Alteration of gene expression profiles in the brain of Japanese medaka (Oryzias latipes) exposed to KC-400 or PCB126

Polychlorinated biphenyls (PCBs) are known as neurotoxic chemicals and possibly alter animal behavior. We previously reported that PCB-exposure induced abnormal schooling behavior in Japanese medaka (Oryzias latipes). This abnormal behavior might be caused by the functional alteration of central or terminal nervous system. To understand the mechanism(s) of behavioral change by PCB-exposure, we analyzed the gene expression profiles in the brain of medaka exposed to 3,3',4,4',5-pentachlorobiphenyl (PCB126) or a PCB mixture (Kanechlor-400: KC-400) using a cDNA microarray that we constructed. Twelve FLF-II strain medaka (six individuals per treatment) were dietary exposed to PCB126 (0.01 microg/g b.w./day) or KC-400 (1 microg/g b.w./day) for three weeks. For the control, six fish were fed a control diet. After the exposure period, fish were dissected, and the brain samples were collected. The samples from control fish were pooled and used as a common reference in the microarray experiment. Microarray data were normalized by the LOWESS method, and we screened the genes whose expression levels were altered more than 1.5-fold. Gene expression profiling showed 97 down-regulated and 379 up-regulated genes in the brain of medaka exposed to PCB126. KC-400 exposure suppressed 15 genes and induced 266 genes in medaka brain. Among these genes, the expression levels of 7 and 188 genes were commonly down- or up-regulated, respectively in both treatment groups. On the other hand, 31 gene expressions were significantly different between PCB126 and KC-400 treatment groups, and three out of 31 genes were received opposite effects. In addition, the microarray data showed that thyroid hormone-responsive genes were up-regulated by PCB-exposure, which may imply that PCBs or their metabolites mimic thyroid hormone effects in the brain of PCB-exposed medaka.     

Alteration of gene expression profiles in the brain of Japanese medaka (Oryzias latipes) exposed to KC-400 or PCB126

Polychlorinated biphenyls (PCBs) are known as neurotoxic chemicals and possibly alter animal behavior. We previously reported that PCB-exposure induced abnormal schooling behavior in Japanese medaka (Oryzias latipes). This abnormal behavior might be caused by the functional alteration of central or terminal nervous system. To understand the mechanism(s) of behavioral change by PCB-exposure, we analyzed the gene expression profiles in the brain of medaka exposed to 3,3′,4,4′,5-pentachlorobiphenyl (PCB126) or a PCB mixture (Kanechlor-400: KC-400) using a cDNA microarray that we constructed. Twelve FLF-II strain medaka (six individuals per treatment) were dietary exposed to PCB126 (0.01 μg/g b.w./day) or KC-400 (1 μg/g b.w./day) for three weeks. For the control, six fish were fed a control diet. After the exposure period, fish were dissected, and the brain samples were collected. The samples from control fish were pooled and used as a common reference in the microarray experiment. Microarray data were normalized by the LOWESS method, and we screened the genes whose expression levels were altered more than 1.5-fold. Gene expression profiling showed 97 down-regulated and 379 up-regulated genes in the brain of medaka exposed to PCB126. KC-400 exposure suppressed 15 genes and induced 266 genes in medaka brain. Among these genes, the expression levels of 7 and 188 genes were commonly down- or up-regulated, respectively in both treatment groups. On the other hand, 31 gene expressions were significantly different between PCB126 and KC-400 treatment groups, and three out of 31 genes were received opposite effects. In addition, the microarray data showed that thyroid hormone-responsive genes were up-regulated by PCB-exposure, which may imply that PCBs or their metabolites mimic thyroid hormone effects in the brain of PCB-exposed medaka.     

