Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.3.5 (5'-nucleotidase)
 3,167 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In the previous paper we presented findings which indicated that enzyme heterogeneity exists among PMN leukocyte granules. From histochemical staining of bone marrow smears, we obtained evidence that azurophil and specific granules differ in their enzyme content. Moreover, a given enzyme appeared to be restricted to one of the two types. Clear results were obtained with alkaline phosphatase, but those with a number of other enzymes were suggestive rather than conclusive. Since the approach used previously was indirect, it was of interest to localize the enzymes directly in the granules. Toward this end, we carried out cytochemical procedures for five enzymes on normal rabbit bone marrow cells which had been fixed and incubated in suspension. The localization of reaction product in the granules was determined by electron microscopy. In accordance with the results obtained on smears, azurophil granules were found to contain peroxidase and three lysosomal enzymes: acid phosphatase, arylsulfatase, and 5'-nucleotidase; specific granules were found to contain alkaline phosphate. Specific granules also contained small amounts of phosphatasic activity at acid pH. Another finding was that enzyme activity could not be demonstrated in mature granules with metal salt methods (all except peroxidase); reaction product was seen only in immature granules. The findings confirm and extend those obtained previously, indicating that azurophil granules correspond to lysosomes whereas specific granules represent a different secretory product.
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PMID:Differences in enzyme content of azurophil and specific granules of polymorphonuclear leukocytes. II. Cytochemistry and electron microscopy of bone marrow cells. 569 83

Two human monocyte subsets from the peripheral blood of healthy donors have been isolated in greater than 90% purity by countercurrent centrifugal elutration and human serum albumin gradients and their functional capabilities have been assessed. We have demonstrated that one subset ("regular" monocytes, RM) showed intense cytoplasmic peroxidase staining and contained substantial peroxidase activity. In contrast, another subset ("intermediate" monocytes, IM) stained poorly for peroxidase and had low peroxidase activity. By electron microscopic analysis combined with peroxidase localization, it was found that IM had fewer peroxidase-positive granules per cell than did RM. IM coelutriated with some lymphocytes and by cell sizing analysis were shown to be slightly smaller than RM. Functional and cytochemical analysis of these subsets indicated that IM had less activity than RM in assays such as accessory cell function for mitogen-induced T lymphocyte proliferation and antibody-dependent cellular cytotoxicity, and that fewer IM expressed OKM1 antigen and pokeweed mitogen (PWM) receptors on their membranes than did RM. The subset of IM not bearing either the PWM receptor or the OKM1 antigen had very low peroxidase activity. IM also were found to have a greater sensitivity to polyriboinosinic and polyribocytidilic acid (100 micrograms/ml)-induced secretion of interferon. There was no significant difference in the phagocytic capability, the percentage of Fc receptor-positive cells, 5'-nucleotidase activity, DR antigen expression, or the responsiveness to migration inhibitory factor of IM as compared with RM. Furthermore, it was found that the ratio of IM to RM increased after prolonged cytapheresis, which suggests that IM are more mobilizable than RM from the extravascular reservoirs of human monocytes.
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PMID:Characterization of a human blood monocyte subset with low peroxidase activity. 619 41

The use of the Romanowsky staining technique, Sudan Black B, the periodic acid-Schiff reaction and methods for revealing peroxidase, acid and alkaline phosphatases and butyrate, acetate or chloroacetate esterases for identifying and discriminating subvarieties of acute leukaemia at the light microscope level is reviewed and the results of their application in a recent study of the first 720 cases admitted to the Medical Research Council's 8th Acute Myeloid Leukaemia trial summarized. The distribution of varieties of acute myeloid leukaemia and the relevance of age and cytochemical findings to clinical prognosis is presented. Identification of the predominant primitive cell - myeloblast, promyelocyte, monoblast, and others - appears to have little prognostic significance. In fact, the presence of periodic acid-Schiff positive erythroblasts is a bad prognostic sign. The association of certain cytochemical findings with the 15;17 translocation and acute promyelocytic leukaemia, especially the patterns of esterase positivity in Auer rods and the Sudan Black, peroxidase, periodic acid-Schiff and esterase findings characteristic of the 8;21 translocation are illustrated. Cytochemical features helpful in distinguishing acute megakaryoblastic leukaemia, notably the periodic acid-Schiff reaction, differential esterase reactivities and 5'-nucleotidase, are discussed and illustrated. Brief reference is made to the cytochemical differentiation of lymphoblastic leukaemias. Details of a technical method for the demonstration of 5'-nucleotidase are given.
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PMID:Cytochemistry of the acute leukaemias. 620 45

