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PubMed1988

The generation of alloreactive cytotoxic T lymphocytes requires the expression of ecto-5'nucleotidase activity.

Ecto-5'-nucleotidase (ecto-5'NT) activity is highly expressed by CTL precursor cells. The aim of this work was to investigate whether ecto-5'NT is involved in their functional activation. The inhibition of ecto-5'NT activity by biochemical (alpha,beta-methyleneadenosine 5-diphosphate; AOPCP) or immunologic inhibitors (polyclonal anti-ecto-5'NT serum) suppressed the proliferative and cytotoxic activation of CTL against a lymphoblastic B cell line in primary one-way MLC. The kinetic analysis of suppression showed that AOPCP and anti-5'NT serum had different effects: AOPCP suppressed only the activation of the cytotoxic program whereas anti-5'NT serum suppressed also recognition and binding to the stimulatory/target cell. Inhibition of CTL generation was not a metabolic consequence of the purine salvage blockade. Incubation of AOPCP-treated MLC with hypoxanthine restored proliferation but not cytotoxicity. The ecto-5'NT inhibitors used had no general toxic effect on cell numbers or viability. AOPCP selectively affected CTL generation and displayed no activity against mitogen-induced proliferation, activation of Ts cells, and generation of IL-2-activated killer cells. These data indicate that ecto-5'NT has a critical role in the functional activation of alloreactive CTL.

5'-Nucleotidase↗
PubMed1985

Distribution of 5'-nucleotidase in the tissues of sheep and the effect of kidney and liver lesions on the activity of the enzyme in plasma and urine.

The distribution of 5'-nucleotidase (5'-NT) activity in the tissues of the sheep differs from that of gamma glutamyl transferase (GGT). Nevertheless, both enzymes are released into the plasma of sheep which have been infected with Fasciola hepatica or in which the bile duct has been ligated. In a sheep dosed with sporidesmin, a fungal toxin which damages the biliary tract, there was an increase in GGT activity in the plasma and urine but no change in 5'-NT activity. Neither enzyme was released into the plasma or urine of sheep dosed with carbon tetrachloride. In sheep with renal tubular necrosis and hepatocellular necrosis caused by dosage with hexachlorobutadiene, both enzymes were released into the plasma, and GGT, but not 5'-NT activity was found in urine. In sheep with tubular necrosis of the kidney caused by the administration of mercuric chloride, GGT activity, but not 5'-NT, increased in the plasma and urine.

5'-Nucleotidase↗
PubMed1988

Further studies on the diagnostic value of gamma-glutamyl transpeptidase and 5'-nucleotidase in cattle, sheep and horses.

The distribution of 5'-nucleotidase (5'-NT) and gamma-glutamyl transpeptidase (gamma-GT) is similar in the tissues of the sheep, calf and horse, except that there is relatively less gamma-GT in calf liver than in the liver of the other two species. The liver lesion produced by the oral administration of chloroform is similar in the three species and is accompanied by the release of 5'-NT into the plasma of the sheep and calf but not of the horse. Conversely, gamma-GT is released into plasma of the horse but not of the sheep or calf. This difference is not related to the tissue distribution of the two enzymes. The kidney lesion in sheep produced by the intravenous administration of mercuric chloride is accompanied by a reduction in the rate of excretion of an injected dose of inulin and by an increase in the concentration of urea in plasma and in the activity of gamma-GT in plasma and urine. There was no increase in 5'-NT activity in plasma or urine.

5'-Nucleotidase↗
PubMed1985

Decreased ecto-5'nucleotidase activity of peripheral blood lymphocytes in human monoclonal gammopathies: correlation with tumor cell kinetics.

Ecto-5'nucleotidase (5'NT) activity of peripheral blood (PB) lymphocytes was determined in 31 patients with serum monoclonal gammopathies (MG). Twenty-one patients had a diagnosis of multiple myeloma (MM), and ten patients had monoclonal gammopathy of undetermined significance (MGUS). The proliferative activity of the bone marrow plasma cells (LI%) was investigated in 28 of these MG patients by means of tritiated thymidine uptake evaluated by simultaneous autoradiography and cytoplasmic immunofluorescence. 5'NT activity was significantly lower in MG patients as compared with normal controls. MM patients had lower 5'NT activity than MGUS patients, but the difference was not significant. By contrast, MM had significantly higher LI% than MGUS patients. There was a linear regression of 5'NT on LI% which was statistically significant: the higher the LI%, the lower the 5'NT. Because the LI% is an accurate prognostic and monitoring factor in MG, this correlation indicates that 5'NT may be of assistance in predicting the clinical progress of MG patients. In seven MGs, the PB T and B lymphocytes were studied separately. The T cell subpopulation was 5'NT deficient compared to the normal controls, shown as a significant linear regression of T cell 5'NT on the LI%. This suggests that in MG there may be an alteration of nonneoplastic T lymphocytes correlated with tumor growth. The OKT8+ lymphocytes were mainly responsible for the 5'NT deficiency of unseparated T lymphocytes.

