Modulation of steroidogenesis by coastal waters and sewage effluents of Hong Kong, China, using the H295R assay

BACKGROUND, AIM, AND SCOPE: The presence of a variety of pollutants in the aquatic environment that can potentially interfere with the production of sex steroid hormones in wildlife and humans has been of increasing concern. The aim of the present study was to investigate the effects of extracts from Hong Kong marine waters, and influents and effluents from wastewater treatment plants on steroidogenesis using the H295R cell bioassay. After exposing H295R cells to extracts of water, the expression of four steroidogenic genes and the production of three steroid hormones were measured. MATERIALS AND METHODS: Water samples were collected during the summer of 2005 from 24 coastal marine areas and from the influents and effluents of two major waste water treatment plants (WWTPs) in Hong Kong, China. Samples were extracted by solid phase extraction (SPE). H295R cells were exposed for 48 h to dilutions of these extracts. Modulations of the expression of the steroidogenic genes CYP19, CYP17, 3betaHSD2, and CYP11beta2 were determined by measuring mRNA concentrations by real-time polymerase chain reaction (Q-RT-PCR). Production of the hormones progesterone (P), estradiol (E2), and testosterone (T) was quantified using enzyme linked immunosorbent assays (ELISA). RESULTS: Extracts from samples collected in two fish culture areas inhibited growth and proliferation of H295R cells at concentrations greater or equal to 10(5) L equivalents. The cells were exposed to the equivalent concentration of active substances in 10,000 L of water. Thus, to observe the same level of effect as observed in vitro on aquatic organisms would require a bioaccumulation factor of this same magnitude. None of the other 22 marine samples affected growth of the cells at any dilution tested. Twelve of the marine water samples completely inhibited the expression of CYP19 without affecting E2 production; inhibition of CYP17 expression was observed only in one of the samples while expression of CYP11beta2 was induced as much as five- and ninefold after exposure of cells to extracts from two locations. The expression of the progesterone gene 3betaHSD2 was not affected by any of the samples; only one sample induced approximately fourfold the production of E2. Although more than twofold inductions were observed for P and T production, none of these values were statistically significant to conclude effects on the production of these two hormones. While influents from WWTPs did not affect gene expression, an approximately 30% inhibition in the production of E2 and a 40% increase in P occurred for the exposure with influents from the Sha Tin and Stonecutters WWTPs, respectively. Effluents from WWTPs did not affect the production of any of the studied hormones, but a decrement in the expression of the aldosterone gene CYP11beta2 was observed for the Sha Tin WWTP exposure. No direct correlation could be established between gene expression and hormone production. DISCUSSION: Observed cytotoxicity in the two samples from fish culture areas suggest the presence of toxic compounds; chemical analysis is required for their full identification. Although effluents from WWTPs did not affect hormone production, other types of endocrine activity such as receptor-mediated effects cannot be ruled out. Interactions due to the complexity of the samples and alternative steroidogenic pathways might explain the lack of correlation between gene expression and hormone production results. CONCLUSIONS: Changes observed in gene expression and hormone production suggest the presence in Hong Kong coastal waters of pollutants with endocrine disruption potential and others of significant toxic effects. The aromatase and aldosterone genes seem to be the most affected by the exposures, while E2 and P are the hormones with more significant changes observed. Results also suggest effectiveness in the removing of compounds with endocrine activity by the WWTPs studied, as effluent samples did not significantly affect hormone production. The H295R cell showed to be a valuable toll in the battery required for the analysis of endocrine disrupting activities of complex environmental samples. RECOMMENDATIONS AND PERSPECTIVES: Due to the intrinsic complexity of environmental samples, a combination of analytical tools is required to realistically assess environmental conditions, especially in aquatic systems. In the evaluation of endocrine disrupting activities, the H295R cell bioassay should be used in combination with other genomic, biological, chemical, and hydrological tests to establish viable modes for endocrine disruption and identify compounds responsible for the observed effects.     

