Coordinate induction of glutathione biosynthesis and glutathione-metabolizing enzymes is correlated with salt tolerance in tomato

The acclimation of reduced glutathione (GSH) biosynthesis and GSH-utilizing enzymes to salt stress was studied in two tomato species that differ in stress tolerance. Salt increased GSH content and GSH:GSSG (oxidized glutathione) ratio in oxidative stress-tolerant Lycopersicon pennellii (Lpa) but not in Lycopersicon esculentum (Lem). These changes were associated with salt-induced upregulation of gamma-glutamylcysteine synthetase protein, an effect which was prevented by preincubation with buthionine sulfoximine. Salt treatment induced glutathione peroxidase and glutathione-S-transferase but not glutathione reductase activities in Lpa. These results suggest a mechanism of coordinate upregulation of synthesis and metabolism of GSH in Lpa, that is absent from Lem.     

The influence of intracellular idarubicin and daunorubicin levels on drug cytotoxicity in childhood acute leukemia

Uptake and efflux of two anthracyclines, idarubicin (IDA) and daunorubicin (DNR), was studied in childhood acute leukemia samples. A comparison of IDA and DNR transport phenomena in relation to drug cytotoxicity and expression of P-glycoprotein (PGP) was made. Intracellular content of IDA/DNR was determined by flow cytometry using the fluorescent properties of the drugs. In vitro drug cytotoxicity was measured by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT) assay. PGP expression was analysed by flow cytometry. The uptake and efflux rates were non-significantly higher for IDA than DNR. There were no differences between three types of leukemia with respect to drug content during accumulation and retention. After correction for the cell volume, intracellular concentration of both drugs in each moment of uptake and efflux was significantly lower in relapsed ALL and AML samples in comparison with initial ALL cells. Efflux, but not uptake, of both drugs was inversely correlated with PGP expression and IDA, but not DNR, cytotoxicity. The cytotoxicity was correlated with drug accumulation for both drugs and with drug retention for IDA. In conclusion, it seems that (1) intracellular content was related to the lipophilic properties of the drugs rather than to the type of leukemia, (2) decreased intracellular concentration of both drugs might have an impact on compromised therapy results in AML and relapsed ALL children, (3) IDA presents higher cytotoxicity, which possibly might be decreased by the presence of PGP. These results might have a practical impact on the rational design of new chemotherapy protocols.     

Comparison of multiplex PCR,and an immunochromatographic method sensitivity for the detection of Escherichia coli O157:H7 in minced beef

In this study, the sensitivities of multiplex PCR and an immuno-chromatographic methods to detect Escherichia coli O157:H7 in minced beef were compared. The detection of Escherichia coli O157:H7 in minced beef inoculated with 1-100 cells of this bacterium was possible after enrichment of culture and subsequent analysis by either of the two methods. Enrichment conditions were eight hours of incubation at 37 degrees C or 42 degrees C in a non-selective medium (Buffered Peptone Water). Multiplex PCR analysis was performed using three primer sets with analysis by gel electrophoresis. The Quix immuno-chromatographic assay which is a new kit being marketed by New Horizons Diagnostics, Columbia, MD, was used for immunological analysis of the enriched broths.The sensitivity of both tests was similar. The results depended on the concentration of the specific bacterium in the culture since the influence of the proportion of other bacteria to the E. coli O157:H7 was not observed. The data suggests that either method or used together, when coupled with an enrichment technique, could provide a rapid mean to detect the presence of this pathogen in minced meat samples.     

Salt stress induces up-regulation of an efficient chloroplast antioxidant system in the salt-tolerant wild tomato species Lycopersicon pennellii but not in the cultivated species

