Myeloperoxidase is involved in H2O2-induced apoptosis of HL-60 human leukemia cells

We examined the mechanism of H(2)O(2)-induced cytotoxicity and its relationship to oxidation in human leukemia cells. The HL-60 promyelocytic leukemia cell line was sensitive to H(2)O(2), and at concentrations up to about 20-25 micrometer, the killing was mediated by apoptosis. There was limited evidence of lipid peroxidation, suggesting that the effects of H(2)O(2) do not involve hydroxyl radical. When HL-60 cells were exposed to H(2)O(2) in the presence of the spin trap alpha-(4-pyridyl-1-oxide)-N-tert-butylnitrone (POBN), we detected a 12-line electron paramagnetic resonance spectrum assigned to the POBN/POBN(.) N-centered spin adduct previously described in peroxidase-containing cell-free systems. Generation of this radical by HL-60 cells had the same H(2)O(2) concentration dependence as initiation of apoptosis. In contrast, studies with the K562 human erythroleukemia cell line, which is often used for comparison with the HL-60, and with high passaged HL-60 cells (spent HL-60) studied under the same conditions failed to generate POBN(.). Cellular levels of antioxidant enzymes superoxide dismutase, glutathione peroxidase, and catalase did not explain the differences between these cell lines. Interestingly, the K562 and spent HL-60 cells, which did not generate the radical, also failed to undergo H(2)O(2)-induced apoptosis. Based on this we reasoned that the difference in H(2)O(2)-induced apoptosis might be due to the enzyme myeloperoxidase. Only the apoptosis-manifesting HL-60 cells contained appreciable immunoreactive protein or enzymatic activity of this cellular enzyme. When HL-60 cells were incubated with methimazole or 4-aminobenzoic acid hydrazide, which are inhibitors of myeloperoxidase, they no longer underwent H(2)O(2)-induced apoptosis. Hypochlorous acid stimulated apoptosis in both HL-60 and spent HL-60 cells, indicating that another oxidant generated by myeloperoxidase induces apoptosis and that it may be the direct mediator of H(2)O(2)-induced apoptosis. Taken together these observations indicate that H(2)O(2)-induced apoptosis in the HL-60 human leukemia cell is mediated by myeloperoxidase and is linked to a non-Fenton oxidative event marked by POBN(.).     

Evidence that proximal multiple mutations in Big Blue transgenic mice are dependent events

To characterize the nature of multiple mutations in the tissues of an intact animal, the Big Blue transgenic mouse mutation detection system was used to examine 1459 mutants from eight normal tissues and 507 mutants from 11 tumors. Multiple mutations occurred and predominantly doublet mutants were identified (i.e. two mutations within one mutant lacI gene), but multiplets of up to five mutations were observed. The frequency of doublets in normal tissues and spontaneous tumors from p53-deficient mice was enhanced to the same degree (660 and 667 fold, respectively) over that expected for two independent mutational events. Doublets, multiplets and singlets have similar patterns of mutation. The distance between mutations in doublets fits an exponential distribution, not that expected for randomly spaced events, suggesting that many doublets occur in rapid succession within the same cell cycle.     

Decorticate, decerebrate and opisthotonic posturing and seizures in Kenyan children with cerebral malaria

Background

Intermittent preventive treatment of malaria during pregnancy (IPTp) is a key intervention in the national strategy for malaria control in Tanzania. SP, the current drug of choice, is recommended to be administered in the second and third trimesters of pregnancy during antenatal care (ANC) visits. To allow for a proper design of planned scaling up of IPT services in Tanzania it is useful to understand the IPTp strategy's acceptability to health managers, ANC service providers and pregnant women. This study assesses the knowledge, attitudes and practices of these groups in relation to malaria control with emphasis on IPTp services.

Methods

The study was conducted in February 2004, in Korogwe District, Tanzania. It involved in-depth interviews with the district medical officer (DMO), district hospital medical officer in charge and relevant health service staff at two peripheral dispensaries, and separate focus group discussions (FGDs) with district Council Health Management Team members at district level and pregnant women at dispensary and community levels.

