Psammaplin A and Its Analogs Attenuate Oxidative Stress in Neuronal Cells through Peroxisome Proliferator-Activated Receptor γ Activation.

Psammaplins are sulfur containing bromotyrosine alkaloids that have shown antitumor activity through the inhibition of class I histone deacetylases (HDACs). The cytotoxic properties of psammaplin A (1), the parent compound, are related to peroxisome proliferator-activated receptor γ (PPARγ) activation, but the mechanism of action of its analogs psammaplin K (2) and bisaprasin (3) has not been elucidated. In this study, the protective effects against oxidative stress of compounds 1-3, isolated from the sponge Aplysinella rhax, were evaluated in SH-SY5Y cells. The compounds improved cell survival, recovered glutathione (GSH) content, and reduced reactive oxygen species (ROS) release at nanomolar concentrations. Psammaplins restored mitochondrial membrane potential by blocking mitochondrial permeability transition pore opening and reducing cyclophilin D expression. This effect was mediated by the capacity of 1-3 to activate PPARγ, enhancing gene expression of the antioxidant enzymes catalase, nuclear factor E2-related factor 2 (Nrf2), and glutathione peroxidase. Finally, HDAC3 activity was reduced by 1-3 under oxidative stress conditions. This work is the first description of the neuroprotective activity of 1 at low concentrations and the mechanism of action of 2 and 3. Moreover, it links for the first time the previously described effects of 1 in HDAC3 and PPARγ signaling, opening a new research field for the therapeutic potential of this compound family.

First description of adenosine production by Gnomoniopsis smithogilvyi, causal agent of chestnut brown rot.

Gnomoniopsis smithogilvyi (Gnomoniaceae, Diaporthales) is the main causal agent of chestnut brown rot on sweet chestnut worldwide. The rotting of nuts leads to alterations in the organoleptic qualities and decreased fruit production, resulting in significant economic losses. In 2021, there was an important outbreak of chestnut rot in southern Galicia (Spanish northwest). The profile of secondary metabolites from G. smithogilvyi was studied, especially to determine its capability for producing mycotoxins, as happens with other rotting fungi, due to the possible consequences on the safety of chestnut consumption. Secondary metabolites produced by isolates of G. smithogilvyi growing in potato dextrose agar (PDA) medium were identified using liquid chromatography coupled with high-resolution mass spectrometry. Three metabolites with interesting pharmacological and phyto-toxicological properties were identified based on their exact mass and fragmentation patterns, namely adenosine, oxasetin, and phytosphingosine. The capacity of G. smithogilvyi to produce adenosine in PDA cultures was assessed, finding concentrations ranging from 176 to 834 µg/kg. Similarly, the production of mycotoxins was ruled out, indicating that the consumption of chestnuts with necrotic lesions does not pose a health risk to the consumer in terms of mycotoxins.

Analytical and functional profiles of paralytic shellfish toxins extracted from Semele proficua and Senilia senilis from Angola.

Paralytic shellfish poisoning is a foodborne illness that typically derive from the consumption of shellfish contaminated with saxitoxin-group of toxins produced by dinoflagellates of the genus Gymnodinium, Alexandrium and Pyrodinium. N-sulfocarbamoyl, carbamate and dicarbamoyl are the most abundant. In 2007 and 2008 some episodes of PSP occurred in Angola where there is not monitoring program for shellfish contamination with marine biotoxins. Therefore, ten samples extracted from Semele proficua from Luanda Bay and Senilia senilis from Mussulo Bay, were analyzed by HPLC finding saxitoxin, decarbamoylsaxitoxin and other three compounds that have an unusual profile different to the known hydrophilic PSP toxins were found in different amounts and combinations. These new compounds were not autofluorescent, and they presented much stronger response after peroxide oxidation than after periodate oxidation. The compounds appear as peaks eluted at 2.5 and 5.6 min after periodate oxidation and 8.2 min after peroxide oxidation. Electrophysiological studies revealed that none of the three unknown compounds had effect at cellular level by decreasing the maximum peak inward sodium currents by blocking voltage-gated sodium channels. Thus, not contributing to PSP intoxication. The presence in all samples of saxitoxin-group compounds poses a risk to human health and remarks the need to further explore the presence of new compounds that contaminate seafood, investigating their activity and developing monitoring programs.

