Palytoxin Toxicity Research Articles

Toxicity of palytoxin, purified ovatoxin-a, ovatoxin-d and extracts of Ostreopsis cf. ovata on the Caco-2 intestinal barrier model

Human intoxications in the Mediterranean Sea have been linked to blooms of the dinoflagellate Ostreopsis cf. ovata, producer of palytoxin (PlTX)-like toxins called ovatoxins (OVTXs). Exposure routes include only inhalation and contact, although PlTX-poisoning by seafood has been described in tropical regions. To address the impact of OVTXs on the intestinal barrier, dinoflagellate extracts, purified OVTX-a and -d and PlTX were tested on differentiated Caco-2 cells. Viability, inflammatory response and barrier integrity were recorded after 24 h treatment. OVTX-a and -d were not cytotoxic up to 20 ng/mL but increased IL-8 release, although to a lesser extent compared to PlTX. While PlTX and OVTX-a (at 0.5 and 5 ng/mL respectively) affected intestinal barrier integrity, OVTX-d up to 5 ng/mL did not. Overall, OVTX-d was shown to be less toxic than OVTX-a and PlTX. Therefore, oral exposure to OVTX-a and -d could provoked lower acute toxicity than PlTX.

In Vivo Evaluation of the Chronic Oral Toxicity of the Marine Toxin Palytoxin.

Palytoxin (PLTX) is one of the most poisonous substances known to date and considered as an emergent toxin in Europe. Palytoxin binds to the Na+-K+ ATPase, converting the enzyme in a permeant cation channel. This toxin is known for causing human fatal intoxications associated with the consumption of contaminated fish and crustaceans such as crabs, groupers, mackerel, and parrotfish. Human intoxications by PLTX after consumption of contaminated fishery products are a serious health issue and can be fatal. Different reports have previously explored the acute oral toxicity of PLTX in mice. Although the presence of palytoxin in marine products is currently not regulated in Europe, the European Food Safety Authority expressed its opinion on PLTX and demanded assessment for chronic toxicity studies of this potent marine toxin. In this study, the chronic toxicity of palytoxin was evaluated after oral administration to mice by gavage during a 28-day period. After chronic exposure of mice to the toxin, a lethal dose 50 (LD50) of 0.44 µg/kg of PLTX and a No-Observed-Adverse-Effect Level (NOAEL) of 0.03 µg/kg for repeated daily oral administration of PLTX were determined. These results indicate a much higher chronic toxicity of PLTX and a lower NOAEL than that previously described in shorter treatment periods, pointing out the need to further reevaluate the levels of this compound in marine products.

Reevaluation of the acute toxicity of palytoxin in mice: Determination of lethal dose 50 (LD50) and No-observed-adverse-effect level (NOAEL)

Palytoxin is an emergent toxin in Europe and one of the most toxic substances know to date. The toxin disrupts the physiological functioning of the Na+/K+-ATPase converting the enzyme in a permeant cation channel. Human intoxications by PLTX after consumption of contaminated fishery products are a serious health issue and can be fatal. Several reports have previously investigated the oral and intraperitoneal toxicity of PLTX in mice. However, in all cases short observation periods (24 and 48 h) after toxin administration were evaluated. In this work, single oral or intraperitoneal doses of PLTX were administered to healthy mice and surviving animals were followed up for 96 h. The data obtained here allowed us to calculate the oral and intraperitoneal lethal doses 50 (LD50) which were in the range of the values previously described. Surprisingly, the oral NOAEL for PLTX was more than 10 times lower than that previously described, a fact that indicates the need for the reevaluation of the levels of the toxin in edible fishery products.

A new ecotoxicity tolerance index of macrobenthos associated with Zoanthus sansibaricus in the littoral zone of Hormuz Island, Persian Gulf, Iran.

Palytoxin (PTX), one of the most potent marine natural products, which has been produced in zoanthids, also occurred in various other marine macrobenthos living in close association with zoanthid colonies, e.g. molluscs, accumulate high toxin concentrations in their organs, where PTX is stored in its active form. In littoral of Hormuz Island, among identified zoanthids, the highest number of toxic colonies per unit area is related to Zoanthus sansibaricus which can have a significant impact on assossiated macrobenthos with it. The purpose of this study was to collect more information on the toxicity of PTX in Z. sansibaricus and the consequences of exposure to PTX on associated macrobenthos with it using aqatic snails and the calculation of ecotoxicity tolerance index (ETI). Hence, zoanthid colonies were collected during reef walk at low tide in April 2016. Also, among assossiated gastropods, Planaxis sulcatus, were collected from the rock for experimental purpose as they are very abundant on and around Hormuz Island reef. In this model for each transect, 36 snails were divided into 4 set, each set had 3 replicates and working mucus solution, as PTX exist in the mucus of zoanthids, was injected in 2 different doses. Results showed that no snails were dead during the study period. However, calculation of Dara index (DI) was indicated that PTX accumulate in high toxin concentrations in snail’s foot and thus snail can be introduced as an indicator of ecotoxicity conditions. Results of ETI, also were shown moderate poisoning in all transects. Hence, tolerance of marine animals to the toxin may enable it to enter food chains and to be followed by potential exposure for humans. However, due to lack of data on PTX of zoanthids research for the entire Persian Gulf, further investigation is needed to clearly ascertain this matter.