Effects of polychlorinated biphenyls on liver function and sexual characteristics in Japanese medaka (Oryzias latipes)

The effects of polychlorinated biphenyls (PCBs) on liver function and their differences between sexes were analyzed in Japanese medaka (Oryzias latipes) exposed to PCB126 or Kanechlor-400 (KC-400) using microarray. PCB exposure induced vitellogenin 1 expression in female medaka while suppressing choriogenin genes, which suggests that the effects of PCBs on estrogen-responsive genes do not occur directly through an estrogen receptor-mediated pathway. Reduction of androgen receptor alpha expression was also observed, and the gene expression pattern in PCB-exposed males changed to become more similar to that of females. Furthermore, changes in glycolysis-related genes indicate that PCB exposure might enhance glucose production via gluconeogenesis in the liver of medaka. Taken together, our results suggest that PCBs disrupt the endocrine system, especially androgen function, and may have the potential to cause demasculinizing effects. Additionally, induction of gluconeogenesis might be a response to maintain glucose levels consumed as a result of PCB exposures.     

Effects of polychlorinated biphenyls on liver function and sexual characteristics in Japanese medaka (Oryzias latipes)

The effects of polychlorinated biphenyls (PCBs) on liver function and their differences between sexes were analyzed in Japanese medaka (Oryzias latipes) exposed to PCB126 or Kanechlor-400 (KC-400) using microarray. PCB exposure induced vitellogenin 1 expression in female medaka while suppressing choriogenin genes, which suggests that the effects of PCBs on estrogen-responsive genes do not occur directly through an estrogen receptor-mediated pathway. Reduction of androgen receptor alpha expression was also observed, and the gene expression pattern in PCB-exposed males changed to become more similar to that of females. Furthermore, changes in glycolysis-related genes indicate that PCB exposure might enhance glucose production via gluconeogenesis in the liver of medaka. Taken together, our results suggest that PCBs disrupt the endocrine system, especially androgen function, and may have the potential to cause demasculinizing effects. Additionally, induction of gluconeogenesis might be a response to maintain glucose levels consumed as a result of PCB exposures.     

Distribution and source apportionments of polychlorinated biphenyls (PCBs) in mariculture sediments from the Pearl River Delta,South China

Surface and core sediments collected from six mariculture farms in the Pearl River Delta (PRD) were analyzed to evaluate contamination levels of polychlorinated biphenyls (PCBs). The ∑PCBs (37 congeners) concentrations ranged from 5.10 to 11.0 ng g⁻¹ (mean 7.96 ng g⁻¹) in surface and 3.19 to 22.1 ng g⁻¹ (mean 7.75 ng g⁻¹) in core sediments, respectively. The concentrations were significantly higher than that measured in the sediments of their corresponding reference sites, whereby the average enrichment percentages were 62.0% and 42.7% in surface and core sediments, respectively. Significant correlations (R² = 0.77, p < 0.05) of PCB homologue group proportions between fish feeds and surface mariculture sediments suggested that fish feed input was probably the main source for the enrichment of PCBs. Due to the fact that PCBs could be transferred along food chains, PCB contamination in fish feeds and mariculture sediments should not be overlooked.     

A whole life cycle assessment on effects of waterborne PBDEs on gene expression profile along the brain–pituitary–gonad axis and in the liver of zebrafish

Polybrominated diphenyl ethers (PBDEs) are now found ubiquitously in the aquatic environment and biota, and there is a growing concern that PBDEs may disrupt endocrine systems, leading to reproductive impairments of aquatic animals. In our study, zebrafish (Danio rerio) were exposed to the 5 ng/L, 1 μg/L and 50 μg/L of DE-71 for the duration of the whole life cycle (120 days, from eggs to adults). The expression of selected genes along the brain–pituitary–gonadal (BPG) axis and liver, and the levels of plasma sex hormones were examined.In male fish, up-regulation of GnRH in brain, FSHβ and LHβ in pituitary, FSH-receptor, LH-receptor, and CYP19a in testis was clearly evident, while down-regulation of CYP11a and 3β-HSD was found in testis. In female fish, a 2.4-fold up-regulation of 3β-HSD was found in ovary upon exposure to 50 μg/L of DE-71. GnRH in brain, FSHβ and LHβ in pituitary were also up-regulated, while ERβ, TH and TPH in brain and GnRH-receptor in pituitary were significantly down-regulated. Hepatic ERα, AR and VTG in males were all down-regulated, while hepatic ERα and AR in female were up-regulated.Serum estradiol (E₂) was reduced in both male and female upon exposure to DE-71, while significant increases in serum testosterone (T) and 11-keto-testosterone (11-KT) were only found in male but not female fish. The ratio of T/E₂ as well as the ratio of 11-KT/E₂ in male fish increased in a dose-dependent manner upon exposure to DE-71. Our overall results showed that whole life exposure of DE-71 altered the expression of regulatory genes and receptors at all three levels of the BPG axis in zebrafish, and the responses are sex dependent. The observed disruption of GnRH and GtHs can be further related to the subsequent disruption in both levels and balance sex steroid hormones.     

