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Characterization of Tru-Trax in-vitro penetration testing of cervical mucus.

To determine the distribution of results of an in-vitro cervical mucus penetration system employing both human and bovine mucus (Tru-Trax) in a general infertility population, 133 couples prospectively underwent in-vitro and post-coital testing of cervical mucus. The distribution of Tru-Trax results in couples with normal semen analyses and Insler scores showed significantly greater penetration in bovine (22.3 +/- 6.0 mm) than in human mucus (20.3 +/- 5.4 mm) (P < 0.001). However, the lower limit of the 95% confidence interval of the normal population with either type of mucus was approximately 10 mm. This limit was significantly lower than that described by the kit manufacturer. The predictive value for post-coital tests using human mucus in the Tru-Trax system was good (> 90%) in all groups. The overall penetration into either human or bovine mucus was significantly correlated with the percentage of motile spermatozoa in the semen sample (P < 0.001). In conclusion, in-vitro cervical mucus penetration tests with the Tru-Trax system are related to in-vivo post-coital tests, although the lower limit of the range of anticipated results in the normal population was significantly lower than that described by the manufacturer. The ultimate value of this type of testing will await clinical trials which evaluate clinical end-points such as pregnancy rates.

Adult↗

Eradication of small intestinal bacterial overgrowth in the cystic fibrosis mouse reduces mucus accumulation.

OBJECTIVES: Mucus accumulation in cystic fibrosis (CF) is involved in blockage of the distal small intestine. Because expression of mucin genes and mucus secretion can be increased by infection and previous work indicated that small intestinal bacterial overgrowth occurs in CF, we tested whether reduction of bacterial load by antibiotic treatment would reduce mucin gene expression and mucus accumulation in the CF mouse small intestine. METHODS: CF transmembrane conductance regulator null (cftr (tm1UNC)) and wild type littermates were treated with ciprofloxacin and metronidazole for 3 weeks. Muc2 and Muc3 gene expression were measured by quantitative reverse-transcriptase polymerase chain reaction. Periodic acid Schiff (PAS) staining and morphometry were used to measure the size of mucus droplets within goblet cells and dilation of the intestinal crypt lumen, as estimates of mucus secretion and accumulation. RESULTS: Antibiotic treatment did not significantly affect Muc2 and Muc3 gene expression in CF mice. In untreated CF mice, the crypt lumen was almost sevenfold wider than wild type. Antibiotic treatment of CF mice reduced the intensity of PAS crypt lumen staining, and the lumen width was decreased by approximately 25%. The area occupied by PAS-positive material in goblet cells was significantly greater in tissues from antibiotic treated mice. CONCLUSIONS: Eradication of bacterial overgrowth in CF mice significantly decreased mucus secretion and accumulation in intestinal crypts without an effect on mucin gene expression. It is proposed that bacterial overgrowth stimulates mucus secretion, which contributes to its accumulation in the small intestine. Control of bacterial overgrowth is expected to reduce mucus accumulation and may improve intestinal function and overall health in CF.

Animals↗

Immunoglobulin levels and phagocytes in the cervical mucus plug at term of pregnancy.

BACKGROUND: To characterize the potential for adaptive immune protection in cervical mucus plugs with respect to immunoglobulin isotypes and effector cells (phagocytes). METHODS: Thirty-one cervical mucus plugs were collected from healthy women in labor at term. The cervical mucus plugs were allocated either to analysis of immunoglobulins by enzyme-linked immunosorbent assay (ELISA), gel chromatography and Western blotting (n = 20) or to microscopical, including immunocytochemical, analyses. The levels of immunoglobulin in the plugs were compared to the levels in 10 samples of ovulatory cervical mucus from nonpregnant women. RESULTS: In the cervical mucus plugs, levels of immunoglobulin G (IgG) [median 3270 microg/mL (100-14 500)] and IgA [540 (22-2820)], but not IgM [30.5 (1.0-160)], were significantly elevated compared to cervical mucus from nonpregnant women (p < 0.02 for IgG and IgA). The IgG : IgA ratio in the plugs was also elevated (p < 0.02). The proportion of secretory immunoglobulin A (SIgA) relative to total IgA in the plugs ranged from 16 to 65% (n = 5). IgA and IgG were largely intact. Microscopically, the vagina-proximal part of the cervical mucus plugs contained bacteria and was rich in cells, mainly phagocytes. Conversely, the uterine part contained few cells. CONCLUSION: The high immunoglobulin levels in combination with the presence of phagocytes suggest a potential for adaptive immune defense in the cervical mucus plug, which, together with innate immune factors, may act as an immunological gatekeeper protecting the fetomaternal unit against infection from the vagina.

