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Udgivelsesdato: 2009-Jun 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 x 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.

Mannan-binding lectin-associated serine protease 2 (MASP-2) is an enzyme of the innate immune system. MASP-2 forms complexes with the pattern recognition molecules mannan-binding lectin (MBL), H-ficolin, L-ficolin, or M-ficolin, and is activated when one of these proteins recognizes microorganisms and subsequently cleaves complement factors C4 and C2, thus initiating the activation of the complement system. Missense polymorphisms of MASP-2 exist in different ethnic populations. To further characterize the nature of these, we have produced and characterized rMASP-2s representing the following naturally occurring polymorphisms: R99Q, D120G, P126L, H155R, 156_159dupCHNH (CHNHdup), V377A, and R439H. Only very low levels of CHNHdup were secreted from the cells, whereas quantities similar to wild-type MASP-2 were found intracellularly, indicating that this mutation results in a misfolded protein. We found that D120G and CHNHdup could not associate with MBL, whereas R99Q, P126L, H155R, V377A, R439H, and wild-type MASP-2 bound equally well to MBL. Accordingly, when D120G and CHNHdup were mixed with MBL, no activation of complement factor C4 was observed, whereas R99Q, P126L, and V377A cleaved C4 with an activity comparable to wild-type MASP-2 and H155R slightly better. In contrast, the R439H variant was deficient in this process despite its normal binding to MBL. This variant was also not able to autoactivate in the presence of MBL and mannan. We find the R439H variant is common in Sub-Saharan Africans with a gene frequency of 10%. Our results indicate that individuals with different types of MASP-2 defects may be identified through genotyping.

Candida albicans is the major cause of systemic fungal infections in immunocompromised patients. We investigated the susceptibility of mice deficient in complement factor B and C2 (Bf/C2-/-), C1q (C1qa-/-), and mannan-binding lectin (MBL)-A (MBL-A) and MBL-C (MBL-A/C-/-) to systemic infection with C. albicans. Animals were infected i.p. with 10(8)C. albicans blastoconidia and monitored for mortality. Bf/C2-/- mice showed high mortality (over 90%) within the study period of 3 weeks. In contrast, mortality in C1qa-/- mice was below 15% whereas that of MBL-A/C-/- mice was 40% (P0.001). Intravenous infection of mice with 8x10(5) blastoconidia resulted in the same trend with Bf/C2-/- mice being highly susceptible compared to the other strains. Histology of kidney sections of infected Bf/C2-/- mice showed widespread mycelia confirming the high CFU counts from cultured tissue homogenates. In C1qa-/-, MBL-A/C-/- and wild type C57BL/6 mice hyphal growth was limited. However, massive inflammatory infiltration was apparent, which was not seen in Bf/C2-/- mice. The ability of the mouse sera to opsonize C. albicans was determined by quantification of phagocytosis of C. albicans by peritoneal phagocytes. Whilst phagocytosis mediated by Bf/C2-/- mouse serum was low (10.6%), more phagocytosis could be seen in MBL-A/C-/- (19.9%), C1qa-/- mice (23.9%) and wild type mice (29%). Deficiency of classical pathway activation has only a low impact whereas the lectin pathway contributes to the host defence against candidosis. The more pronounced lack of complement activation in Bf/C2-/- mice leads to uncontrolled infection due to an opsonophagocytic defect.

This study retrospectively investigated the association between pre-transplant levels of mannose-binding lectin (MBL) plus the associated serine protease (MASP)-2 and the occurrence of cytomegalovirus (CMV) infection and symptomatic CMV disease during the first 12 weeks after kidney transplantation. Materials and methods. Altogether 159 consecutive single kidney transplant recipients were included. The patients were screened for CMV pp65 antigenaemia every second week. No CMV prophylaxis or pre-emptive treatment was given. MBL and MASP-2 were measured in samples taken at transplantation and 10 weeks later.CMV infection, defined as at least one positive test, was found in 95 patients (59.8%). MBL and MASP-2 measured at transplantation were similar in patients with and without CMV infection. The incidence of CMV infection was also similar in 36 patients (58.3%) with pre-transplant MBL levels below the reference level (500 microg/L) and in patients with higher MBL levels (60.2%). Symptomatic CMV disease was diagnosed in 35 patients (22%), and MASP-2 levels at transplantation in the lower quartile range (or=148 microg/L) was significantly associated with CMV disease during the first 12 weeks, P = 0.028. MBL levels decreased significantly from transplantation to 10 weeks later, and median (interquartile range) fell from 2597 (526-4939) microug/L to 1520 (270-3069) microg/L (P0.001). In contrast, MASP-2 levels increased significantly from 252 (148-382) microg/L to 380 (302-492) microg/L (P0.001).Pre-transplant MBL levels do not influence the incidence of any CMV infection or symptomatic CMV disease during the first 12 weeks after kidney transplantation. However, low MASP-2 levels may play a role in the development of symptomatic CMV disease.

peer-reviewed The amount and complexity of software in automotive systems is constantly increasing. Today’s luxury cars include numerous electronic control units. A large part of the functionality of these units is driven by software. In the future even more software-intensive automotive systems are expected as automotive manufacturers and suppliers tend to integrate and combine applications on more powerful platforms. The increasing amount and complexity of software in these platforms has led to the situation where software engineering has become an essential discipline within automotive systems development. This paper identifies essential areas of software engineering that will have a significant impact on future automotive systems and systems development. Particularly, it discusses how the software engineering discipline can be developed in the context of overall automotive systems development while considering essential requirements and constraints that are or will become prevalent in this domain.

Product line approaches are well-known in many manufacturing industries, such as consumer electronics, medical systems and automotive [1]. In recent years, approaches with a similar background have rapidly emerged within Software Engineering, so called Software Product Line (SPL) approaches [2], [3]. As automotive manufacturers and suppliers design and implement complex applications, such as driver assistance [4], they strive for mechanisms that allow them to implement such functionality on integrated platforms. This offers the opportunity to build a variety of similar systems with a minimum of technical diversity and thus allows for strategic reuse of components. This has resulted in a growing interest in SPL approaches both in the software engineering and the automotive systems domain. This paper discusses the increasing importance that SPL approaches could play within the context of Automotive Systems Engineering. To accomplish this, we first provide an overview of the major challenges faced by Automotive Systems Engineering [5]. We then present a selection of SPL approaches, which could provide solutions for the described challenges. To complement this we make the case for empirical evaluation as a basis for well-founded decisions and selection of techniques. Finally, we present and in-depth discussion of how the approaches and techniques outlined can be used to address the identified challenges. The paper concludes with an overview of open research questions and expected benefits for the development of automotive systems.

