Vitellogenin Production Research Articles

Effects of Wastewater Treatment Plant Effluent in a Receiving Stream on Reproductive Behavior of Fathead Minnows (Pimephales promelas)

Wastewater treatment plant effluents contain a variety of endocrine disrupting chemicals (EDCs), including chemicals with estrogenic activity such as 17β-estradiol (E2), 17α-ethinyl estradiol (EE2), and nonylphenols. These substances can affect both behavior and physiology in vertebrate animals. To explore the presence and effects of these EDCs in a natural setting, juvenile and adult male fathead minnows, Pimephales promelas, were held in cages upstream and downstream of the effluent site of a wastewater treatment plant for 21 days and subsequently tested for changes in reproductive behaviors and production of vitellogenin. Additionally, estrogenic activity in the stream was measured using a yeast bioassay. Estrogenicity was found to be significantly higher downstream of the wastewater effluent when compared to levels upstream. Vitellogenin levels did not show a correlational pattern with levels of estrogenicity in the water, but two measures of reproductive behaviors occurred significantly less often in downstream males than upstream males. This suggests that a brief (three-week) exposure to stream water containing wastewater treatment plant effluent can bring about changes in reproductive behavior of fish and that behavior may be more sensitive to low levels of environmental endocrine disruptors than vitellogenin production.

Assessing effects of aromatase inhibition on fishes with group-synchronous oocyte development using western mosquitofish (Gambusia affinis) as a model

Exposure to certain anthropogenic chemicals can inhibit the activity to cytochrome P450 aromatase (CYP19) in fishes leading to decreased plasma 17β-estradiol (E2), plasma vitellogenin (VTG), and egg production. Reproductive dysfunction resulting from exposure to aromatase inhibitors has been extensively investigated in several laboratory model species of fish. These model species have ovaries that undergo asynchronous oocyte development, but many fishes have ovaries with group-synchronous oocyte development. Fishes with group-synchronous oocyte development have dynamic reproductive cycles which typically occur annually and are often triggered by complex environmental cues. This has resulted in a lack of test data and uncertainty regarding sensitivities to and adverse effects of aromatase inhibition. The present study used the western mosquitofish (Gambusia affinis) as a laboratory model to investigate adverse effects of chemical aromatase inhibition on group-synchronous oocyte development. Adult female western mosquitofish were exposed to either 0, 2, or 30 μg/L of the model nonsteroidal aromatase inhibiting chemical, fadrozole, for a complete reproductive cycle. Fish were sampled at four time-points representing pre-vitellogenic resting, early vitellogenesis, late vitellogenesis/early ovarian recrudescence, and late ovarian recrudescence. Temporal changes in numerous reproductive parameters were measured, including gonadosomatic index (GSI), plasma sex steroids, and expression of selected genes in the brain, liver, and gonad that are important for reproduction. In contrast to fish from the control treatment, fish exposed to 2 and 30 μg/L of fadrozole had persistent elevated expression of cyp19 in the ovary, depressed expression of vtg in the liver, and a low GSI. These responses suggest that completion of a group-synchronous reproductive cycle was unsuccessful during the assay in fish from either fadrozole treatment. These adverse effects data show that exposure to aromatase inhibitors has the potential to cause reproductive dysfunction in a wide range of fishes with both asynchronous and group-synchronous reproductive strategies.

Effectiveness of Curcumin Supplementation in feed on liver productivity and reproductive performance of female carp Cyprinus carpio L.

The availability of quality seeds, on time with large amounts is a major problem in freshwater aquaculture. Supplementation of curcumin in female carp aims to increase the reproductive productivity of fish through increased activity of the liver in the synthesis of vitellogenin, a precursor of egg yolk, that play a role in the growth and development of eggs until larvae. This study used 25 female carp, which supplemented with curcuminat several doses: 0; 2.5; 5; 10; and 20 g of curcumin/kg of feed. The collected data was analyzed using a completely randomized design (CRD). The results showed that supplementation of curcumin female carp significantly (p<0.05) affected the malondialdehyde(MDA) concentration of liver, estradiol hormone in the blood, gonadosomatic index, relative fecundity, and larval survival, but did not significantly (p>0.05) affect the superoxide dismutase (SOD), vitellogenin concentration in blood and egg diameter. The study showed that the supplementation of curcumin prevents liver damage showed by a decreased level of liver MDA. The supplementation of curcumin with a dose of 5 g/kg of feed optimizes the productivity and reproductive performance of carp by increasing the production of vitellogenin, the diameter of egg, and the number of survival larva through optimizing the performance of the fish liver.

