RESUMO
There is a close cross-talk between complement, Toll-like receptors (TLRs) and coagulation. The role of the central complement component 5 (C5) in physiological and pathophysiological hemostasis has not, however, been fully elucidated. This study examined the effects of C5 in normal hemostasis and in Escherichia coli-induced coagulation and tissue factor (TF) up-regulation. Fresh whole blood obtained from six healthy donors and one C5-deficient individual (C5D) was anti-coagulated with the thrombin inhibitor lepirudin. Blood was incubated with or without E. coli in the presence of the C5 inhibitor eculizumab, a blocking anti-CD14 monoclonal antibody (anti-CD14) or the TLR-4 inhibitor eritoran. C5D blood was reconstituted with purified human C5. TF mRNA was measured by quantitative polymerase chain reaction (qPCR) and monocyte TF and CD11b surface expression by flow cytometry. Prothrombin fragment 1+2 (PTF1·2) in plasma and microparticles exposing TF (TF-MP) was measured by enzyme-linked immunosorbent assay (ELISA). Coagulation kinetics were analyzed by rotational thromboelastometry and platelet function by PFA-200. Normal blood with eculizumab as well as C5D blood with or without reconstitution with C5 displayed completely normal biochemical hemostatic patterns. In contrast, E. coli-induced TF mRNA and TF-MP were significantly reduced by C5 inhibition. C5 inhibition combined with anti-CD14 or eritoran completely inhibited the E. coli-induced monocyte TF, TF-MP and plasma PTF1·2. Addition of C5a alone did not induce TF expression on monocytes. In conclusion, C5 showed no impact on physiological hemostasis, but substantially contributed to E. coli-induced procoagulant events, which were abolished by the combined inhibition of C5 and CD14 or TLR-4.
Assuntos
Células Sanguíneas/fisiologia , Complemento C5/metabolismo , Infecções por Escherichia coli/imunologia , Escherichia coli/fisiologia , Hemostasia/fisiologia , Sepse/imunologia , Receptor 4 Toll-Like/metabolismo , Anticorpos Bloqueadores/farmacologia , Anticorpos Monoclonais Humanizados/farmacologia , Células Sanguíneas/efeitos dos fármacos , Coagulação Sanguínea , Células Cultivadas , Dissacarídeos/farmacologia , Feminino , Hirudinas/farmacologia , Humanos , Receptores de Lipopolissacarídeos/imunologia , Masculino , Testes de Função Plaquetária , Receptor Cross-Talk , Proteínas Recombinantes/farmacologia , Fosfatos Açúcares/farmacologia , Tromboelastografia , Tromboplastina/genética , Tromboplastina/metabolismo , Receptor 4 Toll-Like/antagonistas & inibidoresRESUMO
Kidney transplanted patients still have significantly higher mortality compared with the general population. The innate immune system may play an important role during periods, with suppression of the adaptive immune system. In the present study, two soluble pattern recognition molecules of the innate immune system were investigated, collectin liver 1 (CL-L1) and collectin kidney 1 (CL-K1). Potential associations of their pretransplant levels and long-term graft and recipient survival were examined. The levels of CL-L1 and CL-K1 were measured at the time of transplantation in 382 patients (≥17 years) transplanted in 2000-2001. The cohort was subsequently followed until December 31, 2014. Data on patient and graft survival were obtained from the Norwegian Renal Registry. Both high CL-L1 (≥376 ng/mL) and high CL-K1 (≥304 ng/mL) levels were significantly associated with overall mortality in multivariate Cox analyses with hazard ration (HR) 1.50, 95% confidence interval (CI) 1.09-2.07, p = 0.013 and HR 1.43, 95% CI 1.02-1.99, p = 0.038, respectively. Moreover, high CL-K1 levels were significantly associated with cardiovascular mortality. No association between measured biomarkers and death-censored graft loss was found. Finally, there was a significant correlation between these two collectins, r = 0.83 (95% CI 0.80-0.86). In conclusion, CL-L1 and CL-K1 were significantly associated with mortality in kidney transplant recipients.