Blotting, Western↗

Adhesion to sole, Solea senegalensis Kaup, mucus of microorganisms isolated from farmed fish, and their interaction with Photobacterium damselae subsp. piscicida.

Abstract Most studies carried out to select microorganisms as candidate probiotics have focused on in vitro antagonism tests, such as the production of inhibitory compounds against pathogenic microorganisms. However, attachment to mucous surfaces could be another criterion to be considered when selecting potential probiotics for aquaculture. Nineteen isolates obtained from farmed Senegalese sole, Solea senegalensis Kaup, and gilthead sea bream, Sparus aurata L., have been evaluated for their capacity to adhere to skin and intestinal mucus of Senegalese sole, and their antagonistic effect against Photobacterium damselae subsp. piscicida, an important pathogen for farmed sole. The isolates from gilthead sea bream showed the highest percentage of adhesion to sole mucus, whilst the pathogenic microorganisms assayed and the isolates from sole showed, in general, a lower ability to adhere to sole mucus. The results suggest that the adhesion to fish mucus was more dependent on the isolate tested than on the host mucus. The isolates from gilthead sea bream also showed a higher antagonistic activity against P. damselae subsp. piscicida than those from Senegalese sole. Four isolates were selected, on the basis of their adhesive ability and antagonistic effect on P. damselae subsp. piscicida, to study their interactions with the pathogen in respect of adhesion to skin and intestinal mucus under exclusion, competition and displacement conditions. The results obtained show the ability of three isolates to reduce the adhesion of P. damselae subsp. piscicida to sole mucus under displacement and competition conditions. The adhesion of the pathogen to sole intestinal mucus was also significantly reduced when three isolates were assayed under exclusion conditions.

Animals↗

Relationship between gastric mucus synthesis, secretion and surface gel erosion measured in amphibian stomach in vitro.

1. The layer of adherent mucus that protects the surface of the stomach reflects a dynamic balance between biosynthesis of glycoprotein, secretion of preformed mucus and erosion of the adherent gel layer. The present study is the first in which all these processes have been measured concomitantly and was undertaken to define interrelationships between the three parameters. 2. A chambered sac preparation of amphibian gastric mucosa is described. Biosynthesis was determined by specific incorporation of radiolabelled sugars into purified glycoprotein. Mucus secretion was determined by measuring the thickness of the adherent gel and erosion of the surface layer was assessed from the appearance of soluble mucin in the luminal solution. 3. 16,16-Dimethyl-prostaglandin (PG) E2 stimulated glucosamine incorporation by 10-fold, but did not alter the rate of incorporation of galactose. There was a rapid two-fold increase in the thickness of the adherent mucus layer but no change in the rate of erosion. Dibutyryl-cAMP also stimulated mucus release but, unlike PG, increased glycoprotein labelling by galactose. 4. Both distention or the application of a cholinergic agonist increased adherent mucus thickness. Stimulation of mucus release in response to carbachol was accompanied by a decrease in glycoprotein labelling by galactose. In contrast, the adrenergic agent noradrenaline decreased secretion but did not influence labelling. 5. These results indicate that biosynthesis and secretion of gastric mucus are subject to differential regulation. Moreover, the profile of incorporation of sugars in response to secretagogues also differs, indicating the need for caution when interpreting effects on glycoprotein biosynthesis.

16,16-Dimethylprostaglandin E2↗

Cervical mucus prolactin levels in normal fertile and infertile women.

