Metabolic impact of adenovirus-mediated overexpression of the glucose-6-phosphatase catalytic subunit in hepatocytes

Glucose-6-phosphatase (G6Pase) catalyzes the hydrolysis of glucose 6-phosphate (Glu-6-P) to free glucose and, as the last step in gluconeogenesis and glycogenolysis in liver, is thought to play an important role in glucose homeostasis. G6Pase activity appears to be conferred by a set of proteins localized to the endoplasmic reticulum, including a glucose-6-phosphate translocase, a G6Pase phosphohydrolase or catalytic subunit, and glucose and inorganic phosphate transporters in the endoplasmic reticulum membrane. In the current study, we used a recombinant adenovirus containing the cDNA encoding the G6Pase catalytic subunit (AdCMV-G6Pase) to evaluate the metabolic impact of overexpression of the enzyme in primary hepatocytes. We found that AdCMV-G6Pase-treated liver cells contain significantly less glycogen and Glu-6-P, but unchanged UDP-glucose levels, relative to control cells. Further, the glycogen synthase activity state was closely correlated with Glu-6-P levels over a wide range of glucose concentrations in both G6Pase-overexpressing and control cells. The reduction in glycogen synthesis in AdCMV-G6Pase-treated hepatocytes is therefore not a function of decreased substrate availability but rather occurs because of the regulatory effects of Glu-6-P on glycogen synthase activity. We also found that AdCMV-G6Pase-treated-cells had significantly lower rates of lactate production and [3-3H]glucose usage, coupled with enhanced rates of gluconeogenesis and Glu-6-P hydrolysis. We conclude that overexpression of the G6Pase catalytic subunit alone is sufficient to activate flux through the G6Pase system in liver cells. Further, hepatocytes treated with AdCMV-G6Pase exhibit a metabolic profile resembling that of liver cells from patients or animals with non-insulin-dependent diabetes mellitus, suggesting that dysregulation of the catalytic subunit of G6Pase could contribute to the etiology of the disease.     

Quality of subcutaneous,intermuscular and intramuscular fat tissue using elevated quantities of medium chain fatty acids in pig fattening

The present study comprised fat tissue samples of 46 (partly 23) pig carcasses randomly obtained from each one of four production systems: common fattening, pigs fattened on a low-fat diet and pigs grown on diets enriched with medium chain fatty acids (MCFA) in low or high amounts (0.3% and 3.6% C₈ with C₁₄, respectively). As models for subcutaneous, intermuscular and intramuscular fat tissues, back fat, dissected belly fat tissue and belly meat were used. In all fat tissues, the contents of MCFA were significantly elevated only with the high dietary content of MCFA, with a preferential retention of the MCFA in the storage tissues. In the MCFA supplemented groups, linoleic acid contents were slightly lower in subcutaneous and intermuscular fat as compared to the control group, in the group with the low-fat diet linoleic acid was considerably lower in all tissues. In spite of the only marginal differences in fatty acid pattern, the penetrometrically assessed firmness of backfat as well as the oxidative resistance of back and belly fat were almost twice as high in the high-MCFA group as in the other groups. In the low-fat group, water content of the back fat (16.9%) was higher than the average of the other groups (14.5%). The implications for routing assessment of fat tissue properties at slaughter plants are discussed.     

Reaction of neuronal nitric-oxide synthase with oxygen at low temperature. Evidence for reductive activation of the oxy-ferrous complex by tetrahydrobiopterin

The reaction of reduced NO synthase (NOS) with molecular oxygen was studied at -30 degreesC. In the absence of substrate, the complex formed between ferrous NOS and O2 was sufficiently long lived for a precise spectroscopic characterization. This complex displayed similar spectral characteristics as the oxyferrous complex of cytochrome P450 (lambda max = 416.5 nm). It then decomposed to the ferric state. The oxidation of the flavin components was much slower and could be observed only at temperatures higher than -20 degreesC. In the presence of substrate (L-arginine), another, 12-nm blue-shifted, intermediate spectrum was formed. The breakdown of the latter species resulted in the production of Nomega-hydroxy-L-arginine in a stoichiometry of maximally 52% per NOS heme. This product formation took place also in the absence of the reductase domain of NOS. Both formation of the blue-shifted intermediate and of Nomega-hydroxy-L-arginine required the presence of tetrahydrobiopterin (BH4). We propose that the blue-shifted intermediate is the result of reductive activation of the oxygenated complex, and the electron is provided by BH4. These observations suggest that the reduction of the oxyferroheme complex may be the main function of BH4 in NOS catalysis.     

Isochoric specific heat of sulfur hexafluoride at the critical point: Laboratory results and outline of a spacelab experiment for the d1-mission in 1985

The specific heat at constant volume c_v shows a weak singularity at the critical point. Renormalization group techniques have been applied, predicting a universal critical behavior which has to be experimentally confirmed for different systems. In this paper an experiment is presented to measure the specific heat of SF₆ along the critical isochore (_c=0.737 g·cm^–3), applying a continuous heating method. The results cover a temperature span of –1.5×10^–2< <1.70×10^–2 [=(T–T _c)/T _c] and were strongly affected by gravity effects that emerge in the sample of 1-mm hydrostatic height near the critical point. Using regression analysis, data were fitted with functions of the form c _v/R=A × ¦¦^– + B for the one-phase state and c _v/R=A × ¦¦^– + B for the twophase state. Within their error bounds the critical values (==0.098, A/A=1.83) represent the measurements for the temperature span 3.5×10^{–5< ¦¦<2.0×10 –3}, in good agreement with theoretical predictions. In order to exclude density profiles in the specimen, which are unavoidable in terrestrial experiments due to the high compressibility of fluids at the critical point and the gravity force, a space-qualified scanning ratio calorimeter has been constructed, which will permit long-term c_v measurements under microgravity (-g) conditions. The experiment will be part of the German Spacelab mission in October 1985. The significant features of the apparatus are briefly sketched.Paper presented at the Ninth Symposium on Thermophysical Properties, June 24–27, 1985, Boulder, Colorado, U.S.A.     

Mediator-Related Symptoms and Anaphylaxis in Children with Mastocytosis

Mastocytosis is characterized by the pathological accumulation of mast cells (MC) in various organs. In these patients, MC may degranulate and thereby contribute to clinical symptoms, especially when a concomitant allergy is present. However, MC activation can not only be induced by high-affinity receptors for IgE, but also by anaphylatoxins, neuropeptides, IgG immune complexes, complement-components, drugs, products of bacteria or parasites, as well as physical factors such as heat, cold, vibration, stress, sun, or physical effort. Symptoms due to mediators released by activated MC may develop in adults suffering from systemic mastocytosis, but also evolve in children who usually have cutaneous mastocytosis (CM). Clinically, CM is otherwise characterized by typical brown, maculopapular skin lesions or mastocytoma associated with a positive Darier’s sign. Pruritus and flushing are common and blistering may also be recorded, especially in diffuse CM (DCM). Pediatric patients with mastocytosis may also have gastrointestinal, respiratory, and neurologic complaints. Although anaphylaxis is not a typical finding, pediatric patients with massive skin involvement and high tryptase levels have a relatively high risk to develop anaphylaxis. This paper reviews MC mediator-related symptoms and anaphylaxis in children with mastocytosis, with special emphasis on risk factors, triggers, and management.