Decision-making about hormone replacement therapy by women in England and Scotland.

OBJECTIVE: The aim of this study was to explore women's decision-making about the balance of risks and benefits of taking hormone replacement therapy (HRT) based on the latest evidence from the Women's Health Initiative (WHI) trial of combined HRT. METHODS: Women aged 50-69 years, who were eligible for the Women's International Study of long Duration Oestrogen after Menopause (WISDOM) trial, were invited to participate in one of eight focus groups. Participants were asked to discuss their views about taking HRT based on the latest international evidence. RESULTS AND CONCLUSIONS: Eighty-two women participated overall. Qualitative content analysis was applied to the discussion transcripts. Women regarded the decisions they make about taking HRT as highly personal, and, for women currently taking HRT, the overwhelming reason for continuation was perceived improvement in quality of life regardless of either the risks or the benefits in the longer term.

Anaerobic initial reaction of n-alkanes in a denitrifying bacterium: evidence for (1-methylpentyl)succinate as initial product and for involvement of an organic radical in n-hexane metabolism.

A novel type of denitrifying bacterium (strain HxN1) with the capacity to oxidize n-alkanes anaerobically with nitrate as the electron acceptor to CO(2) formed (1-methylpentyl)succinate (MPS) during growth on n-hexane as the only organic substrate under strict exclusion of air. Identification of MPS by gas chromatography-mass spectrometry was based on comparison with a synthetic standard. MPS was not formed during anaerobic growth on n-hexanoate. Anaerobic growth with [1-(13)C]n-hexane or d(14)-n-hexane led to a 1-methylpentyl side chain in MPS with one (13)C atom or 13 deuterium atoms, respectively. This indicates that the 1-methylpentyl side chain originates directly from n-hexane. Electron paramagnetic resonance spectroscopy revealed the presence of an organic radical in n-hexane-grown cells but not in n-hexanoate-grown cells. Results point at a mechanistic similarity between the anaerobic initial reaction of n-hexane and that of toluene, even though n-hexane is much less reactive; the described initial reaction of toluene in anaerobic bacteria is an addition to fumarate via a radical mechanism yielding benzylsuccinate. We conclude that n-hexane is activated at its second carbon atom by a radical reaction and presumably added to fumarate as a cosubstrate, yielding MPS as the first stable product. When 2,3-d(2)-fumarate was added to cultures growing on unlabeled n-hexane, 3-d(1)-MPS rather than 2,3-d(2)-MPS was detected, indicating loss of one deuterium atom by an as yet unknown mechanism.

A prospective study of faecal bile acids and colorectal cancer.

A prospective study of 7079 people aged 45-74 recruited through general practices in South Wales, Herefordshire and Edinburgh, Scotland was undertaken to test the hypothesis that faecal bile acids are implicated in the causation of large bowel cancer. The population was recruited between 1974 and 1980 and the response rate for stool collection was 67%. Bile acid analyses were performed on those cases that presented by 1990. It was decided in advance to examine the hypothesis separately for left- and right-sided bowel cancer because of known epidemiological differences between the two sites and to exclude the cases presenting within 2 years of the stool sample from the analyses because the cancer could have been present at recruitment and might have possibly affected faecal bile acid concentrations. Each case (n = 51 left-sided and 8 right-sided) was matched with three controls by age (within 5 years), sex, place of residence and time of providing the stool sample (within 3 months). Statistical analyses using conditional logistic regression showed no significant differences between the left-sided cases and controls for any of the concentrations of individual bile acids, total bile acid concentrations, faecal neutral steroids, percentage bacterial conversion and the ratio of lithocholic acid to deoxycholic acid concentrations. There was a statistically significant (P = 0.021) association of the presence of chenodeoxycholic acid (5/8 samples) in the right-sided cases compared with the controls (3/23), odds ratio 6.26 (95% confidence interval 1.19, 32.84). A high proportion of primary bile acids has also been found in other studies of patients with a genetic predisposition to proximal bowel cancer, however this pattern may also occur in low risk groups, such as Indian vegetarians, suggesting that they may predispose to right-sided bowel cancer only in the presence of other, as yet unknown factors. If bile acids are involved in the causation of large bowel cancer, they may be part of a complex set of interacting factors.

