The OECD validation program of the H295R steroidogenesis assay: Phase 3. Final inter-laboratory validation study

Background, goals, and scope

In response to increasing concerns regarding the potential of chemicals to interact with the endocrine system of humans and wildlife, various national and international programs have been initiated with the aim to develop new guidelines for the screening and testing of these chemicals in vertebrates. Here, we report on the validation of an in vitro assay, the H295R steroidogenesis assay, to detect chemicals with the potential to inhibit or induce the production of the sex steroid hormones testosterone (T) and 17??-estradiol (E2) in preparation for the development of an Organization for Economic Cooperation and Development (OECD) test guideline.

Methods

A previously optimized and pre-validated protocol was used to assess the potential of 28 chemicals of diverse structures and properties to validate the H295R steroidogenesis assay. These chemicals are comprised of known endocrine-active chemicals and ??negative?? chemicals that were not expected to have effects on the targeted endpoints, as well as a number of test chemicals with unknown modes of action at the level of the steroidogenic pathway. A total of seven laboratories from seven countries participated in this effort. In addition to effects on hormone production, confounding factors, such as cell viability and possible direct interference of test substances with antibody-based hormone detection assays, were assessed. Prior to and during the conduct of exposure experiments, each laboratory had to demonstrate that they were able to conduct the assay within the margin of predefined performance criteria.

Results

With a few exceptions, all laboratories met the key quality performance parameters, and only 2% and 7% of all experiments for T and E2, respectively, were excluded due to exceedance of these parameters. Of the 28 chemicals analyzed, 13 and 14 tested affected production of T and E2, respectively, while 11 and 8 did not result in significant effects on T and E2 production, respectively. Four and six chemicals produced ambiguous results for effects on T and E2 production, respectively. However, four of these cases each for T and E2 were associated with only one laboratory after a personnel change occurred. Significant interference of test chemicals with some of the antibody-based hormone detection systems occurred for four chemicals. Only one of these chemicals, however, significantly affected the ability of the detection system to categorize the chemical as affecting E2 or T production.

Discussion and conclusions

With one exception, the H295R steroidogenesis assay protocol successfully identified the majority of chemicals with known and unknown modes of interaction as inducers or inhibitors of T and E2 production. Thus it can be considered a reliable screen for chemicals that can alter the production of sex steroid hormones. One of the remaining limitations associated with the H295R steroidogenesis assay protocol is the relatively small basal production of E2 and its effect on quantifying the decreased production of this hormone with regard to the identification of weak inhibitors. An initial comparison of the data produced in this study with those from in vivo studies from the literature demonstrated the potential of the H295R steroidogenesis assay to identify chemicals affecting hormone homeostasis in whole organisms. Particularly promising was the lack of any false negatives during the validation and the very low number of false positives (1 out of 28 chemicals for each T and E2).

Perspectives

Based on the results obtained during this validation study and the accordingly revised test protocols, an OECD draft test guideline was developed and submitted to the OECD working group of the national coordinators of the test guidelines program (WNT) for comments in December 2009.     

Atmospheric composition change – global and regional air quality

Air quality transcends all scales with in the atmosphere from the local to the global with handovers and feedbacks at each scale interaction. Air quality has manifold effects on health, ecosystems, heritage and climate. In this review the state of scientific understanding in relation to global and regional air quality is outlined. The review discusses air quality, in terms of emissions, processing and transport of trace gases and aerosols. New insights into the characterization of both natural and anthropogenic emissions are reviewed looking at both natural (e.g. dust and lightning) as well as plant emissions. Trends in anthropogenic emissions both by region and globally are discussed as well as biomass burning emissions. In terms of chemical processing the major air quality elements of ozone, non-methane hydrocarbons, nitrogen oxides and aerosols are covered. A number of topics are presented as a way of integrating the process view into the atmospheric context; these include the atmospheric oxidation efficiency, halogen and HO_x chemistry, nighttime chemistry, tropical chemistry, heat waves, megacities, biomass burning and the regional hot spot of the Mediterranean. New findings with respect to the transport of pollutants across the scales are discussed, in particular the move to quantify the impact of long-range transport on regional air quality. Gaps and research questions that remain intractable are identified. The review concludes with a focus of research and policy questions for the coming decade. In particular, the policy challenges for concerted air quality and climate change policy (co-benefit) are discussed.     

Visitor perceptions of rural landscapes: a case study in the Peak District National Park, England

Maintaining national parks is an integral policy tool to conserve rare habitats. However, because national parks are funded by taxpayers, they must also serve the needs of the general public. Increasingly, and thanks to today's diverse society, there is evidence that this creates challenges for park managers who are pulled in two opposing directions: to conserve nature on the one hand and to meet different visitor expectations on the other. This tension was explored in the Peak District National Park, a rural landscape dominated by heather moorland and sheep farming in Northern England where research was conducted to determine how social class and ethnicity shaped perceptions of the park. Results uncovered that social class played a very strong role in shaping perceptions of this region with 'middle class' respondents reacting far more favourably to the park than people from more working class backgrounds. We observed ethnicity playing a similar role, though our results are less significantly different.     