Synergistic Effect of Brevetoxin BTX-3 and Ciguatoxin CTX3C in Human Voltage-Gated Nav1.6 Sodium Channels.

Emerging marine biotoxins such as ciguatoxins and brevetoxins have been widely and independently studied as food pollutants. Their maximum levels in food components were set without considering their possible synergistic effects as consequence of their coexistence in seafood and their action at the same cellular target. The absolute lack of data and regulations of the possible combined effects that both marine biotoxins may have raised the need to analyze their direct in vitro effects using electrophysiology techniques. The results presented in this study indicate that ciguatoxins and brevetoxins had a synergistic effect on human Nav1.6 voltage-gated sodium channels by hyperpolarizing their activation and inactivation states. The results presented here indicate that brevetoxin 3 (BTX-3) acts as partial agonist of human sodium channels, while ciguatoxin 3C (CTX3C) was a full agonist, explaining the differences in the effect of each toxin in the channel. Therefore, this work sets the cellular basis to further apply this type of studies to other food toxicants that may act synergistically and thus implement the corresponding regulatory limits considering their coexistence and the risks to human and animal health derived from it.

Acute toxicology report of the emerging marine biotoxin Brevetoxin 3 in mice: Food safety implications.

Brevetoxins (PbTxs) are emerging marine toxins that can lead to Neurotoxic Shellfish Poisoning in humans by the ingestion of contaminated seafood. Recent reports on brevetoxin detection in shellfish in regions where it has not been described before, arise the need of updated guidelines to ensure seafood consumers safety. Our aim was to provide toxicological data for brevetoxin 3 (PbTx3) by assessing oral toxicity in mice and comparing it with intraperitoneal administration. We followed an Up-and-Down procedure administering PbTx3 to mice and registering clinical signs, neuromuscular function, histopathology, and blood changes. Neuromuscular dysfunction like seizures and ataxia, as well as loss of limb strength were observed at 6 h. Performance and clinical signs largely improved at 24 h, time at which no blood biochemical or histological alterations were detected independently of the administration route. However, PbTx3 oral administration results in lower toxicity than intraperitoneal administration. Mortality was only observed at 4000 µg/kg bw PbTx3 administered via oral, but we still found toxicity clinical signs at low toxin doses. We could stablish an oral Lowest-Observable-Adverse-Effect-Level for PbTx3 of 100 µg/kg bw and an oral No-Observable-Adverse-Effect-Level of 10 µg/kg bw in mice. The data here reported should be considered in the evaluation of risks of PbTxs for human health.

Mouse N2a Neuroblastoma Assay: Uncertainties and Comparison with Alternative Cell-Based Assays for Ciguatoxin Detection.

The growing concern about ciguatera fish poisoning (CF) due to the expansion of the microorganisms producing ciguatoxins (CTXs) increased the need to develop a reliable and fast method for ciguatoxin detection to guarantee food safety. Cytotoxicity assay on the N2a cells sensitized with ouabain (O) and veratridine (V) is routinely used in ciguatoxin detection; however, this method has not been standardized yet. This study demonstrated the low availability of sodium channels in the N2a cells, the great O/V damage to the cells and the cell detachment when the cell viability is evaluated by the classical cytotoxicity assay and confirmed the absence of toxic effects caused by CTXs alone when using the methods that do not require medium removal such as lactate dehydrogenase (LDH) and Alamar blue assays. Different cell lines were evaluated as alternatives, such as human neuroblastoma, which was not suitable for the CTX detection due to the greater sensitivity to O/V and low availability of sodium channels. However, the HEK293 Nav cell line expressing the α1.6 subunit of sodium channels was sensitive to the ciguatoxin without the sensitization with O/V due to its expression of sodium channels. In the case of sensitizing the cells with O/V, it was possible to detect the presence of the ciguatoxin by the classical cytotoxicity MTT method at concentrations as low as 0.0001 nM CTX3C, providing an alternative cell line for the detection of compounds that act on the sodium channels.

Cyclophilin B serum levels present variations across the menstrual cycle.

