The influence of temperature and incubation time on deformability of human erythrocytes

Human erythrocytes have been heated and stressed in a novel and controlled manner using rectangular microcapillaries. Heated cells attached to the capillary wall were stressed by liquid flow. Under particular conditions of stress, temperature and incubation time the body of the cell could be pulled in the flow, retaining a connection with the glass by means of a narrow process or tether. The tethers appear as: regularly beaded, irregularly beaded or without beads depending upon the incubation conditions. We have outlined the incubation regimes necessary to achieve these different responses in the temperature range 48–55°C. The cells become less deformable as the incubation is continued beyond an optimum time. The behaviour of the tether is compared with that of a viscoelastic liquid. Circular dichroism studies of ghost membranes show that the denaturation of membrane proteins is partially reversible when incubation times are similar to those required to bring about a loss of deformability.     

Characterization of a glycoprotein isolated from a continuous tumor-cell line of human lung carcinoma

A glycoprotein with a molecular weight of 62 000 has been isolated from a tumor-cell line, A549, and purified to homogeneity by gel chromatography. The glycoprotein contained sialic acid, galactose, mannose, N-acetylglucosamine and a relatively high amount of glutamic acid and proline. The data indicated that the overall composition of this glycoprotein was different from that of the glycoprotein of M_r 62 000 isolated from lung lavage of patients with alveolar proteinosis. The glycoprotein did not react with the antiserum from lung lavage of patients with alveolar proteinosis. The glycoprotein did not react with the antiserum raised against glycoprotein of M_r 62 000 isolated from lung lavage of patients with alveolar proteinosis.     

The nature of sialic acids in human lymphocytes

Analysis of the sialic acids obtained by mild acid hydrolysis of B lymphocytes reveals the presence of N-acetylneuraminic acid and 9-O-acetyl-N-acetylneuraminic acid. For T lymphocytes only N-acetylneuraminic acid has been demonstrated to occur. The applied methods include quantitative colorimetry, thin-layer chromatography and combined gas-liquid chromatography-mass spectrometry.     

Study of the carbohydrate moiety of human serum sex hormone-binding globulin

Sex hormone-binding globulin from human blood serum contains two biantennary N-linked oligosaccharide chains of the N-acetyllactosamine type and one O-linked oligosaccharide per one molecule of the glycoprotein. These conclusions have been based on the results of methylation analysis of the whole glycoprotein and investigation of the structures of its glycopeptides prepared using pronase digestion.     

Role of commercial probiotic strains against human pathogen adhesion to intestinal mucus

AIMS: The aims of this study present were to assess and to evaluate in vitro the abilities of commercial probiotic strains derived from fermented milk products and related sources currently marketed in European countries, to inhibit, compete and displace the adhesion of selected potential pathogens to immobilized human mucus. METHODS AND RESULTS: The adhesion was assessed by measuring the radioactivity of bacteria adhered to the human mucus. We tested 12 probiotic strains against eight selected pathogens. All strains tested were able to adhere to mucus. All probiotic strains tested were able to inhibit and displace (P<0.05) the adhesion of Bacteroides, Clostridium, Staphylococcus and Enterobacter. In addition, the abilities to inhibit and to displace adhered pathogens depended on both the probiotic and the pathogen strains tested suggesting that several complementary mechanisms are implied in the processes. CONCLUSIONS: Our results indicate the need for a case-by-case assessment in order to select strains with the ability to inhibit or displace a specific pathogen. Probiotics could be useful to correct deviations observed in intestinal microbiota associated with specific diseases and also, to prevent pathogen infections. SIGNIFICANCE AND IMPACT OF THE STUDY: The competitive exclusion properties of probiotics as well as their ability to displace and inhibit pathogens are the most importance for therapeutic manipulation of the enteric microbiota. The application of such strategies could contribute to expand the beneficial properties on human health against pathogen infection.     

Structural organization of the carbohydrate moiety of human transcortin as determined by methylation analysis of the whole glycoprotein

Methylation analysis of human transcortin showed that this glycoprotein contains N-glycosidically linked oligosaccharide chains of N-acetyllactosamine type, most of the chains being biantennary and others tri- and/or tetraantennary. The carbohydrate chains of transcortin are also heterogenous with respect to the content of fucose and the position of the glycosidic linkages.     

