菲律宾蛤仔(Ruditapes philippinarum)金属硫蛋白基因克隆及Cd~(2+)胁迫下的表达分析

金属硫蛋白(Metallothionein,MT)是一类富含半胱氨酸的小分子蛋白质,参与机体重金属解毒和金属元素代谢等生理过程。本研究采用RACE技术,克隆获得了菲律宾蛤仔(Ruditapes philippinarum)金属硫蛋白(RpMT)的全长cDNA序列。RpMT的cDNA全长为570bp,编码75个氨基酸,包含15个MT所特有的Cys-Xn-Cys结构。采用实时荧光定量PCR技术,分析了两种壳色菲律宾蛤仔(白蛤和斑马蛤)RpMT基因在Cd2+暴露后的表达变化。结果发现:Cd2+急性和亚慢性暴露均可导致两种壳色蛤仔消化腺和鳃组织RpMT基因表达量的显著上调;暴露后两种壳色蛤仔鳃组织RpMT基因表达量的增加幅度均高于消化腺组织,且以白蛤鳃组织基因表达水平的上调幅度较高。上述结果表明,RpMT可能在菲律宾蛤仔抵御Cd2+胁迫过程中发挥了重要作用。     

Response of antioxidant systems to copper in the gills of the clam Ruditapes decussatus

Copper (Cu) is an essential element for biological systems, however, when present in excess, is toxic. Metallothioneins can play an important role in Cu homeostasis and detoxification. Moreover, Cu can catalyse the production of toxic hydroxyl radicals that cause lipid peroxidation but defence systems in the cells can limit the oxidative damage. The present study was performed to investigate the effect of three Cu concentrations (0.5, 2.5 and 25 μg l⁻¹) on the response of antioxidant enzyme activities (superoxide dismutase (SOD), catalase (CAT), selenium-dependent glutathion peroxidase and total glutathion peroxidase), total proteins, metallothioneins (MT), malondialdehyde (MDA) concentrations in the gills of the clam, Ruditapes decussatus. The activity of antioxidant enzymes and total proteins, MT and MDA concentrations were measured in the gills of the clams after 1, 3, 7, 14, 21 and 28 days of Cu exposure. Results indicate that Cu only induces an imbalance in the oxygen metabolism during the first week of Cu exposure due to a decrease in mitochondrial SOD and CAT, selenium-dependent and total glutathion peroxidase activities. Cu also causes lipid peroxidation, measured by the MDA formation, that was Cu dependent. In the gills of clams exposed to 25 μg Cu/l, the excess of Cu triggers the induction of MT synthesis after 3 days of exposure.     

Cadmium accumulation by the blue crab, callinectes sapidus: Involvement of hemocyanin and characterization of cadmium-binding proteins

The accumulation of cadmium from seawater by the blue crab, Callinectes sapidus, was studied as a function of metal concentration and exposure time, with special emphasis on cadmium-binding proteins. Cadmium was found, in decreasing order of magnitude, in gills, digestive gland and hemolymph. When exposed to 0·5 ppm cadmium for 2–24h, virtually all of the cadmium in the cytosolic fraction of the gill was associated with a low molecular weight (LMW) cadmium-binding protein (MW 8000). However, after 48h of exposure only 50% of the cadmium in the cytosol was bound to this protein. The rest wasfound to be associated with proteins of a molecular weight of 300 000 and 60 000. This pattern of cadmium distribution did not change over a 12-day depuration period. Similar results were obtained upon exposure to 0·1 ppm cadmium. The pattern of cadmium accumulation in the cytosolic fraction of the digestive gland was in marked contrast to that observed for the gill. Initially, the cadmium was distributed over three low molecular weight fractions. During depuration the distribution of cadmium changed and all of the metal became bound to a low molecular weight protein (MW 9000). The cadmium concentrations in the gill and digestive gland remained essentially constant during depuration (12 days). The LMW cadmium-binding proteins were purified by a combination of gelpermeation and ion-exchange chromatography. Their molecular weight, spectral properties and amino acid composition are characteristic of the vertebrate metallothioneins. During exposure to cadmium the metal rapidly appeared in the hemolymph, mainly associated with hemocyanin. During depuration cadmium was transferred from the hemolymph to the digestive gland, demonstrating that hemocyanin acts as a carrier in trace metal transport.     