Analysis of gamma-hydroxybutyrate (GHB) in urine by gas chromatography-mass spectrometry.

A simple method for the direct analysis of gamma-hydroxybutyrate (GHB) from human urine is described. The method uses solid-phase extraction, liquid-liquid extraction, and silyl-derivatization, then gas chromatographic-mass spectrometric analysis using GHB-d6 as the internal standard. The method was linear from 5 to 500 mg/L, and coefficients of variation were less than 10%. Twenty-six urine specimens previously analyzed by an existing method were analyzed and yielded GHB concentrations ranging from 0 to 6100 mg/L; the results correlated between the two methods. Compared with existing methods, the method described here is superior because it is specific to GHB and can discriminate between GHB and gamma-butyrolactone.

Anaerobic utilization of alkylbenzenes and n-alkanes from crude oil in an enrichment culture of denitrifying bacteria affiliating with the beta-subclass of Proteobacteria.

Denitrifying bacteria were enriched from freshwater sediment with added nitrate as electron acceptor and crude oil as the only source of organic substrates. The enrichment cultures were used as laboratory model systems for studying the degradative potential of denitrifying bacteria with respect to crude oil constituents, and the phylogenetic affiliation of denitrifiers that are selectively enriched with crude oil. The enrichment culture exhibited two distinct growth phases. During the first phase, bacteria grew homogeneously in the aqueous phase, while various C1-C3 alkylbenzenes, but no alkanes, were utilized from the crude oil. During the second phase, bacteria also grew that formed aggregates, adhered to the crude oil layer and emulsified the oil, while utilization of n-alkanes (C5 to C12) from the crude oil was observed. During growth, several alkylbenzoates accumulated in the aqueous phase, which were presumably formed from alkylbenzenes. Application of a newly designed, fluorescently labelled 16S rRNA-targeted oligonucleotide probe specific for the Azoarcus/Thauera group within the beta-subclass of Proteobacteria revealed that the majority of the enriched denitrifiers affiliated with this phylogenetic group.

Degradative capacities and 16S rRNA-targeted whole-cell hybridization of sulfate-reducing bacteria in an anaerobic enrichment culture utilizing alkylbenzenes from crude oil.

A mesophilic sulfate-reducing enrichment culture growing anaerobically on crude oil was used as a model system to study which nutritional types of sulfate-reducing bacteria may develop on original petroleum constituents in oil wells, tanks, and pipelines. Chemical analysis of oil hydrocarbons during growth revealed depletion of toluene and o-xylene within 1 month and of m-xylene, o-ethyltoluene, m-ethyltoluene, m-propyltoluene, and m-isopropyltoluene within approximately 2 months. In anaerobic counting series, the highest numbers of CFU (6 x 10(6) to 8 x 10(6) CFU ml-1) were obtained with toluene and benzoate. Almost the same numbers were obtained with lactate, a substrate often used for detection of the vibrio-shaped, incompletely oxidizing Desulfovibrio sp. In the present study, however, lactate yielded mostly colonies of oval to rod-shaped, completely oxidizing, sulfate-reducing bacteria which were able to grow slowly on toluene or crude oil. Desulfovibrio species were detected only at low numbers (3 x 10(5) CFU ml-1). In agreement with this finding, a fluorescently labeled, 16S rRNA-targeted oligonucleotide probe described in the literature as specific for members of the Desulfovibrionaceae (suggested family) hybridized only with a small portion (< 5%) of the cells in the enrichment culture. These results are consistent with the observation that known Desulfovibrio species do not utilize aromatic hydrocarbons, the predominant substrates in the enrichment culture. All known sulfate-reducing bacteria which utilize aromatic compounds belong to a separate branch, the Desulfobacteriaceae (suggested family). Most members of this family are complete oxidizers. For specific hybridization with members of this branch, the probe had to be modified by a nucleotide exchange. Indeed, this modified probe hybridized with more than 95% of the cells in the enrichment culture. The results show that completely oxidizing, alkylbenzene-utilizing sulfate-reducing bacteria rather than Desulfovibrio species have to be considered in attempts to understand the microbiology of sulfide production in oil wells, tanks, and pipelines when no electron donors other than the indigenous oil constituents are available.