5'-Nucleotidase↗
PubMed1985

Erythrocyte pyrimidine-5'-nucleotidase activity as a sensitive indicator of lead exposure.

Since the red cell enzyme pyrimidine-5'-nucleotidase (PN) is inhibited by lead, we examined the feasibility of using the activity of this enzyme as a measure of lead exposure. Erythrocyte PN activity was measured in 110 blood samples obtained from subjects working in industries which utilize lead and 40 control subjects. The measurements were then compared with a number of traditional indices of lead poisoning. These included blood and urine lead concentrations and free erythrocyte protoporphyrin, erythrocyte zinc protoporphyrin, urinary coproporphyrin and urinary delta-aminolaevulinic acid levels. There was a highly significant negative correlation between erythrocyte PN activity and blood lead concentration (-0,83; P less than 0,0001), which was greater than that for any of the other measurements. It was therefore concluded that erythrocyte PN activity is an excellent indicator of lead exposure.

5'-Nucleotidase↗
PubMed1985

Reduced ecto-5'-nucleotidase activity and enhanced OKT10 and HLA-DR expression on CD8 (T suppressor/cytotoxic) lymphocytes in the acquired immune deficiency syndrome: evidence of CD8 cell immaturity.

Markedly reduced ecto-5'-nucleotidase activity was found in peripheral blood lymphocytes from 27 out of 30 homosexual men with the acquired immune deficiency syndrome (AIDS) in association with Kaposi's sarcoma (AIDS-KS; 2.67 +/- 1.70 U/10(6) cells; n = 13), opportunistic infections (AIDS-OI; 9.29 +/- 7.32; n = 7), or the AIDS-related complex (ARC; 9.82 +/- 6.12; n = 10). These values were significantly different from healthy controls (22.70 +/- 4.58; p less than 0.001). In AIDS-KS patients, both T cells and non-T cells exhibited significantly reduced ecto-5'-NT activity (p less than 0.001). AIDS-KS CD8 cells contained 20% of the mean ecto-5'-NT activity (7.04 +/- 3.53) displayed by control CD8 cells (34.07 +/- 4.86; p less than 0.001). No significant difference in enzyme level was observed between control and AIDS-KS CD4 cells (11.93 +/- 4.98 vs 7.98 +/- 3.28, respectively). In AIDS patients, lymphocyte ecto-5'-NT activity was inversely related (r = -0.518; p less than 0.01) to the absolute number of OKT10+ cells, but no correlation was found with the number of HLA-DR+ cells (r =-0.224). Two-color analysis of lymphocytes from AIDS-KS patients revealed that 75 +/- 12% of circulating CD8 cells expressed the OKT10 antigen, whereas only 10 +/- 6% of control CD8 cells did. HLA-DR antigens, which are not normally found on circulating resting T cells, were expressed in AIDS-KS CD8 cells, although to a lesser extent than OKT10. These data demonstrate that most AIDS CD8 cells differ from control CD8 cells. Although it has been suggested that these cells are activated cytotoxic or suppressor cells, the data presented here support the hypothesis they are immature. Reduced T cell ecto-5'-NT activity and enhanced expression of OKT10 and HLA-DR antigens on circulating CD8 cells, in conjunction with lack of transferrin receptor-(OKT9) and IL 2 receptor-(Tac) bearing lymphocytes, sustain this latter hypothesis. The correlation of the numerical reduction of CD4 cells with the reduced levels of ecto-5'-NT (r = 0.606; p less than 0.01) suggests that the abnormal maturation of CD8 cells seen in AIDS might be a consequence of the CD4 deficiency characteristic of this syndrome.

5'-Nucleotidase↗
PubMed1985

Isozymes of human erythrocyte pyrimidine 5'-nucleotidase--chromatographic separation and their properties.

