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An alien metabolite vs. a synthetic chemical hazard: An ecotoxicological comparison in the Mediterranean blue mussel
Abstract
Bioactive natural products from marine invasive species may dramatically impact native communities, while many synthetic pharmaceutical drugs are released into the marine environment and have long-lasting harmful effects on aquatic life. Sometimes, metabolites from alien species and synthetic compounds share similar mechanisms of action, suggesting comparable ecotoxicological impacts. This applies to the alkaloid caulerpin (CAU) from the green alga Caulerpa cylindracea, highly invasive in the Mediterranean Sea, and to the synthetic lipid-lowering drug fenofibrate (FFB), both acting as agonists of peroxisome proliferator-activated receptors (PPARs). Analogies with FFB, which is widely considered hazardous to the aquatic environment, have led to concerns about the ecotoxicological potential of CAU. The problem has implications for public health as CAU is well known to enter the food web accumulating in fish of commercial importance. Here, we compared the effects of FFB and CAU through biochemical and histopathological analysis on a relevant bioindicator molluscan species, the mussel Mytilus galloprovincialis. Under laboratory conditions, mussels were fed with food enriched with CAU or FFB. After treatment, biochemical markers were analyzed revealing metabolic capacity impairments, cellular damage, and changes in acetylcholinesterase activity in mussels fed with FFB-enriched food. NMR-based metabolomic studies also showed significant alterations in the metabolic profiles of FFB-treated mussels. In addition, dietary administration of FFB produced morphological alterations in the mussels' gills and digestive tubules. Obtained results confirm that FFB is harmful to aquatic life and that its release into the environment should be avoided. Conversely, dietary treatment with CAU did not produce any significant alterations in the mussels. Overall, our results pave the way for the possible valorization of the huge biomass from one of the world's worst invasive species to obtain CAU, a natural product of interest in drug discovery.
... Our results agree with previous ones in bivalves which found a capacity of some PhACs such as fibrates or anti-inflammatories to increase of the LPO levels in bivalves (Aguirre-Martinez, 2021;Gonzalez-Rey and Bebianno, 2012). This process affected the balance between ROS production and elimination capacity, with elevated lipid peroxidation levels and morphological alterations in gills and digestive tubules that increased the cellular damage (Quinn et al., 2011;Russo et al., 2023). ...
... For instance, Canesi et al. (2007) noted increased levels of GST and CAT activities in the digestive gland after BEZ and GEM exposure. However, in agreement with our findings, other pharmaceuticals, such as FF, did not alter GST and SOD in the soft tissues of M. galloprovincialis (Russo et al., 2023), and GST and GPx activities remained unaffected following FLX exposure in the digestive gland of the bivalve Perna perna (Cortez et al., 2019). ...
... The co-occurrence of introduced allochthons green algae belonging to the genus Caulerpa J.V. Lamourox, 1809 (Ceccherelli and Cinelli, 1999;Meinesz et al., 2001;Ceccherelli et al., 2002;Mollo et al., 2008;Jongma et al., 2013) and the native seagrass P. oceanica in the Mediterranean Sea provided an opportunity to explore the hypothesis that allelopathy influence interspecific competition. Among the bioactive algal metabolites that may exhibit allelopathic effects, the sesquiterpene caulerpenyne (CYN) and the bisindole alkaloid caulerpin (CAU) are particularly abundant in Caulerpa taxifolia (M.Vahl) C. Agardh, 1817 and Caulerpa cylindracea Sonder, 1845, respectively (Defranoux and Mollo, 2020;Mollo et al., 2015;Russo et al., 2023). These metabolites fulfill diverse functions in algal biology, such as serving in defense mechanisms and participating in signaling pathways (Dumay et al., 2002a;2002b). ...
... The digestive gland of bivalves is also commonly used for ecotoxicological studies (Bignell et al., 2008;Marigómez et al., 2013) because is the major organ for contaminants biotransformation (Livingstone et al., 2006). For the evaluation of histopathological alterations, histopathological indices (I h ) have been used in several studies assessing the effects of pollutants in mussels (El Ayari et al., 2024;Pinto et al., 2019;Russo et al., 2023;Tresnakova et al., 2023). The present results on I h for gills revealed that any of the tested treatments induced significant histopathological changes in mussels. ...
... Control food was prepared by soaking a combination of microalgae and probiotics (RotiBomb dry food, Algova) in acetone (Sigma-Aldrich, suitable for HPLC, ≥99.9 %) and then evaporating the organic solvent under reduced pressure, while treated food was made in the same manner but after dissolving caulerpin in an equal volume of acetone. The dose of CAU (1 mg/g dry food) was adopted according to Russo et al. (2023). This procedure led to a uniform mixture of CAU with the food since it is almost insoluble in water. ...
The invasive green algae Caulerpa taxifolia (M. Vahl) C. Agardh, 1817 and Caulerpa cylindracea Sonder, 1845 are
widely diffused in the Mediterranean Sea, where they compete for space with the endemic seagrass Posidonia
oceanica (Linnaeus) Delile, 1813, a keystone species in Mediterranean marine biodiversity. The present study
aims to explore the possible effects of bioactive metabolites from the invasive algae on the seagrass, which may
imply an allelopathic action. In particular, the study focuses on the effects of the algal alkaloid caulerpin and the
sesquiterpene caulerpenyne. Changes in leaf growth, chlorophyll content, and leaf protein expression of
P. oceanica genets under treatments were evaluated after 28 days of cultivation in mesocosms. Caulerpenyne
strongly inhibited the growth of adult leaves and the formation of new ones, while inducing the elongation of the
intermediate leaves and increased total chlorophyll content; on the contrary, caulerpin did not significantly
influence leaf growth and the formation of new ones. A total of 107 differentially accumulated proteins common
to the two treatments were also identified using the proteomic approach. Both molecules induced cells to
maintain homeostasis, enhancing the amino acid metabolism or fatty acid biosynthesis. Despite these disruptions,
P. oceanica demonstrated a more efficient response to stress induced by caulerpin, stimulating the
biosynthesis of essential amino acids to maintain cellular homeostasis and mitigate damage caused by reactive
oxygen species (ROS). Overall, obtained results supports the possible role of caulerpenyne, and not caulerpin, as
an effector in allelopathic interactions among invasive Caulerpa species and P. oceanica in the Mediterranean
Inflammation contributes to the pathogenesis of several diseases. Fenofibrate, known as a peroxisome proliferator-activated receptor - α (PPAR-α) agonist, is a classic drug for treating hyperlipidemia. In addition to its lipid-lowering effect, fenofibrate has also been reported to exert anti-inflammatory effects with complicated underlying mechanisms of action. In general, the anti-inflammatory effect of fenofibrate is secondary to its lipid-lowering effect, especially for the inflammation caused by hyperlipidemia in the circulatory system. Some anti-inflammatory actions may also come from its regulatory effects on intracellular lipid metabolism by activating PPAR-α. In addition, some roles in anti-inflammation might be mediated by its direct regulation of inflammatory signaling pathways. In order to understand anti-inflammatory activities and the underlying mechanisms of fenofibrate action in disease better, we herein reviewed and discussed the anti-inflammatory roles and its subserving mechanisms in various diseases of different organ systems. Thus, this review offers insights into the optimal use of fenofibrate in the clinical setting.
The invasive macroalga Caulerpa cylindracea has spread widely in the Mediterranean Sea, becoming a favorite food item for native fish for reasons yet unknown. By using a combination of behavioral, morphological, and molecular approaches, herein we provide evidence that the bisindole alkaloid caulerpin, a major secondary metabolite of C. cylindracea, significantly increases food intake in the model fish Danio rerio, influencing the regulation of genes involved in the orexigenic pathway. In addition, we found that the compound improves fish reproductive performance by affecting the hypothalamus–pituitary–gonadal axis. The obtained results pave the way for the possible valorization of C. cylindracea as a sustainable source of a functional feed additive of interest to face critical challenges both in aquaculture and in human nutrition.