Effects of tributyltin on the energy metabolism of pen shell (Atrina pectinata japonica)

We examined the effects of tributyltin (TBT) on aerobic and anaerobic energy metabolism of pen shell (Atrina pectinata japonica). We exposed pen shells to TBT at nominal concentrations of 0 (control) and 1.0microg/l for 72h under aerobic condition. At the end of the exposure, half of the pen shells in each treatment were wrapped in plastic wrap to simulate exposure to hypoxia and held at 25 degrees C for another 12h. The concentrations of the products of energy metabolism, namely lactate, pyruvate, fumarate and succinate, in adductor muscle were measured. The exposure to TBT under aerobic condition significantly elevated lactate, pyruvate and fumarate concentrations (p<0.001). After subsequent exposure to anaerobic condition, the mean concentration of succinate in the TBT treatment group was 64% of that in the control group, but there were no significant differences. Our results suggest that the energy metabolism of pen shell is disrupted by exposure to TBT.     

Effects of acetaminophen in <Emphasis Type="Italic">Brassica juncea</Emphasis> L. Czern.: investigation of uptake,translocation, detoxification,and the induced defense pathways

Purpose  

Besides classical organic pollutants and pesticides, pharmaceuticals and their residues have nowadays become recognized as relevant environmental contaminants. The risks of these chemicals for aquatic ecosystems are well known, but information about the pharmaca-plant interactions and metabolic pathways is scarce. Therefore, we investigate the process of uptake of acetaminophen (N-Acetyl-4-aminophenol) by Brassica juncea, drug-induced defense responses and detoxification mechanisms in different plant parts.     

The influence of size-specific indirect interactions in predator-prey systems

Nonlethal indirect interactions between predators often lead to nonadditive effects of predator number on prey survival and growth. Previous studies have focused on systems with at least two different predator species and one prey species. However, most predators undergo extreme ontological changes in phenotype such that interactions between different-sized cohorts of a predator and its prey could lead to nonadditive effects in systems with only two species. This may be important since different-sized individuals of the same species can differ more in their ecology than similar-sized individuals of different species. This study examined trait-mediated indirect effects in a two-species system including a cannibalistic predator with different-sized cohorts and its prey. I tested for these effects using larvae of two stream salamanders, Gyrinophilus porphyriticus (predator) and Eurycea cirrigera (prey), by altering the densities and combinations of predator size classes in experimental streams. Results showed that the presence of large individuals can significantly reduce the impact of density changes of smaller conspecifics on prey survival through nonlethal means. In the absence of large conspecifics, an increase in the relative frequency of small predators significantly increased predation rates, thereby reducing prey survival. However, with large conspecifics present, increasing the density of small predators did not decrease prey survival, resulting in a 14.3% lower prey mortality than predicted from the independent effects of both predator size classes. Small predators changed their microhabitat use in the presence of larger conspecifics. Prey individuals reduced activity in response to large predators but did not respond to small predators. Both predators reduced prey growth. These results demonstrate that the impact of a predator can be significantly altered by two different types of trait-mediated indirect effects in two-species systems: between different-sized cohorts and between different cohorts and prey. This study demonstrates that predictions based on simple numerical changes that assume independent effects of different size classes or ignore size structure can be strongly misleading. We need to account for the size structure within predator populations in order to predict how changes in predator abundance will affect predator-prey dynamics.     