Tissue slice technology for assessing alterations in fish hepatic phase I and phase II XME activity

Alterations to hepatic xenobiotic metabolizing enzymes (XMEs) is an important biomarker of contaminant exposure in aquatic toxicology. Measurement of XMEs in fish liver slices in vitro is an emerging tool for examining enzyme activity and response within the intact 3-D architecture of the liver tissue. We examined integrated phase I/phase II, and phase II metabolism of XMEs from liver slices in control and B[a]P-treated rainbow trout and channel catfish. Fluorescent assay substrates to measure rates of metabolism included 7-methoxycoumarin (7-MC), 7-ethoxycoumarin (7-EC) and 7-hydroxycoumarin (7-HC). Time-dependent increases in metabolism, and a lower rate of 7-MC metabolism compared with 7-EC metabolism, were observed at all time points for both fish species. In rainbow trout, B[a]P pretreatment caused a 10-fold increase in phase I metabolism of both 7-MC and 7-EC, and a 1.6-fold increase in phase II metabolism of 7-HC. Phase I activity in channel catfish was not notably altered by B[a]P pretreatment. However, B[a]P pretreatment in channel catfish caused a 48% decrease in phase II metabolism of 7-HC. These results indicate differences in baseline and B[a]P-altered XME profiles between rainbow trout and channel catfish.     

Distributions of dissolved and particulate biogenic thiols in the subartic Pacific Ocean

Dissolved and particulate concentrations of the biogenic thiols cysteine (Cys), arginine–cysteine (Arg–Cys), glutamine–cysteine (Gln–Cys), γ-glutamate–cysteine (γ-Glu–Cys) and glutathione (GSH) were measured in the subartic Pacific Ocean in the summer of 2003 using high performance liquid chromatography (HPLC) with precolumn derivatization as reported in previous work. In this study, a preconcentration protocol for the derivatized thiols was utilized to extend detection limits of dissolved thiols to picomolar levels. The measured concentrations of particulate and dissolved thiols were uncoupled, with distinctive depth profiles and large differences in the particulate to dissolved ratios between individual compounds. Glutathione was the most abundant particulate thiol whereas the most abundant dissolved thiol was γ-Glu–Cys, with concentrations as high as 15 nM. Given the relatively small pool of intracellular γ-Glu–Cys and the very low dissolved concentrations of GSH, we hypothesize that glutathione released from cells is rapidly converted to the potentially degradation resistant γ-Glu–Cys outside the cell. The relatively high concentrations of other dissolved thiols compared to particulate concentrations implies both biological exudation and slow degradation rates. Some thiols appear to vary with changes in nutrient availability but this effect is difficult to decouple from changes in community structure inferred from pigment analyses. Dissolved thiol concentrations also exceed typical metal concentrations in the subartic Pacific, supporting previous arguments that they may be important in metal speciation.     

The correlation between bioaccumulation and pattern of stress-related genes expression of black sea bream (<Emphasis Type="Italic">Acanthopagrus schlegeli</Emphasis>) by cadmium exposure

In order to correlate the expression of detoxifying enzyme genes and Cd accumulation in black sea bream, we analyzed four tissues (brain, gills, liver, and muscle) from black sea breams that were exposed to four different concentrations of Cd (0, 2, 13, and 25 mg/L) for various durations (0, 24, 48, 72, and 96 h). The highest level of Cd was accumulated in the liver, followed by the gills, brain, and muscle. The accumulation of Cd was significantly correlated with the duration of exposure and the concentration in brain, gill, and liver tissue, but not in muscle tissue, and the rate of accumulation increased with Cd concentration. The expression of metallothionein II (MT II) mRNA exhibited a similar pattern as Cd accumulation, especially in that the expression of MT II mRNA decreased in muscle tissue with increases in exposure duration. In contrast, the expression of cytochrome P450 1A (CYP1A) mRNA was highest in the liver, followed by brain, muscle, and gill tissues, and in gills and muscle tissue of Cd-exposed fish, the expression of CYP1A mRNA fell below that of the control fish. Overall, the liver of black sea bream was the most sensitive to Cd exposure, and the expression of MT II mRNA was 200-fold greater than the control fish. These findings indicate that the detoxification mechanisms of black sea bream are influenced by both MT II and CYP1A and that the genes participate in the detoxification of different tissues.     

