Mannan-binding lectin deficiency modulates the humoral immune response dependent on the genetic environment

Mannan-binding lectin (MBL) is a plasma protein implicated in innate immune defence against a broad range of microorganisms, including viruses. It is also thought that MBL plays a role in the recruitment of the specific clonal immune response. This was studied by injecting soluble hepatitis B surface antigen (HBsAg) intravenously into mice deficient in both MBL-A and MBL-C (MBL DKO mice). The MBL DKO animals on mixed genetic background (SV129EvSv × C57BL/6) produced higher antibody titres than the wild-type littermates. After primary challenge with the antigen the immunoglobulin M anti-HBsAg antibody titres were threefold higher in the MBL DKO mice than in the wild-type mice. Following the boost, the immunoglobulin G anti-HBsAg antibody titres were 10-fold higher in the MBL DKO mice, suggesting that MBL plays a role in a negative feedback regulation of adaptive immunity. However, the modulating effect of MBL was dependent on the genetic environment. The MBL DKO mice backcrossed on a C57BL/6 background showed the opposite response with the MBL DKO mice now producing fewer antibodies than the wild-type animals, whereas MBL deficiency in mice with the SV129EvSv background did not show any effect in antibody production. These findings indicate that the modifying effect of MBL on the humoral immune response is influenced by the genetic environment.     

Complement activation in infective endocarditis: correlation with extracardiac manifestations and prognosis

In an infectious process complement activation is necessary for a proper immune and inflammatory response, but when exacerbated may cause tissue injuries. In infective endocarditis (IE) patients tend to develop high titres of circulating immune complexes (CIC) that activate complement. The aim of this study was to evaluate for the first time complement activation in IE for possible correlation with extracardiac manifestations and clinical prognosis. Twenty patients with IE, 14 healthy controls and 15 patients presenting mitral and aortic valve lesions (with no signs of either infection or other associated diseases), were studied. Plasma levels of C3adesArg, SC5b-9, C1rs-C1Inh and C3b(Bb)P were determined by ELISA and C3d by double decker immunoelectrophoresis. C3 and C4 levels were assayed by turbidimetry and CIC by ELISA. Elevation of plasma levels of all complement activation products, with the exception of C3b(Bb)P, indicated a significant classical pathway activation in IE patients when compared to controls (C3d: P < 0.00004; C3adesArg: P < 0.03, SC5b-9: P < 0.01, C1rs-C1Inh: P < 0.00007). CIC levels were significantly increased (P < 0.005) and C3 reduced in IE patients (P < 0.05). Elevated C3d (P < 0.02) and C3adesArg (P < 0.03) levels were associated with pulmonary manifestations. In addition, C3d was significantly elevated in the patients who died when compared to those who had a good recovery (P < 0.02). Our data demonstrate the activation of the complement classical pathway, most probably mediated by CIC, in IE and suggests C3d and C3adesArg as possible markers for extracardiac lesion and severity of the disease.     

甘露聚糖结合凝集素诱导人白血病细胞系U937细胞凋亡

目的研究甘露聚糖结合凝集素（MBL）对白血病细胞系U937凋亡的影响。方法不同浓度MBL处理U937细胞后,应用CCK-8法分析细胞增殖情况,Hoechst33258染色观察细胞核及染色质,AnnexinV/PI双染流式细胞术分析细胞凋亡率,real-timePCR和免疫印迹分析凋亡相关基因及蛋白表达。结果 30～50μg/mlMBL培养72h,U937细胞增殖明显受抑,出现不同程度核固缩、核碎裂;随MBL浓度升高或作用时间延长,凋亡细胞数逐渐增多;Fas、Caspase-3mRNA、Fas和FasL蛋白表达量升高,Caspase-3和多腺苷二磷酸多聚酶（PARP）蛋白被剪切激活或失活。结论 MBL可诱导白血病细胞U937细胞凋亡,其机制与上调Fas表达、剪切Caspase-3和PAPR有关。     

Catheter-related bacteraemia and infective endocarditis caused by Kocuria species

We describe five patients with positive blood culture for Kocuria species. Three patients had catheter-related bacteraemia and one had infective endocarditis caused by Kocuria kristinae, and one had a K. marina isolate, which was considered to be a contaminant. Identification of the isolates was further confirmed by 16S rRNA gene sequence analysis. In conclusion, Kocuria species are an unusual cause of infection in immunocompromised patients. Accurate identification with molecular methods is imperative for the diagnosis of these unusual pathogens.     