Degradation of Chlorinated Dibenzofurans and Dibenzo-p-Dioxins by Sphingomonas sp. Strain RW1.

The ability of the dibenzofuran- and dibenzo-p-dioxin-mineralizing bacterium Sphingomonas sp. strain RW1 (R.-M. Wittich, H. Wilkes, V. Sinnwell, W. Francke, and P. Fortnagel, Appl. Environ. Microbiol. 58:1005-1010, 1992) to oxidize chlorinated derivatives of dibenzofuran and dibenzo-p-dioxin was analyzed. Strain RW1 degraded several mono- and dichlorinated dibenzofurans and dibenzo-p-dioxins, but it did not degrade more highly chlorinated congeners. Most mono- and dichlorinated dibenzofurans and dibenzo-p-dioxins investigated in this study were degraded to the corresponding mono- and dichlorinated salicylates and catechols, respectively, together with salicylate and catechol. This indicates an initial dioxygenolytic attack on the substituted as well as on the nonsubstituted aromatic nucleus of most of the target compounds. Strain RW1 could not grow at the expense of monochlorinated dibenzo-p-dioxins and dibenzofurans as carbon sources, with the exception of 4-chlorodibenzofuran, which was stoichiometrically converted to 3-chlorosalicylate.

Metabolism of Hydroxydibenzofurans, Methoxydibenzofurans, Acetoxydibenzofurans, and Nitrodibenzofurans by Sphingomonas sp. Strain HH69.

The metabolism of 11 substituted dibenzofurans by the dibenzofuran-degrading Sphingomonas sp. strain HH69 was investigated. Strain HH69 utilizes 2-, 3-, and 4-acetoxydibenzofuran as well as 2-, 3-, and 4-hydroxydibenzofuran as sole sources of carbon and energy. The degradation of acetoxydibenzofurans is initiated by hydrolysis of the ester bonds, yielding the corresponding hydroxydibenzofurans and acetate. Strain HH69 grew on 2-methoxydibenzofuran only after it was adapted to the utilization of 5-methoxysalicylic acid, whereas 3- and 4-methoxydibenzofuran as well as 2- and 3-nitrodibenzofuran were only cooxidized. During the breakdown of all eight hydroxy-, methoxy-, and nitrodibenzofurans studied here, the corresponding substituted salicylic acids accumulated in the culture broth. In the cases of 2- and 3-hydroxydibenzofuran as well as 2- and 3-nitrodibenzofuran, salicylic acid was also formed. Those four dibenzofurans which did not serve as carbon sources for strain HH69 were converted to a nonutilizable salicylic acid derivative. From turnover experiments with the mutant HH69/II, which is deficient in meta-cleavage, 2,2(prm1),3,4(prm1)-tetrahydroxybiphenyl, 2,2(prm1),3-trihydroxy-5(prm1)-methoxybiphenyl, 2,2(prm1),3-trihydroxy-5(prm1)-nitrobiphenyl, and 2,2(prm1),3-trihydroxy-4(prm1)-nitrobiphenyl were isolated as the main products formed from 3-hydroxydibenzofuran, 2-methoxydibenzofuran, and 2- and 3-nitrodibenzofuran, respectively. These results indicate significant regioselectivity for the dioxygenolytic cleavage of the ether bond of these monosubstituted dibenzofurans, with a preference for the nonsubstituted aromatic nucleus. Substituted trihydroxybiphenyls are converted further by meta-cleavage followed by the removal of the side chain of the resulting product. A stepwise degradation of this side chain was found to be involved in the metabolism of 2-hydroxydibenzofuran.

Biotransformation of diphenyl ether by the yeast Trichosporon beigelii SBUG 752.

