Influenza Viruses Display High-Affinity Binding to Human Polyglycosylceramides Represented on a Solid-Phase Assay Surface

AbstractPolyglycosylceramides (PGCs), complex glycolipids containing up to 50 or more sugar residues, are recognized as the minor components of the cell-surface membranes, but a knowledge on their tissue distribution, structure, and function is limited. In this study, the binding of influenza viruses to preparations of PGCs was investigated using a TLC overlay assay and a microwell adsorption assay. The ability of PGCs to bind influenza virus was dependent on the source from which they were derived. Preparations of PGCs from human erythrocytes were found to support binding of A and B influenza virus strains at a much lower concentration than sialyl-6-paragloboside and to be somewhat better receptors for these viruses compared to the sialylglycoprotein fetuin. A high virus-binding activity of PGCs suggests that these species could potentially serve as biologically important cell-surface receptors for influenza viruses

Discovery of Species-unique Peptide Biomarkers of Bacterial Pathogens by Tandem Mass Spectrometry-based Proteotyping

Mass spectrometry (MS) and proteomics offer comprehensive characterization and identification of microorganisms and discovery of protein biomarkers that are applicable for diagnostics of infectious diseases. The use of biomarkers for diagnostics is widely applied in the clinic and the use of peptide biomarkers is increasingly being investigated for applications in the clinical laboratory. Respiratory-tract infections are a predominant cause for medical treatment, although, clinical assessments and standard clinical laboratory protocols are time-consuming and often inadequate for reliable diagnoses. Novel methods, preferably applied directly to clinical samples, excluding cultivation steps, are needed to improve diagnostics of infectious diseases, provide adequate treatment and reduce the use of antibiotics and associated development of antibiotic resistance. This study applied nano-liquid chromatography (LC) coupled with tandem MS, with a bioinformatics pipeline and an in-house database of curated high-quality reference genome sequences to identify species-unique peptides as potential biomarkers for four bacterial pathogens commonly found in respiratory tract infections (RTIs): Staphylococcus aureus; Moraxella catarrhalis; Haemophilus influenzae and Streptococcus pneumoniae. The species-unique peptides were initially identified in pure cultures of bacterial reference strains, reflecting the genomic variation in the four species and, furthermore, in clinical respiratory tract samples, without prior cultivation, elucidating proteins expressed in clinical conditions of infection. For each of the four bacterial pathogens, the peptide biomarker candidates most predominantly found in clinical samples, are presented. Data are available via ProteomeXchange with identifier PXD014522. As proof-of-principle, the most promising species-unique peptides were applied in targeted tandem MS-analyses of clinical samples and their relevance for identifications of the pathogens, i.e. proteotyping, was validated, thus demonstrating their potential as peptide biomarker candidates for diagnostics of infectious diseases

Helicobacter suis binding to carbohydrates on human and porcine gastric mucins and glycolipids occurs via two modes

Helicobacter suis colonizes the stomach of most pigs and is the most prevalent non-Helicobacter pylori Helicobacter species found in the human stomach. In the human host, H. suis contributes to the development of chronic gastritis, peptic ulcer disease and MALT lymphoma, whereas in pigs it is associated with gastritis, decreased growth and ulcers. Here, we demonstrate that the level of H. pylori and H. suis binding to human and pig gastric mucins varies between individuals with species dependent specificity. The binding optimum of H. pylori is at neutral pH whereas that of H. suis has an acidic pH optimum, and the mucins that H. pylori bind to are different than those that H. suis bind to. Mass spectrometric analysis of mucin O-glycans from the porcine mucin showed that individual variation in binding is reflected by a difference in glycosylation; of 109 oligosaccharide structures identified, only 14 were present in all examined samples. H. suis binding to mucins correlated with glycans containing sulfate, sialic acid and terminal galactose. Among the glycolipids present in pig stomach, binding to lactotetraosylceramide (Gal beta 3GlcNAc beta 3Gal beta 4Glc beta 1Cer) was identified, and adhesion to Gal beta 3GlcNAc beta 3Gal beta 4Glc at both acidic and neutral pH was confirmed using other glycoconjugates. Together with that H. suis bound to DNA (used as a proxy for acidic charge), we conclude that H. suis has two binding modes: one to glycans terminating with Gal beta 3GlcNAc, and one to negatively charged structures. Identification of the glycan structures H. suis interacts with can contribute to development of therapeutic strategies alternative to antibiotics