Insulin/IGF signaling and TORC1 promote vitellogenesis via inducing juvenile hormone biosynthesis in the American cockroach.

Vitellogenesis, including vitellogenin (Vg) production in the fat body and Vg uptake by maturing oocytes, is of great importance for the successful reproduction of adult females. The endocrinal and nutritional regulation of vitellogenesis differs distinctly in insects. Here, the complex crosstalk between juvenile hormone (JH) and the two nutrient sensors insulin/IGF signaling (IIS) and target of rapamycin complex1 (TORC1), was investigated to elucidate the molecular mechanisms of vitellogenesis regulation in the American cockroach, Periplaneta americana Our data showed that a block of JH biosynthesis or JH action arrested vitellogenesis, in part by inhibiting the expression of doublesex (Dsx), a key transcription factor gene involved in the sex determination cascade. Depletion of IIS or TORC1 blocked both JH biosynthesis and vitellogenesis. Importantly, the JH analog methoprene, but not bovine insulin (to restore IIS) and amino acids (to restore TORC1 activity), restored vitellogenesis in the neck-ligated (IIS-, TORC1- and JH-deficient) and rapamycin-treated (TORC1- and JH-deficient) cockroaches. Combining classic physiology with modern molecular techniques, we have demonstrated that IIS and TORC1 promote vitellogenesis, mainly via inducing JH biosynthesis in the American cockroach.

Screening of estrogenic endocrine-disrupting chemicals in meat products based on the detection of vitellogenin by enzyme-linked immunosorbent assay

Vitellogenin (VTG) is typically produced by females but when present in males can indicate contamination with estrogenic endocrine-disrupting chemicals (EDCs). Here we used primary hepatocytes isolated from male crucian carp uncontaminated with estrogenic EDCs as a culture model. Nine EDCs were used for validation, and an enzyme-linked immunosorbent assay (ELISA) was used to quantify VTG production by primary hepatocytes. The lower limit of detection of 17β-E2, DES, and HES using the method was 10−12 M. Fresh pork was roasted, and non-hydrolytic acetonitrile-vortexing was used to extract potential estrogenic EDCs. The extracted substances were separated by thin-layer chromatography (TLC), and dimethyl terephthalate present in roast pork was found to induce high VTG concentrations. Therefore, the detection of VTG by ELISA provides a sensitive and reliable method for detecting known and unknown estrogenic EDCs in food products.

Tyramine and its receptor TYR1 linked behavior QTL to reproductive physiology in honey bee workers (Apis mellifera)

Honey bees (Apis mellifera) provide an excellent model for studying how complex social behavior evolves and is regulated. Social behavioral traits such as the division of labor have been mapped to specific genomic regions in quantitative trait locus (QTL) studies. However, relating genomic mapping to gene function and regulatory mechanism remains a big challenge for geneticists. In honey bee workers, division of labor is known to be regulated by reproductive physiology, but the genetic basis of this regulation remains unknown. In this case, QTL studies have identified tyramine receptor 1 (TYR1) as a candidate gene in region pln2, which is associated with multiple worker social traits and reproductive anatomy. Tyramine (TA), a neurotransmitter, regulates physiology and behavior in diverse insect species including honey bees. Here, we examine directly the effects of TYR1 and TA on worker reproductive physiology, including ovariole number, ovary function and the production of vitellogenin (VG, an egg yolk precursor). First, we used a pharmacology approach to demonstrate that TA affects ovariole number during worker larval development and increases ovary maturation during the adult stage. Second, we used a gene knockdown approach to show that TYR1 regulates vg transcription in adult workers. Finally, we estimated correlations in gene expression and propose that TYR1 may regulate vg transcription by coordinating hormonal and nutritional signals. Taken together, our results suggest TYR1 and TA play important roles in regulating worker reproductive physiology, which in turn regulates social behavior. Our study exemplifies a successful forward-genetic strategy going from QTL mapping to gene function.

Investigation on Endocrine Disruption of the Larval Lampricide 3-Trifluoromethyl-4-Nitrophenol: Short-Term Reproduction Assay with Fathead Minnow (Pimephales promelas).

