Mucin Binding to Moraxella catarrhalis during Airway Inflammation Is Dependent on Sialic Acid.

Chronic obstructive pulmonary disease (COPD) is associated with colonization by bacterial pathogens and repeated airway infections, leading to exacerbations and impaired lung function. The highly glycosylated mucins in the mucus lining the airways are an important part of the host defense against pathogens. However, mucus accumulation can contribute to COPD pathology. Here, we examined whether inflammation is associated with glycosylation changes that affect interactions between airway mucins and pathogens. We isolated mucins from lower airway samples (n = 4-9) from long-term smokers with and without COPD and from never-smokers. The most abundant terminal glycan moiety was N-acetylneuraminic acid (Neu5Ac) among smokers with and without COPD and N-acetyl-hexoseamine among never-smokers. Moraxella catarrhalis bound to MUC5 mucins from smokers with and without COPD. M. catarrhalis binding correlated with inflammatory parameters and Neu5Ac content. M. catarrhalis binding was abolished by enzymatic removal of Neu5Ac. Furthermore, M. catarrhalis bound to α2,6 sialyl-lactose, suggesting that α2,6 sialic acid contributes to M. catarrhalis binding to mucins. Furthermore, we detected more M. catarrhalis binding to mucins from patients with pneumonia than to those from control subjects (n = 8-13), and this binding correlated with C-reactive protein and Neu5Ac levels. These results suggest a key role of inflammation-induced Neu5Ac in the adhesion of M. catarrhalis to airway mucins. The inflammation-induced ability of MUC5 mucins to bind M. catarrhalis is likely a host defense mechanism in the healthy lung, although it cannot be excluded that impaired mucociliary clearance limits the effectiveness of this defense in patients with COPD.

Effects of Size and Geographical Origin on Atlantic salmon, Salmo salar, Mucin O-Glycan Repertoire.

Diseases cause ethical concerns and economic losses in the Salmonid industry. The mucus layer comprised of highly O-glycosylated mucins is the first contact between pathogens and fish. Mucin glycans govern pathogen adhesion, growth and virulence. The Atlantic salmon O-glycome from a single location has been characterized and the interindividual variation was low. Because interindividual variation is considered a population-based defense, hindering the entire population from being wiped out by a single infection, low interindividual variation among Atlantic salmon may be a concern. Here, we analyzed the O-glycome of 25 Atlantic salmon from six cohorts grown under various conditions from Sweden, Norway and Australia (Tasmania) using mass spectrometry. This expanded the known Atlantic salmon O-glycome by 60% to 169 identified structures. The mucin O-glycosylation was relatively stable over time within a geographical region, but the size of the fish affected skin mucin glycosylation. The skin mucin glycan repertoires from Swedish and Norwegian Atlantic salmon populations were closely related compared with Tasmanian ones, regardless of size and salinity, with differences in glycan size and composition. The internal mucin glycan repertoire also clustered based on geographical origin and into pyloric cecal and distal intestinal groups, regardless of cohort and fish size. Fucosylated structures were more abundant in Tasmanian pyloric caeca and distal intestine mucins compared with Swedish ones. Overall, Tasmanian Atlantic salmon mucins have more O-glycan structures in skin but less in the gastrointestinal tract compared with Swedish fish. Low interindividual variation was confirmed within each cohort. The results can serve as a library for identifying structures of importance for host-pathogen interactions, understanding population differences of salmon mucin glycosylation in resistance to diseases and during breeding and selection of strains. The results could make it possible to predict potential vulnerabilities to diseases and suggest that inter-region breeding may increase the glycan diversity.

Exploring the Arctic Charr Intestinal Glycome: Evidence of Increased N-Glycolylneuraminic Acid Levels and Changed Host-Pathogen Interactions in Response to Inflammation.

Disease outbreaks are a limiting factor for the sustainable development of the aquaculture industry. The intestinal tract is covered by a mucus layer mainly comprised by highly glycosylated proteins called mucins. Mucins regulate pathogen adhesion, growth, and virulence, and the glycans are vital for these functions. We analyzed intestinal mucin O-glycans on mucins from control and full-fat extruded soy-bean-fed (known to cause enteritis) Arctic charr using liquid chromatography-tandem mass spectrometry. In total, 56 glycans were identified on Arctic charr intestinal mucins, with a high prevalence of core-5-type and sialylated O-glycans. Disialic-acid-epitope-containing structures including NeuAcα2,8NeuAc, NeuAc(Gc)α2,8NeuGc(Ac), and NeuGcα2,8NeuGc were the hallmark of Arctic charr intestinal mucin glycosylation. Arctic charr fed with soy bean meal diet had lower (i) number of structures detected, (ii) interindividual variation, and (iii) N-glycolylneuraminic-acid-containing glycans compared with control Arctic charr. Furthermore, Aeromonas salmonicida grew less in response to mucins from inflamed Arctic charr than from the control group. The Arctic charr glycan repertoire differed from that of Atlantic salmon. In conclusion, the loss of N-glycolylneuraminic acid may be a biomarker for inflammation in Arctic char, and inflammation-induced glycosylation changes affect host-pathogen interactions.