Isolation and partial characterization of glycolipids and a carbohydrate from the secondary granules and plasma membrane of polymorphonuclear leukocytes

Chloroform/methanol extracts of the secondary granule and plasma membrane fractions of polymorphonuclear leukocytes have been shown to contain both non-polar and polar carbohydrate-containing materials. The ratio of the polar to the non-polar material was much higher in the plasma membrane than the secondary granule fraction. The non-polar material contains at least two ceramide-like glycolipids and accounts for most of the broad band of periodic acid/Schiff-positive material which migrates at the dye front in sodium dodecyl sulfate electrophoretic gels of granule and plasma membrane extracts. The polar material appears to be a single substance containing no fatty acids or sialic acid and is composed of glucose, hexosamine and a carboxylic acid derivative of pentose. Expressed on a per mg of protein basis, the amount of carbohydrate associated with the polar material in the plasma membrane fraction was about five times that of the secondary granule fraction.     

Evaluation of ram semen quality using polyacrylamide gel instead of cervical mucus in the sperm penetration test

Fertility is a very complex biological function that depends on several properties of the spermatozoa, including sperm motility. Two objectives are analyzed in this study: (1) Replace the cervical mucus by a synthetic medium in a sperm penetration test, and (2) evaluating the results of this test objectively analyzing the sperm number that migrates. In experiment 1, we have tested eight concentrations of acrylamide (1%-2%). Rheological properties of media were analyzed. The plastic straws, loaded with acrylamide, were placed vertically on the semen sample tube for 15 min at 39 °C. After, the acrylamides were placed, by segments of 5 mm, into wells of a 24-well plate, dyed with Hoechst 33342 and the number of spermatozoa were calculated by automated microscopy analysis. The 1.55% and 1.6% acrylamide gel showed a number of spermatozoa emigrating closer to that seen with natural mucus. In experiment 2, we applied the sperm penetration in acrylamide 1.6% and 1.55% using fresh semen and cooled semen at 15 °C and 5 °C. The spermatozoa counts were performed for each segment of 10 mm. Semen chilled at 15 °C presented intermediate values of sperm counts in comparison with fresh semen (higher) and 5 °C chilled semen. The sperm counts do not differ between acrylamides but the rheological properties of acrylamide 1.6% were more similar to those of the natural cervical mucus. In experiment 3, we have observed significant correlations between the number of spermatozoa and several sperm quality parameters (positive: progressive motility and velocity according to the straight path; negative: damaged acrosomes and apoptotic cells) in 1.6% acrylamide media. We conclude that the size of the cell subpopulation, objectively calculated, that migrate beyond 20 mm in 0.5-mL straws filled with acrylamide is a useful parameter in ram sperm quality assessment and further studies are needed to evaluate its relationship with field fertility.     

Adhesion of four Bifidobacterium strains to human intestinal mucus from subjects in different age groups

The number of bifidobacteria in faeces and intestinal contents has been shown to be reduced with increasing age of the subject. The adhesion of four Bifidobacterium strains was tested to mucus isolated from subjects of different age. All strains bound significantly less to mucus isolated from elderly subjects, compared to mucus from the other age groups. Two of the tested strains also showed decreased adhesion to mucus isolated from 6-month-old and adult subjects compared to the adhesion to mucus from 2-month-old subjects. The results suggest that reduced adhesion may be a factor involved in the decreasing colonisation of elderly subjects by bifidobacteria.     

A search for mixotrophy and mucus trap production in Alexandrium spp. and the dynamics of mucus trap formation in Alexandrium pseudogonyaulax

Recently, a hitherto unknown feeding strategy, the toxic mucus trap, was discovered in the dinoflagellate Alexandrium pseudogonyaulax. In this study, over 40 strains of 8 different Alexandrium species (A. ostenfeldii, A. tamarense, A. catenella, A. taylorii, A. margalefii, A. hiranoi, A. insuetum and A. pseudogonyaulax) were screened for their ability to ingest prey and/or to form mucus traps. The mucus trap feeding strategy, where a mucus trap is towed by the longitudinal flagellum remains unique to A. pseudogonyaulax. In additional experiments, details of the trap were examined and quantified, such as speed and frequency of trap formation as well as what happens to the trap after the A. pseudogonyaulax cell detaches from it. The percentage of A. pseudogonyaulax cells producing a mucus trap and the number of prey cells caught increased with increasing prey concentration, whereas the physical size of the traps was independent of prey concentration. In one strain given an excess of prey, within 1 h over 90% of individual A. pseudogonyaulax cells had formed a trap, each containing an average of 45 prey cells. Individual A. pseudogonyaulax cells steadily produced traps and up to 5 traps were produced by a single A. pseudogonyaulax cell after only 24 h. The attachment of an A. pseudogonyaulax cell to the trap only ceased during, and just following, cell division. Prey cells were, to some extent, capable of escaping from the mucus trap, but the trap remained sticky and continued catching prey for up to 48 h after the trap had been abandoned by the A. pseudogonyaulax cell. These results reveal that the effects of the mucus trap extend far beyond the removal of prey through ingestion, and the potential impact of this strategy on surrounding cells is high.     

