The role of NADPH in the regulation of glucose-6-phosphate and 6-phosphogluconate dehydrogenases in rat adipose tissue

Summary Previous studies examining the regulation of the synthesis of G6PDH and 6PGDH in rat liver and adipose tissue have focused on the induction of these enzymes by different diets and some hormones. In rat liver these enzymatic activities seem to be regulated by a mechanism involving changes in the NADPH requirements. In this paper we have studied the effect of changes in the flux through different NADPH-consuming pathways on G6PDH and 6PGDH levels in adipose tissue and on the NADPH/NADP ratio. The results show that: I) an increase in the consumption of NADPH, caused by the activation of either fatty acid synthesis or detoxification systems which consume NADPH, is paralleled by an increase in the levels of these enzymes; II) when the increase in consumption of NADPH is prevented, the G6PDH and 6PGDH levels do not change.Abbreviations G6PDH Glucose-6-Phosphate Dehydrogenase - 6PGDH 6-Phosphogluconate Dehydrogenase - GR Glutathione Reductase - ME Malic Enzyme - tBHP t-Butyl Hydroperoxide - NF Nitrofurantoin - CumOOH Cumene Hydroperoxide     

Elutriation of human keratinocytes and melanocytes fromin vitro cultured epithelium

Summary Human epidermal keratinocytes grown in culture and at different stages of differentiation are shown to be viably separated by elutriation. A specific fraction enriched in melanocytes was obtained. Elutriation of cells obtained fromin vitro cultured epithlium could prove useful in studies concerning the biochemistry and molecular markers of cells isolated from normal epithelium and from different pathologies.     

Polyamine Metabolism and Osmotic Stress : II. Improvement of Oat Protoplasts by an Inhibitor of Arginine Decarboxylase

We have attempted to improve the viability of cereal mesophyll protoplasts by pretreatment of leaves with dl-α-difluoromethylarginine (DFMA), a specific `suicide' inhibitor of the enzyme (arginine decarboxylase) responsible for their osmotically induced putrescine accumulation. Leaf pretreatment with DFMA before a 6 hour osmotic shock caused a 45% decrease of putrescine and a 2-fold increase of spermine titer. After 136 hours of osmotic stress, putrescine titer in DFMA-pretreated leaves increased by only 50%, but spermidine and spermine titers increased dramatically by 3.2- and 6-fold, respectively. These increases in higher polyamines could account for the reduced chlorophyll loss and enhanced ability of pretreated leaves to incorporate tritiated thymidine, uridine, and leucine into macromolecules. Pretreatment with DFMA significantly improved the overall viability of the protoplasts isolated from these leaves. The results support the view that the osmotically induced rise in putrescine and blockage of its conversion to higher polyamines may contribute to the lack of sustained cell division in cereal mesophyll protoplasts, although other undefined factors must also play a major role.     

Differentiation of U-937 human monocyte-like cell line by 1α,25-dihydroxyvitamin D3 or by retinoic acid : Opposite effects on insulin receptors

The monocyte-like human cell line U-937 has been differentiated in vitro by incubation with either 1 alpha,25-dihydroxyvitamin D3 or retinoic acid (RA) plus dibutyryl cyclic AMP (db-cAMP). Both methods were effective in inducing the appearance of maturation markers. Their actions on insulin receptors were the opposite, however; 1 alpha,25-dihydroxyvitamin D3 increased the binding of the hormone, while RA plus db-cAMP decreased the binding. These effects were specific for insulin, since the transferrin receptors were reduced by both methods of differentiation. Thus, the changes in insulin receptors during maturation in vitro depend on the inducing agent and are not causally related to the differentiation process.     

Inhibition of transferrin receptor expression by interferon-alpha in human lymphoblastoid cells and mitogen-induced lymphocytes

125I-Transferrin binding to lymphoblastoid K562 and Daudi cells markedly increased after exposure of the cells to culture conditions that stimulated proliferation. Treatment of these cells with interferon-alpha (IFN-alpha) resulted in concurrent inhibition of cell growth and of the rise in transferrin binding. Scatchard analyses revealed that IFN reduced the number of transferrin receptors without altering the binding constant. When 125I-transferrin binding was measured using permeabilized cells, the IFN-induced reduction of binding was comparable to that observed with intact cells, indicating that IFN diminished the total number of cellular transferrin receptors. We also found that addition of IFN-alpha to phytohemagglutinin-stimulated human lymphocytes inhibited the mitogen-induced enhancement of [3H]thymidine incorporation as well as surface binding of 125I-transferrin. Our findings suggest that the decrease in transferrin receptor expression on IFN-alpha-treated cells may be one of the mechanisms responsible for the antiproliferative action of IFN.     

Rapid method for evaluating intra-erythrocyte Na+ activity

The measurements of intracellular "Na+ activity" was performed in 10 ml of heparinized venous blood. First the blood was three times washed in isotonic magnesium chloride solution (114 mmol/l). Thereby the buffy coat was removed. Then the microhematrocrit was taken for packet cell volume determination. After the erythrocytes were lysed by ultrasound. Sodium "Na+ Activity" is measured in the hemolysate by Ion-Selective electrode. With this method all "pipetting" operations are eliminated and for the "Na+ activity" determination was used ion-selective electrode with an indirect measurements, which is less influenced by the matrix. Reference intervals determined for a healthy population were 7.3 +/- 0.6 mmol/l.     

Modulation of natural killer activity by thymosin alpha 1 and interferon

Summary A single injection of -interferon (-IFN) (30 000 units/mouse), a major biological modifier of natural killer (NK) cytolytic activity, strongly stimulated NK activity in normal mice, as expected, while the same treatment did not statistically alter the NK response in cyclophosphamide (CY)-suppressed animals.We investigated the possibility of thymosin ₁ cooperating with -IFN in boosting NK activity in CY-suppressed animals.The results show that treatment with thymosin ₁ (200 g/kg) for 4 days, followed by a single injection of -IFN 24 h before testing, strongly restored NK activity in CY-suppressed mice. Thymosin ₁ was, moreover, able to accelerate the recovery rate of NK activity in bone marrow reconstituted murine chimeras.Taken together the data support the concept that the synergic effect between thymosin ₁ and -IFN could be the result of effects on differentiation of the NK lineage at different levels.     

Action ofN-methyl-N′-nitro-N-nitrosoguanidine inRhizobium trifolii

Rhizobium trifolii was highly resistant to the lethal effect ofN-methyl-N-nitro-N-nitrosoguanidine (MNNG), but it was sensitive to the mutagenic action of this chemical. A concentration of 500g/ml yields a survival of between 1% and 10%, which allows us to obtain a higher number of mutants than lower concentrations that yield higher survival rates. Lethal damage produced by nitrosoguanidine was repaired, and repair is inhibited by acriflavine.