Seasonal variation of oxidative biomarkers in gills and digestive gland of green-lipped mussel Perna viridis from Arabian Sea

Investigations on seasonal variation in oxidative stress biomarkers were carried out on the natural population of green-lipped mussel Perna viridis collected from Bambolim beach area of Goa. Oxidative stress indices such as lipid peroxidation (LPX), hydrogen peroxide (H₂O₂), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX), glutathione transferase (GST), glutathione reductase (GR), reduced glutathione (GSH) and ascorbic acid (ASA) were measured in gills and digestive gland of P. viridis during February, May, August and November. The present study reveals two important aspects regarding the antioxidant defence status of tissues of P. viridis. Firstly, antioxidant capacity of tissues of P. viridis exhibits seasonal variation. Secondly, various components of antioxidant capacity such as oxidative stress markers, levels of antioxidant enzymes and small antioxidant molecules vary differently in tissues with respect to different seasons. Although the oxidative stress status of gills and digestive gland of P. viridis expressed in terms of LPX and H₂O₂ was the lowest in February, its level was maximal in gills and digestive gland during May and November, respectively. While activities of SOD and GPX of tissues of P. viridis were found to be low in August, activities of CAT and GR were recorded to be low in February. GST activity in gills although remained high in February, in digestive gland elevated values were recorded in August and November. A seasonal variation in the levels of small antioxidant molecules was also noticed. Among non enzymatic antioxidants ASA content of tissues was maximal in May and August in comparison to February and November, but GSH remained high in November. It therefore appears that environmental factors may play a crucial role in regulating the oxidative stress capacity of tissues of P. viridis.     

Municipal wastewater contamination in the Southern California Bight. Part II. Cytosolic distribution of contaminants and biochemical effects in fish livers

The objective of this study was to examine the cytosolic distribution of metals and oxygenated organic metabolites (MTBs), and biochemical effects, in livers of fish collected from both highly contaminated and less contaminated southern California coastal sites. Cytosolic extracts were separated by Sephadex G-75 column chromatography into high molecular weight (> 20 000 daltons) enzyme-containing (ENZ) pools, medium molecular weight (3000–20000 daltons) metallothionein- or metallothionein-like-containing (MT) pools, and low molecular weight (< 3000 daltons) glutathione-containing (GSH) pools.Concentrations of Cd, Cu and Zn were frequently lower in cytosolic pools of longspine combfish, yellowchin sculpin, and California tonguefish from highly contaminated Palos Verdes (PV) relative to those from less contaminated Santa Monica Bay (SMB) despite much higher concentrations of these metals in sediments at PV. Patterns of cytosolic metal distribution differed more between metals than between species or sampling locations. Most Cd, Cu and Zn occurred in the MT pools of these three species, with the exception of Zn in California tonguefish which occurred predominately in the ENZ pool. In all three species, ENZ-Cu showed positive slopes when regressed against total cytosolic Cu, while ENZ-Cd showed no significant slopes when regressed against total cytosolic Cd. Patterns for Zn were the least consistent among species, with higher ENZ-Zn slopes occurring in fish livers with lower cytosolic Zn concentrations.The largest portion of DDT and PCB oxygenated MTBs occurred in GSH pools of scorpionfish livers from PV or less contaminated Cortes Bank (CB). Concentrations of MTBs in ENZ- and MT-pools of CB scorpionfish livers showed positive slopes when regressed against total cytosolic- and GSH-MTBs.Positive slopes for regressions of ENZ-Cu, -Zn and -MTBs against total cytosolic concentrations are consistent with the model of an equilibrium-dependent exchange of these among cytosolic pools.The lower metal concentrations, higher glutathione concentrations, and higher catalase activities found in fish from PV relative to those from SMB are in accordance with effects known to result from exposure to organic contaminants.     

