Reproductive factors, exogenous hormone use and risk of hepatocellular carcinoma among US women: results from the Liver Cancer Pooling Project.

BACKGROUND: Hepatocellular carcinoma (HCC) occurs less commonly among women than men in almost all regions of the world. The disparity in risk is particularly notable prior to menopause suggesting that hormonal exposures during reproductive life may be protective. Exogenous oestrogenic exposures such as oral contraceptives (OCs), however, have been reported to increase risk, suggesting that estrogens may be hepatocarcinogenic. To examine the effects of reproductive factors and exogenous hormones on risk, we conducted a prospective analysis among a large group of US women. METHODS: In the Liver Cancer Pooling Project, a consortium of US-based cohort studies, data from 799,500 women in 11 cohorts were pooled and harmonised. Cox proportional hazards regression models were used to generate hazard ratios (HRs) and 95% confidence intervals (CIs) for the associations of reproductive factors and exogenous hormones with HCC (n=248). RESULTS: Bilateral oophorectomy was associated with a significantly increased risk of HCC (HR=2.67, 95% CI=1.22-5.85), which did not appear to be related to a shorter duration of exposure to endogenous hormones or to menopausal hormone therapy use. There was no association between OC use and HCC (HR=1.12, 95% CI=0.82-1.55). Nor were there associations with parity, age at first birth, age at natural menopause, or duration of fertility. CONCLUSIONS: The current study suggests that bilateral oophorectomy increases the risk of HCC but the explanation for the association is unclear. There was no association between OC use and HCC risk. Examination of endogenous hormone levels in relation to HCC may help to clarify the findings of the current study.

Metabolic syndrome and insulin resistance in relation to biliary tract cancer and stone risks: a population-based study in Shanghai, China.

BACKGROUND: Serum lipids, diabetes, and obesity, individual components of metabolic syndrome, are associated with biliary tract cancer and stone risk, but the associations of metabolic syndrome or insulin resistance with biliary tract cancers and stones are not well studied. METHODS: In this population-based case-control study in Shanghai, China (627 biliary tract cancers, 1037 biliary stones, and 959 controls), metabolic syndrome was defined as the presence of any three of the five components, including high waist circumference, high triglycerides, low high-density lipoprotein cholesterol (HDL), high blood pressure, and diabetes. Insulin resistance and ß-cell function were assessed, using homeostasis assessment models. RESULTS: Metabolic syndrome was significantly associated with gallbladder cancer (odds ratio (OR)=2.75, 95% confidence interval (95% CI)=1.82-4.15) and biliary stones (OR=1.64, 95% CI=1.24-2.16), with a significant dose effect with increasing number of metabolic syndrome components (P trend <0.0001). The observed association persisted among subjects without a history of diabetes. The association between insulin resistance and gallbladder cancer was borderline (P trend=0.06). There was a significant inverse association between ß-cell function and gallbladder cancer risk (P trend <0.001). CONCLUSION: Our findings suggest that metabolic syndrome and insulin resistance have a role in the aetiology of biliary tract cancers and biliary stones, and if confirmed, they imply that lifestyle control of these factors may lower the risk of biliary stones and biliary tract cancer.

Diabetes in relation to biliary tract cancer and stones: a population-based study in Shanghai, China.

BACKGROUND: Biliary tract cancers are rare but fatal malignancies. Diabetes has been related to biliary stones, but its association with biliary tract cancers is less conclusive. METHODS: In a population-based case-control study of 627 cancers, 1037 stones, and 959 controls in Shanghai, China, we examined the association between diabetes and the risks of biliary tract cancer and stones, as well as the effect of potential mediating factors, including serum lipids and biliary stones (for cancer), contributing to the causal pathway from diabetes to biliary diseases. RESULTS: Independent of body mass index (BMI), diabetes was significantly associated with gallbladder cancer and biliary stones ((odds ratio (OR) (95% confidence interval)=2.6 (1.5-4.7) and 2.0 (1.2-3.3), respectively). Biliary stones and low serum levels of high-density lipoprotein (HDL) were significant mediators of the diabetes effect on gallbladder cancer risk, accounting for 60 and 17% of the diabetes effect, respectively. High-density lipoprotein was also a significant mediator of the diabetes effect on biliary stones, accounting for 18% of the diabetes effect. CONCLUSIONS: Independent of BMI, diabetes is a risk factor for gallbladder cancer, but its effect is mediated in part by biliary stones and serum HDL levels, suggesting that gallbladder cancer risk may be reduced by controlling diabetes, stones, and HDL levels.

