Biomedical subjects

Bernd Stahl

Publications and source records attributed to Bernd Stahl.

17 recordsLinked to original sources

PubMed2006

A specific prebiotic oligosaccharide mixture stimulates delayed-type hypersensitivity in a murine influenza vaccination model.

Analogous to reported immunomodulatory effects of probiotics, this study was performed to analyse the immunomodulatory properties of prebiotic oligosaccharides that share chemical characteristics with human milk oligosaccharides. A mixture containing galacto- and fructo-oligosaccharides (GOS/FOS; ratio 9:1) was tested at dietary doses between 1% and 10% (w/w of total diet) in an influenza vaccination model, using 10 C56BL/6JolaHsd mice per group. The modulation of vaccine specific delayed-type hypersensitivity (DTH) responses was studied as a marker of T-helper 1 (Th1) immunity, as well as other immune parameters. GOS/FOS enhanced DTH responses dose-dependently (optimum at 5% w/w of total diet; 41.4+/-14.1% increased compared to controls, p<0.05). No significant changes were detected on splenocyte proliferation or vaccine-specific antibody concentrations. Simultaneously, GOS/FOS dose-dependently increased the proportion of faecal bifidobacteria and lactobacilli (maximal effect at 10% w/w of total diet; 16.8+/-2.4% and 5.8+/-1.3% increased compared to controls respectively, p<0.01 for both parameters). In a comparative experiment, GOS/FOS and FOS/inulin (both at 2% w/w of total diet) induced similar significant effects on the gut microbiota. In contrast to GOS/FOS, FOS/inulin did not enhance DTH responses, indicating that an increase in the proportions of bifidobacteria and lactobacilli is not sufficient for an immunomodulatory effect in this model. The use of GOS/FOS in dietary products might provide an opportunity to stimulate the adaptive immune response in a Th1-direction and subsequently inhibit infections and Th2-related immune disorders in humans, for instance allergies. Clinical studies are being performed to confirm this.

Animals↗

PubMed2006

Human milk oligosaccharides affect P-selectin binding capacities: in vitro investigation.

OBJECTIVE: In the initial phase of cellular immune response, selectins mediate the emigration of leukocytes from the blood stream into inflammatory regions. Human milk oligosaccharides (HMOs) possess binding epitopes of selectin ligands such as sialyl Lewis(x) and sialyl Lewis(a) and therefore might impair the interaction of selectins with cellular ligands. Neutral, acidic, sialylated, or fucosylated HMO fractions with polymerization degrees of 3 to 50 were investigated regarding this interaction in a dynamic flow chamber model that considers physiologic shear stress conditions. METHODS: Human milk oligosaccharides were compared with kappa-carrageenans and pectin oligosaccharides to deduce structure-activity relations. Fucoidan and sialyl Lewis(x) served as positive controls. RESULTS: All HMO fractions affected P-selectin ligand binding capacity but were not comparable to fucoidan. The activity of the acidic HMO fraction resembled sialyl Lewis(x) in decreasing the binding of the ligand to P-selectin. CONCLUSION: Human milk oligosaccharides modulate rather than block the function of P-selectin.

Binding Sites↗

PubMed2005

Separation of lactose from human milk oligosaccharides with simulated moving bed chromatography.

The successful separation of the disaccharide lactose from a complex mixture of human milk oligosaccharides (HMOS) with the continuous chromatography of simulated moving bed (SMB) technique is described. Since lactose is the main carbohydrate in human milk with well-known functions for the infant, it is necessary to separate it from the rest of the oligosaccharides to divide them into less complex fractions and analyse their partial unknown functions. For separation of lactose from HMOS two different stationary phases (size-exclusion gel as well as ion-exchange gel) were used. As the main result, it is shown that a size-exclusion gel with the particle size of 50-100 microm and porosity of 50 A was the preferred stationary phase for our separation process with almost complete lactose separation and stable conditions.

Chromatography, Gel↗

PubMed2005

Prebiotic carbohydrates in human milk and formulas.