Maternal-to-fetal transfer and concentration profiles of PCB congeners for Steller sea lions (Eumetopias jubatus) from Hokkaido,Japan

The concentrations of PCB congeners in the blubber and liver of mother Steller sea lions (Eumetopias jubatus; SSLs) and their fetuses from the coast of Hokkaido, Japan in 2008, 2010 and 2012 were analyzed by HRGC–HRMS, in order to elucidate PCB congener profiles and maternal-to-fetal transfer of PCBs in SSLs. ΣPCBs in the fetuses were 1400 ± 660 (the mean ± SD) ng/g-fat in the blubber and 570 ± 320 ng/g-fat in the liver, respectively. There was a concern that SSLs had been contaminated by PCBs during the fetal period. The concentrations of the major congeners in the blubber and liver were a correlation between the fetus and mother (blubber: r = 0.9934, liver: r = 0.9160; P ≦ 0.05). The results indicated that PCBs in the fetuses came from the mothers. #177 and #199 showed no correlations between the fetus and the mother in the blubber and liver. This indicated a selective capture by some natural protector such as the placenta.     

Toxicogenomic analysis of immune system-related genes in Japanese flounder (Paralichthys olivaceus) exposed to heavy oil

Heavy oil contamination is one of the most important environmental issues. Toxicities of polycyclic aromatic hydrocarbons (PAHs), including immune toxicities, are well characterized, however, the immune toxic effects of heavy oil, as a complex mixture of PAHs, have not been investigated. In the present study, we selected Japanese flounder (Paralichthys olivaceus) as a model organism, and observed alteration of immune function by the exposure to heavy oil. To analyze the expression profiles of immune system-related genes, we selected 309 cDNAs from our flounder EST library, and spotted them on a glass slide. Using this cDNA array, alteration of gene expression profiles was analyzed in the kidneys of flounders exposed to heavy oil. Six Japanese flounders (mean body weight: 197 g) were acclimated to laboratory conditions at 19–20 °C. Three fish were exposed to heavy oil C (bunker C) at a concentration of 3.8 g/L for 3 days, and the others were kept in seawater without heavy oil and used as the control. After the exposure period, the fish were transferred into control seawater and maintained for 4 days, and then they were dissected and their kidneys were removed. Total RNA was extracted from the kidney samples to use in gene expression analyses. The microarray detected alteration of immune system-related genes in the kidneys of heavy oil-exposed flounders, including down-regulation of immunoglobulin light chain, CD45, major histocompatibility complex class II antigens and macrophage colony-stimulating factor precursor, and up-regulation of interleukin-8 and lysozyme. These results suggest that pathogen resistance may be weakened in heavy oil-exposed fish, causing a subsequent bacterial infection, and then proinflammatory genes may be induced as a defensive response against the infection. Additionally, we found candidate genes for use as biomarkers of heavy oil exposure, such as N-myc downstream regulated gene 1 and heat shock cognate 71 kDa proteins.     

Organic pollutants (PAHs,PCBs) in sediments from the Mar Piccolo in Taranto (Ionian Sea,Southern Italy)

Polychlorinated biphenyls (PCBs) and 17 parent polycyclic aromatic hydrocarbons (PAHs) were determined in surface sediments from nine stations in the Mar Piccolo of Taranto (Ionian Sea, Southern Italy). Total PAH concentrations ranged from 380 to 12,750 μg/kg d.w., while total PCB levels ranged from 2 to 1684 μg/kg d.w.; this values were higher than those found in others marine coastal areas of the Mediterranean Sea. For PAHs, low molecular weight/high molecular weight, phenanthrene/anthracene and fluoranthene/pyrene ratio were used for discriminating between pyrolitic and petroleum origin. Results showed that PAHs were mainly of pyrolitic origin. PCB and PAH levels in sediments were compared with Sediments Quality Guidelines (ERM–ERL, TEL–PEL indexes) for evaluation probable toxic effects on marine organism. Finally, ERM and PEL quotients were used to evaluate the degree to which chemicals exceed guidelines. Results suggest an ecotoxicological risk for benthic organisms mainly in the first inlet, where high concentrations of PCBs were found in sediments influenced by harbour activities.     