Reproductive factors and risks of biliary tract cancers and stones: a population-based study in Shanghai, China.

BACKGROUND: Parity has been linked to gallbladder cancer and gallstones, but the effects of other reproductive factors are less clear. METHODS: We examined 361 incident biliary tract cancer cases, 647 biliary stone cases, and 586 healthy women in a population-based study in Shanghai. RESULTS: The effects of parity (odds ratios, OR(> or =3 vs 1 child)=2.0, 95% confidence interval (CI) 0.7-5.1), younger age at first birth (OR(per 1-year decrease)=1.2, 95% CI 0.99-1.6), and older age at menarche (OR(per 1-year increase)=1.4, 95% CI 1.1-1.8) on gallbladder cancer risk were more pronounced among women with stones, but the interactions were not significant. CONCLUSION: Our results provide support for high parity, younger age at first birth, and late age at menarche in the development of gallbladder cancer, particularly among women with biliary stones.

Indole-3-glycerol-phosphate synthase from Sulfolobus solfataricus as a model for studying thermostable TIM-barrel enzymes.

Indole-3-glycerol-phosphate synthase, a thermophilic and thermostable enzyme from the archaeon Sulfolobus solfataricus, was purified and characterized. The sequence of the thermophilic enzyme was compared to the sequence of a homologous mesophilic enzyme from Escherichia coli. The secondary structure of the thermophilic enzyme was predicted taking into account the patterns of hydropathy, chain flexibility and amphipathicity and the CD spectrum. From this analysis it turned out that indole-3-glycerol-phosphate synthase from S. solfataricus can be considered a model for studying thermostable TIM-barrel enzymes. Some peculiarities of the amino-acid sequence of indole-3-glycerol-phosphate synthase from S. solfataricus are discussed in relation to the thermostability of the enzyme.

An extremely thermostable aromatic aminotransferase from the hyperthermophilic archaeon Pyrococcus furiosus.

Pyrococcus furiosus is a strictly anaerobic archaeon (formerly archaebacterium) that grows optimally at 100 degrees C by the fermentation of peptides. Cell-free extracts were found to contain two distinct aromatic aminotransferases (ArAT, EC 2.6.1.57), one of which was purified to electrophoretic homogeneity. P. furiosus ArAT is a homodimer with a subunit M(r) value of 44,000 +/- 1000. Using 2-ketoglutarate as the amino acceptor, the purified enzyme catalyzed the pyridoxal 5'-phosphate (PMP)-dependent transamination of phenylalanine, tyrosine and tryptophan with respective kcat values of 253, 72 and 62 (s-1 at 80 degrees C) under saturating conditions. The Km values for all three amino acids were between 1.1 and 2.1 mM and the optimum temperature for catalysis was above 95 degrees C. The melting point for the pure enzyme was also above 95 degrees C as determined by the change in ellipticity at 220 nm. Irreversible denaturation of the pure enzyme was not apparent after 6 h at 80 degrees C in the presence of PMP and 2-ketoglutarate and the time required for a 50% loss in activity at 95 degrees C was approx. 16 h. This decreased to approx. 12 h if cofactor and substrate were not added. In contrast, the apoenzyme (lacking PMP) lost most (70%) of its activity (measured after reconstitution) after 6 h at 80 degrees C, indicating that both PMP and 2-ketoglutarate stabilize the enzyme at extreme temperatures. Although few ArATs have been characterized to date, the molecular properties and substrate specificity of P. furiosus ArAT more resemble those of the ArAT from Escherichia coli than those of the analogous enzyme from rat liver. Moreover, the P. furiosus enzyme is by far the most thermostable aminotransferase of any type to be purified so far.

The active site of Sulfolobus solfataricus aspartate aminotransferase.