3-Trifluoromethyl-4-nitrophenol (TFM) has been used for more than 60 yr to control the invasive parasitic sea lamprey (Petromyzon marinus) in the Great Lakes Basin (USA/Canada). In the early 1990s, researchers reported that TFM induced vitellogenin in fish and that TFM was an agonist for the rainbow trout estrogen receptor. To support continued registration of TFM for sea lamprey control, regulatory agencies required further testing to evaluate potential endocrine disruption effects. Fathead minnow (Pimephales promelas) were exposed to TFM at measured concentrations of 0.0659, 0.181, 0.594, 1.79, and 5.11 mg active ingredient (a.i.)/L for 21 d. No-observable- and lowest-observable-effect concentrations (NOEC and LOEC, respectively) were determined to be 1.79 mg/L or greater for each endpoint. Male survival in the highest treatment group was reduced relative to the controls. Percentage of egg fertility was reduced in the highest treatment group, resulting in an estimated NOEC of 1.79 mg/L. Whereas no effect on the gonadosomatic index (GSI) was observed for males, female GSI was increased in the 5.11-mg/L treatment. Vitellogenin production was not altered relative to the controls for all TFM treatment groups. However, female testosterone was elevated in the 5.11-mg/L treatment. The results suggest that prolonged exposure to TFM at concentrations exceeding 1.79 mg/L has the potential to disrupt endocrine function. Biologically relevant effects were found at the highest exposure concentration following a 21-d exposure. However, the duration of exposure in our study is not consistent with typical treatment durations (12 h) for sea lamprey control. Environ Toxicol Chem 2020;39:1599-1607. © 2020 SETAC.

Analysis of vitellogenin by histochemical method as an indicator of estrogenic effect in male Danio rerio exposed to metals.

Several chemicals present in the aquatic environment have the ability to alter the endocrine system of aquatic organisms, including the metals cadmium (Cd) and zinc (Zn). In males, a tool to analyze this effect is the vitellogenin (VTG) detection, a protein with defined function in the yolk production and subsequent aid to embryo-larval development. This protein is produced just by females and can be detected by simple and cheap methods such as histochemical method. Thus, this study aimed to analyze the capacity of Cd and Zn to induce VTG production in Danio rerio males and determine whether the histochemical labeling method is efficient to study estrogenic effects in this species. For this, D. rerio adult males were exposed chronically (21days) to 0.25 and 1μg/L of Cd and 120 and 180μg/L of Zn, values allowed by the Brazilian and American legislation to aquatic life protection. After which, the organisms were submitted to a histological process as preparation to liver VTG marking by histochemical method and subsequent qualitative analysis of this protein in zebrafish's liver. After analyzes, it was possible observed that Cd and Zn are capable of inducing VTG production in D. rerio malesandthat the histochemistry method is efficient for detection of estrogenic effects in this species.

English

Aquatic organisms in the coastal regions are often exposed to many anthropogenic effluents. Kuwait Bay receives considerable quantities of sewage and industrial effluents, and thus, naturally, many endocrine disrupting chemicals (EDCs) find their way into the coastal waters. These EDCs could influence the physiology of the aquatic organisms inhabiting the coastal region. Response of two important local fish of aquaculture importance, bluefin bream (Sparidentax hasta) and yellowfin bream (Acanthopagrus latus) was evaluated by injecting a standard vitellogenin inducer, 17 β-estradiol (17 BEST).   The inducer was used at 2, 4 and 6 µg/fish (injected intraperitoneally) to induce vitellogenin production in juvenile fish (150-200 g).  There was a dose-dependent Vtg induction response as evidenced by a qualitative immune-dot assay. All levels significantly enhanced Vtg gene expression, as early as one-hour post injection. The expression was noticed to be the highest in liver tissue at 6 h post injection. The bluefin bream was used for amplification of the Vtg gene yielding 282 bp product which was sequenced. This indicated a 97% sequence similarity with the Vtg gene sequence of European seabream, S. aurata. Key words: Induction, Vtg, EDCs, Kuwait, fish, effluent. &nbsp

Adverse Outcome Pathway Network–Based Assessment of the Interactive Effects of an Androgen Receptor Agonist and an Aromatase Inhibitor on Fish Endocrine Function

