Cost-effectiveness of clopidogrel in STEMI patients in the Netherlands: a model based on the CLARITY trial.

OBJECTIVE: This study assesses the costs and effects of combination treatment with clopidogrel and aspirin in comparison to aspirin alone in patients with an ST-segment elevation myocardial infarction (STEMI) in a Dutch setting. METHODS: A decision tree model is used to combine data from different sources about efficacy, epidemiology and costs. In the short-run, cost-effectiveness is based on efficacy data derived from the CLARITY trial. The cost-effectiveness of continued treatment is addressed by analysing which conditions need to be fulfilled to deem the strategy 'cost-effective', and discussing whether it is likely that it is. Estimates concerning the benefits of preventing events are derived from Swedish registries. Approximations of both direct and indirect costs are derived from the literature. Effects are expressed as life years gained and Quality Adjust Life Years (QALYs). Uncertainties are addressed by uni- and multivariate sensitivity analyses with and without taking account of the dependency between the separate ischaemic events. RESULTS: A treatment regimen similar to that of the CLARITY trial, including patients similar to those in the trial, is estimated to result in 0.05 additional life years and 0.062 additional quality adjusted life years for a cost that is euro1929 lower than aspirin therapy. Continuation of treatment outside the trial period is expected to result in ICERs of below euro20,000 per QALY as long as the real risk reduction of combination treatment is greater than 0.487% per year. CONCLUSION: The results indicate that clopidogrel therapy combined with aspirin, according to the regimen seen in CLARITY, and using data from Swedish registries to inform the model, is cost-effective. Sensitivity analyses suggest that the model is robust to a wide range of parameter estimates, including those based on data from Swedish registries. Continued treatment is very likely to be cost effective in light of all the indirect evidence.

Complete sequence of a 184-kilobase catabolic plasmid from Sphingomonas aromaticivorans F199.

The complete 184,457-bp sequence of the aromatic catabolic plasmid, pNL1, from Sphingomonas aromaticivorans F199 has been determined. A total of 186 open reading frames (ORFs) are predicted to encode proteins, of which 79 are likely directly associated with catabolism or transport of aromatic compounds. Genes that encode enzymes associated with the degradation of biphenyl, naphthalene, m-xylene, and p-cresol are predicted to be distributed among 15 gene clusters. The unusual coclustering of genes associated with different pathways appears to have evolved in response to similarities in biochemical mechanisms required for the degradation of intermediates in different pathways. A putative efflux pump and several hypothetical membrane-associated proteins were identified and predicted to be involved in the transport of aromatic compounds and/or intermediates in catabolism across the cell wall. Several genes associated with integration and recombination, including two group II intron-associated maturases, were identified in the replication region, suggesting that pNL1 is able to undergo integration and excision events with the chromosome and/or other portions of the plasmid. Conjugative transfer of pNL1 to another Sphingomonas sp. was demonstrated, and genes associated with this function were found in two large clusters. Approximately one-third of the ORFs (59 of them) have no obvious homology to known genes.

Identification and sequence analysis of a 27-kilobase chromosomal fragment containing a Salmonella pathogenicity island located at 92 minutes on the chromosome map of Salmonella enterica serovar typhimurium LT2.

Using a genomic approach, we have identified a new Salmonella pathogenicity island, SPI-4, which is the fourth Salmonella pathogenicity island to be identified. SPI-4 was located at 92 min on the chromosome map and was flanked by the ssb and soxSR loci. The DNA sequence covering the entire SPI-4 and both boundaries was determined. The size of SPI-4 was about 25 kb and it contains 18 putative open reading frames (ORFs). Three of these ORFs encode proteins that have significant homology with proteins involved in toxin secretion. Another five ORFs encode proteins that have significant homology with hypothetical proteins from Synechocystis sp. strain PCC6803 or Acinetobacter calcoaceticus. The rest of the ORFs encode novel proteins, one of which has five membrane-spanning domains. SPI-4 is likely to carry a type I secretion system involved in toxin secretion. Furthermore, a previously identified locus (ims98), which is required for intramacrophage survival, was also mapped within the SPI-4 region. These findings suggested that SPI-4 is needed for intramacrophage survival.

Power frequency magnetic fields do not contribute to transformation of JB6 cells.

The potential for power frequency magnetic fields to enhance neoplastic transformation has been investigated in vitro using promotion-sensitive mouse epidermal JB6 cells. In a soft agar assay, 60-Hz magnetic fields of 0.01, 0.1, 1.0, or 1.1 mT flux density did not induce anchorage-independent growth. In addition, these magnetic fields did not enhance tumor promoter-induced transformation showing no increase in the maximum number of transformed colonies and no shift in the dose-response curve. Thus, these data do not support the notion that environmental exposures to magnetic fields contribute to transformation.

Short exposures to 60 Hz magnetic fields do not alter MYC expression in HL60 or Daudi cells.

Analysis of changes in gene expression induced by 60 Hz magnetic fields has been considered to support an association between exposure to magnetic fields and cancer risk. Several reports have indicated that these fields rapidly activate many genes in mammalian cells. However, previous studies in this area have not provided sufficient information to support the conclusions drawn. To clarify this controversial research, we have attempted to validate, under rigorously controlled conditions, key experiments on induction of gene expression by magnetic fields. An extensive series of experiments, incorporating critical improvements in experimental design, most notably blind exposures and internal standards, was performed with human HL60 and Daudi cells. Exposure conditions covered a range of flux densities (5.7 microT to 10 mT) and times (20-60 min). No alteration in the human MYC gene, commonly referred to as c-myc, or beta-actin steady-state mRNA levels was observed. The lack of an effect was not attributable to exposure geometry, timing of RNA preparation, or serum lot and concentration. To eliminate any remaining variables, exact replication was performed in one of the laboratories previously reporting gene expression effects; again, no evidence for altered MYC expression was found. Finally, differential display PCR indicated that extremely low-frequency magnetic field-induced changes in gene expression were not prevalent.