Nuclear receptors (NRs) form a large family of ligand-dependent transcription factors that control the expression of a multitude of genes involved in diverse, vital biological processes[…]
The current perception that climate change is the principal threat to biodiversity is at best premature. Although highly relevant, it detracts focus and effort from the primary threats: habitat destruction and overexploitation. We collated causes of vertebrate extinctions since 1900, threat information for amphibia, birds, and mammals from the IUCN Red List, and scrutinized others’ attempts to compare climate change with commensurate anthropogenic threats. In each analysis, none of the arguments founded on climate change's wide-ranging effects are as urgent for biodiversity as those for habitat loss and overexploitation. Present conservation efforts must refocus on these issues. Conserving ecosystems by focusing on these major threats not only protects biodiversity but is the only available, economically viable, global strategy to reverse climate change.
Plastic pollution and invasive species are recognised as pervasive threats to marine biodiversity. However, despite the extensive on-going research on microplastics’ effects in the biota, knowledge on their combination with additional stressors is still limited. This study investigates the effects of polyamide microplastics (PA-MPs, 1 mg/L), alone and in combination with the toxic exudate from the invasive red seaweed Asparagopsis armata (2%), after a 96 h exposure, in the mussel Mytilus galloprovincialis. Biochemical responses associated with oxidative stress and damage, neurotoxicity, and energy metabolism were evaluated in different tissues (gills, digestive gland, and muscle). Byssus production and PA-MP accumulation were also assessed. Results demonstrated that PA-MPs accumulated the most in the digestive gland of mussels under PA-MP and exudate co-exposure. Furthermore, the combination of stressors also resulted in oxidative damage at the protein level in the gills as well as in a significant reduction in byssus production. Metabolic capacity increased in both PA-MP treatments, consequently affecting the energy balance in mussels under combined stress. Overall, results show a potential increase of PA-MPs toxicity in the presence of A. armata exudate, highlighting the importance of assessing the impact of microplastics in realistic scenarios, specifically in combination with co-occurring stressors, such as invasive species.
Mercury (Hg) is one of the most hazardous pollutants, due to its toxicity, biological magnification and worldwide persistence in aquatic systems. Thus, new efficient nanotechnologies (e.g. graphene oxide functionalized with polyethyleneimine (GO-PEI)) have been developed to remove this metal from the water. Aquatic environments, in particular transitional systems, are also subjected to disturbances resulting from climate change, such as salinity shifts. Salinity is one of the most relevant factors that influences the distribution and survival of aquatic species such as mussels. To our knowledge, no studies assessed the ecotoxicological impairments induced in marine organisms exposed to remediate seawater (RSW) under different salinity levels. For this, the focus of the present study was to evaluate the effects of seawater previously contaminated with Hg and remediated with GO-PEI, using the species Mytilus galloprovincialis, maintained at three different salinities (30, 20 and 40). The results obtained demonstrated similar histopathological and metabolic alterations, oxidative stress and neurotoxicity in mussels under RSW treatment at stressful salinity conditions (20 and 40) in comparison to control salinity (30). On the other hand, the present findings revealed toxicological effects including cellular damage and histopathological impairments in mussels exposed to Hg contaminated seawater in comparison to non-contaminated ones, at each salinity level. Overall, these results confirm the high efficiency of GO-PEI to sorb Hg from water with no noticeable toxic effects even under different salinities, leading to consider it a promising eco-friendly approach to remediate contaminated water.
The identification of novel strategies to control Helicobacter pylori (Hp)-associated chronic inflammation is, at present, a considerable challenge. Here, we attempt to combat this issue by modulating the innate immune response, targeting formyl peptide receptors (FPRs), G-protein coupled receptors that play key roles in both the regulation and the resolution of the innate inflammatory response. Specifically, we investigated, in vitro, whether Caulerpin—a bis-indole alkaloid isolated from algae of the genus Caulerpa—could act as a molecular antagonist scaffold of FPRs. We showed that Caulerpin significantly reduces the immune response against Hp culture filtrate, by reverting the FPR2-related signaling cascade and thus counteracting the inflammatory reaction triggered by Hp peptide Hp(2–20). Our study suggests Caulerpin to be a promising therapeutic or adjuvant agent for the attenuation of inflammation triggered by Hp infection, as well as its related adverse clinical outcomes.
Inosine monophosphate (IMP) is the intracellular precursor for both adenosine monophosphate and guanosine monophosphate and thus plays a central role in intracellular purine metabolism. IMP can also serve as an extracellular signaling molecule, and can regulate diverse processes such as taste sensation, neutrophil function, and ischemia‐reperfusion injury. How IMP regulates inflammation induced by bacterial products or bacteria is unknown. In this study, we demonstrate that IMP suppressed tumor necrosis factor (TNF)‐α production and augmented IL‐10 production in endotoxemic mice. IMP exerted its effects through metabolism to inosine, as IMP only suppressed TNF‐α following its CD73‐mediated degradation to inosine in lipopolysaccharide‐activated macrophages. Studies with gene targeted mice and pharmacological antagonism indicated that A2A, A2B, and A3 adenosine receptors are not required for the inosine suppression of TNF‐α production. The inosine suppression of TNF‐α production did not require its metabolism to hypoxanthine through purine nucleoside phosphorylase or its uptake into cells through concentrative nucleoside transporters indicating a role for alternative metabolic/uptake pathways. Inosine augmented IL‐β production by macrophages in which inflammasome was activated by lipopolysaccharide and ATP. In contrast to its effects in endotoxemia, IMP failed to affect the inflammatory response to abdominal sepsis and pneumonia. We conclude that extracellular IMP and inosine differentially regulate the inflammatory response.
The purification of homogeneous glutathione S-transferases B and C from rat liver is described. Kinetic and physical properties of these enzymes are compared with those of homogeneous transferases A and E. The letter designations for the transferases are based on the reverse order of elution from carboxymethylcellulose, the purification step in which the transferases are separated from each other. Transferase B was purified on the basis of its ability to conjugate iodomethane with glutathione, whereas transferase C was purified on the basis of conjugation with 1,2-dichloro-4-nitrobenzene. Although each of the four enzymes can be identified by its reactivity with specific substrates, all of the enzymes are active to differing degrees in the conjugation of glutathione with p-nitrobenzyl chloride. Assay conditions for a variety of substrates are included.
All four glutathione transferases have a molecular weight of 45,000 and are dissociable into subunits of approximately 25,000 daltons. Despite the similar physical properties and overlapping substrate specificities of these enzymes, only transferases A and C are immunologically related.
Peroxisome proliferator-activated receptor gamma (PPARγ), belonging to the nuclear receptor superfamily, is a ligand-dependent transcription factor involved in a variety of pathophysiological conditions such as inflammation, metabolic disorders, cardiovascular disease, and cancers. In this latter context, PPARγ is expressed in many tumors including breast cancer, and its function upon binding of ligands has been linked to the tumor development, progression, and metastasis. Over the last decade, much research has focused on the potential of natural agonists for PPARγ including fatty acids and prostanoids that act as weak ligands compared to the strong and synthetic PPARγ agonists such as thiazolidinedione drugs. Both natural and synthetic compounds have been implicated in the negative regulation of breast cancer growth and progression. The aim of the present review is to summarize the role of PPARγ activation in breast cancer focusing on the underlying cellular and molecular mechanisms involved in the regulation of cell proliferation, cell cycle, and cell death, in the modulation of motility and invasion as well as in the cross-talk with other different signaling pathways. Besides, we also provide an overview of the in vivo breast cancer models and clinical studies. The therapeutic effects of natural and synthetic PPARγ ligands, as antineoplastic agents, represent a fascinating and clinically a potential translatable area of research with regards to the battle against cancer.