Aspartate aminotransferase from the archaebacterium Sulfolobus solfataricus binds pyridoxal 5' phosphate, via an aldimine bond, with Lys-241. This residue has been identified by reducing the enzyme in the pyridoxal form with sodium cyanoboro[3H]hydride and sequencing the specifically labeled peptic peptides. The amino acid sequence centered around the coenzyme binding site is highly conserved between thermophilic aspartate aminotransferases and differs from that found in mesophilic isoenzymes. An alignment of aspartate aminotransferase from Sulfolobus solfataricus with mesophilic isoenzymes, attempted in spite of the low degree of similarity, was confirmed by the correspondence between pyridoxal 5' phosphate binding residues. Using this alignment it was possible to insert the archaebacterial aspartate aminotransferase into a subclass, subclass I, of pyridoxal 5' phosphate binding enzymes comprising mesophilic aspartate aminotransferases, tyrosine aminotransferases and histidinol phosphate aminotransferases. These enzymes share 12 invariant amino acids most of which interact with the coenzyme or with the substrates. Some enzymes of subclass I and in particular aspartate aminotransferase from Sulfolobus solfataricus, lack a positively charged residue, corresponding to Arg-292, which in pig cytosolic aspartate aminotransferase interacts with the distal carboxylate of the substrates (and determines the specificity towards dicarboxylic acids). It was confirmed that aspartate aminotransferase from Sulfolobus solfataricus does not possess any arginine residue exposed to chemical modifications responsible for the binding of omega-carboxylate of the substrates. Furthermore, it has been found that aspartate aminotransferase from Sulfolobus solfataricus is fairly active when alanine is used as substrate and that this activity is not affected by the presence of formate. The KM value of the thermophilic aspartate aminotransferase towards alanine is at least one order of magnitude lower than that of the mesophilic analogue enzymes.

The nature, incidence, and impact of eye injuries among US military personnel: implications for prevention.

OBJECTIVES: To assess incidence rates of eye injuries in the US Armed Forces and to identify demographic and occupational correlates of risk. DESIGN: Retrospective population-based study. SETTING: US military medical facilities worldwide. PARTICIPANTS: All individuals in the US Armed Forces during 1998. MAIN OUTCOME MEASURES: Incidence rates of hospitalizations and ambulatory visits for eye injuries. RESULTS: The incidence rate of ambulatory visits (983 per 100 000 person-years) for eye injuries was 58 times higher than the incidence rate of hospitalizations (17 per 100 000 person-years) for eye injuries. Orbital floor fractures, contusions, and open wounds to the ocular adnexa and orbit accounted for 85% of eye injuries resulting in hospitalization, while 80% of ambulatory visits were for superficial wounds and foreign bodies. Hospitalization rates varied widely across demographic subgroups. Men had twice the incidence rate as women, and the youngest age group (17-24 years) had 6 times the incidence rate of the oldest age group (35-65 years). Together, motor vehicle crashes and fights caused nearly half of the hospitalizations. Ambulatory rates varied significantly in relation to occupation but not to demography. Tradespeople (eg, metal body machinist, welder, and metalworker) had incidence rates 3 to 4 times higher than the overall population rate. CONCLUSIONS: Hospitalization and ambulatory data provide different views of the morbidity associated with eye injuries. General safety precautions and behavior modification, rather than eye-specific interventions, are indicated to prevent the most serious eye injuries. However, the consistent use of eye protection during known hazardous occupational activities could prevent much of the morbidity associated with the less serious, yet more common, eye injuries.

Single-well reactive tracer test and stable isotope analysis for determination of microbial activity in a fast hydrocarbon-contaminated aquifer.

Single-well reactive tracer tests, such as the push-pull test are useful tools for characterising in-situ bioattenuation processes in contaminated aquifers. However, the analytical models that are used to interpret push-pull data may be over-simplified, and potentially overlook important processes responsible for the frequent discrepancy between predicted and observed results obtained from push-pull tests. In this study, the limitations underlying the push-pull test methodology were investigated and were supported with results from a push-pull test conducted in a sulphate-reducing aquifer contaminated by crude oil. Poor (<7%) mass recovery was achieved when the push-pull test was performed in a fast-flowing aquifer, preventing a quantifiable reaction rate to be determined. Breakthrough curve data were unexplainable using simplified analytical models, but exhibited trends analogous with tests conducted by others, when >20% mass recoveries were achieved. Push-pull test data collected from sulphate-reducing aquifers indicate that the assumption of a well-mixed batch reactor system is incorrect and that reaction rates obtained from push-pull tests in such systems may be affected by the extraction regime implemented. Evidence of microbial respiration of the reactive tracer was provided by stable sulphur isotope analysis, from which an isotope fractionation factor of +9.9 +/- 8.1 per thousand was estimated. The stable isotope data support the argument that reaction rates calculated using push-pull tests are not uniformly distributed in space and time and are likely to be influenced by heterogeneities in the flow field.