A high molecular mass constituent of cranberry juice inhibits helicobacter pylori adhesion to human gastric mucus

Because previous studies have shown that a high molecular mass constituent of cranberry juice inhibited adhesion of Escherichia coli to epithelial cells and coaggregation of oral bacteria, we have examined its effect on the adhesion of Helicobacter pylori to immobilized human mucus and to erythrocytes. We employed three strains of H. pylori all of which bound to the mucus and agglutinated human erythrocytes via a sialic acid-specific adhesin. The results showed that a high molecular mass constituent derived from cranberry juice inhibits the sialic acid-specific adhesion of H. pylori to human gastric mucus and to human erythrocytes.     

Comparison of isolated peanut agglutinin receptor glycoproteins from human,bovine and porcine erythrocyte membranes

Affinity chromatography has been used to isolate and compare the peanut agglutinin receptors from neuraminidase-treated human, bovine and porcine erythrocyte membranes. Passage of Triton X-100-solubilised membrane material through either Sepharose- or acrylamide-based affinity columns resulted in the reversible binding of receptor molecules to the immobilised lectin. Elution with 0.2M galactose released specifically bound glycoprotein fractions, the composition and molecular weights of which were determined by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate.Carbohydrate analysis by gas chromatography identified these bound glycoprotein fractions as the major sources of the $\text{O-}\text{glycosidic-linked}$ disaccharide $\text{galactosyl}\text{-β-(1 → 3)-N-}\text{acetylgalactosamine}$ in these membranes. It is suggested that these isolated fractions represent a discrete population of glycoproteins within the membranes studied, which possess both $\text{O-}\text{glycosidic}$- and $\text{N-}\text{glycosidic-linked}$ carbohydrates.     

Studies on bonnet monkey cervical mucus the effect of proteases on mucus glycoproteins of macaca radiata

The influence of proteinases on monkey cervical glycoproteins was investigated to assess their effect on cervical mucus and, thereby, on sperm penetration. The major component of periovulatory cervical mucus, a high molecular weight glycoprotein, was treated with Pronase, trypsin, chymotrypsin, papain, and bovine seminal peptidase, and the enzyme-resistant glycoprotein was purified by gel filtration on Sepharose 2B. A macromolecular component in high yield was recovered containing carbohydrate and protein moieties. Asialoglycoprotein, on treatment with Pronase, trypsin, and bovine seminal peptidase released more than one glycoprotein fragment. The carbohydrate and amino acid components of the native and degraded glycoproteins were similar in composition with variations in proportions. The structure of the carbohydrate-rich, pronase-resistant glycoprotein, further purified on Sepharose 2B, was examined. Sequential Smith degradation and methylation of the degraded glycoprotein fragment established a structure that shows some differences to that of the native glycoprotein. The influence of proteinases on cervical-mucus glycoproteins and a possible mechanism of sperm penetration through Pronase-treated glycoproteins is discussed.     

Effects of oral treatment with N-acetylcysteine on the viscosity of intrauterine mucus and endometrial function in estrous mares

Persistent breeding-induced endometritis is ranked as the third most common medical problem in the adult mare and leads to enormous economic loss in horse breeding. In mares suffering from persistent breeding-induced endometritis, increased amounts of intrauterine (i.u.) fluid or viscous mucus in estrus or after breeding may act as a barrier for sperm and can contribute to low fertility. Current therapies of these mares aim to eliminate i.u. fluid and mucus by uterine lavage and/or administration of ecbolic drugs. Recently, i.u. administration of N-acetylcysteine (NAC) has been shown to support therapy in mares with endometritis. It was the objective of the present study to investigate effects of an oral administration of NAC on the viscosity of i.u. fluid in estrous mares. It was hypothesized that oral treatment with NAC reduces the viscosity of i.u. fluid and has a positive effect on the inflammatory response of the endometrium. Mares (n = 12) were included in the study as soon as estrus was detected (ovarian follicle >3.0 cm and endometrial edema), which was defined as Day 1. They were randomly assigned to a treatment (10 mg/kg NAC on Days 1-4) or a control group (no treatment). On days 1 and 5 i.u. mucus was collected and its rheologic properties were accessed. On Day 5, endometrial biopsies were obtained and evaluated for integrity of the luminal epithelium, number of polymorphonuclear neutrophils (PMN), staining for cyclooxygenase 2 (COX2), staining with Kiel 67 antigen (Ki-67), lectins and periodic acid Schiff (PAS). In the treatment group, viscosity of i.u. mucus increased significantly between Days 1 and 5 (P < 0.05), while no differences were found in control mares (n.s.). At no time were significant differences between treated and control mares seen. Integrity of epithelium was not affected. After NAC treatment the mean number of PMN in endometrial biopsies was significantly lower compared to mares of the control group (1.9 ± 0.3 vs. 4.8 ± 0.4; P < 0.05). Nuclear immunostaining for COX2 was significantly lower after NAC treatment compared to control mares (P < 0.05). Score for PAS and Alcain staining of mucus in deep uterine glands differed significantly between groups (both P < 0.05). We conclude that oral NAC treatment does not reduce viscosity of uterine mucus but has an antiinflammatory effect on the equine endometrium.     