Marine organisms are frequently exposed to pollutants, including trace metals, derived from natural and anthropogenic activities. In order to prevent environmental pollution, different approaches have been applied to remove pollutants from waste water and avoid their discharge into aquatic systems. However, organisms in their natural aquatic environments are also exposed to physico-chemical changes derived from climate change-related factors, including temperature increase. According to recent studies, warming has a negative impact on marine wildlife, with known effects on organisms physiological and biochemical performance. Recently, a material based on graphene oxide (GO) functionalized with polyethyleneimine (PEI) proved to be effective in the remediation of mercury (Hg) contaminated water. Nevertheless, no information is available on the toxic impacts of such remediated water towards aquatic systems, neither under actual nor predicted temperature conditions. For this, the present study assessed the toxicity of seawater, previously contaminated with Hg and remediated by GO-PEI, using the clam species Ruditapes philippinarum exposed to actual and a predicted temperature conditions. The results obtained demonstrated that seawater contaminated with Hg and/or Hg+GO-PEI induced higher toxicity in clams exposed to 17 and 22 °C compared to organisms exposed to remediated seawater at the same temperatures. Moreover, similar histological and biochemical results were observed between organisms exposed to control and remediated seawater, independently of the temperatures (17 and 21 °C), highlighting the potential use of GO-PEI to remediate Hg from seawater without significant toxicity issues to the selected marine species.
In the present study, the occurrence of 40 pharmaceuticals belonging to several therapeutic groups was investigated for the first time in hospital effluent, wastewater treatment plant influent and effluent, and seawater in Mahdia, Tunisia. Forty-six samples were collected within a 6-month sampling period. Pharmaceuticals were analyzed using solid-phase extraction followed by ultra-performance liquid chromatography-triple quadrupole mass spectrometry. Thirty-three out of the forty target compounds were detected over a wide concentration of ranges, from nanograms per liter to micrograms per liter, depending on the type of sample. Maximum values were detected for caffeine at 902 μgL⁻¹ in hospital wastewater. This compound, as well as salicylic acid, sulfadiazine, and sulfamethizole, were detected in all samples. The average concentration of total pharmaceuticals in hospital wastewater (340 μgL⁻¹) was higher than those detected in influent and effluent wastewater and seawater (275.11 and 0.2 μgL⁻¹, respectively). Risk quotients (RQs) were also estimated to provide a preliminary environmental risk assessment and results revealed that sulfadiazine, sulfamethoxazole, and fluoxetine could pose medium/high risk to the tested aquatic organisms for maximum measured concentrations in wastewater (including hospital and WWTP samples). Although the measured environmental concentrations (MECs) detected in seawater samples might not pose a toxic effect to the aquatic organisms (except for salicylic acid, sulfamethoxazole and fluoxetine), further researches are needed due to the continuous release of wastewater in the environment and the limited efficiency of wastewater treatment processes.
Due to widespread occurrence of lipid lowering drugs such as statins, fibrates and their metabolites in the aquatic environments, there is a worldwide growing concern in their role in water quality and aquatic biota. However, this concern is limited by ability to address their occurrence, distribution, fate and eco-toxicological effects. This study focuses on the quantification of the levels of statins, fibrates and their metabolites in the aquatic environments using Ultra-High Performance Liquid Chromatography coupled to high resolution quadrupole time-of-flight mass spectrometry (UHPLC-QTOF-MS). The developed UHPLC–QTOF–MS based method was successfully applied to the analysis of statins, fibrates and metabolites in real water samples collected from Daspoort WWWs influent and effluent and Apies River. A series of statin compounds (mevastatin, simvastatin, pravastatin, rosuvastatin, fluvastatin, atorvastatin), fibrates (gemfibrozil, fenofibrate) and the corresponding metabolites (clofibric and fenofibric acids) were detected and quantified in the range of 0.56–19.90 µg/L in both waste and River water samples. In general, the results of the present study are an indication of pollution hazards from wastewater treatment processes and these levels poses a huge risk to the growth and reproduction of aquatic organisms. Thus, regulating the limit levels of statins, fibrates and metabolites in any type of water is paramount as it will provide the vital information on the toxic risks associated with organic pollutants of pharmaceutical origin. Keywords: Aquatic ecosystem, Cholesterol/lipid lowering drugs, UHPLC-QTOF-MS, Eco-toxicological, Spatial distribution
Recent studies have suggested that Mediterranean indigenous fish species are affected by bioactive metabolites coming from marine invasive species via food web interactions. In particular, both physiological and behavioural changes in the white sea bream Diplodus sargus were related to caulerpin (CAU), a bisindolic alkaloid particularly abundant in the invasive alga Caulerpa cylindracea, on which the fish actively feed. Dietary administration of CAU decreased aggressiveness in D. sargus, suggesting an anxiolytic-like effect of CAU possibly mediated by endogenous anxiolytic agents. This hypothesis is supported here by the finding of a significant increase of NPY transcriptional expression in the brain of fish fed with CAU enriched food, shedding more light on the neural mechanisms behind the altered behaviour of D. sargus.
Although the chemical warfare between invasive and native species has become a central problem in invasion biology, the molecular mechanisms by which bioactive metabolites from invasive pests influence local communities remain poorly characterized. This study demonstrates that the alkaloid caulerpin (CAU)—a bioactive component of the green alga Caulerpa cylindracea that has invaded the entire Mediterranean basin—is an agonist of peroxisome proliferator-activated receptors (PPARs). Our interdisciplinary study started with the in silico prediction of the ligand-protein interaction, which was then validated by in vivo, ex vivo and in vitro assays. On the basis of these results, we candidate CAU as a causal factor of the metabolic and behavioural disorders observed in Diplodus sargus, a native edible fish of high ecological and commercial relevance, feeding on C. cylindracea. Moreover, given the considerable interest in PPAR activators for the treatment of relevant human diseases, our findings are also discussed in terms of a possible nutraceutical/pharmacological valorisation of the invasive algal biomasses, supporting an innovative strategy for conserving biodiversity as an alternative to unrealistic campaigns for the eradication of invasive pests.
The biological invasion of the green algae Caulerpa cylindracea represents a serious scientific and public issue in the Mediterranean Sea, essentially due to strong modifications both to habitat structure and native benthic communities. Although alterations in health status and changes in flesh quality of some marine species (dietary exposed to C. cylindracea) have been observed, no studies on cause-effect relationships have been carried out. Here, for the first time, through a controlled feeding experiment followed by ¹H NMR Spectroscopy and multivariate analysis (PCA, OPLS-DA), we showed that caulerpin taken with diet is directly responsible of changes observed in metabolic profile of fish flesh, including alteration of lipid metabolism, in particular with a reduction of ω3 PUFA content. The potential of caulerpin to directly modulate lipid metabolism opens up new questions about causal mechanism triggered by algal metabolite also in view of a possible exploitation in the nutraceutical/medical field.
