Identification of capillaries in sections from skeletal muscle by use of lectins and monoclonal antibodies reacting with histo-blood group ABH antigens

This study was performed to evaluate the application of different lectins and monoclonal antibodies against ABH antigens to detect and characterize carbohydrate structures in capillaries of skeletal muscle from humans and laboratory animals. Blood group specific lectins (Griffonia simplicifolia, Griffonia simplicifolia isolectin B4,Lotus tetragonlobus, Ulex europaeus, andDolichos biflorus) and monoclonal antibodies reacting with histo-blood group carbohydrate antigens belonging to type 1 (Le^a) and type 2 (H, A and Le^y) chains were used as histological markers for capillaries in sections from skeletal muscle. The material consisted of 20 human masseter muscle biopsies from individuals with known blood types: (eight blood group O, nine blood group A, two blood group B, and one blood group AB) and masseter muscles specimens from different laboratory animals (mouse, rat, rabbit, cat, dog, pig, cow, and macaca monkey). Unfixed sections and an avidin alkaline phosphatase method were used to visualize the specific reaction.Ulex lectin stained capillaries in all human biopsies either strongly or moderately. Strong muscle capillary reaction was observed in biopsies from O, B and AB individuals while capillaries from A individuals were only moderately stained.Griffonia simplicifolia marked capillaries in A, B, and AB individuals andGriffonia simplicifolia isolectin B4 stained capillaries in muscle biopsies from B and AB donors.Dolichos biflorus was a weak marker of muscle capillaries from A individuals. Only capillaries from O individuals were stained with the antibody against H type 2. Capillary reaction was not observed with the other antibodies used.Girffonia simplicifolia was an excellent marker for capillaries in mouse muscle whileGriffonia simplicifolia isolectin B4 is recommended for rat muscles. Periodic acid treatment and subsequentLotus tetragonolobus staining is suitable to visualize capillaries in mouse, rat and pig muscle. Using a sensitive histochemical technique for staining with lectins and monoclonal antibodies reacting with blood group related antigens the microvascular density in human skeletal muscle may be estimated. Further, the carbohydrate compounds in the muscle capillaries reflect the individual blood type. A selection of lectins is suitable for demonstration of capillaries in animal skeletal muscle.     

Topological Analysis of Chlamydia trachomatis L2 Outer Membrane Protein 2

Using monospecific polyclonal antisera to different parts of Chlamydia trachomatis L2 outer membrane protein 2 (Omp2), we show that the protein is localized at the inner surface of the outer membrane. Omp2 becomes immunoaccessible when Chlamydia elementary bodies are treated with dithiothreitol, and protease digestions indicate that Omp2 has a possible two-domain structure.     

Denitrification, Dissimilatory Reduction of Nitrate to Ammonium, and Nitrification in a Bioturbated Estuarine Sediment as Measured with 15N and Microsensor Techniques

Nitrogen and oxygen transformations were studied in a bioturbated (reworked by animals) estuarine sediment (Norsminde Fjord, Denmark) by using a combination of ¹⁵N isotope (NO₃^-), specific inhibitor (C₂H₂), and microsensor (N₂O and O₂) techniques in a continuous-flow core system. The estuarine water was NO₃^- rich (125 to 600 μM), and NO₃^- was consistently taken up by the sediment on the four occasions studied. Total NO₃^- uptake (3.6 to 34.0 mmol of N m^-2 day^-1) corresponded closely to N₂ production (denitrification) during the experimental steady state, which indicated that dissimilatory, as well as assimilatory, NO₃^- reduction to NH₄⁺ was insignificant. When C₂H₂ was applied in the flow system, denitrification measured as N₂O production was often less (58 to 100%) than the NO₃^- uptake because of incomplete inhibition of N₂O reduction. The NO₃^- formed by nitrification and not immediately denitrified but released to the overlying water, uncoupled nitrification, was calculated both from ¹⁵NO₃^- dilution and from changes in NO₃^- uptake before and after C₂H₂ addition. These two approaches gave similar results, with rates ranging between 0 and 8.1 mmol of N m^-2 day^-1 on the four occasions. Attempts to measure total nitrification activity by the difference between NH₄⁺ fluxes before and after C₂H₂ addition failed because of non-steady-state NH₄⁺ fluxes. The vertical distribution of denitrification and oxygen consumption was studied by use of N₂O and O₂ microelectrodes. The N₂O profiles measured during the experimental steady state were often irregularly shaped, and the buildup of N₂O after C₂H₂ was added was much too fast to be described by a simple diffusion model. Only bioturbation by a dense population of infauna could explain these observations. This was corroborated by the relationship between diffusive and total fluxes, which showed that only 19 to 36 and 29 to 62% of the total O₂ uptake and denitrification, respectively, were due to diffusion-reaction processes at the regular sediment surface, excluding animal burrows.     

Histochemical studies on genetical control of hormonal enzyme inducibility in the mouse. III. Beta-glucuronidase distribution pattern of epididymis in different genotypes.