Mannan-binding lectin insufficiency in children with recurrent infections of the respiratory system

Blood samples were collected over a 4-year period from 335 children (aged 1-16 years) suffering from recurrent respiratory infections and 78 controls. The patients were subdivided into four groups: I, children with no immune system defects detected (n = 101); II, children with allergies (n = 94); III, children with humoral response defects (n = 93); and IV, children with disturbances of cellular immunity (n = 66). Nineteen patients had both humoral and cellular abnormalities. All patients and controls were investigated to determine the exon 1 and promoter region variants of the mbl-2 gene. MBL serum concentrations were also determined in samples from 291 patients and 75 controls. The proportion of O (B, D or C) alleles was significantly higher in the patient group compared to controls, and this association was strongest for subgroup III. The promoter LX variant frequency was also commoner in the patients as a whole, and significantly so in subgroups II and IV. Genotypes markedly influenced MBL concentrations in all groups, and correlated with ability to activate the lectin pathway of complement activation. The strongest and most significant inverse correlations between serum MBL and respiratory disease were found in patient group III and in 17 patients with multiple humoral and/or cellular abnormalities. Among nine patients with unexpectedly low LP activity in view of their MBL concentrations, one person was found to be MASP-2 deficient. Our results indicate that mannan-binding lectin insufficiency, with or without a coexisting immune defect, is associated with the occurrence of recurrent respiratory infections in childhood, and this relationship is particularly strong and statistically significant in children with concomitant impairments of humoral immunity.     

Case clustering in infective endocarditis: the role of availability bias

Limited data exist regarding the impact of variations in clinical practice and physicians' cognitive bias on the diagnosis of infective endocarditis (IE). As an illustration of these effects, unexpected clustering of IE diagnosis was encountered in a prospectively studied cohort. Transoesophageal echocardiography examinations for suspected IE were performed more frequently following a diagnosis of IE, and were associated with a subsequent cluster of IE cases. The cognitive bias of physicians resulting from a recent case of IE can lead to a transient increase in diagnosing additional cases of IE.     

Healthcare-associated infective endocarditis: an undesirable effect of healthcare universalization

Invasive medical technology has led to an increase in the incidence of healthcare-associated infective endocarditis (HAIE). A prospective multicentre cohort study was conducted at seven hospitals in Andalusia, Spain, to establish the characteristics of HAIE and to compare them with those of community-acquired infective endocarditis (CAIE). HAIE was defined as either infective endocarditis (IE) manifesting >48 h after admission to hospital, or IE associated with a significant invasive procedure performed in the 6 months before diagnosis. Seven hundred and ninety-three cases of IE were investigated, and HAIE accounted for 127 (16%). As compared with patients with CAIE, patients with HAIE were older (60.1 ± 14.4 years vs. 53.6 ± 17.5 years) and had more comorbidities (Charlson index 3.3 ± 2.3 vs. 1.8 ± 2.3) and staphylococcal infections (58.3% vs. 24.8%). Vascular manipulation was the main cause of bacteraemia responsible for HAIE (63%). Peripheral vein catheter-associated bacteraemia accounted for 32.8% of the catheter-related bacteraemias. In-hospital mortality (44.9% vs. 24.2%) was higher in the HAIE group. Septic shock (OR 2.2, 95% CI 2.9–30.2) and surgery not performed because of high surgical risk (OR 1.6, 95% CI 1.2–20) were independent predictors of mortality in HAIE. The present study demonstrates that HAIE is a growing health problem associated with high mortality. Careful management of vascular devices is essential to minimize the risk of bacteraemias leading to HAIE.     

Inflammatory biomarkers in infective endocarditis: machine learning to predict mortality

Infective endocarditis (IE) is the cardiac disease with the highest rates of mortality. New biomarkers that are able to identify patients at risk for death are required to improve patient management and outcome. This study aims to investigate if cytokines, chemokines and growth factors measured at IE diagnosis can predict mortality. Patients with definite IE, according to the Duke’s modified criteria, were included. Using high-performance Luminex assay, 27 different cytokines, chemokines and growth factors were analyzed. Machine learning techniques were used for the prediction of death and subsequently creating a decision tree, in which the cytokines, chemokines and growth factors were analyzed together with C-reactive protein (CRP). Sixty-nine patients were included, 41 (59%) male, median age 54 [interquartile range (IQR) = 41–65 years] and median time between onset of the symptoms and diagnosis was 12 days (IQR = 5–30 days). The in-hospital mortality was 26% (n = 18). Proinflammatory cytokines interkeukin (IL)-15 and C-C motif chemokine ligand (CCL4) were found to predict death, adding value to CRP levels. The decision tree predicted correctly the outcome of 91% of the patients at hospital admission. The high-risk group, defined as CRP ≥ 72 mg/dL, IL-15 ≥ 5·6 fg/ml and CCL4 ≥ 6·35 fg/ml had an 88% in-hospital mortality rate, whereas the patients classified as low-risk had a mortality rate of 8% (P = < 0·001). Cytokines IL-15 and CCL4 were predictors of mortality in IE, adding prognostic value beyond that provided by CRP levels. Assessment of cytokines has potential value for clinical risk stratification and monitoring in IE patients.     

