Purine nucleoside phosphorylase in chronic lymphocytic leukemia (CLL). (1/439)

Purine nucleoside phosphorylase (PNP), the enzyme schematically next to adenosine deaminase in the purine salvage pathway, has been demonstrated cytochemically in peripheral blood lymphocytes of healthy subjects and chronic lymphocytic leukemia (CLL) patients. The enzyme activity is confined to the cytosol. In healthy subjects the majority of lymphocytes are strongly reactive for PNP, whereas the rest are devoid of cytochemically demonstrable activity. The percentage of PNP-positive cells largely corresponds to the number of E rosette-forming cells and is inversely proportional to the number of Ig-bearing cells. In six of seven CLL patients studied only a minor percentage of the lymphocytes showed strong PNP activity, whereas the large majority (88%--98%) possessed trace activity. Such patients have a high number of Ig-bearing cells and a low number of E rosette-forming cells. A different pattern of markers was found in the lymphocytes of the seventh CLL patient: 66% were strongly reactive for PNP, an important number formed E rosettes, and a minor percentage were Ig bearing. These data indicate that PNP can be useful as a "nonmembrane" marker in the differentiation of the B and T cell origin in CLL and deserves to be studied in other lymphoproliferative disorders.  (+info)

Use of alanosine as a methylthioadenosine phosphorylase-selective therapy for T-cell acute lymphoblastic leukemia in vitro. (2/439)

Methylthioadenosine phosphorylase (MTAP) is an important enzyme for the salvage of adenine and methionine and is deficient in a variety of cancers including T-cell acute lymphocytic leukemia (T-ALL). Previously, we reported that the MTAP gene was deleted in over 30% of T-ALL patients at both diagnosis and relapse. We now report that MTAP-primary T-ALL cells are more sensitive to the toxicity of L-alanosine, an inhibitor of de novo AMP synthesis, than are MTAP+ primary T-ALL cells. As measured by [3H]thymidine incorporation, DNA synthesis in all seven MTAP-primary T-ALL cells was inhibited by L-alanosine with a mean IC50 of 4.8+/-5.3 ILM (range, 0.3-11.3 microM). On the other hand, the IC50 for 60% (12 of 20) of MTAP+ primary T-ALL was 19+/-18 microM (range, 1.7-67 microM; P = 0.02), whereas the remaining 40% (8 of 20) had an IC50 of >80 microM4. Furthermore, normal lymphocytes and MTAP+ primary T-ALL cells were rescued from L-alanosine toxicity by the MTAP substrate 5'-deoxyadenosine, but MTAP-T-ALL cells were not. These results indicate that normal cells, which are intrinsically MTAP+, would be protected from L.-alanosine toxicity, whereas MTAP-tumor cells would be killed. Thus, our results support the use of L-alanosine alone or in combination with a salvage agent as a MTAP-selective therapy and therefore lay the foundation for the initiation of clinical trials for the treatment of T-ALL and other MTAP-deficient malignancies with L-alanosine.  (+info)

Effects of a phosphate buffered extracellular (Ep4) solution in preservation and reperfusion injury in the canine liver. (3/439)

The Ep4 solution, a phosphate buffered extracellular-type solution, is effective in canine lung transplantation following a 96-hour hypothermic (4 degrees C) preservation. In this experiment, we used this solution for liver preservation followed by transplantation. We compared the Ep4 solution with the lactated Ringer's (LR) and the Collins' M (CM) solution (a phosphate buffered intracellular-type solution) in two studies, 1) 48-hour liver preservation, and 2) orthotopic liver transplantation after 5-hour preservation. In the preservation study, purine nucleoside phosphorylase (PNP) levels as a marker of endothelial damage, and alanine aminotransferase (ALT) levels were significantly lower in the livers immersed into the Ep4 solution than in those immersed into other solutions at 36 and 48 hours after preservation. Microscopically, the endothelial injury occurred 24 hours after preservation in the CM solution, and 36 hours after preservation in the LR and Ep4 solutions. In the transplantation study, serum PNP and ALT levels in the livers immersed in Ep4 solution showed a lower tendency compared with those in other solutions at the time of reperfusion, but the histological differences among three groups were not apparent. The present study suggests that the liver can be stored better for a longer time using Ep4 solution than using LR and CM solutions.  (+info)

Prevention of Kupffer cell-induced oxidant injury in rat liver by atrial natriuretic peptide. (4/439)

The generation of reactive oxygen species (ROS) by activated Kupffer cells contributes to liver injury following liver preservation, shock, or endotoxemia. Pharmacological interventions to protect liver cells against this inflammatory response of Kupffer cells have not yet been established. Atrial natriuretic peptide (ANP) protects the liver against ischemia-reperfusion injury, suggesting a possible modulation of Kupffer cell-mediated cytotoxicity. Therefore, we investigated the mechanism of cytoprotection by ANP during Kupffer cell activation in perfused rat livers of male Sprague-Dawley rats. Activation of Kupffer cells by zymosan (150 microgram/ml) resulted in considerable cell damage, as assessed by the sinusoidal release of lactate dehydrogenase and purine nucleoside phosphorylase. Cell damage was almost completely prevented by superoxide dismutase (50 U/ml) and catalase (150 U/ml), indicating ROS-related liver injury. ANP (200 nM) reduced Kupffer cell-induced injury via the guanylyl cyclase-coupled A receptor (GCA receptor) and cGMP: mRNA expression of the GCA receptor was found in hepatocytes, endothelial cells, and Kupffer cells, and the cGMP analog 8-bromo-cGMP (8-BrcGMP; 50 microM) was as potent as ANP in protecting from zymosan-induced cell damage. ANP and 8-BrcGMP significantly attenuated the prolonged increase of hepatic vascular resistance when Kupffer cell activation occurred. Furthermore, both compounds reduced oxidative cell damage following infusion of H2O2 (500 microM). In contrast, superoxide anion formation of isolated Kupffer cells was not affected by ANP and only moderately reduced by 8-BrcGMP. In conclusion, ANP protects the liver against Kupffer cell-related oxidant stress. This hormonal protection is mediated via the GCA receptor and cGMP, suggesting that the cGMP receptor plays a critical role in controlling oxidative cell damage. Thus ANP signaling should be considered as a new pharmacological target for protecting liver cells against the inflammatory response of activated Kupffer cells without eliminating the vital host defense function of these cells.  (+info)