Although human C-reactive protein (CRP) becomes upregulated during septicemia, its role remains unclear, since purified CRP showed no binding to many common pathogens. Contrary to previous findings, we show that purified human CRP (hCRP) binds to Salmonella enterica, and that binding is enhanced in the presence of plasma factors. In the horseshoe crab, Carcinoscorpius rotundicauda, CRP is a major hemolymph protein. Incubation of hemolymph with a range of bacteria resulted in CRP binding to all the bacteria tested. Lipopolysaccharide-affinity chromatography of the hemolymph co-purified CRP, galactose-binding protein (GBP) and carcinolectin-5 (CL5). Yeast two-hybrid and pull-down assays suggested that these pattern recognition receptors (PRRs) form pathogen recognition complexes. We show the conservation of PRR crosstalk in humans, whereby hCRP interacts with ficolin (CL5 homologue). This interaction stabilizes CRP binding to bacteria and activates the lectin-mediated complement pathway. We propose that CRP does not act alone but collaborates with other plasma PRRs to form stable pathogen recognition complexes when targeting a wide range of bacteria for destruction.

Burn injury destroys skin, the second largest innate immune organ in the body, and triggers chaotic immune and inflammatory responses. The pattern recognition molecule, mannan-binding lectin (MBL), plays an important role in the first-line host defense against infectious agents. MBL initiates the lectin complement pathway and acts as an opsonin. Recent studies suggest that MBL also modulates inflammatory responses. We report that local responses after burn in MBL null mice differ from those found in wild-type (WT) mice in the following important biological markers: spontaneous eschar separation, thinned epidermis and dermis, upregulation of soluble factors including cytokines, chemokines, cell adhesion molecules, a growth factor-binding protein, and matrix metalloproteinases. Mice lacking C1q, C4, or C3 did not show the lack of eschar separation seen in MBL null-burn phenotype. These findings implicate MBL as an important molecule in the maintenance of the homeostatic balance.

Aim The lectin complement pathway, initiated by mannan-binding-lectin (MBL) plays a role in tissue destruction following ischemia/reperfusion, and MBL deficiency has been associated with favorable outcome in stroke patients. MAp44 is produced in the heart and may theoretically function as an endogenous inhibitor of MBL-mediated activities. The aim of this study was to investigate the possible association between MBL deficiency, MAp44 levels and outcome in comatose survivors of out-of-hospital cardiac arrest (OHCA). Methods In a single center post hoc analysis of the prospective multicenter randomized Target Temperature Management (TTM) trial, we measured MBL and MAp44 levels at baseline, 24, 48 and 72 h after OHCA in 169 consecutive patients randomly assigned to TTM at 33 °C or 36 °C for 24 h. Primary outcome was 30 days mortality and secondary outcome was favorable neurological outcome assessed by Cerebral Performance Category (CPC1-2) and modified Rankin Scale (mRS0-3) 180 days after OHCA. Results Patients with MBL deficiency (defined as plasma levels ≤100 ng ml−1 at baseline) (n = 22) carried a 30-day mortality of 41% compared to 32% in MBL sufficient patient (n = 147), p = 0.55. Baseline MAp44 levels were not associated with mortality, p = 0.25. There was no significant difference in neurological outcome between the two MBL groups assessed by CPC (p = 0.69) and mRS (p = 0.91). In multivariable models, baseline MBL (OR = 1.0, p = 0.70), (OR = 1.5, p = 0.30) and MAp44 levels (OR = 1.0, p = 0.99), (OR = 1.6, p = 0.21) were not associated with favorable neurological outcome assessed by CPC and mRS, respectively. Conclusions In comatose survivors after cardiac arrest, neither MBL deficiency nor levels of MBL and MAp44 were associated with mortality or neurological outcome.

The complement system is an essential element of the innate immune response that becomes activated upon recognition of molecular patterns associated with microorganisms, abnormal host cells, and modified molecules in the extracellular environment. The resulting proteolytic cascade tags the complement activator for elimination and elicits a pro-inflammatory response leading to recruitment and activation of immune cells from both the innate and adaptive branches of the immune system. Through these activities, complement functions in the first line of defense against pathogens but also contributes significantly to the maintenance of homeostasis and prevention of autoimmunity. Activation of complement and the subsequent biological responses occur primarily in the extracellular environment. However, recent studies have demonstrated autocrine signaling by complement activation in intracellular vesicles, while the presence of a cytoplasmic receptor serves to detect complement-opsonized intracellular pathogens. Furthermore, breakthroughs in both functional and structural studies now make it possible to describe many of the intricate molecular mechanisms underlying complement activation and the subsequent downstream events, as well as its cross talk with, for example, signaling pathways, the coagulation system, and adaptive immunity. We present an integrated and updated view of complement based on structural and functional data and describe the new roles attributed to complement. Finally, we discuss how the structural and mechanistic understanding of the complement system rationalizes the genetic defects conferring uncontrolled activation or other undesirable effects of complement.

OBJECTIVE Mannan-binding lectin (MBL) is a complement-activating carbohydrate-recognizing molecule associated with diabetic nephropathy. MBL is associated with all-cause mortality in type 2 diabetes, but whether MBL is associated with mortality in type 1 diabetes remains unknown. We therefore aimed to investigate this. RESEARCH DESIGN AND METHODS We studied an existing 12-year prospective cohort with type 1 diabetes with 198 patients with diabetic nephropathy (121 men, age 41 years [95% CI 40–42], estimated glomerular filtration rate [eGFR] 67 mL/min/1.73 m2 [95% CI 63–70]) and 174 normoalbuminuric patients (103 men, age 43 years [95% CI 41–44], eGFR 93 mL/min/1.73 m2 [95% CI 91–95]). Mortality rates were compared according to the concentration-determining MBL2 genotype or the MBL concentration. Patients were classified as having high or low MBL expression genotypes. The effect of MBL concentration was estimated by comparing patients with MBL concentrations above or below the median. RESULTS Ninety-eight patients died during follow-up. The unadjusted hazard ratio (HR) for all-cause mortality was 1.61 (95% CI 1.07–2.43) for patients with high MBL expression genotypes versus patients with low MBL expression genotypes (P = 0.023). All-cause mortality was higher in patients with MBL concentrations above the median than in patients with MBL concentrations below the median (unadjusted HR 1.90 [95% CI 1.26–2.87], P = 0.002). CONCLUSIONS High MBL expression genotypes and high MBL concentrations are both associated with increased mortality rates in type 1 diabetes compared with low MBL expression genotypes and low MBL concentrations.

