Antiadhesive property of microalgal polysaccharide extract on the binding of Helicobacter pylori to gastric mucin

The emergence of antibiotic-resistant Helicobacter pylori is of concern in the treatment of H. pylori-associated gastroduodenal diseases. As the organism was reported to bind gastric mucin, we used porcine gastric mucin as substrate to assess the antiadhesive property of polysaccharides derived from Spirulina (PS), a commercially available microalga, against the binding of H. pylori to gastric mucin. Results show that polysaccharides prevented H. pylori from binding to gastric mucin optimally at pH 2.0, without affecting the viability of either bacteria or gastric epithelial cells, thus favouring its antiadhesive action in a gastric environment. Using ligand overlay analysis, polysaccharide was demonstrated to bind H. pylori alkyl hydroperoxide reductase (AhpC) and urease, which have shown here to possess mucin-binding activity. An in vivo study demonstrated that bacteria load was reduced by >90% in BALB/c mice treated with either Spirulina or polysaccharides. It is thus suggested that polysaccharides may function as a potential antiadhesive agent against H. pylori colonization of gastric mucin.     

Folding of pig gastric mucin non-glycosylated domains: a discrete molecular dynamics study

Mucin glycoproteins consist of tandem-repeating glycosylated regions flanked by non-repetitive protein domains with little glycosylation. These non-repetitive domains are involved in polymerization of mucin and play an important role in the pH-dependent gelation of gastric mucin, which is essential for protecting the stomach from autodigestion. We examine folding of the non-repetitive sequence of PGM-2X (242 amino acids) and the von Willebrand factor vWF-C1 domain (67 amino acids) at neutral and low pH using discrete molecular dynamics (DMD) in an implicit solvent combined with a four-bead peptide model. Using the same implicit solvent parameters, folding of both domains is simulated at neutral and low pH. In contrast to vWF-C1, PGM-2X folding is strongly affected by pH as indicated by changes in the contact order, radius of gyration, free-energy landscape, and the secondary structure. Whereas the free-energy landscape of vWF-C1 shows a single minimum at both neutral and low pH, the free-energy landscape of PGM-2X is characterized by multiple minima that are more numerous and shallower at low pH. Detailed structural analysis shows that PGM-2X partially unfolds at low pH. This partial unfolding is facilitated by the C-terminal region GLU236-PRO242, which loses contact with the rest of the domain due to effective “mean-field” repulsion among highly positively charged N- and C-terminal regions. Consequently, at low pH, hydrophobic amino acids are more exposed to the solvent. In vWF-C1, low pH induces some structural changes, including an increased exposure of CYS at position 67, but these changes are small compared to those found in PGM-2X. For PGM-2X, the DMD-derived average β-strand propensity increases from 0.26 ± 0.01 at neutral pH to 0.38 ± 0.01 at low pH. For vWF-C1, the DMD-derived average β-strand propensity is 0.32 ± 0.02 at neutral pH and 0.35 ± 0.02 at low pH. The DMD-derived structural information provides insight into pH-induced changes in the folding of two distinct mucin domains and suggests plausible mechanisms of the aggregation/gelation of mucin.     

Establishment of monoclonal antibodies against carbohydrate moiety of gastric mucins distributed in the different sites and layers of rat gastric mucosa

Eight monoclonal antibodies (MAbs), designated RGM21 RGM42, were generated against mucin purified from the rat gastric mucosa. By applying ELISA, all of these MAbs were proved to react not only with the purified mucin, but also with the oligosaccharide mixture obtained from the antigenic mucin by alkaline borohydride treatment. Treatment of the mucin-attached ELISA well with trypsin, sodium periodate or galactose oxidase prior to the addition of the MAb was applied to characterize these MAbs. Histochemical observation indicated that all these MAbs were able to stain the formalin fixed-paraffin embedded sections of the rat gastroduodenal mucosa. Although each of these MAbs reacted with distinct mucus-producing cells localized in particular regions of the gastroduodenal mucosa, their staining specificity could generally be classified into four groups. These MAbs might be useful for estimating the physiological and pathological changes of mucins in the gastric mucosa.     

