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- ItemMulti-capillary column-ion mobility spectrometry (MCC-IMS) as a new method for the quantification of occupational exposure to sevoflurane in anaesthesia workplaces: an observational feasibility study(London : BioMed Central, 2015) Kunze, Nils; Weigel, Cathrin; Vautz, Wolfgang; Schwerdtfeger, Katrin; JĂ¼nger, Melanie; Quintel, Michael; Perl, ThorstenBackground: Occupational exposure to sevoflurane has the potential to cause health damage in hospital personnel. Workplace contamination with the substance mostly is assessed by using photoacoustic infrared spectrometry with detection limits of 10 ppbv. Multi-capillary column-ion mobility spectrometry (MCC-IMS) could be an alternative technology for the quantification of sevoflurane in the room air and could be even more accurate because of potentially lower detection limits. The aim of this study was to test the hypothesis that MCC-IMS is able to detect and monitor very low concentrations of sevoflurane (<10 ppbv) and to evaluate the exposure of hospital personnel to sevoflurane during paediatric anaesthesia and in the post anaesthesia care unit (PACU). Methods: A MCC-IMS device was calibrated to several concentrations of sevoflurane and limits of detection (LOD) and quantification (LOQ) were calculated. Sevoflurane exposure of hospital personnel was measured at two anaesthesia workplaces and time-weighted average (TWA) values were calculated. Results: The LOD was 0.0068 ppbv and the LOQ was 0.0189 ppbv. During paediatric anaesthesia the mean sevoflurane concentration was 46.9 ppbv (8.0 - 314.7 ppbv) with TWA values between 5.8 and 45.7 ppbv. In the PACU the mean sevoflurane concentration was 27.9 ppbv (8.0 – 170.2 ppbv) and TWA values reached from 8.3 to 45.1 ppbv. Conclusions: MCC-IMS shows a significantly lower LOD and LOQ than comparable methods. It is a reliable technology for monitoring sevoflurane concentrations at anaesthesia workplaces and has a particular strength in quantifying low-level contaminations of sevoflurane. The exposure of the personnel working in these areas did not exceed recommended limits and therefore adverse health effects are unlikely.
- ItemEmission fluxes and atmospheric degradation of monoterpenes above a boreal forest: Field measurements and modelling(Milton Park : Taylor & Francis, 2001) Spanke, Jörg; Rannik, Ăœllar; Forkel, Renate; Nigge, Walter; Hoffman, ThorstenThe contribution of monoterpenes to aerosol formation processes within and above forests is not well understood. This is also true for the particle formation events observed during the BIOFOR campaigns in Hyytiälä, Finland. Therefore, the diurnal variation of the concentrations of several biogenic volatile organic compounds (BVOCs) and selected oxidation products in the gas and particle phase were measured on selected days during the campaigns in Hyytiälä, Finland. α-pinene and Δ3-carene were found to represent the most important monoterpenes above the boreal forest. A clear vertical gradient of their concentrations was observed together with a change of the relative monoterpene composition with height. Based on concentration profile measurements of monoterpenes, their fluxes above the forest canopy were calculated using the gradient approach. Most of the time, the BVOC fluxes show a clear diurnal variation with a maximum around noon. The highest fluxes were observed for α-pinene with values up to 20 ng m−2 s−1 in summer time and almost 100 ng m−2 s−1 during the spring campaign. Furthermore, the main oxidation products from α-pinene, pinonaldehyde, and from β-pinene, nopinone, were detected in the atmosphere above the forest. In addition to these more volatile oxidation products, pinic and pinonic acid were identified in the particle phase in a concentration range between 1 and 4 ng m−3. Beside these direct measurement of known oxidation products, the chemical sink term in the flux calculations was used to estimate the amount of product formation of the major terpenes (α-pinene, β-pinene, Δ3-carene). A production rate of very low volatile oxidation products (e.g., multifunctional carboxylic) from ·OH- and O3-reaction of monoterpenes of about 1.3·104 molecules cm−3 s−1 was estimated for daylight conditions during summer time. Additionally, model calculations with the one-dimensional multilayer model CACHE were carried out to investigate the diurnal course of BVOC fluxes and chemical degradation of terpenes.