Effect of mercury and cadmium on early life stages of Java medaka (Oryzias javanicus): A potential tropical test fish

Several organisms have been used as indicators, bio-monitoring agents or test organisms in ecotoxicological studies. A close relative of the well established Japanese medaka, the Java medaka (Oryzias javanicus), has the potential to be a test organism. The fish is native to the estuaries of the Malaysian Peninsula, Thailand, Indonesia and Singapore. In this study, newly fertilised eggs were exposed to different concentrations of Cd and Hg. Observations were done on the development of the embryos. Exposure to low levels of Cd and Hg (0.01–0.05 ppm) resulted in several developmental disorders that led to death. Exposure to ⩾1.0 ppm Cd resulted in immediate developmental arrest. The embryos of Java medaka showed tolerance to a certain extent when exposed to ⩾1.0 ppm Hg compared to Cd. Based on the sensitivity of the embryos, Java medaka is a suitable test organism for ecotoxicology in the tropical region.     

Distributions of polychlorinated biphenyls in the Daliao River estuary of Liaodong Bay,Bohai Sea (China)

The distributions of 41 polychlorinated biphenyls (PCBs) were determined in the aqueous phase, suspended particulate matter (SPM), and sediment of the Daliao River estuary in Liaodong Bay, Bohai Sea (China). The total PCB concentrations ranged from 5.51 to 40.28 ng L⁻¹ in the surface water, from 6.78 to 66.55 ng L⁻¹ dry weight in the SPM, and from 0.83 to 7.29 ng g⁻¹ dry weight in the sediment. The PCB concentrations in water, SPM, and sediment were moderate relative to those reported for other estuary and marine systems around the world. Sedimentary PCB concentrations decreased offshore due to the active deposition of laterally transported river-borne particles. The predominance of the highly chlorinated congeners for the water, SPM, and sediment samples are an indication of either a lack of degradation or the presence of nearby or recent releases into the environment.     

Regulation of iodothyronine deiodinases and sodium iodide symporter mRNA expression by perchlorate in larvae and adult Chinese rare minnow (Gobiocypris rarus)

Perchlorate is a widespread contaminant in the aquatic environment. In the present work, the expressions of deiodinase enzymes (d1, d2, and d3) and sodium iodide symporter (nis) genes were determined after larval and adult rare minnow (Gobiocypris rarus) exposed to 5 and 50 μg/L perchlorate for 21 days. The results showed that deflation of swim bladder development was observed in larvae at 50 μg/L perchlorate treatment. An up-regulation of the d2 and nis mRNA levels were observed in the larve and in brain of adults. Meanwhile the expressions of d3 mRNA levels were significantly down-regulated in the liver. These results indicate the changes in d2, nis, and d3 mRNA expression brings about increased outer-ring deiodination, idodine uptake, and a further decrease of inner-ring deiodination, respectively reflecting auto-regulation of hypothalamic-pituitary-thyroid (HPT) axis in adult after perchlorate exposure. The larval fish development could be affected by perchlorate at environmentally relevant concentrations.     

Monitoring of organic micropollutants in Ghana by combination of pellet watch with sediment analysis: E-waste as a source of PCBs

Plastic resin pellets collected at 11 beaches covering the whole Ghanaian coastline were analyzed for polychlorinated biphenyls (PCBs). PCB concentrations (∑13 congeners) were higher in Accra, capital city, and Tema (39–69 ng/g-pellets) than those in rural coastal towns (1–15 ng/g-pellets) which are close to global background, indicating local inputs of PCBs. River sediments were also analyzed for PCBs together with molecular markers. Sedimentary PCBs concentrations were highest at a site (AR02) downstream of an electronic waste (e-waste) scrapyard. At the site (AR02), concentration of linear alkylbenzenes (LABs), a marker of municipal wastewater, was lower than another site (AR03) which is located at the downstream of downtown Accra. This result suggests that PCBs are introduced more to the river from the e-waste site than from activities in downtown Accra. PAHs concentrations were relatively higher in urban areas with strong petrogenic signature. Abundance of triphenylbenzenes suggested plastic combustion near e-waste scrapyard.     