Effect of interferon-gamma on purine catabolic and salvage enzyme activities in rats. (5/439)

To determine whether interferon-gamma affects rat purine catabolic and salvage enzyme activities, rats were injected with interferon-gamma (600000 U/kg, i.p.) and, similarly to a vehicle-injected control group, killed before or after injection at 6, 12, and 24 h. Organ homogenates were prepared and enzymatic reactions with substrates were carried out, after which the products were measured either chromatographically or spectrophotometrically. Western and Northern blotting also were performed. In contrast to the vehicle-injected rats, interferon-gamma-injected rats showed a significant rise in xanthine oxidoreductase activity in the liver, while enzyme activity was unchanged in the spleen, kidney, and lung. Western analysis of hepatic xanthine oxidoreductase showed an increased concentration of this protein 12 and 24 h after interferon-gamma injection. Northern analysis disclosed an enhanced mRNA expression coding for this enzyme, peaking 12 h after injection. Contrastingly, the activities of adenosine deaminase, purine nucleoside phosphorylase, hypoxanthine guanine phosphoribosyltransferase, and adenine phosphoribosyltransferase were not affected by interferon-gamma in any organ tested. While interferon-gamma causes an increased hepatic biosynthesis of xanthine oxidoreductase, the physiologic role of this enzyme induction remains undetermined.  (+info)

Isolation and characterization of mutations in the Escherichia coli regulatory protein XapR. (6/439)

In this work, the LysR-type protein XapR has been subjected to a mutational analysis. XapR regulates the expression of xanthosine phosphorylase (XapA), a purine nucleoside phosphorylase in Escherichia coli. In the wild type, full expression of XapA requires both a functional XapR protein and the inducer xanthosine. Here we show that deoxyinosine can also function as an inducer in the wild type, although not to the same extent as xanthosine. We have isolated and characterized in detail the mutants that can be induced by other nucleosides as well as xanthosine. Sequencing of the mutants has revealed that two regions in XapR are important for correct interactions between the inducer and XapR. One region is defined by amino acids 104 and 132, and the other region, containing most of the isolated mutations, is found between amino acids 203 and 210. These regions, when modelled into the three-dimensional structure of CysB from Klebsiella aerogenes, are placed close together and are most probably directly involved in binding the inducer xanthosine.  (+info)

High IL-18 (interferon-gamma inducing factor) concentration in a purine nucleoside phosphorylase deficient patient. (7/439)

The plasma concentration of IL-18 (interferon-gamma inducing factor) was severely increased in a 3 year old boy with purine nucleoside phosphorylase (PNP) deficiency. The presence and activity of IL-18 were confirmed by immunoblotting and bioassay, respectively. These results suggest that IL-18 may be abundantly produced and secreted into plasma by PNP deficient macrophages in PNP deficiency.  (+info)

Nucleosides as a carbon source in Bacillus subtilis: characterization of the drm-pupG operon. (8/439)

In Bacillus subtilis, nucleosides are readily taken up from the growth medium and metabolized. The key enzymes in nucleoside catabolism are nucleoside phosphorylases, phosphopentomutase, and deoxyriboaldolase. The characterization of two closely linked loci, drm and pupG, which encode phosphopentomutase (Drm) and guanosine (inosine) phosphorylase (PupG), respectively, is reported here. When expressed in Escherichia coli mutant backgrounds, drm and pupG confer phosphopentomutase and purine-nucleoside phosphorylase activity. Northern blot and enzyme analyses showed that drm and pupG form a dicistronic operon. Both enzymes are induced when nucleosides are present in the growth medium. Using mutants deficient in nucleoside catabolism, it was demonstrated that the low-molecular-mass effectors of this induction most likely were deoxyribose 5-phosphate and ribose 5-phosphate. Both Drm and PupG activity levels were higher when succinate rather than glucose served as the carbon source, indicating that the expression of the operon is subject to catabolite repression. Primer extension analysis identified two transcription initiation signals upstream of drm; both were utilized in induced and non-induced cells. The nucleoside-catabolizing system in B. subtilis serves to utilize the base for nucleotide synthesis while the pentose moiety serves as the carbon source. When added alone, inosine barely supports growth of B. subtilis. This slow nucleoside catabolism contrasts with that of E. coli, which grows rapidly on a nucleoside as a carbon source. When inosine was added with succinate or deoxyribose, however, a significant increase in growth was observed in B. subtilis. The findings of this study therefore indicate that the B. subtilis system for nucleoside catabolism differs greatly from the well-studied system in E. coli.  (+info)