Objective.The complement system plays a crucial role in the pathogenesis of inflammatory processes. The lectin pathway of the complement system is activated through the recognition of pathogens by soluble pattern recognition molecules (PRM), i.e., mannan-binding lectin (MBL), collectin-LK, and the ficolins. PRM are reportedly correlated to disease activity in rheumatoid arthritis (RA). The aim was to evaluate the pathogenic role of PRM in juvenile idiopathic arthritis (JIA).Methods.We measured MBL, M-ficolin, H-ficolin, MBL-associated serine proteases (MASP) 1, MASP-2, MASP-3, and 2 alternative splice products, MBL-associated protein (MAp) 44 and MAp19, in plasma and synovial fluid (SF) of children with persistent oligoarticular (n = 109 in plasma, n = 38 in SF) and systemic JIA (n = 19 in plasma, n = 11 in SF). The concentrations of the proteins were measured by in-house time-resolved immunofluorometric assays.Results.We observed significantly higher levels of M-ficolin, MASP-1, MASP-2, and MASP-3 in plasma and SF from patients with systemic JIA compared with persistent oligoarticular JIA (p < 0.001). In paired samples of plasma and SF from 47 patients with oligoarticular and systemic JIA, we observed higher concentrations in plasma for both subtypes for 7 of the measured proteins while the reverse relationship was observed for MASP-3. M-ficolin and MASP-2 correlated to erythrocyte sedimentation rate, C-reactive protein, white blood cell count, and platelet count (p < 0.001). M-ficolin was in addition related to the number of active joints and inversely related to hemoglobin levels.Conclusion.Our results point to plasma M-ficolin and MASP-2 as inflammatory markers in JIA. The levels of all proteins are higher in plasma than in SF, except for MASP-3, indicating that MASP-3 may be produced locally in joints.

Summary There is increasing evidence that the complement system plays an important role in diabetes and the development of diabetic vascular complications. In particular, mannan-binding lectin (MBL) levels are elevated in diabetes patients, and diabetes patients with diabetic nephropathy have higher MBL levels than diabetes patients with normal renal function. The MBL-associated serine proteases (MASPs) MASP-1, MASP-2 and MASP-3 and MBL-associated protein MAp44 have not yet been studied in diabetes patients. We therefore measured plasma levels of MASP-1, MASP-2, MASP-3 and MAp44 in 30 children with type 1 diabetes mellitus (T1DM) and 17 matched control subjects, and in 45 adults with T1DM and 31 matched control subjects. MASP-1 and MASP-2 levels were significantly higher in children and adults with T1DM than in their respective control groups, whereas MASP-3 and MAp44 levels did not differ between patients and controls. MASP-1 and MASP-2 levels correlated with HbA1c, and MASP levels decreased when glycaemic control improved. Because MASP-1 and MASP-2 have been shown to interact directly with blood coagulation, elevated levels of these proteins may play a role in the enhanced thrombotic environment and consequent vascular complications in diabetes.

BACKGROUND: Recent studies have shown that activation of the complement system may be associated with long-term graft function. The aim of this retrospective study was to assess the impact of the pattern recognition molecules of the lectin pathway on long-term graft survival after kidney transplantation.METHODS: Patients transplanted in 2000 to 2001 were included. Mannose-binding lectin, Ficolin-1, and Ficolin-3 were measured in serum at the time of transplantation. Data on death-censored graft loss were obtained from the Norwegian Renal Registry. Competing risks regression was used to investigate the association between time to graft loss and the explanatory variables. The variables were: high Ficolin-3 (upper quartile, ≥33.3 μg/mL) versus low Ficolin-3 (RESULTS: A total of 382 patients with a median follow-up of 9.8 years were included. Sixty-six patients (17%) had death-censored graft loss, and 116 (30%) patients died. In a final competing risks model, high Ficolin-3 (subhazard ratio [SHR] = 1.95, P = 0.009), acute rejection (one vs. none) (SHR = 1.93, P = 0.033), acute rejection (two vs. none) (SHR = 5.45, P < 0.001), and age (SHR = 0.98, P = 0.006) were associated with death-censored graft loss. Basiliximab induction was associated with improved graft survival (SHR = 0.50, P = 0.016). No associations between mannose-binding lectin or Ficolin-1 and graft loss were found.CONCLUSION: High Ficolin-3 level at the time of transplantation was an independent significant risk factor for shorter graft survival, even when adjusted for other covariates.

Objective.To examine whether proteins of the lectin pathway of the complement system are involved in systemic lupus erythematosus (SLE) pathogenesis.Methods.Using time-resolved immunofluorometric assays, plasma levels of mannan-binding lectin (MBL)-associated serine proteases 1 (MASP-1), MASP-2, MASP-3, MBL-associated protein of 19 kDa (MAp19), and MAp44 were determined in 58 patients with SLE and 65 healthy controls (HC).Results.Plasma concentrations in patients with SLE were higher than HC regarding MASP-1, MASP-3, and MAp44 (p < 0.0001, 0.0003, and 0.0013). Complement factor 3 correlated negatively and anti-dsDNA positively with levels of MAp19 (p = 0.0035, p = 0.0133).Conclusion.In SLE, plasma levels of MASP and MAp are altered and associated with SLE characteristics, supporting a role in SLE pathogenesis.

Background.The innate immune system may have adverse effects in diabetes and cardiovascular disease. The complement system seems to play a key role through erroneous complement activation via hyperglycaemia-induced neoepitopes. Recently mannan-binding lectin (MBL) was shown to worsen diabetic kidney changes. We hypothesize that mouse ficolin B exerts detrimental effects in the diabetic kidney as seen for MBL.Methods.We induced diabetes with streptozotocin in female wild-type mice and ficolin B knockout mice and included two similar nondiabetic groups. Renal hypertrophy and excretion of urinary albumin and creatinine were quantified to assess diabetic kidney damage.Results.In the wild-type groups, the kidney weighed 24% more in the diabetic mice compared to the controls. The diabetes-induced increase in kidney weight was 29% in the ficolin B knockout mice, that is, equal to wild-type animals (two-way ANOVA,P=0.60). In the wild-type mice the albumin-to-creatinine ratio (ACR) was 32.5 mg/g higher in the diabetic mice compared to the controls. The difference was 62.5 mg/g in the ficolin B knockout mice, but this was not significantly different from the wild-type animals (two-way ANOVA,P=0.21).Conclusions.In conclusion, the diabetes-induced effects on kidney weight and ACR were not modified by the presence or absence of ficolin B.