pH Effect of the Sphingomyelin Membrane Interfacial Tension

The effect of pH on the interfacial tension of a sphingomyelin membrane in aqueous solution has been studied. Three models describing H⁺ and OH⁻ ion adsorption on the bilayer lipid surface are presented. In models I and II, the membrane surface is continuous, with uniformly distributed functional groups as centers of H⁺ and OH⁻ ion adsorption. In model III, the membrane surface is composed of lipid molecules, with and without adsorbed H⁺ and OH⁻ ions. The contribution of each individual lipid molecule to the overall interfacial tension of the bilayer was assumed to be additive in models I and II. In model III, the Gibbs isotherm was used to describe adsorption of H⁺ and OH⁻ ions at the bilayer surface. Theoretical equations are derived to describe the interfacial tension as a function of pH for all three models. Maximum interfacial tension was observed experimentally at the isoelectric point.     

Strain-dependent proliferation in response to human gastric mucin and adhesion properties of Helicobacter pylori are not affected by co-isolated Lactobacillus sp

Background: Helicobacter pylori colonize the mucus layer that covers the gastric epithelium and can cause gastritis, ulcers, and gastric cancer. Recently, Lactobacillus sp. have also been found to reside in this niche permanently. This study compares adhesive properties and proliferation of co‐isolated lactobacilli and H. pylori in the presence of mucins and investigates possibilities for lactobacilli‐mediated inhibition of H. pylori. Materials and methods: Binding and proliferation of four H. pylori and four Lactobacillus strains, simultaneously isolated after residing in the stomachs of four patients for >4 years, to human gastric mucins were investigated using microtiter‐based methods. Results: The H. pylori strains co‐isolated with lactobacilli exhibited the same mucin binding properties as demonstrated for H. pylori strains previously. In contrast, no binding to mucins was detected with the Lactobacillus strains. Proliferation of mucin‐binding H. pylori strains was stimulated by the presence of mucins, whereas proliferation of non‐binding H. pylori and Lactobacillus strains was unaffected. Associative cultures of co‐isolated H. pylori and Lactobacillus strains showed no inhibition of H. pylori proliferation because of the presence of whole bacteria or supernatant of lactobacilli. Conclusions: The presence of lactobacilli in the stomach did not select for different mucin binding properties of H. pylori, and Lactobacillus sp. did neither compete for binding sites nor inhibit the growth of co‐isolated H. pylori. The effects of human gastric mucins on H. pylori proliferation vary between strains, and the host–bacteria interaction in the mucus niche thus depends on both the H. pylori strain and the microenvironment provided by the host mucins.     

Phase behavior of aqueous gelatin/oligosaccharide mixtures

The phase behavior of aqueous mixtures of gelatin and oligosaccharides above their gelation temperature is investigated experimentally, and rationalized according to a simple multicomponent Flory-Huggins model. When the gelatin is only weakly charged, entropic considerations dominate and it is found that the cloud point curve of the mixtures is extremely sensitive to the molecular weight distribution of the oligosaccharide. Even very small quantities of long-chain oligosaccharides present in an otherwise short-chain oligosaccharide population can radically reduce the compatibility. Added salt does not significantly affect the phase diagram, although a strong effect on the kinetics of phase separation is seen. Lowering the pH increases the electrostatic charge on the gelatin and strongly enhances the compatibility. Because the kinetics of gelation and phase separation are different, gelation can freeze in nonequilibrium states. Therefore, all phase diagrams were determined well above the gelation temperature (about 37°C). © 1997 John Wiley & Sons, Inc. Biopoly 41: 607–622, 1997     

