Quantitative analysis of constitutive and 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced cytochrome P450 1B1 expression in human lymphocytes.

Exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD or dioxin) results in a broad spectrum of biological responses, including altered metabolism, disruption of normal hormone signaling pathways, reproductive and developmental effects, and cancer. Cytochrome P450 1B1 (CYP1B1) is a dioxin-inducible gene that is active in the formation of 4-hydroxyestradiol, a potentially genotoxic catechol estrogen. Therefore, the analysis of CYP1B1 in humans may be useful in establishing relationships between dioxin exposure and adverse health effects. In this study, we examined the expression of CYP1B1 in human peripheral blood lymphocytes of unexposed individuals using a quantitative reverse transcription-PCR method. Absolute CYP1B1 RNA levels varied more than 30-fold in uncultured mononuclear cells obtained from 10 individuals. In vitro treatment of mitogen-stimulated lymphocytes with TCDD for 1-5 days of culture resulted in a peak induction of CYP1B1 after 3 days. The induction of CYP1B1 RNA levels after 3 days of culture was dose-dependent, exhibited a maximum response above 10 nM TCDD, and varied greatly among different individuals. However, the half maximal dose required for this induction was similar between individuals and comparable to that observed in the MCF-7 and HepG2 human cell lines. These observations indicate that CYP1B1 exhibits variable constitutive expression and is inducible in vitro by TCDD in human lymphocytes and that the magnitude of induction varies within the population. These data define the suitability of CYP1B1 for use as a mechanistically based biomarker in ongoing molecular epidemiological studies of human populations exposed to dioxins and related chemicals that bind the aromatic hydrocarbon receptor.

Animal models of human response to dioxins.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is the most potent member of a class of chlorinated hydrocarbons that interact with the aryl hydrocarbon receptor (AhR). TCDD and dioxinlike compounds are environmentally and biologically stable and as a result, human exposure is chronic and widespread. Studies of highly exposed human populations show that dioxins produce developmental effects, chloracne, and an increase in all cancers and suggest that they may also alter immune and endocrine function. In contrast, the health effects of low-level environmental exposure have not been established. Experimental animal models can enhance the understanding of the effects of low-level dioxin exposure, particularly when there is evidence that humans respond similarly to the animal models. Although there are species differences in pharmacokinetics, experimental animal models demonstrate AhR-dependent health effects that are similar to those found in exposed human populations. Comparisons of biochemical changes show that humans and animal models have similar degrees of sensitivity to dioxin-induced effects. The information gained from animal models is important for developing mechanistic models of dioxin toxicity and critical for assessing the risks to human populations under different circumstances of exposure.

Obtaining information about susceptibility from the epidemiological literature.

Whether people become ill after encountering environmental pollutants depends on the magnitude of their exposure and their capacity to respond. Exposure and intrinsic response capabilities vary within the population. Those that become ill when the general population remains largely unaffected are considered to be highly susceptible. The U.S. Environmental Protection Agency (USEPA), responsible for protecting the public from environmental pollutants, has developed risk assessment procedures to assist in evaluating the likelihood of health effects. However, the Agency's ability to evaluate the risk faced by highly susceptible populations is often hindered by the paucity of adequate health effects data. Response variability can be assessed with animal models and human epidemiological studies. Although animal models are useful when evaluating the effect of gender and developmental stage on susceptibility, inbred rodent strains underestimate the genetic and lifestyle-induced variability in susceptibility found in human populations. Epidemiological approaches are the preferred source of information on variability. This paper reviews the epidemiological literature from the perspective of a risk assessor seeking data suitable for estimating the risk to highly susceptible populations. Epidemiological approaches do not measure the full range of population response variability. Rather, "susceptibility factors" are evaluated either as risk factors or by focusing on the susceptible population, e.g. children. Susceptibility factors due to genetics, developmental stage, gender, ethnicity, disease state and lifestyle are most frequently encountered. Often, the information describing the health impact of the susceptibility factor is incomplete due to, (1) a failure to consider factors modifying susceptibility; (2) inadequate exposure data; (3) a failure to evaluate the health impact of the susceptibility factor. In addition, for a given exposure agent, several susceptibility factors may be relevant. While incomplete data describing susceptibility factors limits the opportunity for quantitative estimations of risk, available information can supplement qualitative evaluations and risk management.

Immunochemical techniques in biological monitoring.

Immunoassays are analytical methods that detect interactions between antibodies and antigens. Immunoassays were used originally to detect large biological molecules. The new generation of these antibody-based assays can detect small synthetic compounds. As a result, immunoassays are being developed specifically for biomarkers of exposure and effect to environmentally prevalent chemicals. Immunochemical detection of parent compounds in blood and tissues, metabolites in excreta, and adducts with DNA and protein have been successfully performed by several investigators. Although there is great potential for use of immunoassays in biological monitoring studies, the limitations of these analyses must be fully understood to prevent improper evaluation of the acquired data. This review will cover some of the background material necessary to understand how an antibody-based assay is developed. The differences between polyclonal and monoclonal antibody-based assays and the importance of antibody class, affinity, specificity, and cross-reactivity must be considered in both study design and data analysis.