We are grateful to the participating families. This study was partially supported by NIH R01DC009645 and the John T. and Winifred M. Hayward Foundation to M. T., Background: 3MC1 syndrome is a rare autosomal recessive disorder characterized by intellectual disability, short stature and distinct craniofacial, umbilical, and sacral anomalies. Five mutations in MASP1, encoding lectin complement pathway enzymes MASP-1 and MASP-3, have thus far been reported to cause 3MC1 syndrome. Only one previously reported mutation affects both MASP-1 and MASP-3, while the other mutations affect only MASP-3. Methods: We evaluated six unrelated individuals with 3MC1 syndrome and performed Sanger sequencing for all coding exons of MASP1. We also measured complement lectin and alternative pathway activities in an affected individual's serum. Results: We found two novel splice site mutations, c.1012-2A > G in one and c.891 + 1G > T in two probands, and three novel missense mutations, c.1451G > A (p.G484E), c.1657G > A (p.D553N), and c.1987G > T (p.D663Y). Missense mutations affect only MASP-3, while splice site mutations affect both MASP-1 and MASP-3. In a proband who is homozygous for c.891 + 1G > T, we detected a total lack of lectin complement pathway activity and a 2.5-fold lower alternative pathway activity. The phenotype observed in patients whose both MASP-1 and MASP-3 are affected and in those whose only MASP-3 is affected does not appear to be different. We observed structural brain abnormalities, neonatal tooth, a vascular anomaly and a solid lesion in liver as novel phenotypic features of 3MC1 syndrome. Conclusion: Novel mutations and additional phenotypic features expand the genotypic and phenotypic spectrum of 3MC1 syndrome. Although patients with MASP-1 dysfunction in addition to disrupted MASP-3 have an altered complement system, their disease phenotype is not different from those having only MASP-3 dysfunction., NIH [R01DC009645]; John T. and Winifred M. Hayward Foundation

Severe infections related to treatment are common in patients with multiple myeloma (MM). Genetic polymorphisms of the immune system may influence the risk of infections. Mannan-binding lectin (MBL) is part of the innate immune system, and individuals homozygous for wild-type MBL encoding gene (MBL2) have a well-functioning MBL pathway of complement activation, in contrast to individuals carrying one or two variant alleles. We evaluated 113 courses of high-dose melphalan and autologous stem cell transplantation (ASCT) in patients with MM. Patients homozygous for wild-type MBL2 had a significantly reduced risk of septicaemia during the ASCT procedure compared with patients carrying variant MBL2: Odds Ratio (OR) 0.19 (95% CI: 0.04-0.77), (P=0.02) in multivariate analysis. The risk of Common Toxicity Criteria grade 3-4 infections in general was not affected by wild-type MBL2: OR 1.20 (95% CI: 0.52-2.78), (P=0.67). The findings indicate that MBL to some extent protects against the most severe infections during ASCT.

Mannan-binding lectin (MBL) is a vital and versatile component of innate immunity. It is present in serum and may bind to a plethora of microbial pathogens and mediate opsonization of these by complement-dependent and/or independent mechanisms. Low-MBL levels in serum, attributed to certain genetic polymorphisms, constitute a major factor predisposing to several infectious diseases. However, recent studies propose that MBL extends beyond its classic role as a first-line host-defense molecule to a modulator of inflammation. In this review, we summarize and explore this potential and a possible novel role of MBL in asthma and allergy.

Mannan-binding lectin (MBL) is attracting considerable interest due to its role in the immune defense. The high frequency of congenital MBL deficiency makes it feasible to evaluate clinical relevance through epidemiological investigations on fairly limited numbers of patients. MBL deficiency is determined by three mutant allotypes termed B, C and D in the coding region as well as mutations in the promoter region. It has been suggested that individuals, with deficiency-associated allotypes, may present significant amounts of low molecular weight MBL. We have compared the quantification of MBL by four commercially available assays with results obtained by our own in-house assays. Most assays are selectively sensitive for the wild type MBL (allotype A), but special combinations of antibodies also detect mutant forms of MBL. Thus a sandwich-type time-resolved immunoflourometric assay (TRIFMA), with a mouse monoclonal antibody (93C) as the catching and detecting antibody, shows B/B and D/D homozygous individuals to present signals corresponding to up to 500 ng MBL per ml (with plasma from an A/A individual as standard) as compared to less than 50 ng/ml and 200 ng/ml, respectively, when measured in other assays. In GPC at isotonic conditions the MBL in B/B and D/D individuals showed a Mr of 450 kDa. This MBL cannot bind to mannan. We further present a new method for quantifying the amount of MBL polypeptide chain. By applying plasma samples on SDS-PAGE at reducing conditions followed by Western blotting and quantification by chemiluminescense, this approach presents single polypeptide chains to the antibody independent of allotype differences in the collagen-like region. Titrations of recombinant MBL served as standard. In sera from homozygous mutants (O/O) the MBL concentrations estimated on Western blot were in the range of 100 to 500 ng/ml and correlated with that measured in the 93C-based TRIFMA.

The primary objectives of this analysis were to examine the effects of early posttransplantation (10 wk) serum levels of osteoprotegerin (OPG), mannose-binding lectin (MBL), and MBL-associated serine proteases (MASP; MASP-2 and MASP-3) on long-term (8-yr) patient survival, graft survival, and cardiovascular (CV) death. During a period of 16 mo (1995 to 1996), a total of 173 consecutive renal transplant recipients without diabetes before transplantation were included in a prospective study that was designed to address the impact of metabolic CV risk factors on survival and CV end points. Baseline sera from 172 patients were available for analysis. Follow-up data until January 1, 2004, were obtained from a national renal registry. Patients with high (fourth quartile) serum levels of OPG had significantly higher all-cause mortality (hazard ratio [HR] 6.3; 95% confidence interval [CI] 3.3 to 11.8; P < 0.001) and CV death (HR 10.8; 95% CI 3.8 to 30.4; P < 0.001) than patients with lower OPG concentrations. After multiple Cox regression analysis, high serum levels of OPG remained an independent predictor of all-cause mortality (HR 6.0; 95% CI 3.1 to 11.6, P < 0.001) and CV death (HR 8.2; 95% CI 2.5 to 26.4; P < 0.001). Multiple linear regression analysis revealed that age, creatinine clearance, and high-sensitivity C-reactive protein all were independently associated with OPG (R 2 = 0.42). No significant association between OPG and death-censored graft loss was revealed. Serum levels of MBL, MASP-2, and MASP-3 were not significantly associated with patient survival, CV death, or graft loss. Early measured posttransplantation serum OPG is a highly significant independent predictor of death from any cause or CV death in white renal transplant recipients.