Cancer is a leading cause of death throughout the world, and cancer therapy remains a big medical challenge in terms of both its therapeutic efficacy and safety. Therefore, to find out a safe anticancer drug has been long goal for oncologist and medical scientists. Among clinically used medicines with no or little toxicity, fenofibrate is a drug of the fibrate class that plays an important role in lowering the levels of serum cholesterol and triglycerides while elevating the levels of high-density lipoproteins. Recently, several studies have implied that fenofibrate may exert anticancer effects via a variety of pathways involved in apoptosis, cell-cycle arrest, invasion, and migration. Given the great potential that fenofibrate may have anticancer effects, this review was to investigate all published works which directly or indirectly support the anticancer activity of fenofibrate. These studies provide evidence that fenofibrate exerted antitumor effects in several human cancer cell lines, such as breast, liver, glioma, prostate, pancreas, and lung cancer cell lines. Among these studies some have further confirmed the possibility and efficacy of fenofibrate anticancer in xenograft mouse models. In the last part of this review, we also discuss the potential mechanisms of action of fenofibrate based on the available information. Overall, we may repurpose fenofibrate as an anticancer drug in cancer treatment, which urgently need further and comprehensively investigated.
Intertidal species face multiple stressors on a daily basis due to their particular habitat. The submergence at high tide in the aquatic environment and emergence at low tide to the aerial environment, associated with a wide variation of abiotic parameters, along with anthropogenic contamination are some of the daily stresses that these organisms are exposed to. With such a dynamic environment, organisms developed strategies that allow them to avoid or tolerate these stressors. Among these species, bivalves are some of the most hypoxia tolerant, being commonly used as a biomonitoring tool due to their capacity to accumulate pollutants from the environment and reflect the imposed toxic impacts. However, when evaluating the response ability of organisms to different stressors under laboratory conditions, it is not common to consider the fact that exposure to tides can act as a confounding factor. The present study assessed the effects of air exposure on the biochemical (metabolic capacity, energy reserves, and oxidative stress related biomarkers) performance of intertidal Mytilus galloprovincialis mussels. Specimens of M. galloprovincialis were submitted once every 24 h to different periods of air exposure (3 and 6 h) for 14 days, under constant air and seawater temperature (19 ± 1 °C). Results obtained revealed that air exposure can cause biochemical changes in mussels. The present findings demonstrated that individuals exposed to air induced superoxide dismutase (SOD) and catalase (CAT) activity as mechanisms to withstand the abiotic changes while mobilizing lipid content as the principal source of energy, and increasing protein content possibly as a result of an increase in the number of antioxidant defense enzymes. Moreover, individuals under air exposure suffered higher oxidative damage while showing higher metabolic rate. Results demonstrated that longer periods of air exposure induced more injuries, since individuals emerged during 6 h presented higher oxidative stress than individuals under 3 h of air exposure.
The invasive green alga Caulerpa cylindracea has become an important component of the diet of the Mediterranean white seabream Diplodus sargus. As a consequence of this “exotic diet”, the algal bisindolic alkaloid caulerpin accumulates in the fish tissues. Although the compound shows structural similarity to endogenous indolamines that modulate animal behaviour, the potential impact of caulerpin on fish behaviour still remains unexplored. In this report, behavioural experiments both on groups and on single fish responding towards a mirror were performed under different doses of dietary caulerpin. Differences between treated and control groups for each behaviour and for the overall aggressive pattern during the different experimental phases showed that the aggressiveness of D. sargus decreased with the administration of caulerpin. These results call the attention to a still unexplored potential ability of bioactive metabolites from marine invasive species, to alter the behaviour on native species, with putative negative effects on patterns of fish growth and population dynamics.
In the process of a paradigm shift in toxicity testing, many efforts have focused on how to integrate and interpret information for biological events occurring at molecular and cellular level to be predictive of adverse effects at organism or population level to be useful, for example, for regulatory decision-making. The adverse outcome pathway (AOP) concept provides such a framework of knowledge-based safety assessment of chemicals that links mechanistic information with an apical endpoint. Here we outline an AOP that links the activation of peroxisome proliferator-activated receptor α (PPARα) to reproductive tract malformations and impaired fertility in males. The development of this AOP relies on evidence collected from rodent models and incorporates human mechanistic and epidemiological data. Interest in PPARα action as a mechanistic basis for effects on the reproductive system arises from the relationships between activation of this receptor and impairment of steroidogenesis leading to reproductive toxicity. The PPARα-initiated AOP is a first step for structuring current knowledge about mode of action (MoA), which is neither androgen receptor-mediated nor via direct aromatase inhibition. In the current form, the pathway lays a strong basis for linking an endocrine MoA with an adverse effect, a prerequisite requirement for the identification of endocrine disrupting chemicals.
Fibrates, which are widely used lipidaemic-modulating drugs, are emerging environmental pollutants. However, fibrate concentrations in the environment have not been thoroughly surveyed. Here, we determined concentrations of the most commonly used fibrates and their metabolites in source water and drinking water samples from ten drinking water treatment plants in Shanghai and Zhejiang, China, using solid-phase extraction and liquid chromatography–tandem mass spectrometry. All the target compounds were detected in at least some of the source water samples, at concentrations ranging from 0.04 ng/L (fenofibrate) to 1.53 ng/L (gemfibrozil). All the compounds except fenofibrate were also detected in at least some of the drinking water samples, at recoveries ranging from 35.5% to 91.7%, suggesting that these compounds are poorly removed by typical drinking water treatment processes. In a peroxisome proliferator-activated receptor α agonistic activity assay, the target compounds showed no significant activity at nanogram per litre concentrations; therefore, our results suggest that the fibrate concentrations in drinking water in Shanghai and Zhejiang, China do not significantly affect human health. However, because of the increasing westernization of the Chinese diet, fibrate use may increase, and thus monitoring fibrate concentrations in aquatic environments and drinking water in China will become increasingly important.
Recent studies have showed that the Mediterranean white sea bream Diplodus sargus has included the invasive green alga Caulerpa cylindracea in its diet. As a result of the novel alimentary habits, the fish accumulates the bioactive algal red pigment caulerpin in its tissues with consequent metabolic and enzymatic alterations. However, this may not be an isolated case: other fish species have been described to feed on C. cylindracea, while the possible accumulation of caulerpin in their tissues were never investigated before. In this report, we analysed stomach contents and caulerpin levels in the native sparid species Spondyliosoma chantarus, Sarpa salpa, and Diplodus vulgaris, and in the scarid Sparisoma cretense, in comparison with the lessepsian siganid Siganus luridus. C. cylindracea was found in the stomachs of all studied fish species, with the only exception of S. cretense, where prey items could not be determined due to the high degree of digestion. Chemical analysis on fish tissues revealed that only S. cantharus and S. salpa accumulated caulerpin, while no traces of the compound were detected in the other species. Despite of intense research efforts on natural products from C. cylindracea, a complete picture of the impact caused by the entering of this alga in the fish diet is far to be elucidated. The identification of the accumulation of caulerpin in other Mediterranean native fish, suggests the need to further research, also in order to assess the possible transfer of such molecules to humans through seafood consumption.
Cadmium (Cd) is one of the most important metal contaminants in the Bohai Sea. In this work, NMR-based metabolomics was used to investigate the toxicological effects of Cd at an environmentally relevant concentration (50 µg L(-1)) in three different life stages (D-shape larval, juvenile and adult) of mussels Mytilus galloprovincialis. Results indicated that the D-shape larval mussel was the most sensitive life stage to Cd. The significantly different metabolic profiles meant that Cd induced differential toxicological effects in three life stages of mussels. Basically, Cd caused osmotic stress in all the three life stages via different metabolic pathways. Cd exposure reduced the anaerobiosis in D-shape larval mussels and disturbed lipid metabolism in juvenile mussels, respectively. Compared with the D-shape larval and juvenile mussels, the adult mussels reduced energy consumption to deal with Cd stress.