Cyclophilins are a family of chaperones involved in inflammation and cell death. Cyclophilin B is released by inflammatory cells and acts through the receptor CD147, affecting matrix metalloproteases release, whilst cyclophilin D participates in hypoxia-induced apoptosis. Previous studies related hormones like estradiol or prolactin to these proteins, however, their blood concentrations across the menstrual cycle have not been determined. In this work, eleven healthy women (BMI: 21.8 kg/m2) were monitored during a single menstrual cycle, making blood extractions at follicular, periovulatory and mid-luteal phases. Hormone and cyclophilin levels were determined in each phase. Statistical differences were determined by repeated measures ANOVA and estimated marginal means tests, or by Friedman and Dunn-Bonferroni tests for parametric and non-parametric variables, respectively. Bivariate correlations were evaluated with the Spearman coefficient. Cyclophilin B concentrations presented significant differences during the menstrual cycle (p = 0.012). The highest levels of this protein were found at follicular extraction, followed by a decrease at periovulatory phase and a slight increase at mid-luteal phase. Cyclophilin D showed the same profile, although statistical significance was not reached. This immunophilin exhibited a positive correlation with luteinizing hormone at periovulatory phase (r = 0.743, p = 0.009) and with follicle stimulating hormone at mid-luteal phase (r = 0.633, p = 0.036). This is the first study describing the changes in cyclophilin B concentrations across the menstrual cycle, as well as the association of luteinizing and follicle stimulating hormones with cyclophilin D. These results suggest a role of these proteins in the cyclic inflammatory events that affect female reproductive system that should be explored.

Proteomic and toxicological analysis of the response of dinoflagellate Alexandrium catenella to changes in NaNO3 concentration.

Dinoflagellates of the genus Alexandrium cause Harmful Algal Blooms (HABs) in coastal waters worldwide, damaging marine environments, aquaculture, and human health. They synthesize potent neurotoxic alkaloids known as PSTs (i.e., Paralytic Shellfish Toxins), the etiological agents of PSP (i.e., Paralytic Shellfish Poisoning). In recent decades, the eutrophication of coastal waters with inorganic nitrogen (e.g., nitrate, nitrite, and ammonia) has increased the frequency and scale of HABs. PSTs concentrations within Alexandrium cells can increase by up to 76% after a nitrogen enrichment event; however, the mechanisms that underlie their biosynthesis in dinoflagellates remains unclear. This study combines mass spectrometry, bioinformatics, and toxicology and investigates the expression profiles of PSTs in Alexandrium catenella grown in 0.4, 0.9 and 1.3 mM NaNO3. Pathway analysis of protein expression revealed that tRNA amino acylation, glycolysis, TCA cycle and pigment biosynthesis were upregulated in 0.4 mM and downregulated in 1.3 mM NaNO3 compared to those grown in 0.9 mM NaNO3. Conversely, ATP synthesis, photosynthesis and arginine biosynthesis were downregulated in 0.4 mM and upregulated in 1.3 mM NaNO3. Additionally, the expression of proteins involved in PST biosynthesis (sxtA, sxtG, sxtV, sxtW and sxtZ) and overall PST production like STX, NEO, C1, C2, GTX1-6 and dcGTX2 was higher at lower nitrate concentrations. Therefore, increased nitrogen concentrations increase protein synthesis, photosynthesis, and energy metabolism and decrease enzyme expression in PST biosynthesis and production. This research provides new clues about how the changes in the nitrate concentration can modulate different metabolic pathways and the expression of PST biosynthesis in toxigenic dinoflagellates.

Cyclophilins modify their profile depending on the organ or tissue in a murine inflammatory model.

