Microphotometric analysis of NADH-tetrazolium reductase and alpha-glycerophosphate dehydrogenase in human quadriceps muscle.

Synopsis In serial cross-sections of human skeletal muscles stained for either NADH-tetrazolium reductase (NADH-TR) or -glycerophosphate dehydrogenase (-GPD), a linear relation was found between the total content of enzyme in a cell (expressed as the thickness of the section) and the absorbance of the formazan reaction product formed. Little variation (<4.8%) was found in the concentration of formazan (absorbance per unit thickness) when the same cell was measured in serial cross-sections of various thicknesses (2–10 m) along a longitudinal distance of at least 200 m along the cell. The reduction in enzyme activity was found to be negligible after aqueous preincubation. A maximum of 10–12% of the formazan produced in the NADH-TR reaction might be the result of nothing dehydrogenase activity, whereas this unspecific reaction might account for up to 20% of the formazan deposited in the -GPD reactions after 30 min incubation. The diffusion of Nitro BT into the tissue during the incubation period was found to be unhindered. The rates of formazan production decreased with increasing incubation time, especially in the -GPD reaction in both fibre types. The ratio of the mean absorbance of the formazan in type I fibres to that in type II fibres (in the same section) was 1.41 (coefficient of variation, 2.5%) in the NADH-TR reaction and 0.68 (coefficient of variation, 3.8%) in the -GPD reaction. These values were not affected either by variations in the incubation time (5–40 min) or by the thickness of the section (2–8 m). The concentrations of NADH-TR and -GPD seem to be constant along the length of the muscle fibre. The histochemical reactions reported, together with measurements of the thickness of the sections, seem suitable for the microphotometric quantification of the two enzymes in single fibres of human skeletal muscles.     

Effects of con A and other lectins on pure 5'nucleotidase isolated from lymphocyte plasma membranes.

Inhibition of purified or membrane-bound 5′nucleotidase by various lectins was studied in lymphocytes from pig mesenteric lymph nodes. Con A or $\text{Lens culinaris}$ lectin LcH inhibited (75 %) purified 5′nucleotidase by a non-competitive process without cooperativity. Inhibition by these lectins of 5′ nucleotidase activity in whole lymphocytes, plasma membranes (untreated or solubilized) and LcH-receptor fraction displayed high positive cooperativity, reached higher level (90 %) and was of mixed type. An interaction between lectin receptors and 5′nucleotidase accounted for these differences. Wheat germ agglutinin (WGA) and divalent Con A which are not mitogenic for T lymphocytes had no effect on 5′nucleotidase; pokeweed mitogen (PWM), mitogen of T and B cells, was not inhibitor. When membrane proteins were cross-linked by glutaraldehyde, Con A inhibition of whole lymphocyte 5′nucleotidase presented the same properties as the purified enzyme. Possible correlation between 5′nucleotidase inhibition and lymphocyte stimulation is discussed.     

Host plant discrimination within Cruciferae: Feeding responses of four leaf beetles (Coleoptera: Chrysomelidae) to glucosinolates, cucurbitacins and cardenolides

Feeding responses of four Chrysomelidae to six less acceptable plants and to compounds from them were investigated by means of leaf disc tests. Significant differences were found between responses of different species, and plants containing potent feeding inhibitors were always rejected. Cucurbitacins are potent feeding inhibitors to Phyllotreta nemorum, and this species does not eat Iberis species containing these compounds. Cardenolides are potent feeding inhibitors to P. undulata, P. tetrastigma and Phaedon cochleariae, and these three species do not eat the cardenolide containing Cheiranthus and Erysimum.Six different glucosinolates all proved to be stimulatory when applied to pea leaf discs. Although the glucosinolates differed somewhat in their ability to stimulate feeding, no correlation is found between content of glucosinolates and acceptability of the investigated plants. Application of sinigrin to Iberis and Cheiranthus did not improve their acceptability. The presence of glucosinolates is necessary for feeding to occur, but it is less important which glucosinolates are present.Cardenolides and cucurbitacins are suggested to be a second generation of protective compounds in Cruciferae, glucosinolates being the first.

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Zusammenfassung Der Einfluss einiger sekundärer Pflanzenstoffe aus Cruciferen auf die Futteraufnahme von vier Chrysomeliden, die auf dieser Pflanzenfamilie vorkommen, wurde mittels Blattscheiben-Tests untersucht. Cucurbitacine sind starke Frasshemmstoffe für Phyllotreta nemorum, weniger starke Hemmstoffe für P. undulata und schwache Hemmstoffe für P. tetrastigma und Phaedon cochleariae. Iberis-Arten, die Cucurbitacine enthalten, werden von P. nemorum und P. undulata abgelehnt, von den beiden anderen Arten aber akzeptiert. Cardenolid-Glykoside vom Strophanthidin-Typ sind starke Frasshemmstoffe für P. undulata, P. tetrastigma und Phaedon cochleariae. Diese Arten lehnen Cheiranthus-und Erysimum-Arten, die solche Stoffe enthalten, ab. Die Futteraufnahme von P. nemorum wird von diesen Stoffen nicht beeinflusst; P. nemorum akzeptiert Cheiranthus- und Erysimum-Arten.Futteraufnahme fand bei Abwesenheit von Senfölglukosiden nicht statt. Sechs verschiedene Senfölglukoside waren alle imstande, das Aufnehmen von Erbsen-Blattscheiben zu stimulieren. Gewisse Unterschiede in der stimulierenden Wirkung der einzelnen Glukoside wurden gefunden. Das Vorkommen bestimmter Glukoside und die Akzeptabilität der Pflanzen zeigten aber keine Korrelation. Anwesenheit oder Abwesenheit von Frasshemmstoffen beeinflusst die Akzeptabilität der Pflanzenarten mehr als die Anwesenheit bestimmter Senfölglukoside.Wenn Senfölglukoside als eine erste Generation von Abwehrstoffen in Cruciferen aufgefasst werden, können Cucurbitacine in Iberis und Cardenolid-Glykoside in Cheiranthus und Erysimum als eine zweite betrachtet werden.

