Single-step,rapid separation of acidic and basic isoenzymes from commercial horseradish peroxidase

A simple and sensitive high-performance liquid chromatography method with electrochemical detector is described for the determination of free 3-methoxy-4-hydroxyphenylacetic acid (HVA) in human plasma. The method does not involve any extraction, is specific and reproducible, and has the potential to measure serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) simultaneously. The plasma concentration of free HVA in eight normal, healthy adult volunteers was 10.9 ± 4.6 (mean ± SD). In a preliminary study, in one schizophrenic patient the plasma HVA increased twofold after neuroleptic treatment.     

Impact of salt adaptation on esterified fatty acids and cytochrome oxidase in plasma and thylakoid membranes from the cyanobacterium Anacystis nidulans

During adaptation of photoautotrophically growing fresh water cyanobacterium Anacystis nidulans to high salinity the cells showed a pronounced increase of proton-sodium antiporter activity, and of cytochrome c oxidase in isolated and purified plasma membrane. At the same time the concentrations of plasma membrane-bound EDTA-resistant copper and iron (determined by inductively coupled plasma atomic emission spectrometry) rose proportionately, accompanied by an increase in whole cell respiration. In plasma membranes from salt adapted cells lipid/protein ratios were markedly higher than in control cells, levels of esterified saturated and long-chain fatty acids being significantly higher than the respective levels of unsaturated and short-chain fatty acids which explains the higher lipid-phase transition temperatures derived from Arrhenius plots. Immunoblotting of the membrane proteins with antisera raised against the cytochrome c oxidases from Paracoccus denitrificans and A. nidulans gave two cross-reacting bands with apparent molecular weights around 50000 and 30000 (subunits I and II, respectively) which were more pronounced in plasma membranes from salt adapted cells when compared to control cells. The protein pattern of plasma membranes from salt adapted cells also showed the appearance of bands at apparent molecular weights of 44000–48000 and 54000–56000 which might stem from the proton/sodium-antiporter in this membrane.Abbreviations CM cytoplasmic or plasma membrane - ICM intracytoplasmic or thylakoid membrane - cyt cytochrome - DCCD N,N-dicyclohexylcarbodiimide - Hepes N-2-hydroxyethylpiperazine-N-2-ethanesulfonate - ICP-AES inductively coupled plasma atomic emission spectrometry - SDS-PAGE sodium dodecylsulfate polyacrylamide gel electrophoresis - EPR electron paramagnetic resonance spectrometry     

Die Photosynthese von Meerespflanzen in ihrer Beziehung Zum Salzgehalt

Zusammenfassung Werden Blattstücke vonPosidonia oceanica oder Thallusteile vonUlva lactuca unter sonst optimalen Assimilationsbedingungen aus Meerwasser in Süßwasser, überführt, so sinkt hier die apparente Assimilation beiPosidonia bis zu Null oder gar zu negativen Werten ab, beiUlva bis zu etwa 23% der Leistung im Meerwasser. Bei Rückführung in Meerwasser wird die Assimilationsrate ebenso schnell wieder erhöht, die Ausgangswerte werden allerdings innerhalb der Versuchszeit — einige 20-min-Perioden — meist nicht in voller Höhe wieder erreicht.Folgt der Wechsel zwischen Meer- und Süßwasser im Wechsel von 20 min, so reagieren die Versuchspflanzen mit der gleichen Präzision wie auf das Aus- und Einschalten einer Lichtquelle.Wird anPosidonia die photosynthetische Rate in verschiedenen Stufen verdünnten Meerwassers (Verdünnung nicht mit dest. Wasser, sondern mit Leitungswasser von bekanntem Bicarbonatgehalt und p_H) bestimmt und in Prozenten der Leistung in Meerwasser ausgedrückt, so ergibt sich die in Abb. 3 dargestellte Kurve. Innerhalb der Versuchszeit scheint jeder Meerwasserkonzentration ein bestimmter Photosynthesewert zugeordnet zu sein, ein Verhalten, das auf das Vorhandensein von Gleichgewichtszuständen hindeutet, die zwischen dem photosynthetischen System und dem Zustand des Außenmediums bestehen.Infolge der überaus raschen Reaktion der Pflanzen auf Änderungen im Außenmedium wird als Arbeitshypothese die Vermutung ausgesprochen, daß an der Änderung der photosynthetischen Rate bei Übertragung in und Rückführung aus Süßwasser in Meerwasser nicht Zustandsveränderungen imZellinneren, sondern Zustände der Membranen verantwortlich sind, welche das Ausmaß der CO₂-Permeation bestimmen und damit einen begrenzenden Faktor der Photosynthese darstellen.Mit 4 Textabbildungen     