Summary The mannan-binding lectin (MBL) pathway of complement activation is important in host defence against pathogens and possibly against cancer. We investigated the effect of major surgery on two central components of the MBL pathway; MBL and the MBL-associated serine protease MASP-2, and for comparison also measured the interleukin (IL)-6 and C-reactive protein (CRP) levels. Serial blood samples were obtained from patients belonging to two different cohorts. Cohort 1 comprised 60 patients undergoing open or laparoscopic colectomy for benign disease (n = 12) or colon cancer (n = 48). Cohort 2 comprised 27 patients undergoing elective, open surgery for colorectal cancer, and was included in order to cover blood sampling between days 2 and 6. As expected, the surgical stress induced a marked acute phase response, as evidenced by a large increase in IL-6 (18-fold) and CRP (13-fold) levels with maximum at 12 h and 2 days, respectively. However, in both cohorts the levels of MBL and MBL-associated serine protease 2 (MASP-2) were largely unaffected, except for a minor but significant increase around day 8 in cohort 1. The preoperative levels of IL-6 and CRP were correlated significantly in both cohorts (r = 0·71, P

Events in the mannan-binding lectin (MBL) pathway of complement activation involve the binding of MBL to patterns of carbohydrate structures presented on the surface of microorganisms and the following activation of the proenzymes of the complement system. The protocols for MBL antigen assay are described in this chapter and are useful for routine quantification of MBL and evaluation of the MBL pathway activity in clinical samples. The assays are based on the robust, highly sensitive and reproducible time-resolved immunofluorometric assay (TRIFMA). TRIFMAs are similar to enzyme-linked immunosorbent assays (ELISAs), with the only difference being the type of labeling of detecting molecules. Different approaches have been used to measure the MBL concentration in plasma or serum. The first is based on a modification of the conventional sandwich ELISA, in which microtiter wells are coated with anti-MBL antibody and then incubated with dilutions of plasma or serum and the amount of bound MBL is measured by using europium-labeled anti-MBL antibody (MBL antigen assay). The second assay quantifies MBL on the basis of its lectin-binding activity (lectin assay), in which microtiter wells are coated with mannan instead of anti-MBL antibody. The MBL antigen assay is more sensitive (down to 2 ng of MBL/ml of plasma) than the lectin assay (10 ng/ml).

SummaryMannan-binding lectin (MBL), an important component of innate immunity, binds to a range of foreign antigens and initiates the lectin complement pathway. Earlier studies have reported high plasma MBL levels in allergic patients in comparison to healthy controls. In view of varied plasma MBL levels being determined by genetic polymorphisms in its collagen region, we investigated the association of single nucleotide polymorphisms (SNPs) in the collagen region of human MBL with respiratory allergic diseases. The study groups comprised patients of bronchial asthma with allergic rhinitis (n = 49) and allergic bronchopulmonary aspergillosis (APBA) (n = 11) and unrelated age-matched healthy controls of Indian origin (n = 84). A novel intronic SNP, G1011A of MBL, showed a significant association with both the patient groups in comparison to the controls (P

Mannan-binding lectin (MBL) is a plasma protein of the innate immune system with the ability to initiate antimicrobial and inflammatory actions. MBL deficiency is common. More than 10% of the general population may, depending on definition, be classified as MBL deficient, underlining the redundancy of the immune system. Ongoing research attempt to illuminate at which conditions MBL deficiency may lead to disease. With examples, this review illustrates the diversity of results obtained so far.

Diabetes mellitus is a major and increasing health problem worldwide. One of the most serious consequences of diabetes is the development of diabetic angiopathy, which includes cardiovascular disease, neuropathy, retinopathy and nephropathy. Diabetic nephropathy alone affects 15-25% of patients with type 1 diabetes and 30-40% of patients with type 2 diabetes and is the single-most important cause of end-stage renal failure in the Western World. Existing research has demonstrated the involvement of glycation factors, growth factors/cytokines, hemodynamic factors and intracellular changes in the pathogenesis of diabetic kidney disease. An emerging amount of recent data suggests that the complement system, especially the MBL pathway, plays an important role in the pathogenesis of diabetic vascular complications. Although the numerous therapeutic interventions available today may delay the development and progression of diabetes vascular complications, there is an ongoing need for new therapeutic strategies. In this article the evidence for a connection between the complement system and vascular dysfunction will be reviewed, with a special focus on the relation to diabetic kidney disease. Several ways of specifically manipulating the complement system already exist. However, whether or not these drugs provide new targets for intervention on diabetic vascular complications is still unknown.

Inflammation and complement activation via the mannose-binding lectin (MBL) pathway have been suggested to play a role in the pathogenesis of diabetic microvascular complications. The association between the complement-activating protein MBL and the development of persistent microalbuminuria was evaluated in an inception cohort of 286 newly diagnosed type 1 diabetic patients consecutively admitted to the Steno Diabetes Center between 1 September 1979 and 31 August 1984. Serum MBL was measured with an immunofluorometric assay in 270 of the patients (159 men) after 3 years of diabetes duration. During the median (range) follow-up period of 18.0 (1.0–21.8) years, 75 patients subsequently progressed to persistent micro- or macroalbuminuria (urinary albumin excretion rate >30 mg/24 h). In patients with MBL levels above the median (1,597 μg/l), the cumulative incidence of persistent micro- or macroalbuminuria was 41% (CI 31–50) as compared with 26% (CI 17–34) in patients with MBL levels below the median (log-rank test, P = 0.003). In a Cox proportional hazard model with sex and age as fixed covariates, MBL was independently associated with later development of persistent micro- or macroalbuminuria (hazard ratio 1.21 [CI 1.02–1.42] per 1,000 μg/l increase in MBL; P = 0.03) after adjusting for possible confounders. In our study, high levels of MBL early in the course of type 1 diabetes was significantly associated with later development of persistent micro- or macroalbuminuria, suggesting that complement activation initiated by MBL may be involved in the pathogenesis of diabetic microvascular complications.