Metabolites in bile of fish from São Sebastião Channel, São Paulo, Brazil as biomarkers of exposure to petrogenic polycyclic aromatic compounds

This study reports the analysis of polycyclic aromatic compound (PAC) metabolites, as biomarkers of exposure to PACs in marine environment. PAC metabolites were measured in bile samples from 14 species of demersal fish caught in the São Sebastião Channel (SSC), SE Brazilian coastline. Naphthalene (NPH) equivalents, phenanthrene (PHN) equivalents, and benzo[a]pyrene (BaP) equivalents were quantified using a reverse-phase high-performance liquid chromatography coupled with fluorescence detection method. For all samples, the means (±standard deviation, n = 37) of concentrations obtained for NPH, PHN, and BaP equivalents were, respectively, 290,000 ± 200,000 ng/g, 18,000 ± 14,000 ng/g, and 970 ± 1900 ng/g. These results indicate recent exposure of these fish to PACs in their environment. In addition, two species (Cyclichthys spinosus and Prionotus nudigula) of fish were analyzed in order to investigate local sources of PAC contamination in the SSC and the influence of the petroleum terminal in fish caught in remote areas. The results showed that these fish species potentially migrate along the channel, especially P. nudigula. Correlations among groups of PAC metabolites indicate the same petrogenic source for NPH and PHN equivalents and a combustion source (e.g., automobile, ships) for BaP equivalents. The ratio BaP/PHN equivalents (0.05 ± 0.07, n = 37) confirms the predominance of petrogenic PACs for contamination by these chemicals in this region.     

Occurrence and distribution of organochlorine residues in surface sediments of the Candarli Gulf (Eastern Aegean)

The residual levels of organochlorine pesticides (OCPs) and polychlorinated biphenyls (PCBs) were determined in surface sediments collected from Candarli Gulf. Total concentrations of OCPs and PCBs in sediments ranged from 10.2 to 57.3 and 2.8 to 205 ng g⁻¹ dwt, respectively. DDTs in sediments were derived from the aged and weathered agricultural soils in 61% of the sampling stations while 39% of the sites originated from the recent DDT inputs. Their concentrations appeared to be relatively low by global standards and only sediments receiving the impact from the Bakircay River and petrochemical industry approached the sediment quality guidelines for PCBs and DDTs. Based on ERL/ERM guidelines, DDT and PCBs posed ecological risk to the bottom-dwelling consumers.     

Atmospheric polychlorinated biphenyl (pcb) inputs to a coastal city near the marmara sea

Recent studies have shown that polychlorinated biphenyl (PCB) levels are high in coastal sites. Various types of deposition events, including dry deposition, air–water exchange and wet deposition, were analyzed as part of the study to illustrate the pollution level. The atmospheric levels and deposition mechanisms of 82 PCBs were investigated in Mudanya, a coastal city in Turkey. A total of 175 samples and 112 blanks were collected. Air–water exchange and particle phase dry deposition fluxes were 120 ± 90 ng/m²-d and 125 ± 160 ng/m²-d, respectively. The wet-dry deposition sampler (WDDS) consisted of two reservoirs, wet and dry. Collected with the WDDS, the dry deposition flux was 21 ± 20 ng/m²-d, and the wet deposition fluxes (during the rainy period) were 145 ± 130 ng/m²-d and 195 ± 270 ng/m²-d for the dissolved and particulate phases, respectively. Mass transfer coefficients (MTCs), dry deposition velocities and washout ratios were calculated and compared with the literature values.     

Does heavy oil pollution induce bacterial diseases in Japanese flounder Paralichthys olivaceus?

