Anthony P. Corfield

Mucins and mucosal protection in the gastrointestinal tract: new prospects for mucins in the pathology of gastrointestinal disease

The luminal surface of the gastrointestinal tract is covered by a viscoelastic mucous gel layer that acts as a protective barrier against the harsh luminal environment. The structural characteristics of this barrier are primary indicators of its physiological function and changes to its composition have long been identified in gastrointestinal pathologies. During the past decade significant improvements in analytical techniques coupled with detailed knowledge of the genes coding for the mucin proteins have provided exciting new insights into the role of the mucous layer and its relevance to gastrointestinal disease. The high molecular weight mucins are responsible for the viscoelastic properties of the mucous barrier. They are widely expressed in epithelial tissues and are characterised by variable number tandem repeat peptide sequences rich in serine, threonine, and proline which carry large numbers of O -linked oligosaccharide chains.1 2 At present, 12 genes have been described, shown in table 1.1 3 Secreted and membrane associated forms have been found based on their function as extracellular viscous secretions or viscoelastic polymer gels or location as membrane anchored molecules in the glycocalyx.3 4 Two clusters have been reported, the secretory mucin genes MUC2, MUC5AC, MUC5B, and MUC6 on chromosome 11p15.5, and MUC3, MUC11, and MUC12 on chromosome 7q22.3 View this table: Table 1 Mucin genes and their location in the human gastrointestinal tract Histochemical techniques for mucin detection rely on the ability to detect carbohydrate or negative charge and were widely used for classification of changes in disease.5 6 The use of lectins and anticarbohydrate antibodies has greatly improved the specific detection of mucins histochemically and biochemically.5 6 A group of mucin oligosaccharide antigens, including Tn, sialyl-Tn, T, Lewisx and Lewisy, sialyl and sulpho-Lewisx and -Lewisa, and the blood group ABH antigens, have been identified …

Mucins: A biologically relevant glycan barrier in mucosal protection

BACKGROUND The mucins found as components of mucus gel layers at mucosal surfaces throughout the body play roles in protection as part of the defensive barrier on an organ and tissue specific basis. SCOPE OF THE REVIEW The human MUC gene family codes up to 20 known proteins, which can be divided into secreted and membrane-associated forms each with a typical protein domain structure. The secreted mucins are adapted to cross link in order to allow formation of the extended mucin networks found in the secreted mucus gels. The membrane-associated mucins possess membrane specific domains which enable their various biological functions as part of the glycocalyx. All mucins are highly O-glycosylated and this is tissue specific and linked with specific biological functions at these locations. Mucin biology is dynamic and the processes of degradation and turnover are well integrated with biosynthesis to maintain a continuous mucosal protection against all external aggressive forces. Interaction of mucins with microflora plays an important role in normal function. Mucins are modified in a variety of diseases and this may be due to abberant mucin peptide or glycosylation. MAJOR CONCLUSIONS Mucins represent a family of glycoprotein having fundamental roles in mucosal protection and communication with external environment. GENERAL SIGNIFICANCE The review emphasises the nature of mucins as glycoproteins and their role in presenting an array of glycan structures at the mucosal cell surface.

Mucinases and sialidases: their role in the pathogenesis of sexually transmitted infections in the female genital tract

Background: Mucinases and sialidases contribute to the process of invasion and colonisation in many conditions and infections of the female reproductive tract by degrading the protective cervical mucus. The role of hydrolytic enzymes in the pathogenesis of sexually transmitted diseases and their effect on cervical mucus are discussed in this review. Methods: Articles were searched for using the keywords “sialidase,” “mucinase,” “protease,” and “sexually transmitted infections.” As well as review and other articles held by our group, searches were conducted using PubMed, Grateful Med, and the University of Bath search engine, BIDS. Results: Numerous publications were found describing the production of hydrolytic enzymes in sexually transmitted diseases. Because the number of publications exceeded the restrictions imposed on the size of the review, the authors selected and discussed those which they considered of the most relevance to sexually transmitted infections.