Cervical mucus and serum samples were obtained from 42 anovular women and 20 normal ovular fertile women (controls) for the determination of prolactin concentration by a solid phase radioimmunoassay. The former group was subgrouped into 14 galactorrhoeic (9 with oligohypomenorrhea and 5 with amenorrhea) and 28 non-galactorrhoeic (19 with oligohypomenorrhea and 9 with amenorrhea). The level of prolactin in cervical mucus of normally menstruating women was significantly higher than that of serum at P less than 0.0125 (11.68 +/- 0.77 ng/ml and 16.09 +/- 1.65 ng/ml, mean +/- SE, respectively). Serum prolactin level in galactorrhoeic amenorrhea cases was 8.2 times that of controls, while cervical mucus prolactin amounted to 32 times its control value. The rise of cervical mucus prolactin in cases of galactorrhea oligohypomenorrhea was 21 times its control value which is markedly higher than that of serum (1.9 times). The average rise of serum prolactin in the whole group of galactorrhea with abnormal menstrual function was about 4 times the control value, while the average rise of cervical mucus prolactin was about 41 times the corresponding control value. In the cases of galactorrhea with menstrual dysfunction, serum prolactin level may be normal in 50% of cases, while cervical mucus prolactin is strikingly elevated in 100% of cases. The possibility of pituitary adenoma should be considered if the level of serum or cervical mucus prolactin exceeds 100 ng/ml and 600 ng/ml respectively. The possible physiological role of cervical mucus prolactin and its source are discussed.

Cervix Mucus↗

Interactions of Giardia lamblia with human intestinal mucus: enhancement of trophozoite attachment to glass.

Giardia lamblia trophozoites frequently are associated with mucus in vivo. We investigated the effects of human intestinal mucus on parasite attachment and survival in vitro. All samples of mucus from the duodenum and ileum (from four humans and two rabbits) enhanced attachment at 100 micrograms/ml. Attachment increased with mucus concentrations from 1 to 1000 micrograms/ml but declined toward the unstimulated level at concentrations above 1000 micrograms/ml. Mucus from the small intestine also promoted the survival of the parasites during the 2-h incubation. In contrast, colonic mucus promoted survival, but inhibited attachment. Fractionation of mucus from the human small intestine by cesium chloride equilibrium density gradient ultracentrifugation revealed that both attachment- and survival-promoting activities were in the low density, protein-rich fraction. The high density fractions containing the mucins were devoid of activity. Thus, a non-mucin fraction of mucus from the human small intestine may promote colonization by G. lamblia.

Adhesiveness↗

Intestinal mucus protects Giardia lamblia from killing by human milk.

We have previously shown that nonimmune human milk kills Giardia lamblia trophozoites in vitro. Killing requires a bile salt and the activity of the milk bile salt-stimulated lipase. We now show that human small-intestinal mucus protects trophozoites from killing by milk. Parasite survival increased with mucus concentration, but protection was overcome during longer incubation times or with greater milk concentrations. Trophozoites preincubated with mucus and then washed were not protected. Protective activity was associated with non-mucin CsCl density gradient fractions. Moreover, it was heat-stable, non-dialyzable, and non-lipid. Whereas whole mucus inhibited milk lipolytic activity, protective mucus fractions did not inhibit the enzyme. Furthermore, mucus partially protected G. lamblia trophozoites against the toxicity of oleic acid, a fatty acid which is released from milk triglycerides by lipase. These studies show that mucus protects G. lamblia both by inhibiting lipase activity and by decreasing the toxicity of products of lipolysis. The ability of mucus to protect G. lamblia from toxic lipolytic products may help to promote intestinal colonization by this parasite.

Animals↗

Modulation of Pseudomonas aeruginosa adherence to the corneal surface by mucus.

To gain access to the corneal epithelium and cause infections keratitis, bacterial pathogens must first interact with ocular surface factors that could affect bacterial adherence. In this study, we demonstrated that the mucus layer, and, in particular, the mucin fraction of mucus, modulated adherence to intact corneal epithelium of Pseudomonas aeruginosa but not that of Staphylococcus aureus or Streptococcus pyogenes. Removal of endogenous mucus from rat or rabbit eyes increased the adherence of P. aeruginosa by 3- to 10-fold. Ocular mucus obtained from rat eyes, porcine stomach mucin, or bovine submaxillary gland mucin inhibited adherence of P. aeruginosa to uninjured corneal epithelium. The mucin fraction of ocular mucus, purified by ultracentrifugation, was found to contain the inhibitory activity, and inhibition was demonstrated at concentrations of mucin as low as 35 micrograms/ml. Ocular mucin was the only material tested that inhibited adherence of P. aeruginosa to an injured cornea. However, the binding of P. aeruginosa to immobilized substrates in vitro did not predict which fraction would possess antiadherence activity: bacteria bound well to whole ocular mucus, mucin, the nonmucin fraction of ocular mucus, and dilute human tears as well as to porcine stomach mucin and bovine submaxillary gland mucin. The effectiveness of the mucin fraction of ocular mucus at inhibiting the binding of P. aeruginosa to the cornea implies that this material is a barrier that protects the surface of the eye from P. aeruginosa adherence.

