to improved strength and function. We studied the clinical and laboratory features, and treatment response, in patients with motor-sensory axonal polyneuropathies who were found to have C5b-9 complement staining on endoneurial microvessels. METHODS: Retrospective review of 16 consecutive adults with motor-sensory axonal polyneuropathies who were then found to have C5b-9 staining of endoneurial microvessels on nerve biopsy, and subsequently treated with intravenous corticosteroids (1 g methylprednisolone for 5 consecutive days, and then weekly). Strength measurements were done using quantitative handheld dynamometry. Nerve biopsy analysis included frozen and fixed tissue. RESULTS: Patients (mean onset age, 59 ± 4 years; range, 34-83 years; 12 of 16 were males; 9 of 16 had diabetes) had progressive (median duration, 2 years), asymmetric, distal weakness, in the lower extremities (16 of 16) and/or upper extremities (7 of 16), and panmodal sensory loss. Electrodiagnostic studies showed axon loss. Nerve pathology showed abnormal C5b-9 staining on endoneurial microvessels. Axon loss was present in all nerves, often varied among fascicles. Inflammation was uncommon. Distal strength usually improved (mean improvement of 34 ± 6% of normal strength; P = .0003) with corticosteroid treatment. DISCUSSION: Motor-sensory axonal polyneuropathies having noninflammatory, humoral immune pathology with C5b-9 staining of endoneurial microvessels (HIEM) frequently manifest progressive asymmetric, distal, lower extremity with or without upper extremity weakness that improves rapidly during corticosteroid treatment. HIEM may represent a new class of noninflammatory-vasculopathic, treatable axonal motor-sensory neuropathies.


Assuntos
Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Neuropatias Diabéticas/fisiopatologia , Microvasos/patologia , Nervos Periféricos/metabolismo , Nervos Periféricos/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Axônios/metabolismo , Axônios/patologia , Neuropatias Diabéticas/patologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos
3.
Blood ; 136(18): 2080-2089, 2020 10 29.
Artigo em Inglês | MEDLINE | ID: mdl-32877502

RESUMO

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a highly contagious respiratory virus that can lead to venous/arterial thrombosis, stroke, renal failure, myocardial infarction, thrombocytopenia, and other end-organ damage. Animal models demonstrating end-organ protection in C3-deficient mice and evidence of complement activation in humans have led to the hypothesis that SARS-CoV-2 triggers complement-mediated endothelial damage, but the mechanism is unclear. Here, we demonstrate that the SARS-CoV-2 spike protein (subunit 1 and 2), but not the N protein, directly activates the alternative pathway of complement (APC). Complement-dependent killing using the modified Ham test is blocked by either C5 or factor D inhibition. C3 fragments and C5b-9 are deposited on TF1PIGAnull target cells, and complement factor Bb is increased in the supernatant from spike protein-treated cells. C5 inhibition prevents the accumulation of C5b-9 on cells, but not C3c; however, factor D inhibition prevents both C3c and C5b-9 accumulation. Addition of factor H mitigates the complement attack. In conclusion, SARS-CoV-2 spike proteins convert nonactivator surfaces to activator surfaces by preventing the inactivation of the cell-surface APC convertase. APC activation may explain many of the clinical manifestations (microangiopathy, thrombocytopenia, renal injury, and thrombophilia) of COVID-19 that are also observed in other complement-driven diseases such as atypical hemolytic uremic syndrome and catastrophic antiphospholipid antibody syndrome. C5 inhibition prevents accumulation of C5b-9 in vitro but does not prevent upstream complement activation in response to SARS-CoV-2 spike proteins.


Assuntos
Betacoronavirus , Fator D do Complemento/antagonistas & inibidores , Inativadores do Complemento/farmacologia , Via Alternativa do Complemento/efeitos dos fármacos , Glicoproteína da Espícula de Coronavírus/farmacologia , Linhagem Celular , Ativação do Complemento/efeitos dos fármacos , Complemento C3/metabolismo , Complemento C5/antagonistas & inibidores , Fator H do Complemento/metabolismo , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Humanos , Glicoproteína da Espícula de Coronavírus/fisiologia
4.
J Clin Invest ; 130(11): 6151-6157, 2020 11 02.
Artigo em Inglês | MEDLINE | ID: mdl-32759504

RESUMO

Emerging data indicate that complement and neutrophils contribute to the maladaptive immune response that fuels hyperinflammation and thrombotic microangiopathy, thereby increasing coronavirus 2019 (COVID-19) mortality. Here, we investigated how complement interacts with the platelet/neutrophil extracellular traps (NETs)/thrombin axis, using COVID-19 specimens, cell-based inhibition studies, and NET/human aortic endothelial cell (HAEC) cocultures. Increased plasma levels of NETs, tissue factor (TF) activity, and sC5b-9 were detected in patients. Neutrophils of patients yielded high TF expression and released NETs carrying active TF. Treatment of control neutrophils with COVID-19 platelet-rich plasma generated TF-bearing NETs that induced thrombotic activity of HAECs. Thrombin or NETosis inhibition or C5aR1 blockade attenuated platelet-mediated NET-driven thrombogenicity. COVID-19 serum induced complement activation in vitro, consistent with high complement activity in clinical samples. Complement C3 inhibition with compstatin Cp40 disrupted TF expression in neutrophils. In conclusion, we provide a mechanistic basis for a pivotal role of complement and NETs in COVID-19 immunothrombosis. This study supports strategies against severe acute respiratory syndrome coronavirus 2 that exploit complement or NETosis inhibition.