Response of antioxidant systems to copper in the gills of the clam Ruditapes decussatus

Copper (Cu) is an essential element for biological systems, however, when present in excess, is toxic. Metallothioneins can play an important role in Cu homeostasis and detoxification. Moreover, Cu can catalyse the production of toxic hydroxyl radicals that cause lipid peroxidation but defence systems in the cells can limit the oxidative damage. The present study was performed to investigate the effect of three Cu concentrations (0.5, 2.5 and 25 microg l(-1)) on the response of antioxidant enzyme activities (superoxide dismutase (SOD), catalase (CAT), selenium-dependent glutathion peroxidase and total glutathion peroxidase), total proteins, metallothioneins (MT), malondialdehyde (MDA) concentrations in the gills of the clam, Ruditapes decussatus. The activity of antioxidant enzymes and total proteins, MT and MDA concentrations were measured in the gills of the clams after 1, 3, 7, 14, 21 and 28 days of Cu exposure. Results indicate that Cu only induces an imbalance in the oxygen metabolism during the first week of Cu exposure due to a decrease in mitochondrial SOD and CAT, selenium-dependent and total glutathion peroxidase activities. Cu also causes lipid peroxidation, measured by the MDA formation, that was Cu dependent. In the gills of clams exposed to 25 microg Cu/l, the excess of Cu triggers the induction of MT synthesis after 3 days of exposure.     

Colonization of Sulfurovum sp. on the gill surfaces of Alvinocaris longirostris, a deep-sea hydrothermal vent shrimp

Deep‐sea hydrothermal vents are unique light‐independent ecosystems that are sustained by chemosynthetic bacteria. For many of the invertebrates inhabiting in such environments, bacteria play essential roles in both energy acquisition and detoxification of potentially toxic gases such as H₂S. In this study, the bacterial flora present on the gills of Alvinocaris longirostris (Bresiliidae: Caridea), a shrimp inhabiting hydrothermal vents (1532 m depth) at the Hatoma Knoll of the Okinawa Trough, was investigated. Bacterial 16S rDNA fragments were successfully amplified from the gills and 70% of these fragments showed an identical pattern in the restriction fragment length polymorphism analysis. These fragments were assigned to the ribotype AL‐1. Phylogenetic analyses suggest that AL‐1 forms a monophyletic clade with Sulfurovum spp. (ε‐Proteobacteria). Fluorescence in situ hybridization for AL‐1 and electron microscopy showed the presence of short‐rod bacteria lining up on the cuticular layer of the surface of the gill filaments. These results suggest that bacterial association with gills also occurs in bresiliid shrimps.     

Effect of cadmium, copper and mercury on antioxidant enzyme activities and lipid peroxidation in the gills of the hydrothermal vent mussel Bathymodiolus azoricus

Metals are known to influence lipid peroxidation and oxidative status of marine organisms. Hydrothermal vent mussels Bathymodiolus azoricus live in deep-sea environments with anomalous conditions, including high metal concentrations. Although B. azoricus are aerobic organisms they possess abundant methano and thioautotrophic symbiotic bacteria in the gills. The enzymatic defences (superoxide dismutase (SOD), catalase (CAT), total glutathione peroxidase (Total GPx) and selenium-dependent glutathione peroxidase (Se–GPx)) and lipid peroxidation were determined in the gills of B. azoricus exposed to Cd (0.9 μM), Cu (0.4 μM) and Hg (0.1 μM) with different times of exposure. The experiments were performed in pressurized containers at 9 ± 1 °C and 85 bars.Results show that vent mussels possess antioxidant enzymatic protection in the gills. Cd and Cu had an inhibitory effect in the enzymatic defence system, contrarily to Hg. These enzymatic systems are not completely understood in the B. azoricus, since reactive oxygen species might be produced through other processes than natural redox cycling, due to hydrogen sulphide and oxygen content present. Also the symbiotic bacteria may play an important contribution in the antioxidant protection of the gills.     