Assuntos
Doenças Cardiovasculares/mortalidade , Colectinas/metabolismo , Rejeição de Enxerto/mortalidade , Falência Renal Crônica/cirurgia , Transplante de Rim/mortalidade , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Doenças Cardiovasculares/etiologia , Criança , Pré-Escolar , Estudos de Coortes , Lectina de Ligação a Manose da Via do Complemento , Feminino , Seguimentos , Rejeição de Enxerto/etiologia , Sobrevivência de Enxerto , Humanos , Lactente , Testes de Função Renal , Transplante de Rim/efeitos adversos , Masculino , Pessoa de Meia-Idade , Prognóstico , Fatores de Risco , Taxa de Sobrevida , Adulto JovemRESUMO
This study aimed to examine whether acute intermittent porphyria (AIP) is associated with systemic inflammation and whether the inflammation correlates with disease activity. A case-control study with 50 AIP cases and age-, sex- and place of residence-matched controls was performed. Plasma cytokines, insulin and C-peptide were analysed after an overnight fast using multiplex assay. Long pentraxin-3 (PTX3) and complement activation products (C3bc and TCC) were analysed using enzyme-linked immunosorbent assay (ELISA). Urine porphobilinogen ratio (U-PBG, µmol/mmol creatinine), haematological and biochemical tests were performed using routine methods. Questionnaires were used to register AIP symptoms, medication and other diseases. All 27 cytokines, chemokines and growth factors investigated were increased significantly in symptomatic AIP cases compared with controls (P < 0·0004). Hierarchical cluster analyses revealed a cluster with high visfatin levels and several highly expressed cytokines including interleukin (IL)-17, suggesting a T helper type 17 (Th17) inflammatory response in a group of AIP cases. C3bc (P = 0·002) and serum immunoglobulin (Ig)G levels (P = 0·03) were increased significantly in cases with AIP. The U-PBG ratio correlated positively with PTX3 (r = 0·38, P = 0·006), and with terminal complement complex (TCC) levels (r = 0·33, P = 0·02). PTX3 was a significant predictor of the biochemical disease activity marker U-PBG in AIP cases after adjustment for potential confounders in multiple linear regression analyses (P = 0·032). Prealbumin, C-peptide, insulin and kidney function were all decreased in the symptomatic AIP cases, but not in the asymptomatic cases. These results indicate that AIP is associated with systemic inflammation. Decreased C-peptide levels in symptomatic AIP cases indicate that reduced insulin release is associated with enhanced disease activity and reduced kidney function.
Assuntos
Inflamação/sangue , Porfiria Aguda Intermitente/sangue , Biomarcadores/sangue , Peptídeo C/sangue , Estudos de Casos e Controles , Citocinas/sangue , Feminino , Humanos , Imunoglobulina G/sangue , Inflamação/imunologia , Inflamação/metabolismo , Insulina/sangue , Rim/imunologia , Rim/fisiopatologia , Masculino , Pessoa de Meia-Idade , Porfiria Aguda Intermitente/imunologia , Porfiria Aguda Intermitente/metabolismo , Pré-Albumina/metabolismo , Linfócitos T Auxiliares-Indutores/imunologiaRESUMO
Inhibition of complement factor 5 (C5) reduced myocardial infarction in animal studies, while no benefit was found in clinical studies. Due to lack of cross-reactivity of clinically used C5 antibodies, different inhibitors were used in animal and clinical studies. Coversin (Ornithodoros moubata complement inhibitor, OmCI) blocks C5 cleavage and binds leukotriene B4 in humans and pigs. We hypothesized that inhibition of C5 before reperfusion will decrease infarct size and improve ventricular function in a porcine model of myocardial infarction. In pigs (Sus scrofa), the left anterior descending coronary artery was occluded (40 min) and reperfused (240 min). Coversin or placebo was infused 20 min after occlusion and throughout reperfusion in 16 blindly randomized pigs. Coversin significantly reduced myocardial infarction in the area at risk by 39% (p = 0.03, triphenyl tetrazolium chloride staining) and by 19% (p = 0.02) using magnetic resonance imaging. The methods correlated significantly (R = 0.92, p < 0.01). Tissue Doppler echocardiography showed increased systolic displacement (31%, p < 0.01) and increased systolic velocity (29%, p = 0.01) in coversin treated pigs. Interleukin-1ß in myocardial microdialysis fluid was significantly reduced (31%, p < 0.05) and tissue E-selectin expression was significantly reduced (p = 0.01) in the non-infarcted area at risk by coversin treatment. Coversin ablated plasma C5 activation throughout the reperfusion period and decreased myocardial C5b-9 deposition, while neither plasma nor myocardial LTB4 were significantly reduced. Coversin substantially reduced the size of infarction, improved ventricular function, and attenuated interleukin-1ß and E-selectin in this porcine model by inhibiting C5. We conclude that inhibition of C5 in myocardial infarction should be reconsidered.
Assuntos
Complemento C5/antagonistas & inibidores , Infarto do Miocárdio/patologia , Animais , Proteínas de Artrópodes/farmacologia , Proteínas de Transporte/farmacologia , Modelos Animais de Doenças , Ecocardiografia , Imunofluorescência , Imageamento por Ressonância Magnética , Distribuição Aleatória , Sus scrofaRESUMO
The 22q11.2 deletion syndrome (22q11.2 DS), also known as DiGeorge syndrome, is a genetic disorder with an estimated incidence of 1:4000 births. These patients may suffer from affection of many organ systems with cardiac malformations, thymic hypoplasia or aplasia, hypoparathyroidism, palate anomalies and psychiatric disorders being the most frequent. The incidence of autoimmune diseases is increased in older patients. The aim of the present study was to examine a cytokine profile in patients with 22q11.2 DS by measuring a broad spectrum of serum cytokines. Patients with a proven deletion of chromosome 22q11.2 (n = 55) and healthy individuals (n = 54) recruited from an age- and sex-comparable group were included in the study. Serum levels of 27 cytokines, including chemokines and growth factors, were analysed using multiplex technology. Interferon-inducible protein 10 (IP-10) was also measured by ELISA to confirm the multiplex results. The 22q11.2 DS patients had distinctly and significantly raised levels of pro-inflammatory and angiostatic chemokine IP-10 (P < 0.001) compared to controls. The patients with congenital heart defects (n = 31) had significantly (P = 0.018) raised serum levels of IP-10 compared to patients born without heart defects (n = 24). The other cytokines investigated were either not detectable or did not differ between patients and controls.
