Pase Activity Research Articles

The effects of glutathione and adenosine on plasma membrane ATPases of the corneal endothelium. An hypothesis on the stimulatory mechanism of perfused glutathione upon deturgescence.

Reduced glutathione (0.3 mM) stimulates the activity of sodium-potassium activated ATPase (Na+K+ATPase) by 54% in plasma membranes prepared from bovine corneal endothelial cells. Oxidized glutathione, however, has no effect on Na+K+ATPase activity in the same tissue, although it does inhibit magnesium activated ATPase (Mg++ATPase) by approximately 30%. Adenosine neither stimulates nor inhibits either Na+K+ATPase or Mg++ATPase in these plasma membranes. It is postulated that the stimulatory effect of glutathione on deturgescence stems from the direct reaction of the reduced form of the tripeptide on sulfhydryl groups located on plasma membranes of corneal endothelial cells. It is highly probable that these sulfhydryl groups are part of the Na+k+ATPase complex itself.

N-alkaline phosphatase activity in the peripheral blood lymphocytes and serum of a patient with malignant lymphoma and with cytochemically demonstrable lymphocyte alkaline phosphatase

Twenty per cent of the circulating blood lymphocytes of a patient with non-Hodgkin's lymphoma expressed a positive alkaline phosphatase activity on cytochemical examination. The demonstrable alkaline phosphatase activity was localised on large lymphocytes which showed simultaneously scattered butyrate esterase and acid phosphatase positivity, surface membrane Ig, with multiple heavy chain isotypes but predominating κ light chains, and negative mouse erythrocyte rosetting. Alkaline phosphatase activity of the lymphocyte extracts of this patient was elevated 10-fold compared to that of normal lymphocyte extracts and did not catalyse the hydrolysis of cysteamine-S-phosphate, whereas the normal lymphocyte extracts did so with similar efficiency to the hydrolysis of p-nitrophenyl phosphate. These findings suggest that the lymphocytes of this patient contain an alkaline phosphatase (APase) isoenzyme different from that present in normal lymphocyte extracts. A Pase isoenzyme with similar catalytic properties was denoted previously N-phosphatase, N-A Pase. The extract of the patient's lymphocytes migrated on polyacrylamide gel to produce two A Pase activity bands; a slow-migrating weak band corresponding to the A Pase of normal lymphocyte extracts and a second faster-moving band, containing the major part of activity, corresponding to purified N-A Pase derived from Burkitt lymphoma cells. The serum of the same patient had a two-fold elevated A Pase activity compared to the upper limit of normal. N-A Pase composed 65–75% of the total A Pase activity.

Electron Microscopical and Enzyme Histochemical Changes in the Rat Myocardium During Prolonged Autolysis

The effect on rat myocardium of autolysis at 19 degrees C, for up to 20 days, was studied by electron microscopy and enzyme histochemistry. The enzymes studied included monoamine oxidase (MAO), sytochrome oxidase (CytO), non-specific esterase ((Ns.E.), phosphorylase (P-ase), succinate dehydrogenase (SDH) and various NAD- and NADP-linked dehydrogenases. The myocardium lost its histochemical P-ase activity within a few hours of autolysis, whereas the activity of all other enzymes remained quite normal for at least about 4 days, except that of MAO and SDH, which were normal for about 8 and 12 days, respectively. The myocardial cells lost activity of various enzymes in a patchy manner during prolonged autolysis and practically all histochemical enzyme activity disappeared within 20 days. The early period of autolysis was accompanied by rapid ultrastructural changes of myocardial cells. During prolonged autolysis the gross architecture of the myocardium was lost gradually by the 12th to 20th days. Mitochondria were the organelles most resistant to the effects of autolysis, and numerous mitochondria with morphologically solid inner and outer membranes were seen among the totally disintegrated myocardium 20 days after death. The loss of P-ase activity coincided with the loss of glycogen. The loss of MAO, SDH and CytO activities was not closely related to the morphological preservation of mitochondria, but, in accordance with other enzymes, was more closely related to the disintegration of the over-all myocardial structure. The present results showed that the architecture of the myocardium, and especially that of the mitochondria, was surprisingly resistant to the effects of autolysis at room temperature. Also several enzymes of the myocardium other than those examined so far maintained quite stable histochemically demonstrable activity during prolonged autolysis. These observations give support to the possibility of making the diagnosis of myocardial infarction at postmortem more accurate than with the present morphological and histochemical routine methods.