Diabetic nephropathy is associated with insulin resistance, and low-grade inflammation and activation of the complement system may contribute to this cascade. Mannan-binding lectin (MBL) activates the complement system, and elevated MBL concentrations have been observed in normoalbuminuric type 1 diabetic patients. The aim of this study was to assess whether MBL is associated with diabetic nephropathy in type 1 diabetes, and whether there is an association between MBL and low-grade inflammatory markers or insulin resistance.A total of 191 type 1 diabetic patients from the Finnish Diabetic Nephropathy Study were divided into three groups based upon their AER. Patients with normal AER (n=67) did not take antihypertensive medication, while patients with microalbuminuria (n=62) or macroalbuminuria (n=62) were all treated with an ACE inhibitor. As a measure of insulin sensitivity we used estimated glucose disposal rate. MBL was measured by an immunofluorometric assay, C-reactive protein by a radioimmunoassay and IL-6 by high-sensitivity enzyme immunoassay.Patients with normal AER (median [interquartile range]: 1,154 microg/l [180-2,202 microg/l]) had lower levels of MBL than patients with microalbuminuria (1,713 microg/l [724-2,760 microg/l]; p=0.029) or macroalbuminuria (1,648 microg/l [568-3,394 microg/l]; p=0.019). There was a significant correlation between MBL and estimated glucose disposal rate, but not between MBL and C-reactive protein or IL-6 levels in univariate analysis. However, in a multiple regression analysis, HbA1c was the single variable independently associated with MBL (beta+/-SEM: 0.26+/-0.08; p=0.003).MBL concentrations are increased in type 1 diabetic patients with diabetic nephropathy. MBL was not associated with low-grade inflammatory markers.

L-ficolin and H-ficolin are molecules of the innate immune system. Upon recognition of a suitable target they activate the complement system. The ligand recognition structure of ficolins is contained within a fibrinogen-like domain. We examined the selectivity of the ficolins through inhibiting the binding to bacteria or to beads coupled with N-acetylglucosamine. The binding of L-ficolin to Streptococcus pneumoniae 11F and the beads was inhibited by N-acetylated sugars and not by non-acetylated sugars. However, it was also inhibited by other acetylated compounds. Based on this selectivity L-ficolin is not easily defined as a lectin. The binding of H-ficolin to Aerococcus viridans was not inhibited by any of the sugars or other compounds examined. Based on the selectivity of L-ficolin we developed a new purification procedure involving affinity chromatography on N-acetylcysteine-derivatized Sepharose. The column was loaded in the presence of EDTA and high salt, and L-ficolin was eluted by decreasing the salt concentration. Further purification was achieved by ion exchange chromatography.

Gram-positive organisms like Staphylococcus aureus are a major cause of morbidity and mortality worldwide. Humoral response molecules together with phagocytes play a role in host responses to S. aureus. The mannose-binding lectin (MBL, also known as mannose-binding protein) is an oligomeric serum molecule that recognizes carbohydrates decorating a broad range of infectious agents including S. aureus. Circumstantial evidence in vitro and in vivo suggests that MBL plays a key role in first line host defense. We tested this contention directly in vivo by generating mice that were devoid of all MBL activity. We found that 100% of MBL-null mice died 48 h after exposure to an intravenous inoculation of S. aureus compared with 45% mortality in wild-type mice. Furthermore, we demonstrated that neutrophils and MBL are required to limit intraperitoneal infection with S. aureus. Our study provides direct evidence that MBL plays a key role in restricting the complications associated with S. aureus infection in mice and raises the idea that the MBL gene may act as a disease susceptibility gene against staphylococci infections in humans.

The hepatic protein mannan-binding lectin (MBL) activates the complement system on binding to carbohydrate patterns and is involved in first-line defense against invading microorganisms. Emerging evidence indicates that in some situations MBL may cause inexpedient complement activation and tissue injury through binding to endothelial glycosylations. MBL levels are suppressed by insulin treatment in critically ill patients, and, hypothetically, hepatic portal hypoinsulinemia could lead to increased levels of MBL in patients with type 1 diabetes. We measured MBL and C-reactive protein (CRP) levels in 132 normoalbuminuric type 1 diabetic patients and 66 healthy age- and sex-matched controls. The median MBL concentration was higher in diabetic patients than in healthy controls [1290 micro g/liter (interquartile range, IQR 354-2961 micro g/liter) vs. 970 micro g/liter (IQR 277-1607 micro g/liter), P = 0.025], whereas CRP concentrations were similar among patients and controls [1.42 mg/liter (IQR 0.95-2.21) vs. 1.21 mg/l (IQR 0.74-2.13), NS]. In diabetic subjects, CRP levels correlated with poor glycemic control as indicated by hemoglobin A(1c) and daily insulin dose, which was not the case with MBL. MBL concentrations were positively correlated with urinary albumin excretion (r = 0.22; P = 0.013) and increased with increasing urinary albumin excretion tertile (P = 0.036). In conclusion, our data demonstrate that circulating MBL concentrations are significantly elevated in patients with type 1 diabetes and suggest a possible role of MBL in the pathogenesis of renovascular complications in diabetes.

Background Evidence links the lectin pathway of complement activation to diabetic kidney disease. Upon carbohydrate-recognition by pattern-recognition molecules, eg, mannan-binding lectin (MBL), the MBL-associated serine protease (MASP-2) is activated and initiates the complement cascade. The MASP2 gene encodes MASP-2 and the alternative splice product MBL-associated protein 19 (MAp19). Both MAp19 and MASP-2 circulate in complex with MBL. We tested the hypothesis that MAp19 and MASP-2 concentrations predict the risk of incident microalbuminuria. Methods Baseline MAp19 and MASP-2 were measured in 270 persons with newly diagnosed type 1 diabetes tracked for incidence of persistent microalbuminuria in a prospective observational 18-year-follow-up study. Results Seventy-five participants (28%) developed microalbuminuria during follow-up. MBL-associated protein 19 concentrations were higher in participants that later progressed to microalbuminuria as compared with those with persistent normoalbuminuria (268 ng/mL [95% CI, 243-293] vs 236 ng/mL [95% CI, 223-250], P = .02). Participants with MAp19 concentration within the highest quartile of the cohort had an increased risk of microalbuminuria as compared with participants with MAp19 concentration within the combined lower 3 quartiles in unadjusted Cox analysis, hazard ratio 1.86 ([95% CI, 1.17-2.96], P = .009). This remained significant in adjusted models, eg, adjusting for age, sex, HbA1c , systolic blood pressure, urinary albumin excretion, smoking, serum creatinine, and serum cholesterol. MBL-associated serine protease concentration was not associated with incidence of microalbuminuria. Conclusions In conclusion, the results show an association between baseline MAp19 concentration and the incidence of microalbuminuria in an 18-year-follow-up study on persons with newly diagnosed type 1 diabetes.