The Effect of Nucleating Particulates on Photochemical Aerosol Formation

The role of nucleating particulates in the formation of photochemical aerosols has been studied in a steady, laminar flow of ultrafiltered air containing NO₂ and octene-1 in the concentration range of (30 to 170 ppm) when subjected to intense irradiation under isothermal conditions. The particulates consisted of monodisperse polystyrene latex (d = 0.36 μ.) in concentrations similar to those in the atmosphere (6 × 10¹ to 3 × 10³ cm^–3); the irradiation intensity varied between (6 to 40 × 10³ lumen/liter) and the mean exposure duration between 30 and 180 sec. Samples of the flow prior to and after its photoactivation were withdrawn either by an Aerosol Spectrometer (AS) or by a Royco Aerosol Photometer (PH). While these indications refer thus to the same system, they differ, because the photometric data include all colloidal components in the airborne state, whereas the counts obtained from the AS deposits refer only to the nucleated latex particles. The following pattern becomes evident: The photochemical reaction yields fractional products (less than three percent) which have the tendency to agglomerate (or polymerize) due to their relatively low volatility—independent of the presence or absence of nucleating particulates. In their presence, this reaction becomes kinetically more probable and thus faster, hence the accumulant formation occurs preferably on the nuclei and causes their growth such that, e.g., a 10-fold higher nuclei concentration will produce under the same conditions 10 times the accumulant mass while autonucleation is suppressed. The growth process appears thus principally different from that of fog formation by H₂O-condensation, whereas for identical super saturation it is inversely proportional to the nuclear concentration. In the absence of nuclei autonucleation, i.e., self-agglomeration, occurs at a much lesser reaction rate and higher photon demand. The growth rate of the nuclei, when present, depends on the concentration of the oxidation catalyst (NO₂), its interaction with the nuclei surface is indicated. Under identical conditions the mass of nuclear accumulant is directly proportional to the concentration of the reactive hydrocarbon, while the growth rate depends on the light intensity and the exposure duration. The findings indicate that density and nature of particulate matter present in an air mass prior or during photo-activation are—aside from the chemical reactant levels—of major significance in aerosol formation.     

Mercury accumulation in sediment and fish from rivers affected by alluvial gold mining in the Brazilian Madeira River basin,Amazon

The degree and distribution of mercury (Hg) accumulation in sediment and fish from a tributary affected by alluvial small-scale gold mining in the Madeira River/Amazon is studied, in relation to a reference site. The results obtained so far agree well with previous studies and confirm that a tremendous contamination of main food web compartments occurs in these highly exposed, but vulnerable tropical waters. An essential part of the released metallic Hg may still exist as macroscopic liquid Hg drops in the sediment. Both global (0.4 mg/kg of Hg) and local (0.1 mg/kg of Hg) background sediment values as well as safety levels for fish (0.5 mg/kg of Hg) are considerably exceeded by a factor of up to 25, 100, and 4, respectively, and give rise to serious concern, not least with regard to the formation of the very toxic monomethyl-Hg. It is further discussed that atmospheric transport and deposition of Hg in water reservoirs built for hydroelectric power generation may act as critical pathways for longterm Hg accumulation, even in unexposed riverine systems.     

Predicting the mating system from phenotypic correlations between life-history and sperm quality traits in the Alpine whitefish <Emphasis Type="Italic">Coregonus zugensis</Emphasis>

The mating behavior and reproductive strategies of Alpine whitefish like Coregonus zugensis (Nüsslin) are poorly understood, probably because they spawn in deep water where direct observations are difficult. In this study, we interpret life-history and sperm quality traits of fish that we caught from their spawning place. We found that males invest heavily into gonadal tissue (up to 5.6% of their body weight), which is, in comparison to other fish, consistent with external fertilization, distinct pairing and moderate to high communal spawning, or no pairing and low to moderate communal spawning. Sperm competition theory and recent experimental studies on other salmonids predict that males optimize ejaculate characteristics in relation to the costs of sperm and the level of competition they have to expect: dominant males are predicted to invest less into ejaculate quality and to have slower spermatozoa than subdominant males. We found that spermatozoa of older males are slower than those of younger males. Moreover, older males have larger breeding tubercles, a secondary sexual trait that has, in some previous studies, been found to be linked to good condition and to good genetic quality. Our results suggest that C. zugensis has age-linked reproductive strategies, that multimale spawning is common, i.e., that sperm competition plays a significant role, and that older males are on average dominant over younger males at the spawning place.     

Work stress and the stress-buffering roles of work and family resources

This paper describes the development of the Work Environment Scale, which includes measures of perceived stressful and supportive aspects of the job milieu. This scale is used to examine stressful effects of the work environment on personal functioning and the stress-buffering value of work and family social resources among a representative community group of men and women. Work stressors had a greater impact on men, but supportive social resources provided more attenuation of such effects among men than among women. Although work stressors generally had less effect on men whose wives were employed, high stress in the women's work settings had some indirect negative consequences for their husbands. Reprint requests should be directed to the authors, Social Ecology Laboratory, Department of Psychiatry and Behavioral Sciences, Stanford University Medical School, Stanford, California 94305, U.S.A.