Characterization of the plasma lipoproteins of the genetically obese hyperlipoproteinemic Zucker fatty rat

The plasma lipoproteins of the Zucker fatty rat were characterized with respect to lipid and apoprotein composition, and results were compared with those obtained from lean controls. Information on apoproteins was obtained from gel filtration experiments and electrophoresis on polyacrylamide gels. Very low density lipoproteins (VLDL) were increased several-fold in fatties, and 78% of their mass was triglycerides compared with 60% in the controls. Low density (LDL) and high density (HDL) lipoproteins were increased by a factor of 2, although their compositions were similar to those of the controls. Levels of apoVLDL, apoLDL, and apoHDL were five, two and two times higher, respectively, in the fatties, and the two most rapidly moving subunit peptides on polyacrylamide gels were disproportionately elevated in the apoproteins. The slower of these two bands was present in relatively greater amounts than the faster one in fatties. If the slower peptide is an activator of lipoprotein lipase, analogous to the comparable subunit peptides of human apolipoproteins, plasmas of fatties could contain up to 10 times more lipase activator activity than control plasma. This finding, and the fact that adipose tissue lipoprotein lipase activity of fatties was about 150% of controls, suggests that fatties have increased capacities for VLDL catabolism. We have previously shown that hepatic VLDL secretory rates are higher than normal in these animals. The increased capacity for catabolism may be a response to the altered secretory rates.

Histochemical observations on the enzymes of chicken yolk sac membrane.

Synopsis In the endodermal cells of the yolk sac membrane of chicken embryos incubated for 12 to 18 d, the activity of glucose‐6‐phosphatase (G‐6‐Pase) was high, the activity of acid phosphatase (Acid Pase) was moderate, the activities of lactate dehydrogenase (LDH) and of glyceraldehyde‐3‐phosphate dehydrogenase (G‐3‐PDH) were low, and succinate dehydrogenase (SDH) and alkaline phosphatase (Alk Pase) activities were not detected. On the day before hatching, however, SDH became evident with low activity, and the activities of Acid Pase, LDH and G‐3‐PDH increased and remained higher until the day after. Five days after hatching, there were marked decreases in the activities of all the enzymes except SDH. Alk Pase was not detected at any time. The function of the yolk sac endo‐derm in the absorption of yolk is discussed in the light of these observations.

Histochemical Study of So-called “Marker Enzymes of Cholestasis” During Extrahepatic Bile Duct Obstruction in the Rat

A histochemical study was performed of the enzymes gamma glutamyl transpeptidase, leucine aminopeptidase and non specific alkaline phosphatase in livers of normal rats and of rats with ligated common bile duct. In normal rat liver the activity of GGT is present at the apical border of the lining cells of bile ducts and ductules. During the first days of cholestasis, original bile ducts as well as proliferating ductular structures show a strongly positive reaction; enzyme activity is concentrated at the apical pole and at the lateral membranes of the cells. After obstruction of more than 1 week periportal hepatocytes also present a slight enzyme activity. In normal rat liver LAP shows a weakly positive canalicular pattern. Some portal biliary structures are also positive. After bile duct ligation, the canalicular activity decreases rapidly, whereas the ductular activity shows a strong increase. After incubation for Alk. Pase, normal rat liver shows positivity of some canaliculi in a narrow periportal area. 1 day after obstruction, all the sinusoidal, canalicular and lateral membranes of the parenchymal cells are positive. This positivity decreases from the third day on, but during all the experiment Alk. Pase activity remains higher than in control animals. The correlation of these histochemical results with the changes in serum concentrations of these enzymes described in the literature, further support the hypothesis that the variations in serum levels during extrahepatic obstruction, are directly dependent from variations in liver enzymatic activity; the latter in turn are a result of a stimulating effect of cholestasis, with overproduction and regurgitation of these enzymes into the blood stream. Es wurden histochemische Untersuchungen an der Leber von normalen Ratten und von Ratten nach Unterbindung des Ductus choledochus vorgenommen. Folgende Enzyme wurden untersucht: Gamma-Glutamyl-Transpeptidase (GGT), Leuzin-Aminopeptidase und unspezifische alkalische Phosphatase. In der normalen Rattenleber findet man eine Aktivität der GGT im apikalen Bereich der Epithelien der Gallengänge und Ductuli. Während der ersten Tage der Cholestase zeigen die normalen Gallengänge wie auch die proliferierenden duktulären Strukturen eine stark positive Reaktion. Die Aktivität ist aber nicht nur am apikalen Pol der Zellen sondern auch an den seitlichen Membranen der Zellen nachweisbar. Nach einer Unterbindung von länger als einer Woche zeigen auch die periportalen Leberzellen eine leichte Enzymaktivität. In der normalen Rattenleber ist der Leuzin-Aminopeptidase schwach positiv in den Gallenkanälchen. Auch einige Gallengänge in den Periportalfeldern sind positiv. Nach Gallengangsunterbindung zeigen die Gallenkanälchen eine Abnahme der Aktivität, während die Ductuli eine starke Zunahme der Aktivität aufweisen. Die alkalische Phosphatase zeigt in den Gallenkanälchen eine positive Reaktion in kleinen Periportalfeldern. 1 Tag nach der Unterbindung sind alle sinusoidalen, kanalikulären und lateralen Membranen der Parenchymzellen positiv. Nach 3 Tagen nimmt die Aktivität ab, aber die Aktivität ist immer noch höher als in den Kontrolltieren. Die Korrelation dieser histochemischen Ergebnisse mit Veränderungen der Aktivitäten der Enzyme im Blutserum, wie sie in der Literatur beschrieben wurden, unterstützt die Hypothese, daß die Variationen in den Serumwerten während extrahepatischer Cholestase direkt abhängig sind von der Variation der Aktivität der Enzyme in der Leber. Die Aktivität der Enzyme im Serum ist abhängig von dem Grad der Cholestase, die mit Überproduktion und Abfluß der Enzyme in den Blutstrom einhergeht.

