Medicin, narkotika og færdselsuheld

Projektet IMMORTAL - “Impaired Motorists, Methods of Roadside Testing and Assessment for Licensing” har indhentet ny viden om forskellige forhold, der kan påvirke køreegenskaberne. Det drejer sig bla. om påvirkning af medicin, narkotika og alkoholI ét af delprojekterne blev der taget blod- eller spytprøver fra førere af motorkøretøjer, der blev behandlet efter færdselsuheld på Odense Universitetshospital eller Glostrup sygehus. Prøverne blev analyseret for forskellige typer af medicin, narkotika samt alkohol. Patienter, der blev fundet positive eller selv havde oplyst brug af medicin eller narkotika blev efterfølgende kontaktet med henblik på at gennemføre et kvalitativt interview om uheldet og dets konsekvenser. Baseret på interviewene kan det sluttes, at i de fleste uheld var der tale om forskellige uheldsfaktorer, der kunne relateres til interviewpersonen, såsom for høj hastighed, ulovlig overhaling, manglende opmærksomhed, misopfattelse af situationen samt psykisk uligevægt. For halvdelen af de førere, der var positive for et eller flere stoffer, da uheldet skete, kan det ud fra koncentrationen antages, at påvirkningen af dette eller disse stoffer også havde medvirket til uheldets opståen. Interviewene viste, at de påvirkede førere kunne inddeles i 3 grupper: 1. Unge, velfungerende mænd, som tog illegale stoffer (overvejende hash eller amfetamin), enten om aftenen eller i weekenden. 2. Midaldrende mænd og kvinder (35-54 år), der var gået på førtidspension, givet på grund af deres afhængighed af alkohol og/eller lægeordineret medicin. 3. Førere over 55 år, som stadig var aktive på arbejdsmarkedet eller pensionerede, og som anvendte ordineret medicin eller håndkøbsmedicin

Alkohol, medicin og narkotikaforekomst hos alvorligt tilskadekomne bilister

Denne artikel omhandler forekomsten af alkohol, narkotika og medicin hos danske bilister, som er blevet alvorligt skadet i færdselsuheld. Undersøgelsen er baseret på anonyme blodprøver fra 840 førere af person- og varebiler, der blev indbragt til hospitalerne i Odense, Kolding, Vejle, Viborg og Aalborg i perioden oktober 2007 - marts 2010. Blodprøverne blev taget som led i de blodprøver, der rutinemæssigt tages i forbindelse med behandlingen og de blev analyseret for alkohol og 21 misbrugs- og lægemiddelstoffer samt enkelte omdannelsesprodukter (metabolitter). Til brug for undersøgelsen blev der desuden indhentet oplysning om patientens alder og køn, typen af køretøj, uheldsstedet, typen af færdselsuheld, tidspunkt for færdselsuheldet og prøvetagningen samt eventuel medicin, der var blevet givet før udtagning af blodprøven. Alkohol alene eller alkohol sammen med andre stoffer var til stede hos knap 20 % af bilisterne. I alt var 30,5 % af bilisterne positive for én eller flere stofgrupper. Blandt medicinske stoffer alene var forekomsten højest for medicinske opioider (2,5 %) efterfulgt af benzodiazepiner (1,6 %). For de narkotiske stoffer alene var forekomsten højest for cannabis (2,3 %) efterfulgt af amfetamin (1,1 %). I alt var 41,6 % af de tilskadekomne i eneuheld positive for stoffer, heraf 30 % for alkohol alene eller alkohol sammen med andre stoffer. For bilister i flerpartsuheld var i alt 19 % positive, heraf knap 9 % for alkohol alene eller alkohol sammen med andre stoffer. Forekomsten af alkohol og andre stoffer var mere end dobbelt så stor hos de tilskadekomne mænd som hos de tilskadekomne kvinder. For begge køn var forekomsten størst hos aldersgruppen 25-34 år. Forekomsten af alkohol var størst i weekenden, mens den for alle andre stoffer var størst på hverdage. Resultaterne viste, at alkohol var væsentlig hyppigere til stede hos alvorligt skadede bilister i forhold til medicin og narkotika

Probing the O-glycoproteome of gastric cancer cell lines for biomarker discovery

Circulating O-glycoproteins shed from cancer cells represent important serum biomarkers for diagnostic and prognostic purposes. We have recently shown that selective detection of cancer-associated aberrant glycoforms of circulating O-glycoprotein biomarkers can increase specificity of cancer biomarker assays. However, the current knowledge of secreted and circulating O-glycoproteins is limited. Here, we used the COSMC KO "Simple- Cell" (SC) strategy to characterize the O-glycoproteome of two gastric cancer SimpleCell lines (AGS, MKN45) as well as a gastric cell line (KATO III) which naturally expresses at least partially truncated O-glycans. Overall, we identified 499 O-glycoproteins and 1236 O-glycosites in gastric cancer SimpleCells, and a total 47 O-glycoproteins and 73 O-glycosites in the KATO III cell line. We next modified the glycoproteomic strategy to apply it to pools of sera from gastric cancer and healthy individuals to identify circulating O-glycoproteins with the STn glycoform. We identified 37 O-glycoproteins in the pool of cancer sera, and only nine of these were also found in sera from healthy individuals. Two identified candidate O-glycoprotein biomarkers (CD44 and GalNAc-T5) circulating with the STn glycoform were further validated as being expressed in gastric cancer tissue. A proximity ligation assay was used to show that CD44 was expressed with the STn glycoform in gastric cancer tissues. The study provides a discovery strategy for aberrantly glycosylated O-glycoproteins and a set of O-glycoprotein candidates with biomarker potential in gastric cancer.This work was supported by The Danish Research Councils, The Mizutani Foundation, The Danish National Research Foundation (DNRF107) and Fundacão para a Ciência e a Tecnologia (FCT) and COMPETE (Programa Operacional Temático Factores de Competitividade, comparticipado pelo fundo comunitário europeu FEDER) in the framework of the projects: PTDC/BBB-EBI/0786/2012; EXPL/CTM-BIO/0762/2013. Grants were received from FCT (SFRH/BD/73717/2010 to DC), (SFRH/BPD/75871/2011 to AM), (SFRH/BPD/96510/2013 to CG) and (SFRH/BPD/66288/2009 to JAF). IPATIMUP is an Associate Laboratory of the Portuguese Ministry of Science, Technology and Higher Education, and is partially supported by FCT