Animals↗

Helicobacter pylori infection does not reduce the viscosity of human gastric mucus gel.

The mechanism by which Helicobacter pylori undermines host defence mechanisms is unclear. Several in vitro studies using soluble mucins have suggested that H pylori may compromise mucus function. Gastric mucus gel was obtained from 13 H pylori infected patients; six untreated subjects and seven after eradication of the infection. Gastric mucus is a non-Newtonian substance in that its viscosity changes with changing rates of shear, requiring mucus viscosity to be measured in a rotational cone-plate microviscometer. Viscosity was measured at shear rates varying from 1.15 s-1 to 46 s-1. The gastric mucus viscosity was significantly higher in patients infected with H pylori compared with mucus gel obtained after eradication of the infection. The results of our study suggest that the previous studies using in vitro methods involving soluble mucins or its components may have lead to erroneous conclusions about the in vivo interactions of H pylori and gastric mucus gel. The present findings argue against the hypothesis that degradation of gastric mucus by H pylori is important in the pathogenesis of peptic ulcer.

Female↗

Adhesion of probiotic lactobacilli to chick intestinal mucus.

In the present work, interactions between three Lactobacillus strains (Lactobacillus fermentum CRL1015, Lactobacillus animalis CRL1014, and Lactobacillus fermentum CRL1016) and chicken small intestinal mucus were determined. Three lactobacilli isolated from chicken and selected by their potentially probiotic properties were able to grow in mucus preparations. Three peaks from gel filtration chromatography of intestinal mucus were obtained. The adhesion to three mucus fractions (I, II, and III), especially fraction III, was higher (P < 0.01) in L. fermentum CRL1015 than L. animalis CRL1014. Pretreatment of this fraction with proteases and metaperiodate showed lower (P < 0.01) adhesion values than that of the control, suggesting that a glycoprotein from the mucus acts as a receptor for L. fermentum CRL1015. Highest adhesion values were obtained at pH 7 and 42 degrees C, and neither the removal of divalent cations with ethylenediaminetetraacetic acid (EDTA) nor the addition of calcium produced significant variation from the adhesion values of the control (P > 0.01). This adhesion was only inhibited by N-acetyl-glucosamine. Salmonella pullorum and Salmonella gallinarum showed high (P < 0.01) values of adhesion to chick intestinal mucus. The results obtained from assays of the inhibition of adherence of Salmonella spp. to mucus, immobilized in polystyrene tissue culture wells, indicated that the pathogen adhesion was not reduced by lactobacilli (P > 0.05) or their spent culture supernatants (P > 0.05), suggesting that these strains did not interfere with the binding sites for Salmonella spp. adhesion to the small intestinal mucus.

Animals↗

CD4(+) T cell-dependent airway mucus production occurs in response to IL-5 expression in lung.

The potential role of airway interleukin-5 (IL-5) expression in eliciting mucus production was demonstrated in a pulmonary IL-5 transgenic mouse model (NJ.1726) in which naive transgenic mice display comparable levels of airway mucus relative to allergen-sensitized and -challenged wild-type mice. The intrinsic mucus accumulation of NJ.1726 was abolished in compound transgenic-gene knockout mice deficient of either CD4(+) cells [NJ.1726/CD4(-/-)] or alphabeta T cell receptor-positive (TCR(+)) cells [NJ.1726/alphabeta TCR(-/-)]. In addition, mucus production in naive NJ.1726 was inhibited by >90% after administration of the soluble anti-IL-4 receptor alpha-subunit antagonist. The loss of mucus production in NJ.1726/CD4(-/-), NJ.1726/alphabeta TCR(-/-), and anti-IL-4 receptor alpha-subunit antagonist-treated mice occurred notwithstanding the significant pulmonary eosinophilia and expansion of airway B cells induced by ectopic IL-5 expression. Furthermore, the loss of mucus accumulation occurred in these mice despite elevated levels of airway and peripheral IL-5, indicating that IL-5 does not directly induce goblet cell metaplasia and mucus production. Thus pulmonary expression of IL-5 alone is capable of inducing CD4(+) T cell-dependent goblet cell metaplasia, apparently mediated by IL-4 receptor alpha-subunit-ligand interactions, and represents a previously unrecognized novel pathway for augmenting allergen-induced mucus production.