Erythrocyte and Porcine Intestinal Glycosphingolipids Recognized by F4 Fimbriae of Enterotoxigenic Escherichia coli

Enterotoxigenic F4-fimbriated Escherichia coli is associated with diarrheal disease in neonatal and postweaning pigs. The F4 fimbriae mediate attachment of the bacteria to the pig intestinal epithelium, enabling an efficient delivery of diarrhea-inducing enterotoxins to the target epithelial cells. There are three variants of F4 fimbriae designated F4ab, F4ac and F4ad, respectively, having different antigenic and adhesive properties. In the present study, the binding of isolated F4ab, F4ac and F4ad fimbriae, and F4ab/ac/ad-fimbriated E. coli, to glycosphingolipids from erythrocytes and from porcine small intestinal epithelium was examined, in order to get a comprehensive view of the F4-binding glycosphingolipids involved in F4-mediated hemagglutination and adhesion to the epithelial cells of porcine intestine. Specific interactions between the F4ab, F4ac and F4ad fimbriae and both acid and non-acid glycosphingolipids were obtained, and after isolation of binding-active glycosphingolipids and characterization by mass spectrometry and proton NMR, distinct carbohydrate binding patterns were defined for each fimbrial subtype. Two novel glycosphingolipids were isolated from chicken erythrocytes, and characterized as GalNAcα3GalNAcß3Galß4Glcß1Cer and GalNAcα3GalNAcß3Galß4GlcNAcß3Galß4Glcß1Cer. These two compounds, and lactosylceramide (Galß4Glcß1Cer) with phytosphingosine and hydroxy fatty acid, were recognized by all three variants of F4 fimbriae. No binding of the F4ad fimbriae or F4ad-fimbriated E. coli to the porcine intestinal glycosphingolipids occurred. However, for F4ab and F4ac two distinct binding patterns were observed. The F4ac fimbriae and the F4ac-expressing E. coli selectively bound to galactosylceramide (Galß1Cer) with sphingosine and hydroxy 24:0 fatty acid, while the porcine intestinal glycosphingolipids recognized by F4ab fimbriae and the F4ab-fimbriated bacteria were characterized as galactosylceramide, sulfatide (SO3-3Galß1Cer), sulf-lactosylceramide (SO3-3Galß4Glcß1Cer), and globotriaosylceramide (Galα4Galß4Glcß1Cer) with phytosphingosine and hydroxy 24:0 fatty acid. Finally, the F4ad fimbriae and the F4ad-fimbriated E. coli, but not the F4ab or F4ac subtypes, bound to reference gangliotriaosylceramide (GalNAcß4Galß4Glcß1Cer), gangliotetraosylceramide (Galß3GalNAcß4Galß4Glcß1Cer), isoglobotriaosylceramide (Galα3Galß4Glcß1Cer), and neolactotetraosylceramide (Galß4GlcNAcß3Galß4Glcß1Cer)

Sulfatide Recognition by Colonization Factor Antigen CS6 from Enterotoxigenic Escherichia coli