The GalNAc-type O-Glycoproteome of CHO Cells Characterized by the SimpleCell Strategy

The Chinese hamster ovary cell (CHO) is the major host cell factory for recombinant production of biological therapeutics primarily because of its “human-like” glycosylation features. CHO is used for production of several O-glycoprotein therapeutics including erythropoietin, coagulation factors, and chimeric receptor IgG1-Fc-fusion proteins, however, some O-glycoproteins are not produced efficiently in CHO. We have previously shown that the capacity for O-glycosylation of proteins can be one limiting parameter for production of active proteins in CHO. Although the capacity of CHO for biosynthesis of glycan structures (glycostructures) on glycoproteins are well established, our knowledge of the capacity of CHO cells for attaching GalNAc-type O-glycans to proteins (glycosites) is minimal. This type of O-glycosylation is one of the most abundant forms of glycosylation, and it is differentially regulated in cells by expression of a subset of homologous polypeptide GalNAc-transferases. Here, we have genetically engineered CHO cells to produce homogeneous truncated O-glycans, so-called SimpleCells, which enabled lectin enrichment of O-glycoproteins and characterization of the O-glycoproteome. We identified 738 O-glycoproteins (1548 O-glycosites) in cell lysates and secretomes providing the first comprehensive insight into the O-glycosylation capacity of CHO (http://glycomics.ku.dk/o-glycoproteome_db/)

Structural and biochemical characterization of the exopolysaccharide deacetylase Agd3 required for Aspergillus fumigatus biofilm formation

The exopolysaccharide galactosaminogalactan (GAG) is an important virulence factor of the fungal pathogen Aspergillus fumigatus. Deletion of a gene encoding a putative deacetylase, Agd3, leads to defects in GAG deacetylation, biofilm formation, and virulence. Here, we show that Agd3 deacetylates GAG in a metal-dependent manner, and is the founding member of carbohydrate esterase family CE18. The active site is formed by four catalytic motifs that are essential for activity. The structure of Agd3 includes an elongated substrate-binding cleft formed by a carbohydrate binding module (CBM) that is the founding member of CBM family 87. Agd3 homologues are encoded in previously unidentified putative bacterial exopolysaccharide biosynthetic operons and in other fungal genomes. The exopolysaccharide galactosaminogalactan (GAG) is an important virulence factor of the fungal pathogen Aspergillus fumigatus. Here, the authors study an A. fumigatus enzyme that deacetylates GAG in a metal-dependent manner and constitutes a founding member of a new carbohydrate esterase family.Bio-organic Synthesi

Substrate‐Guided Front‐Face Reaction Revealed by Combined Structural Snapshots and Metadynamics for the Polypeptide N‐Acetylgalactosaminyltransferase 2

The retaining glycosyltransferase GalNAc-T2 is a member of a large family of human polypeptide GalNActransferases that is responsible for the post-translational modification of many cell-surface proteins. By the use of combined structural and computational approaches, we provide the first set of structural snapshots of the enzyme during the catalytic cycle and combine these with quantum-mechanics/molecular-mechanics (QM/MM) metadynamics to unravel the catalytic mechanism of this retaining enzyme at the atomicelectronic level of detail. Our study provides a detailed structural rationale for an ordered bi–bi kinetic mechanism and reveals critical aspects of substrate recognition, which dictate the specificity for acceptor Thr versus Ser residues andenforce a front-face SNi-type reaction in which the substrate Nacetyl sugar substituent coordinates efficient glycosyl transfer

The conserved metalloprotease invadolysin is present in invertebrate haemolymph and vertebrate blood

We identified invadolysin, a novel essential metalloprotease, for functions in chromosome structure, cell proliferation and migration. Invadolysin also plays an important metabolic role in insulin signalling and is the only protease known to localise to lipid droplets, the main lipid storage organelle in the cell. In silico examination of the protein sequence of invadolysin predicts not only protease and lipase catalytic motifs, but also post-translational modifications and the secretion of invadolysin. Here we show that the protease motif of invadolysin is important for its role in lipid accumulation, but not in glycogen accumulation. The lipase motif does not appear to be functionally important for the accumulation of lipids or glycogen. Post-translational modifications likely contribute to modulating the level, localisation or activity of invadolysin. We identified a secreted form of invadolysin in the soluble fraction of invertebrate hemolymph (where we observe sexually dimorphic forms) and also vertebrate plasma, including in the extracellular vesicle fraction. Biochemical analysis for various post-translational modifications demonstrated that secreted invadolysin is both N- and O-glycosylated, but not apparently GPI-linked. The discovery of invadolysin in the extracellular milieu suggests a role for invadolysin in normal organismal physiology