SummaryThe proteolytic cascade of the complement system is initiated when pattern-recognition molecules (PRMs) bind to ligands, resulting in the activation of associated proteases. In the lectin pathway of complement, the complex of mannan-binding lectin (MBL) and MBL-associated serine protease-1 (MASP-1) initiates the pathway by activating a second protease, MASP-2. Here we present a structural study of a PRM/MASP complex and derive the overall architecture of the 450 kDa MBL/MASP-1 complex using small-angle X-ray scattering and electron microscopy. The serine protease (SP) domains from the zymogen MASP-1 dimer protrude from the cone-like MBL tetramer and are separated by at least 20 nm. This suggests that intracomplex activation within a single MASP-1 dimer is unlikely and instead supports intercomplex activation, whereby the MASP SP domains are accessible to nearby PRM-bound MASPs. This activation mechanism differs fundamentally from the intracomplex initiation models previously proposed for both the lectin and the classical pathway.

Defining mechanisms governing translation of molecular binding events into immune activation is central to understanding immune function. In the lectin pathway of complement, the pattern recognition molecules (PRMs) mannan-binding lectin (MBL) and ficolins complexed with the MBL-associated serine proteases (MASP)-1 and MASP-2 cleave C4 and C2 to generate C3 convertase. MASP-1 was recently found to be the exclusive activator of MASP-2 under physiological conditions, yet the predominant oligomeric forms of MBL carry only a single MASP homodimer. This prompted us to investigate whether activation of MASP-2 by MASP-1 occurs through PRM-driven juxtaposition on ligand surfaces. We demonstrate that intercomplex activation occurs between discrete PRM/MASP complexes. PRM ligand binding does not directly escort the transition of MASP from zymogen to active enzyme in the PRM/MASP complex; rather, clustering of PRM/MASP complexes directly causes activation. Our results support a clustering-based mechanism of activation, fundamentally different from the conformational model suggested for the classical pathway of complement.

Previous studies using mannose-binding lectin (MBL) and complement C4–deficient mice have suggested that the lectin pathway (LP) is not required for the development of inflammatory arthritis in the collagen Ab–induced arthritis (CAIA) model. MBL, ficolins and collectin-11 are key LP pattern recognition molecules that associate with three serine proteases—MASP-1, MASP-2, and MASP-3—and with two MBL-associated proteins designated sMAP and MBL-associated protein of 44kDA (MAp44). Recent studies have shown that MAp44, an alternatively spliced product of the MASP-1/3 gene, is a competitive inhibitor of the binding of the recognition molecules to all three MASPs. In these studies, we examined the effect of treatment of mice with adenovirus (Ad) programmed to express human MAp44 (AdhMAp44) on the development of CAIA. AdhMAp44 and Ad programming GFP (AdGFP) expression were injected i.p. in C57BL/6 wild type mice prior to the induction of CAIA. AdhMAp44 significantly reduced the clinical disease activity (CDA) score by 81% compared with mice injected with AdGFP. Similarly, histopathologic injury scores for inflammation, pannus, cartilage and bone damage, as well as C3 deposition in the cartilage and synovium, were significantly reduced by AdhMAp44 pretreatment. Mice treated with AdmMAp44, programming expression of mouse MAp44, also showed significantly decreased CDA score and histopathologic injury scores. In addition, administration of AdhMAp44 significantly diminished the severity of Ross River virus–induced arthritis, an LP-dependent model. Our study provides conclusive evidence that an intact complement LP is essential to initiate CAIA, and that MAp44 may be an appropriate treatment for inflammatory arthritis.

Background: Mannan-binding lectin (MBL) and mannan-binding lectin-associated serine protease-2 (MASP-2) represent important arms of the innate immune system, and different deficiencies may result in infections or autoimmune diseases. Both bipolar and panic disorders are associated with increased inflammatory response, infections and mutual comorbidity. However, associations with MBL, MASP-2 or the gene, MBL2, coding for MBL, have not been investigated thoroughly. Methods: 100 patients with bipolar disorder, 100 with panic disorder and 349 controls were included. Serum concentrations of MBL and MASP-2 were measured and seven single nucleotide polymorphisms (SNPs) influencing these concentrations were genotyped. Disease association with genetic markers and serum levels were investigated. Results: In panic disorder, we observed a large proportion (30%) of MBL deficient (MBL2 YA two-marker haplotype. Bipolar disorder was associated with the MBL2 LXPA haplotype and lower MASP-2 levels. Limitations: No information on course or severity of disorders was included, and only MBL and MASP-2 were measured, excluding other components from the complement pathway. Restrictions defined by ethnical committees preclude information of control’s ethnic origin. Conclusions: Significant differences in MBL and MASP-2 concentrations were observed between cohorts, especially an intriguing finding associating panic disorder with MBL deficiency. These differences could not be fully explained by allele or haplotype frequency variations. Since MBL deficiency is highly heterogeneous and associated with both infectious and autoimmune states, more research is needed to identify which complement pathway components could be associated with bipolar respectively panic disorder.

BACKGROUND: Spaceflight studies and ground-based analogues of microgravity indicate a weakening of human immunity. Mannan-binding lectin (MBL) and H-, L-, and M-ficolin together constitute the lectin pathway and mediate the clearance of pathogens through complement activation. We hypothesized that simulated microgravity may weaken human innate immune functions and studied the impact of 6° head-down tilted bed rest (HDT) for 21 d on MBL and ficolin levels.METHODS: Within a 6-mo period, seven men underwent two periods of HDT. Blood samples were analyzed for MBL, H-, L-, and M-ficolin, mannose-binding lectin-associated protein of 44 kDa (MAp44), and collectin liver 1 (CL-L1) by time-resolved immunofluorometric assays (TRIFMA).RESULTS: We observed well-defined individual preintervention levels of MBL and ficolins. Remarkably similar intraindividual changes occurred for MBL and MBL levels decreased (mean 282 ng · ml(-1)) in the recovery phase. Conversely, CL-L1, a protein with MBL-like properties, increased (mean 102 ng · ml(-1)) during the recovery phase. M-ficolin increased (mean 79 ng · ml(-1)) within the first 2 d of HDT, followed by a decrease (mean 112 ng · ml(-1)) during the recovery phase. L-ficolin increased (mean 304 ng · ml(-1)) during HDT, while H-ficolin was essentially unaffected. MAp44, a down-regulator of the lectin pathway, decreased initially (mean 78 ng · ml(-1)) in the recovery phase followed by an increase (mean 131 ng · ml(-1)).DISCUSSION: Alterations in MBL and ficolin levels were modest and with our current knowledge do not lead to overt immunodeficiency. Pronounced changes occurred when the subjects resumed the upright position. In selected individuals, these changes appear to be a conserved response to HDT.