Assuntos
Quimiocina CXCL10/metabolismo , Síndrome de DiGeorge/imunologia , Cardiopatias Congênitas/imunologia , Adolescente , Adulto , Células Cultivadas , Quimiocinas/sangue , Criança , Pré-Escolar , Estudos de Coortes , Síndrome de DiGeorge/complicações , Síndrome de DiGeorge/diagnóstico , Feminino , Cardiopatias Congênitas/diagnóstico , Cardiopatias Congênitas/etiologia , Humanos , Lactente , Mediadores da Inflamação/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/sangue , Masculino , Pessoa de Meia-Idade , Adulto JovemRESUMO
Complement activation and low complement levels are common in systemic lupus erythematosus (SLE). Antiphospholipid antibodies (aPL) are found in about 30-40% of patients with SLE. This study aimed to investigate the association between aPL and complement levels in patients with SLE. Serum samples were collected from 269 patients with SLE enrolled in the Norwegian Systemic Connective Tissue and Vasculitis Registry (NOSVAR) during 2003-2009, and from 353 controls. All samples were analysed for anti-ß2 glycoprotein 1 (anti-ß2GP1) and anticardiolipin antibodies (aCL), C-reactive protein (CRP) and complement components C3 and C4. Median CRP level was significantly higher in cases than in controls (2.06 versus 0.90 mg/l; P < 0.0001). No significant difference in CRP was found between SLE patients with or without aPL (2.09 versus 1.89; P = 0.665). Median C3 levels were similar in cases (1.03 g/l) and controls (1.00 g/l), whereas median C4 levels were 0.16 g/l in cases versus. 0.19 in controls (P < 0.0001). However, aPL-positive SLE patients had significantly lower median C3 levels (0.92 versus. 1.07 g/l; P = 0.001) and C4 levels (0.11 versus 0.16 g/l; P < 0.0001) compared to aPL-negative patients. Lower C3 and C4 values in aPL-positive SLE patients may reflect a higher consumption of C3 and C4 due to more pronounced complement activation in aPL-positive SLE patients compared to SLE patients without aPL.
Assuntos
Anticorpos Antifosfolipídeos/sangue , Complemento C3/metabolismo , Complemento C4/metabolismo , Lúpus Eritematoso Sistêmico/imunologia , Adolescente , Adulto , Idoso , Proteína C-Reativa/metabolismo , Ativação do Complemento , Feminino , Humanos , Masculino , Noruega , Adulto JovemRESUMO
The complement system can be activated via the lectin pathway by the recognition molecules mannose-binding lectin (MBL) and the ficolins. Ficolin-2 exhibits binding against a broad range of ligands, including biomaterials in vitro, and low ficolin-2 levels are associated with increased risk of infections. Thus, we investigated the biocompatibility of the recognition molecules of the lectin pathway in two different types of cardiopulmonary bypass circuits. Bloods were drawn at five time-points before, during and postoperatively from 30 patients undergoing elective cardiac surgery. Patients were randomized into two groups using different coatings of cardiopulmonary bypass circuits, Phisio® (phosphorylcholine polymer coating) and Bioline® (albumin-heparin coating). Concentrations of MBL, ficolin-1, -2 and -3 and soluble C3a and terminal complement complex (TCC) in plasma samples were measured. Ficolin-3-mediated complement activation potential was evaluated with C4, C3 and TCC as output. There was no significant difference between the two circuit materials regarding MBL, ficolin-1 and -3. In the Bioline® group the ficolin-2 levels decreased significantly after initiation of surgery (P < 0.0001) and remained reduced throughout the sampling period. This was not seen for Phisio®-coated circuits. Ficolin-3-mediated complement activation potential was reduced significantly in both groups after start of operation (P < 0.0001), whereas soluble C3a and TCC in the samples were increased (P < 0.0001). Ficolin-2 was depleted from plasma during cardiac surgery when using heparin-coated bypass circuits and did not reach baseline level 24 h postoperation. These findings may have implications for the postoperative susceptibility to infections in patients undergoing extracorporeal circulation procedures.
Assuntos
Anticoagulantes , Ponte Cardiopulmonar , Lectina de Ligação a Manose da Via do Complemento , Stents Farmacológicos , Heparina , Lectinas/sangue , Proteínas do Sistema Complemento/metabolismo , Feminino , Glicoproteínas/sangue , Humanos , Masculino , Período Pós-Operatório , Fatores de Tempo , FicolinasRESUMO
The complement system and the Toll-like (TLR) co-receptor CD14 play important roles in innate immunity and sepsis. Tissue factor (TF) is a key initiating component in intravascular coagulation in sepsis, and long pentraxin 3 (PTX3) enhances the lipopolysaccharide (LPS)-induced transcription of TF. The aim of this study was to study the mechanism by which complement and CD14 affects LPS- and Escherichia coli (E. coli)-induced coagulation in human blood. Fresh whole blood was anti-coagulated with lepirudin, and incubated with ultra-purified LPS (100 ng/ml) or with E. coli (1 × 10(7) /ml). Inhibitors and controls included the C3 blocking peptide compstatin, an anti-CD14 F(ab')2 antibody and a control F(ab')2 . TF mRNA was measured using quantitative polymerase chain reaction (qPCR) and monocyte TF surface expression by flow cytometry. TF functional activity in plasma microparticles was measured using an amidolytic assay. Prothrombin fragment F 1+2 (PTF1.2) and PTX3 were measured by enzyme-linked immunosorbent assay (ELISA). The effect of TF was examined using an anti-TF blocking antibody. E. coli increased plasma PTF1.2 and PTX3 levels markedly. This increase was reduced by 84->99% with compstatin, 55-97% with anti-CD14 and > 99% with combined inhibition (P < 0·05 for all). The combined inhibition was significantly (P < 0·05) more efficient than compstatin and anti-CD14 alone. The LPS- and E. coli-induced TF mRNA levels, monocyte TF surface expression and TF functional activity were reduced by > 99% (P < 0·05) with combined C3 and CD14 inhibition. LPS- and E. coli-induced PTF1.2 was reduced by 76-81% (P < 0·05) with anti-TF antibody. LPS and E. coli activated the coagulation system by a complement- and CD14-dependent up-regulation of TF, leading subsequently to prothrombin activation.