Impact of haplotypes of TNF in the natural course of infective endocarditis

Based on previous findings for the role of single nucleotide polymorphisms (SNPs) of TNF for the predisposition for bloodstream infections, this study investigates the role of these SNPs at the promoter positions –376, –308, –238 in infective endocarditis (IE). In a case–control study, 83 patients with IE and 83 controls were enrolled. Blood genotyping for the presence of G or A alleles of the three SNPs was carried out using restriction fragment length polymorphisms. Haplotypes were calculated. Patients were mostly infected by Staphylococcus aureus (32.5%) and by species of enterococci (14.3%) and streptococci (14.3%). Carriage of the minor frequency A alleles at –238 of the promoter region of TNF was greater than in controls (8.4% versus 1.2%, p 0.003). The presence of any of the three GGA/GAA/AGA haplotypes was more frequent in patients with IE (OR 8.22, 95CI% 1.8–37.4, p 0.001). After multivariate logistic regression analysis, it was found that the only factor related to fatal outcome was carriage of the wild-type GGG haplotype (OR, 3.29, 95CI%, 1.05–10.29, p 0.04). GGA, AGA and GAA haplotypes were more frequent in patients with IE than in controls, suggesting a predisposition for IE and a potential protective role against fatal outcome, as the wild-type GGG haplotype was independently related with death.     

MBL对树突状细胞体外分化成熟的影响

目的: 探讨甘露聚糖结合凝集素 (MBL)对人外周血单核细胞来源的树突状细胞 (MoDC)分化成熟的影响。方法: 以天然人MBL刺激MoDC, 在倒置显微镜下观察DC的形态; 用FACS分析DC的表型; 用 3H- TdR掺入法测定DC刺激同种异体T细胞增殖的能力; 以酵母多糖颗粒吞噬试验评估DC的抗原摄取能力; 用ELISA检测DC培养上清中IL- 12和TNF- α的含量。结果: MBL刺激的DC表面分子CD1a、CD83、CD40、CD80、CD86和MHC DR的表达均上调,摄取酵母多糖颗粒的能力降低, 激发初始T细胞增殖的能力加强, 分泌的IL- 12增多但几乎不分泌TNF- α。结论: MBL能诱导DC分化成熟, 提示其可能通过调节DC的功能而参与获得性免疫应答。     

Mechanism of complement-dependent haemolysis via the lectin pathway: role of the complement regulatory proteins.

Mannan-binding lectin (MBL) is an acute phase protein which activates the classical complement pathway at the level of C4 and C2 via two novel serine proteases homologous to C1r and C1s. We recently reported that haemolysis via this lectin pathway requires alternative pathway amplification. The present experiments sought to establish the basis for this requirement, and hence focused on the activity and regulation of the C3 convertases. Complement activation was normalized between the lectin and classical pathways such that identical amounts of bound C4 and of haemolytically active C4,2 sites were present on the indicator cells. Under these conditions, there was markedly less haemolysis, associated with markedly less C3 and C5 deposited, via the lectin pathway than via the classical pathway, particularly when alternative pathway recruitment was blocked by depletion of factor D. Lectin pathway activation was associated with enhanced binding in the presence of MBL of complement control proteins C4bp and factor H to C4b and C3b, respectively, with decreased stability of the C3-converting enzyme C4b,2a attributable to C4bp. Immunodepletion of C4bp and/or factor H increased lectin pathway haemolysis and allowed lysis to occur in absence of the alternative pathway. Thus, the lectin pathway of humans is particularly susceptible to the regulatory effects of C4bp and factor H, due at least in part to MBL enhancement of C4bp binding to C4b and factor H binding to C3b.     