Assuntos
Betacoronavirus , Complexo de Ataque à Membrana do Sistema Complemento , Infecções por Coronavirus , Armadilhas Extracelulares , Neutrófilos , Pandemias , Pneumonia Viral , Tromboplastina , Trombose , Idoso , Betacoronavirus/imunologia , Betacoronavirus/metabolismo , Ativação do Complemento/efeitos dos fármacos , Complexo de Ataque à Membrana do Sistema Complemento/imunologia , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Infecções por Coronavirus/sangue , Infecções por Coronavirus/imunologia , Armadilhas Extracelulares/imunologia , Armadilhas Extracelulares/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Neutrófilos/imunologia , Neutrófilos/metabolismo , Peptídeos Cíclicos/farmacologia , Pneumonia Viral/sangue , Pneumonia Viral/imunologia , Receptor da Anafilatoxina C5a/antagonistas & inibidores , Receptor da Anafilatoxina C5a/sangue , Receptor da Anafilatoxina C5a/imunologia , /imunologia , Trombina/imunologia , Trombina/metabolismo , Tromboplastina/imunologia , Tromboplastina/metabolismo , Trombose/sangue , Trombose/imunologia , Trombose/virologia
5.
J Clin Invest ; 130(11): 5967-5975, 2020 11 02.
Artigo em Inglês | MEDLINE | ID: mdl-32730233

RESUMO

BACKGROUNDInitial reports from the severe acute respiratory coronavirus 2 (SARS-CoV-2) pandemic described children as being less susceptible to coronavirus disease 2019 (COVID-19) than adults. Subsequently, a severe and novel pediatric disorder termed multisystem inflammatory syndrome in children (MIS-C) emerged. We report on unique hematologic and immunologic parameters that distinguish between COVID-19 and MIS-C and provide insight into pathophysiology.METHODSWe prospectively enrolled hospitalized patients with evidence of SARS-CoV-2 infection and classified them as having MIS-C or COVID-19. Patients with COVID-19 were classified as having either minimal or severe disease. Cytokine profiles, viral cycle thresholds (Cts), blood smears, and soluble C5b-9 values were analyzed with clinical data.RESULTSTwenty patients were enrolled (9 severe COVID-19, 5 minimal COVID-19, and 6 MIS-C). Five cytokines (IFN-γ, IL-10, IL-6, IL-8, and TNF-α) contributed to the analysis. TNF-α and IL-10 discriminated between patients with MIS-C and severe COVID-19. The presence of burr cells on blood smears, as well as Cts, differentiated between patients with severe COVID-19 and those with MIS-C.CONCLUSIONPediatric patients with SARS-CoV-2 are at risk for critical illness with severe COVID-19 and MIS-C. Cytokine profiling and examination of peripheral blood smears may distinguish between patients with MIS-C and those with severe COVID-19.FUNDINGFinancial support for this project was provided by CHOP Frontiers Program Immune Dysregulation Team; National Institute of Allergy and Infectious Diseases; National Cancer Institute; the Leukemia and Lymphoma Society; Cookies for Kids Cancer; Alex's Lemonade Stand Foundation for Childhood Cancer; Children's Oncology Group; Stand UP 2 Cancer; Team Connor; the Kate Amato Foundations; Burroughs Wellcome Fund CAMS; the Clinical Immunology Society; the American Academy of Allergy, Asthma, and Immunology; and the Institute for Translational Medicine and Therapeutics.


Assuntos
Betacoronavirus/metabolismo , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Infecções por Coronavirus , Citocinas/sangue , Pandemias , Pneumonia Viral , Síndrome de Resposta Inflamatória Sistêmica , Adolescente , Criança , Pré-Escolar , Infecções por Coronavirus/sangue , Infecções por Coronavirus/epidemiologia , Feminino , Humanos , Masculino , Pneumonia Viral/sangue , Pneumonia Viral/epidemiologia , Estudos Prospectivos , Índice de Gravidade de Doença , Síndrome de Resposta Inflamatória Sistêmica/sangue , Síndrome de Resposta Inflamatória Sistêmica/epidemiologia
6.
PLoS Pathog ; 16(6): e1008606, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32569291