The first step in the pathogenesis of enterotoxigenic Escherichia coli (ETEC) infections is adhesion of the bacterium to the small intestinal epithelium. Adhesion of ETEC is mediated by a number of antigenically distinct colonization factors, and among these, one of the most commonly detected is the non-fimbrial adhesin coli surface antigen 6 (CS6). The potential carbohydrate recognition by CS6 was investigated by binding of recombinant CS6-expressing E. coli and purified CS6 protein to a large number of variant glycosphingolipids separated on thin-layer chromatograms. Thereby, a highly specific binding of the CS6-expressing E. coli, and the purified CS6 protein, to sulfatide (SO3-3Galβ1Cer) was obtained. The binding of the CS6 protein and CS6-expressing bacteria to sulfatide was inhibited by dextran sulfate, but not by dextran, heparin, galactose 4-sulfate or galactose 6-sulfate. When using recombinantly expressed and purified CssA and CssB subunits of the CS6 complex, sulfatide binding was obtained with the CssB subunit, demonstrating that the glycosphingolipid binding capacity of CS6 resides within this subunit. CS6-binding sulfatide was present in the small intestine of species susceptible to CS6-mediated infection, e.g. humans and rabbits, but lacking in species not affected by CS6 ETEC, e.g. mice. The ability of CS6-expressing ETEC to adhere to sulfatide in target small intestinal epithelium may thus contribute to virulence

T-Cell Protein Tyrosine Phosphatase, a Regulator of the PDGF Signaling Pathway

Platelet-derived growth factor (PDGF) is a potent stimulator of cell growth, survival and motility. PDGF exerts its function by binding to specific tyrosine kinase receptors, initiating receptor auotphosphorylation and initiation of specific signaling pathways that regulates the cellular response. It is critical that these signals can be modulated and terminated, since over-activation of signaling pathways are often found in diseases, such as cancer. Protein tyrosine phosphatases (PTPs) counteract the tyrosine kinases by dephosphorylating proteins, thereby playing a crucial role in the control of signaling events. The aim of this thesis has been to study the regulation of PDGF receptor signaling by the T-cell protein tyrosine phosphatase (TC-PTP). In the first two studies, we demonstrated that loss of TC-PTP specifically redirected the PDGF β-receptor towards a rapid Rab4a-dependent recycling after ligand-induced internalization. Furthermore, we found that the sorting of activated PDGF β-receptor into the recycling pathway was dependent on sequential PKCα and Rab4a activation. Since the PDGF α-receptor did not recycle in the absence of TC-PTP, this study displays the first evidence of differences in trafficking of the PDGF receptor family members. PDGF β-receptor recycling was also induced by activating PKCα through the LPA receptor. The LPA-induced PDGF β-receptor recycling correlated with increased receptor phosphorylation and cell migration at low concentrations of PDGF-BB. The data suggests that PKCα activation could serve as a point of cross-talk between receptor families, regulating the duration and magnitude of PDGF β-receptor signaling. In the last study, we searched for novel substrates for TC-PTP downstream of the PDGF β-receptor, and identified the pyruvate kinase M2, PK-M2, as a possible substrate. PK-M2 is expressed in cells that proliferate rapidly, including tumor cells. Our data suggests that TC-PTP can interact with the glycolytic complex, affecting the activity of PK-M2 and hence, altering the glucose metabolism for proliferating tumor cells

Sjuksköterskors möjligheter att främja hälsoprocesser hos patienter med diabetes typ 2 : En litteraturöversikt

Bakgrunden till studien är att diabetes typ 2 är ett utbrett problem i samhället. Det kräver mycket resurser från vården och därför är det viktigt att främja hälsa hos patienter med diabetes typ 2. Förutom resurser från vården är det många patienter med diabetes typ 2 som upplever ohälsa och inte har motivation till livsstilsförändringar. Om patienter genomgår nödvändiga livsstilsförändringar och kan identifiera vad som får dem att uppleva hälsa bidrar det till bättre folkhälsa. Syftet är att undersöka vad som kan bidra till att stödja och stärka hälsoprocesser hos patienter som lever med typ 2 diabetes från sjuksköterskors och patienters perspektiv. Litteraturöversikten har fördjupat sig i forskningen hos patienter med diabetes typ 2 och de processer som främjar hälsa för att identifiera problem och möjligheter som finns i livsstilsförändringar. Artiklar söktes i olika databaser med olika sökord för att hitta passande information. Etiska överväganden och tankar på hållbar utveckling har även varit i fokus. Resultatet visar att patienter med diabetes typ 2 är i behov av ett stöd för kunskap, motivation och hjälp i sina livsstilsförändringar. Resultatet formade två huvudteman, Betydelsen av kunskap och Vikten av en vårdande relation. Utifrån dessa formades fem subteman, Betydelsen av att dela kunskap om diabetes typ 2, Kunskap om livsstilens inverkan på hälsa, Kunskap om kost, Sjuksköterskors och vårdpersonalens betydelse och Stödja egenvård. Slutsatserna betonar att sjuksköterskor kan bidra i patienters hälsoprocess med kunskap, stöd, motivation genom goda relationer samt tillämpa personcentrerad vård och främja delaktighet