Predictive approaches to assessing the toxicity of contaminant mixtures have been largely limited to chemicals that exert effects through the same biological molecular initiating event. However, by understanding specific pathways through which chemicals exert effects, it may be possible to identify shared "downstream" nodes as the basis for forecasting interactive effects of chemicals with different molecular initiating events. Adverse outcome pathway (AOP) networks conceptually support this type of analysis. We assessed the utility of a simple AOP network for predicting the effects of mixtures of an aromatase inhibitor (fadrozole) and an androgen receptor agonist (17β-trenbolone) on aspects of reproductive endocrine function in female fathead minnows. The fish were exposed to multiple concentrations of fadrozole and 17β-trenbolone individually or in combination for 48 or 96 h. Effects on 2 shared nodes in the AOP network, plasma 17β-estradiol (E2) concentration and vitellogenin (VTG) production (measured as hepatic vtg transcripts) responded as anticipated to fadrozole alone but were minimally impacted by 17β-trenbolone alone. Overall, there were indications that 17β-trenbolone enhanced decreases in E2 and vtg in fadrozole-exposed fish, as anticipated, but the results often were not statistically significant. Failure to consistently observe hypothesized interactions between fadrozole and 17β-trenbolone could be due to several factors, including lack of impact of 17β-trenbolone, inherent biological variability in the endpoints assessed, and/or an incomplete understanding of interactions (including feedback) between different pathways within the hypothalamic-pituitary-gonadal axis. Environ Toxicol Chem 2020;39:913-922. © 2020 SETAC.

Implications of increasing pollution levels on commercially important fishes in Lake Victoria

Lake Victoria receives huge quantities of effluent from domestic, agricultural and industrial sources. We used fish condition factor (K), vitellogenin (VTG) production and liver lesions as biomarkers to assess pollution levels in the lake. We tested the hypothesis that pollution levels do not affect the selected biomarkers. Beach seine and cast nets were used to collect Oreochromis niloticus (n = 230), Lates niloticus (n = 99) and Protopterus aethiopicus (n = 37) in areas presumed to be less or more polluted, both inshore and offshore. K was lower in more polluted compared to less polluted areas of the lake. VTG production was high in both less and more polluted areas for O. niloticus (0.77 ± 0.08 µg/L), L. niloticus (0.73 ± 0.09 µg/L) and P. aethiopicus (0.55 ± 0.06 µg/L). Liver tissue showed lesions such as vacuolations, cellular degeneration, sinusoidal dilation, focal necrosis, increased Küpffer cells and congestion of sinusoids. The prevalence of liver tissue alteration showed normal lesion (19.9%, n = 73), slight (8.2%, n = 30), moderate (41.5%, n = 152), severe (18.6%, n = 68) alterations and irreparable damage (11.8%, n = 43). Severe liver alterations in O. niloticus, L. niloticus and P. aethiopicus were higher in more polluted compared to less polluted areas. Chemical contamination of Lake Victoria caused liver lesions and other changes in fishes, possibly leading to adverse effects on the lake’s fisheries resources. Overtime, such chemical contamination could lead to negative impacts on the consumers of fish if actions are not taken to mitigate the risks.

RNA interference-mediated knockdown of the transcription factor Krüppel homologue 1 suppresses vitellogenesis in Chilo suppressalis.

Vitellogenesis in holometabolous insects involves the production and secretion of vitellogenin (Vg) and other yolk protein precursors in developing oocyte by the fat body, all of which is predominantly orchestrated by juvenile hormone (JH). Krüppel homologue 1 (Kr-h1) is a zinc finger transcription factor that has been demonstrated to be a JH-early inducible gene and to contribute to reproduction. However, the exact molecular function of Kr-h1 in insect reproduction is poorly understood. In the current study, we used the notorious pest Chilo suppressalis as a model system to investigate the role of Kr-h1 in female reproduction. Cloning and sequencing C. suppressalis Kr-h1 revealed that it shares high identity with its homologues from other lepidopteran insects. Moreover, RNA interference-mediated knockdown of CsKr-h1 substantially reduced the transcription of Vg in the fat body, dramatically decreased yolk protein deposition and also impaired oocyte maturation and ovarian development, indicating that Kr-h1 is indispensable for normal vitellogenesis in C. suppressalis. Based on these results, we conclude that Kr-h1 is crucial to reproduction in insects and that targeting this gene could potentially be a new way to suppress rice pests.