Results

Knowledge of malaria risks during pregnancy was high among pregnant women although some women did not associate coma and convulsions with malaria. Contacting traditional healers and self-medication with local herbs for malaria management was reported to be common. Pregnant women and ANC staff were generally aware of SP as the drug recommended for IPTp, albeit some nurses and the majority of pregnant women expressed concern about the use of SP during pregnancy. Some pregnant women testified that sometimes ANC staff allow the women to swallow SP tablets at home which gives a room for some women to throw away SP tablets after leaving the clinic. The DMO was sceptical about health workers' compliance with the direct observed therapy in administering SP for IPTp due to a shortage of clean water and cups at ANC clinics. Intensified sensitization of pregnant women about the benefits of IPTp was suggested by the study participants as an important approach for improving IPTp compliance.

Conclusion

The successful implementation of the IPTp strategy in Tanzania depends on the proper planning of, and support to, the training of health staff and sustained sensitization of pregnant women at health facility and community levels about the benefits of IPTp for the women and their unborn babies.     

Thymidylate synthase polymorphism and microsatellite instability: association in colorectal cancer

5-Fluorouracil (5FU) is the main drug used for the treatment of colorectal cancer (CRC) and Thymidilate Synthase (TS) is its target enzyme. TS gene has regulatory tandemly repeated sequences in its 5' and 3'untraslated region (5'-3' UTR). CRC often shows a kind of genomic instability called Microsatellite Instability (MSI) that is associated with TS levels and survival. Our data show that the genotype 2R/2R (homozygosity for 2 tandem repeat sequences in the 5'UTR) is more frequently associated with MSI+ and lower TS levels. More over we did not find any significant association between the 2R/3R (heterozygosity for 2 and 3 tandem repeat sequences in the 5'UTR) and 3R/3R (homozygosity for 3 tandem repeat sequences in the 5' UTR) genotypes with the MSI+ and MSI-, while these genotypes were associated with a higher TS expression. As a consequence we can hypothesise that patients bearing CRC with the MSI+, the 2R/2R genotype and with low TS levels could have a better prognosis and they could not be drug resistant.     

Characterization and expression pattern of the novel MIA homolog TANGO

A novel human gene, TANGO, encoding a MIA ('melanoma inhibitory activity') homologous protein was identified by a gene bank search. TANGO, together with the homologous genes MIA, OTOR (FPD, MIAL) and MIA2 define a novel gene family sharing important structural features, significant homology at both the nucleotide and protein level, and similar genomic organization. The four members share 34-45% amino acid identity and 47-59% cDNA sequence identity. TANGO encodes a mature protein of 103 amino acids in addition to a hydrophobic secretory signal sequence. Sequence homology confirms the highly conserved SH3 structure present also in MIA, OTOR and MIA2. Thus, it appears that there are a number of extracellular proteins with SH3-fold like structures. Interestingly, in situ hybridization, RT-PCR and Northern Blots revealed very broad TANGO expression patterns in contrast to the highly restricted expression patterns previously determined for the other members of the MIA gene family. The only cells lacking TANGO expression are cells belonging to the hematopoetic system. High levels of TANGO expression were observed both during embryogenesis and in adult tissues.     

Dissemination of the entomopathogenic fungus verticillium lecanii (Zimmermann) Viégas (Hyphomycetales: Moniliaceae) in a population of Frankliniella occidentalis (Pergande, 1895) (Thysanoptera: Thripidae)

Tests were carried out to investigate the dissemination of the entomopathogenic fungus Verticillium lecanii (Zimmermann) Viégas in a population of Frankliniella occidentalis. The tested factors, which influence the efficacy of the fungus against the pest insect, have been the population density of the thrips at the application moment as well as the temperature. The population density influenced the dissemination of the fungal spores in the population. The higher the density has been, the higher the insetting control effect has been as well. The temperature influenced the speed of the fungal effect, too. The higher the temperature has been, the earlier the control effect started. However, an increase of the natural mortality was found as well.