Human milk oligosaccharides play an important role, as prebiotic soluble fibres, in the postnatal development of the intestinal flora. Infant formulas are virtually free of prebiotic oligosaccharides. As a consequence, formula-fed infants develop an intestinal flora significantly different to the flora of breastfed infants. Due to the complexity of human milk oligosaccharides, it is necessary to use alternative sources of prebiotic ingredients as components of infant formulas. The present review summarizes the data of experimental research and clinical studies with a prebiotic mixture containing 90% short-chain galacto-oligosaccharides and 10% long-chain fructo-oligosacchrides are summarized. The data demonstrate that, with this prebiotic mixture, the growth of bifidobacteria and lactobacilli can be stimulated, the faecal pH can be decreased, and the presence of pathogens can be reduced to levels similar to those of breastfed infants. Thus, prebiotic oligosaccharides such as the studied mixture provide beneficial effects for formula-fed infants.

Breast Feeding↗

PubMed2005

Galacto-oligosaccharides and long-chain fructo-oligosaccharides as prebiotics in infant formulas: a review.

UNLABELLED: The present review summarizes clinical and experimental data concerning the possible effects of a prebiotic mixture of short-chain galacto-oligosaccharides and long-chain fructo-oligosaccharides. The results from several studies, made up of over 400 preterm and term infants, clearly demonstrate that the prebiotic mixture under examination specifically stimulates the growth of bifidobacteria and lactobacilli and reduces the growth of pathogens. As a consequence of the changed intestinal flora by the dietary galacto-oligosaccharides and fructo-oligosaccharides, the faecal pH values and the short-chain fatty acid pattern were similar to those found in breastfed infants. In addition, the stool consistency was the same as in breastfed infants. In vitro experiments have demonstrated that the specific short-chain fatty acid pattern, at a pH similar to that found in faecal samples of breastfed infants, reduces the growth of pathogens in a dose-dependent manner but does not influence the growth of bifidobacteria and lactobacilli. In an animal vaccination model, the prebiotic mixture improved the response to vaccination. In an allergy model (sensitization by ovalbumin), the allergic reaction was reduced by the prebiotic mixture. The data obtained from animal experiments are in agreement with preliminary data from clinical trials which indicate a reduced allergic response (reduced plasma IgE/IgG4 ratio) and reduced episodes of upper airway infection during the first year of life. CONCLUSION: Experimental evidence demonstrates that the prebiotic mixture employed in these studies modulates the intestinal flora and modulates the immune system as human milk does. There are sufficient experimental data to put forward the hypothesis that substances like the prebiotic mixture under study will substantially contribute to the improvement of the protective properties of infant formulas.

Feces↗

PubMed2005

Dietary prebiotic oligosaccharides are detectable in the faeces of formula-fed infants.

UNLABELLED: Human milk oligosaccharides are not digested during intestinal passage and can be detected in stools. In this study it was investigated whether a prebiotic mixture of low-molecular-weight galacto-oligosaccharides (GOS) and high-molecular-weight fructo-oligosaccharides (FOS) can be detected in stool samples of formula-fed infants. The test formula was supplemented with 0.8 g/dl oligosaccharides (GOS+FOS). In the control formula, maltodextrins were used as placebo. Fecal flora was assessed at the beginning (day 1) and at the end of a 28-d feeding period (day 2). At day 2 the content of galacto- and fructo-oligosaccharides in the stool samples were measured. On study day 1, the number of bifidobacteria was not different among the groups (supplemented group: 7.7 (6.2) CFU/g; placebo group: 8.0 (6.0) CFU/g). At the end of the 28-d feeding period, the number of bifidobacteria was significantly higher in the group fed the supplemented formula when compared to placebo (supplemented group: 9.8 (0.7) CFU/g stool; placebo group: 7.1 (4.7) CFU/g stool; p<0.001). In all infants fed the supplemented formula, GOS and FOS could be identified in the stool samples. That was not the case in infants fed the non-supplemented formula. CONCLUSION: The present data confirm the bifidogenicity of oligosaccharides and indicate that dietary galacto-oligosaccharides and long chain fructo-oligosaccharides remain during the whole passage in the lumen of the gastrointestinal tract, similarly to human milk oligosaccharides.

Anthropometry↗

PubMed2005

Increase of faecal bifidobacteria due to dietary oligosaccharides induces a reduction of clinically relevant pathogen germs in the faeces of formula-fed preterm infants.