[Clinical assessment of a new oral bacterial treatment for children with acute diarrhea]. / Valutazione clinica di un nuovo batterioterapico orale in pazienti di età pediatrica con diarrea acuta.

This study was carried out to evaluate the efficacy of a new oral bacteriotherapeutic drug in 63 children under 4 years old affected by infective diarrhoea or by extraintestinal infective pathology caused by oral antibiotic therapy. The patients were divided into three groups: the first and the second included subjects affected by acute diarrhoea enrolled in a comparative and controlled study; the third included subjects in chemotherapeutic treatment because suffering from infective extraintestinal pathologies, these patients were enrolled in a not controlled study. The first group underwent the rehydratant treatment for the acute enteritis together with the therapy with inactivated Lactobacillus acidophilus LB to value its antidiarrheic activity. To the second group (control group) was administered a no bacteriotherapeutic drug, while to the third group was administered the antibiotic therapy for extra-intestinal pathologies together with inactivated Lactobacillus acidophilus to avoid possible phenomena due to intestinal dysmicrobism. For all the patients included in the study, were reported; the number, the characteristics, the consistency of the motions and the description of the symptomatology during the treatment and daily. The greater part of the patients affected by the intestinal infective pathology subdued bacteriologic tests by coproculture. The obtained results showed a statistically significant clinical improvement of the first group patients compared with those of the second group, while those of the third group showed that the drug activity prevents the intestinal dysmicrobism affections due to the antibiotic therapy.

Determinants of high pesticide exposure events in the agricultural health cohort study from enrollment (1993-1997) through phase II (1999-2003).

We conducted an analysis of the determinants of high pesticide exposure events (HPEEs), which are defined as self-reported incidents of high exposure to pesticides, fertilizers, or other chemicals in the Agricultural Health Study, a cohort of private applicators and their spouses residing in North Carolina or Iowa, and commercial applicators residing in Iowa. We examined the risk of HPEEs occurring between enrollment (phase 1: 1993-1997) and follow-up (phase II: 1999-2003) among participants who completed the phase II questionnaire (n=43,149) by calculating hazard rate ratios and 95% confidence intervals using Cox proportional-hazard regression. During the followup period, 1,582 HPEEs were reported (3.8%). HPEE risk was significantly higher among Iowa residents, younger participants, those with a hearing deficit, a risk-taking personality, and an HPEE prior to enrollment. Among private applicators (n=30,102), larger farm size, higher frequency and duration of pesticide use, spraying pesticides with open cab windows, using a tractor cab without a charcoal filter, repairing spray equipment, wearing work clothing more than two days without changing, not removing work boots before entering the home, and storing pesticides in the home were associated with significantly higher HPEE risk. Among commercial applicators (n=2326), higher frequency of pesticide use was associated with a significantly higher HPEE risk. Among spouses (n=10,721), higher frequency of pesticide use, using an application vehicle with a cab, and storing pesticides in the home were associated with a significantly higher HPEE risk. Our findings indicate that HPEEs were associated with several modifiable pesticide handling procedures that can be targeted in safety training and education.

Milk identification of different species: 13C-NMR spectroscopy of triacylglycerols from cows and buffaloes' milks.

Triacylglycerols from cows and buffaloes' milk fat were investigated by 13C nuclear magnetic resonance (NMR) spectroscopy. By the addition of pure triacylglycerols standards, we identified the resonances of both milk fats, and the peaks were used for qualitative and quantitative analysis of acyl groups. Multivariate analysis treatment of triacylglycerols distribution and composition parameters enabled us to identify milk. This study shows that NMR can safely be used to quantitate milk fatty acid content, providing unique information for milk identification of different animal species.

Stability of a thermophilic TIM-barrel enzyme: indole-3-glycerol phosphate synthase from the thermophilic archaeon Sulfolobus solfataricus.

