Glycolipid- and glycoprotein-based blood group A antigen expression in human thrombocytes. A1/A2 difference

Total non-acid glycolipid fractions and total sodium dodecylsulphate (SDS) solubilized protein fractions were isolated from human thrombocytes obtained from single human donors having different blood group A₁/A₂ phenotypes. The blood group A glycolipid antigens were characterized by immunostaining of thin layer plates with different monoclonal anti-A antibodies. The glycoproteins carrying blood group A epitopes were identified by SDS-PAGE and Western blot analysis using a monoclonal anti-A antibody. Blood group A glycolipid antigens were found in both A₁ and A₂ thrombocytes but the A₂ individuals expressed at least ten times less A glycolipids compared to the A₁ individuals. Expression of A type 3/4 chain and small amounts of A type 1 chain glycolipids were seen in thrombocytes of both A₁ and A₂ individuals, while the type 2 chain A glycolipids appeared to be missing from the A₂ thrombocytes. Blood group A reactive glycoproteins were only found in thrombocytes of A₁ individuals and could not be detected in A₂ individuals or a blood group O individual. The major blood group A glycoprotein were found as a double band migrating in the 130 kDa region.Abbreviations SDS sodium dodecyl sulfate - PAGE polyacrylamide gel electrophoresis - HPTLC high performance thin layer chromatography - CBB Coomassie brilliant blue - GVH graft versus host Part of this work was presented at the Xth International Symposium on Glycoconjugates, Jerusalem, Israel. September, 1989.In the short hand designation for glycolipids, the letter indicate blood group determinant, the first numeral, the number of sugar residues, and the second numeral, the type of carbohydrate chain. Thus, A-6-1 means a hexaglycosylceramide with a blood group A determinant based on the type 1 carbohydrate chain.     

Neonatal screening for congenital adrenal hyperplasia using 17-hydroxyprogesterone assay in filter paper blood spots

Screening of infants for congenital adrenal hyperplasia (CAH) using filter paper blood samples collected on the 5th day of life was performed with a radioimmunoassay for 17-hydroxyprogesterone without extraction with organic solvents. A total of 153,000 newborns were screened and 12 cases of CAH were detected (1:12,800). With recall levels related to gestational age, the recall rate could be lowered to 0.05%.     

A method for the quantitation of intestinal metaplasia of the stomach by morphometry

A novel method to quantitate the extent of intestinal metaplasia in gastrectomy specimens is presented. Morphometric measurements of histochemically labeled mucin-producing goblet cells were done in three selected gastrectomies (one having a peptic ulcer, one with adenocarcinoma of the intestinal type, and one with adenocarcinoma of the diffuse type). The sectioning of the gastrectomy specimens was standardized. The results indicated that the intestinal metaplasia was significantly higher in the specimen with adenocarcinoma of the intestinal type (as compared to the other two specimens), both along the lesser and greater curvatures as well as in the fundic area. These quantitative results confirm nonquantitative reports based on subjective visual impressions. This quantitative histochemical method for measuring the actual length as well as the topographical distribution of intestinal metaplasia in resected stomachs will be used in future studies of intestinal metaplasia with the aim of disclosing possible differences among populations with disparate incidences of gastric carcinoma.     

Estimates of hierarchical variation in flower morphology in natural populations ofScleranthus annuus (Caryophyllaceae), an inbreeding annual