UNLABELLED: In a previous study on formula-fed preterm infants, we were able to demonstrate that dietary oligosaccharides (a mixture of 90% galacto-oligosaccharides and 10% fructo-oligosaccharides in a concentration of 1 g/dl) stimulate the growth of faecal bifidobacteria. In the present explorative analysis of this study, we focus on the effect of the dominance of bifidobacteria on the presence of clinically relevant pathogens (Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus haemolyticus, Pseudomonas aeruginosa, Enterobacter, Klebsiella, Proteus, Streptococcus group B, Clostridium difficile, Bacillus subtilis and Acinetobacter). CONCLUSION: The data demonstrate that stimulation of bifidobacteria by prebiotic oligosaccharides reduces the presence of clinically relevant pathogens in the faecal flora, indicating that prebiotic substances might have the capacity to protect against enteral infections.

Bacterial Infections↗

PubMed2004

Human milk--derived oligosaccharides and plant-derived oligosaccharides stimulate cytokine production of cord blood T-cells in vitro.

Human milk contains large amounts of free oligosaccharides (HMOs). HMOs have been shown to exert antiinflammatory properties, and evidence for their immunomodulatory effects is increasing. The purpose of this study was to evaluate influences of two human breast milk-derived oligosaccharide samples (neutral and acidic oligosaccharides), and of a low-molecular-weight fucoidan on cytokine production and activation of cord blood mononuclear cells. Cord blood mononuclear cells from randomly chosen healthy newborns were co-cultured with the oligosaccharide samples. By means of flow cytometry, intracellular cytokine production (d 20) and surface marker expression of T cells (d 5) were measured. In vitro-induced Ig levels were quantified nephelometrically (total IgG1) and by ELISA (total IgE) in the supernatant of cell cultures. The acidic oligosaccharide fraction increased the percentage of interferon-gamma producing CD3+CD4+ and CD3+CD8+ cells (p < 0.05) and the IL-13 production in CD3+CD8+ cells (p < 0.05). In acidic oligosaccharide cultures, CD25+ expression on CD3+CD4+ cells was significantly elevated (p < 0.05). Low-molecular-weight fucoidan induced IL-4 production in CD3+CD4+ T cells (p < 0.05) and IL-13 production in CD3+CD8+ T cells (p < 0.05), whereas interferon-gamma production remained unaffected in both T-cell populations. Ig production (total IgE and total IgG1) remained unaffected. Human milk-derived oligosaccharides and plant-derived oligosaccharides affect the cytokine production and activation of cord blood derived T cells in vitro. Therefore, oligosaccharides and, in particular, acidic oligosaccharides may influence lymphocyte maturation in breast-fed newborns.

B-Lymphocytes↗

PubMed2004

2,5-Dihydroxybenzoic acid butylamine and other ionic liquid matrixes for enhanced MALDI-MS analysis of biomolecules.

The performance of the new ionic liquid MALDI-MS matrix 2,5-dihydroxybenzoic acid butylamine (DHBB) was assessed and compared to results obtained with the ionic liquid MALDI-MS matrixes alpha-cyano-4-hydroxycinnamic acid butylamine (CHCAB), 3,5-dimethoxycinnamic acid triethylamine (SinTri), and the frequently used solid MALDI matrixes 2,5-dihydroxybenzoic acid (DHB) and alpha-cyano-4-hydroxycinnamic acid (CHCA). The vacuum-stable, liquid consistency of ionic liquid matrix sample preparations considerably enhanced MALDI-MS analysis in terms of shot-to-shot reproducibility. Consequently, relative standard deviations serving as a measure for reproducibility of intensity-values acquired from 90 different spots on one MALDI-MS preparation were approximately one-half as high when solid DHB was replaced by the ionic liquid DHBB and eight times lower after exchange of solid CHCA by ionic liquid CHCAB. Interestingly, the ionic liquid MALDI matrix DHBB conserved the broad applicability of its solid analogue DHB, reduced MALDI induced fragmentation of monosialylated glycans and gangliosides, and was the superior ionic liquid matrix for MALDI-MS analysis of oligosaccharides and polymers, such as poly(ethylene glycol). It also worked well with glycoconjugates, peptides, and proteins; however, the tendency of DHBB to form multiple alkali adduct ions with peptides and proteins made CHCAB the ionic liquid matrix of choice for peptides. SinTri was the best ionic liquid matrix for proteins of high molecular weight, such as IgG. Furthermore, it was demonstrated for the first time that solvent properties and MALDI matrix properties of ionic liquids, such as DHBB, can be combined to enable fast, direct screening of an enzymatic reaction. This was proven by the desialylation of sialylactose with sialidase from Clostridium perfringens in the presence of diluted aqueous DHBB and subsequent direct MALDI-MS analysis of the reaction mixture.