The stability and activity of indole-3-glycerol phosphate synthase from Sulfolobus solfataricus were studied as a function of pH and temperature. In this paper we focus on three points: (1) the long-term stability of the protein to irreversible denaturation at high temperature; (2) the short-term stability of the protein to reversible temperature-driven unfolding; and (3) the dependence of its activity on temperature. Results can be summarized as follows: (a) the same first-order kinetic constant (0.020+/-0.003 min-1) was determined at different pH values (6.5, 8.0 and 9.5) from long-term stability experiments at 80 degrees C; (b) short-term stability experiments revealed different behaviour in two different pH ranges (6.5-8.0, 8.5-9.5), suggesting that the melting temperature is higher at alkaline than at neutral pH; (c) the dependence of activity on temperature was investigated at pH 7.0 and 9.0, and a discontinuity was observed in the Arrhenius plot of kcat values at pH 9.0. We also investigated the stability in the presence of guanidinium chloride at 20 degrees C either at pH 7.0 or at pH 9.0, and we present data that indicate that the unfolding mechanism closely approaches a two-state model at pH 7.0 and a more complex mechanism at pH 9.0. Satisfactory fitting of the equilibrium unfolding transition obtained by fluorescence measurements at pH 9.0 required a model that involves a stable intermediate in addition to the native and unfolded forms. At 20 degrees C the folded conformation is more stable than the unfolded conformation by (14. 7+/-1.2) kJ/mol at pH 7.0 and by (25.5+/-1.8) kJ/mol at pH 9.0.

Solution structure of human calcitonin in membrane-mimetic environment: the role of the amphipathic helix.

The 32 amino acid hormone human calcitonin was studied at pH 3.7 and 7.4 by multidimensional NMR spectroscopy in sodium dodecyl sulfate micelles at 310K. The secondary structure was obtained from nuclear Overhauser enhancement spectroscopy (NOESY), 3JHNalpha coupling constants, and slowly exchanging amide data. Three-dimensional structures consistent with NMR data were generated by using distance geometry calculations. A set of 265 interproton distances derived from NOESY experiments, hydrogen-bond constraints obtained from amide exchange, and coupling constants were used. From the initial random conformations, 30 distance geometry structures with minimal violations were selected for further refinement with restrained energy minimization. In micelles, at both pHs, the hormone assumes an amphipathic alpha-helix from Leu9 to Phel6, followed by a type-I beta-turn between residues Phel6 and Phel9. From His20 onward the molecule is extended and no interaction with the helix was observed. The relevance of the amphipathic helix for the structure-activity relationship, the possible mechanisms of interaction with the receptor, as well as the formation of fibrillar aggregates, is discussed.

Characterization of aromatic aminotransferases from the hyperthermophilic archaeon Thermococcus litoralis.

The hyperthermophilic archaeon (formerly archaebacterium) Thermococcus litoralis grows at temperatures up to 98 degrees C using peptides and proteins as the sole sources of carbon and nitrogen. Cell-free extracts of the organism contained two distinct types of aromatic aminotransferases (EC 2.6.1.57) which were separated and purified to electrophoretic homogeneity. Both enzymes are homodimers with subunit masses of approximately 47 kDa and 45 kDa. Using 2-oxoglutarate as the amino acceptor, each catalyzed the pyridoxal-5'-phosphate-dependent transamination of the three aromatic amino acids but showed virtually no activity towards aspartic acid, alanine, valine or isoleucine. From the determination of Km and kcat values using 2-oxoglutarate, phenylalanine, tyrosine and tryptophan as substrates, both enzymes were shown to be highly efficient at transaminating phenylalanine (kcat/Km approximately 400 s-1 mM-1); the 47-kDa enzyme showed more activity towards tyrosine and tryptophan compared to the 45-kDa one. Kinetic analyses indicated a two-step mechanism with a pyridoxamine intermediate. Both enzymes were virtually inactive at 30 degrees C and exhibited maximal activity between 95-100 degrees C. They showed no N-terminal sequence similarity with each other (approximately 30 residues), nor with the complete amino acid sequences of aromatic aminotransferases from Escherichia coli and rat liver. The catalytic properties of the two enzymes are distinct from bacterial aminotransferases, which have broad substrate specificities, but are analogous to two aromatic aminotransferases which play a biosynthetic role in a methanogenic archaeon. In contrast, it is proposed that one or both play a catabolic role in proteolytic T. litoralis in which they generate glutamate and an arylpyruvate. These serve as substrates for glutamate dehydrogenase and indolepyruvate ferredoxin oxidoreductase in a novel pathway for the utilization of aromatic amino acids.