Protective effects of beta-cyclodextrins vs. zearalenone-induced toxicity in HeLa cells and Tg(vtg1:mCherry) zebrafish embryos

Zearalenone is a xenoestrogenic mycotoxin produced by Fusarium species. High exposure with zearalenone induces reproductive disorders worldwide. Cyclodextrins are ring-shaped host molecules built up from glucose units. The apolar cavity of cyclodextrins can entrap so-called guest molecules. The formation of highly stable host-guest type complexes with cyclodextrins can decrease the biological effect of the guest molecule. Therefore, cyclodextrins may be suitable to decrease the toxicity of some xenobiotics even after the exposure. In this study, the protective effect of beta-cyclodextrins against zearalenone-induced toxicity was investigated in HeLa cells and zebrafish embryos. Fluorescence spectroscopic studies demonstrated the formation of stable complexes of zearalenone with sulfobutyl-, methyl-, and succinyl-methyl-substituted beta-cyclodextrins at pH 7.4 (K = 1.4–4.7 × 104 L/mol). These chemically modified cyclodextrins considerably decreased or even abolished the zearalenone-induced loss of cell viability in HeLa cells and mortality in zebrafish embryos. Furthermore, the sublethal effects of zearalenone were also significantly alleviated by the co-treatment with beta-cyclodextrins. To test the estrogenic effect of the mycotoxin, a transgenic bioindicator zebrafish model (Tg(vtg1:mCherry)) was also applied. Our results suggest that the zearalenone-induced vitellogenin production is partly suppressed by the hepatotoxicity of zearalenone in zebrafish. This study demonstrates that the formation of stable zearalenone-cyclodextrin complexes can strongly decrease or even abolish the zearalenone-induced toxicity, both in vitro and in vivo. Therefore, cyclodextrins appear as promising new mycotoxin binders.

Role of neuroparsin 1 in vitellogenesis in the mud crab, Scylla paramamosain

Neuroparsin (NP) is an important neuropeptide in invertebrates. It is well-known that NP displays multiple biological activities, including antidiuretic and inhibition of vitellogenesis in insects. However, the information about its effect in crustaceans is scarce. In this study, the sequence of Sp-NP1 was selected from the transcriptome database from the mud crab, Scylla paramamosain. Sequence analyses indicate that the Sp-NP1 amino acid (AA) sequences consist of a 27 AA signal peptide and a 74 AA mature peptide, which contains 12 cysteine residues. qRT-PCR analysis has revealed that the expressions of Sp-NP1 gene are high in the nervous tissues and extremely low in the ovary and hepatopancreas. In situ hybridization has shown that the positive signals are localized in cell cluster 6 of protocerebrum and cell clusters 10 and 11 of deutocerebrum. The presence of Sp-NP1 in the haemolymph has been detected in S. paramamosain through western blot, which indicates that Sp-NP1 serves as an endocrine factor in the regulation of physiological activities. In vitro experiments have further shown that the mRNA level of vitellogenin in the hepatopancreas notably decreases following administration of recombinant Sp-NP1, while the mRNA level of vitellogenin receptor and cyclin B in the ovary shows no significant differences. Collectively, Sp-NP1 possibly can inhibit the production of vitellogenin in the hepatopancreas and has no direct effect on the ovary in S. paramamosain.

Does hepatotoxicity interfere with endocrine activity in zebrafish (Danio rerio)?

Vitellogenin (VTG), a well-established biomarker for the diagnosis of endocrine activity in fish, is used in multiple OECD test guidelines (TG) to identify activities of chemicals on hormonal pathways. However, the synthesis of VTG may not only be modified by typical endocrine-related pathways, but also through non-endocrine-mediated processes. In particular, hepatotoxicity, i.e. toxicant-induced impairment of liver structure and function, might influence VTG as a biomarker, since VTG is synthesized in hepatocytes. An intimate understanding of the interplay between endocrine-related and non-endocrine-related pathways influencing VTG production is crucial for the avoidance of erroneous diagnoses in hazard assessment for regulatory purposes of chemical compounds. In order to investigate whether hepatotoxicity may interfere with hepatic VTG synthesis, adult zebrafish (Danio rerio) were exposed to three well-known hepatotoxicants, acetaminophen, isoniazid and acetylsalicylic acid, according to OECD TG 230. Various hepatotoxicity- and endocrine system-related endpoints were recorded: mRNA expression of selected endocrine- and hepatotoxicity-related marker genes in the liver; VTG levels in head/tail homogenates; and liver histopathology. All three test compounds induced significant, but mild single cell necrosis of hepatocytes and transcriptional changes of hepatotoxicity-related marker genes, thus confirming hepatotoxic effects. A positive correlation between hepatotoxicity and reduced hepatic VTG synthesis was not observed, with the single exception of a weak increase in female zebrafish exposed to APAP. This suggests that – in studies conducted according to OECD TG 229 or 230 − it is unlikely that hepatotoxic chemicals will interfere with the hepatic capacity for VTG synthesis.