Assuntos
Coagulação Sanguínea/imunologia , Proteína C-Reativa/imunologia , Escherichia coli/imunologia , Receptores de Lipopolissacarídeos/imunologia , Componente Amiloide P Sérico/imunologia , Tromboplastina/imunologia , Antitrombinas/farmacologia , Complemento C3/antagonistas & inibidores , Complemento C3/imunologia , Hirudinas/farmacologia , Humanos , Receptores de Lipopolissacarídeos/metabolismo , Lipopolissacarídeos , Fragmentos de Peptídeos/imunologia , Peptídeos Cíclicos/farmacologia , Protrombina/imunologia , RNA Mensageiro/genética , Proteínas Recombinantes/farmacologia , Sepse/imunologia , Sepse/microbiologia , Tromboplastina/biossíntese , Tromboplastina/genética , Regulação para CimaRESUMO
Atypical haemolytic uraemic syndrome (aHUS) is associated with (genetic) alterations in alternative complement pathway. Nevertheless, comprehensive evidence that the complement system in aHUS patients is more prone to activation is still lacking. Therefore, we performed a thorough analysis of complement activation in acute phase and in remission of this disease. Complement activation patterns of the aHUS patients in acute phase and in remission were compared to those of healthy controls. Background levels of complement activation products C3b/c, C3bBbP and terminal complement complex (TCC) were measured using enzyme-linked immunosorbent assay (ELISA) in ethylenediamine tetraacetic acid (EDTA) plasma. In vitro-triggered complement activation in serum samples was studied using zymosan-coating and pathway-specific assay. Furthermore, efficiencies of the C3b/c, C3bBbP and TCC generation in fluid phase during spontaneous activation were analysed. Patients with acute aHUS showed elevated levels of C3b/c (P < 0·01), C3bBbP (P < 0·0001) and TCC (P < 0·0001) in EDTA plasma, while values of patients in remission were normal, compared to those of healthy controls. Using data from a single aHUS patient with complement factor B mutation we illustrated normalization of complement activation during aHUS recovery. Serum samples from patients in remission showed normal in vitro patterns of complement activation and demonstrated normal kinetics of complement activation in the fluid phase. Our data indicate that while aHUS patients have clearly activated complement in acute phase of the disease, this is not the case in remission of aHUS. This knowledge provides important insight into complement regulation in aHUS and may have an impact on monitoring of these patients, particularly when using complement inhibition therapy.
Assuntos
Síndrome Hemolítico-Urêmica Atípica/imunologia , Ativação do Complemento , Complemento C3b/metabolismo , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Doença Aguda , Adolescente , Adulto , Síndrome Hemolítico-Urêmica Atípica/sangue , Síndrome Hemolítico-Urêmica Atípica/patologia , Síndrome Hemolítico-Urêmica Atípica/terapia , Estudos de Casos e Controles , Criança , Pré-Escolar , Ativação do Complemento/efeitos dos fármacos , Fator B do Complemento/metabolismo , Fator H do Complemento/metabolismo , Via Alternativa do Complemento/efeitos dos fármacos , Feminino , Humanos , Lactente , Masculino , Pessoa de Meia-Idade , Plasma Rico em Plaquetas , Isoformas de Proteínas/sangue , Indução de Remissão , Diálise Renal , Zimosan/farmacologiaRESUMO
Congestive heart failure is associated with increased levels of several inflammatory mediators, and animal studies have shown that infusion of a number of cytokines can induce heart failure. However, several drugs with proven efficacy in heart failure have failed to affect inflammatory mediators, and anti-inflammatory therapy in heart failure patients has thus far been disappointing. Hence, to what extent heart failure is caused by or responsible for the increased inflammatory burden in the patient is still unclear. Over the past couple of decades, resynchronization therapy with a biventricular pacemaker has emerged as an effective treatment in a subset of heart failure patients, reducing both morbidity and mortality. Such treatment has also been shown to affect the inflammation associated with heart failure. In this study, we review recent data on the association between heart failure and inflammation, and in particular how resynchronization therapy can affect the inflammatory process.