Peroxisome proliferator-activated receptor alpha (PPARα) is a nuclear hormone receptor involved in the transcriptional regulation of lipid metabolism, fatty acid oxidation, and glucose homeostasis. Its activation stimulates antioxidant enzymes such as catalase, whose expression is decreased in aged human skin. Here we investigated the expression of PPARα in aged and ultraviolet (UV)-irradiated skin, and whether PPARα activation can modulate expressions of matrix metalloproteinase (MMP)-1 and procollagen through catalase regulation. We found that PPARα mRNA level was significantly decreased in intrinsically aged and photoaged human skin as well as in UV-irradiated skin. A PPARα activator, Wy14643, inhibited UV-induced increase of MMP-1 and decrease of procollagen expression and caused marked increase in catalase expression. Furthermore, production of reactive oxygen species (ROS) was suppressed by Wy14643 in UV-irradiated and aged dermal fibroblasts, suggesting that the PPARα activation-induced upregulation of catalase leads to scavenging of ROS produced due to UV irradiation or aging. PPARα knockdown decreased catalase expression and abolished the beneficial effects of Wy14643. Topical application of Wy14643 on hairless mice restored catalase activity and prevented MMP-13 and inflammatory responses in skin. Our findings indicate that PPARα activation triggers catalase expression and ROS scavenging, thereby protecting skin from UV-induced damage and intrinsic aging.
Publisher Summary Glutathione reductase is a flavoprotein catalyzing the NADPH-dependent reduction of glutathione disulfide (GSSG) to glutathione (GSH). The reaction is essential for the maintenance of glutathione levels. Glutathione has a major role as a reductant in oxidation–reduction processes, and serves in detoxication and several other cellular functions of great importance. A purification method of this enzyme from calf liver and rat liver is described in this chapter. Similar methods are used for the purification of the enzyme from yeast, porcine, and human erythrocytes. All the steps are carried out at about 5 ° . The purification method from calf liver consists of various steps including preparation of cytosol fraction, chromatography on DEAE-sephadex, precipitation with ammonium sulfate, and chromatography on hydroxyapatite. The purification of glutathione reductase from rat liver is usually combined with the preparation of glutathione transferases, thioltransferase, and glyoxalase I.
Inflammation contributes to the pathogenesis of several diseases. Fenofibrate, known as a peroxisome proliferator-activated receptor - α (PPAR-α) agonist, is a classic drug for treating hyperlipidemia. In addition to its lipid-lowering effect, fenofibrate has also been reported to exert anti-inflammatory effects with complicated underlying mechanisms of action. In general, the anti-inflammatory effect of fenofibrate is secondary to its lipid-lowering effect, especially for the inflammation caused by hyperlipidemia in the circulatory system. Some anti-inflammatory actions may also come from its regulatory effects on intracellular lipid metabolism by activating PPAR-α. In addition, some roles in anti-inflammation might be mediated by its direct regulation of inflammatory signaling pathways. In order to understand anti-inflammatory activities and the underlying mechanisms of fenofibrate action in disease better, we herein reviewed and discussed the anti-inflammatory roles and its subserving mechanisms in various diseases of different organ systems. Thus, this review offers insights into the optimal use of fenofibrate in the clinical setting.
The increasing use of rare earth elements (REEs) in electric and electronic equipment has been associated with the presence of these elements in aquatic systems. The present study aimed to evaluate the toxicity of two REEs, Lanthanum (La) and Gadolinium (Gd), towards the mussel species Mytilus galloprovincialis. For this, the toxicity was assessed after a short-term exposure (14 days) to an environmentally relevant concentration of each element (10 μg/L), followed by a recovery period (14 days) in the absence of any contaminant. The measured biomarkers included energy-related parameters, activity of antioxidant and biotransformation enzymes, indicators of oxidative damage, levels of oxidized glutathione and neurotoxicity. After exposure mussels accumulated more La (0.54 μg/g) than Gd (0.15 μg/g). After recovery higher concentration decrease was observed for Gd (≈40% loss) compared to La exposed mussels (≈30% loss) which may be associated with lower detoxification capacity of mussels previously exposed to La. Mussels increased their metabolism (i.e., higher electron transport system activity) only after the exposure to Gd. Exposure to La and Gd resulted into lower energy expenditure, while when both elements were removed glycogen and protein concentrations decreased to values observed in non-contaminated mussels. Antioxidant and biotransformation capacity was mainly increased in the presence of Gd. This defense response avoided the occurrence of cellular damage but still loss of redox balance was found regardless the contaminant, which was re-established after the recovery period. Neurotoxicity was only observed in the presence of Gd with no effects after the recovery period. Results showed that a short-term exposure to La and especially to Gd can exert deleterious effects that may compromise specific biochemical pathways in aquatic species, such as M. galloprovincialis, but under low concentrations organisms can be able to re-establish their biochemical status to control levels after a recovery period.
Peroxisome proliferator activated receptors (PPAR) are kinds of key transcriptional factors in regulating LC-PUFA biosynthesis. Until now, little is known about PPAR in marine molluscs as well as in other invertebrates. Sinonovacula constricta is the first marine mollusc that is proved to possess the complete LC-PUFA biosynthetic pathway, and it can be a perfect representative to clarify this situation. In this study, the molecular properties of S. constricta PPAR were characterized, and corresponding potential regulatory roles in S. constricta Δ6 fatty acyl desaturase (Fad) transcription were estimated by dual luciferase assay. Results showed that two PPAR homologs (PPAR_a and _b) were identified in S. constricta. They both contain typical features of vertebrate PPAR, suggesting highly conserved functional regions in PPAR. By phylogenetic comparison, they are clearly different with vertebrate PPAR and can be divided into two distinctive sub-groups. Moreover, they show a high expression level in gill, labial palps, mantle and intestine. Their down-regulated expressions in trochophore larva and veliger larva might be attributed to food-deprivation. Additionally, the transcriptional activity of S. constricta Δ6 Fad promoter was significantly activated by both PPAR_a and _b, indicating that S. constricta PPAR might play a role in the regulation of LC-PUFA biosynthesis. To our knowledge, this is the first systematic analysis of PPAR in a marine mollusc. The results will provide a valuable reference for further researches on the function of marine molluscan PPAR and their underlying mechanisms in regulating LC-PUFA biosynthesis.
A simple, rapid UHPLC‐MS/MS method has been developed and optimised for the quantitation of a range of pharmaceuticals, metabolites and related bioactive compounds in the bivalve mollusc species mussels (Mytilus edulis) and Pacific oysters (Crassostrea gigas). Shellfish tissues were extracted using a simple solvent‐based extraction method prior to concentration and purification by pass‐through SPE and quantified using stable isotope dilution tandem mass spectrometry. The analytes covered a range of therapeutic classes including antidepressants, anticonvulsants, beta‐blockers and antiplatelets. Of the 34 compounds which were included in the study initially, 28 compounds were found to demonstrate acceptable performance. Performance was assessed by examining extraction efficiencies, matrix effects, sensitivity, and within‐ and between‐batch precision. The results presented here show that as indicated by acceptable HorRat and accuracy values the method is fit for purpose. Application of this method to environmental mussel and oyster samples revealed the presence of 12 compounds at quantifiable concentrations, with the antidepressant sertraline being present at the highest level, reaching a concentration of 6.12 ng/g in mussel tissue. This article is protected by copyright. All rights reserved.