The variation in two sets of morphological characters of the flowers of the highly inbreedingScleranthus annuus (Caryophyllaceae) was assessed using 15–20 plants from each of 20 natural populations from the southernmost region of Sweden. The stamen fertility data set consisted of 10 characters describing the degree of fertility of the ten stamens/staminoids, while the sepal/gynoecium data set comprised 10 sepal characters, together with style and stigma length. Substantial variation was found in both the degree of development and the fertility of the stamens, the degree of variability in stamen fertility being related to stamen position within the flower. Considerable variation was found in the characters of the sepal and the gynoecium. Hierarchical analyses of variance indicated that 29% of the variation in total male reproductive effort was distributed among populations, 28% among plants within populations and 43% represented within-plant variation. The corresponding averages for the characters from the sepal/gynoecium data set are 26, 38, and 35%: a greater proportion of the total variance in female reproductive characters is accounted for by among individual variation than is the case with the male reproductive characters. Significance tests of Mahalanobis distances derived by canonical variate analyses indicated that all populations were significantly separated using the sepal/gynoecium data set, while only 50% of the pairwise comparisons on the basis of the stamen fertility data set were significant. Cluster analysis did not reveal any aggregation of the populations. The incongruence of the two data sets and their ability to discriminate between the populations is discussed.     

Symmetric infection of rotavirus on polarized human intestinal epithelial (Caco-2) cells.

When rotavirus infects the mature villus tip cells of the small intestine, it encounters a highly polarized epithelium. In order to understand this virus-cell interaction more completely, we utilized a cell culture-adapted rhesus rotavirus (RRV) to infect human intestinal (Caco-2) and Madin-Darby canine kidney (MDCK-1) polarized epithelial cells grown on a permeable support. Filter-grown Caco-2 cells and MDCK-1 cells, producing a transepithelial resistance of 300 to 500 and greater than 1,000 omega . cm2, respectively, were infected from either the apical or basolateral domain with RRV or Semliki Forest virus. Whereas Semliki Forest virus infection only occurred when input virions had access to the basolateral domain of MDCK-1 or Caco-2 cells, RRV infected MDCK-1 and Caco-2 monolayers in a symmetric manner. The effect of rotavirus infection on monolayer permeability was analyzed by measuring the transepithelial electrical resistance. Rotavirus infection on filter-grown Caco-2 cells caused a transmembrane leak at 18 h postinfection, before the development of the cytopathic effect (CPE) and extensive virus release. Electrical resistance was completely abolished between 24 and 36 h postinfection. Although no CPE could be detected on RRV-infected MDCK cells, the infection caused a transmembrane leak that totally abolished the electrical resistance at 18 to 24 h postinfection. Cell viability and the CPE analysis together with immunohistochemistry and immunofluorescence data indicated that the abolishment of resistance across the monolayer was due not to an effect on the plasma membrane of the cells but to an effect on the paracellular pathway limited by tight junctions. Attachment and penetration of rotavirus onto Caco-2 cells caused no measurable transmembrane leak during the first hour of infection.     

Simulation of field scale denitrification losses from soils under grass ley and barley

Denitrification losses from soils under barley and grass ley crops were simulated. The model, which includes the major processes determining inputs, transformations and outputs of nitrogen in arable soils, represents a scale compatible with information generally available in agricultural field research. The denitrification part of the model includes a field potential denitrification rate and functions for the effect of soil aeration status, soil temperature and soil nitrate content. Easily metabolizable organic matter is assumed not to limit denitrification. Simulated values were compared with denitrification measurements made during two growing seasons in the barley and grass ley treatments of a field experiment in central Sweden.Calibration revealed that the optimal parameter values describing the effect of soil aeration on denitrification rates were similar for both treatments. The response function derived agreed well with two data sets found in the literature. The potential denitrification rate constant, derived in the simulations, was higher for grass ley than for barley, which was consistent with the differences in overall rates of carbon and nitrogen turnover found between treatments.The simulated mean denitrification rates for the two seasons were within 20% of the mean of the measured values. However, simulated denitrification showed less temporal variability and a less skewed frequency distribution than measured denitrification. Some of the measured denitrification events not explained by the model could have been due to the stimulating effects of soil drying/wetting and freezing/thawing on microbial activity.     