Role of biotransformation in determining aldicarb toxicity in fish

The carbamate pesticide, aldicarb, demonstrates significant acute toxicity in fish, and is readily biotransformed by most organisms studied. In fish, both the cytochrome P450 (CYP) and the flavin monooxygenase systems (FMO) are involved in bioactivating aldicarb to aldicarb sulfoxide, which is a more potent inhibitor of acetylcholinesterase (AChE). Channel catfish (Ictalurus punctatus), along with many other fresh water species, do not express FMO and are relatively resistant to the effects of aldicarb. This project examined the toxicity, AChE inhibition, metabolism, and toxicokinetic of aldicarb in channel catfish, and compared these values with an aldicarb-sensitive species, rainbow trout, which expresses FMO. Studies of in vitro and in vivo aldicarb biotransformation in catfish suggest that a low rate of bioactivation (10 times slower V_max), resulting in less initial conversion to the activated metabolite, aldicarb sulfoxide, may be a contributing factor to resistance of channel catfish to aldicarb toxicity. These data are supported by toxicokinetic and enzyme inhibition studies. This work demonstrates that differences in FMO expression among fish species may have significant influence on toxicity resulting from exposure to some xenobiotics.     

不同长牡蛎群体对海水酸化的生理响应差异分析

海水酸化暴露可对海洋生物产生多层面的影响。本研究以潮间带野生与潮下带养殖长牡蛎(不同生境背景)的不同组织(鳃、外套膜及消化腺)为研究对象,分析在室内调控p CO2模拟海水酸化暴露条件下,其基础代谢活动、能量代谢以及氧化应激相关指标的变化情况。结果显示:海水酸化暴露后,两种长牡蛎(Crassostrea gigas)的基础代谢过程均受到了一定抑制作用且受影响程度差异明显。潮间带野生与潮下带养殖长牡蛎的关键生理过程(能量代谢及氧化应激)对海水酸化暴露存在不同的响应变化,表明两种长牡蛎应对海水酸化的调节机制可能存在差异。依据PLS-DA分析结果显示,在所有生理指标中,对样本的差异贡献较高(VIP值>1)的指标为:SDH、AST、ATPase、ATP含量、糖原含量、CAT、GST及SOD,表明海水酸化暴露后,在两种长牡蛎的3种组织中上述指标的响应变化程度更大。综合评价分析多个生理指标的整体变化揭示:在海水酸化暴露条件下,潮间带野生长牡蛎比潮下带养殖长牡蛎对海水酸化的生理响应更为剧烈;相比于鳃及消化腺组织,长牡蛎外套膜组织可能受影响更大。     

Role of metallothioneins in Cu and Cd accumulation and elimination in the g gill and digestive gland cells of mytilus galloprovincialis lam.

The data presented in this paper demonstrate that exposure of mussels to Cu (40 μg/liter) or Cd (200 μg/liter) for a period of 2 days is sufficient to induce Cu or Cd thioneins in the gills of the metal-exposed animals. The concentrations of Cu and Cd thioneins increase during the period of metal exposure (3–4 weeks), confirming the fact that metallothioneins play a fundamental role in the accumulation of these metals in the tissues examined (gills and digestive gland). Moreover, it has been demonstrated that when metal-loaded mussels are returned to the field for the recovery period, Cu is rapidly eliminated from the gill and digestive gland cells, showing a biological half-life of 9–10 days, whereas Cd is released much more slowly from the tissues, only about 50% of the total metal being lost after 4 months of detoxification. Interestingly enough, during the detoxification period the concentration of the metal bound to thioneins also follows the same pattern as the total metal present in the tissues, in fact the concentration of Cu thioneins decreases near the control level in about 24 days but, in contrast, the Cd-thionein concentration in the cytisol of the gill and digestive gland cells remaains high, decreasing only by about 40% at the end of the 4 month recovery period.The data presented also demonstrate that when Cd-loaded, detoxified mussels are exposed to Cu (100 μg/liter) for 6 h, the metal taken up by the gill cells is able to displace the Zn which is always present in the Cd-thionein fraction, but it does not, however, displace the Cd present. In this case also, period (20 days), whereas the Cd-thionein content decreases slightly. Such data seem to indicate that the metabolic characteristics of thioneins are closely related to the metal bound to them, particularly where the elimination rate of the metals from the cells is concerned.     