Inflammation is the leading subjacent cause of many chronic diseases. Despite several studies in the last decades, the molecular mechanism involving its pathophysiology is not fully known. Recently, the implication of cyclophilins in inflammatory-based diseases has been demonstrated. However, the main role of cyclophilins in these processes remains elusive. Hence, a mouse model of systemic inflammation was used to better understand the relationship between cyclophilins and their tissue distribution. To induce inflammation, mice were fed with high-fat diet for 10 weeks. In these conditions, serum levels of interleukins 2 and 6, tumour necrosis factor-α, interferon-ϒ, and the monocyte chemoattractant protein 1 were elevated, evidencing a systemic inflammatory state. Then, in this inflammatory model, cyclophilins and CD147 profiles in the aorta, liver, and kidney were studied. The results demonstrate that, upon inflammatory conditions, cyclophilins A and C expression levels were increased in the aorta. Cyclophilins A and D were augmented in the liver, meanwhile, cyclophilins B and C were diminished. In the kidney, cyclophilins B and C levels were elevated. Furthermore, CD147 receptor was also increased in the aorta, liver, and kidney. In addition, when cyclophilin A was modulated, serum levels of inflammatory mediators were decreased, indicating a reduction in systemic inflammation. Besides, the expression levels of cyclophilin A and CD147 were also reduced in the aorta and liver, when cyclophilin A was modulated. Therefore, these results suggest that each cyclophilin has a different profile depending on the tissue, under inflammatory conditions.

Association of cyclophilins and cardiovascular risk factors in coronary artery disease.

Cyclophilins are chaperone proteins that play important roles in signal transduction. Among them, cyclophilins A, B, C, and D were widely associated with inflammation and cardiovascular diseases. Cyclophilins A and C have been proposed as coronary artery disease biomarkers. However, less is known about their relationship with cardiovascular risk factors. Therefore, this study aimed to determine the association between cyclophilin A, B, C, and D and cardiovascular risk factors in coronary artery disease. Serum levels of cyclophilins were measured in 167 subjects (subdivided according to cardiovascular risk factors presence). This study reveals that cyclophilin A and C are elevated in patients regardless of the risk factors presence. Moreover, cyclophilin B is elevated in male patients with hypertension, type 2 diabetes, or high glucose levels. In addition, cyclophilins A, B, and C were significantly correlated with cardiovascular risk factors, but only cyclophilin B was associated with type 2 diabetes. The multivariate analysis strengthens the predictive value for coronary artery disease presence of cyclophilin A (>8.2 ng/mL) and cyclophilin C (>17.5 pg/mL) along with the cardiovascular risk factors tobacco, hypertension, dyslipidemia, and high glucose and cholesterol levels. Moreover, the risk of coronary artery disease is increased in presence of cyclophilin B levels above 63.26 pg/mL and with hypertension or dyslipidemia in male patients. Consequently, cyclophilins A and C serum levels are reinforced as useful coronary artery disease biomarkers, meanwhile, cyclophilin B is a valuable biomarker in the male population when patients are also suffering from hypertension or dyslipidemia.

Detection of mycotoxins in cheese using an optimized analytical method based on a QuEChERS extraction and UHPLC-MS/MS quantification.

Mycotoxins can produce toxic effects on humans; hence, it is of high importance to determine their presence in food products. This work presents a reliable method for the quantification of 32 mycotoxins in cheese. The analysis procedure was optimized based on a QuEChERS extraction process and the ultra-high performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS) detection. The analysis method was validated for four cheese varieties (emmental, blue, brie and camembert) in terms of linearity, sensitivity, matrix effect, accuracy and precision. Satisfactory precision and accuracy values were achieved, with recoveries above 70% for most mycotoxins. The developed method was applied to the analysis of 38 commercial cheese samples. A high occurrence of beauvericin and enniatins were found, ranging from 31% for enniatin A to 100% for enniatin B. The ochratoxin A was detected in three samples at concentrations that may pose a risk to human health.

Synergistic effect of environmental food pollutants: Pesticides and marine biotoxins.

Emerging marine biotoxins such as ciguatoxins and pyrethroid compounds, widely used in agriculture, are independently treated as environmental toxicants. Their maximum residue levels in food components are set without considering their possible synergistic effects as consequence of their interaction with the same cellular target. There is an absolute lack of data on the possible combined cellular effects that biological and chemical pollutants, may have. Nowadays, an increasing presence of ciguatoxins in European Coasts has been reported and these toxins can affect human health. Similarly, the increasing use of phytosanitary products for control of food plagues has raised exponentially during the last decades due to climate change. The lack of data and regulation evaluating the combined effect of environmental pollutants with the same molecular target led us to analyse their in vitro effects. In this work, the effects of ciguatoxins and pyrethroids in human sodium channels were investigated. The results presented in this study indicate that both types of compounds have a profound synergistic effect in voltage-dependent sodium channels. These food pollutants act by decreasing the maximum peak inward sodium currents and hyperpolarizing the sodium channels activation, effects that are boosted by the simultaneous presence of both compounds. A fact that highlights the need to re-evaluate their limits in feedstock as well as their potential in vivo toxicity considering that they act on the same cellular target. Moreover, this work sets the cellular basis to further apply this type of studies to other water and food pollutants that may act synergistically and thus implement the corresponding regulatory limits taking into account its presence in a healthy diet.

