glutaminase and Chronic-Disease

In chronic metabolic acidosis in the rat, there is increased ammoniagenesis, gluconeogenesis and renal extraction of glutamine with induction of renal phosphate-dependent glutaminase (PDG). Because the stimulus for these changes is not yet clear and also because acute acidosis is the more common clinical problem, the present study deals mainly with the metabolism of glutamine in acute metabolic acidosis. When acute metabolic acidosis is produced in rats by administration of mineral acid or by causing them to swim, thus inducing a severe lactic acidosis, a factor is found in the plasma which stimulates renal glutamine uptake and ammoniagenesis in vivo as well as in vitro. Acute acidosis does not induce synthesis of PDG in the kidney but causes a change in enzyme kinetics. The plasma factor not only enhances glutamine entry into cells, but apparently causes a conformational change in PDG, as shown by an increase in V1.0mM/Vmax. Intestinal metabolism of glutamine is also stimulated in vivo and in vitro by the plasma factor of acute acidosis.

Topics: Acidosis; Acute Disease; Ammonia; Animals; Chronic Disease; gamma-Glutamyltransferase; Glutamate Dehydrogenase; Glutaminase; Glutamine; Intestinal Mucosa; Kidney; Kidney Tubules; Mitochondria; Physical Exertion

Topics: Acidosis; Ammonia; Animals; Biological Transport; Chronic Disease; Dactinomycin; Deamination; Diazooxonorleucine; Enzyme Inhibitors; Glutamate Synthase; Glutaminase; Glutamine; Humans; Kidney; Mitochondria; Phosphates

Proximal tubules were isolated from control and acidotic rats by collagenase digestion and Percoll density gradient centrifugation. Western blot analysis indicated that the tubules were approximately 95% pure. The samples were analyzed by two-dimensional difference gel electrophoresis (DIGE) and DeCyder software was used to quantify the temporal changes in proteins that exhibit enhanced or reduced expression. The mass-to-charge ratios and the amino acid sequences of the recovered tryptic peptides were determined by MALDI-TOF/TOF mass spectrometry and the proteins were identified using Mascot software. This analysis confirmed the well-characterized adaptive responses in glutaminase (GA), glutamate dehydrogenase (GDH), and phosphoenolpyruvate carboxykinase (PEPCK). This approach also identified 17 previously unrecognized proteins that are increased with ratios of 1.5 to 5.6 and 16 proteins that are decreased with ratios of 0.67 to 0.03 when tubules from 7-day acidotic vs. control rats were compared. Some of these changes were confirmed by Western blot analysis. Temporal studies identified proteins that were induced either with rapid kinetics similar to PEPCK or with more gradual profiles similar to GA and GDH. All of the mRNAs that encode the latter proteins contain an AU sequence that is homologous to the pH response element found in GA mRNA. Thus selective mRNA stabilization may be a predominant mechanism by which protein expression is increased in response to acidosis.

Topics: Acidosis; Adaptation, Physiological; Animals; Blotting, Western; Chronic Disease; Electrophoresis, Gel, Two-Dimensional; Glutamate Dehydrogenase; Glutaminase; Hydrogen-Ion Concentration; In Vitro Techniques; Kidney Tubules, Proximal; Kinetics; Male; Phosphoenolpyruvate Carboxykinase (GTP); Proteins; Proteomics; Rats; Rats, Sprague-Dawley; RNA, Messenger; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

Increased rat renal ammoniagenesis is sustained during chronic metabolic acidosis by the cell-specific induction of the regulatory enzymes of glutamine catabolism and of gluconeogenesis. A glutaminase-specific cDNA hybridizes to 6.0- and 3.4-kb mRNAs that are contained in total or poly(A)+ RNA isolated from rat kidney. When translated in a rabbit reticulocyte lysate, each of the fractionated mRNAs produces the 72-kDa precursor of the mitochondrial glutaminase. The levels of both mRNAs are increased 5-fold within 1 day following onset of chronic acidosis and reach a maximum (8-fold) after 5 days. During recovery from chronic acidosis, the levels of the glutaminase mRNAs are returned to normal within 1 day. The observed changes in mRNA levels correlate with equivalent changes in the relative levels of translatable glutaminase mRNA. Nuclear run-on assays indicate that the rate of transcription of the renal phosphoenolpyruvate carboxykinase gene is increased and decreased in response to onset and recovery from chronic acidosis, respectively. In contrast, the rates of transcription of the glutaminase and beta-actin genes are unaffected by alterations in acid-base balance. Thus, the increase in renal glutaminase activity during chronic acidosis results from an equivalent increase in the levels of total and translatable glutaminase mRNAs which apparently results from an increased stability of the glutaminase mRNA.