Effect of pH on the phase behavior of DMPC bilayers

We have studied the effect of acidic pH on the phase behavior of the zwitterionic lipid 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) using differential scanning calorimetry and x-ray scattering. Dispersions of DMPC in HCl solutions of pH = 4 and 3 behave identical to dispersions in water. The main transition temperature increases sharply and the pre-transition disappears at lower pH. An untilted gel phase is observed at pH = 2 and 1, in contrast to the tilted gel phase found at higher pH. The relatively large periodicity of the untilted gel phase, in comparison to that of the tilted gel phase occurring near neutral pH, clearly demonstrates the simultaneous charging and dehydration of the headgroups as the pH approaches the pK of the phosphate group. Headgroup dehydration at low pH also leads to the formation of DMPC crystallites and the inverted hexagonal phase at low and high temperatures, respectively, after a few days of incubation. These results show the significant effect of acidic pH on the phase behavior of zwitterionic lipids.     

PFG-NMR diffusometry: a tool for investigating the structure and dynamics of noncommercial purified pig gastric mucin in a wide range of concentrations

For the first time, Pulsed Field Gradient-Nuclear Magnetic Resonance, a powerful noninvasive tool for studying the dynamics and structure of complex gels, has been used to measure diffusion of probe molecules in aqueous solutions/gels of noncommercial purified pig gastric mucin (PGM), in a concentration range up to 5 wt %. Complementary data were obtained from rheology measurements. The combination of techniques revealed a strong pH dependency of the structure of the PGM samples while changes in concentration, ionic strength, and temperature appeared to induce less pronounced alterations. Viscosity was found to vary in a nonmonotonous way with pH, with the more viscous solutions found at intermediate pH. We propose that this finding is due to a reduced charge density at lower pH, which is expected to continuously increase the relative importance of hydrophobic associations. The results suggest a loose network of expanded fully charged PGM molecules with considerable mobility at neutral pH (pH 7.4). At intermediate pH (pH 4), a three-dimensional expanded network is favored. At pH 1, the charge density is low and microphase separation occurs since hydrophobic associations prevail. This leads to the formation of clusters concentrated in PGM molecules separated by regions depleted in PGM. The results obtained increase our knowledge about the gastric mucosal layer, which in vivo contains mucin in the same concentration range as that of the samples investigated here.     

Effect of pH on the interfacial tension of bilayer lipid membrane formed from phosphatidylcholine or phosphatidylserine

The effect of pH of an electrolyte solution on the interfacial tension of lipid membrane formed from phosphatidylcholine (PC) or phosphatidylserine (PS) was studied. The relationships were well described by an equation presented earlier based on the Gibbs isotherm but only in the proximity of the isoelectric point. Therefore, in this work models have been derived to describe the adsorption of the H⁺ and OH⁻ ions at lipid surfaces formed from PC or PS, which would reproduce changes in interfacial tension more correctly, particularly in the ranges distant from the isoelectric point. In one model, the surface is continuous with uniformly distributed functional groups constituting the centres of H⁺ and OH⁻ ion adsorption while in the other the surface is built of lipid molecules, free or with attached H⁺ and OH⁻ ions. In both models, the contributions of the individual lipid molecule forms to the interfacial tension of the bilayer were assumed to be additive.     

Microstructure and aggregation behavior of methylcelluloses prepared in NaOH/urea aqueous solutions

Three water-soluble methylcelluloses (MCs) were prepared through homogeneous reaction in NaOH/urea aqueous solution, using dimethyl sulfate as a methylation reagent. The microstructure of the MC samples was characterized by IR, GC/MS, NMR, while dilute solution properties were measured by SEC–LLS, DLS and viscometer. The total degrees of substitution (DS) of the MC samples were 1.09, 1.42 and 1.56, respectively. However, we found that the relative DS value varies with the position of the hydroxyl group, i.e., C-2 > C-3 ≈ C-6, indicating the difference of reaction activity of different hydroxyl groups. In aqueous solution, MC has a trendency to form aggregates and hard to form actual solution, even at low concentration and low temperature, which was confirmed by the SEC–LLS and DLS result that isolated MC chains and large aggregates coexisted in the dilute aqueous solution. MC aqueous solutions showed two-stage temperature dependence of hydrodynamic radius. In the first stage, i.e., the temperature ranges from 20 to 65 °C, the hydrodynamic radius of MC displayed bimodal distribution, corresponding to the single chains and large aggregates. While in the second stage, i.e., the temperature higher than 70 °C, only large aggregates appeared. The results also proved that the microstructure of MC had a great influence on its physical properties.     