We describe a one-step kinetic method for the determination of 5'-nucleotidase (EC 3.1.3.5). Inosine is formed by the hydrolysis of inosine 5'-monophosphate which is catalyzed by seric 5'-nucleotidase, and then is converted to hypoxanthine by nucleoside phosphorylase. Two moles of hydrogen peroxide are formed for each mole of hypoxanthine oxidized to urate by xanthine oxidase. The rate formation of hydrogen peroxide is monitored at 510 nm using the oxidation of the chromogenic system 3,5-dichloro-2-hydroxybenzenesulfonic acid/4-aminophenazone in the presence of peroxidase. beta-Glycerophosphate inhibits the unspecific cleavage of the substrate by alkaline phosphatases. Inorganic phosphate is added to improve the reagent stability, and ferrocyanide to reduce bilirubin interference. Automation of the technique requiring 20 microliter of serum on a centrifugal analyzer is also described.
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PMID:A one-step determination of serum 5'-nucleotidase using a centrifugal analyzer. 627 35

Rat embryo fibroblasts cultured in the presence of monensin exhibited an inhibited uptake of horseradish peroxidase. The inhibition was detected after 3 h, after which time the cells became increasingly vacuolated; the concentration of monensin required to inhibit pinocytosis (0.4 microM for half-maximum inhibition at 18 h) was similar to that found by others to inhibit secretion. Both the exchange of 5'-nucleotidase between the membranes of cytoplasmic organelles and the cell surface and the internalization of anti-5'-nucleotidase bound to the cell surface were inhibited by approximately 90% in monensin-treated cells. The effects of monensin were reversible: cells cultured first with monensin, and then in fresh medium, exhibited control levels of horseradish peroxidase uptake, exchange of 5'-nucleotidase, and internalization of anti-5'-nucleotidase bound to the cell surface. After monensin treatment, the median density of both galactosyl transferase and 5'-nucleotidase increased from 1.128 to 1.148, and the median density of both N-acetyl-beta-glucosaminidase and horseradish peroxidase taken up by endocytosis decreased from 1.194 to 1.160. The results indicate that monensin is a reversible inhibitor of pinocytosis and, presumably, therefore, of membrane recycling. They suggest that the inhibition of membrane recycling occurs at a step other than the fusion of pinocytic vesicles with lysosomes and is perhaps a consequence of an effect of the ionophore on the Golgi complex.
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PMID:Inhibition of pinocytosis in rat embryo fibroblasts treated with monensin. 628 96

Alkaline phosphatase (AP) is one of the enzymes which is highly active in the plasmalemma of endothelial cells (ECs) of BBB-type microvessels. In the ECs of non-BBB type vessels, the reaction for AP (and other phosphatases) is negative (e.g. choroid plexus, area postrema, hypophysis). After BBB damage, the leakage of the vessels can be demonstrated by the use of horseradish peroxidase (HRP). Concomitantly, changes in polar distribution of AP in the ECs occur, paralleled by the appearance of numerous pinocytic vesicles, deep invaginations of the plasmalemma and channel-like structures. The delimiting membranes of these structures possess AP, 5'-nucleotidase, nucleoside diphosphatase and Na+, K+-ATPase activities. These observations suggest that the redistribution of plasmalemma bound enzymes from luminal to abluminal surface results from membrane flow associated with formation of pinocytic vesicles and channel-like structures in affected ECs. In the area of brain where the process of resolution of brain edema occurs, the shift of the enzymatic activity from luminal to abluminal plasmalemma of the ECs is observed probably because of the need to remove various solutes present in the edematous fluid. The appearance of positive reaction for AP in the abluminal side of the EC can be a reflection of the changed functional polarity of these cells associated with reverse transport of solutes from brain, back into the blood stream.
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PMID:Enzyme cytochemistry of blood-brain barrier (BBB) disturbances. 630 82

After stimulation of the mouse peritoneal cavity with newborn calf serum (NBCS), four types of monocyte and macrophage were distinguished on the basis of peroxidase (PO) patterns. Cytochemically, these cells showed strong heterogeneity in 5'-nucleotidase (5'N) activity. Monocytes and monocyte-derived macrophages with PO activity in granules lacked 5'N activity. Resident macrophages (with PO activity in RER and nuclear envelope) generally had significant 5'N activity on the plasma membrane, the pattern showing close correlation with the biochemical findings. The group of PO-negative macrophages comprised both 5'N-negative and 5'N-positive cells. These findings suggest two possibilities, i.e., that monocytes (5'N-)transform via PO-negative cells (5'N -/+) into resident macrophages (5'N +), or that the monocytes and monocyte-derived macrophages and the resident macrophages represent separate lineages. The fourth type of macrophage, the exudate-resident cell (with PO activity both in granules and in the RER and nuclear envelope), occurred only in low numbers and very late after NBCS stimulation, and is therefore considered not to be a transitional cell between monocytes and resident macrophages.
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PMID:Heterogeneity in 5'-nucleotidase activity of mouse peritoneal macrophages. An EM-cytochemical and biochemical study. 631 16