Butylamines↗

PubMed2004

Progressive decrease in chaperone protein levels in a mouse model of Huntington's disease and induction of stress proteins as a therapeutic approach.

The manipulation of chaperone levels has been shown to inhibit aggregation and/or rescue cell death in Saccharomyces cerevisiae, Caenorhabditis elegans, Drosophila melanogaster and cell culture models of Huntington's disease (HD) and other polyglutamine (polyQ) disorders. We show here that a progressive decrease in Hdj1, Hdj2, Hsp70, alphaSGT and betaSGT brain levels likely contributes to disease pathogenesis in the R6/2 mouse model of HD. Despite a predominantly extranuclear location, Hdj1, Hdj2, Hsc70, alphaSGT and betaSGT were found to co-localize with nuclear but not with extranuclear aggregates. Quantification of Hdj1 and alphaSGT mRNA levels showed that these do not change and therefore the decrease in protein levels may be a consequence of their sequestration to aggregates, or an increase in protein turnover, possibly as a consequence of their relocation to the nucleus. We have used genetic and pharmacological approaches to assess the therapeutic potential of chaperone manipulation. Ubiquitous overexpression of Hsp70 in the R6/2 mouse (as a result of crossing to Hsp70 transgenics) delays aggregate formation by 1 week, has no effect on the detergent solubility of aggregates and does not alter the course of the neurological phenotype. We used an organotypic slice culture assay to show that pharmacological induction of the heat shock response might be a more useful approach. Radicicol and geldanamycin could both maintain chaperone induction for at least 3 weeks and alter the detergent soluble properties of polyQ aggregates over this time course.

Animals↗

PubMed2004

Role of Lewis A and Lewis B blood group antigens in Helicobacter pylori infection.

BACKGROUND AND AIMS: We investigated the prevalence of Helicobacter pylori infection in a large group of women to determine whether there was an association of current infection status with Lewis blood group antigen A and B phenotype. METHODS: Between November 2000 and November 2001, mothers were recruited after delivery of their offspring at the Department of Gynecology and Obstetrics at the University of Ulm, Ulm, Germany. The H. pylori infection status of the women was determined by 13C urea breath test. Their Lewis A and Lewis B phenotype was determined using standard laboratory techniques. RESULTS: In total, 22.2% of the 712 women included in the study (mean age 30.7 years) had a current H. pylori infection. The prevalence of infection varied from 15.5% in women of German nationality to 75.0% in women of Turkish nationality (p < .001). Most women (68.1%) had a Le(a-b+) phenotype. The prevalence of H. pylori infection in women with Le(a-b+) phenotypes was lower than in other women (p = .02). In multivariate analysis, the odds ratio (OR) for a current H. pylori infection given Le(a-b+) was 0.56 [95% confidence interval (CI) 0.33-0.95] compared to women with Le(a-b-). CONCLUSION: Le(a-b+) blood group phenotype in combination with secretor status may hinder colonization of H. pylori in the population studied.

Adolescent↗

PubMed2003

A brain-specific isoform of small glutamine-rich tetratricopeptide repeat-containing protein binds to Hsc70 and the cysteine string protein.