Synopsis This article reports the application of Hayashi's histochemical technique for -glucoronidase to mouse epididymis. A methodological study. which established optimal conditions for demonstrating the enzyme in this organ, is reported. The distribution pattern of -glucuronidase is described and correlated with previous data for -naphtyl acetate esterase. Differences between sites of granular and diffuse reaction product for these two enzymes are recorded. Possible interpretations of these findings in terms of intracellular localization of enzymes are discussed. Studies on different strains reveal regular differences in histochemical organization between mice of various genotypes. Histochemical data which imply androgen inducibility of -glucuronidase in mouse epididymis are preliminarily noted. Present address: Department of Anatomy, Faculty of Medicine, Sir Charles Tupper Medical Building, Dalhousie University, Halifax, Nova Scotia, Canada.     

Gastric inflammatory markers and interleukins in patients with functional dyspepsia, with and without Helicobacter pylori infection

Helicobacter pylori is the most important cause of gastritis, peptic ulcers and the development of gastric cancer. The chronic active inflammation is dominated by neutrophils, macrophages, lymphocytes and plasma cells. Several interleukins (IL-8, IL-10 and IFN-gamma) are involved in the inflammatory process in the gastric mucosa. The aim of this study was to investigate the gastric inflammation in patients with functional dyspepsia. Fifty-three consecutive patients were included and antral biopsies were obtained for histology, culture and immunohistochemistry. The sections were examined for the interleukins IL-4, IL-6, IL-8, IL-10 and IFN-gamma as well as for the cell markers CD4, CD8, CD14, Cd19, CD25 and CD30. Only CD4 and CD19 were significantly increased in patients with increased gastric inflammation and increased density of H. pylori. However, several of the examined markers (IFN-gamma, IL-8, IL-10 and CD14) showed a non-significant trend to be increased in patients with extensive gastric inflammation and high density of H. pylori. Therefore, an arbitrary index (IM11) for all the 11 immunological markers was made as an average value for each of the four morphological groups. For the four morphologically different groups of patients the values were 0.49, 0.77, 0.86 and 1.25, respectively. Significant increases in the index from none to moderate antral inflammation as well as the density of H. pylori were found (p<0.001). By using an index of inflammatory markers trends can be summarized and thereby significant which may be of importance when gastric inflammation is investigated in children and patients with functional dyspepsia.     

(S)-Carboxy-3-Hydroxyphenylglycine, an Antagonist of Metabotropic Glutamate Receptor (mGluR)1a and an Agonist of mGluR2, Protects Against Audiogenic Seizures in DBA/2 Mice

Abstract: The in vivo anticonvulsant effects and in vitro metabo-tropic glutamate receptor selectivity of ( S )-4-carboxy-3-hydroxy-phenylglycine [(S)-4C3HPG] were examined. Intracerebroventricular injection of (S)-4C3HPG dose-dependently antagonized audiogenic-induced clonic and tonic convulsions in DBA/2 mice with ED₆₀ values of 76 and 110-nmol per mouse, respectively. (S)-4C3HPG dose-dependently inhibited the spontaneously evoked epileptic spikes in a cingulate cortex-corpus callosum slice preparation. (SJ-4C3HPG displaced the binding of [³H]glutamate in membranes prepared from baby hamster kidney (BHK) cells expressing the metabotropic glutamate receptor mGluR1a with an EC₅₀ of 5 β 1 u M. ( S )-4C3HPG dose-dependently antagonized glutamate-stimulated phosphoinositide hydrolysis in BHK cells expressing mGluR 1a with an IC₅₀ of 15 β 3 μ M. ( S )-4C3HPG was, however, an agonist at mGluR2 with an EC₆₀ of 21 β 4 μ M for inhibition of forskolin-stimulated cyclic AMP formation in BHK cells expressing the mGluR2. ( S )-4C3HPG had no effects at mGluR4a. These data suggest that the anticonvulsant action of ( S )-4C3HPG is mediated by combined antagonism of mGluRIa and agonism of mGluR2. These results suggest the importance of mGluR1a and/or mGluR2 in the control of epileptic activity.     

Utilization of side-chain precursors for penicillin biosynthesis in a high-producing strain of Penicillium chrysogenum

Utilization of the side-chain precursors phenoxyacetic acid (POA) and phenylacetic acid (PA) for penicillin biosynthesis by Penicillium chrysogenum was studied in shake flasks. Precursor uptake and penicillin production were followed by HPLC analysis of precursors and products in the medium and in the cells. P. chrysogenum used both POA and PA as precursors, producing phenoxymethylpenicillin (penicillin V) and benzylpenicillin (penicillin G), respectively. If both precursors were present simultaneously, the formation of penicillin V was blocked and only penicillin G was produced. When PA was added at different times to cells that were induced initially for POA utilization and were producing penicillin V, the POA utilization and penicillin V formation were blocked, whereas the cells started utilizing PA and produced penicillin G. The blocking of the POA turnover lasted for as long as PA was present in the medium. If POA was added to cultures induced initially for PA utilization and producing penicillin G, this continued irrespective of the presence of POA. Utilization of POA increased concomitant with depletion of PA from the medium. Analysis of cellular pools from a growing cell system with POA as precursor to which PA was added after 48 h showed that the cellular concentration of POA was kept high without production of penicillin V and at a concentration comparable to the concentration in the medium. The cellular concentration of POA was higher than the concentration of PA that was utilized for penicillin G production. Correspondence to: S. Havn Eriksen