Assuntos
Dispositivos de Terapia de Ressincronização Cardíaca , Terapia de Ressincronização Cardíaca/métodos , Citocinas/uso terapêutico , Insuficiência Cardíaca/terapia , Inflamação/terapia , Anti-Inflamatórios/uso terapêutico , Citocinas/metabolismo , Insuficiência Cardíaca/fisiopatologia , Humanos , Mediadores da Inflamação/imunologia , Resultado do TratamentoRESUMO
Microdialysis is an excellent tool to assess tissue inflammation in patients, but in vitro systems to evaluate recovery of inflammatory mediators have not been standardized. We aimed to develop a reference plasma preparation and evaluate different perfusion fluids with respect to recovery of metabolic and inflammatory markers. The reference preparation was produced by incubation of human blood with lipopolysaccharide and cobra venom factor to generate cytokines and activate complement, respectively. Microdialysis with 100 kDa catheters was performed using different colloid and crystalloid perfusion fluids (hydroxyethyl starch (HES) 130/0.4, HES 200/0.5, hyperosmolar HES 200/0.5, albumin 200 g/l, T1 perfusion fluid and Ringer's acetate) compared to today's recommended dextran 60 solution. Recovery of glucose, glycerol and pyruvate was not significantly different between the perfusion fluids, whereas lactate had lower recovery in HES 200/0.5 and albumin perfusion fluids. Recovery rates for the inflammatory proteins in comparison with the concentration in the reference preparation differed substantially: IL-6 = 9%, IL-1ß = 18%, TNF = 0.3%, MCP-1 = 45%, IL-8 = 48%, MIG = 48%, IP-10 = 25%, C3a = 53% and C5a = 12%. IL-10 was not detectable in microdialysis dialysate. HES 130/0.4 and HES 200/0.5 yielded a recovery not significantly different from dextran 60. Hyperosmolar HES 200/0.5 and albumin showed significantly different pattern of recovery with increased concentration of MIG, IP-10, C3a and C5a and decreased concentration of IL-1ß, TNF, MCP-1 and IL-8 in comparison with dextran 60. In conclusion, microdialysis perfusion fluid dextran 60 can be replaced by the commonly used HES 130/0.4, whereas albumin might be used if specific immunological variables are in focus. The present reference plasma preparation is suitable for in vitro evaluation of microdialysis systems.
Assuntos
Citocinas/metabolismo , Inflamação/diagnóstico , Leucócitos Mononucleares/imunologia , Microdiálise/métodos , Perfusão , Albuminas/metabolismo , Células Cultivadas , Proteínas do Sistema Complemento/metabolismo , Venenos Elapídicos/metabolismo , Humanos , Derivados de Hidroxietil Amido/metabolismo , Inflamação/imunologia , Mediadores da Inflamação/metabolismo , Lipopolissacarídeos/metabolismo , Microdiálise/normas , Plasma/metabolismo , Padrões de ReferênciaRESUMO
Complement convertases are enzymatic complexes that play a central role in sustaining and amplification of the complement cascade. Impairment of complement function leads directly or indirectly to pathological conditions, including higher infection rate, kidney diseases, autoimmune- or neurodegenerative diseases and ischaemia-reperfusion injury. An assay for direct measurement of activity of the convertases in patient sera is not available. Existing assays testing convertase function are based on purified complement components and, thus, convertase formation occurs under non-physiological conditions. We designed a new assay, in which C5 blocking compounds enabled separation of the complement cascade into two phases: the first ending at the stage of C5 convertases and the second ending with membrane attack complex formation. The use of rabbit erythrocytes or antibody-sensitized sheep erythrocytes as the platforms for convertase formation enabled easy readout based on measurement of haemolysis. Thus, properties of patient sera could be studied directly regarding convertase activity and membrane attack complex formation. Another advantage of this assay was the possibility to screen for host factors such as C3 nephritic factor and other anti-complement autoantibodies, or gain-of-function mutations, which prolong the half-life of complement convertases. Herein, we present proof of concept, detailed description and validation of this novel assay.
Assuntos
Convertases de Complemento C3-C5/análise , Eritrócitos/enzimologia , Imunoensaio/métodos , Animais , Autoanticorpos/imunologia , Fator Nefrítico do Complemento 3/imunologia , Convertases de Complemento C3-C5/imunologia , Via Alternativa do Complemento/imunologia , Proteínas do Sistema Complemento/imunologia , Eritrócitos/imunologia , Cobaias , Meia-Vida , Humanos , Coelhos , OvinosRESUMO
Both the complement system and tissue factor (TF), a key initiating component of coagulation, are activated in sepsis, and cross-talk occurs between the complement and coagulation systems. C1-inhibitor (C1-INH) can act as a regulator in both systems. Our aim in this study was to examine this cross-talk by investigating the effects of C1-INH on Escherichia coli-induced haemostasis and inflammation. Fresh human whole blood collected in lepirudin was incubated with E. coli or ultrapurified E. coli lipopolysaccharide (LPS) in the absence or presence of C1-INH or protease-inactivated C1-INH. C3 activation was blocked by compstatin, a specific C3 convertase inhibitor. TF mRNA was measured using reverse transcription-quantitative polymerase chain reaction (RT-qPCR), and TF surface expression was measured by flow cytometry. In plasma, the terminal complement complex, prothrombin F1·2 (PTF1·2) and long pentraxin 3 (PTX3) were measured by enzyme-linked immunosorbent assay (ELISA). Cytokines were analysed using a multiplex kit. C1-INH (1·25-5 mg/ml) reduced both LPS- and E. coli-induced coagulation, measured as a reduction of PTF1·2 in plasma, efficiently and dose-dependently (P < 0·05). Both LPS and E. coli induced marked up-regulation of TF mRNA levels and surface expression on whole blood monocytes. This up-regulation was reduced efficiently by treatment with C1-INH (P < 0·05). C1-INH reduced the release of PTX3 (P < 0·05) and virtually all cytokines measured (P < 0·05). Complement activation was inhibited more efficiently with compstatin than with C1-INH. C1-INH inhibited most of the other readouts more efficiently, consistent with additional non-complement-dependent effects. These results indicate that complement plays a role in activating coagulation during sepsis and that C1-INH is a broad-spectrum attenuator of the inflammatory and haemostatic responses.