中华哲水蚤抗氧化系统对海水酸化的响应

海洋浮游动物可以对因二氧化碳浓度升高导致的海水酸化做出敏感的响应,然而关于海水酸化对其生理机能的研究却较少。因此,我们研究了暴露于不同二氧化碳浓度（0.08,0.20,0.50 and 1.00%）的酸化条件下中华哲水蚤主要抗氧化酶以及2种解毒酶的活性变化情况。结果表明,暴露于酸化海水中的中华哲水蚤的GPx酶活性显著高于对照组,然而,其它抗氧化组分,包括GST、SOD活性,GSH水平和GSH/GSSG比值均被显著抑制。中华哲水蚤的ATPase活性被海水酸化显著刺激,而AchE活性却显著被抑制。此外,主成分分析结果表明,各指标变化的75.93%可以用第一和第二主成分解释。因二氧化碳浓度升高导致的海水酸化可以通过影响桡足类体内一些酶类的活性来影响其新陈代谢及存活。在未来的研究工作中,需要对海水酸化和其它环境因素的协同作用对桡足类的影响作进一步研究。     

原油污染对栉孔扇贝抗氧化酶活性的影响

以原油水溶性成分(water soluble fraction of crude oil,WSF)为污染物,采用暴露实验法,研究了栉孔扇贝(Chlamys farreri)鳃和消化腺组织中超氧化物歧化酶(Superoxide Dismutase,SOD)和过氧化氢酶(Catalase,CAT)活性的变化.结果表明,WSF污染下,鳃和消化腺组织SOD和CAT酶活性随暴露时间增加一般表现为降低-升高-降低的趋势,酶活性达到最高的时间随浓度不同而变化.第1天时消化腺SOD在0.08 mg/L浓度下被诱导,而后随时间增加表现为抑制效应;浓度为0.21和0.88 mg/L时消化腺SOD酶活性被抑制,随暴露时间延长而活性增加.暴露时间为4d时,石油烃浓度在0.08和0.88 mg/L时鳃组织SOD酶活性均被抑制,而浓度为0.21 mg/L时被诱导.消化腺和鳃组织SOD可以作为扇贝被污染胁迫的指标.     

海水酸化和磺胺甲恶唑复合暴露对大马蹄螺抗氧化酶活性和脂质过氧化的影响

为探讨海水酸化和磺胺甲恶唑环境残留对珊瑚礁区生物的影响,本文以南海珊瑚礁区关键礁栖生物大马蹄螺（Trochus niloticus） 为研究对象,探究了两种环境胁迫因素对大马蹄螺的氧化胁迫作用。分别采用两个 pH 水平 （pH 8.1、 pH 7.6） 和三个磺胺甲恶唑浓度水平 （0 ng/L、100 ng/L 和 1 000 ng/L） 单独和复合暴露 28 天,测定其对大马蹄螺鳃组织的超氧化物歧化酶 （SOD）、过氧化氢酶 （CAT） 和谷胱甘肽巯基转移酶 （GST） 活性以及谷胱甘肽 （GSH） 含量和脂质过氧化水平 （LPO） 的影响。结果发现：海水酸化和磺胺甲恶唑单独暴露能够导致大马蹄螺鳃组织出现不同程度的氧化应激现象,表现为抗氧化酶 （CAT、SOD 和 GST） 活性和谷胱甘肽含量的升高。在复合暴露条件下,大马蹄螺鳃组织的抗氧化酶（CAT、SOD 和 GST） 活性和谷胱甘肽含量均受到不同程度的抑制,同时伴随着脂质过氧化水平的显著升高。这表明短期海水酸化和磺胺甲恶唑复合暴露可导致大马蹄螺鳃组织出现氧化损伤现象,进而可能对个体存活和种群繁衍造成负面影响。     