Role of the pH on hyaluronan behavior in aqueous solution

In this paper, we have examined the behavior of hyaluronan solutions at different pH values. A slight degradation is observed in acidic conditions (pH = 1.6) and basic medium (pH = 12.6) from molecular weight distribution analysis, but the rheological behavior is relatively not influenced much by the pH at the exclusion of two domains: around pH = 2.5, a gel-like behavior is shown and is attributed to cooperative interchain interactions due to the reduction of the polymer net charge and may be the protonation of the acetamido groups; for pH > 12, the decrease of viscosity is mainly attributed to a reduction of the stiffness of the polymeric backbone in alkaline conditions due to the partial breakage of the H-bond network.     

pH对不同富集能力植物根际土壤溶液中镉形态的影响

为了解不同p H对土壤溶液中Cd形态分布的影响,采用Visual MINTEQ 3.0模型和阳极溶出伏安法(ASV)研究土壤p H分别为4.0、5.5、7.0和8.5时镉超富集植物东南景天和非富集植物垂盆草根际土壤溶液中Cd的形态分布。模型计算结果表明:随着p H的升高,2种植物土壤溶液中Cd2+的比例均逐渐减小,其中东南景天根际土壤溶液中Cd2+占总溶解性Cd的比例为46.1%~3.2%,垂盆草为60.9%~9.9%;当土壤p H≥5.5时,土壤溶液中Cd形态主要以有机络合态(Cd-DOM)为主;2种植物相比,4个p H水平下垂盆草土壤溶液中Cd2+比例均高于东南景天,但东南景天中有机络合态Cd所占比例较大。基于ASV法的测定结果表明:东南景天土壤溶液中电活性Cd占总溶解性Cd的比例从最小(p H 8.5时)的5.5%到最大(p H 4.0时)的79.6%,而垂盆草的变化范围为11.8%~86.7%;电活性Cd的测定结果与Visual MINTEQ 3.0模型计算结果显著相关(R2=0.80),从而互相验证了实验测量和模拟计算的可靠性。     

Light-induced pH and K+ changes in the apoplast of intact leaves

The K⁺-sensitive fluorescent dye benzofuran isophthalate (PBFI) and the pH-sensitive fluorescein isothiocyanate dextran (FITC-Dextran) were used to investigate the influence of light/dark transitions on apoplastic pH and K⁺ concentration in intact leaves of Vicia faba L. with fluorescence ratio imaging microscopy. Illumination by red light led to an acidification in the leaf apoplast due to light-induced H⁺ extrusion. Similar apoplastic pH responses were found on adaxial and abaxial sides of leaves after light/dark transition. Stomatal opening resulted only in a slight pH decrease (0.2 units) in the leaf apoplast. Gradients of apoplastic pH exist in the leaf apoplast, being about 0.5–1.0 units lower in the center of the xylem veins as compared with surrounding cells. The apoplastic K⁺ concentration in intact leaves declined during the light period. A steeper light-induced decrease in apoplastic K⁺, possibly caused by higher apoplastic K⁺, was found on the abaxial side of leaves concentration. Simultaneous measurements of apoplastic pH and K⁺ demonstrated that a light-induced decline in apoplastic K⁺ concentration indicative of net K⁺ uptake into leaf cells occurs independent of apoplastic pH changes. It is suggested that the driving force that is generated by H⁺ extrusion into the leaf apoplast due to H⁺-ATPase activity is sufficient for passive K⁺ influx into the leaf cells. Received: 7 March 2000 / Accepted: 12 May 2000     

Dynamic and structural scalings of the complexation between pDNA and bPEI in semidilute and low‐salt solutions