Trichosporon beigelii SBUG 752 was able to transform diphenyl ether. By TLC, HPLC, GC, GC-MS, NMR- and UV-spectroscopy, several oxidation products were identified. The primary attack was initiated by a monooxygenation step, resulting in the formation of 4-hydroxydiphenyl ether, 2-hydroxydiphenyl ether and 3-hydroxydiphenyl ether (48:47:5). Further oxidation led to 3,4-dihydroxydiphenyl ether. As a characteristic product resulting from the cleavage of an aromatic ring, the lactone of 2-hydroxy-4-phenoxymuconic acid was identified. The possible mechanism of ring cleavage to yield this metabolite is discussed.

Hormone replacement therapy and cardiovascular disease: the case for a randomized controlled trial.

The menopause is associated with an increased risk of developing cardiovascular disease. Oestrogen may influence various metabolic pathways which contribute to the pathogenesis of cardiovascular disease, and observational studies suggest that in postmenopausal women oral oestrogen replacement therapy confers some protection against coronary heart disease and to a lesser extent against stroke. What is not clear is the magnitude of the cardioprotective effect and the overall balance of long-term benefits and hazards. Research is also required to establish the relative effects of oestrogen replacement therapy and combined or opposed hormone replacement therapy (HRT) where progestogen is added to counter the proliferative action of oestrogen on the endometrium. A large randomized controlled trial is the only way to provide accurate estimates of the cardioprotective effect of HRT and of other long-term benefits and hazards. Feasibility studies undertaken through the UK Medical Research Council (MRC) General Practice Research Framework show that such a trial is acceptable to patients and their doctors. Recruitment and withdrawal rates indicate that a trial of sufficient size to show a 25% reduction in cardiovascular disease with 90% power at the 1% level would be feasible. The full trial is costly and it is proposed that the UK collaborates with other countries in a major international trial to complement the Women's Health Initiative trial in the USA. Feasibility studies in Europe are underway, the design and scientific rationale for the trial have been approved by the UK MRC and it is hoped that recruitment to the full-scale trial can begin soon.

Anaerobic oxidation of hydrocarbons in crude oil by new types of sulphate-reducing bacteria.

Many crude oil constituents are biodegradable in the presence of oxygen; however, a substantial anaerobic degradation has never been demonstrated. An unusually low content of n-alkanes in oils of certain deposits is commonly attributed to selective utilization of these hydrocarbons by aerobic microorganisms. On the other hand, oil wells and production fluids were shown to harbour anaerobic sulphate-reducing bacteria, but their actual electron donors and carbon sources were unknown. On the basis of nutritional properties of various bacterial isolates it was assumed that fatty acids and H2 are potential electron donors for sulphate reduction in situ. Here we demonstrate that hydrocarbons in crude oil are used directly by sulphate-reducing bacteria growing under strictly anoxic conditions. A moderately thermophilic pure culture selectively utilizes n-alkanes in oil for sulphate reduction to sulphide. In addition, a mesophilic sulphate-reducing enrichment culture is shown to oxidize alkylbenzenes in oil. Thus, sulphate-reducing bacteria utilizing aliphatic and aromatic hydrocarbons as electron donors may present a significant source of sulphide in oil deposits and oil production plants.

The gastrointestinal toxicity of aspirin: an overview of randomised controlled trials.

The proven benefit of aspirin in the secondary prevention of cardiovascular disease and its possible value in primary prevention must be weighted against its potential hazards. This paper is an overview of the gastrointestinal toxicity of aspirin, its most serious complication after intracerebral haemorrhage. Information on toxicity has been drawn only from randomised trials, thus avoiding the potential biases of observational studies. All randomised placebo controlled trials listed in the Anti-platelet Trialists Collaboration where a direct aspirin-placebo comparison was possible were included. Twenty-one trials were included, all but one of secondary prevention. There were over 75,000 person years of aspirin exposure. The pooled odds ratios for categories of gastrointestinal bleeding (e.g. haematemesis, melaena) were between 1.5-2.0; fatal bleeds were very rare. The risk of peptic ulcers was 1.3 and of upper gastrointestinal symptoms 1.7. These risks were lower than those found in observational studies. Attributable disease rates are also presented. For haematemesis for example they varied from 0.2-1.0 per 1000 person years. Toxicity was dose related. Aspirin does have significant gastrointestinal toxicity, although this is rarely fatal. More recent work has demonstrated the efficacy of low doses of aspirin (75 mg daily) but there is limited information yet available on its toxicity.