- ItemProtein O-mannosylation in the murine brain: Occurrence of Mono-O-Mannosyl glycans and identification of new substrates(San Francisco, CA : Public Library of Science, 2016) Bartels, Markus F.; Winterhalter, Patrick R.; Yu, Jin; Liu, Yan; Lommel, Mark; Möhrlen, Frank; Hu, Huaiyu; Feizi, Ten; Westerlind, Ulrika; Ruppert, Thomas; Strahl, SabineProtein O-mannosylation is a post-translational modification essential for correct development of mammals. In humans, deficient O-mannosylation results in severe congenital muscular dystrophies often associated with impaired brain and eye development. Although various O-mannosylated proteins have been identified in the recent years, the distribution of O-mannosyl glycans in the mammalian brain and target proteins are still not well defined. In the present study, rabbit monoclonal antibodies directed against the O-mannosylated peptide YAT(α1-Man)AV were generated. Detailed characterization of clone RKU-1-3-5 revealed that this monoclonal antibody recognizes O-linked mannose also in different peptide and protein contexts. Using this tool, we observed that mono-O-mannosyl glycans occur ubiquitously throughout the murine brain but are especially enriched at inhibitory GABAergic neurons and at the perineural nets. Using a mass spectrometry-based approach, we further identified glycoproteins from the murine brain that bear single O-mannose residues. Among the candidates identified are members of the cadherin and plexin superfamilies and the perineural net protein neurocan. In addition, we identified neurexin 3, a cell adhesion protein involved in synaptic plasticity, and inter-alpha-trypsin inhibitor 5, a protease inhibitor important in stabilizing the extracellular matrix, as new O-mannosylated glycoproteins.
- ItemUsing SRM-MS to quantify nuclear protein abundance differences between adipose tissue depots of insulin-resistant mice(Rockville : American Society for Biochemistry and Molecular Biology, 2015) Ota, Asuka; Kovary, Kyle M.; Wu, Olivia H.; Ahrends, Robert; Shen, Wen-Jun; Costa, Maria J.; Feldman, Brian J.; Kraemer, Fredric B.; Teruel, Mary N.Insulin resistance (IR) underlies metabolic disease. Visceral, but not subcutaneous, white adipose tissue (WAT) has been linked to the development of IR, potentially due to differences in regulatory protein abundance. Here we investigate how protein levels are changed in IR in different WAT depots by developing a targeted proteomics approach to quantitatively compare the abundance of 42 nuclear proteins in subcutaneous and visceral WAT from a commonly used insulin-resistant mouse model, Lepr(db/db), and from C57BL/6J control mice. The most differentially expressed proteins were important in adipogenesis, as confirmed by siRNA-mediated depletion experiments, suggesting a defect in adipogenesis in visceral, but not subcutaneous, insulin-resistant WAT. Furthermore, differentiation of visceral, but not subcutaneous, insulin-resistant stromal vascular cells (SVCs) was impaired. In an in vitro approach to understand the cause of this impaired differentiation, we compared insulin-resistant visceral SVCs to preadipocyte cell culture models made insulin resistant by different stimuli. The insulin-resistant visceral SVC protein abundance profile correlated most with preadipocyte cell culture cells treated with both palmitate and TNFα. Together, our study introduces a method to simultaneously measure and quantitatively compare nuclear protein expression patterns in primary adipose tissue and adipocyte cell cultures, which we show can reveal relationships between differentiation and disease states of different adipocyte tissue types.
- ItemThe influence of cadmium stress on the content of mineral nutrients and metal-binding proteins in arabidopsis halleri(Heidelberg : Springer, 2012) Przedpełska-Wąsowicz, Ewa; Polatajko, Aleksandra; Wierzbicka, MałgorzataWe investigated the influence of cadmium stress on zinc hyperaccumulation, mineral nutrient uptake, and the content of metal-binding proteins in Arabidopsis halleri. The experiments were carried out using plants subjected to long-term cadmium exposure (40 days) in the concentrations of 45 and 225 μM Cd2+. Inductively coupled plasma-mass spectrometry, size exclusion chromatography coupled with plasma-mass spectrometry, and laser ablation inductively coupled plasma-mass spectrometry used for ablation of polyacylamide gels were employed to assess the content of investigated elements in plants as well as to identify metal-binding proteins. We found that A. halleri is able to translocate cadmium to the aerial parts in high amounts (translocation index >1). We showed that Zn content in plants decreased significantly with the increase of cadmium content in the growth medium. Different positive and negative correlations between Cd content and mineral nutrients were evidenced by our study. We identified more than ten low-molecular-weight (<100 kDa) Cd-binding proteins in Cd-treated plants. These proteins are unlikely to be phytochelatins or metallothioneins. We hypothesize that low-molecular-weight Cd-binding proteins can be involved in cadmium resistance in A. halleri.
- ItemInfrared ellipsometric study of hydrogen-bonded long-chain thiolates on gold: Towards resolving structural details(Basel : MDPI, 2011) Tsankov, Dimiter; Philipova, Irena; Kostova, Kalina; Hinrichs, KarstenA set of newly synthesized aryl-substituted amides of 16-mercaptohexadecanoic acid (R = 4-OH; 3,5-di-OH) are self-assembled on Au(111) substrate. Self assembled monolayers (SAMs) formed by these molecules are studied by ellipsometry from infrared to visible spectral range. Best fit calculations based on the three-phase optical model are employed in order to determine the average tilt angle of the hydrocarbon chains. The data revealed that the SAMs reside in a crystalline-like environment as the long methylene chains predominantly exist in all-trans conformation. The calculated tilt angle of the hydrocarbon chain is decreased by approximately 12° in comparison with the one for the correspondent long-chain n-alkyl thiols. Strong hydrogen bonded networks were detected between the amide proton and the carbonyl oxygen as well as between hydroxyl groups in the end aryl substituents. The transition dipole moments of the C=O, N-H and O-H modes are oriented almost parallel to the gold surface.