The response of the chloroplastic antioxidant system of the cultivated tomato Lycopersicon esculentum (Lem) and its wild salt-tolerant related species L. pennellii (Lpa) to NaCl stress was studied. An increase in H₂O₂ level and membrane lipid peroxidation was observed in chloroplasts of salt-stressed Lem. In contrast, a decrease in these indicators of oxidative stress characterized chloroplasts of salt-stressed Lpa plants. This differential response of Lem and Lpa to salinity, correlates with the activities of the antioxidative enzymes in their chloroplasts. Increased activities of total superoxide dismutase (SOD), ascorbate peroxidase (APX), monodehydroascorbate reductase (MDHAR), glutathione-S-transferase (GST), phospholipid hydroperoxide glutathione peroxidase (PHGPX) and several isoforms of non-specific peroxidases (POD) were found in chloroplasts of salt-treated Lpa plants. In these chloroplasts, in contrast, activity of lipoxygenase (LOX) decreased while in those of salt-stressed Lem it increased. Although total SOD activity slightly increased in chloroplasts of salt-treated Lem plants, differentiation between SOD types revealed that only stromal Cu/ZnSOD activity increased. In contrast, in chloroplasts of salt-treated Lpa plants FeSOD activity increased while Cu/ZnSOD activity remained unchanged. These data indicate that salt-dependent oxidative stress and damage, suffered by Lem chloroplasts, was effectively alleviated in Lpa chloroplasts by the selective up-regulation of a set of antioxidative enzymes. Further support for the above idea was supplied by leaf discs experiments in which pre-exposure of Lpa plants to salt-treatment conferred cross-tolerance to paraquat-induced oxidative stress while increased oxidative damage by paraquat-treatment was found in salt-stressed Lem plants.     

In vitro activity of oxazaphosphorines in childhood acute leukemia: preliminary report

Glufosfamide (beta-D-glucosyl-ifosfamide mustard) is a new agent for cancer chemotherapy. Its pharmacology is similar to commonly used oxazaphosphorines, but it does not require activation by hepatic cytochrome P-450 and preclinically demonstrates lower nephrotoxicity and myelosuppression than ifosfamide. The aim of the study was a comparison of the drug resistance profiles of glufosfamide and other oxazaphosphorines in childhood acute leukemias. Leukemic cells, taken from children with ALL on diagnosis (n = 41), ALL on relapse (n = 12) and AML on diagnosis (n = 13) were analyzed by means of the MTT assay. The following drugs were tested: glufosfamide (GLU), 4-HOO-ifosfamide (IFO), 4-HOO-cyclophosphamide (CYC) and mafosfamide cyclohexylamine salt (MAF). In the group of initial ALL samples median cytotoxicity values for GLU, IFO, CYC and MAF were 15.5, 33.8, 15.7 and 7.8 microM, respectively. In comparison with initial ALL samples, the relative resistance for GLU and IFO in relapsed ALL samples was 1.9 (p = 0.049) and 1.3 (ns), and in initial AML samples 31 (p < 0.001) and 5 (p = 0.001), respectively. All oxazaphosphorines presented highly significant cross-resistance. Glufosfamide presented high activity against lymphoblasts both on diagnosis and on relapse.     

Effect of phage therapy on the turnover and function of peripheral neutrophils

The aim of this investigation was to establish the impact of phage therapy on the turnover and function of circulating neutrophils in 37 patients with suppurative bacterial infections. We determined the levels of circulating neutrophils and their precursors before therapy, after 3 weeks of therapy, and at a distant time interval (3 months) following the beginning of therapy. In addition, we measured the ability of neutrophils to phagocytize Staphylococcus aureus in vitro. Eight healthy blood donors served as a control group. The results showed that, among the studied parameters, the significant changes involved neutrophil precursor count and the ability of neutrophils to phagocytize bacteria. The percentage of neutrophils in patients before therapy was lower than in healthy donors (mean 58.0, versus 61.4). This value dropped further in patients after 3 months of following the therapy (mean 55.6). The content of neutrophil precursors, on the other hand, was lower in healthy donors than in patients before therapy (mean 2.5, versus 3.8). After 3 weeks of the therapy and after 3 months, the levels of neutrophil precursors were significantly higher (mean 4.8 and 4.9, respectively) than in control donors. The phagocytic index was lower in patients before therapy than in control donors (mean 66.3, versus 70.1) and decreased further after 3 weeks of therapy (mean 59.0) and after 3 months (mean 59.6). The results of this investigation indicate that successful phage therapy accelerates the turnover of neutrophils, accompanied by a decrease in their ability to phagocytize bacteria.     

ShiftDetector: detection of shift mutations

MOTIVATION: Sequencing of a bi-allelic PCR product, which contains an allele with a deletion/insertion mutation results in a superimposed tracefile following the site of this shift mutation. A trace file of this type hampers the use of current computer programs for base calling. ShiftDetector analyses a sequencing trace file in order to discover if it is a superimposed sequence of two molecules that differ in a shift mutation of 1 to 25 bases. The program calculates a probability score for the existence of such a shift and reconstructs the sequence of the original molecule. AVAILABILITY: ShiftDetector is available from http://cowry.agri.huji.ac.il