Effects of β-naphthoflavone on hepatic biotransformation and glutathione biosynthesis in largemouth bass (Micropterus salmoides)

We are investigating the effects of in vivo exposure of prototypical enzyme inducing agents on hepatic biotransformation enzyme expression in largemouth bass (Micropterus salmoides), a predatory game fish found throughout the United States and Canada. The current study targeted those genes involved in biotransformation and oxidative stress that may be regulated by Ah-receptor-dependent pathways. Exposure of bass to β-naphthoflavone (β-NF, 66 mg/kg, i.p.) elicited a 7–9-fold increase in hepatic microsomal cytochrome P4501A-dependent ethoxyresorufin O-deethylase (EROD) activities, but did not affect cytosolic GST catalytic activities toward 1-chloro-2,4-dinitrobenzene (CDNB) or 5-androstene-3,17-dione (ADI). Glutathione S-transferase A (GST-A) mRNA expression exhibited a transient, but non-significant increase following exposure to β-NF, and generally tracked the minimal changes observed in GST–CDNB activities. Expression of the mRNA encoding glutamate-cysteine ligase catalytic subunit (GCLC), the rate-limiting enzyme in glutathione (GSH) biosynthesis, was increased 1.7-fold by β-NF. Changes in GCLC mRNA expression were paralleled by increases in intracellular GSH. In summary, largemouth bass hepatic CYP1A-dependent and GSH biosynthetic pathways, and to a lesser extent GST, are responsive to exposure to β-NF.     

A multi-biomarker approach to assess effects of Triclosan in the clam Ruditapes philippinarum

The effects of 7 days’ exposure to differing Triclosan (TCS) concentrations (300, 600, and 900 ng/L) were investigated in the clam Ruditapes philippinarum. Vitellogenin (Vg)-like protein levels in haemolymph and digestive gland from males and females, gill acetylcholinesterase (AChE) activity, superoxide dismutase (SOD) and catalase (CAT) activities in gills and digestive gland, and gill lipid peroxidation (LPO) were measured. The highest TCS concentrations decreased significantly Vg levels in male haemolymph and digestive gland, whereas no significant variations were found in females. The highest TCS concentrations increased significantly SOD activity in gills, but decreased it in digestive gland. No changes in CAT activity were observed. In gills, TCS reduced significantly AChE activity, but it did not induce significant variations in LPO. Our study demonstrates that TCS alters biochemical parameters in R. philippinarum, even at environmentally realistic concentrations, and suggests differing modes of action of the contaminant, in clams at least.     

Conjugation of 4-hydroxynonenal by largemouth bass (Micropterus salmoides) glutathione S-transferases

The glutathione S-transferases (GST) are a major group of conjugative enzymes involved in the detoxification of electrophilic compounds and products of oxidative stress. We have previously described the kinetics of hepatic GST conjugation in largemouth bass using a variety of synthetic GST reference substrates. In the present study, we investigated the ability of largemouth bass hepatic GSTs to conjugate 4-hydroxynon-2-enal (4HNE), a mutagenic and cytotoxic alpha-beta-unsaturated aldehyde produced during oxidative injury. Hepatic cytosolic fractions from largemouth bass rapidly catalyzed GSH-dependent 4HNE conjugation, with the rate of GST-4HNE conjugation in bass liver exceeding those of several other mammalian and aquatic species. No apparent sex-related differences in GST-4HNE activity were observed among adult bass. SDS-PAGE and Western blotting analysis of GSH affinity-purified bass liver cytosolic GST revealed the presence of two major GST subunits of approximately 30 and 27 KDa that exhibited slight cross-reactivity when probed with a rat alpha class GST antibody, but not to rat mu, pi or theta class GST. The rapid conjugation of 4HNE by hepatic GST suggests an important role for GSTs in protecting against peroxidation of polyunsaturated fatty acids in bass liver.