Behaviour of pendimethalin in tropical and mediterranean plain field condition

Abstract

Behaviour of the herbicide pendimethalin [N‐(l‐ethylpropyl)‐3,4‐dimethyl‐2,6‐dinitrobenzenamine] was evaluated in plain field under wet tropical conditions (Martinique, FWI) and in the Mediterranean area (Languedoc, South of France). Simultaneously, adsorption parameters were discussed with Freundlich equation, and desorption evaluated using a model with two compartments corresponding to two different energy levels. Pendimethalin showed a strong adsorption in all three soils and was very difficult to desorb. Higher clay content in tropical vertisol and ferrisol involved adsorption parameters (Kfa) higher than the one found in the Mediterranean fluvisol. Although heavy rainfalls in the tropics intervene on migration, pendimethalin propensity to leaching is levelled by its strong adsorption potential and appears very limited under the conditions chosen during this work. Half‐life of the compound (in the 0–25 cm soil layer) varied from 13 to 17 days. Pendimethalin was also classified as a “non‐leacher” compound using the “Gustafson model”.     

Persistence Factors for Mobile Source (Roadway) Carbon Monoxide Modeling

A critical step in the modeling of the carbon monoxide (CO) impacts of mobile sources is predicting an 8-hour CO concentration given a modeled "worst-case" 1-hour concentration. Often, this is done by a multiplicative persistence factor. A meteorological persistence factor (MPF) accounts for the variability over 8 hours of wind speed, wind direction, stability class, and temperature. A vehicular persistence factor (VPF) reflects the lower traffic volumes during the off-peak hours.

Hourly meteorological data for ten years for four cities in Florida were obtained from the National Climatic Data Center. The CALINE3 model was used to obtain hourly CO concentrations, which were combined to derive MPFs for each city. Similarly, VPFs were derived from hourly vehicle counts from one busy roadway in each city. The mean VPF multiplied by the second highest MPF was defined as the worst-case total persistence factor (TPF). These worst-case TPFs increased significantly as more hours of nighttime were included in the 8- hour averaging time, but were fairly consistent from city to city. In general, the results suggest worst-case TPFs in the range of 0.4 to 0.5, lower than has been recommended by EPA in the past.     

Hydrogen sulfide flux measurements from construction and demolition debris (C&D) landfills

Hydrogen sulfide (H2S) has been identified as a principal odorous component of gaseous emissions from construction and demolition debris (C&D) landfills. Although several studies have reported the ambient concentrations of H2S near C&D landfills, few studies have quantified emission rates of H2S. One of the most widely used techniques for measuring surface gas emission rates from landfills is the flux chamber method. Flux measurements using the flux chamber were performed at five different C&D landfills from April to August, 2003. The flux rates of H2S measured in this research were between 0.192 and 1.76 mg/(m2-d).     

The pathway of dechlorination of PCB congener by a photochemical chain process in 2-propanol: the role of medium and quenching

As part of a program aimed at developing a field process for cleanup of PCB contaminated soils using photochemistry in basic 2-propanol, additional details of the dechlorination pathway are presented. The mechanism involves a chain reaction with both homolytic photochemical C-Cl bond fission and electron transfer steps producing PCB anion radicals. Kinetics of dechlorination of various congeners show patterns of relative rates associated with the basic 2-propanol medium that are not found in other media because both electron transfer and photochemical homolysis steps determine overall rates of dechlorination and govern the pathways and relative concentrations of intermediates. The electron transfer steps display opposite structure-reactivity correlations to the photo-homolysis, C-Cl bond fission steps. Oxygen quenching is shown to differentially affect both types of steps. In contrast to the suggestion that inter system crossing can be highly efficient with reaction originating from a PCB triplet, oxygen quenching data suggest that a significant minimum of the quantum yield is non-quenchable, presumably because of a reaction path from the PCB singlet. This may help to explain why exclusion of air is not entirely necessary in practice.     

Mitigation assessment of vegetated drainage ditches for collecting irrigation runoff in California

Widespread contamination of California water bodies by the organophosphate insecticides diazinon and chlorpyrifos is well documented. While their usage has decreased over the last few years, a concomitant increase in pyrethroid usage (e.g., permethrin) (replacement insecticides) has occurred. Vegetated agricultural drainage ditches (VADD) have been proposed as a potential economical and environmentally efficient management practice to mitigate the effects of pesticides in irrigation and storm runoff. Three ditches were constructed in Yolo County, California for a field trial. A U-shaped vegetated ditch, a V-shaped vegetated ditch, and a V-shaped unvegetated ditch were each amended for 8 h with a mixture of diazinon, permethrin, and suspended sediment simulating an irrigation runoff event. Water, sediment, and plant samples were collected spatially and temporally and analyzed for diazinon and permethrin concentrations. Pesticide half-lives were similar between ditches and pesticides, ranging from 2.4 to 6.4 h. Differences in half-distances (distance required to reduce initial pesticide concentration by 50%) among pesticides and ditches were present, indicating importance of vegetation in mitigation. Cis-permethrin half-distances in V ditches ranged from 22 m (V-vegetated) to 50 m (V-unvegetated). Half-distances for trans-permethrin were similar, ranging from 21 m (V-vegetated) to 55 m (V-unvegetated). Diazinon half-distances demonstrated the greatest differences (55 m for V-vegetated and 158 m for V-unvegetated). Such economical and environmentally successful management practices will offer farmers, ranchers, and landowners a viable alternative to more conventional (and sometimes expensive) practices.