Renal enzymes during experimental diabetes mellitus in the rat. Role of insulin, carbohydrate metabolism, and ketoacidosis

The activities of various ammoniagenic, gluconeogenic, and glycolytic enzymes were measured in the renal cortex and also in the liver of rats made diabetic with streptozotocin. Five groups of animals were studied: normal, normoglycemic diabetic (insulin therapy), hyperglycemic, ketoacidotic, and ammonium chloride treated rats. Glutaminase I, glutamate dehydrogenase, glutamine synthetase, phosphoenolpyruvate carboxykinase (PEPCK), hexokinase, phosphofructokinase, fructose-1,6-diphosphatase, malate dehydrogenase, malic enzyme, and lactate dehydrogenase were measured. Renal glutaminase I activity rose during ketoacidosis and ammonium chloride acidosis. Glutamate dehydrogenase in the kidney rose only in ammonium chloride treated animals. Glutamine synthetase showed no particular variation. PEPCK rose in diabetic hyperglycemic animals and more so during ketoacidosis and ammonium chloride acidosis. It also rose in the liver of the diabetic animals. Hexokinase activity in the kidney rose in diabetic insulin-treated normoglycemic rats and also during ketoacidosis. The same pattern was observed in the liver of these diabetic rats. Renal and hepatic phosphofructokinase activities were elevated in all groups of experimental animals. Fructose-1,6-diphosphatase and malate dehydrogenase did not vary significantly in the kidney and the liver. Malic enzyme was lower in the kidney and liver of the hyperglycemic diabetic animals and also in the liver of the ketoacidotic rats. Lactate dehydrogenase fell slightly in the liver of diabetic hyperglycemic and NH4Cl acidotic animals. The present study indicates that glutaminase I is associated with the first step of increased renal ammoniagenesis during ketoacidosis. PEPCK activity is influenced both by hyperglycemia and ketoacidosis, acidosis playing an additional role. Insulin appears to prevent renal gluconeogenesis and to favour glycolysis. The latter would seem to remain operative in hyperglycemic and ketoacidotic diabetic animals.     

Human immunodeficiency virus-specific cytotoxic responses of seropositive individuals: distinct types of effector cells mediate killing of targets expressing gag and env proteins.

By using target cells that expressed isolated env, gag, p27nef, or p23vif molecules introduced by recombinant vaccinia viruses containing genes encoding these polypeptides, it was possible to identify env, gag, p27nef, and p23vif as cytolytic target antigens for freshly isolated blood cells from human immunodeficiency virus 1 (HIV-1) seropositive patients. Most of the patients tested (95%) manifested a specific cytotoxic activity against vaccinia virus-env-infected target cells. The env-specific cytotoxic activity was not restricted by the major histocompatibility complex and was not mediated by T lymphocytes, as shown by the absence of blocking effect with an anti-CD3 monoclonal antibody and by the inefficiency of CD3+, CD8+, or CD4+ and CD8+ depletion to reduce the cytotoxic activity against the env-expressing target cells. In the same conditions, the cytotoxic activity specific for gag was abrogated and gag major histocompatibility complex-restricted cytotoxic T lymphocytes were detected in 85% of the subjects tested. Therefore, in a HIV-1 seropositive subject, distinct types of effector cells mediate the lysis of target cells expressing gag and env proteins.     

Substituted N-(2-aminophenyl)-benzamides, (E)-N-(2-aminophenyl)-acrylamides and their analogues: novel classes of histone deacetylase inhibitors

Inhibition of histone deacetylases (HDACs) is emerging as a new strategy in human cancer therapy. Novel 2-aminophenyl benzamides and acrylamides, that can inhibit human HDAC enzymes and induce hyperacetylation of histones in human cancer cells, have been designed and synthesized. These compounds selectively inhibit proliferation and cause cell cycle arrest in various human cancer cells but not in normal cells. The growth inhibition of 2-aminophenyl benzamides and acrylamides against human cancer cells in vitro is reversible and is dependent on the induction of histone acetylation. Compounds of this class can significantly reduce tumor growth in human tumor xenograft models.     

Bird pollination in an angraecoid orchid on Reunion Island (Mascarene Archipelago, Indian Ocean)