Prevalence and development of two Sarcocystis spp. in the horse (author's transl)

The prevalence of Sarcocystis spp. in horses was investigated in a survey at the Munich abattoir during 1978/79. Muscle specimens (oesophagus, diaphragm, sublingual muscle, myocardium) were examined using tryptic digestion. Out of 200 horses 31 (15.5%) were found to be carriers of sarcocysts. No parasites were found in the myocardium. In three animals sarcocysts could be isolated and differentiated in fresh preparations. Cysts with 5 to 11 microns by less than 0.5 microns hairlike, unstable protrusions were classified as Sarcocystis equicanis, whereas those with 2.5 to 4.5 microns by 0.8 to 1.0 microns fingerlike, stabile protrusions were assigned to be S. fayeri. Histologically S. equicanis cysts were thin-walled and S. fayeri cysts were thick-walled and often striated. For both species the dog acts as final host. A mixture of sporocysts of both species measured: 12.0--14.4 (13.4 +/- 0.7) X 9.3--10.5 (9.8 +/- 0.4) microns. The prepatent period is 11 to 17 days. Two ponies experimentally infected with 100,000 sporocysts each did not show clinical signs. In fresh preparations and in histopathological examinations of biopsied (111th, 130th, 152th, and 165th day post-infection (p.i.) and postmortem material (167th and 189th day p.i.) different developmental stages of sarcocysts of both species were seen and the following pathological alterations observed: circumscribed non-purulent inflammation, moderate Zenker's degeneration of muscle fibres, and degenerated cysts, of which sometimes only parts of the cyst wall were left. In fresh preparations S. equicanis and S. Fayeri could be differentiated 111 days p.i. The observed disappearance of the sarcocysts is suggested to be a self-cleaning process.     

Molecular Phylogeny of Metazoan Intermediate Filament Proteins

We have cloned cytoplasmic intermediate filament (IF) proteins from a large number of invertebrate phyla using cDNA probes, the monoclonal antibody IFA, peptide sequence information, and various RT-PCR procedures. Novel IF protein sequences reported here include the urochordata and nine protostomic phyla, i.e., Annelida, Brachiopoda, Chaetognatha, Echiura, Nematomorpha, Nemertea, Platyhelminthes, Phoronida, and Sipuncula. Taken together with the wealth of data on IF proteins of vertebrates and the results on IF proteins of Cephalochordata, Mollusca, Annelida, and Nematoda, two IF prototypes emerge. The L-type, which includes 35 sequences from 11 protostomic phyla, shares with the nuclear lamins the long version of the coil 1b subdomain and, in most cases, a homology segment of some 120 residues in the carboxyterminal tail domain. The S-type, which includes all four subfamilies (types I to IV) of vertebrate IF proteins, lacks 42 residues in the coil 1b subdomain and the carboxyterminal lamin homology segment. Since IF proteins from all three phyla of the chordates have the 42-residue deletion, this deletion arose in a progenitor prior to the divergence of the chordates into the urochordate, cephalochordate, and vertebrate lineages, possibly already at the origin of the deuterostomic branch. Four phyla recently placed into the protostomia on grounds of their 18S rDNA sequences (Brachiopoda, Nemertea, Phoronida, and Platyhelminthes) show IF proteins of the L-type and fit by sequence identity criteria into the lophotrochozoic branch of the protostomia. Received: 2 April 1998 / Accepted: 19 June 1998     

Characterization of a Dopamine D1 Receptor from Apis mellifera: Cloning, Functional Expression, Pharmacology, and mRNA Localization in the Brain