Aeromonas salmonicida binds differentially to mucins isolated from skin and intestinal regions of Atlantic salmon in an N-acetylneuraminic acid-dependent manner.

Aeromonas salmonicida subsp. salmonicida infection, also known as furunculosis disease, is associated with high morbidity and mortality in salmonid aquaculture. The first line of defense the pathogen encounters is the mucus layer, which is predominantly comprised of secreted mucins. Here we isolated and characterized mucins from the skin and intestinal tract of healthy Atlantic salmon and studied how A. salmonicida bound to them. The mucins from the skin, pyloric ceca, and proximal and distal intestine mainly consisted of mucins soluble in chaotropic agents. The mucin density and mucin glycan chain length from the skin were lower than were seen with mucin from the intestinal tract. A. salmonicida bound to the mucins isolated from the intestinal tract to a greater extent than to the skin mucins. The mucins from the intestinal regions had higher levels of sialylation than the skin mucins. Desialylating intestinal mucins decreased A. salmonicida binding, whereas desialylation of skin mucins resulted in complete loss of binding. In line with this, A. salmonicida also bound better to mammalian mucins with high levels of sialylation, and N-acetylneuraminic acid appeared to be the sialic acid whose presence was imperative for binding. Thus, sialylated structures are important for A. salmonicida binding, suggesting a pivotal role for sialylation in mucosal defense. The marked differences in sialylation as well as A. salmonicida binding between the skin and intestinal tract suggest interorgan differences in the host-pathogen interaction and in the mucin defense against A. salmonicida.

Gill Mucus and Gill Mucin O-glycosylation in Healthy and Amebic Gill Disease-Affected Atlantic Salmon.

Amoebic gill disease (AGD) causes poor performance and death in salmonids. Mucins are mainly comprised by carbohydrates and are main components of the mucus covering the gill. Since glycans regulate pathogen binding and growth, glycosylation changes may affect susceptibility to primary and secondary infections. We investigated gill mucin O-glycosylation from Atlantic salmon with and without AGD using liquid chromatography-mass spectrometry. Gill mucin glycans were larger and more complex, diverse and fucosylated than skin mucins. Confocal microscopy revealed that fucosylated mucus coated sialylated mucus strands in ex vivo gill mucus. Terminal HexNAcs were more abundant among O-glycans from AGD-affected Atlantic salmon, whereas core 1 structures and structures with acidic moieties such as N-acetylneuraminic acid (NeuAc) and sulfate groups were less abundant compared to non-infected fish. The fucosylated and NeuAc-containing O-glycans were inversely proportional, with infected fish on the lower scale of NeuAc abundance and high on fucosylated structures. The fucosylated epitopes were of three types: Fuc-HexNAc-R, Gal-[Fuc-]HexNAc-R and HexNAc-[Fuc-]HexNAc-R. These blood group-like structures could be an avenue to diversify the glycan repertoire to limit infection in the exposed gills. Furthermore, care must be taken when using skin mucus as proxy for gill mucus, as gill mucins are distinctly different from skin mucins.

Obesity abrogates the concentration-dependent effect of leptin on endogenous cholesterol synthesis in human monocytes.

Leptin the cytokine-like hormone is involved not only in local inflammations, but it regulates cholesterol biosynthesis in human monocytes. Since, monocyte-membrane composition in obesity shows considerable difference from control cells, our aim was to elucidate the concentration dependence of the effect of leptin in OW monocytes, and the downstream signaling of high and low leptin concentrations. Control and OW monocytes were stimulated with leptin in the presence or absence of different inhibitors. Our results are as follows: a concentration-dependent biphasic effect could only be detected in control monocytes whereas in OW cells only elevated cholesterol synthesis was found. The signal pathway of 50 ng/mL leptin stimulation involves Ca(2+) signal, activation of PI3K, MAPK and HMG CoA reductase. In the 500 ng/mL leptin-stimulated control monocytes the suppression of cholesterol synthesis was dependent on the Ca(2+) signal, the H-7 sensitive cPKC and PI3K activation, whereas in OW monocytes only PI3K was involved in increased cholesterol synthesis. We conclude that leptin-signaling in OW monocytes is characterized by Ca(2+) influx, abrogation of H-7 sensitive cPKC activation, and by PI3K mediated PKC activation.