Toxic potential of palytoxin.

This review briefly describes the origin, chemistry, molecular mechanism of action, pharmacology, toxicology, and ecotoxicology of palytoxin and its analogues. Palytoxin and its analogues are produced by marine dinoflagellates. Palytoxin is also produced by Zoanthids (i.e. Palythoa), and Cyanobacteria (Trichodesmium). Palytoxin is a very large, non-proteinaceous molecule with a complex chemical structure having both lipophilic and hydrophilic moieties. Palytoxin is one of the most potent marine toxins with an LD50 of 150 ng/kg body weight in mice exposed intravenously. Pharmacological and electrophysiological studies have demonstrated that palytoxin acts as a hemolysin and alters the function of excitable cells through multiple mechanisms of action. Palytoxin selectively binds to Na(+)/K(+)-ATPase with a Kd of 20 pM and transforms the pump into a channel permeable to monovalent cations with a single-channel conductance of 10 pS. This mechanism of action could have multiple effects on cells. Evaluation of palytoxin toxicity using various animal models revealed that palytoxin is an extremely potent neurotoxin following an intravenous, intraperitoneal, intramuscular, subcutaneous or intratracheal route of exposure. Palytoxin also causes non-lethal, yet serious toxic effects following dermal or ocular exposure. Most incidents of palytoxin poisoning have manifested after oral intake of contaminated seafood. Poisonings in humans have also been noted after inhalation, cutaneous/systemic exposures with direct contact of aerosolized seawater during Ostreopsis blooms and/or through maintaining aquaria containing Cnidarian zoanthids. Palytoxin has a strong potential for toxicity in humans and animals, and currently this toxin is of great concern worldwide.

Toxicity of palytoxin after repeated oral exposure in mice and in vitro effects on cardiomyocytes

Palytoxin (PLTX) is a highly toxic hydrophilic polyether detected in several edible marine organisms from intra-tropical areas, where seafood poisoning were reported. Symptoms usually start with gastro-intestinal malaise, often accompanied by myalgia, muscular cramps, dyspnea and, sometimes, arrhythmias. Monitoring programs in the Mediterranean Sea have detected PLTX-like molecules in edible mollusks and echinoderms. Despite the potential exposure of the human population and its high toxic potential, the toxicological profile of the molecule is still an issue. Thus, the effects of repeated oral administration of PLTX in mice were investigated. Seven days of PLTX administration caused lethality and toxic effects at doses ≥30 μg/kg/day. A NOAEL was estimated equal to 3 μg/kg/day, indicating a quite steep dose–response curve. This value, due to the limited number of animal tested, is provisional, although represents a sound basis for further testing. Macroscopic alterations at gastrointestinal level (gastric ulcers and intestinal fluid accumulation) were observed in mice dead during the treatment period. Histological analysis highlighted severe inflammation, locally associated with necrosis, at pulmonary level, as well as hyper-eosinophilia and fiber separation in myocardium. A cardiac damage was supported by the in vitro effect of the toxin on cardiomyocytes, indicating a severe and irreversible impairment of their electrical properties: electrophysiological recordings detected a progressive cell depolarization, arrest of action potentials and beating.

Head and neck cancer cells and xenografts are very sensitive to palytoxin: decrease of c-jun n-terminale kinase-3 expression enhances palytoxin toxicity