Ocular mucins: Purification, metabolism and functions

Abstract Mucins are present at the ocular surface in both secreted and membrane-bound forms. Mucins are produced in partby the conjunctial goblet cells, and are complemented by non-globet secretions. This review focuses on secreted ocular mucins. They are present in the tear film, probably both in gel and soluble form, and play a role in lubrication and ocular defense. It is apparent that mucins are highly adapted to their functions. State of the art techniques for mucin purification and analysis are presented. Density gradient centrifugation, gel filtration, ion-exchange chromatography and agarose gel electrophoresis are discussed, together with methods of oliogosaccharide analysis. Reagents for the detection of mucin are considered in conjunction with these methods, which we have employed in the analysis of human and canine ocular mucins. The general structure of mucins is reviewed. The biosyntheas and glycosylation of ocular mucins are not yet fully understood, and are discussed in relation to currently established concepts. The impaact of disease on the nature and secretion of mucins is considered, as well as the physiological and pathological significance of mucus degradation.

The roles of enteric bacterial sialidase, sialate O-acetyl esterase and glycosulfatase in the degradation of human colonic mucin

Sialidase activity in normal faecal extracts showed a preference for mucin-related glycoprotein and oligosaccharide substrates, but the presence of two or moreO-acetyl esters at positions C7–C9 on the sialic acids retarded the rate of hydrolysis. A specific sialateO-acetyl esterase was detected with a lower total activity relative to sialidase with mucin substrates and having a pH optimum of 7.8 and aKM of approximately 1mm sialateO-acetyl ester. A specific glycosulfatase activity was found in faecal extracts using the substrate lactit-[3H]ol 6-O-sulfate with a pH optimum of pH 5.0 and aKM of approximately 1mm.Faecal extracts from ulcerative colitis (UC) patients had higher sialateO-acetyl esterase and glycosulfatase activity, while mucin sialidase activity was unchanged.Metabolically labelled mucin isolated from UC patients contained less sulfate and had lower sialic acidO-acetylation compared with normal mucin. Colonic mucin was degraded more efficiently by faecal extracts from UC patients compared with normal extracts. The UC mucin was degraded more rapidly than the normal mucin by faecal enzyme extracts from both normal and UC subjects.

The role of mucins in host–parasite interactions: Part II – helminth parasites

Some parasites express mucin-like molecules. These have possible roles in attachment and invasion of host cells and in the avoidance of host immune processes. Enzymes of parasite origin might also facilitate infection, either by degrading host mucus barriers or by generating binding sites on host cells. Host mucins have roles in preventing parasite establishment or in parasite expulsion. They, in turn, might be exploited by parasites, either as sources of fuel or binding sites, or as host-finding targets. Here, we describe the biochemical properties of mucins and mucin-like molecules in relation to interactions (established and putative) between helminth parasites and their hosts.

Reduction of sialic acid O-acetylation in human colonic mucins in the adenoma-carcinoma sequence

The oligo-O-acetylation of sialic acids found in normal colonic mucins is greatly reduced in colorectal cancer. Mucins prepared from cancer tissue in adenocarcinoma showed this reduction, while normal O-acetylation was detected in resection margin and control cases and total mucin sialic acid content was significantly decreased in cancer vs control samples. A reduction of the O-acetyl transferase activity catalysing the O-acetylation reaction was also found. A series of cultured human colorectal cell lines derived from the same premalignant adenomatous line, and representative of the adenoma-carcinoma sequence were examined and revealed a depletion of oligo-O-acetylation in the original diploid premalignant line, re-expression in a further premalignant line and reduction in malignant mucinous and adenocarcinoma cell lines. Reduction of sialic acid O-acetylation appears as an early event in the process of malignant transformation in human colorectal cancer.

Colonic mucins in ulcerative colitis: evidence for loss of sulfation.