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The Danish Natural Science Research Council supported the research.     

Glycogenosis in the Dog

Four cases of a generalized form of glycogenosis occurring in German Shepherd dogs, all females, are described. Symptoms could be noticed as early as the age of two months and progressed slowly for months. They appeared as dizziness, muscular weakness, and in two of the cases as poor nutritional state. The abdomen became gradually distended. The main lesion seen at postmortem was a greatly increased liver size with some moderate liver fibrosis. Heavy deposits of a granular substance behaving as glycogen in histochemical tests and at electron microscopy were found in the hepatic cells, muscle fibres of the heart, skeletal and smooth muscles, and in nerve and glial cells of the central nervous system. The substance was lying freely dispersed in the cell cytoplasm without any indication of lysosomal storage. The disease of dogs does not seem to be fully comparable with any of the types observed in man, but is probably much related to Type III or Cori's Disease. Structure analysis of the deposits and enzyme investigations have been done and are published (Čeh et al. 1976).     

The interphase microtubule aster is a determinant of asymmetric division orientation in Drosophila neuroblasts

The mechanisms that maintain the orientation of cortical polarity and asymmetric division unchanged in consecutive mitoses in Drosophila melanogaster neuroblasts (NBs) are unknown. By studying the effect of transient microtubule depolymerization and centrosome mutant conditions, we have found that such orientation memory requires both the centrosome-organized interphase aster and centrosome-independent functions. We have also found that the span of such memory is limited to the last mitosis. Furthermore, the orientation of the NB axis of polarity can be reset to any angle with respect to the surrounding tissue and is, therefore, cell autonomous.     

Caffeine Junkie: an Unprecedented Glutathione S-Transferase-Dependent Oxygenase Required for Caffeine Degradation by Pseudomonas putida CBB5

Caffeine and other N-methylated xanthines are natural products found in many foods, beverages, and pharmaceuticals. Therefore, it is not surprising that bacteria have evolved to live on caffeine as a sole carbon and nitrogen source. The caffeine degradation pathway of Pseudomonas putida CBB5 utilizes an unprecedented glutathione-S-transferase-dependent Rieske oxygenase for demethylation of 7-methylxanthine to xanthine, the final step in caffeine N-demethylation. The gene coding this function is unusual, in that the iron-sulfur and non-heme iron domains that compose the normally functional Rieske oxygenase (RO) are encoded by separate proteins. The non-heme iron domain is located in the monooxygenase, ndmC, while the Rieske [2Fe-2S] domain is fused to the RO reductase gene, ndmD. This fusion, however, does not interfere with the interaction of the reductase with N₁- and N₃-demethylase RO oxygenases, which are involved in the initial reactions of caffeine degradation. We demonstrate that the N₇-demethylation reaction absolutely requires a unique, tightly bound protein complex composed of NdmC, NdmD, and NdmE, a novel glutathione-S-transferase (GST). NdmE is proposed to function as a noncatalytic subunit that serves a structural role in the complexation of the oxygenase (NdmC) and Rieske domains (NdmD). Genome analyses found this gene organization of a split RO and GST gene cluster to occur more broadly, implying a larger function for RO-GST protein partners.     

The holarctic complex Potentilla fruticosa (Rosaceae)

Potentilla fruticosa L. (Rosaceae) is a complex of large morphological, karyological, geographical and ecological amplitudes. A survey of the distribution of the morphological variation is given and the underlying reasons discussed. Migration paths are also discussed. In southeast central Asia there is a complex in need of further investigation. Outside this area two species should be recognized, a diploid and hermaphrodite species, P. floribunda Pursh, in N America, S Europe and Siberia and a tetraploid and dioecious species, P. fruticosa L., in N Europe and Siberia.     

Brewers’ spent grain liquor as a feedstock for lactate production with Lactobacillus delbrueckii subsp. lactis

Brewers’ spent grain (BSG) is a low‐cost by‐product of the brewing process. BSG liquor names the liquid components of BSG, mainly glucose, maltose, and long‐chain α‐1,4‐glycosidic bond glucose oligomers. These substances should be separated in existing BSG biorefineries, as they might lead to an increased formation of microbe‐inhibiting compounds in well‐established hydrothermal/enzymatic saccharification processes. In most cases, this liquid fraction is discarded. The present study presents for the first time an optimized process with BSG liquor for the purpose of producing bulk chemicals (e.g., lactate) in relevant concentrations. The process comprises the application of yeast extract, produced from own brewing processes, as the sole supplemented complex constituent in a simultaneous fermentation and saccharification process. Kinetic parameters for the final optimized process conditions with the organism Lactobacillus delbrueckii subsp. lactis were: maximum specific growth rate µ_max  =  0.47 h^?1, maximum lactate concentration c_Lac, max  =  79.06 g L^?1, process yield Y_PS  =  0.89 g_Lac g_Sugar^?1, lactate production rate q_P  =  4.18 g_Lac g_CDW^?1 h^?1, and productivity P _15 h  =  4.93 g_Lac L^?1 h^?1. BSG liquor, linked with yeast extract from Brewers’ yeast, can be a promising substrate for further bioprocess engineering tasks and contribute to a holistic and sustainable usage of Brewers’ spent grain.