The biochemical properties of two isozymes of human erythrocyte pyrimidine 5'-nucleotidase (P5N) separated by ion-exchange chromatography were studied. These two isozymes, P5N-I and P5N-IIB, can be distinguished from each other by substrate specificity, pH dependent activity and thermostability. P5N-I is thermolabile and dephosphorylates various pyrimidine 5'-monophosphates optimally around pH 7. UMP and CMP were found to be most effective substrates. P5N-IIB is thermostable and characterized by its high Michaelis constant and maximum velocity for dUMP or dTMP and acidic pH optima. The molecular weight was estimated to be 39,000 for P5N-I and 44,000 for P5N-IIB by gel filtration. These data suggest that these two isozymes are independent proteins and probably encoded by distinct structural gene loci.

5'-Nucleotidase↗
PubMed1985

[Alkaline phosphatase and 5'-nucleotidase in early osteogenesis in man].

By means of histochemical techniques at light and electron microscopic levels, as well as immunomorphological, biochemical and immunochemical methods localization and dynamics of alkaline phosphatase and 5'-nucleotidase contents have been determined in anlages of long tubular bones in 85 human embryos and prefetuses from the 6th up to 12th week of the intrauterine development, obtained as a result of artificial abortions in healthy women. The greatest activity of the enzymes studied is revealed in areas of an intensive osteogenesis and mineralization. Also, by means of the immunofluorescent method alkaline phosphatase of a placental type is revealed, that is not revealed, however, immunochemically. With increasing time of the intrauterine development, thermostability of alkaline phosphatase increases.

5'-Nucleotidase↗
PubMedDate not supplied

[Age-dependent characteristics of the effect of insulin on 5'-nucleotidase activity in liver plasma membranes].

Vmax of 5'-nucleotidase was found to be similar in liver plasmatic membranes of both 4-5 months old male rats and 24-26 months old animals. At the same time, Km value was higher in the membranes of old rats than in young animals. Insulin (administered at a dose of 2 un per 100 g of body mass) increased the enzyme Vmax without some age-dependent differences and decreased Km value in old animals as compared with controls.

5'-Nucleotidase↗
PubMed1986

Differential expression of ecto-5' nucleotidase activity by functionally and phenotypically distinct subpopulations of human Leu-2+/T8+ lymphocytes.

Subsets of Leu-2+/T8+ cytotoxic/suppressor T lymphocytes were isolated by using various methods of purification and were investigated for expression of ecto-5' nucleotidase (5'NT) enzyme activity by radiochemical, cytochemical, and ultrastructural techniques. By using both the radiochemical and the cytochemical methods. T4-OKM1- cells displayed higher 5'NT activity in comparison with the entire T4- subpopulation. Analyses of the subpopulations of T4- (and predominantly Leu-2+) cells defined by the Leu-15 or Lyt-1 (9.3) monoclonal antibodies demonstrated that T4-Leu-15- and T4-Lyt-1+ cells displayed high 5'NT activity, whereas virtually no activity was present in T4-Leu-15+ and T4-Lyt-1-cells. At the ultrastructural level, the 5'NT reaction product was detected on the plasma membrane of a proportion of nongranular Leu-2+/T8+ lymphocytes, but no activity was found on cells with a granular lymphocyte (GL) morphology. 5'NT activity was also analyzed in peripheral blood mononuclear cells from one patient with expanded numbers of GL and two patients with GL leukemia. The enzymatic activity was significantly lower in these patients than in normal controls. This study provides new cytochemical evidence demonstrating the heterogeneity of Leu-2+/T8+ cells, and indicates that the population with the suppressor phenotype and function (Leu-15+/Lyt-1-, GL morphology) displays low or absent 5'NT activity, whereas the population composed of cytotoxic cell precursors (Leu-15-/Lyt-1+, nongranular morphology) has high 5'NT activity. Implications of these data for the interpretation of low 5'NT activity described in several immunodeficiency states and lymphoproliferative disorders are discussed.

5'-Nucleotidase↗
PubMed1984

Cytochemical localisation of 5'-nucleotidase in amoeboid microglial cells in postnatal rats.

The activity of the enzyme 5'-nucleotidase has been studied in postnatal rats. In 5 days old rats, under the light microscope, round amoeboid microglial cells were stained for the enzyme. The cytoplasm was stained dark brown, with a clear nucleus. In older rats, the reactive amoeboid microglial cells assumed an oval or flattened form. In the electron microscope, in 5 days old rat, reaction product was deposited at various sites in the cytoplasm of the amoeboid microglial cells: over lysosomes in subplasmalemmal vacuoles as well as in saccules of the Golgi apparatus. In older rats the cells showed reaction over the lysosomes. The cells became oval and elongated.