Bromoditerpenes from the Red Seaweed Sphaerococcus coronopifolius as Potential Cytotoxic Agents and Proteasome Inhibitors and Related Mechanisms of Action.

Seaweeds are a great source of compounds with cytotoxic properties with the potential to be used as anticancer agents. This study evaluated the cytotoxic and proteasome inhibitory activities of 12R-hydroxy-bromosphaerol, 12S-hydroxy-bromosphaerol, and bromosphaerol isolated from Sphaerococcus coronopifolius. The cytotoxicity was evaluated on malignant cell lines (A549, CACO-2, HCT-15, MCF-7, NCI-H226, PC-3, SH-SY5Y, and SK-MEL-28) using the MTT and LDH assays. The ability of compounds to stimulate the production of hydrogen peroxide (H2O2) and to induce mitochondrial dysfunction, the externalization of phosphatidylserine, Caspase-9 activity, and changes in nuclear morphology was also studied on MCF-7 cells. The ability to induce DNA damage was also studied on L929 fibroblasts. The proteasome inhibitory activity was estimated through molecular docking studies. The compounds exhibited IC50 values between 15.35 and 53.34 µM. 12R-hydroxy-bromosphaerol and 12S-hydroxy-bromosphaerol increased the H2O2 levels on MCF-7 cells, and bromosphaerol induced DNA damage on fibroblasts. All compounds promoted a depolarization of mitochondrial membrane potential, Caspase-9 activity, and nuclear condensation and fragmentation. The compounds have been shown to interact with the chymotrypsin-like catalytic site through molecular docking studies; however, only 12S-hydroxy-bromosphaerol evidenced interaction with ALA20 and SER169, key residues of the proteasome catalytic mechanism. Further studies should be outlined to deeply characterize and understand the potential of those bromoditerpenes for anticancer therapeutics.

Extracellular cyclophilins A and C induce dysfunction of pancreatic microendothelial cells.

Extracellular cyclophilins (eCyps) A and B are chemotactic mediators in several illnesses in which inflammation plays an important role such as diabetes and cardiovascular diseases. Recently, eCypC has been reported as a potential biomarker for coronary artery disease but its effect in endothelium has not been determined. Moreover, there is a lack of studies with all these proteins in the same model, which makes difficult a direct comparison of their effects. In this work, MS1 pancreatic microendothelial cells were treated with eCyps A, B and C and their impact on endothelial function was analysed. eCyps A and C stimulated the release of IL-6 and MCP-1 and increased the expression of the receptor CD147, but eCypB did not affect these pro-inflammatory markers. Moreover, eCypC activated the translocation of NFkB-p65 to the nucleus. All these effects were reversed by pre-treatment with cyclosporine A. eCyps also produced endothelial dysfunction, as evidenced by the decrease in eNOS activation. Finally, the crosstalk among eCyps addition and their protein and gene expression was evaluated. eCypA generated a depletion in its protein and gene levels, whilst eCyps B and C upregulated their own protein expression. Moreover, each eCyp altered the intracellular expression of other Cyps, including cyclophilin D. This work is the first report of eCyps influence on iCyps expression, as well as the first description of eCypC as an activator of CD147 receptor and a mediator of endothelial dysfunction, which points to a potential role of this protein in vascular complications associated to diabetes.

Intestinal secretory mechanisms in Okadaic acid induced diarrhoea.