Using a combination of static and dynamic laser light scattering, we investigated the complexation of a supercoiled plasmid DNA (pDNA, 10⁴ bp) and a branched polyethyleneimine (bPEI, M_w = 25 kD) in semidilute and low‐salt aqueous solutions. Our results unearth some scaling laws for dynamic and structural properties of the resultant complexes (polyplexes) with different bPEI:pDNA (N:P) molar ratios. Namely, the average scattering intensity (<I>) and the average linewidth of the Rayleigh peak (<Γ>) are scaled to the scattering vector (q) as <I> ∝ q or <Γ> ∝ q, where α_S and α_D are two N:P dependent scaling exponents. The N:P ratio strongly affects the complexation. When N:P < 2.0, the motions of the negatively charged and extended pDNA chains and the polyplexes are highly correlated so that they behave like a transient network with a fractal dimension. As the N:P ratio increases, nearly all of pDNA chains condensed and the overall charge of the polyplexes reverses to slightly positive, resulting in a turbid dispersion of large loose aggregates made of smaller, but more compact, polyplexes. Further increase of N:P finally disrupts large loose aggregates, leading to a homogeneous transparent dispersion of the polyplexes. © 2010 Wiley Periodicals, Inc. Biopolymers 93: 571–577, 2010. This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com     

X-ray and neutron scattering studies of the hydration structure of alkali ions in concentrated aqueous solutions

The presence of ions in water provides a rich and varied environment in which many natural processes occur with important consequences in biology, geology and chemistry. This article will focus on the structural properties of ions in water and it will be shown how the 'difference' methods of neutron diffraction with isotopic substitution (NDIS) and anomalous X-ray diffraction (AXD) can be used to obtain direct information regarding the radial pair distribution functions of many cations and anions in solution. This information can subsequently be used to calculate coordination numbers and to determine ion-water conformation in great detail. As well as enabling comparisons to be made amongst ions in particular groups in the periodic table, such information can also be contrasted with results provided by molecular dynamics (MD) simulation techniques. To illustrate the power of these 'difference' methods, reference will be made to the alkali group of ions, all of which have been successfully investigated by the above methods, with the exception of the radioactive element francium. Additional comments will be made on how NDIS measurements are currently being combined with MD simulations to determine the structure around complex ions and molecules, many of which are common in biological systems.     

Current information on the association of Helicobacter pylori with autophagy and gastric cancer

Helicobacter pylori (H. pylori) is a Gram-negative bacterium and causative agent of gastric cancer. H. pylori induce defective autophagy or inhibit it by means of CagA and vacuolating cytotoxin A (VacA) toxins leading to the gastric cancer induction. Impaired or defective autophagy leads to the accumulation of cytotoxic materials, such as ROS and P62 that lead to increased mutations in the DNA, genome instability, and risk of cancer formation. H. pylori CagA may inhibit autophagy through the c-Met-PI3k/Akt-mTOR signaling pathway. However, VacA induces autophagy by some signaling pathways. In the gastric epithelial cells, VacA is a necessary and sufficient factor for the creation of autophagy. While CagA is a negative regulator of this phenomenon, the elimination of this gene from H. pylori has increased autophagy and the production of inflammatory cytokines is reduced. In gastrointestinal cancers, some of the microRNAs (miRNAs) act as tumor suppressors and some other are oncogenes by regulating various genes expression. H. pylori can also modify autophagy through a mechanism that includes the function of miRNAs. In autophagy, oncogenic miRNAs inhibit activation of some tumor suppressor signaling pathways (e.g., ULK1 complex, Beclin-1 function, and Atg4 messaging), whereas tumor suppressor miRNAs can block the activation of oncogenic signaling pathways. For instance, Beclin-1 is negatively regulated by miRNA-376b (oncogenic miRNA) and miRNA-30a (tumor suppressor miRNA). Similarly, Atg4 by miRNA-376b (oncogenic miRNA) and miRNA-101 (tumor suppressor miRNA). So, this apparent paradox can be explained as that both Beclin-1 and Atg4 play different roles in a particular cell or tissue.