RESUMO

An important effector function of the human complement system is to directly kill Gram-negative bacteria via Membrane Attack Complex (MAC) pores. MAC pores are assembled when surface-bound convertase enzymes convert C5 into C5b, which together with C6, C7, C8 and multiple copies of C9 forms a transmembrane pore that damages the bacterial cell envelope. Recently, we found that bacterial killing by MAC pores requires local conversion of C5 by surface-bound convertases. In this study we aimed to understand why local assembly of MAC pores is essential for bacterial killing. Here, we show that rapid interaction of C7 with C5b6 is required to form bactericidal MAC pores on Escherichia coli. Binding experiments with fluorescently labelled C6 show that C7 prevents release of C5b6 from the bacterial surface. Moreover, trypsin shaving experiments and atomic force microscopy revealed that this rapid interaction between C7 and C5b6 is crucial to efficiently anchor C5b-7 to the bacterial cell envelope and form complete MAC pores. Using complement-resistant clinical E. coli strains, we show that bacterial pathogens can prevent complement-dependent killing by interfering with the anchoring of C5b-7. While C5 convertase assembly was unaffected, these resistant strains blocked efficient anchoring of C5b-7 and thus prevented stable insertion of MAC pores into the bacterial cell envelope. Altogether, these findings provide basic molecular insights into how bactericidal MAC pores are assembled and how bacteria evade MAC-dependent killing.


Assuntos
Atividade Bactericida do Sangue , Membrana Celular/metabolismo , Parede Celular/metabolismo , Complemento C5/metabolismo , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Escherichia coli/metabolismo , Proteínas do Sistema Complemento/metabolismo , Células HEK293 , Humanos
7.
Int J Mol Sci ; 21(5)2020 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-32121610

RESUMO

Several studies have suggested that there is a link between membrane attack complex (MAC) deposition in the retina and the progression of diabetic retinopathy (DR). Our recent investigation demonstrated that circulating IgG-laden extracellular vesicles contribute to an increase in retinal vascular permeability in DR through activation of the complement system. However, the mechanism through which extracellular vesicle-induced complement activation contributes to retinal vascular cytolytic damage in DR is not well understood. In this study, we demonstrate that IgG-laden extracellular vesicles in rat plasma activate the classical complement pathway, and in vitro Streptozotocin (STZ)-induced rat diabetic plasma results in MAC deposition and cytolytic damage in human retinal endothelial cells (HRECs). Moreover, removal of the plasma extracellular vesicles reduced the MAC deposition and abrogated cytolytic damage seen in HRECs. Together, the results of this study demonstrate that complement activation by IgG-laden extracellular vesicles in plasma could lead to MAC deposition and contribute to endothelium damage and progression of DR.


Assuntos
Ativação do Complemento/imunologia , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Células Endoteliais/patologia , Vesículas Extracelulares/metabolismo , Retina/patologia , Animais , Morte Celular , Sobrevivência Celular , Complemento C1/metabolismo , Diabetes Mellitus Experimental/sangue , Diabetes Mellitus Experimental/imunologia , Diabetes Mellitus Experimental/patologia , Vesículas Extracelulares/ultraestrutura , Humanos , Imunoglobulinas/metabolismo , Masculino , Ratos Sprague-Dawley
8.
Am J Pathol ; 190(6): 1138-1150, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32194049

RESUMO

The complement membrane attack complex (MAC) is classically known as a cytolytic effector of innate and adaptive immunity that forms pores in the plasma membrane of pathogens or targeted cells, leading to osmolysis. Nucleated cells resist MAC-mediated cytolysis by expression of inhibitors that block MAC assembly or by rapid removal of MAC through endocytosis or shedding. In the absence of lysis, MAC may induce intracellular signaling and cell activation, responses implicated in a variety of autoimmune, inflammatory, and transplant disease settings. New discoveries into the structure and biophysical properties of MAC revealed heterogeneous MAC precursors and conformations that provide insights into MAC function. In addition, new mechanisms of MAC-mediated signaling and its contribution to disease pathogenesis have recently come to light. MAC-activated cells have been found to express proinflammatory proteins-often through NF-κB-dependent transcription, assemble inflammasomes, enabling processing, and facilitate secretion of IL-1ß and IL-18, as well as other signaling pathways. These recent insights into the mechanisms of action of MAC provide an updated framework to therapeutic approaches that can target MAC assembly, signaling, and proinflammatory effects in various complement-mediated diseases.