Assuntos
Proteínas Inativadoras do Complemento 1/metabolismo , Escherichia coli/imunologia , Monócitos/imunologia , Sepse/imunologia , Tromboplastina/metabolismo , Coagulação Sanguínea , Proteína C-Reativa/metabolismo , Células Cultivadas , Técnicas de Cocultura , Proteínas Inativadoras do Complemento 1/genética , Proteína Inibidora do Complemento C1 , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Lipopolissacarídeos/imunologia , Masculino , Monócitos/efeitos dos fármacos , Fragmentos de Peptídeos/sangue , Protrombina , RNA Mensageiro/análise , Sepse/tratamento farmacológico , Componente Amiloide P Sérico/metabolismo , Tromboplastina/genéticaRESUMO
Atrial fibrillation is highly prevalent, and affected patients are at an increased risk of a number of complications, including heart failure and thrombo-embolism. Over the past years, there has been increasing interest in the role of inflammatory processes in atrial fibrillation, from the first occurrence of the arrhythmia to dreaded complications such as strokes or peripheral emboli. As the standard drug combination which aims at rate control and anticoagulation only offers partial protection against complications, newer agents are needed to optimize treatment. In this paper, we review recent knowledge regarding the impact of inflammation on the occurrence, recurrence, perpetuation and complications of the arrhythmia, as well as the role of anti-inflammatory therapies in the treatment for the disease.
Assuntos
Arritmias Cardíacas/fisiopatologia , Fibrilação Atrial/fisiopatologia , Insuficiência Cardíaca/fisiopatologia , Acidente Vascular Cerebral/fisiopatologia , Tromboembolia/fisiopatologia , Animais , Antiarrítmicos/uso terapêutico , Anti-Inflamatórios/uso terapêutico , Anticoagulantes/uso terapêutico , Arritmias Cardíacas/tratamento farmacológico , Arritmias Cardíacas/etiologia , Fibrilação Atrial/complicações , Fibrilação Atrial/tratamento farmacológico , Insuficiência Cardíaca/tratamento farmacológico , Insuficiência Cardíaca/etiologia , Humanos , Inflamação/tratamento farmacológico , Acidente Vascular Cerebral/tratamento farmacológico , Acidente Vascular Cerebral/etiologia , Tromboembolia/tratamento farmacológico , Tromboembolia/etiologiaRESUMO
BACKGROUND: The complement pathway and CD14 play essential roles in inflammation, but little is known about the relative roles of complement and CD14 in E. coli-induced tissue factor (TF) mRNA upregulation, expression by monocytes, and functional activity in human whole blood. METHODS: Whole E. coli bacteria were incubated for up to 4 h in human whole blood containing the anticoagulant lepirudin, which does not affect complement activation. TF mRNA levels were analyzed using reverse transcription, quantitative real-time PCR (RT-qPCR), and the expression of TF on the cell surface was analyzed using flow cytometry. Complement was selectively inhibited using the C3 convertase inhibitor compstatin or a C5a receptor antagonist (C5aRa), while CD14 was blocked by an anti-CD14 F(ab')2 monoclonal antibody. RESULTS: The E. coli-induced TF mRNA upregulation was reduced to virtually background levels by compstatin, whereas anti-CD14 had no effect. Monocyte TF expression and TF activity in plasma microparticles were significantly reduced by C5aRa. Anti-CD14 alone only slightly reduced E. coli-induced monocyte TF expression but showed a modest additive effect when combined with the complement inhibitors. Inhibiting complement and CD14 efficiently reduced the expression of the E. coli-induced cytokines IL-1beta, IL-6, IL-8, and platelet-derived growth factor bb. CONCLUSION: Our results indicate that E. coli-induced TF mRNA upregulation is mainly dependent on complement activation, while CDI4 plays a modest role in monocyte TF expression and the plasma TF activity in human whole blood.