Okadaic acid (OA) is an important marine lipophilic phycotoxin responsible for diarrhetic shellfish poisoning (DSP). This toxin inhibits protein phosphatases (PPs) like PP2A and PP1, though, this action does not explain OA-induced toxicity and symptoms. Intestinal epithelia comprise the defence barrier against external agents where transport of fluid and electrolytes from and to the lumen is a tightly regulated process. In some intoxications this balance becomes dysregulated appearing diarrhoea. Therefore, we evaluated diarrhoea in orally OA-treated mice as well as in mice pre-treated with several doses of cyproheptadine (CPH) and then treated with OA at different times. We assessed stools electrolytes and ultrastructural alteration of the intestine, particularly evaluating tight and adherens junctions. We detected increased chloride and sodium faecal concentrations in the OA-exposed group, suggesting a secretory diarrhoea. Pre-treatment with CPH maintains chloride concentration in values similar to control mice. Intestinal cytomorphological alterations were observed for OA mice, whereas CPH pre-treatment attenuated OA-induced damage in proximal colon and jejunum at 2 h. Conversely, tight junctions' distance was only affected by OA in jejunum at the moment diarrhoea occurred. In this study we found cellular mechanisms by which OA induced diarrhoea revealing the complex toxicity of this compound.

Enniatins A1 and B1 alter calcium homeostasis of neuronal cells leading to apoptotic death.

Enniatins (ENNs) A1 and B1 are non-regulated mycotoxins produced by Fusarium spp. that commonly occur in different types of food. These toxins are cytotoxic in several cell lines, but their mechanism of action is unclear. In this study, the cytotoxic effects of ENNs A1 and B1 in SH-SY5Y human neuroblastoma cells were analysed. Moreover, to better understand their mechanism of action, mitochondrial function, reactive oxygen species (ROS) levels and calcium fluxes were monitored. ENNs A1 and B1 reduced cell viability, presenting IC50 values of 2.0 and 2.7 µM, respectively. Both toxins induced caspase-dependent apoptosis, but only ENN A1 increased ROS production. Apoptotic cell death seems to be triggered by the increase in cytosolic calcium produced by both ENNs, since the toxins altered Ca2+ homeostasis by depleting intracellular reservoirs. Finally, binary combinations of ENN A1, ENN B1, ENN A and ENN B were tested. All mixtures resulted in an antagonistic effect, with the exception of ENN A and ENN B1 combination, which produced an additive effect. The results presented in this study provide the first evidence of ENNs A1 and B1 effects on calcium fluxes, providing new insights into the mechanism of action of these mycotoxins.

NeuroTorp, a lateral flow test based on toxin-receptor affinity for in-situ early detection of cyclic imine toxins.

The emergent cyclic imine toxins produced by marine dinoflagellates are potent antagonists of nicotinic acetylcholine receptors. Shellfish accumulate cyclic imine toxins following filter-feeding on toxic dinoflagellates vectoring them to humans. Herein is presented a lateral flow test for the detection of cyclic imine toxins based on three new concepts for test strips: i) the immobilization of lipoprotein vesicles in the test-line, ii) the high affinity of neurotoxins for their receptor targets and iii) the use of high porosity glass fiber filter membranes as support for the fabrication of the lateral flow test NeuroTorp (WO2017108115). Purified electrocyte membrane vesicles from Torpedo marmorata were used as a source of receptor and were immobilized in the test-line. Biotin-α-bungarotoxin was used as toxin tracer for the NeuroTorp LFT given its high affinity for nicotinic acetylcholine receptors while neutravidin nanogold particle conjugates enable its visual detection. Herein is reported for the first time the use of GF/C glass fiber membranes as the stationary phase for a lateral flow test. The GF/C filter ensures both: the immobilization of a complex lipoprotein in the test-line and the capillary migration of the mobile phase. Scanning electron microscopy studies shed light into the mechanism by which Torpedo-electrocyte membranes vesicles are immobilized in the GF/C glass microfiber. The electrocyte membrane vesicles anchor in neighboring microfibers randomly disposed in the same plane of the GF/C filter forming stable microfilm structures ensuring the functionality of nicotinic acetylcholine receptors. NeuroTorp is a ready-to-use low-cost early warning device for rapid detection of cyclic imine toxins in shellfish by end-users.

Furanoditerpenes from Spongia (Spongia) tubulifera Display Mitochondrial-Mediated Neuroprotective Effects by Targeting Cyclophilin D.