Allergens↗

Effect of pentobarbital anesthesia on rheology and transport of canine tracheal mucus.

Tracheal mucus samples were collected from six tracheostomized dogs before and after induction of pentobarbital anesthesia. The dynamic mechanical properties of the mucus were determined by the magnetic microrheometer technique. In 15 experiments, the mean value of elasticity at 1 rad/s rose from 67 dyn/cm2 before anesthesia to 212 dyn/cm2 in the period 15-90 min after induction. At the same time, mean viscosity at 1 rad/s rose from 24 to 79 P. The mucociliary transportability of some of the mucus samples was assayed by means of in vitro frog palate technique. In 10 experiments, the relative transport rate for dog tracheal mucus went from 98% of frog palate control befor anesthesia to 79% during pentobarbital anesthesia. A negative relationship between from palate transport rate and mucus elasticity was found. A negative correlation between transport and mucus viscosity at constant elasticity was also discovered. Finally, the mucus collection rate was reduced by a factor of about 8 during anesthesia, suggesting a reduction in the average depth of mucus.

Animals↗

Mucus-depleted frog palate as a model for the study of mucociliary clearance.

To better understand the frog palate model of mucociliary transport, we measured the transport rate of mucus (MTR) from the leopard frog, Rana pipiens, and from the bullfrog, R. catesbeiana, recorded the stability of the MTR over a period of hours and days and over the course of 1 yr, and measured the viscoelasticity, percent solid composition, and spinnability (filance) of mucus from both species. Bullfrog mucus was less rigid than leopard frog mucus (log G* at 1 rad/s 2.09 vs. 2.61; P less than 0.01) and had a higher viscosity-to-elasticity ratio (tan delta at 1 rad/s 0.36 vs. 0.26; P less than 0.05). It also had a lower solids content (8.71 vs. 13.72%; P = 0.02), and there was a trend to lower spinnability for bullfrog mucus (filance 26.7 vs. 33.5 mm). These data suggest that bullfrog mucus has viscoelastic properties similar to normal mammalian respiratory mucus and leopard frog mucus has viscoelasticity similar to sputum samples. MTR was significantly slower in the winter than in the summer months (17 vs. 30 mm/min; P less than 0.0001). Although the leopard frog palate could be used for at least 7 consecutive days without exhaustion, bullfrog palates could be used for only 5 days. Palates of either species could generally be tested for 6 h/day without a significant decrease in MTR. These data clarify some of the sources of variability in the use of this system and suggest methods of standardization.

Animals↗

Work of adhesion of respiratory tract mucus.

A method was devised to measure the work of adhesion (WA) to a substrate of mucus, a viscoelastic gel, from the measured contact angle of glycerol on a mucus substrate and the known physical properties of a Teflon surface. Fifteen sputum samples from cystic fibrosis (CF) patients were compared with 25 mucus samples from canine tracheal pouches (CP), studied in the hydrated and partially dehydrated states. Apparent viscosity (eta A) and recoverable shear strain (SR) were measured by fluxgate magnetometry, and water content was inferred from vapor pressure osmometry. Na+, K+, and Ca2+ concentrations were measured with specific ion electrodes and Cl- with a chloridimeter. The Cl- concentration of the CP mucus was inversely proportional to its osmolality, and the Cl- concentration of the CP mucus was 102.5 +/- 1.6 meq/l compared with 55.6 +/- 2.5 meq/l for CF sputum. When CP mucus osmolality was increased from 316.0 +/- 5.5 to 430.0 +/- 7.5 mosmol/kg, WA increased from 25.1 +/- 1.8 to 31.1 +/- 1.2 ergs/cm2 and eta A increased from 391 +/- 55 to 622 +/- 121 P, respectively. CF sputum WA was 30.2 +/- 0.6 ergs/cm2, eta A was 1,110 +/- 316 P, and osmolality was 466.0 +/- 14.0 mosmol/kg. The increased WA and eta A of mucus in CF patients may thus be dependent on the hydration of mucus, which is related to the documented Cl- transport defect.