Assuntos
Inativadores do Complemento/uso terapêutico , Infecções por Escherichia coli/tratamento farmacológico , Infecções por Escherichia coli/metabolismo , Receptores de Lipopolissacarídeos/efeitos dos fármacos , Monócitos/metabolismo , RNA Mensageiro/biossíntese , Tromboplastina/biossíntese , Adulto , Anticoagulantes/farmacologia , Anticoagulantes/uso terapêutico , Proteínas do Sistema Complemento/metabolismo , Citocinas/sangue , Relação Dose-Resposta a Droga , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Hirudinas/farmacologia , Humanos , Monócitos/efeitos dos fármacos , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Proteínas Recombinantes/farmacologia , Proteínas Recombinantes/uso terapêutico , Sepse/tratamento farmacológico , Sepse/microbiologia , Regulação para Cima/efeitos dos fármacosRESUMO
OBJECTIVES: To assess the relationship between interleukin (IL)-1-related molecules, infarct size and left ventricular (LV) remodelling following acute myocardial infarction (MI). METHODS: Forty-two patients with first-time diagnosis of ST segment elevation MI (STEMI), with a single occluded vessel successfully revascularized by primary percutaneous coronary intervention (PCI), were recruited to this observational study conducted at a university teaching hospital and followed for 1 year. MAIN OUTCOME MEASURES: Plasma levels of IL-1ß, IL-1 receptor antagonist (IL-1Ra), IL-18 and caspase-1 were analysed before and 2 days, 1 week and 2 months after PCI. Serial cardiac magnetic resonance imaging (CMR) was used for the assessment of infarct size and LV remodelling. CMR findings at 1 year was the primary outcome variable. RESULTS: Univariate analysis showed that IL-1-related mediators were strongly (IL-1 ß), moderately (caspase-1) and weakly (IL-1Ra) associated with impaired myocardial function and noninfarct mass, but not infarct size, 1 year after reperfused STEMI. In multivariate analyses, troponin T predicted LV ejection fraction (LVEF), infarct size and LV end-diastolic (LVEDVi) and end-systolic volume index (LVESVi). However, significant additional variance was explained by IL-1ß, IL-18 and caspase-1. IL-1ß levels at 2 months, IL-18 at 2 days and pre-PCI caspase-1 were predictors of LVEF. Caspase-1 and in particular IL-1ß at 2 days were the only predictors of noninfarct mass. IL-1ß and IL-18 at 2 days were predictors of LVEDVi, whilst pre-PCI levels of IL-1ß contributed to prediction of LVESVi. By contrast, pro-B-type natriuretic peptide, C-reactive protein, IL-6 and transforming growth factor-ß1 (TGF-ß1) had no or only a weak (TGF-ß1) association with these CMR parameters in multivariate analyses. CONCLUSIONS: IL-1ß levels after STEMI were strongly associated with impaired myocardial function and noninfarct LV mass after 1 year, suggesting a potential role for IL-1ß as a predictor of maladaptive myocardial remodelling following reperfused MI.
Assuntos
Angioplastia Coronária com Balão , Hipertrofia Ventricular Esquerda/sangue , Interleucina-1beta/sangue , Infarto do Miocárdio/terapia , Remodelação Ventricular , Proteína C-Reativa/análise , Caspase 1/sangue , Feminino , Seguimentos , Ventrículos do Coração/patologia , Humanos , Hipertrofia Ventricular Esquerda/patologia , Proteína Antagonista do Receptor de Interleucina 1/sangue , Interleucina-1/sangue , Interleucina-18/sangue , Imagem Cinética por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Infarto do Miocárdio/sangue , Infarto do Miocárdio/patologia , Peptídeo Natriurético Encefálico/sangue , Fragmentos de Peptídeos/sangue , Volume Sistólico , Fator de Crescimento Transformador beta1/sangue , Troponina T/sangueRESUMO
Low-density lipoprotein (LDL) apheresis is an extracorporeal treatment modality used in high-risk patients when LDL cholesterol levels cannot be reduced adequately with medication. The treatment is highly effective, but could be affected by potential unwanted effects on pro- and anti-inflammatory biomarkers. In this paper, we review the literature regarding the effect of LDL apheresis on pro- and anti-inflammatory biomarkers important in atherosclerosis, also as patients in LDL apheresis have high risk for atherosclerotic complications. We discuss the effect of LDL apheresis on complement, cytokines and finally a group of other selected pro- and anti-inflammatory biomarkers. The complement system is affected by LDL apheresis, and there are differences between different LDL apheresis systems. The plasma separation columns seem to trigger the formation of proinflammatory complement factors including C3a and C5a, while the same anaphylatoxins are adsorbed by the LDL apheresis columns, however, to varying degree. Proinflammatory cytokines are to some extent adsorbed by the LDL apheresis columns, while some of the anti-inflammatory cytokines increase during treatment. Finally, we discuss the effect of apheresis on different biomarkers including C-reactive protein, fibrinogen, adhesion molecules, myeloperoxidase and HDL cholesterol. In conclusion, even if there are differences between pro- and anti-inflammatory biomarkers during LDL apheresis, the consequences for the patients are largely unknown and larger studies need to be performed. Preferably, they should be comparing the effect of different LDL apheresis columns on the total inflammatory profile, and this should be related to clinical endpoints.