Firebugs fight infection with egg yolk protein

Fighting off an infection is never easy, but insects are prepared. By releasing antimicrobial peptides (mini-proteins) from the fat body – the insect equivalent of the liver – and mobilising specialised cells that target invading infections in the haemolymph (insect blood), insects are able to defend themselves when wounded or at risk of infection. However, Dalibor Kodrík and colleagues from the Biology Centre of the Czech Academy of Sciences, wondered whether a group of proteins known as vitellogenins – which are usually produced by female animals and go on to create egg yolk proteins – might also contribute to insect immunity. Kodrík says, ‘Studies in the silkworm and honeybee reported strong antibacterial activity of vitellogenins’. Could the proteins contribute to the immunity of his favourite insects, firebugs (Pyrrhocoris apterus)? Knowing that the nematode Steinernema carpocapsae, which infects insects with toxic Xenorhabdus bacteria, and the lethal Isaria fumosorosea fungus are used to control insect numbers, Kodrík decided to investigate whether either biological control agent triggers vitellogenin production in firebugs and whether the proteins offered any protection from these infections.After injecting one group of firebugs with the toxic nematodes and a second group with the fungus, Emad Ibrahim and Umesh Guatam collected the infected insects’ haemolymph and fat bodies several days later to find out whether the insects were producing vitellogenins to combat the infections. When they measured the levels of vitellogenin mRNA and protein, Radmila Čapková Frydrychová, Andrea Bednářová and Pavel Jedlička discovered that the nematode infection significantly boosted the males’ vitellogenin mRNA production 7.7-fold and protein 1.5-fold. However, the infection caused the females’ vitellogenin mRNA levels in the fat body to crash, dramatically dropping 150-fold, while their haemolymph vitellogenin protein levels also plummeted.In contrast, the male firebugs’ response to the slower developing fungal infection was only modest, while the females experienced the same haemolymph and fat body reductions in vitellogenin that the scientists had discovered in the nematode-infected insects. In addition, when the team tested the mortality rates from both infections, it seemed that the female firebugs were better protected than their male counterparts. And when Václav Krištu˚fek treated the nematode's toxic Xenorhabdus lodgers with vitellogenin, the bacteria succumbed, although the protein made little impact on the fungal infection.So, it appears that vitellogenin does contribute to the insect's immunity, offering some protection from nematode and fungal infections. Kodrík also suspects that the reduction in vitellogenin levels experienced by infected females may not be as unexpected as it first appears. ‘One can speculate that the vitellogenin level sufficient for effective defence against pathogens might be much lower than that required for nutritional supply of developing eggs’, says Čapková Frydrychová, who suggests that infected females may shut down non-essential processes, such as egg development, at times of crisis to invest energy in fighting the infection. And Kodrík is keen to discover whether the essential egg protein is also a key player in the immune response of other insects, including Drosophila, which could allow him to begin unravelling vitellogenin's antibiotic mechanism.

The expression of cockroach insulin-like peptides is differentially regulated by physiological conditions and affected by compensatory regulation

In insects, the insulin receptor (InR) pathway is involved in regulating key physiological processes, including juvenile hormone (JH) synthesis, vitellogenin production, and oocyte growth. This raises the question about which ligand (or ligands) binds to InR to trigger the above effects. We have cloned seven insulin-like peptides (BgILP1 to 7) from female Blattella germanica cockroaches and found that the brain expresses BgILP1 to 6, the fat body BgILP7, and the ovary BgILP2. Starvation induces the reduction of BgILP3, 5, and 6 mRNA levels in the brain, and the various BgILPs are differentially expressed during the gonadotrophic cycle. In addition, by knocking down the BgILPs we were able to identify compensatory regulation at transcriptional level between the different BgILPs, although none of the BgILP knockdown assays, including the knockdown of the seven BgILPs, produced the same phenotypes that we achieved by depleting InR. Taken together, the results indicate that B. germanica ILPs are differentially expressed in tissues and in response to physiological conditions, and that they are affected by compensatory regulation.