Histochemical evidence for phosphorylase, branching enzyme and glycogen synthetase activities in rat brain

The brains of young adult Sprague-Dawley rats were frozen and sectioned in a cryostat, or cut into blocks 2 mm thick, then incubated to demonstrate activities of phosphorylase ( Pase), phosphorylase and branching enzyme ( Pase + B) and glycogen synthetase (GS). The blocks were then embedded in paraffin and sections from them, as well as cryostat sections, were stained with dilute Gram's iodine or alcoholic PAS. Cryostat sections of brains exposed to combined unilateral ischaemia and hypoxia were either treated in the same way or pre-incubated in the medium for Pase lacking substrate and glycogen primer. Results showed that: 1. (1) Controversy over localization of Pase activity in brain is due to the use of various incubation media which, if devoid of ethanol, demonstrate Pase + B rather than Pase activity. 2. (2) Although localization of enzymic activity is sharp in en bloc incubation, it differs from that in cryostat sections and is not present in all cellular elements; the medium even when perfused does not penetrate sufficiently. Localization in cryostat sections is poor but more complete. 3. (3) GS activity can be demonstrated in cryostat and paraffin sections, and like Pase and Pase + B activities has the same distribution as glycogen. 4. (4) From evidence obtained by pre-incubation of ischaemic-hypoxic brain sections and of exhausted cat gastrocnemius and soleus muscles, it was concluded that criticisms directed towards the validity of demonstration of Pase, Pase + B and GS activities by techniques used by Takeuchi and co-workers and others are unwarranted.

A modification of the serum glucose-6-phosphatase assay

The difficulties in estimating G-6- Pase activity (distinct from non-specific acid and alkaline phosphatases) in serum are discussed. Experiments with pregnant human serum and with rat liver microsomes suggest that G-6- Pase activity can be estimated specifically in serum buffered to pH 6.5, provided 1.4 × 10 −3 BeSO 4 is added as an inhibitor of alkaline phosphatase. A second portion of the same serum, preincubated at pH 5.0 to destroy G-6- Pase activity, is used to estimate acid phosphatase activity. Subtraction of this from the first value results in an estimate of true G-6- Pase activity.

Clinical and experimental studies of acid phosphatase in renal failure

Recently, a high level of acid phosphatase (Acid Pase) activity was observed in diseased tissues of brain, liver, lung and other organs in pathological cases. Also, we detected chemically that Acid Pase activity was elevated in every part of damaged kidneys (cortex, medulla, papilla), especially at pH 5. Kidney tissue supernatant was separated into Enzyme I and Enzyme II components by liquid chromatography on Sephadex G-75. The activity of the Enzyme I component increased to a great extent in every part of the contracted human kidney and in rabbit kidneys damaged by potassium chromate. This enzyme originated from the prostate, its activity was inhibited by tartrate and sodium fluoride, and its molecular weight was more than 70000. Enzyme II derived from erythrocytes, its activity was inhibited only by formaldehyde, and its molecular weight was 10000–20000. Also the Acid Pase activity was increased in the erythrocytes and in the urine of patients suffering from chronic renal failure (BUN more than 100 mg/dl). The Acid Pase activity in the erythrocyte of rabbits after removal of both kidneys was increased. Hence Acid Pase activity seems to be an important factor in the metabolism of the damaged kidney.