Interactions between central glutamatergic,catecholaminergic and cholinergic systems with regard to locomotor activity in monoamine-depleted mice

Summary Following injection of 5µg of the competitive NMDA receptor antagonist AP-5 into the nucleus accumbens, but not following injection of the same dose into the dorsal striatum, a pronounced locomotor stimulation in monoamine-depleted mice was produced; the-adrenoceptor agonist clonidine (1 mg/kg) administered ip caused a marked potentiation of an intraaccumbens AP-5 (2.5µg) injection.On the other hand, 10µg of AP-5 combined with an ip injection of clonidine (1 mg/kg) caused a marked locomotor stimulation following local application into the dorsal striatum but not following application into the prefrontal cortex. Likewise, in combination with systemically administered clonidine, a substantial locomotor stimulation was observed after application of the muscarine receptor antagonist methscopolamine (62µg) into the dorsal striatum but not into the prefrontal cortex.This study suggests that NMDA receptors in the nucleus accumbens exert an inhibitory influence on locomotor activity. The dorsal striatum may also be involved in such control via NMDA and muscarinic receptors.     

Structure of phosphate-free ribonuclease A refined at 1.26 A

The structure of phosphate-free bovine ribonuclease A has been refined at 1.26-A resolution by a restrained least-squares procedure to a final R factor of 0.15. X-ray diffraction data were collected with an electronic position-sensitive detector. The final model consists of all atoms in the polypeptide chain including hydrogens, 188 water sites with full or partial occupancy, and a single molecule of 2-methyl-2-propanol. Thirteen side chains were modeled with two alternate conformations. Major changes to the active site include the addition of two waters in the phosphate-binding pocket, disordering of Gln-11, and tilting of the imidazole ring of His-119. The structure of the protein and of the associated solvent was extensively compared with three other high-resolution, refined structures of this enzyme.     

Hypothermic effects on action potential and force production of hedgehog and guinea pig papillary muscles

Action potentials and isometric force were recorded in papillary muscles from guinea pigs and summer hedgehogs at different temperatures between 37 and 0 degrees C. The action potential of the hedgehog was of a lower amplitude (mean 83 +/- 6 mV) than that of the guinea pig (mean 110 +/- 5 mV). The action potential duration at 50% repolarization was 22 +/- 2 msec in the hedgehog as compared to 105 +/- 11 msec in the guinea pig. Moreover, there was no distinct plateau phase of the hedgehog action potential. Lowering temperature prolonged the action potential duration in the two preparations by about the same percentage. However, the guinea pig preparation became progressively less excitable below 20 degrees C. Lowered temperature produced a positive inotropic effect in the guinea pig, whereas this effect was very slight in the hedgehog heart. Postextrasystolic potentiation was seen in the guinea pig but not in the hedgehog preparation. It is suggested that this difference between the preparations may be due to a greater relative amount of activator calcium in the hedgehog heart. The difference in cold tolerance between the preparations may reflect a difference in chemical composition of the sarcolemma.     

ATP induces microsecond rotational motions of myosin heads crosslinked to actin.

We have used saturation transfer electron paramagnetic resonance (ST-EPR) to study the effect of ATP on the rotational dynamics of spin-labeled myosin heads crosslinked to actin (XLAS1). We have previously shown that ATP induces microsecond rotational motions in activated myofibrils or muscle fibers, but the possibility remained that the motion occurred only in the detached phase of the cross-bridge cycle. The addition of ATP to the crosslinked preparation has been shown to be a model system for active cross-bridges, presumably providing an opportunity to measure the motion in the attached state, without interference from unattached heads. In the absence of ATP, XLAS1 had very little microsecond rotational mobility, yielding a spectrum identical to that observed for uncrosslinked acto-S1. This suggests that all of the labeled S1 forms normal rigor complexes when crosslinked to actin. The addition of 5 mM ATP greatly increased the microsecond rotational mobility of XLAS1, and the effects were reversed upon depletion of ATP. The most plausible explanation for these results is that myosin heads undergo microsecond rotational motion while attached actively to actin during steady state ATPase activity. These results have important implications for the interpretation of spectroscopic data obtained during muscle contraction.