Assuntos
Imunidade Adaptativa/fisiologia , Ativação do Complemento/fisiologia , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Imunidade Inata/fisiologia , Animais , Humanos , Interleucinas/metabolismo , NF-kappa B/metabolismo , Transdução de Sinais/fisiologia
9.
Dev Comp Immunol ; 102: 103486, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31473265

RESUMO

The complement system is a crucial component of the innate immune system that links innate and adaptive immunity. CL-11, a protein similar to Mannose-binding lectin (MBL), plays significant role in the innate immune system in mammals and fish, serving as an initiator of the lectin pathway of complement activation. In this study, a CL-11 homolog (TfCol-11) was identified in roughskin sculpin (Trachidermus fasciatus), and its expression and role in immune responses were characterized. The open reading frame of TfCol-11 is 795 bp long, encoding a 264 amino acid polypeptide. The deduced amino acid sequence of this protein is highly homologous to sequences in other teleosts, and is similar to vertebrate CL-11, containing a canonical collagen-like region, a carbohydrate recognition domain, and a neck region. Recombinant TfCol-11 purified from Escherichia coli(E.coli) was able to bind to different microbes in a Ca2+-independent manner. Meanwhile, a 993 bp-long of partial MASP cDNA with a 96 bp 5' untranslated region (UTR) was also cloned from roughskin sculpin, containing 299 amino acids and consisting of three domains (CUB-EGF-CUB). qRT-PCR indicated that TfCol-11 and MASP mRNAs were predominately co-expressed in the liver. The temporal expression of TfCol-11 and MASP were both drastically up-regulated in the liver, skin, and blood by LPS challenge. Recombinant TfCol-11 purified from E.coli BL21(DE3) was able to agglutinate some bacteria in a Ca2+-dependent manner. In addition, an in vitro pull-down experiment demonstrated that TfCol-11 was able to bind to MASP, and in vivo experiments showed that TfCol-11 was associated with increased membrane attack complex (MAC) levels. It is therefore possible that TfCol-11 may plays a role in activating the complement system and in the defense against invading microorganisms in roughskin sculpin.


Assuntos
Colectinas/metabolismo , Ativação do Complemento , Proteínas de Peixes/metabolismo , Perciformes/imunologia , Testes de Aglutinação , Sequência de Aminoácidos , Animais , Sequência de Bases , Colectinas/química , Colectinas/genética , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Proteínas de Peixes/química , Proteínas de Peixes/genética , Regulação da Expressão Gênica , Imunidade Inata , Serina Proteases Associadas a Proteína de Ligação a Manose/química , Serina Proteases Associadas a Proteína de Ligação a Manose/genética , Serina Proteases Associadas a Proteína de Ligação a Manose/metabolismo , Fases de Leitura Aberta , Filogenia , Domínios Proteicos , Alinhamento de Sequência , Distribuição Tecidual
10.
Microbes Infect ; 22(1): 19-30, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31473336

RESUMO

The Gram-negative bacterium Klebsiella pneumoniae is an opportunistic pathogen, which can cause life-threatening infections such as sepsis. Worldwide, emerging multidrug resistant K. pneumoniae infections are challenging to treat, hence leading to increased mortality. Therefore, understanding the interactions between K. pneumoniae and the immune system is important to develop new treatment options. We characterized ten clinical K. pneumoniae isolates obtained from blood of bacteremia patients. The interaction of the isolates with human serum was investigated to elucidate how K. pneumoniae escapes the host immune system, and how complement activation by K. pneumoniae changed the capsule structure. All K. pneumoniae isolates activated the alternative complement pathway despite serum resistance of seven isolates. One serum sensitive isolate activated two or all three pathways, and this isolate was lysed and had numerous membrane attack complexes in the outer membrane. However, we also found deposition of complement components in the capsule of serum resistant isolates resulting in morphological capsule changes and capsule shedding. These bacteria did not lyse, and no membrane attack complex was observed despite deposition of C5b-9 within the capsule, indicating that the capsule of serum resistant K. pneumoniae isolates is a defense mechanism against complement-mediated lysis.


Assuntos
Cápsulas Bacterianas/imunologia , Proteínas do Sistema Complemento/imunologia , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/imunologia , Bacteriemia/imunologia , Bacteriemia/microbiologia , Cápsulas Bacterianas/metabolismo , Atividade Bactericida do Sangue , Ativação do Complemento , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Proteínas do Sistema Complemento/deficiência , Interações Hospedeiro-Patógeno , Humanos , Infecções por Klebsiella/imunologia , Klebsiella pneumoniae/isolamento & purificação
11.
Eur J Ophthalmol ; 30(5): 1061-1068, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31203676

RESUMO

PURPOSE: To examine the role of systemic activation of the complement system (assessed by levels of circulating C3a, Ba, and sC5b-9) in patients (n = 122) with advanced age-related macular degeneration, geographic atrophy, and neovascular age-related macular degeneration, compared with cataract controls (n = 27). METHODS: Plasma complement factors were measured using enzyme-linked immunosorbent assays. Statistical analysis included univariate and multivariate logistic regression (p < 0.05). RESULTS: Adjusted for age, the odds ratios of C3a and sC5b-9 for any advanced age-related macular degeneration were 1.78 (95% confidence interval = 1.16-2.73, p < 0.01) and 1.20 (95% confidence interval = 1.04-1.39, p = 0.01), respectively. We found a significantly elevated adjusted odds ratio of C3a (adjusted odds ratio = 1.71, 95% confidence interval = 1.12-2.60, p = 0.01) and sC5b-9 (adjusted odds ratio = 1.22, 95% confidence interval = 1.04-1.43, p = 0.01) for neovascular age-related macular degeneration. Adjusted for age, neither C3a, sC5b-9, nor Ba were associated with geographic atrophy. CONCLUSION: We suggest a role for elevated plasma levels of C3a and sC5b-9 in patients with neovascular age-related macular degeneration. The study's results reinforce the need for more investigation to assess the impact of therapeutic interventions targeted at the complement signaling pathways in age-related macular degeneration.