Biodegradation of diphenyl ether and its monohalogenated derivatives by Sphingomonas sp. strain SS3.

The bacterium Sphingomonas sp. strain SS3, which utilizes diphenyl ether and its 4-fluoro, 4-chloro, and (to a considerably lesser extent) 4-bromo derivatives as sole sources of carbon and energy, was enriched from soil samples of an industrial waste deposit. The bacterium showed cometabolic activities toward all other isomeric monohalogenated diphenyl ethers. During diphenyl ether degradation in batch culture experiments, phenol and catechol were produced as intermediates which were then channeled into the 3-oxoadipate pathway. The initial step in the degradation follows the recently discovered mechanism of 1,2-dioxygenation, which yields unstable phenolic hemiacetals from diphenyl ether structures. Oxidation of the structure-related dibenzo-p-dioxin yielded 2-(2-hydroxyphenoxy)-muconate upon ortho cleavage of the intermediate 2,2',3-trihydroxydiphenyl ether. Formation of phenol, catechol, halophenol, and halocatechol from the conversion of monohalogenated diphenyl ethers gives evidence for a nonspecific attack of the dioxygenating enzyme system.

Genetic factors determining thrombosis and fibrinolysis.

Raised plasma levels of fibrinogen, factor VIIc, and plasminogen activator inhibitor-1 (PAI-1) are associated with an increased risk of ischemic heart disease. Levels of these proteins are determined in part by environmental influences such as smoking and dietary fat intake. However, genetic variation explains much of the interindividual variation in plasma levels of these proteins not accounted for by environmental factors. We previously investigated the DNA variation at the fibrinogen gene locus and showed that BclI restriction fragment length polymorphism (RFLP) of the beta-fibrinogen gene is associated with between-person differences in plasma fibrinogen levels. This RFLP is unlikely to be the functional base change itself, since it lies downstream of the gene. The rate-limiting step in the production of the mature fibrinogen molecule in the human hepatoma cell-line HepG2 is the synthesis of the beta-polypeptide chain, which in turn is influenced by the amount of messenger (mRNA) available. One possibility is that BclI RFLP is in linkage disequilibrium with a base change in the region of the beta-gene controlling synthesis of its mRNA and ultimately of fibrinogen protein. We identified a base change in the 5'-flanking region of the beta-fibrinogen gene that is in linkage disequilibrium with the BclI RFLP, that is associated with plasma fibrinogen levels, and that may be involved in control of fibrinogen gene expression. For the factor VII gene, we identified a polymorphism, detected after Msp I digestion of polymerase chain reaction (PCR)-amplified genomic DNA, that is strongly associated with factor VII coagulant activity (factor VIIc). The base change that creates the Msp I polymorphism is a G to A substitution, leading to the replacement of arginine (Arg) with glutamine (Gln) in the protein product of the M2 allele. In a sample of 284 men from the United Kingdom the frequency of the Gln allele (M2 loss of cutting site) is 0.1, and individuals of genotype Arg/Gln have factor VIIc levels 22% below the sample mean. In this sample, the Msp I genotype was found to be the strongest predictor of factor VIIc, accounting for 20.2% of the variance, with cholesterol accounting for an additional 3.5%. Three individuals homozygous for the Gln allele had both low factor VIIc and low factor VII protein concentrations. The conformation of the factor VII Gln may be different from that of the Arg protein, affecting its intracellular processing, secretion, turnover in plasma, or activity.(ABSTRACT TRUNCATED AT 400 WORDS)

Extra-cranial bleeding and other symptoms due to low dose aspirin and low intensity oral anticoagulation.