Effect of age on alkaline phosphomonoesterase activity in the adrenals of male mice.

AbstractThis investigation was undertaken to examine the effects of aging on alkaline phosphomonoesterase (Alk Pase) activity in the adrenals of a highly inbred strain of C57BL/10 male mice. A total of 140 male mice were assigned to seven main groups and sacrificed at 1, 4, 8, 12, 16, 20 and 24 months of age for biochemical and morphologic evaluations of adrenal Alk Pase activity. The biochemical findings indicated that while aging may result in a decrease of serum and liver Alk Pase activity, enzyme activity in the adrenals of male mice increases to a maximum level at approximately eight months of age and subsequently decreases at each successive age level. The histochemical findings revealed that the highest concentrations of enzyme activity occurred in the fascicular and reticular zones of sexually mature male mice. There were no major variations in zonal distribution with advancing age. Electron microscopy showed Alk Pase activity along membranes of cortical cells and within the subendothelial space.The progressive increase in Alk Pase enzyme activity up to eight months of age, and the subsequent fall in activity during senescence as well as its absence in the adrenals of female mice provides further support for a role of androgen in mobilizing cortical alkaline phosphomonoesterase activity.

Histochemical observation on phosphatase activities of degenerating and regenerating taste buds

AbstractUsing rat's circumvallate papillae, ATPase, alk. Pase and acid Pase of taste buds were observed after the transection of the glossopharyngeal nerve.The taste buds began to disappear after the nerve was cut and were completely lost after ten days. Following the regeneration of the glossopharyngeal nerve, taste buds reappeared from the bottom of the gutters of circumvallate papillae about 25 days after the operation.ATPase was strongly present on the cell membrane of taste bud cells as far as they existed during degeneration and regeneration. Alk. Pase, which is normally localized on the superficial layers of the epithelium overlying the gutters of circumvallate papilla, gradually diminished as the taste buds degenerated and reappeared as the taste buds regenerated; that is, the activity began to diminish three days after the operation, became feeble after ten days and reappeared after 25 days. It is concluded that taste bud cells secrete alk. Pase in the gutters of circumvallate papillae. Acid Pase activity, usually found in the supranuclear portion of taste bud cells, was intensely reactive during degeneration but did not reappear at the early stage of regeneration of taste bud cells.

ATPase Activity of the Erythrocytic Membrane in Anaplasmosis And of Anaplasma marginale.

SummaryThe erythrocytic AT Pase activity of uninfected calves and calves infected with Anaplasma marginale was determined. Samples from infected animals showed increased ATPase activity during the initial appearance of marginal bodies in the erythro-cytes and at the peak of infection. In view of this increase the possibility of enzyme activity in the Anaplasma bodies per se was investigated. Marginal bodies were purified and subjected to sucrose density-gradient sedimentation procedures. It was demonstrated that fractions containing purified marginal bodies possessed significant ATPase activity. The possible relationship between this activity and the pathology of anaplasmosis was discussed.

The effect of cortisone on ribonucleic acid polymerase and ribonuclease during development: Coincidental evidence for the identity of ribonucleic acid polymerase with the “operator” gene

Three maxima in RNA Pase activity occurred during the 4th instar (days 11–20 after hatching) in the larval salivary gland of Sciara coprophila. They occured at days 12–13, 16 and 20. Minima in cytoplasmic RNAase activity occurred on the same days. Feeding cortisone acetate enhanced the 12–14 day maximum but the development of peaks at days 16 and 20 did not occur. Cortisone administration depressed cytoplasmic RNAse activity. Specific nuclear RNA content paralleled RNA Pase activity. The temporal and quantitative parallels between RNA Pase activity and glucose-6-phosphatase activity suggest that RNA Pase serves as the “operator” (in the terms of the Jacob Monod models) for the enzymes hydrolyzing glucose-6-phosphate. The correspondence of RNA Pase maxima with RNAase minima suggests that the control of RNAase is linked to the activity of RNA Pase. Both enzymes apparently contribute to cellular control of RNA and resulting protein synthesis.

Adenosine triphosphatase activity of the vascular system.

SummaryThe AT Pase activity of the vascular system of the dog has been studied, lit has been shown that the enzyme content of -the different arteries enjoys wide variation. The venous system does not contain the enzyme. ATPase activity of the aorta of the rat was not influenced by age or captivity. The aortic ATPase activity of various species of animals is presented.