Chronic antiplatelet therapy and mortality among patients with infective endocarditis

Whether antiplatelet therapy is associated with better outcomes among patients with infective endocarditis (IE) remains controversial. A retrospective study was conducted concerning all patients with IE, treated in a tertiary-care centre of Canada between 1991 and 2006, who satisfied the modified Duke criteria for a definite or possible IE. The primary outcome was all-cause mortality within 90 days of diagnosis. A secondary outcome was the development of major systemic embolism. In total, 241 patients satisfied the inclusion criteria, 75 of whom had been on chronic antiplatelet therapy prior to developing endocarditis. Seventy-one (29.5%) patients died. According to multivariate analysis, age, a high Charlson score, aortic valve involvement, myocardial infarction and presence of a perivalvular abscess were strongly associated with mortality. Undergoing valvular replacement (adjusted OR (AOR) 0.28, 95% CI  0.09–0.84) and chronic antiplatelet therapy before IE (AOR  0.27, 95% CI  0.11–0.64) correlated with lower mortality. There was a trend for lower mortality among patients started on antiplatelet drugs after admission (AOR  0.29, 95% CI  0.08–1.13). The effect of aspirin on mortality was much the same in patients who received 325 or 80 mg daily. Chronic antiplatelet therapy was not associated with a significantly lower risk of major embolism. In conclusion, chronic antiplatelet therapy was associated with lower mortality among patients with IE, independently of any effect on major embolism. Whether or not a beneficial effect could be replicated by initiating antiplatelet therapy at the time of diagnosis remains unproven.     

Mannan-binding lectin may facilitate the clearance of circulating immune complexes--implications from a study on C2-deficient individuals

Deficiency of both mannan-binding lectin (MBL) and complement components C4 and C2 has been associated with increased risk of systemic lupus erythematosus (SLE). MBL can activate the complement system either through C4 and C2 or directly through C3. Circulating immune complexes (CICs) are believed to play a pathogenic role in SLE and MBL has been shown to bind certain forms of immunoglobulins, including IgM, IgG and IgA. Thus, MBL might promote CIC clearance. In order to evaluate this, six individuals with non-functional classical pathway due to the rare homozygous C2 deficiency were chosen, as the classical pathway is known to have a fundamental role in CIC clearance. Four of the six C2-deficient individuals had SLE, two of whom also had MBL deficiency. MBL serum levels and genotypes were compared with the serum levels of CICs, as measured by their content of kappa, lambda, IgM, IgA, IgG and C3 opsonization. The C2-deficient individuals had higher serum levels of CICs than 16 healthy controls (P < 0.0001). Furthermore, an inverse association was observed between MBL and CIC levels in the C2-deficient individuals, which was strongest for IgM-CICs (r = - 0.84, P = 0.037). Moreover, C3 opsonization of the CICs correlated positively with MBL levels in the C2-deficient individuals (r = 0.89, P = 0.017). In conclusion, individuals with C2 deficiency have increased levels of CICs and MBL may facilitate their clearance. Defective CIC clearance might partly explain the increased risk of SLE associated with low MBL.     

Mannan-binding lectin MBL2 gene polymorphism in chronic hepatitis C: association with the severity of liver fibrosis and response to interferon therapy

Hepatitis C virus (HCV) is a major cause of hepatic disease and of liver transplantation worldwide. Mannan-binding lectin (MBL), encoded by the MBL2 gene, can have an important role as an opsonin and complement activating molecule in HCV persistence and liver injury. We assessed the MBL2 polymorphism in 102 Euro-Brazilian patients with moderate and severe chronic hepatitis C, paired for gender and age with 102 HCV seronegative healthy individuals. Six common single nucleotide polymorphisms in the MBL2 gene, three in the promoter (H/L, X/Y and P/Q) and three in exon 1 (A, the wild-type, and B, C or D also known as O) were evaluated using real-time polymerase chain reaction with fluorescent hybridization probes. The concentration of MBL in plasma was measured by enzyme-linked immunosorbent assay. The frequency of the YA/YO genotype was significantly higher in the HCV patients compared with the controls (P = 0.022). On the other hand, the genotypes associated with low levels of MBL (XA/XA, XA/YO and YO/YO) were decreased significantly in the patients with severe fibrosis (stage F4), when compared with the patients with moderate fibrosis (stage F2) (P = 0.04) and to the control group (P = 0.011). Furthermore, MBL2 genotypes containing X or O mutations were found to be associated with non-responsiveness to pginterferon and ribavirin treatment (P = 0.023). MBL2 polymorphisms may therefore be associated not only with the development of chronic hepatitis C, but also with its clinical evolution and response to treatment.     