Breast cancerpatients experiences from the nursing encounter : a litterature study

År 2000 fick 6 348 kvinnor diagnosen bröst cancer. Bröst cancer är därmed den vanligaste formen av cancer hos kvinnor. Personer som har fått diagnosen cancer upplever stora existentiella förändringar, osäkerhet och ensamhet. Syftet med denna studie var att beskriva bröst cancerpatienters upplevelser i mötet med sjuksköterskan. Studien är en litteraturstudie och författarna har använt sig av Polit & Hunglers modell. I resultatet framgick det att kvinnornas upplevelser i mötet med sjuksköterskan var både positiva och negativa. Resultatet indelades i fyra olika teman: sjuksköterskans bemötande, tillgänglighet, sjuksköterskans kompetens och kommunikation ? information. Beroende på hur kvinnorna upplevde mötet med sjuksköterskan ledde det till antigen lidande eller välbefinnande. Positiva upplevelser ökade kvinnornas välbefinnande medan negativa upplevelser orsakade lidande. Ingen av kvinnorna lämnades dock oberörd efter mötet med sjuksköterskan. Resultatet visade att sjuksköterskans bemötande var avgörande för hur kvinnorna klarade av sin sjukdomssituation. Om kvinnorna var ångestfyllda, förnekande eller hade accepterat sin situation färgade deras upplevelser av mötet med sjuksköterskan.In year 2000 6 348 women were diagnosed with breast cancer, breast cancer is thereby the most common form of cancer among women. People who are diagnosed with cancer experience great existentiell changes, uncertainty and loneliness. The aim of this study was to describe the breast cancerpatients experiences from the nursing encounter. This study is a litterature study and the authors have used Polit & Hunglers modell. The findings showed that the womens experiences were both positive and negative from the nursing encount. The findings were sorted into four different themes: the nurses treatment, accessibility, the nurses competance and communication ? information. Depending on how the women experienced the nursing encounter it resulted in wellbeing or suffering. Positive experiences increased the womens wellbeing and negative experiences caused suffering. Still none of the women were left uneffected after the nursing encounter. The findings showed that the nurses treatment of the patients were of vital importance for how the women managed their illness and their situation. Whether the women were afraid, in denial or accepted the situation also effected the nursing encounter

Lärares planering - en kvalitativ studie om hur lärare i Geografi och Idrott och hälsa uppfattar planering

Syftet med arbetet är att undersöka lärares syn på planering och hur ämnesplanering för kurser och lektioner fungerar i ämnena Geografi och Idrott och hälsa. För att uppnå syftet har en kvalitativ metod med fenomenografisk inriktning använts. Intervjuer har genomförts med fyra geografilärare och fyra idrottslärare på fyra olika skolor. Svaren delges som utvalda citat i resultatdelen. De följs av en fenomenografisk analys av resultatet, där svaren delas in i kategorier. Det vi kommit fram till är att de lärare vi intervjuat har samma syn på planering som styrdokumenten och de planerar bland annat för att få en stomme och känna sig trygga. De börjar med att grovplanera för att sedan detaljplanera och de anser att en röd tråd bör finnas i undervisningen. Det finns en vilja hos lärarna att samarbeta med andra lärare och många av dem gör det. Eleverna har inflytande i planeringsarbetet. Resultatet visar även att lärarna känner sig tryggare nu än när de var nya i yrket