Data from the early stages of the thrombosis prevention trial (TPT) have been used to establish and quantify the risk of extracranial bleeding due to low dose aspirin (75 mg) and low intensity oral anticoagulation with warfarin (international normalised ratio, INR, 1.5) singly or in combination, in men aged between 45 and 69 who are at high risk of ischaemic heart disease (IHD). The design of the trial is factorial, the four treatments being combined low dose aspirin and low intensity anticoagulation (WA), low intensity anticoagulation alone (W), low dose aspirin alone (A) and double placebo treatment (P). The trial is being carried out through the Medical Research Council's General Practice Research Framework, with participating practices throughout the United Kingdom. Results are based on the first 3,667 men entered. The risk of major gastrointestinal bleeding due to active treatment is probably about 1 in 500 man-years of treatment, there currently being no difference between the three active regimes (WA, W, A). Intermediate and minor bleeding episodes occur more frequently with WA than with W or A on their own, the excess being mainly due to minor nose bleeds and bruises. In turn, both W and A on their own cause more such minor episodes than placebo treatment, P. There is no evidence that any of the three active regimens increases the risk of peptic ulceration, nor do they increase reports of indigestion. Aspirin increases reports of constipation and reduces reports of blurred vision. Minor bleeding occurs less frequently in smokers than in non-smokers but is not influenced by age. The antithrombotic regimes used are feasible and acceptable.

Gemfibrozil reduces plasma prothrombin fragment F1 + 2 concentration, a marker of coagulability, in patients with coronary heart disease.

The effects of gemfibrozil on several indices of haemostatic activity were explored in male patients with coronary heart disease (CHD). Sixty-three of 71 patients completed a crossover study in which gemfibrozil 1,200 mg/day and matching placebo were each taken in randomised order for 2 months in a double-blind manner, separated by a 2-month washout period. Serum cholesterol decreased by an average (95% confidence interval) of 12 (9 to 15)% and non-fasting triglyceride concentration by 43 (34 to 51)% during active treatment. Plasma prothrombin fragment F1 + 2 concentration, a marker of the in vivo rate of generation of thrombin, was 25 (12 to 37)% lower on average while on gemfibrozil than during the placebo phase. Factor VII coagulant activity (VIIc) and antigen concentration, and fibrinopeptide A concentration were not influenced by gemfibrozil in the group overall. However, the VIIc response appeared to be dependent upon the untreated cholesterol level. Hypercholesterolaemic men (cholesterol greater than 6.5 mmol/l) experienced a significant reduction in VIIc averaging 6% of standard during active therapy. Other effects of gemfibrozil were a 5 (2 to 9)% increase in plasma fibrinogen by a gravimetric method, an 11 (8 to 13)% increase in platelet count, and a 6 (2 to 10)% reduction in white cell count. The reduced incidence of CHD following gemfibrozil therapy in hyperlipidaemic patients may arise in part through a reduction in procoagulant activity and thus the risk of an occlusive coronary thrombosis.

Metabolism of dibenzo-p-dioxin by Sphingomonas sp. strain RW1.

In the course of our screening for dibenzo-p-dioxin-utilizing bacteria, a Sphingomonas sp. strain was isolated from enrichment cultures inoculated with water samples from the river Elbe. The isolate grew with both the biaryl ethers dibenzo-p-dioxin and dibenzofuran (DF) as the sole sources of carbon and energy, showing doubling times of about 8 and 5 h, respectively. Biodegradation of the two aromatic compounds initially proceeded after an oxygenolytic attack at the angular position adjacent to the ether bridge, producing 2,2',3-trihydroxydiphenyl ether or 2,2',3-trihydroxybiphenyl from the initially formed dihydrodiols, which represent extremely unstable hemiacetals. Results obtained from determinations of enzyme activities and oxygen consumption suggest meta cleavage of the trihydroxy compounds. During dibenzofuran degradation, hydrolysis of 2-hydroxy-6-oxo-6-(2-hydroxyphenyl)-hexa-2,4-dienoate yielded salicylate, which was branched into the catechol meta cleavage pathway and the gentisate pathway. Catechol obtained from the product of meta ring fission of 2,2',3-trihydroxydiphenyl ether was both ortho and meta cleaved by Sphingomonas sp. strain RW1 when this organism was grown with dibenzo-p-dioxin.