Adolescent↗

Effects of verapamil, carbenoxolone and N-acetylcysteine on gastric wall mucus and ulceration in stressed rats.

The effects of verapamil on gastric wall mucus and ulceration were studied in rats which were restrained and exposed to 4 degrees C (stress). Stress for 2 h significantly depleted stomach wall mucus and produced marked gastric glandular ulcers. Verapamil pretreatment (2, 4, 8 or 16 mg/kg), injected intraperitoneally 30 min before experimentation, significantly prevented stress-induced mucus depletion and gastric ulceration; however, it did not itself influence stomach wall mucus levels in nonstressed animals. Intragastric administration of carbenoxolone (100 or 200 mg/kg), also given 30 min before stress, exhibited similar actions as verapamil. A 15% solution of N-acetylcysteine (10 ml/kg), given orally, strongly decreased the mucus content in both nonstress and stress conditions; it induced ulcers in nonstressed rats, and worsened stress ulceration. These effects were not reversed by verapamil pretreatment. The influence of multiple-dose pretreatment with verapamil or carbenoxolone on mucus content and ulceration in the gastric glandular mucosa during stress is also discussed. It is concluded that gastric wall mucus depletion is likely to play an important role in stress ulcer formation; the antiulcer action of verapamil could partly be due to the preservation of mucus.

Acetylcysteine↗

Mucus glycoprotein content of human cholesterol gallstones.

Cholesterol gallstones were obtained from patients undergoing cholecystectomy and the mucus glycoprotein extracted. The biliary mucus glycoprotein was separated from other contaminants by Sepharose 4B gel filtration and the PAS staining excluded volume used to estimate mucus glycoprotein content of the gallstones. Hexosamine and sialic acid analysis of the glycoprotein indicated it was compositionally similar to the human mucus glycoprotein from bile. The mucus glycoprotein content of the nine stones analysed individually varied between 0.75 and 2.3 mg (a 3-fold variation) (1.27 +/- 0.16 mg, mean +/- SEM), whereas stone weight varied between 0.076 and 5.885 g (a 77-fold variation) (1.27 +/- 0.63 g, mean +/- SEM). When a pool of smaller stones, average weight 47 mg, was extracted, only 1.73 mg of glycoprotein was isolated, an average of 0.01 mg/stone. Analysis of the mucus glycoprotein by gel filtration on Sepharose 2B showed the majority of the glycoprotein was excluded as is the case with the mucus glycoprotein in bile. These results are consistent with biliary mucus glycoprotein being involved in the initial stages of gallstone formation but not in subsequent growth.

Cholelithiasis↗

Improvement of cystic fibrosis airway mucus transportability by recombinant human DNase is related to changes in phospholipid profile.

The aim of this study was to test whether changes in mucus surface properties by rhDNase treatment could be related to an increased recovery of phospholipids. Purulent sputa from 18 patients with cystic fibrosis (CF) were incubated with either rhDNase (4 microg/ml) or control excipient. The incubation of mucus samples with rhDNase induced a significant increase (p < 0.002) in the sol phase proportion (33.7 +/- 24.0%) compared with that obtained with excipient (12.6 +/- 12.4%). Phospholipids were recovered in significantly (p < 0.05) greater amounts from both mucus gel and sol phases after incubation with rhDNase. The phosphatidylglycerol content of mucus sol phase was significantly increased by rhDNase (p < 0.03), as well as the mucus gel phase surface properties and transport by ciliary activity and by cough (p < 0.05). The improvement of mucus gel surface properties and transport capacity by ciliary activity were significantly related to the increased recovery of phosphatidylglycerol (r = -0.74, p < 0.03 and r = 0.94, p < 0.05, respectively). These results suggest that rhDNase is able to increase the free water content and alter the phospholipid profile of mucus, with a related improvement in CF mucus transportability.

Adult↗