Topics: Acidosis; Actins; Animals; Blotting, Northern; Chronic Disease; Enzyme Induction; Glutaminase; Kidney; Kinetics; Male; Phosphoenolpyruvate Carboxykinase (GTP); Rats; Rats, Inbred Strains; RNA, Messenger; Transcription, Genetic

Phosphate- independent glutaminase (PIG) and gamma-glutamyl transpeptidase (gamma GT) activities were evaluated in renal cortex homogenates of rats under normal acid-base balance or chronic metabolic acidosis (CMA). An incubation medium containing glutamine 2 mM, no phosphate, pH 7,40, was used. PIG activity was measured as glutamate production and total gamma GT activity as ammonia production. In normal rats gamma GT activity was significantly higher (0,84 +/- 0,05 mumol/min/g wet wt.) than PIG activity (0,48 +/- 0,06 mumol/min/g wet wt.) (p less than 0,01). In CMA there was a significant increase in PIG activity and an even higher increase in gamma GT activity (p less than 0,01 compared to controls in both cases). The glutamate production/ammonia production ratio was 0,57 +/- 0,05 in normal rats, and 0,37 +/- 0,03 in CMA ( p less than 0,025). These data suggest that the increase in PIG activity and, to a further extent, the increase in gamma GT activity may play an important role in augmenting renal ammonia production in CMA.

Topics: Acidosis; Ammonia; Animals; Chronic Disease; gamma-Glutamyltransferase; Glutaminase; Glutamine; Kidney Cortex; Male; Phosphates; Rats; Rats, Inbred Strains

Topics: Acidosis; Acute Disease; Adaptation, Physiological; Ammonia; Animals; Bicarbonates; Chronic Disease; Female; Glutaminase; Kidney; Phosphates; Rats

1. Slices of duodenum and jejunum produce ammonia from glutamine in vitro. 2. Ammoniagenesis does not increase in response to acidosis or potassium deficiency, two conditions known to cause enhanced ammoniagenesis in the kidney. 3. Gut contains glutaminase 1 as well as gamma-glutamyl transpeptidase. 4. These enzymes do not show any increase during starvation.

Topics: Acidosis; Acute Disease; Ammonia; Animals; Chronic Disease; Duodenum; gamma-Glutamyltransferase; Glutaminase; Glutamine; In Vitro Techniques; Intestine, Small; Jejunum; Organ Specificity; Rats; Starvation

For more exact knowledge on the pathogenesis of pyelonephritis a series of large scale histochemical and biochemical investigations were performed. In this study the author reports his observations on the renal glutaminase I activities in experimental and human pyelonephritis.. Details on materials and methods were given in a recent paper [PHILIPPSON, Exp. Path. 8, 182-193 (1973)]. Glutaminase I activities (L-glutamine amidohydrolase, EC 3.5.1.2.) were quantitatively analyzed according to MATTENHEIMER and DEBRUIN (7) in which the reaction was not only produced by cooling but also by simultaneous inhibition by means of 15% trichloric acetic acid. Ammonia was released and determined by Berthelot's reaction as modified by WELLER (18).. The results in rabbit kidneys are compiled in the tables 1-3, the observations on human kidneys are represented in table 4. 139 rabbits showed unilateral experimental pyelonephritis, in 19 rabbits pyelonephritis did not develop or "healed up spontaneously". 31 samples of pyelonephritic nephrocirrhotic human kidneys (obtained by surgical operations) and 20 samples of healthy human kidneys were investigated. After separation of renal cortex and medulla the cortex was separated into glomerula and renal tubules by differential centrifugation. Significance was tested by universal comparison of the group mean values by t-test (rabbit and human kidneys separately). 1. In the rabbit glomerula glutaminase I (glut I) activities were increased significantly from the beginning of the experiment up to 100 days. They decreased markedly in the subsequent nephrocirrhotic stage (fig. 1). 2. The glut I activities in the rabbit cortical tubules decreased immediately, intensively and succesively up to the late-chronic nephrocirrhotic stage. 3. The rabbit medullary glut I showed likewise intensive decrease in the acute and subacute phase progressing to very low activities in the nephrocirrhotic stages. 4. In all samples from human pyelonephritic nephrocirrhoses (glomerula, cortical tubules and medulla) extremely decreased glut I activities were demonstrated. 5. The enzyme activities observed and the results of the semiquantitative histological analysis [SORGER and co-workers, Exp. Path. 9, 280-287 (1974)] are closely correlated. 6. In the glomerula of the "spontaneously healed up" tissue the glut I activities were markedly increased whereas they were moderately decreased in the cortical tubules and in the medulla. 7. The dry weights and values of protein content were equal to those earlier reported (PHILIPPSON 1973). 8. In the significance test (universally applied t-test) the overwhelming majority of the values showed high significance (p less than or equal to 0.001). The results are correlated with the observations of the semiquantitative histological analysis (SORGER et al. 1974) and discussed on the basis of data reported in literature.

Topics: Animals; Chronic Disease; Glutaminase; Humans; Kidney; Kidney Cortex; Kidney Glomerulus; Pyelonephritis; Rabbits

Topics: Adult; Chronic Disease; Female; Glutaminase; Glutamine; Humans; Male; Middle Aged; Neoplasms; Pregnancy; Transaminases

Topics: Acidosis; Ammonia; Animals; Chronic Disease; Glutamates; Glutaminase; Glutamine; Kidney; Rats

Topics: Acute Disease; Amino Acids; Animals; Asparaginase; Blood; Body Weight; Chromatography, Ion Exchange; Chronic Disease; Diet; Escherichia coli; Female; Glutaminase; Glutamine; L-Lactate Dehydrogenase; Leukemia, Experimental; Mice; Neoplasm Transplantation; RNA Viruses; Virus Diseases