Assuntos
Neovascularização de Coroide/sangue , Ativação do Complemento/fisiologia , Complemento C3a/metabolismo , Fator B do Complemento/metabolismo , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Atrofia Geográfica/sangue , Degeneração Macular Exsudativa/sangue , Idoso , Idoso de 80 Anos ou mais , Neovascularização de Coroide/diagnóstico , Ensaio de Imunoadsorção Enzimática , Feminino , Angiofluoresceinografia , Atrofia Geográfica/diagnóstico , Humanos , Masculino , Razão de Chances , Tomografia de Coerência Óptica , Degeneração Macular Exsudativa/diagnóstico
12.
Eur J Pediatr ; 179(1): 133-140, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31691001

RESUMO

Individuals with celiac disease (CD) are at increased risk of invasive pneumococcal disease (IPD). The aim of this study was to explore whether the complement response to Streptococcus pneumoniae differed according to CD status, and could serve as an explanation for the excess risk of IPD in CD. Twenty-two children with CD and 18 controls, born 1999-2008, were included at Kalmar County Hospital, Sweden. The degree of complement activation was evaluated by comparing levels of activation products C3a and sC5b-9 in plasma incubated for 30 min with Streptococcus pneumoniae and in non-incubated plasma. Complement analyses were performed with enzyme-linked immunosorbent assay (ELISA). Pneumococcal stimulation caused a statistically significant increase in C3a as well as sC5b-9 in both children with CD and controls but there was no difference in response between the groups. After incubation, C3a increased on average 4.6 times and sC5b-9 22 times in both the CD and the control group (p = 0.497 and p = 0.724 respectively).Conclusion: Complement response to Streptococcus pneumoniae seems to be similar in children with and without CD and is thus unlikely to contribute to the increased susceptibility to invasive pneumococcal disease in CD.What is Known:• An excess risk of pneumococcal infections has been demonstrated in individuals with celiac disease.• Infectious complications can depend on hyposplenism but alternative mechanisms are sparsely examined.What is New:• Complement activation in response to Streptococcus pneumoniae was examined in children with and without celiac disease but no differences could be demonstrated.


Assuntos
Doença Celíaca/complicações , Ativação do Complemento , Infecções Pneumocócicas/etiologia , Adolescente , Biomarcadores/sangue , Estudos de Casos e Controles , Doença Celíaca/imunologia , Doença Celíaca/microbiologia , Criança , Complemento C3a/metabolismo , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Suscetibilidade a Doenças , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Masculino , Infecções Pneumocócicas/imunologia , Fatores de Risco
13.
Clin J Am Soc Nephrol ; 14(12): 1719-1732, 2019 12 06.
Artigo em Inglês | MEDLINE | ID: mdl-31694864

RESUMO

BACKGROUND AND OBJECTIVES: Atypical hemolytic uremic syndrome is a form of thrombotic microangiopathy caused by dysregulation of the alternative complement pathway. There is evidence showing complement activation in other thrombotic microangiopathies. The aim of this study was to evaluate complement activation in different thrombotic microangiopathies and to monitor treatment response. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: Complement activation was assessed by exposing endothelial cells to sera or activated-patient plasma-citrated plasma mixed with a control sera pool (1:1)-to analyze C5b-9 deposits by immunofluorescence. Patients with atypical hemolytic uremic syndrome (n=34) at different stages of the disease, HELLP syndrome (a pregnancy complication characterized by hemolysis, elevated liver enzymes, and low platelet count) or severe preeclampsia (n=10), and malignant hypertension (n=5) were included. RESULTS: Acute phase atypical hemolytic uremic syndrome-activated plasma induced an increased C5b-9 deposition on endothelial cells. Standard and lower doses of eculizumab inhibited C5b-9 deposition in all patients with atypical hemolytic uremic syndrome, except in two who showed partial remission and clinical relapse. Significant fibrin formation was observed together with C5b-9 deposition. Results obtained using activated-plasma samples were more marked and reproducible than those obtained with sera. C5b-9 deposition was also increased with samples from patients with HELLP (all cases) and preeclampsia (90%) at disease onset. This increase was sustained in those with HELLP after 40 days, and levels normalized in patients with both HELLP and preeclampsia after 6-9 months. Complement activation in those with malignant hypertension was at control levels. CONCLUSIONS: The proposed methodology identifies complement overactivation in patients with atypical hemolytic uremic syndrome at acute phase and in other diseases such as HELLP syndrome and preeclampsia. Moreover, it is sensitive enough to individually assess the efficiency of the C5 inhibition treatment.