感染性心内膜炎合并急性肾损伤的临床特点和手术预后分析

目的分析感染性心内膜炎(IE)合并急性肾损伤(AKI)的诊治和预后情况,评判手术治疗对预后的影响。方法回顾性分析北京协和医院2010年1月至2016年5月45例IE合并AKI患者的临床资料,其中包括8例肾脏病理组织学改变;将患者分为手术(22例)和非手术组(23例),比较两组的临床资料和预后。结果本组资料男女比2.46∶1,发病年龄(48.3±16.6)岁。35.6%有基础瓣膜疾病,先天性瓣膜疾病最常见。好发感染瓣膜依次为二尖瓣(46.7%)、主动脉瓣(28.9%)和人工瓣膜(8.9%)。常见的感染病原为链球菌(46.7%)和葡萄球菌(35.6%),感染病原中也可见部分少见及特殊类型。肾脏表现中,尿潜血和尿蛋白的比例分别为82.2%和64.4%;8例肾活检病理中,新月体肾炎3例,局灶增生性肾炎和系膜增生性肾炎各2例,急性间质性肾炎1例;免疫荧光最常见的形式是C3沉积。手术和非手术患者基线资料、手术存活率无显著差别,但手术组肾功能恢复显著优于非手术组(P0.05),血清肌酐在术后7 d和30 d即较术前显著下降(P0.05和P0.01)。结论 IE的基础疾病和感染病原学较传统记载存在变化,IE导致肾实质损害时,新月体肾炎并非少见;适时手术有利于改善IE合并AKI患者的肾脏预后。     

Low mannose-binding lectin complement activation function is associated with predisposition to Legionnaires' disease

Innate immune system deficiency may predispose to severe infections such as Legionnaires' disease. We have investigated the role of mannose-binding lectin (MBL) deficiency in the Melbourne Aquarium Legionnaires' disease outbreak. Serum samples from patients and controls that were exposed but shown to be uninfected from the Melbourne Aquarium Legionnaires' disease outbreak were tested for MBL function (C4 deposition) and level (mannan-binding). MBL function was lower in Legionnaires' disease cases than in age- and sex-matched uninfected, exposed controls. The frequency of MBL deficiency with C4 deposition < 0.2 U/microl was significantly higher in Legionnaires' disease cases than in controls. This also applied to Legionnaires' disease cases requiring hospital care. There was no difference in MBL mannan-binding levels between Legionnaires' disease patients and controls. There was no significant interval change in MBL function or level after a mean of 46 days. MBL complement activation functional deficiency appears to predispose to Legionnaires' disease.     

Mannan-binding lectin (MBL)-associated serine protease-1 (MASP-1), a serine protease associated with humoral pattern-recognition molecules: normal and acute-phase levels in serum and stoichiometry of lectin pathway components

The pattern-recognition molecules mannan-binding lectin (MBL) and the three ficolins circulate in blood in complexes with MBL-associated serine proteases (MASPs). When MBL or ficolin recognizes a microorganism, activation of the MASPs occurs leading to activation of the complement system, an important component of the innate immune system. Three proteins are produced from the MASP1 gene: MASP-1 and MASP-3 and MAp44. We present an assay specific for MASP-1, which is based on inhibition of the binding of anti-MASP-1-specific antibody to MASP-1 domains coated onto microtitre wells. MASP-1 was found in serum in large complexes eluting in a position corresponding to ～600 kDa after gel permeation chromatography in calcium-containing buffer and as monomers of ～75 kDa in dissociating buffer. The concentration of MASP-1 in donor sera (n = 105) was distributed log-normally with a median value of 11 μg/ml (range 4-30 μg/ml). Serum and citrate plasma levels were similar, while the values in ethylenediamine tetraacetic acid plasma were slightly lower and in heparin plasma were 1·5 times higher than in serum. MASP-1 was present at adult level at 1 year of age, while it was 60% at birth. In normal healthy individuals the level of MASP-1 was stable throughout a 2-month period. After induction of an acute-phase reaction by operation we found an initial short decrease, concomitant with an increase in C-reactive protein levels, followed by an increase, doubling the MASP-1 concentration after 2 days. The present data prepare the ground for studies on the associations of MASP-1 levels with disease.     

Mesangio-capillary glomerulonephritis associated with Q-fever endocarditis

A patient with nephrotic syndrome and Q-fever endocarditis (confirmed serologically and ultrastructurally) was found to have mesangio-capillary glomerulonephritis with parietal deposits of C3 and IgM and some IgM in the mesangium. Elution studies showed that IgM antibodies reactive against insoluble Coxiella antigens were present in the kidney. Review of the literature suggests that this type of immune complex nephritis may be associated with Q-fever. Possible reasons for the variability of the nephritis associated with infective endocarditis are discussed.