- ItemProteome analyses of hepatocellular carcinoma(Sugar Land, TX : Xia & He Publishing, 2014) Megger, Dominik A.; Naboulsi, Naboulsi; Meyer, Helmut E.; Sitek, BarbaraProteomics has evolved into a powerful and widely used bioanalytical technique in the study of cancer, especially hepatocellular carcinoma (HCC). In this review, we provide an up to date overview of feasible proteome-analytical techniques for clinical questions. In addition, we present a broad summary of proteomic studies of HCC utilizing various technical approaches for the analysis of samples derived from diverse sources like HCC cell lines, animal models, human tissue and body fluids.
- ItemIdentification of Eps15 as antigen recognized by the monoclonal antibodies aa2 and ab52 of the wuerzburg hybridoma library against Drosophila brain(San Francisco, CA : Public Library of Science, 2011) Halder, Partho; Chen, Yi-chun; Brauckhoff, Janine; Hofbauer, Alois; Dabauvalle, Marie-Christine; Lewandrowski, Urs; Winkler, Christiane; Sickmann, Albert; Buchner, ErichThe Wuerzburg Hybridoma Library against the Drosophila brain represents a collection of around 200 monoclonal antibodies that bind to specific structures in the Drosophila brain. Here we describe the immunohistochemical staining patterns, the Western blot signals of one- and two-dimensional electrophoretic separation, and the mass spectrometric characterization of the target protein candidates recognized by the monoclonal antibodies aa2 and ab52 from the library. Analysis of a mutant of a candidate gene identified the Drosophila homolog of the Epidermal growth factor receptor Pathway Substrate clone 15 (Eps15) as the antigen for these two antibodies.
- ItemBiological pathways modulated by antipsychotics in the blood plasma of schizophrenia patients and their association to a clinical response(London : Nature Publishing Group, 2015) Martins-de-Souza, Daniel; Solari, Fiorella A.; Guest, Paul C.; Zahedi, René P.; Steiner, JohannProteomics is a valuable tool to unravel molecular mechanisms involved in human disorders. Considering the mediocre effectiveness of antipsychotics, which are the main class of drug used to treat schizophrenia, we analyzed a cohort of 58 schizophrenia patients who had blood collected before and after 6 weeks of antipsychotic treatment using a shotgun mass spectrometry proteomic profiling approach. Our aim was to unravel molecular pathways involved with an effective drug response. The results showed that all patients had essentially the same biochemical pathways triggered Independent of the antipsychotic response outcome. However, we observed that these pathways were regulated in different directions in blood samples from those who responded well to antipsychotics, compared with those who had a poorer outcome. These data are novel, timely and may help to guide new research efforts in the design of new treatments or medications for schizophrenia based on biologically relevant pathways.
- ItemThe proteome of human liver peroxisomes: Identification of five new peroxisomal constituents by a label-free quantitative proteomics survey(San Francisco, CA : Public Library of Science, 2013) Gronemeyer, Thomas; Wiese, Sebastian; Ofman, Rob; Bunse, Christian; Pawlas, Magdalena; Hayen, Heiko; Eisenacher, Martin; Stephan, Christian; Meyer, Helmut E.; Waterham, Hans R.; Erdmann, Ralf; Wanders, Ronald J.; Warscheid, BettinaThe peroxisome is a key organelle of low abundance that fulfils various functions essential for human cell metabolism. Severe genetic diseases in humans are caused by defects in peroxisome biogenesis or deficiencies in the function of single peroxisomal proteins. To improve our knowledge of this important cellular structure, we studied for the first time human liver peroxisomes by quantitative proteomics. Peroxisomes were isolated by differential and Nycodenz density gradient centrifugation. A label-free quantitative study of 314 proteins across the density gradient was accomplished using high resolution mass spectrometry. By pairing statistical data evaluation, cDNA cloning and in vivo colocalization studies, we report the association of five new proteins with human liver peroxisomes. Among these, isochorismatase domain containing 1 protein points to the existence of a new metabolic pathway and hydroxysteroid dehydrogenase like 2 protein is likely involved in the transport or b-oxidation of fatty acids in human peroxisomes. The detection of alcohol dehydrogenase 1A suggests the presence of an alternative alcohol-oxidizing system in hepatic peroxisomes. In addition, lactate dehydrogenase A and malate dehydrogenase 1 partially associate with human liver peroxisomes and enzyme activity profiles support the idea that NAD+ becomes regenerated during fatty acid b-oxidation by alternative shuttling processes in human peroxisomes involving lactate dehydrogenase and/or malate dehydrogenase. Taken together, our data represent a valuable resource for future studies of peroxisome biochemistry that will advance research of human peroxisomes in health and disease.