Colonic tissue obtained at surgery from control individuals and patients with ulcerative colitis was used to isolate mucins and to prepare mucin glycopolypeptides by pronase digestion. These were compared with mucins labelled with [35S] sulfate and [3H]-glucosamine after organ culture tissue samples from the same patients. A significant loss of mucin sulfation was detected in the colitis patients by both metabolic labelling and chemical analysis of the glycopolypeptides. A change in the size distribution of purified mucin oligosaccharides fractionated on BioGel P6 after release by β-elimination was seen in both radiolabelled and non-labelled colitis mucins compared with controls. Amino acid analysis of the glycopolypeptides showed a close similarity to the expected ratio of serine:threonine:proline for MUC2 and did not vary between control and colitis groups. Analysis of the mucins confirmed >90% purity in the labelling experiments, characteristic behaviour on density gradient centrifugation and agarose gel electrophoresis in control and ulcerative colitis groups and differences in sulfation and turnover at various sites in the normal colon.

In Vitro and ex Vivo Intestinal Tissue Models to Measure Mucoadhesion of Poly (Methacrylate) and N-Trimethylated Chitosan Polymers

No HeadingPurpose.The adhesion of a range of polymers based on poly(2-(dimethylamino-ethyl) methacrylate (pDMAEMA) was assessed using human mucus-secreting and non mucus-secreting intestinal cell monolayers, HT29-MTX-E12 (E12) and HT29 monolayers, as well as excised non-everted intestinal sacs from rats. Differentiation of mucoadhesion from bioadhesion was achieved by pre-treatment with the mucolytic agent, N-acetyl cysteine (NAC). Adherence of pDMAEMA polymers was compared to that obtained with the mucoadhesive, N-trimethylated chitosan (TMC).Methods.The quantity of adherent coumarin 343-conjugated polymers to HT29, E12, and intestinal sacs was measured by fluorescence. Confocal laser scanning microscopy (CLSM), light microscopy, and fluorescent microscopy were used to provide direct evidence. Measurements of transepithelial electrical resistance (TEER), permeability to FITC-dextran 4000 (FD-4), and the release of lactate dehydrogenase (LDH) were used to assess potential cytotoxicity of polymers.Results.Adherence of unquaternized and of 10%, 24%, and 32% methyl iodide-quaternized pDMAEMA polymers was measured in E12, HT29, and sacs. All pDMAEMA polymers showed significantly higher levels of adhesion to mucus (mucoadhesion) than to epithelium (bioadhesion). Colocalization of pDMAEMA with mucus was confirmed in E12 by microscopy. TMC showed equally high levels of mucoadhesion as unquaternized and 24% quaternized pDMAEMA, but displayed higher levels of bioadhesion. pDMAEMA-based polymers demonstrated lower levels of adherence to E12 and rat sacs in the presence of NAC, whereas adherence of TMC was unchanged. pDMAEMA significantly decreased the permeability of FD-4 across E12 monolayers and sacs and was less cytotoxic in E12 than in HT29. In contrast, TMC increased the permeability of FD-4 across E12 and sacs and was less cytotoxic in E12 than in HT29.Conclusions.Human mucus–producing E12 monolayers can be used to assess polymer mucoadhesion and give similar data to isolated rat intestinal sacs. pDMAEMA displayed similar levels of mucoadhesion and lower levels of bioadhesion than a chitosan derivative and it was not cytotoxic. pDMAEMA decreased FD-4 flux in the presence of mucus, whereas TMC increased it. The combination of mucus and methacrylate polymers appears to increase barrier function of the apical membrane.

Glycan variation and evolution in the eukaryotes

In this review, we document the evolution of common glycan structures in the eukaryotes, and illustrate the considerable variety of oligosaccharides existing in these organisms. We focus on the families of N- and O-glycans, glycosphingolipids, glycosaminoglycans, glycosylphosphatidylinositol (GPI) anchors, sialic acids (Sias), and cytoplasmic and nuclear glycans. We also outline similar and divergent aspects of the glycans during evolution within the groups, which include inter- and intraspecies differences, molecular mimicry, viral glycosylation adaptations, glycosyltransferase specificity relating to function, and the natural dynamism powering these events. Finally, we present an overview of the patterns of glycosylation found within the groups comprising the Eukaryota, namely the Deuterostomia, Fungi, Viridiplantae, Nematoda, and Arthropoda.