5'-Nucleotidase↗
PubMed1984

Decreased 5' nucleotidase activity in lymphocytes from asymptomatic sexually active homosexual men and patients with the acquired immune deficiency syndrome.

5' Nucleotidase (5'NT) is an ectoenzyme associated with the plasma membrane of most mammalian cells. Low 5'NT activity has been observed in peripheral blood lymphocytes from patients with immunodeficiency states. 5'NT activity was measured in null and T-enriched lymphocytes from asymptomatic homosexual men and from 20 men with various degrees of the acquired immune deficiency syndrome (AIDS). Asymptomatic homosexuals were self-referred because of their concern about AIDS and were not necessarily representative of homosexuals in the general population. Enzyme activity was significantly decreased in both null (7.0 +/- 2.4 nmol/10(6) cells/h) and T-enriched (12.0 +/- 6.0 nmol/10(6) cells/h) lymphocytes in homosexuals as compared to lymphocytes from aged-matched heterosexual male and female controls (null = 10.8 +/- 6.5 and T = 22.3 +/- 10.6, P less than .0001 and .008, respectively). Decreased activity was present regardless of whether the patients were asymptomatic, had prodromal symptoms such as fever, lymph node enlargement, weight loss and diarrhea, or had opportunistic infections or Kaposi's sarcoma. Homosexuals had a significantly higher fraction of lymphocytes expressing the activation antigens T10 (20% +/- 3.3%) and Ia (13% +/- 2.9%) than controls (11% +/- 1.8% and 5% +/- 0.8%, respectively, P less than .05). They also had a significantly lower fraction of OKT4-positive helper lymphocytes than controls (22% +/- 3.4% v 35% +/- 2.2%, P less than .05). 5'NT activity in lymphocytes enriched for null cells from homosexuals correlated inversely with the percentage of Ia-positive lymphocytes (r = -.655; P less than .02). There was no correlation between 5'NT activity and the percentage of T4- or T8-positive lymphocytes or the T4/T8 ratio. Moreover, 5'NT activity was significantly decreased in both OKT4 (P less than .025) and OKT8 (P less than .05) enriched lymphocytes in homosexuals compared to controls. The data suggest that decreases in 5'NT may be a generalized defect of the peripheral blood T lymphocytes from active homosexuals that is independent of increases or decreases in specific T subpopulations or clinical status. It may contribute to the pathogenesis of AIDS.

5'-Nucleotidase↗
PubMed1984

Ecto-5'-nucleotidase. I. Expression in human normal and neoplastic lymphoid cell lines representing sequential stages of B-cell differentiation.

The possible association of ecto-5'-nucleotidase (5'-NT) with differentiation of B-cells was explored with the use of normal and neoplastic lymphoblastoid cell lines representing sequential stages of B-cell maturation. There was no relationship between patterns of enzyme expression in the cell lines and immunoglobulin (Ig) secretion, chromosome constitution, proliferative rate, cell volume, or the presence of B1 and B2 antigens. Pre-B-cell lines, which were negative for surface Ig or Ig secretion but positive for cytoplasmic mu-chains, showed the presence of 5'-NT, whereas 9 of 11 lymphoma cell lines, Burkitt's or non-Burkitt's type, both secreting and nonsecreting, did not exhibit enzyme activity. Four myeloma cell lines and 13 of 15 normal B-cell lines were positive for 5'-NT. These results suggested that 5'-NT was present in pre-B-cells and in some very early B-cells. 5'-NT usually disappeared from early and some intermediate B-cells and reappeared in mature B-cells and plasmacytoid cells.

5'-Nucleotidase↗
PubMed1984

Ecto-5'-nucleotidase. II. Effect of 12-O-tetradecanoylphorbol 13-acetate on the expression of enzyme in normal and neoplastic B-cell lines.