Small glutamine-rich tetratricopeptide repeat-containing protein (SGT) is a ubiquitously expressed cochaperone of heat shock cognate protein of 70 kDa (Hsc70). SGT binds to the C terminus of Hsc70, a site used by several tetratricopeptide repeat-containing binding partners to recruit Hsc70 into complexes of diverse function. We describe here an isoform of SGT with 60% amino acid sequence identity that we name betaSGT. In contrast to the previously published alphaSGT, betaSGT is almost exclusively expressed in brain. Both isoforms of SGT possess similar binding properties toward Hsc70 and cysteine string protein, a synaptic vesicle-associated J-domain-containing protein. In addition, SGTs oligomerize without preferences among isoforms. The distribution of protein binding motifs on SGTs reveals a modular structure. The N-terminal domains mediate oligomerization. Binding to Hsc70 is impaired by mutations of basic residues within the central tetratricopeptide repeat domain of betaSGT, indicating a two-carboxylate clamp as the binding mode. The tetratricopeptide repeats are also necessary for binding to the cysteine string protein. However, this binding mode is distinct from the two-carboxylate clamp that is involved in Hsc70 binding. The C-terminal regions of SGTs might constitute independent protein interaction domains. We conclude that betaSGT is likely to cooperate with alphaSGT as co-chaperone of Hsc70 in the brain. The modular structure of SGTs allows them to recruit client proteins to Hsc70 and to direct the resulting complex toward downstream proteins that take over the respective client proteins.

Amino Acid Motifs↗

PubMed2002

RGM is a repulsive guidance molecule for retinal axons.

Axons rely on guidance cues to reach remote targets during nervous system development. A well-studied model system for axon guidance is the retinotectal projection. The retina can be divided into halves; the nasal half, next to the nose, and the temporal half. A subset of retinal axons, those from the temporal half, is guided by repulsive cues expressed in a graded fashion in the optic tectum, part of the midbrain. Here we report the cloning and functional characterization of a membrane-associated glycoprotein, which we call RGM (repulsive guidance molecule). This molecule shares no sequence homology with known guidance cues, and its messenger RNA is distributed in a gradient with increasing concentration from the anterior to posterior pole of the embryonic tectum. Recombinant RGM at low nanomolar concentration induces collapse of temporal but not of nasal growth cones and guides temporal retinal axons in vitro, demonstrating its repulsive and axon-specific guiding activity.

Amino Acid Sequence↗

PubMed2002

Preparative continuous annular chromatography (P-CAC) enables the large-scale fractionation of fructans.

Fructans (fructo-oligosaccharides and inulin) are of increasing physiological and nutritional interest due to their health-promoting effects. Fructans originally extracted from chicory roots were separated by continuous annular and fixed-bed conventional gel chromatography. Both columns were packed with Toyopearl HW 40 (S) and eluted with deionized water. A multicomponent fractionation was established to obtain single oligosaccharides in a low molecular weight range up to a chain length of five and fractions containing an overall size distribution in the high molecular weight range up to a chain length of 90 monosaccharide units. The productivity and resolution of the continuous annular size exclusion chromatograph (40 cm bed height) were investigated and compared with those of the fixed-bed counterpart (2 x 100 cm bed height). The eluting fractions were analyzed by high-pH anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD). The productivity of the annular system was found to be 25-fold higher than the conventional system. Thus, annular chromatography exemplified for the fractionation of fructans is a powerful method for the large-scale and continuous fractionation of oligomeric and polymeric carbohydrates.

Chemical Fractionation↗

PubMed2002

Structural analysis of underivatized neutral human milk oligosaccharides in the negative ion mode by nano-electrospray MS(n) (part 1: methodology).

Underivatized neutral oligosaccharides from human milk were analyzed by nano-electrospray ionization (ESI) using a quadrupole ion trap mass spectrometer (QIT-MS) in the negative-ion mode. Under these conditions neutral oligosaccharides are observed as deprotonated molecules [M-H]- with high intensity. CID-experiments of these species with the charge localized at the reducing end lead to C-type fragment ions forming a "new" reducing end. Fragmentations are accompanied by cross-ring cleavages that yield information about linkages of internal monosaccharides. Several isomeric compounds with distinct structural features, such as different glycosidic linkages, fucosylation and branching sites were investigated. The rules governing the fragmentation behavior of this class of oligosaccharides were elucidated and tested for a representative number of certain isomeric glycoforms using the MS/MS and MS(n) capabilities of the QIT. On the basis of the specific fragmentation behavior of deprotonated molecules, the position of fucoses and the linkage type (Gal beta-->3 GlcNAc or Gal beta1-->4 GlcNAc) could be determined and linear and branched could be differentiated. Rules could be established which can be applied in further investigations of these types of oligosaccharides even from heterogenous mixtures.

