Effects of baculovirus transactivators IE-1 and IE-2 on the Drosophila heat shock 70 promoter in two insect cell lines.

Baculovirus infection of permissive cells proceeds in a cascade fashion with the transcription of early, late and very late genes. The structure of a number of baculovirus early gene promoters has been dissected in detail and the viral factors necessary to stimulate their expression have been identified. Early baculovirus gene promoters in general have a resemblance to host promoters while late and very late gene promoters are different from early baculovirus promoters and are more defined. In this study we investigated whether two key Autographa californica M nucleopolyhedrovirus (AcMNPV) transactivators have the ability to regulate the commonly used cellular promoter from the Drosophila heat shock 70 protein gene, during transient gene expression assays in two insect cell lines permissive for AcMNPV infection, SF-21 and TN-368, or during viral infection. The AcMNPV ie-1 transactivator gene stimulated gene expression of this cellular promoter in both cell lines when the promoter was cis-linked to an enhancer element, but stimulation in the absence of enhancer elements was either undetected or lower than in the presence of enhancer elements in SF-21 and TN-368 cells, respectively. The transactivator ie-2 stimulated gene expression in the presence of cis-linked enhancer elements and ie-1 in SF-21 cells. During viral infection, the heat shock 70 promoter was maximally activated at 12 hours post infection. We discuss how these results affect the interpretation of transient gene expression assays performed in the presence of viral transcription factors.

Meta-analysis of rat lung tumors from lifetime inhalation of diesel exhaust.

Estimating the carcinogenic potential of exposure to diesel-engine exhaust particulates (DEPs) is problematic. In rats, high concentrations of DEPs (> 1,000 microg/m(3)) inhaled over a lifetime result in excess lung tumors. However, data for rats exposed to DEP at concentrations not associated with lung overload are consistent with no tumorigenic effect. Individual rat studies have only a limited number of exposure groups; therefore, we combined the tumor data from eight chronic inhalation studies in a meta-analysis. Statistical analysis identified a threshold of response between 200 and 600 microg/m(3) average continuous lifetime exposure, consistent with biological-effect thresholds reported by other investigators. Our exposure-response analysis of all rats with < 600 microg/m(3) average continuous lifetime exposure found no tumorigenic effect of DEP in these rats. When we evaluated all rat studies, accounted for a threshold and for inhomogeneity between experiments, and expressed the results in terms of human unit risk (UR), we found a negative maximum-likelihood human UR of -32 (times) 10(-6) per microgram per cubic meter (microg/m(3)), but this was not statistically significantly different from zero. Extrapolating the rat upper 95th percentile confidence limit to humans gave an upper-bound human UR of 9.3 (times) 10(-6) per microg/m(3)]. This upper-bound human UR, derived from all of the data points (including 1,087 animals below the estimated threshold and 1,433 in the control groups), falls entirely below the range of estimates derived from lung-overloaded rats or from epidemiology of railroad workers. Our meta-analysis of the low-exposure data in rats does not support a lung cancer risk for DEP exposure at nonoverload conditions. Average ambient concentrations of DEP (0-3 microg/m(3)) are < 1% of the concentration associated here with a threshold of tumor response in the rat bioassay.

Lognormal distributions for body weight as a function of age for males and females in the United States, 1976-1980.

Based on results reported from the NHANES II Survey (the National Health and Nutrition Examination Survey II) for people living in the United States during 1976-1980, we use exploratory data analysis, probability plots, and the method of maximum likelihood to fit lognormal distributions to percentiles of body weight for males and females as a function of age from 6 months through 74 years. The results are immediately useful in probabilistic (and deterministic) risk assessments.

A meta-analysis of the relation between cumulative exposure to asbestos and relative risk of lung cancer.

OBJECTIVES: To obtain summary measures of the relation between cumulative exposure to asbestos and relative risk of lung cancer from published studies of exposed cohorts, and to explore the sources of heterogeneity in the dose-response coefficient with data available in these publications. METHODS: 15 cohorts in which the dose-response relation between cumulative exposure to asbestos and relative risk of lung cancer has been reported were identified. Linear dose-response models were applied, with intercepts either specific to the cohort or constrained by a random effects model; and with slopes specific to the cohort, constrained to be identical between cohorts (fixed effect), or constrained by a random effects model. Maximum likelihood techniques were used for the fitting procedures and to investigate sources of heterogeneity in the cohort specific dose-response relations. RESULTS: Estimates of the study specific dose-response coefficient (kappa 1.i) ranged from zero to 42 x 10(-3) ml/fibre-year (ml/f-y). Under the fixed effect model, a maximum likelihood estimate of the summary measure of the coefficient (k1) equal to 0.42 x 10(-3) (95% confidence interval (95% CI) 0.22 to 0.69 x 10(-3)) ml/f-y was obtained. Under the random effects model, implemented because there was substantial heterogeneity in the estimates of kappa 1.i and the zero dose intercepts (Ai), a maximum likelihood estimate of k1 equal to 2.6 x 10(-3) (95% CI 0.65 to 7.4 x 10(-3)) ml/f-y, and a maximum likelihood estimate of A equal to 1.36 (95% CI 1.05 to 1.76) were found. Industry category, dose measurements, tobacco habits, and standardisation procedures were identified as sources of heterogeneity. CONCLUSIONS: The appropriate summary measure of the relation between cumulative exposure to asbestos and relative risk of lung cancer depends on the context in which the measure will be applied and the prior beliefs of those applying the measure. In most situations, the summary measure of effect obtained under the random effects model is recommended. Under this model, potency, k1, is fourfold lower than that calculated by the United States Occupational Safety and Health Administration.