Assuntos
Ativação do Complemento , Microangiopatias Trombóticas/imunologia , Adulto , Anticorpos Monoclonais Humanizados/uso terapêutico , Síndrome Hemolítico-Urêmica Atípica/tratamento farmacológico , Síndrome Hemolítico-Urêmica Atípica/imunologia , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Feminino , Síndrome HELLP/imunologia , Humanos , Masculino , Pré-Eclâmpsia/tratamento farmacológico , Pré-Eclâmpsia/imunologia , Gravidez , Microangiopatias Trombóticas/tratamento farmacológico
14.
Toxicol Lett ; 317: 110-119, 2019 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-31618666

RESUMO

Trichloroethylene (TCE), a commonly used industrial solvent and degreasing agent, is known to cause trichloroethylene hypersensitivity syndrome (THS) with multi-system damage, including skin, liver and kidney. Clinical evidence have shown that the kidney injury occurs in THS and our previous studies suggested that the terminal complement complex C5b-9 deposited in impaired renal tubules induced by TCE with unclear mechanisms. In the present study, we questioned whether activation of the complement system with renal deposition of C5b-9 contributes to TCE-induced kidney injury in THS. We established a BALB/c mouse model of TCE sensitization with or without pretreatment of exogenous CD59, a C5b-9 inhibitory protein. H&E staining, PAS staining, and biochemical detection of urinary proteins were performed to assess renal function. Deposition of C5b-9 and expression of CD59 were evaluated by immunohistochemistry. Sub-lytic effects of C5b-9 in tubular epithelial cells were assessed by lactate dehydrogenase (LDH) cytotoxicity assay. Expression of endocytosis receptors megalin and cubilin on proximal tubules were assessed by immunofluorescence and qRT-PCR. We found that TCE sensitization induced structural and functional changes of renal tubules in mice, associated with the deposition of sub-lytic C5b-9 on proximal tubular epithelial cells. TCE sensitization decreased proximal tubule uptake of filtered proteins and renal expression of megalin and cubilin, phenotypes that were attenuated by pretreatment with exogenous CD59. Overall, our findings reveal a novel mechanism underlying sub-lytic C5b-9 acting on megalin and cubilin, contributes to the renal tubules damage by TCE exposure.


Assuntos
Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Endocitose , Hipersensibilidade/metabolismo , Nefropatias/metabolismo , Túbulos Renais Proximais/metabolismo , Proteína-2 Relacionada a Receptor de Lipoproteína de Baixa Densidade/metabolismo , Receptores de Superfície Celular/metabolismo , Tricloroetileno , Animais , Células Cultivadas , Ativação do Complemento , Complexo de Ataque à Membrana do Sistema Complemento/imunologia , Modelos Animais de Doenças , Feminino , Hipersensibilidade/imunologia , Hipersensibilidade/patologia , Nefropatias/induzido quimicamente , Nefropatias/imunologia , Nefropatias/patologia , Túbulos Renais Proximais/imunologia , Túbulos Renais Proximais/patologia , Camundongos Endogâmicos BALB C , Transporte Proteico
15.
Am J Reprod Immunol ; 82(6): e13185, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31479579

RESUMO

PROBLEM: As antiphospholipid antibody-positive women with adverse pregnancy outcomes have higher plasma complement activation product levels, and the placentas of women with antiphospholipid syndrome (APS) exhibit C4d complement component deposition, complement activation involvement has been hypothesized in APS pregnancy complications. METHOD OF STUDY: Plasma levels of C5a and C5b-9 complement components of 43 APS non-pregnant patients and 17 pregnant APS women were measured using enzyme-linked immunosorbent assay. The results were compared with those of 16 healthy non-pregnant women and eight healthy pregnant women, respectively. Placenta samples of five APS patients at high risk of pregnancy complications and of five healthy controls were subjected to immunoblotting analysis with specific antibodies to C5b-9 and CD46, CD55, CD59 complement regulators. RESULTS: The mean plasma C5a and C5b-9 levels were significantly higher in the non-pregnant APS patients with previous thrombosis ± pregnancy morbidity (P = .0001 and P = .0034, respectively) and in the pregnant APS women with adverse outcomes (P = .0093 for both). Similarly, C5b-9 amounts were significantly higher in the adverse pregnancy outcome placenta (P = .0115) than in those associated to a favorable outcome. The mean CD46, CD55 and CD59 amounts were, instead, lower, although not always significantly, in the placentas of all the high-risk APS women with respect to the control placentas. CONCLUSION: Data analysis demonstrated that there was significant complement activation in the more severe subset of APS patients and in only the adverse pregnancy outcome APS women. Further studies will clarify whether the lower CD46, CD55, and CD59 expressions in the APS placentas are limited to only high-risk APS patients.