The viscosity and glycoprotein biochemistry of salmonid mucus varies with species,salinity and the presence of amoebic gill disease

Fish mucus has previously been reported to change in appearance and composition among species and in response to changes in salinity and disease status. This study reports on the mucus viscosity and glycoprotein biochemistry of Atlantic salmon (Salmo salar L.), brown trout (Salmo trutta L.) and rainbow trout (Oncorhynchus mykiss Walbaum) in freshwater and seawater, both naïve to and affected by amoebic gill disease (AGD). Cutaneous mucus viscosity was measured over a range of shear rates (11.5, 23, 46 and 115 s^–1), and non-Newtonian behaviour was demonstrated for all three species. Mucus viscosity was significantly greater in seawater than in freshwater for all species, and significantly lower in AGD-affected Atlantic salmon and brown trout. Mucus glucose, total protein and osmolality data indicated that differences in viscosity due to salinity were mostly attributed to changes in mucus hydration, while differences due to disease were mostly attributed to changes in mucus composition. Trends in gill mucus cell histochemistry included shifts in glycoproteins from neutral mucins in freshwater to acidic mucins in seawater, and shifts towards neutral mucins, with an increase in mucus cell numbers, in response to AGD. Results suggested that Atlantic salmon and brown trout are more similar to one another in their mucus profile than to rainbow trout. Atlantic salmon and brown trout both exhibited a whole-body mucus response to AGD, whereas rainbow trout exhibited only a local gill response. Findings hold implications for fish physiology and pathology, and indicate that future fish-disease management strategies should be species and condition specific.Communicated by I.D. HumeThe word mucus has been used in its noun form throughout the paper for clarity

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An erratum to this article can be found at .     

Scanning electron microscope study of human cervical mucus

The ultrastructure of human cervical mucus was studied by scanning electron microscopy. In fertile women, naturally occurring midcycle cervical mucus showed an arrangement of parallel fibers oriented along the main axis of the mucus sample. Sperm migration through a column of cervical mucus in vitro was also studied. It was found that sperm present among the longitudinal fibers were oriented parallel to the them and to the main axis of the mucus sample.     

Diffusion of macromolecules and virus-like particles in human cervical mucus

To determine whether or not large macromolecules and viruses can diffuse through mucus, we observed the motion of proteins, microspheres, and viruses in fresh samples of human cervical mucus using fluorescent recovery after photobleaching and multiple image photography. Two capsid virus-like particles, human papilloma virus (55 nm, approximately 20,000 kDa) and Norwalk virus (38 nm, approximately 10,000 kDa), as well as most of the globular proteins tested (15-650 kDa) diffused as rapidly in mucus as in saline. Electron microscopy of cervical mucus confirmed that the mesh spacing between mucin fibers is large enough (20-200 nm) for small viruses to diffuse essentially unhindered through mucus. In contrast, herpes simplex virus (180 nm) colocalized with strands of thick mucus, suggesting that herpes simplex virus, unlike the capsid virus particles, makes low-affinity bonds with mucins. Polystyrene microspheres (59-1000 nm) bound more tightly to mucins, bundling them into thick cables. Although immunoglobulins are too small to be slowed by the mesh spacing between mucins, diffusion by IgM was slowed by mucus. Diffusion by IgM-Fc(5 mu), the Fc pentamer core of an IgM with all 10 Fab moieties removed, was comparably slowed by mucus. This suggests that the Fc moieties of antibodies make low-affinity bonds with mucins.     