Abstract: The neurotransmitter dopamine is an important regulator of physiological and behavioral functions in both vertebrates and invertebrates. We have isolated a homologue of the vertebrate dopamine D1 receptor subfamily from the honeybee Apis mellifera . [³H]Lysergic acid diethylamide specifically binds to the heterologously expressed receptor with K _D∼5 n M . Dopaminergic receptor ligands compete for this high-affinity binding, with the following order of potency: R (+)-lisuride > chlorpromazine = cis ( Z )-flupentixol > dopamine > S (+)-butaclamol > R (+)-SCH 23390 > haloperidol. Activation of the heterologously expressed receptor of Apis mellifera leads to cyclic AMP production. Receptor mRNA is expressed in perikarya of different brain neuropils, including those of mushroom body intrinsic neurons. These results suggest that this dopamine receptor is involved in signal processing of visual and olfactory information in the honeybee.     

Inequalities of visceral leishmaniasis case-fatality in Brazil: A multilevel modeling considering space,time, individual and contextual factors

BackgroundIn Brazil, case-fatality from visceral leishmaniasis (VL) is high and characterized by wide differences between the various political-economic units, the federated units (FUs). This study was designed to investigate the association between factors at the both FU and individual levels with the risk of dying from VL, after analysing the temporal trend and the spatial dependency for VL case-fatality.MethodologyThe analysis was based on individual and aggregated data of the Reportable Disease Information System-SINAN (Brazilian Ministry of Health). The temporal and spatial distributions of the VL case-fatality between 2007 and 2017 (27 FUs as unit of analysis) were considered together with the individual characteristics and many other variables at the FU level (socioeconomic, demographic, access to health and epidemiological indicators) in a mixed effects models or multilevel modeling, assuming a binomial outcome distribution (death from VL).FindingsA linear increasing temporal tendency (4%/year) for VL case-fatality was observed between 2007 and 2017. There was no similarity between the case-fatality rates of neighboring FUs (non-significant spatial term), although these rates were heterogeneous in this spatial scale of analysis. In addition to the known individual risk factors age, female gender, disease’s severity, bacterial co-infection and disease duration, low level schooling and unavailability of emergency beds and health professionals (the last two only in univariate analysis) were identified as possibly related to VL death risk. Lower VL incidence was also associated to VL case-fatality, suggesting that unfamiliarity with the disease may delay appropriate medical management: VL patients with fatal outcome were notified and had VL treatment started 6 and 3 days later, respectively, in relation to VL cured patients. Access to garbage collection, marker of social and economic development, seems to be protective against the risk of dying from VL. Part of the observed VL case-fatality variability in Brazil could not be explained by the studied variables, suggesting that factors linked to the intra FU environment may be involved.ConclusionsThis study aimed to identify epidemiological conditions and others related to access to the health system possibly linked to VL case-fatality, pointing out new prognostic determinants subject to intervention.     

Caspase-mediated cleavage of the exosome subunit PM/Scl-75 during apoptosis

Recent studies have implicated the dying cell as a potential reservoir of modified autoantigens that might initiate and drive systemic autoimmunity in susceptible hosts. A number of subunits of the exosome, a complex of 3'→5' exoribonucleases that functions in a variety of cellular processes, are recognized by the so-called anti-PM/Scl autoantibodies, found predominantly in patients suffering from an overlap syndrome of myositis and scleroderma. Here we show that one of these subunits, PM/Scl-75, is cleaved during apoptosis. PM/Scl-75 cleavage is inhibited by several different caspase inhibitors. The analysis of PM/Scl-75 cleavage by recombinant caspase proteins shows that PM/Scl-75 is efficiently cleaved by caspase-1, to a smaller extent by caspase-8, and relatively inefficiently by caspase-3 and caspase-7. Cleavage of the PM/Scl-75 protein occurs in the C-terminal part of the protein at Asp369 (IILD³⁶⁹↓G), and at least a fraction of the resulting N-terminal fragments of PM/Scl-75 remains associated with the exosome. Finally, the implications of PM/Scl-75 cleavage for exosome function and the generation of anti-PM/Scl-75 autoantibodies are discussed.