Physical mapping and characterization of a catabolic plasmid from the deep-subsurface bacterium Sphingomonas sp. strain F199.

A supercoiled 180-kb plasmid, pNL1, has been isolated from the deep-subsurface, chemoheterotrophic Sphingomonas sp. strain F199, and a physical map was generated. Analysis of a pNL1-derived cosmid library indicated that catechol 2,3-dioxygenase activity was linked to two distinct regions of the plasmid. Thus, the genes for aromatic catabolism in this Sphingomonas strain are, at least in part, plasmid encoded.

Maps from two interspecific backcross DNA panels available as a community genetic mapping resource.

We established two mouse interspecific backcross DNA panels, one containing 94 N2 animals from the cross (C57BL/6J x Mus spretus)F1 x C57BL/6J, and another from 94 N2 animals from the reciprocal backcross (C57BL/6J x SPRET/Ei)F1 x SPRET/Ei. We prepared large quantities of DNA from most tissues of each animal to create a community resource of interspecific backcross DNA for use by laboratories interested in mapping loci in the mouse. Initial characterization of the genetic maps of both panels has been completed. We used MIT SSLP markers, proviral loci, and several other sequence-defined genes to anchor our maps to other published maps. The BSB panel map (from the backcross to C57BL/6J) contains 215 loci and is anchored by 45 SSLP and 32 gene sequence loci. The BSS panel map (from the backcross to SPRET/Ei) contains 451 loci and is anchored by 49 SSLP loci, 43 proviral loci, and 60 gene sequence loci. To obtain a high density of markers, we used motif-primed PCR to "fingerprint" the panel DNAs. We constructed two maps, each representing one of the two panels. All new loci can be located with a high degree of certainty on the maps at current marker density. Segregation patterns in these data reveal several examples of transmission ratio distortion and permit analysis of the distribution of crossovers on individual chromosomes.

Fingerprinting genomes by use of PCR with primers that encode protein motifs or contain sequences that regulate gene expression.

PCR primers of arbitrary nucleotide sequence have identified DNA polymorphisms useful for genetic mapping in a large variety of organisms. Although technically very powerful, the use of arbitrary primers for genome mapping has the disadvantage of characterizing DNA sequences of unknown function. Thus, there is no reason to anticipate that DNA fragments amplified by use of arbitrary primers will be enriched for either transcribed or promoter sequences that may be conserved in evolution. For these reasons, we modified the arbitrarily primed PCR method by using oligonucleotide primers derived from conserved promoter elements and protein motifs. Twenty-nine of these primers were tested individually and in pairwise combinations for their ability to amplify genomic DNA from a variety of species including various inbred strains of laboratory mice and Mus spretus. Using recombinant inbred strains of mice, we determined the chromosomal location of 27 polymorphic fragments in the mouse genome. The results demonstrated that motif sequence-tagged PCR products are reliable markers for mapping the mouse genome and that motif primers can also be used for genomic fingerprinting of many divergent species.

Localization of the gene for the trans-acting transcription factor Sp1 to the distal end of mouse chromosome 15.

The mouse chromosomal location for the gene (Sp1-1) encoding the trans-acting transcription factor Sp1 has been determined. Analysis of restriction fragment length polymorphisms in recombinant inbred, congenic, and interspecific backcross mice using human and mouse cDNA probes demonstrated that Sp1-1 is a single gene closely linked to the mammary tumor virus integration site-1 (Int-1) on the distal end of chromosome 15. Sp1 is a zinc finger protein, but Sp1-1 is not closely linked to any of the other zinc finger protein genes that have been mapped in mouse. Int-1 and other markers flanking the Sp1-1 locus are part of a conserved linkage group represented on human chromosome 12q.

SV40 stimulates expression of the transacting factor Sp1 at the mRNA level.

Expression of the trans-acting transcription factor Sp1 increased almost 10-fold after infection of cells by simian virus 40. This alteration, attributable to an early viral protein, occurred at the mRNA level beginning at 12 hr postinfection, shortly after the appearance of viral T antigen, and reached a plateau at 20 hr postinfection. The enhanced level of Sp1 message was accompanied by a marked increase in Sp1 protein in the cell nuclei. Furthermore, we have demonstrated that stimulation of Sp1 levels elevates expression from viral and cellular promoters. Enhancing the amount of this trans-acting factor may play a role in aiding the viral life cycle and in neoplastic transformation of infected cells.

Ultraviolet shadowing nucleic acids on nylon membranes.

We describe a method for the direct visualization of nucleic acids on nylon membranes. Nylon is weakly fluorescent under short wave ultraviolet light allowing membrane-bound nucleic acids to be detected with a sensitivity of 10 ng. This procedure involves no staining or destaining of the gels prior to transfer, does not require duplicate sample lanes or blots, and does not interfere with transfer of the nucleic acid to the membrane or subsequent hybridization.

A negative regulatory element with properties similar to those of enhancers is contained within an Alu sequence.

A negative regulatory element has been found within a member of the African green monkey Alu family of interspersed repeated sequences. This "reducer" element decreased transcription in both directions from a cellular simian virus 40-like bidirectional promoter independently of both orientation and position. The reducer was not promoter specific since it also decreased expression from the simian virus 40 early and human beta-globin promoters. In addition, the reducer decreased transcription from a polymerase III promoter. The reducer was contained in 38 base pairs of an Alu family member and interacted specifically with a monkey cell nuclear protein.