An accessory protein of DNA polymerase alpha declines in function with increasing age.

Isoforms of DNA polymerase alpha (pol alpha/primase; pol alpha) were isolated from the livers of C57BL/6 mice either 3 months old (young) or 13 months old (mature). The 13-month-old mice were from two groups, one in which food was available ad libitum (AL), and one in which calories had been restricted to 60% of the AL intake (CR). The polymerases from young vs. mature and CR vs. AL mice differed in total and specific pol alpha activity, with the highest values exhibited by enzymes from 3-month-old mice. A more active isoform of pol alpha was typically isolated from CR animals than from AL animals. Differences in charge were used to chromatographically separate pol alpha into elution peaks exhibiting differing degrees of enzyme activity. DNA pol alpha isolated from tissues of mature mice exhibited a decline in activity which was not associated with decreased recoverable levels of pol alpha protein, but with a decline in the tendency of pol alpha to co-purify with an accessory protein, alpha AP, that binds double-stranded DNA (dsDNA). Low activity pol alpha isoforms which did not co-purify with alpha AP were stimulated by interaction with exogenous alpha AP. Pol alpha isoforms which co-purified with the dsDNA-binding accessory protein exhibited higher specific activity and less enhancement of activity upon interaction with exogenous alpha AP. Calorie restricted animals exhibited a pol alpha isoform that was more like pol alpha from younger animals in that it typically copurified with alpha AP, the DNA-binding accessory protein.

Monte Carlo techniques for quantitative uncertainty analysis in public health risk assessments.

Most public health risk assessments assume and combine a series of average, conservative, and worst-case values to derive a conservative point estimate of risk. This procedure has major limitations. This paper demonstrates a new methodology for extended uncertainty analyses in public health risk assessments using Monte Carlo techniques. The extended method begins as do some conventional methods--with the preparation of a spreadsheet to estimate exposure and risk. This method, however, continues by modeling key inputs as random variables described by probability density functions (PDFs). Overall, the technique provides a quantitative way to estimate the probability distributions for exposure and health risks within the validity of the model used. As an example, this paper presents a simplified case study for children playing in soils contaminated with benzene and benzo(a)pyrene (BaP).

One-hit models of carcinogenesis: conservative or not?

One-hit formulas are widely believed to be "conservative" when used to analyze carcinogenesis bioassays, in the sense that they will rarely underestimate risks of cancer at low exposures. Such formulas are generally applied to the lifetime incidence of cancer at a specific site, with risks estimated from animal data at zero dose (control), and two or more additional doses that are appreciable fractions of a maximum tolerated dose. No empirical study has demonstrated that the one-hit formula is conservative in the sense described. The Carcinogenesis Bioassay Database System contains data on 1212 separate bioassays of 308 chemical substances tested at exactly three evaluable doses. These provided sufficient data to examine 8432 specific combinations of cancer site with sex, species, and chemical. For each of these we fitted a one-hit formula to the zero and maximum dose data points, then examined the relation of the fitted curve to the incidence rate observed at the mid-dose, with and without adjustment for intercurrent mortality. Both underestimates and overestimates of risk at mid-dose occurred substantially more often than expected by chance. We cannot tell whether such underestimates would occur at lower doses, but offer six biological reasons why underestimates might be expected. In a high percentage of animal bioassays, the one-hit formula is not conservative when applied in the usual way to animal data. It remains possible that the one-hit formula may indeed be conservative at sufficiently low doses (below the observational range), but the usual procedure, applied to the usual dose range, can be nonconservative in estimating the slope of the formula at such low doses. Risk assessments for regulation of carcinogens should incorporate some measure of additional uncertainty.

Dose-response relationships for carcinogens: a review.

We review the experimental evidence for various shapes of dose-response relationships for carcinogens and summarize those experiments that give the most information on relatively low doses. A brief review of some models is given to illustrate the shapes of dose-response curve expected from them. Our major interest is in the use of dose-response relationships to estimate risks to humans at low doses, and so we pay special attention to experimentally observed and theoretically expected nonlinearities. There are few experimental examples of nonlinear dose-response relations in humans, but this may simply be due to the limitations in the data. The several examples in rodents, even though for high dose data, suggest that nonlinearity is common. In some cases such nonlinearities may be rationalized on the basis of the pharmacokinetics of the test compound or its metabolites.

Risk assessment and comparisons: an introduction.

Risk assessment is presented as a way of examining risks so that they may be better avoided, reduced, or otherwise managed. Risk implies uncertainty, so that risk assessment is largely concerned with uncertainty and hence with a concept of probability that is hard to grasp. The results of even the simplest risk assessments need to be compared with similar assessments of commonplace situations to give them some meaning. We compare and contrast some risk estimates to display their similarities and differences.

Uncertainties in interspecies extrapolations of carcinogenicity.

The usual classification of results of animal carcinogenicity tests is positive or negative. Attempting to observe correlations between species using such results is complicated by differing test sensitivities. In these circumstances it is helpful to use models to represent the experimental data in a consistent way. Fitting the model parameters to the data allows computation of confidence limits and an assessment of concordance or discordance between different species in a way which accounts for differing test sensitivities. This paper describes this approach in detail for one class of models applied to the carcinogenicity test results of 187 of the NCI bioassay series, allowing comparison between B6C3F1 mice, Fischer 344 rats and Osborne Mendel rats. It is shown that the uncertainties in extrapolating between species are larger than generally acknowledged (a standard deviation of a factor of approximately 4.5), but that within these uncertainties there are few if any discordances.