Neuroprotective properties of five previously described furanoditerpenes 1-5, isolated from Spongia (Spongia) tubulifera, were evaluated in an in vitro oxidative stress model in SH-SY5Y cells. Dose-response treatments revealed that 1-5 improved cell survival at nanomolar concentrations through the restoration of mitochondrial membrane potential and the reduction of reactive oxygen species. Their ability to prevent the mitochondrial permeability transition pore opening was also assessed, finding that 4 and 5 inhibited the channel at 0.001 µM. This inhibition was accompanied by a decrease in the expression of cyclophilin D, the main regulator of the pore, which was also reduced by 1 and 2. However, the activation of ERK and GSK3ß, upstream modulators of the channel, was not affected by compounds. Therefore, their ability to bind cyclophilin D was evaluated by surface plasmon resonance, observing that 2-5 presented equilibrium dissociation constants in the micromolar range. All compounds also showed affinity for cyclophilin A, being 1 selective toward this isoform, while 2 and 5 exhibited selectivity for cyclophilin D. When the effects on the intracellular expression of cyclophilins A-C were determined, it was found that only 1 decreased cyclophilin A, while cyclophilins B and C were diminished by most compounds, displaying enhanced effects under oxidative stress conditions. Results indicate that furanoditerpenes 1-5 have mitochondrial-mediated neuroprotective properties through direct interaction with cyclophilin D. Due to the important role of this protein in oxidative stress and inflammation, compounds are promising drugs for new therapeutic strategies against neurodegeneration.

In vivo subchronic effects of ciguatoxin-related compounds, reevaluation of their toxicity.

Ciguatoxins are marine compounds that share a ladder-shaped polyether structure produced by dinoflagellates of the genus Gambierdiscus and Fukuyoa, and include maitotoxins (MTX1 and MTX3), ciguatoxins (CTX3C) and analogues (gambierone), components of one of the most frequent human foodborne illness diseases known as ciguatera fish poisoning. This disease was previously found primarily in tropical and subtropical areas but nowadays, the dinoflagellates producers of ciguatoxins had spread to European coasts. One decade ago, the European Food Safety Authority has raised the need to complete the toxicological available data for the ciguatoxin group of compounds. Thus, in this work, the in vivo effects of ciguatoxin-related compounds have been investigated using internationally adopted guidelines for the testing of chemicals. Intraperitoneal acute toxicity was tested for maitotoxin 1 at doses between 200 and 3200 ng/kg and the acute oral toxicity of Pacific Ciguatoxin CTX3C at 330 and 1050 ng/kg and maitotoxin 1 at 800 ng/kg were also evaluated showing not effects on mice survival after a 96 h observation period. Therefore, for the following experiments the oral subchronic doses were between 172 and 1760 ng/kg for gambierone, 10 and 102 ng/kg for Pacific Ciguatoxin CTX3C, 550 and 1760 ng/kg for maitotoxin 3 and 800, 2560 and 5000 ng/kg for maitotoxin 1. The results presented here raise the need to reevaluate the in vivo activity of these agents. Although the intraperitoneal lethal dose of maitotoxin 1 is assumed to be 50 ng/kg, without chemical purity identifications and description of the bioassay procedures, in this work, an intraperitoneal lethal dose of 1107 ng/kg was obtained. Therefore, the data presented here highlight the need to use a common procedure and certified reference material to clearly establish the levels of these environmental contaminants in food.

Current Trends and New Challenges in Marine Phycotoxins.

Marine phycotoxins are a multiplicity of bioactive compounds which are produced by microalgae and bioaccumulate in the marine food web. Phycotoxins affect the ecosystem, pose a threat to human health, and have important economic effects on aquaculture and tourism worldwide. However, human health and food safety have been the primary concerns when considering the impacts of phycotoxins. Phycotoxins toxicity information, often used to set regulatory limits for these toxins in shellfish, lacks traceability of toxicity values highlighting the need for predefined toxicological criteria. Toxicity data together with adequate detection methods for monitoring procedures are crucial to protect human health. However, despite technological advances, there are still methodological uncertainties and high demand for universal phycotoxin detectors. This review focuses on these topics, including uncertainties of climate change, providing an overview of the current information as well as future perspectives.