BACKGROUND AND AIMS: Although numerous angraecoid orchids in Madagascar display typical sphingophilous syndrome (i.e. white, nectariferous, long-spurred flowers, producing a strong scent at the crepuscule that is attractive to moths), three species of Angraecum in Reunion, belonging to the endemic section Hadrangis, have atypical unscented and short-spurred flowers. The aim of the study was to investigate the implication of plant-pollinator interaction on the evolution of floral morphology of these peculiar island floral forms. METHODS: The flower morphology of A. striatum (one of the three section Hadrangis species) was investigated by performing a set of floral measures, and the reproductive biology was investigated by a set of hand pollination experiments. Natural pollinators were observed by means of a digital video camera. Pollinator efficiency (pollen removal and deposition) and reproductive success (fruit set) were quantified once a week in natural field conditions during the 2005 flowering season (i.e. from January to March). KEY RESULTS: The orchid is self-compatible but requires a pollinator to achieve fruit set. Only one pollinator was observed, the endemic white-eye Zosterops borbonicus (Zosteropidae). These birds perched on inflorescences, and probed most fresh-looking flowers on each plant for nectar. Nectar was both abundant (averaging 7.7 microL) and dilute (averaging 9.7 % sugar in sucrose equivalents). Birds were mostly active between 0830 and 0930 h. Visits to plants were extremely short, lasting from 9 to 27 s. At the study site, 60.9 % of flowers had pollen removed, and 46.4 % had pollinia deposited on stigmas. The proportion of flowers that initiated a fruit averaged 20.6 % in natural conditions. CONCLUSIONS: For the first time, a bird-pollinated orchid is described from a sub-tribe that is mainly specialized for moth pollination. This study documents a morphological shift in flowers in response to pollinator adaptations in the insular context of the Mascarene Archipelago.     

Natural and recombinant fungal laccases for paper pulp bleaching

Three laccases, a natural form and two recombinant forms obtained from two different expression hosts, were characterized and compared for paper pulp bleaching. Laccase from Pycnoporus cinnabarinus, a well known lignolytic fungus, was selected as a reference for this study. The corresponding recombinant laccases were produced in Aspergillus oryzae and A. niger hosts using the lacI gene from P. cinnabarinus to develop a production process without using the expensive laccase inducers required by the native source. In flasks, production of recombinant enzymes by Aspergilli strains gave yields close to 80 mg l^–1. Each protein was purified to homogeneity and characterized, demonstrating that the three hosts produced proteins with similar physico-chemical properties, including electron paramagnetic resonance spectra and N-terminal sequences. However, the recombinant laccases have higher Michaelian (K _m) constants, suggesting a decrease in substrate/enzyme affinity in comparison with the natural enzyme. Moreover, the natural laccase exhibited a higher redox potential (around 810 mV), compared with A. niger (760 mV) and A. oryzae (735 mV). Treatment of wheat straw Kraft pulp using laccases expressed in P. cinnabarinus or A. niger with 1-hydroxybenzotriazole as redox mediator achieved a delignification close to 75%, whereas the recombinant laccase from A. oryzae was not able to delignify pulp. These results were confirmed by thioacidolysis. Kinetic and redox potential data and pulp bleaching results were consistent, suggesting that the three enzymes are different and each fungal strain introduces differences during protein processing (folding and/or glycosylation).     

Transfer of dysbiotic gut microbiota has beneficial effects on host liver metabolism

Gut microbiota dysbiosis has been implicated in a variety of systemic disorders, notably metabolic diseases including obesity and impaired liver function, but the underlying mechanisms are uncertain. To investigate this question, we transferred caecal microbiota from either obese or lean mice to antibiotic‐free, conventional wild‐type mice. We found that transferring obese‐mouse gut microbiota to mice on normal chow (NC) acutely reduces markers of hepatic gluconeogenesis with decreased hepatic PEPCK activity, compared to non‐inoculated mice, a phenotypic trait blunted in conventional NOD2 KO mice. Furthermore, transferring of obese‐mouse microbiota changes both the gut microbiota and the microbiome of recipient mice. We also found that transferring obese gut microbiota to NC‐fed mice then fed with a high‐fat diet (HFD) acutely impacts hepatic metabolism and prevents HFD‐increased hepatic gluconeogenesis compared to non‐inoculated mice. Moreover, the recipient mice exhibit reduced hepatic PEPCK and G6Pase activity, fed glycaemia and adiposity. Conversely, transfer of lean‐mouse microbiota does not affect markers of hepatic gluconeogenesis. Our findings provide a new perspective on gut microbiota dysbiosis, potentially useful to better understand the aetiology of metabolic diseases.     

Assessment of the Mulder and Van Doorn kinetic procedure and rapid centrifugal analysis of UDP-glucuronosyltransferase activities

The optimal experimental conditions of the enzyme assay described by Mulder and Van Doorn (1975, Biochem J. 151, 131-140) for the measurement of UDP-glucuronosyltransferase activities were tested towards structurally different aglycones. This assessment of this assay revealed that addition of Triton X-100 as enzyme activator was necessary because of its apparent inhibitory effects on interfering reactions. Under these conditions, accordance of the data with results published in the literature was obtained. We present for the first time an UDP-glucuronosyltransferase assay adapted on a fast analyser centrifuge which allows a rapid and sensitive measurement of enzyme activity that is very useful for kinetic constant determination, without consuming a large volume of reagents.