Freshwater snail Parafossarulus striatulus estrogen receptor: Characteristics and expression profiles under lab and field exposure

The modes of action by which putative endocrine disrupting chemicals (EDCs) elicit toxicity in mollusks remains unclear due to our limited understanding of the molluscan endocrine system. We identified and partially characterised the estrogen receptor (ER) of the mollusk Parafossarulus striatulus. The full-length cDNA of the ER of P. striatulus (psER) was isolated and found to have an ORF of 1386 bp which corresponded to 461 amino acids. Phylogenetic analysis revealed that psER is an orthologue of ER of other mollusks. Moreover, the DNA-binding domain, ligand-binding domain, P-box, D-box, and AF2 domain were also identified in psER. Exposure of females and males to 17β-estradiol (E2, 100 ng/L) for 24 h and 72 h did not alter psER transcription, but exposure to 17α-methyltestosterone (MT, 100 μg/L) for 72 h significantly decreased ER transcription in females only (p < 0.05). psER transcription was surveyed in males and females seeded in different regions in Taihu Lake, China. psER transcription were elevated among females and males maintained at site ML. This elevation was statistically significant (p < 0.05) among male snails as compared to snails held at the more pristine site of SZ. This was different to the results from lab, implying that some unknown chemicals or other environmental factors in field could affect psER transcription level in snails. Furthermore, females and males held at site ML also exhibited a significant elevation in vitellogenin transcription as compared to snails held at site SZ, suggesting that vitellogenin production may be directly regulated by psER or co-regulated with psER in this species.

Differences in vitellogenin production between laboratory-raised and wild fathead minnows: Potential consequences for understanding estrogenic exposure in wild populations.

Differences in vitellogenin production between laboratory-raised and wild fathead minnows: Potential consequences for understanding estrogenic exposure in wild populations.

Prolonged exposure of di(2-ethylhexyl) phthalate induces multigenerational toxic effects in Caenorhabditis elegans

The plasticizer di(2-ethylhexyl) phthalate (DEHP) is an emerging organic contaminant that has represented a risk for organisms present in the environment. However, there is still limited information regarding DEHP-induced multigenerational toxicity and the underlying mechanisms. In this study we investigated the multigenerational toxic effects including locomotive behaviors and reproduction upon prolonged DEHP exposure (from larval L1 to adult) and the underlying mechanisms in the nematode Caenorhabditis elegans. The multigenerational effects were examined over 6 generations (F0–F5) with only parental C. elegans (F0) was exposed to DEHP from larval L1 to adults (72h), and the subsequent offsprings (F1–F5) were grown under DEHP-free conditions. The results showed that prolonged exposure (72h) to various concentrations of DEHP caused dose-dependent locomotive impairments and reproduction defects in C. elegans and that a concentration of 0.2mg/L DEHP was enough to cause such sublethal effects. The results showed that after prolonged exposure to DEHP in the F0 generation, abnormal locomotive behaviors such as reduced body bends and head thrashes were observed from generations F0 to F5. Additionally, prolonged exposure to DEHP (20mg/L) in F0 significantly reduced total brood size in F0, and this parental exposure was sufficient to cause multigenerational reproductive toxicity in the offspring generations (F1–F5) as well. Furthermore, the expressions of reproduction-related genes such as vit-2 and vit-6 were down-regulated by about 20% until F3, and the expression of H3Kme2 demethylase, spr-5, was downregulated in F1 by about 40%. Results from this study demonstrate that prolonged exposure to DEHP only at F0 adversely affected reproduction and locomotive behaviors in C. elegans across generations and might be associated with inadequate vitellogenin production and malfunction of H3Kme2 demethylase. This study implies that parentally prolonged exposure to DEHP caused multigenerational defects in both reproduction and locomotive behaviors raising the potential health and ecological risk.