A wide variety of active pharmaceutical ingredients are released into the environment and pose a threat to aquatic organisms. Drug products using micro- and nanoparticle technology can lower these emissions into the environment by their increased bioavailability to the human patients. However, due to this enhanced efficacy, micro- and nanoscale drug delivery systems can potentially display an even higher toxicity, and thus also pose a risk to non-target organisms. Fenofibrate is a lipid-regulating agent and exhibits species-related hazards in fish. The ecotoxic effects of a fenofibrate formulation embedded into a hydroxypropyl methylcellulose microparticle matrix, as well as those of the excipients used in the formulation process, were evaluated. To compare the effects of fenofibrate without a formulation, fenofibrate was dispersed in diluted ISO water alone or dissolved in the solvent DMF and then added to diluted ISO water. The effects of these various treatments were assessed using the fish embryo toxicity test, acridine orange staining and gene expression analysis assessed by quantitative RT polymerase chain reaction. Exposure concentrations were assessed by chemical analysis. The effect threshold concentrations of fenofibrate microparticle precipitates were higher compared to the formulation. Fenofibrate dispersed in 20%-ISO-water displayed the lowest toxicity. For the fenofibrate formulation as well as for fenofibrate added as a DMF solution, greater ecotoxic effects were observed in the zebrafish embryos. The chemical analysis of the solutions revealed that more fenofibrate was present in the samples with the fenofibrate formulation as well as fenofibrate added as a DMF solution compared to fenofibrate dispersed in diluted ISO water. This could explain the higher ecotoxicity. The toxic effects on the zebrafish embryo thus suggested that the formulation as well as the solvent increased the bioavailability of fenofibrate.
Nowadays, functionalized multi-walled carbon nanotubes (f-MWCNTs) are considered to be emerging contaminants and their impact in ecosystem has rose recent concerns, while other contaminants, such as caffeine, have more coverage in literature. Despite this, the effects of a combination of the two has yet to be evaluated, especially considering predicted temperature rise. In the present study a typical bioindicator species for marine environment, the clam Ruditapes decussatus, and classical tools, such as biomarkers and histopathological indices, were used to shed light on species’ response to these contaminants, under actual and predicted warming scenarios. The results obtained showed that both contaminants have a harmful effect at tissue level, as shown by the higher histopathological index, especially in digestive tubules. Temperatures seemed to induce greater biochemical impacts than caffeine and f-MWCNTs when acting alone, namely in terms of antioxidant defences and energy reserves content, which were exacerbated when both contaminants were acting in combination (MIX treatment). Overall, the present findings highlight the complex response of clams to both pollutants, evidencing the role of temperature on clams’ sensitivity, especially to mixture of pollutants.
In the marine environment, organisms are exposed to a high and increasing number of different pollutants that can interact among them. In addition, abiotic factors can change the dynamics between contaminants and organisms, thus increasing or even decreasing the toxic effect of a particular compound. In this study, the effects of caffeine and functionalized multi-walled carbon nanotubes (f-MWCNTs) induced in the clam Ruditapes philippinarum were evaluated, acting alone and in combination, under two temperature levels (18 ºC and 21 ºC). To assess the impact of such compounds, their interaction and the possible influence of temperature, biochemical and histopathological markers were investigated. The effects of f-MWCNTs and caffeine appear to be clearly negative at the control temperature, with lower protein content in contaminated clams and a significant decrease in their metabolism when both pollutants were acting in combination. Also, at control temperature, clams exposed to pollutants showed increased antioxidant capacity, especially when caffeine was acting alone, although cellular damages were still observed at CAF and f-MWCNTs treatments. Increased biotransformation capacity at 18 ºC and MIX treatment may explain lower caffeine concentration observed. At increased temperature differences among treatments were not so evident as at 18 ºC, with a similar biological pattern among contaminated and control clams. Higher caffeine accumulation at MIX treatment under warming conditions may result from clams’ inefficient biotransformation capacity when exposed to increased temperatures.
Today, environmental pollution with pharmaceutical drugs and their metabolites poses a major threat to the aquatic ecosystems. Active substances such as fenofibrate, are processed to pharmaceutical drug formulations before they are degraded by the human body and released into the wastewater. Compared to the conventional product Lipidil® 200, the pharmaceutical product Lipidil 145 One® and Ecocaps take advantage of nanotechnology to improve uptake and bioavailability of the drug in humans.In the present approach, a combination of in vitro drug release studies and physiologically-based biopharmaceutics modeling was applied to calculate the emission of three formulations of fenofibrate (Lipidil® 200, Lipidil 145 One®, Ecocaps) into the environment. Special attention was paid to the metabolized and non-metabolized fractions and their individual toxicity, as well as to the emission of nanomaterials. The fish embryo toxicity test revealed a lower aquatic toxicity for the metabolite fenofibric acid and therefore an improved toxicity profile. When using the microparticle formulation Lipidil® 200, an amount of 126 mg of non-metabolized fenofibrate was emitted to the environment. Less than 0.05% of the particles were in the lower nanosize range. For the nanotechnology-related product Lipidil 145 One®, the total drug emission was reduced by 27.5% with a nanomaterial fraction of approximately 0.5%. In comparison, the formulation prototype Ecocaps reduced the emission of fenofibrate by 42.5% without any nanomaterials entering the environment. In a streamlined life cycle assessment, the lowered dose in combination with a lowered drug-to-metabolite ratio observed for Ecocaps led to a reduction of the full life cycle impacts of fenofibrate with a reduction of 18% reduction in the global warming potential, 61% in ecotoxicity, and 15% in human toxicity. The integrated environmental assessment framework highlights the outstanding potential of advanced modeling technologies to determine environmental impacts of pharmaceuticals during early drug development using preclinical in vitro data.
Due to their widespread use, pharmaceuticals can be metabolized, excreted and ultimately discarded in the environment, thereby affecting aquatic organisms. Lipidregulating drugs are one of the most prescribed medications around the world,
controlling human cholesterol levels, in more than 20 million patients. Despite this
growing use of lipid-regulating drugs, particularly those whose active metabolite is
clofibric acid, the potential toxicological effects of these pharmaceuticals in the
environment is not fully characterized. This work intended to characterize the toxicity
of an acute (120 hours post-fertilization) and chronic (60 days post-fertilization)
exposures to clofibric acid in concentrations of 10.35, 20.7, 41.4, 82.8 and 165.6 μg L-1
in zebrafish (Danio rerio). The concentrations which were implemented in both
exposures were based on predicted environmental concentrations for Portuguese surface
waters. The acute effects were analysed focusing on behavioural endpoints (small and
large distance travelled, swimming time and total distance travelled), biomarkers of
oxidative stress (activity of the enzymes superoxide dismutase, Cu/Zn- and Mn SOD;
catalase, CAT; glutathione peroxidase, Se- and total GPx), biotransformation (activity
of glutathione S-transferases, GSTs) and lipid peroxidation (thiobarbituric acid reactive
substances, TBARS). Chronically exposed individuals were also histologically analysed
for sex determination and gonadal developmental stages. In terms of acute exposure,
significant alterations were reported, in terms of behavioural alterations (hypoactivity),
followed by an overall increase in all tested biomarkers. Chronically exposed organisms
did not show alterations in terms of sex ratio and maturation stages, suggesting that
clofibric acid did not act as an endocrine disruptor. Moreover, the metabolism of
clofibric acid resulted in increased levels of both forms of SOD activity, especially for
animals exposed to higher levels of this drug. An increase of CAT activity was observed
in fish exposed to low levels, and a decrease in those exposed to higher amounts of
clofibric acid. Both GPx forms had their activities increased. The enzyme of
biotransformation GSTs were increased at low levels of clofibric acid but inhibited at
higher amounts of this substance. Lipid peroxidation levels were also changed, with an
induction of this parameter with increasing amounts of clofibric acid. Changes also
occurred in behavioural endpoints and patterns for control organisms and for those
exposed to clofibric acid were significantly distinct, for all types (light and darkness) of
exposure, and for the two analysed endpoints (small and large distance). Results from
this assay allow inferring that clofibric acid can have an ecologically relevant impact in living organisms exposed to this substance, with putative impact on the metabolism of
individuals, affecting their behaviour and ultimately their survival.