As basic research for the effect of heavy oil on the fish immune system, in this study, the number of leukocyte was counted in Japanese flounder Paralichthys olivaceus, after exposure to heavy oil at a concentration of 30 g/8 L for 3 days. To compare the numbers of bacteria in the skin mucus between oil-exposed and control fish, viable bacteria were enumerated by counting colony forming unit (CFU). Compared with 5.79 ± 1.88 × 10⁷ leukocytes/mL in the controls, the exposed fish demonstrated higher counts, averaging 1.45 ± 0.45 × 10⁸ cells/mL. The bacterial numbers of control fish were 4.27 ± 3.68 × 10⁴ CFU/g, whereas they were 4.58 ± 1.63 × 10⁵ CFU/g in the exposed fish. The results suggest that immune suppression of the fish occurred due to heavy oil stressor, and bacteria could invade in the mucus, resulting in the increasing leukocyte number to prevent infectious disease.     

Assessment of risk of PCDD/Fs and dioxin-like PCBs in marine and freshwater fish in Pearl River Delta,China

Fish consumption is known to be beneficial to human health. However since the age of industrialization, the released/disposed chemical pollutants into water systems make fish a source of various environmental toxicants to humans. In oceanic cities with heavy industrial activities, fish products contribute the greatest proportion of exposure to pollutants. In this study, risks and potential effects of dioxins to health of coastal populations in the Pearl River Delta were assessed. Concentrations of polychlorinated dibenzo-p-dioxins (PCDDs), polychlorinated dibenzofurans (PCDFs), and dioxin-like polychlorinated biphenyls (PCBs) were measured in common fish species purchased at local markets. Concentrations of total dioxins in fish ranged from 0.481 to 9.05 pg TEQ/g wet weight were similar to the lesser concentrations reported for fish from other countries. The greatest concentrations of dioxins were measured in mandarin fish, a carnivorous freshwater fish. Exposure of murine primary leydig and ovarian cells to 2,3,7,8-tetrachlorinated dibenzo-p-dioxins (2,3,7,8-TCDD) reduced the synthesis of progesterone, testosterone and/or estrogen. The reductions were probably via inhibitory effects on the expression of the steroidogenic enzymes, steroidogenic acute regulatory protein (StAR) and cytochrome P450 side-chain cleavage enzyme (P450scc). Based on these reproductive parameters, the concentrations of dioxins and dioxin-like residues represent a moderate health risk due to consumption of fish.     

Characteristics of ChgH–GFP transgenic medaka lines,an in vivo estrogenic compound detection system

We previously reported the characteristics of a ChgH–GFP transgenic medaka line that indicates estrogenic compound pollution in environmental water by the green fluorescence of their liver. Recently, we established four more lines. In this study, the characteristics of the five transgenic medaka lines were investigated. The intensity of reporter gene expression varied among transgenic lines and generally correlated well with the amount of integrated transgene in each line. Line-specific ectopic expression was also observed. However, the sensitivity to 17-β estradiol did not differ among transgenic lines. Three transgenic lines are considered to be suitable as bio-indicators of estrogenic activity, due to the ease of observing green fluorescence in their livers. The transgenic lines can also detect the estrogenic activity of testosterone and 17-β trenbolone at the nominal concentration of 30 and 100 μg/l, respectively.     

Toxicity assessments of nanoscale zerovalent iron and its oxidation products in medaka (Oryzias latipes) fish

Iron-based nanotechnologies are increasingly used for environmental remediation; however, toxicologic impacts of iron nanoparticles on the aquatic ecosystem remain poorly understood. We treated larvae of medaka fish (Oryzias latipes) with thoroughly characterized solutions containing carboxymethyl cellulose (CMC)-stabilized nanoscale zerovalent iron (nZVI), aged nanoscale iron oxides (nFe-oxides) or ferrous ion (Fe[II]) for 12–14 days’ aqueous exposure to assess the causal toxic effect(s) of iron NPs on the fish. With the CMC-nZVI solution, the dissolved oxygen level decreased, and a burst of reactive oxygen species (ROS) was generated as Fe(II) oxidized to ferric ion (Fe[III]); with the other two iron solutions, these parameters did not significantly change. CMC-nZVI and Fe(II) solutions caused acute lethally and sublethally toxic effects in medaka larvae, with nFe-oxide-containing solutions causing the least toxic effects. We discuss modes of toxic action of iron NPs and chronic toxic effects in terms of hypoxia, Fe(II) toxicity and ROS-mediated oxidative damage.