Female↗

PubMed2002

Osmoregulated periplasmic glucans of the free-living photosynthetic bacterium Rhodobacter sphaeroides.

The osmoregulated periplasmic glucans (OPGs) produced by Rhodobacter sphaeroides, a free-living organism, were isolated by trichloracetic acid treatment and gel permeation chromatography. Compounds obtained were characterized by compositional analysis, matrix-assisted laser desorption ionization mass spectrometry and nuclear magnetic resonance. R. sphaeroides predominantly synthesizes a cyclic glucan containing 18 glucose residues that can be substituted by one to seven succinyl esters residues at the C6 position of some of the glucose residues, and by one or two acetyl residues. The glucans were subjected to a mild alkaline treatment in order to remove the succinyl and acetyl substituents, analyzed by MALDI mass spectrometry and purified by high-performance anion-exchange chromatography. Methylation analysis revealed that this glucan is linked by 17 1,2 glycosidic bonds and one 1,6 glycosidic bond. Homonuclear and (1)H/(13)C heteronuclear NMR experiments revealed the presence of a single alpha-1,6 glycosidic linkage, whereas all other glucose residues are beta-1,2 linked. The different anomeric proton signals allowed a complete sequence-specific assignment of the glucan. The structural characteristics of this glucan are very similar to the previously described OPGs of Ralstonia solanacearum and Xanthomonas campestris, except for its different size and the presence of substituents. Therefore, similar OPGs are synthesized by phytopathogenic as well as free-living bacteria, suggesting these compounds are intrinsic components of the Gram-negative bacterial envelope.

Amino Acid Sequence↗

PubMed2001

Genetic analysis of alpha-latrotoxin receptors reveals functional interdependence of CIRL/latrophilin 1 and neurexin 1 alpha.

alpha-Latrotoxin triggers massive neurotransmitter release from nerve terminals by binding to at least two distinct presynaptic receptors, neurexin 1 alpha and CIRL1/latrophilin1 (CL1). We have now generated knockout (KO) mice that lack CL1 and analyzed them alone or in combination with neurexin 1 alpha KO mice. Mice lacking only CL1, or both CL1 and neurexin 1 alpha, were viable and fertile. Ca(2+)-independent binding of alpha-latrotoxin to brain membranes was impaired similarly in CL1 single and in CL1/neurexin 1 alpha double KO mice (approximately 75% decrease) but not in neurexin 1 alpha single KO mice. In contrast, Ca(2+)-dependent binding (approximately 2 times above Ca(2+)-independent binding) was altered in both CL1 (approximately 50% decrease) and neurexin 1 alpha single KO mice (approximately 25% decrease) and was decreased further in double KO mice (approximately 75% decrease). Synaptosomes lacking CL1 exhibited the same decrease in alpha-latrotoxin-stimulated glutamate release in the presence and absence of Ca(2+) (approximately 75%). In contrast, synaptosomes lacking neurexin 1 alpha exhibited only a small decrease in alpha-latrotoxin-triggered release in the absence of Ca(2+) (approximately 20%) but a major decrease in the presence of Ca(2+) (approximately 75%). Surprisingly, synaptosomes lacking both CL1 and neurexin 1 alpha displayed a relatively smaller decrease in alpha-latrotoxin-stimulated glutamate release than synaptosomes lacking only CL1 in the absence of Ca(2+) (approximately 50 versus approximately 75%), but the same decrease in the presence of Ca(2+) (approximately 75%). Our data suggest the following two major conclusions. 1) CL1 and neurexin 1 alpha together account for the majority (75%) of alpha-latrotoxin receptors in brain, with the remaining receptor activity possibly due to other CL and neurexin isoforms, and 2) the two receptors act additively in binding alpha-latrotoxin but not in triggering release. Together these data suggest that the two receptors act autonomously in binding of alpha-latrotoxin but cooperatively in transducing the stimulation of neurotransmitter release by alpha-latrotoxin.

Animals↗