The modern technology brought new engineering materials (e.g. nanostructured materials) with advantageous characteristics such as a high capacity to decontaminate water from pollutants (for example metal(loid)s). Among those innovative materials the synthesis of nanostructured materials (NSMs) based on graphene as graphene oxide (GO) functionalized with polyethyleneimine (GO-PEI), had a great success due to their metal removal capacity from water. However, research dedicated to environmental risks related to the application of these materials is still non-existent. To evaluate the impacts of such potential stressors, benthic species can be a good model as they are affected by several environmental constraints. Particularly, the mussel Mytilus galloprovincialis has been identified by several authors as a bioindicator that respond quickly to environmental disturbances, with a wide spatial distribution and economic relevance. Thus, the present work aimed to evaluate the impacts caused in M. galloprovincialis by seawater previously contaminated by Hg and decontaminated using GO-PEI. For this, histopathological and biochemical alterations were examined. This study demonstrated that mussels exposed to the contaminant (Hg), the decontaminant (GO-PEI) and the combination of both (Hg + GO-PEI) presented an increment of histopathological, oxidative stress and metabolic alterations if compared to organisms under remediated seawater and control conditions The present findings highlight the possibility to remediate seawater with nanoparticles for environmental safety purposes.
Mussels, such as the marine bivalve Mytilus galloprovincialis are sentinels for marine pollution but they are also excellent bioindicators under laboratory conditions. For that, in this study we tested the modulation of biochemical responses under realistic concentrations of the toxic metal Lead (Pb) in water for 28 days under different conditions of salinity and temperature, including control condition (temperature 17 ± 1.0 °C and salinity 30 ± 1.0) as well as those within the range expected to occur due to climate change predictions (± 5 in salinity and + 4 °C in temperature). A comprehensive set of biomarkers was applied to search on modulation of biochemical responses in terms of energy metabolism, energy reserves, oxidative stress and damage occurrence in lipids, proteins as well as neurotoxicity signs. The application of an integrative Principal Coordinates Ordination (PCO) tool was successful and demonstrated that Pb caused an increase in the detoxification activity mainly evidenced by glutathione S-transferases and that the salinities 25 and 35 were, even in un-exposed mussels, responsible for cell damage seen as increased levels of lipid peroxidation (at salinity 25) and oxidised proteins (at salinity 35).
Carboxylesterases (CEs) are α,β-hydrolase fold proteins that catalyse the hydrolysis of a wide range of en-dogenous and exogenous compounds. In mammals, these enzymes are involved in the detoxification of certain pesticides and drugs. However, this toxicological role of CEs has received little attention in marine organisms such as the mussel Mytilus galloprovincialis. Therefore, the purpose of this study was to examine whether mussel CE activity is sensitive to inhibition by environmental chemicals of current concern (human pharmaceuticals and personal care products or PPCPs; i.e fluoxetine, loperamide, simvastatin, fenofibrate, nonylphenol and triclosan), so this chemical interaction may be considered as a detoxification mechanism comparable to that of organo-phosphorus pesticides. First, we examined the basal levels of CE activity in multiple tissues of M. galloprovincialis, using a wide range of colorimetric substrates, i.e., p-nitrophenyl acetate (pNPA), p-nitrophenyl butyrate (pNPB), 1-naphthyl acetate (1-NA), 1-naphthyl butyrate (1-NB) and 2-naphtyl acetate (2-NA). Second, we tested for a substrate-dependence of detecting CE inhibition using the model organophosphorus dichlorvos. Finally, some PPCPs were tested for in vitro CE inhibition using multiple substrates. Results showed that long-chain esters (pNPB and 1-NB) provided the highest CE-mediated hydrolysis rates compared with pNPA or 1-NA. Moreover, the digestive gland and gills displayed the highest CE activities compared with haemolymph. As expected, the esterase enzyme was very sensitive to dichlorvos (IC50 s in the nM range), but dose-inhibition relationships were markedly dependent on the type of substrate used for enzyme assay. The in vitro inhibition kinetics identified triclosan as a potential CE inhibitor (IC50 = 7.07-27.2 μM for digestive gland, IC50 = 10.8-104 μM for gill CE activity), although other PPCPs such as simvastatin, fenofibrate and nonylphenol also decreased the enzyme activity to a lesser extent. In-gel esterase staining after non-denaturing polyacrylamide gel electrophoresis confirmed the inhibitory effect of triclosan upon CE activity, which was more pronounced with the substrate 1-NB. These findings suggest that CE inhibition may be a suitable biomarker of PPCP exposure to be incorporated into the battery of sub-individual indicators of PPCP exposure and toxicity. However, the selection of appropriate substrates is a key issue. Our results indicated that pNPB and 1-NB are the most suitable reporters for detecting inhibition of CE by both organophosphorus and PPCPs in mussels.
Organisms in marine systems are exposed to multiple stressors that create a range of associated environmental and ecotoxicological risks. Examples of stressors include alterations related to climate change, such as temperature increase, and the exposure to pollutants arising from human activities. The present study evaluated the impacts of Arsenic exposure (1 mg/L) and warming (21 °C) in Mytilus galloprovincialis, acting alone and in combination. Our results demonstrated that both Arsenic exposure and warming induced oxidative stress and reduced mussels metabolism, with changes becoming more prominent with the exposure time and when mussels were exposed to both stressors in combination. Furthermore, results obtained showed higher As accumulation in organisms exposed to warming treatments. The present study showed that under warming scenarios, the negative impacts induced by As may be enhanced in ecologically and economically relevant bivalves, with potential impacts on population stocks due to increased sensitivity to pollutants, which may eventually result in biodiversity loss and socio-economic impacts.
Recent studies have become increasingly focused on the assessment of pharmaceuticals occurrence in aquatic ecosystems, however the potential toxicity to non-target organisms is still largely unknown. The antihistamine cetirizine is a commonly used pharmaceutical, already detected in surface waters of marine aquatic systems worldwide. In the present study Mytilus galloprovincialis mussels were exposed to a range of cetirizine concentrations (0.3, 3.0, 6.0 and 12.0 μg/L), resembling moderate to highly contaminated areas, over 28 days. The responses of different biochemical markers were evaluated in mussels whole soft tissue, and included energy-related parameters (glycogen content, GLY; protein content, PROT; electron transport system activity, ETS), and oxidative stress markers (superoxide dismutase activity, SOD; catalase activity, CAT; glutathione S-transferases activity, GSTs; lipid peroxidation levels, LPO; reduced (GSH) and oxidized (GSSG) glutathione content). The results obtained demonstrated that with the increase of exposure concentrations mussels tended to increase their energy reserves and maintain their metabolic potential, which was significantly higher only at the highest concentration. Our findings clearly revealed that cetirizine inhibited the activity of GSTs and although induced the activity of antioxidant enzymes (SOD and CAT) mussels were not able to prevent cellular damages observed through the increase of LPO associated to the increase of exposure concentrations. Thus, this study confirmed that cetirizine induces toxic effects in Mytilus galloprovincialis, which, considering their trophic relevance, wide use as bioindicator and wide spatial distribution of this species, can result in ecological and economic negative impacts at a large scale.