Assuntos
Aterosclerose/terapia , Remoção de Componentes Sanguíneos/efeitos adversos , LDL-Colesterol/sangue , Inflamação/etiologia , Animais , Biomarcadores/sangue , Citocinas/sangue , Humanos , Hipercolesterolemia/terapiaRESUMO
The study was designed to investigate whether complement is activated in patients subject to rectal surgery and whether the choice of surgical technique (open or laparoscopic) has any impact on the activation of complement. Our hypothesis is that laparoscopic surgery leads to a lower-level activation of complement than open surgery. Patients (n = 24) subject to rectal surgery owing to rectal cancer were included. The study was prospective and randomized. The patients were randomized to either laparoscopic surgery (n = 12) or open surgery (n = 12). Blood samples for determination of complement activation (C4d, Bb, C3bc and the terminal C5b-9 complex TCC) were drawn before start of surgery (T0) and at the following time-points after start of surgery: 180 min (T1), 360 min (T2), 24 h (T3) and 3-5 days (T4). A significant increase in the alternative pathway activation product Bb and in the terminal pathway activation product TCC was seen over time in both groups (P < 0.001). Bb peaked early (T1) and returned to baseline levels post-operatively, whereas TCC increased steadily with maximum values in the late post-operative period. The plasma concentrations of C4d and C3bc decreased significantly in both groups at T1 and T2 and returned to baseline levels at T4. There was no significant difference between the groups. Rectal surgery causes activation of the complement system. Complement is activated through the alternative pathway. Results mostly showed no significant differences between laparoscopic and open rectal surgery apart from lower levels of factor Bb in the former group in the perioperative period.
Assuntos
Via Alternativa do Complemento/imunologia , Procedimentos Cirúrgicos do Sistema Digestório/métodos , Laparoscopia/métodos , Neoplasias Retais/imunologia , Neoplasias Retais/cirurgia , Idoso , Complemento C3b/imunologia , Complemento C3b/metabolismo , Complemento C4b/imunologia , Complexo de Ataque à Membrana do Sistema Complemento/imunologia , Complexo de Ataque à Membrana do Sistema Complemento/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Fragmentos de Peptídeos/sangue , Fragmentos de Peptídeos/imunologia , Estudos Prospectivos , Estatísticas não ParamétricasRESUMO
BACKGROUND: Sepsis is a major world-wide medical problem with high morbidity and mortality. Gram-negative bacteria are among the most important pathogens of sepsis and their LPS content is regarded to be important for the systemic inflammatory reaction. The CD14/myeloid differentiation factor 2 (MD-2)/TLR4 complex plays a major role in the immune response to LPS . The aim of this study was to compare the effects of inhibiting MD-2 and CD14 on ultra-pure LPS - versus whole E. coli bacteria-induced responses. METHODS: Fresh human whole blood was incubated with upLPS or whole E. coli bacteria in the presence of MD-2 or CD14 neutralizing monoclonal antibodies, or their respective controls, and/or the specific complement-inhibitor compstatin. Cytokines were measured by a multiplex (n = 27) assay. NFκB activity was examined in cells transfected with CD14, MD-2 and/or Toll-like receptors. RESULTS: LPS-induced cytokine response was efficiently and equally abolished by MD-2 and CD14 neutralization. In contrast, the response induced by whole E. coli bacteria was only modestly reduced by MD-2 neutralization, whereas CD14 neutralization was more efficient. Combination with compstatin enhanced the effect of MD-2 neutralization slightly. When compstatin was combined with CD14 neutralization, however, the response was virtually abolished for all cytokines, including IL-17, which was only inhibited by this combination. The MD-2-independent effect observed for CD14 could not be explained by TLR2 signaling. CONCLUSION: Inhibition of CD14 is more efficient than inhibition of MD-2 on whole E. coli-induced cytokine response, suggesting CD14 to be a better target for intervention in Gram-negative sepsis, in particular when combined with complement inhibition.
Assuntos
Infecções por Escherichia coli/imunologia , Escherichia coli/imunologia , Receptores de Lipopolissacarídeos/imunologia , Antígeno 96 de Linfócito/imunologia , Sepse/imunologia , Infecções por Escherichia coli/sangue , Humanos , Receptores de Lipopolissacarídeos/metabolismo , Lipopolissacarídeos/farmacologia , Antígeno 96 de Linfócito/metabolismo , Sepse/metabolismoRESUMO
The innate immune system is rapidly activated during myocardial infarction and blockade of extracellular complement system reduces infarct size. Intracellular complement, however, appears to be closely linked to metabolic pathways and its role in ischemia-reperfusion injury is unknown and may be different from complement activation in the circulation. The purpose of the present study was to investigate the role of intracellular complement in isolated, retrogradely buffer-perfused hearts and cardiac cells from adult male wild type mice (WT) and from adult male mice with knockout of complement component 3 (C3KO). Main findings: (i) Intracellular C3 protein was expressed in isolated cardiomyocytes and in whole hearts, (ii) after ischemia-reperfusion injury, C3KO hearts had larger infarct size (32 ± 9% in C3KO vs. 22 ± 7% in WT; p=0.008) and impaired post-ischemic relaxation compared to WT hearts, (iii) C3KO cardiomyocytes had lower basal oxidative respiration compared to WT cardiomyocytes, (iv) blocking mTOR decreased Akt phosphorylation in WT, but not in C3KO cardiomyocytes, (v) after ischemia, WT hearts had higher levels of ATP, but lower levels of both reduced and oxidized nicotinamide adenine dinucleotide (NADH and NAD+, respectively) compared to C3KO hearts. Conclusion: intracellular C3 protected the heart against ischemia-reperfusion injury, possibly due to its role in metabolic pathways important for energy production and cell survival.