Assuntos
Síndrome Antifosfolipídica/sangue , Ativação do Complemento , Complicações na Gravidez/sangue , Adulto , Antígenos CD55/sangue , Antígenos CD59/sangue , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Feminino , Humanos , Proteína Cofatora de Membrana/sangue , Gravidez
16.
Sci Rep ; 9(1): 13873, 2019 09 25.
Artigo em Inglês | MEDLINE | ID: mdl-31554875

RESUMO

Age Related Macular Degeneration (AMD) is the first cause of social blindness in people aged over 65 leading to atrophy of retinal pigment epithelial cells (RPE), photoreceptors and choroids, eventually associated with choroidal neovascularization. Accumulation of undigested cellular debris within RPE cells or under the RPE (Drusen), oxidative stress and inflammatory mediators contribute to the RPE cell death. The major risk to develop AMD is the Y402H polymorphism of complement factor H (CFH). CFH interacting with oxidized phospholipids on the RPE membrane modulates the functions of these cells, but the exact role of CFH in RPE cell death and survival remain poorly understood. The aim of this study was to analyze the potential protective mechanism of CFH on RPE cells submitted to oxidative stress. Upon exposure to oxidized lipids 4-HNE (4-hydroxy-2-nonenal) derived from photoreceptors, both the human RPE cell line ARPE-19 and RPE cells derived from human induced pluripotent stem cells were protected from death only in the presence of the full length human recombinant CFH in the culture medium. This protective effect was independent from the membrane attack complex (MAC) formation. CFH maintained RPE cells tight junctions' structure and regulated the caspase dependent apoptosis process. These results demonstrated the CFH anti-oxidative stress functions independently of its capacity to inhibit MAC formation.


Assuntos
Fator H do Complemento/farmacologia , Complexo de Ataque à Membrana do Sistema Complemento/efeitos dos fármacos , Epitélio Pigmentado da Retina/efeitos dos fármacos , Aldeídos/farmacologia , Apoptose/efeitos dos fármacos , Western Blotting , Caspases/metabolismo , Morte Celular/efeitos dos fármacos , Linhagem Celular , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Humanos , Células-Tronco Pluripotentes Induzidas/efeitos dos fármacos , Células-Tronco Pluripotentes Induzidas/metabolismo , Microscopia Eletrônica de Transmissão , Estresse Oxidativo/efeitos dos fármacos , Reação em Cadeia da Polimerase em Tempo Real , Proteínas Recombinantes , Epitélio Pigmentado da Retina/metabolismo , Junções Íntimas/efeitos dos fármacos
17.
Mol Microbiol ; 112(4): 1253-1269, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31376198

RESUMO

Proteins of the aegerolysin family have a high abundance in Fungi. Due to their specific binding to membrane lipids, and their membrane-permeabilization potential in concert with protein partner(s) belonging to a membrane-attack-complex/perforin (MACPF) superfamily, they were proposed as useful tools in different biotechnological and biomedical applications. In this work, we performed functional studies on expression of the genes encoding aegerolysin and MACPF-like proteins in Aspergillus niger. Our results suggest the sporulation process being crucial for strong induction of the expression of all these genes. However, deletion of either of the aegerolysin genes did not influence the growth, development, sporulation efficiency and phenotype of the mutants, indicating that aegerolysins are not key factors in the sporulation process. In all our expression studies we noticed a strong correlation in the expression of one aegerolysin and MACPF-like gene. Aegerolysins were confirmed to be secreted from the fungus. We also showed the specific interaction of a recombinant A. niger aegerolysin with an invertebrate-specific membrane sphingolipid. Moreover, using this protein labelled with mCherry we successfully stained insect cells membranes containing this particular sphingolipid. Our combined results suggest, that aegerolysins in this species, and probably also in other aspergilli, could be involved in defence against predators.


Assuntos
Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Proteínas Fúngicas/metabolismo , Proteínas Hemolisinas/metabolismo , Perforina/metabolismo , Aspergillus niger/genética , Aspergillus niger/metabolismo , Complexo de Ataque à Membrana do Sistema Complemento/genética , Proteínas Fúngicas/fisiologia , Regulação Fúngica da Expressão Gênica/genética , Proteínas Hemolisinas/fisiologia , Proteínas de Membrana/metabolismo , Perforina/genética , Esfingolipídeos/metabolismo , Esporos Fúngicos/genética , Esporos Fúngicos/metabolismo
18.
Clin Exp Immunol ; 198(3): 359-366, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31461782