We recently reported in The Journal of Immunology ([1][1]) the presence of a functional alternative pathway in a human patient with Malpuech–Michels–Mingarelli–Carnevale (3MC) syndrome caused by a genetic defect in the MASP1 gene. This defect abolished the production of all three splice

AIMS/HYPOTHESIS: Increasing evidence links complement activation through the lectin pathway to diabetic nephropathy. Adverse complement recognition of proteins modified by glycation has been suggested to trigger complement auto-attack in diabetes. H-ficolin (also known as ficolin-3) is a pattern recognition molecule that activates the complement cascade on binding to glycated surfaces, but the role of H-ficolin in diabetic nephropathy is unknown. We aimed to investigate the association between circulating H-ficolin levels and the incidence of microalbuminuria in type 1 diabetes.METHODS: We measured baseline H-ficolin levels and tracked the development of persistent micro- and macroalbuminuria in a prospective 18 year observational follow-up study of an inception cohort of 270 patients with newly diagnosed type 1 diabetes.RESULTS: Patients were followed for a median of 18 years (range 1-22 years). During follow-up, 75 patients developed microalbuminuria, defined as a persistent urinary albumin excretion rate (UAER) above 30 mg/24 h. When H-ficolin levels were divided into quartile groups an unadjusted Cox proportional hazards regression model showed a significant association with risk of incident microalbuminuria during follow-up (HR, fourth vs first quartile, 2.45; 95% CI 1.24, 4.85) (p = 0.01). This remained significant after adjusting for HbA1c, systolic blood pressure, smoking and baseline UAER (HR 2.09; 95% CI 1.03, 4.25) (p = 0.04).CONCLUSIONS/INTERPRETATION: Our data suggest that high levels of the complement activating molecule H-ficolin are associated with an increased risk of future progression to microalbuminuria in patients with newly diagnosed type 1 diabetes.

Neutrophils are the main proinflammatory cell type in chronically infected lungs of cystic fibrosis (CF) patients; however, they fail to effectively clear the colonizing pathogens. Here, we investigated the molecular composition of non-mucoid and mucoid Pseudomonas aeruginosa-induced neutrophil extracellular traps (NETs) in vitro and compared them to the DNA-protein complexes present in the CF sputum. The protein composition of P. aeruginosa-induced NET fragments revealed that irrespective of the inducing stimuli, NET fragments were decorated with a conserved set of proteins. The DNA-protein complexes derived from CF sputum were consistent with NETosis and shared a similar protein signature, suggesting that the majority of the extracellular DNA was NET derived. The ability of polymorphonuclear leukocytes to produce NETs in response to P. aeruginosa was driven by macrophage migration-inhibitory factor (MIF) by promoting mitogen-activated protein kinase. Analysis of 132 CF patient samples revealed that elevated MIF protein levels correlated with poorer lung function. We suggest that targeting MIF by small molecular inhibitors might reduce the presence of extracellular DNA and serve as an adjunct to the use of antimicrobial drugs that could ultimately reduce bacterial fitness in the lungs during the later stages of CF disease.

The complement system plays an important role in the induction of inflammation. In this study we demonstrate that the initiation complexes of the lectin pathway, consisting of mannose-binding lectin (MBL) and associated serine proteases (MASPs) elicit Ca 2+ signaling in cultured endothelial cells (HUVECs). This is in agreement with our previous results showing that the recombinant catalytic fragment of MASP-1 activates endothelial cells by cleaving protease activated receptor 4. Two other proteases, MASP-2 and MASP-3 are also associated with MBL. Earlier we showed that recombinant catalytic fragment of MASP-2 cannot activate HUVECs, and in this study we demonstrate that the same fragment of MASP-3 has also no effect. We find the same to be the case if we use recombinant forms of the N-terminal parts of MASP-1 and MASP-2 which only contain non-enzymatic domains. Moreover, stable zymogen mutant form of MASP-1 was also ineffective to stimulate endothelial cells, which suggests that in vivo MASP-1 have the ability to activate endothelial cells directly as well as to activate the lectin pathway simultaneously. We show that among the components of the MBL–MASPs complexes only MASP-1 is able to trigger response in HUVECs and the proteolytic activity of MASP-1 is essential. Our results strengthen the view that MASP-1 plays a central role in the early innate immune response.

BACKGROUND & AIMS: The complement system is activated in liver diseases including acute liver failure (ALF); however, the role of the lectin pathway of complement has scarcely been investigated in ALF. The pathway is initiated by soluble pattern recognition molecules: mannan-binding lectin (MBL), M-, L-, and H-ficolin and collectin-liver-1 (CL-L1), which are predominantly synthesized in the liver. We aimed to study lectin levels in ALF patients and associations with clinical outcome.METHODS: Serum samples from 75 patients enrolled by the US ALF Study Group were collected on days 1 and 3. We included 75 healthy blood donors and 20 cirrhosis patients as controls. Analyses were performed using sandwich-type immunoassays (ELISA, TRIFMA).RESULTS: At day 1, the MBL level in ALF patients was 40% lower compared with healthy controls {[median (interquartile range) 0.72 μg/ml(0.91) vs. 1.15 (1.92)(P = 0.02]}, and increased significantly by day 3 [0.83 μg/ml(0.94)(P = 0.01)]. The M-ficolin level was 60% lower [0.54 μg/ml(0.50) vs. 1.48(1.01)(P < 0.0001)]. The CL-L1 level at day 1 was slightly higher compared with healthy controls [3.20 μg/ml(2.37) vs. 2.64(0.72)(P = 0.11)]; this was significant at day 3 [3.35(1.84)(P = 0.006)]. H- and L-ficolin levels were similar to healthy controls. Spontaneous ALF survivors had higher levels of MBL at day 1 [0.96 μg/ml(1.15) vs. 0.60(0.60)(P = 0.02)] and lower levels of L-ficolin by day 3 compared with patients who died or were transplanted [1.61 μg/ml(1.19) vs. 2.17(2.19)(P = 0.02)].CONCLUSION: We observed significant dynamics in lectin levels in ALF patients, which may suggest they play a role in ALF pathogenesis. High MBL and low L-ficolin levels are associated with survival.