Immature B-cells, including B-cell lymphoma lines, are often deficient in ecto-5'-nucleotidase (5'-NT) activity. 12-O-Tetradecanoylphorbol 13-acetate (TPA) was shown to be capable of inducing maturation toward plasmacytoid-like cells in immunoglobulin (Ig)-secreting B-cell lines. An attempt was made to induce the enzyme in 5'-NT-negative B-cell lymphoma lines with TPA to clarify the relationship between 5'-NT and B-cell differentiation. After 3 days in the presence or absence of TPA, these cell lines were examined morphologically, and their 5'-NT activity, Ig secretion, surface Ig, and Ia, B1, and B2 antigens were estimated. Neither Ig secretion nor 5'-NT activity was induced by TPA in any of 4 nonsecreting cell lines studied. Ig secretion was significantly increased in 4 of 5 lg-secreting cell lines. Two of these inducible cell lines, JD 38 and ST 486, became positive for 5'-NT activity and acquired morphologic characteristics of plasma cells after culture with TPA. The lymphoma cell line JD 38 was transplanted into nude mice and gave rise to a solid tumor. Although the tumor cells remained negative for 5'-NT, they could be induced by TPA to express both the enzyme activity and plasmacytoid-like appearance. These data suggested that in the Ig-secreting B-cell lymphoma lines, there was an association between the inducibility of 5'-NT and the capacity of these cell lines to undergo plasma-cytoid-like transformation in response to TPA.

5'-Nucleotidase↗
PubMed1984

Histochemical mapping of ATPase and 5'-nucleotidase in the telencephalon and diencephalon of mouse.

The paper deals with a detailed histochemical mapping of adenosine triphosphatase (ATPase) and 5'-Nucleotidase (5'-N) in the telencephalon and diencephalon of mouse. The majority of brain matter nuclei demonstrated high activity for ATPase while, in contrast, only a few nuclei revealed high intensity of 5'-N. In general, the neuronal population dominated in ATPase activity as compared to the neuropil. The highest ATPase activity was observed in the cerebral cortex, nucleus caudatus-putamen, hippocampus, habenular complex and hypothalamus as compared to the thalamus, subthalamus and metathalamus. The motor dominating regions have indicated, invariably, intense ATPase activity than the sensory dominating or sensory-motor nuclei. The highest 5'-N activity was observed in nucleus caudatusputamen, globus pallidus, induseum griseum, nucleus amygdaloideus anterior, nucleus amygdaloideus centralis and nucleus accumbens. The heterogeneous distribution of 5'-N activity was observed in the different regions of hippocampus ranging from mild through moderate to very intense. The brain matter nuclei which dominated with the myelinated fibers demonstrated very intense 5'-N activity. The significance of ATPase and 5'-N has been discussed in relation to the nature of brain matter nuclei and fiber tracts.

5'-Nucleotidase↗
PubMed1981

Distribution of 5'-nucleotidase and gamma glutamyl transferase activities in the tissues of the horse.

In the horse, 5'-nucleotidase (5'-NT) activity is found mainly in homogenates of lung, kidney, small intestine, mammary gland, liver and pancreas. Lower activities are present in brain and muscle. Activity can be demonstrated histochemically in the glomeruli and tubules of the kidney, in the sinusoidal borders of the hepatocytes and the bile duct epithelium as well as in the blood vessels of all organs. There is no significant difference between the 5'-NT activity in serum and plasma of normal horses and of horses suffering from a range of orthopaedic conditions. Previous findings that gamma glutamyl transferase activity is greatest in kidney and high in liver and pancreas have been confirmed.

5'-Nucleotidase↗
PubMed1983

Inhibition of liver surface membrane Na+, K+-adenosine triphosphatase, Mg+2-adenosine triphosphatase and 5'-nucleotidase activities by protoporphyrin. Observations in vitro and in the perfused rat liver.

To clarify the pathogenesis of protoporphyrin-induced cholestasis, liver surface membrane enzyme activities were determined after (a) isolated rat liver perfusion with protoporphyrin administered by bolus (1.0 mumol) or bolus plus constant infusion (1.0 mumol + 0.05 mumol/min) and (b) combination of liver surface membrane with protoporphyrin (0.9-53.4 nmol/ml) in vitro. The perfusion studies showed that protoporphyrin significantly inhibited Na+, K+- and Mg+2-adenosine triphosphatase activities. In vitro, these adenosine triphosphatase activities and the 5'-nucleotidase activity were inhibited linearly by protoporphyrin up to a concentration of approximately 18 mumol/ml; thereafter, enzyme activity was maintained. Greater inhibition of the adenosine triphosphatase activities occurred with protoporphyrin than was reported for chlorpromazine at similar molar concentrations. This effect was independent of the quantity of membrane protein analyzed and was not reversible with a 50:1 dilution. The inhibitory effect of protoporphyrin on surface membrane enzyme activities was also nonselective. Although the hepatotoxic effects of protoporphyrin may be more generalized, the present data underscore protoporphyrin's toxic interaction with liver surface membranes.

5'-Nucleotidase↗