RESUMO

The complement system is now a therapeutic target for the management of serious and life-threatening conditions such as paroxysmal nocturnal hemoglobinuria, atypical hemolytic uremic syndrome, glomerulonephritis and other diseases caused by complement deficiencies or genetic variants. As complement therapeutics expand into more clinical conditions, monitoring complement activation is increasingly important, as is the baseline levels of complement activation fragments in blood or other body fluid levels. Although baseline complement levels have been reported in the literature, the majority of these data were generated using non-standard assays and with variable sample handling, potentially skewing results. In this study, we examined the plasma and serum levels of the soluble membrane attack complex of complement (sMAC). sMAC is formed in the fluid phase when complement is activated through the terminal pathway. It binds the regulatory proteins vitronectin and/or clusterin and cannot insert into cell membranes, and can serve as a soluble diagnostic marker in infectious disease settings, as previously shown for intraventricular shunt infections. Here we show that in healthy adults, serum sMAC levels were significantly higher than those in plasma, that plasma sMAC levels were similar between in African Americans and Caucasians and that plasma sMAC levels increase with age. Plasma sMAC levels were significantly higher in virally suppressed people living with HIV (PLWH) compared to non-HIV infected healthy donors. More specifically, PLWH with CD4+ T cell counts below 200 had even greater sMAC levels, suggesting diagnostic value in monitoring sMAC levels in this group.


Assuntos
Ativação do Complemento/imunologia , Complexo de Ataque à Membrana do Sistema Complemento/imunologia , Infecções por HIV/imunologia , Reconstituição Imune/imunologia , Adulto , Síndrome Hemolítico-Urêmica Atípica/sangue , Síndrome Hemolítico-Urêmica Atípica/diagnóstico , Síndrome Hemolítico-Urêmica Atípica/imunologia , Biomarcadores/sangue , Clusterina/sangue , Clusterina/imunologia , Complexo de Ataque à Membrana do Sistema Complemento/genética , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Feminino , Infecções por HIV/sangue , Infecções por HIV/metabolismo , Hemoglobinúria Paroxística/sangue , Hemoglobinúria Paroxística/diagnóstico , Hemoglobinúria Paroxística/imunologia , Humanos , Masculino , Vitronectina/sangue , Vitronectina/imunologia , Adulto Jovem
19.
BMC Nephrol ; 20(1): 307, 2019 08 07.
Artigo em Inglês | MEDLINE | ID: mdl-31390992

RESUMO

BACKGROUND: Atypical hemolytic uremic syndrome (aHUS) is a disorder of the microvasculature with hemolytic anemia, thrombocytopenia and acute kidney injury. Nowadays, aHUS is successfully treated with eculizumab, a humanized, chimeric IgG2/4 kappa antibody, which binds human complement C5 and blocks generation of C5a and membrane-attack-complex. CASE PRESENTATION: A 25-year-old woman with end stage renal disease due to relapsing atypical hemolytic uremic syndrome had a relapse of the disease during pregnancy. She was treated with eculizumab. We measured reduced formation of the membrane-attack complex in newborn's umbilical cord vein blood using the sensitive and specific Palarasah-Nielsen-ELISA. CONCLUSIONS: Eculizumab treatment of the mother with end stage renal disease may cause reduced innate immunity which could render newborns more susceptible to infections.


Assuntos
Anticorpos Monoclonais Humanizados/uso terapêutico , Síndrome Hemolítico-Urêmica Atípica/tratamento farmacológico , Inativadores do Complemento/uso terapêutico , Complexo de Ataque à Membrana do Sistema Complemento/efeitos dos fármacos , Complicações na Gravidez/tratamento farmacológico , Adulto , Anticorpos Monoclonais Humanizados/metabolismo , Síndrome Hemolítico-Urêmica Atípica/imunologia , Complemento C3/metabolismo , Complemento C5a/metabolismo , Complemento C9/metabolismo , Inativadores do Complemento/metabolismo , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Ensaio de Imunoadsorção Enzimática/métodos , Feminino , Sangue Fetal/imunologia , Humanos , Recém-Nascido , Falência Renal Crônica/tratamento farmacológico , Gravidez , Recidiva
20.
Bioessays ; 41(10): e1900074, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31452228

RESUMO

The human immune system can directly lyse invading micro-organisms and aberrant host cells by generating pores in the cell envelope, called membrane attack complexes (MACs). Recent studies using single-particle cryoelectron microscopy have revealed that the MAC is an asymmetric, flexible pore and have provided a structural basis on how the MAC ruptures single lipid membranes. Despite these insights, it remains unclear how the MAC ruptures the composite cell envelope of Gram-negative bacteria. Recent functional studies on Gram-negative bacteria elucidate that local assembly of MAC pores by surface-bound C5 convertase enzymes is essential to stably insert these pores into the bacterial outer membrane (OM). These convertase-generated MAC pores can subsequently efficiently damage the bacterial inner membrane (IM), which is essential for bacterial killing. This review summarizes these recent insights of MAC assembly and discusses how MAC pores kill Gram-negative bacteria. Furthermore, this review elaborates on how MAC-dependent OM damage could lead to IM destabilization, which is currently not well understood. A better understanding on how MAC pores kill bacteria could facilitate the future development of novel strategies to treat infections with Gram-negative bacteria.


Assuntos
Membrana Externa Bacteriana , Complexo de Ataque à Membrana do Sistema Complemento/fisiologia , Bactérias Gram-Negativas , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Humanos
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