In the present report the characteristics of nonepithelial phagocytic cells of the murine thymic reticulum are described. Primary cultures were established from thymic fragments. Nonadherent cells with hairy membranes proliferated on the surface of established primary monolayers. These cells were recovered and replated in secondary cultures were they appeared as large adherent cells with dendritic shape. At the electron microscopic level, phagocytic cells of the thymic reticulum in culture (P-TR-C) appear as clear vacuolated cells with an indented nucleus and few lysosomes; this morphological aspect makes them different from the common macrophage, despite their phagocytic capacity. P-TR-C are positive for nonspecific esterase, acid phosphatase which is found in the few lysosomes present, 5'-nucleotidase and alpha-D-mannosidase, but negative for peroxidase. A high proportion of alpha-mannosidase-positive cells is inconsistent with the common macrophage, but in common with other cells with dendritic shape such as Langerhans cells. They are Thy-1-, Ig- and nearly half of them are IA+. P-TR-C can be defined as the stimulator cells for syngeneic stimulation; they are able to induce the proliferation of lymphocytes enriched in mature syngeneic medullary thymocytes, but not in immature cortical ones. Characteristics of P-TR-C make them very similar to the interdigitating cells described in the peripheral lymphoid organs and in the thymus in situ.
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PMID:Thymic reticulum in mice. II. Culture and characterization of nonepithelial phagocytic cells of the thymic reticulum: their role in the syngeneic stimulation of thymic medullary lymphocytes. 660 Oct 9

A membrane fraction denoted N2 upper was isolated from homogenates of rat liver by sucrose gradient centrifugation. This fraction, which was enriched 65-fold over the homogenate in 5'-nucleotidase activity, was used as an immunogen in goats. The antisera obtained contained antibodies to three predominant polypeptides in the N2 upper membrane fraction, as shown by crossed immunoelectrophoresis. These polypeptides had molecular weights of 105,000, 110,000, and 160,000 after recovery from the crossed immunoelectrophoretic gels and are denoted PM105, PM110, and PM160. Each was a distinct polypeptide, as shown by the distinct peptide patterns resulting from limited proteolysis in the presence of detergents. The three polypeptides were synthesized by primary cultures of hepatocytes and were externally oriented at the surface of these cells, as shown by their accessibility in situ to iodination catalyzed by lactoperoxidase. They were not detectable in the serum by crossed immunoelectrophoresis. The three antigens were present at very low (PM110) or nondetectable (PM105, PM160) concentrations in intracellular membrane fractions derived from the Golgi and smooth and rough endoplasmic reticulum of liver. The antigens also were reduced in concentration in a plasma membrane fraction most likely derived from the sinusoidal surface of the hepatocyte. The three membrane antigens bind to concanavalin A; hence, they are probably glycoprotein constituents of a discrete domain of the hepatocyte plasma membrane. Immune complexes were isolated after crossed immunoelectrophoresis and injected into rabbits. Each of the antisera obtained was reactive to one of the membrane polypeptides. Sections of fixed rat livers were reacted with each of the antibodies and then the primary antibody was localized by indirect immunocytochemical methods using horseradish peroxidase or colloidal gold as labels. Each of the three antigens was localized by this method to the bile canalicular domain of the hepatocyte plasma membrane.
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PMID:Establishment of plasma membrane domains in hepatocytes. I. Characterization and localization to the bile canaliculus of three antigens externally oriented in the plasma membrane. 664 79

Rat liver organelles involved in receptor-mediated endocytosis were labeled with a conjugate of galactosylated BSA to horseradish peroxidase [( 3H]galBSA-HRP), injected 10 min before sacrifice. These organelles were recovered at low density (1.11-1.13 g/ml) in sucrose gradients (Quintart, J., P. J. Courtoy, J. N. Limet, and P. Baudhuin, 1983, Eur. J. Biochem., 131:105-112). Upon incubation of such low density fractions in 3,3'-diaminobenzidine (DAB) and H2O2 and equilibration in a second sucrose gradient, galBSA-HRP-containing particles selectively shifted towards heavier densities (Courtoy, P. J., J. Quintart, and P. Baudhuin, 1984, J. Cell Biol., 98:870-876, companion paper), resulting in up to 250-to 300-fold purification with respect to the homogenate. The most purified preparations, wherein DAB-stained structures represented approximately 85% of the total volume of particles, contained only trace activities of enzymes usually regarded as markers for other subcellular entities. These minor activities could reflect either contamination or true enzyme association to the ligand-containing structures. Considering the latter hypothesis, at most 1.0% of alkaline phosphodiesterase I and 2.6% of 5'-nucleotidase (markers for plasma membrane), 3.6% of N-acetyl-beta-glucosaminidase (lysosomes), and 6.0% of galactosyltransferase (Golgi complex) from the homogenate would be associated with the whole population of ligand-containing organelles. After DAB cytochemistry on liver fixed 10 min after galBSA-HRP injection, ligand-containing structures accounted for 0.78-0.89% of the fractional volume of the hepatocytes and displayed a membrane area of 2,100 cm2/cm3, compared with 6,700 cm2/cm3 for the pericellular membrane. Altogether, our data support the hypothesis that these ligand-containing organelles are structurally distinct from plasma membrane, lysosomes, and Golgi complex.
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PMID:Receptor-mediated endocytosis in rat liver: purification and enzymic characterization of low density organelles involved in uptake of galactose-exposing proteins. 669 90


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