Metabolic effects of thiopurine derivatives against human CCRF-CEM leukaemia cells

BACKGROUND and aims. To compare the metabolic effects induced by the anticancer drugs, 6-mercaptopurine (6-MP), 6-thioguanine (6-TG) and 6-methylmercaptopurine riboside (MMPR), which may inhibit the de novo biosynthesis of purine nucleotides or be mis-incorporated into DNA or RNA. METHODS: Leukaemia cells were grown in culture, exposed to a thiopurine and cell extracts were analyzed for NTPs, dNTPs, drug metabolites and P-Rib-PP. RESULTS: In leukaemia cells, 6-MP was converted to MPR-MP, thio-XMP, thio-GMP, thio-GDP and thio-GTP. Metabolites of 6-TG included thio-XMP, thio-GMP, thio-GDP and thio-GTP, while MMPR-MP was the only major metabolite of MMPR, MMPR (25 microM, 4 h) induced a 16-fold increase in P-Rib-PP and 6-MP (25 microM, 4 h) induced a delayed 5.2-fold increase. MPR-MP, thio-GMP and MMPR-MP are inhibitors of amido phosphoribosyltransferase from leukaemia cells with Ki values of 114 +/- 7.10 microM, 6.20 +/- 2.10 microM and 3.09 +/- 0.30 microM, respectively. CONCLUSION: The nucleoside-5'-monophosphate derivatives of the 3 thiopurines inhibit amido phosphoribosyltransferase in growing leukaemia cells but there is also an initial inhibition of the further conversion of IMP in the pathway. In growing cells, MMPR acts solely as an inhibitor of de novo purine biosynthesis while 6-TG and to a lesser extent, 6-MP, are converted to significant concentrations of di- and tri-phosphate derivatives which may have other mechanisms of cytotoxicity.     

Low pressure RF discharge in electronegative gases for plasma processing

An RF plasma discharge in oxygen has been investigated both experimentally and theoretically. The excited species in the oxygen plasma were detected by means of spontaneous luminescence spectroscopy. A self-consistent numerical model was developed for these discharges. The model was confirmed by several theoretical and experimental investigations. The spatial distribution over the discharge gap of both charged and neutral plasma components and their fluxes to electrodes have been calculated.     

Factors influencing sperm-zona pellucida binding in vitro using the intact zona model

The zona pellucida binding assay assesses the ability of spermatozoa to bind to the zona pellucida. The present study investigated the influence of: (i) prior oocyte exposure to spermatozoa on subsequent sperm-zona pellucida binding in vitro; and (ii) cryopreservation of oocytes. Only oocytes obtained from fertile donors were used and the binding capacity of non-inseminated, cryopreserved oocytes was compared with both inseminated/unfertilized, cryopreserved oocytes and inseminated/unfertilized, non-cryopreserved oocytes recovered from in-vitro fertilization cultures on sperm-zona pellucida binding using an intact zona model. There was no statistically significant difference in sperm-zona binding between non-inseminated, cryopreserved oocytes (range 9.6-23.2), inseminated/unfertilized, cryopreserved oocytes (range 15.0-16.0) and inseminated/ unfertilized, non-cryopreserved oocytes (range 3.3-23.0). The coefficient of variation for sperm binding to all oocyte groups was very large (range 37-121%). We conclude that neither prior exposure of human oocytes to human spermatozoa nor cryopreservation of human oocytes influences the subsequent binding of a different population of spermatozoa to the zona pellucida. However, large oocyte-to-oocyte variation of sperm-zona binding may diminish the usefulness of this assay in clinical practice and as a research tool.     

The impact of muscular dystrophy on limb bone growth and scaling in mice

Muscular loading affects bone growth and the factors determining size and shape. However, it is not known what epigenetic impact muscular dystrophy (dystrophia muscularis) has on limb bone growth or ontogenetic scaling. To assess the effects of two types of muscular dystrophy (genotypes dy/dy and dy2J/dy2J) on limb bone growth, we measured lengths and widths of the right humerus, femur and tibia, and lengths of the ulna and radius from dorsal/ventral radiographs of mice taken over a period of 270 days. Radiographs were taken approximately 3 times a week, and the sampling frequency was gradually reduced to once a month. We plotted measurements from each individual against time and fit a Gompertz equation to the growth of each bone. Parameters of the equation were compared using ANOVA across genotypes and between sexes. Slopes of length versus width were calculated for the limb bones of each individual using linear regression. Slope differences among genotypes and between sexes were tested using ANOVA. Control and dy2J values were significantly longer than those of dy mice in all bones, but there was considerable variation across genotypes for the various width measurements. Sexual dimorphism was found in several measurements, where males were always larger than females. There were few significant differences in limb scaling (lengths vs. widths) among genotypes and almost no scaling differences between sexes despite the size differences. Differences among widths suggest that muscular dystrophy affects different parts of limb bones in different ways. This may be the effect of the type and number of muscular attachments, as well as the usage of the limb. The sexually dimorphic measurements suggest that there are size differences in the skeleton between sexes, regardless of the genotype. Our ontogenetic allometry results indicate that size is affected by the muscular dystrophic condition and by sexual dimorphism, while shape remains largely unchanged.     

Lewis and Fischer rat strains display differences in biochemical, electrophysiological and behavioral parameters: studies in the nucleus accumbens and locus coeruleus of drug naive and morphine-treated animals

We investigated the effects of the systemic administration of thymosin alpha 1 plus relatively low doses of human recombinant interleukin-2 or very low doses of interferon alpha,beta in untreated and cyclophosphamide (CY)-treated DBA/2 mice challenged either subcutaneously or intravenously (i.v.) with Friend erythroleukemia cells (FLC). Both treatments resulted in the complete regression of subcutaneous tumor and cured a significative percentage of mice. They also increased the survival time of mice i.v. injected with large numbers of FLC. Neither immunotherapy alone nor CY, alone or in combination with single cytokines, produced similar effects. The antitumor action of these combined chemoimmunotherapy protocols seems to involve activation of the immune response since (a) a synergistic increase of the cytotoxicity of spleen cells was demonstrated in treated mice; (b) selective in vivo depletion of asialo-GM1, CD4, or CD8-positive cells abrogated this antitumor activity; and (c) a high lymphoid cell infiltration was found at the tumor site and in the livers of treated mice.