Caulerpin, a secondary metabolite from the marine invasive green algae Caulerpa cylindracea is known to induce mitochondrial dysfunctions. In this study, the anticancer property of caulerpin was assessed in a panel of colorectal cancer cell lines. We demonstrated that caulerpin inhibited oxidative phosphorylation (OXPHOS) and facilitated an early intervention of the mitochondrial function, via inhibiting mitochondrial complex I, accompanied by the dissipation of mitochondrial membrane potential and a surge of reactive oxygen species (ROS) generation. Moreover, in response to the increment in AMP/ATP ratio, the energy sensor AMP-activated protein kinase (AMPK) was activated by caulerpin treatment in a calcium/calmodulin-dependent protein kinase 2 (CaMKK2)‑dependent manner. Distinguished effect on glycolysis was observed at different time-points after caulerpin treatment. Glycolysis was enhanced after a short time treatment with caulerpin, associated with upregulation of glucose transporter 1 (GLUT1), hexokinase II (HKII) and 6-phosphofructo-2-kinase (PFKFB3) protein expressions. However, long-term activation of AMPK by caulerpin damaged the glycolysis and glucose metabolism in colorectal cells, finally causing cell death. The persistent effect of caulerpin was mediated by AMPKα1, rather than AMPKα2, to abolish cell viability through hindering mTORC1-4E-BP1 axis. Moreover, caulerpin synergized with the glycolytic inhibitor 3BP in inhibiting cellular proliferation both in vitro and in vivo. Our findings on the previously uncharacterized anticancer effects of caulerpin may provide potential therapeutic approaches targeting the colorectal carcinoma metabolism.
The red pigment caulerpin, a secondary metabolite from the marine invasive green algae Caulerpa cylindracea can be accumulated and transferred along the trophic chain, with detrimental consequences on biodiversity and ecosystem functioning. Despite increasing research efforts to understand how caulerpin modifies fish physiology, little is known on the effects of algal metabolites on mammalian cells. Here we report for the first time the mitochondrial targeting activity of both caulerpin, and its closely related derivative caulerpinic acid, by using as experimental model rat liver mitochondria, a system in which bioenergetics mechanisms are not altered. Mitochondrial function was tested by polarographic and spectrophotometric methods. Both compounds were found to selectively inhibit respiratory complex II activity, while complexes I, III, and IV remained functional. These results led us to hypothesize that both algal metabolites could be used as antitumor agents in cell lines with defects in mitochondrial complex I. Ovarian cancer cisplatin-resistant cells are a good example of cell lines with a defective complex I function on which these molecules seem to have a toxic effect on proliferation. This provided novel insight toward the potential use of metabolites from invasive Caulerpa species for the treatment of human ovarian carcinoma cisplatin-resistant cells.
At 726 the number of recorded multicellular non indigenous species (NIS) in the Mediterranean Sea is far higher than in other European Seas. Of these, 614 have established populations in the sea. 384 are considered Erythraean NIS, the balance are mostly ship and culture-introductions. In order to effectively implement EU Regulation on the prevention and management of the introduction and spread of invasive NIS and the Marine Strategy Framework Directive in the Mediterranean Sea it is crucial that this priority pathway is appropriately managed. Three potential impediments – incomplete and inaccurate data; unknown impacts; policy mismatch – hinder implementation. Current geographical, taxonomical and impact data gaps will be reduced only by instituting harmonized standards and methodologies for monitoring NIS populations in all countries bordering the Mediterranean Sea, prioritizing bridgehead sites and dispersal hubs. The option of implementing European environmental policies concerning marine NIS in member states alone may seem expedient, but piecemeal protection is futile. Since only 9 of the 23 states bordering the Mediterranean are EU member states, the crucial element for an effective strategy for slowing the influx of NIS is policy coordination with the Regional Sea Convention (Barcelona Convention) to ensure consistency in legal rules, standards and institutional structures to address all major vectors/pathways.
Organisms experience a wide range of environmental factors such as temperature, salinity and hydrostatic pressure, which pose challenges to biochemical processes. Studies on adaptations to such factors have largely focused on macromolecules, especially intrinsic adaptations in protein structure and function. However, micromolecular cosolutes can act as cytoprotectants in the cellular milieu to affect biochemical function and they are now recognized as important extrinsic adaptations. These solutes, both inorganic and organic, have been best characterized as osmolytes, which accumulate to reduce osmotic water loss. Singly, and in combination, many cosolutes have properties beyond simple osmotic effects, e.g. altering the stability and function of proteins in the face of numerous stressors. A key example is the marine osmolyte trimethylamine oxide (TMAO), which appears to enhance water structure and is excluded from peptide backbones, favoring protein folding and stability and counteracting destabilizers like urea and temperature. Co-evolution of intrinsic and extrinsic adaptations is illustrated with high hydrostatic pressure in deep-living organisms. Cytosolic and membrane proteins and G-protein-coupled signal transduction in fishes under pressure show inhibited function and stability, while revealing a number of intrinsic adaptations in deep species. Yet, intrinsic adaptations are often incomplete, and those fishes accumulate TMAO linearly with depth, suggesting a role for TMAO as an extrinsic 'piezolyte' or pressure cosolute. Indeed, TMAO is able to counteract the inhibitory effects of pressure on the stability and function of many proteins. Other cosolutes are cytoprotective in other ways, such as via antioxidation. Such observations highlight the importance of considering the cellular milieu in biochemical and cellular adaptation.
© 2015. Published by The Company of Biologists Ltd.
Bivalve histopathology has become an important tool in aquatic toxicology, having been implemented in many biomonitoring programmes worldwide. However, there are various gaps in the knowledge of many sentinel organisms and the interference of confounding factors. This work aimed (i) to develop a detailed semi-quantitative histopathological index of the digestive gland and gonad of the Mytilus galloprovincialis mussel collected from five sites contaminated with distinct patterns of organic and inorganic toxicants along the Basque coast (SE Bay of Biscay) and (ii) to investigate whether seasonal variability and parasitosis act as confounding factors. A total of twenty-three histopathological alterations were analysed in the digestive gland and gonad following a weighed condition index approach. The alterations were integrated into a single value for a better understanding of the mussels' health status. The digestive gland was consistently more damaged than the gonad. Mussels from the most impacted sites endured the most significant deleterious effects showing inflammation-related alterations together with digestive tubule atrophy and necrosis. Neoplastic diseases were scarce, with only a few cases of fibromas (benign neoplasia). In contrast, in moderately or little impacted sites, contamination levels did not cause significant tissue damage. However, parasites contributed to overestimating the values of histopathological indices (i.e. more severe tissue damage) in mussels from little impacted sites, whilst the opposite occurred in mussels from highly polluted sites. Accordingly, inter-site differences were more pronounced in autumn when natural physiological responses of advanced maturation stages did not interfere in the histological response. In conclusion, although seasonal variability and parasitosis mask the response of histopathological indices, this biomonitoring approach may provide good sensitivity for assessing the health status of mussels if fluctuations of these confounding factors are considered.
Copyright © 2015 Elsevier B.V. All rights reserved.
Among natural products there are molecules well known to influence the abundance and distribution of marine organisms and to play important roles in their interactions with one another. Recently, chemical ecologists have also started to consider how research on natural products might be useful in understanding marine biological invasions, assessing their impact in the invaded areas, and considering how to deal with them. Their efforts especially focused on the Mediterranean Sea, which is one of the major hotspots of marine biological invasions on earth, showing in what way marine natural products (MNPs) may influence (1) the ability of exotic marine organisms to invade and to get established, (2) how they affect the invaded biota, and (3) public health and the economy. In all cases, the study of such chemical warfare between alien and native species started with the isolation of the pure chemicals required for chemical structure elucidation and subsequent biological testing, implying a central role of natural product chemistry in approaching critical issues in invasion biology. In this position paper we also introduce a theme of possible interest for managing marine invasive species, based on the exploitation of available chemical and biological information on MNPs. We show how a kind of chemical data originally gathered for other objectives—such as obtaining drugs from the sea—might also offer valid alternatives to unrealistic eradication campaigns, becoming the basis for a desirable commercial use of the bioactive compounds obtainable from marine pests, thus paving the way for making the control of invasions profitable. The pests might then be harvested, reducing their impact on marine ecosystems.