Biochemical and immunological characterization and the synthesis of rat intestinal glycoproteins following the induction of rapid synchronous vitamin A deficiency

Glycoproteins and proteins were extracted from segments or scrapings of the intestine in tube-fed, vitamin-A-deficient and control rats on the eight day after withdrawal of retinoic acid from the diet by using either 1% sodium dodecyl sulfate (SDS) or aqueous 5 mM EDTA (pH 7.4). They were then fractionated on columns of Sepharose 4B. Water-soluble peak I material contained large (M_r > 10⁶; S₂₀ = 11.7) glycoprotein aggregates which were rich in hexose, fucose and sialic acid. These aggregates dissociated into several non-identical glycoprotein and protein subunits upon treatment with dithiothreitol. The protein matrix was rich in threonine, valine, proline, serine, glutamate and aspartate. Peak II consisted of smaller proteins and glycoproteins, the latter with much lower carbohydrate content. Some peak II glycoproteins also dissociated into subunits in the presence of dithiothreitol. Peak III consisted mainly of a heterogenous assortment of proteins, including some glycoproteins of low carbohydrate content. Antibodies either to peak II or to peak III reacted both with peaks II and III but not with peak I.The total weight, carbohydrate composition of glycoproteins and the ratio of carbohydrate to protein in the total extract or in each of the three fractions were not significantly affected in vitamin A deficiency despite decreased incorporation of all labeled precursors. Rather, the relatively lower incorporation (approx. 0.8) of radioactive sulfate, D-glucosamine and L-fucose into total SDS-soluble duodenal glycoproteins of vitamin-A-deficient rats could be explained on the basis of a reduced prevalence of goblet cells alone. In contrast, the relative incorporation rate of L-fucose into peak I, but not into peaks II and III, ranged from 0.25 to 0.45, less than expected on the basis of fewer goblet cells alone. The incorporation of radioactive threonine into all protein fractions was reduced to 60% of normal in vitamin A deficiency. Thus, the well established observation that intestinal tissue of vitamin-A-deficient rats synthesizes high molecular weight glycoproteins poorly might be due to several interacting factors: (1) a reduced prevalence of goblet cells, (2) a lower rate of protein synthesis, (3) a lack of retinyl phosphate for the formation of mannosyl or other carbohydrate derivatives, and (4) secondary, and as yet undefined, cellular changes which preferentially reduce the rate of synthesis of high molecular weight fucose- and sialic-acid-enriched glycoproteins.     

Enhancement of mucus accumulation in a human gastric scirrhous carcinoma cell line (KATO-III) by fibroblasttumor cell interaction

Human fibroblasts (WI-38 cells) were found to enhance mucus accumulation by human scirrhous carcinoma cells (KATO-III cells). Coculture of KATO-III with WI-38 cells resulted in enlargement of the KATO-III cells and increases in the proportions of PAS- and colloidal iron-positive KATO-III cells. These morphological alterations were reversed when the KATO-III cells were again cultured without WI-38 cells. Conditioned media from cultures of WI-38 cells or cocultures of KATO-III and WI-38 cells induced the same morphological alterations in KATO-III cells, suggesting that WI-38 cells produce a factor or factors that enhance mucus accumulation in KATO-III cells. This factor seemed to be a protein with a molecular weight of more than 10,000 daltons.     

Influence of snail mucus trails and first snail meal on the behavior of malacophagous sciomyzid larvae

There has been considerable focus on the natural enemies of snails, particularly those of medical and veterinary significance. Much attention has focussed on members of the Family Sciomyzidae (Diptera), the majority of which feed on a range of mollusc species. However, little is known about the influence of first snail meal on subsequent prey choices, an important consideration in biocontrol. We examined neonate larval responses of Ilione albiseta to fresh and aged snail mucus trails of three snail species. Median neonate response rates to aged mucus trails for all three snail species tested were significantly (P < 0.001) weaker than for fresh mucus trails indicating a strategy which enhances the likelihood of reaching prey snail species without expending energy following “cold” trails. More than 78% of first instar larvae, fed on one snail species (Radix peregra or Stagnicola palustris) and subsequently offered a choice of these two snail species for the second snail meal, selected the snail species of the first snail meal suggesting that the first snail meal influences subsequent prey selection. However, the impact of the first snail meal on larval trail-following behavior is less clear-cut. While there may be some preference for the mucus trail of the snail species on which neonate larvae have fed, this does not exclude the larvae from following the mucus trails of other snail species. The results are discussed in the context of the potential use of I. albiseta as a biocontrol agent of vectors of snail borne diseases.