ObjectivesPalytoxin (PTX), a marine toxin isolated from the Cnidaria (zooanthid) Palythoa caribaeorum is one of the most potent non-protein substances known. It is a very complex molecule that presents both lipophilic and hydrophilic areas. The effect of PTX was investigated in a series of experiments conducted in head and neck squamous cell carcinoma (HNSCC) cell lines and xenografts.Materials and methodsCell viability, and gene expression of the sodium/potassium-transporting ATPase subumit alpha1 (ATP1AL1) and GAPDH were analyzed in HNSCC cells and normal epithelial cells after treatment with PTX using cytotoxicity-, clonogenic-, and enzyme inhibitor assays as well as RT-PCR and Northern Blotting. For xenograft experiments severe combined immunodeficient (SCID) mice were used to analyze tumor regression. The data were statistically analyzed using One-Way Annova (SPSS vs20).ResultsSignificant toxic effects were observed in tumor cells treated with PTX (LD50 of 1.5 to 3.5 ng/ml) in contrast to normal cells. In tumor cells PTX affected both the release of LDH and the expression of the sodium/potassium-transporting ATPase subunit alpha1 gene suggesting loss of cellular integrity, primarily of the plasma membrane. Furthermore, strong repression of the c-Jun N-terminal kinase 3 (JNK3) mRNA expression was found in carcinoma cells which correlated with enhanced toxicity of PTX suggesting an essential role of the mitogen activated protein kinase (MAPK)/JNK signalling cascades pathway in the mechanisms of HNSCC cell resistance to PTX. In mice inoculated with carcinoma cells, injections of PTX into the xenografted tumors resulted within 24 days in extensive tumor destruction in 75% of the treated animals (LD50 of 68 ng/kg to 83 ng/kg) while no tumor regression occurred in control animals.ConclusionsThese results clearly provide evidence that PTX possesses preferential toxicity for head and neck carcinoma cells and therefore it is worth further studying its impact which may extend our knowledge of the biology of head and neck cancer.

The Stretch-Activated Channel Blocker Gd3+Reduces Palytoxin Toxicity in Primary Cultures of Skeletal Muscle Cells

Palytoxin (PLTX) is one of the most toxic seafood contaminants ever isolated. Reports of human food-borne poisoning ascribed to PLTX suggest skeletal muscle as a primary target site. Primary cultures of mouse skeletal muscle cells were used to study the relationship between Ca(2+) response triggered by PLTX and the development of myotoxic insult. Ca(2+) imaging experiments revealed that PLTX causes a transitory intracellular Ca(2+) response (transient phase) followed by a slower and more sustained Ca(2+) increase (long-lasting phase). The transient phase is due to Ca(2+) release from intracellular stores and entry through voltage-dependent channels and the Na(+)/Ca(2+) exchanger (reverse mode). The long-lasting phase is due to a massive and prolonged Ca(2+) influx from the extracellular compartment. Sulforhodamine B assay revealed that the long-lasting phase is the one responsible for the toxicity in skeletal muscle cells. Our data analyzed, for the first time, pathways of PLTX-induced Ca(2+) entry and their correlation with PLTX-induced toxicity in skeletal muscle cells. The cellular morphology changes induced by PLTX and the sensitivity to gadolinium suggest a role for stretch-activated channels.

Acute oral toxicity of Palytoxin and Okadaic acid in mice

Acute oral toxicity of Palytoxin and Okadaic acid in mice

Palytoxin toxicology: Animal studies

Palytoxin and its derivatives have been implicated in toxic events in humans following ingestion or inhalation, and many studies on the toxicities of these substances to animals, via various routes of administration, have been described. In this report, the toxicity of palytoxin to animals has been reviewed, with comments on possible mechanisms of action. Information required for the risk assessment of palytoxin and its derivatives is by no means complete, and recommendations for further studies, which may better inform regulatory decisions regarding these substances, are also discussed.

Toxicity of palytoxin and 42-hydroxy-palytoxin on cultured mouse skeletal muscle cells

Toxicity of palytoxin and 42-hydroxy-palytoxin on cultured mouse skeletal muscle cells

Palytoxin toxicity after acute oral administration in mice

The acute oral toxicity of palytoxin (PLTX), a highly toxic compound associated with seafood intoxication in tropical and subtropical areas, was investigated in mice. After gavage administration (300–1697μg/kg) to groups of five female CD-1 mice, signs of toxicity and lethality were recorded for 24h. The LD50 was 767μg/kg (95% confidence limits: 549–1039μg/kg) and the main symptoms observed were scratching, jumping, respiratory distress and paralysis. Hematoclinical analyses showed increased levels of creatine phosphokinase and lactate dehydrogenase at doses of 600μg/kg and above, and aspartate transaminase at 848μg/kg and above. Histological analysis revealed acute inflammation of the forestomach in mice surviving up to 24h after administration (424–1200μg/kg). Other histological alterations were observed in the liver and pancreas, while cardiac and skeletal muscle cells revealed only ultrastructural alterations visible by transmission electron microscopy. Ultrastructural and hematoclinical findings suggest an involvement of skeletal and/or cardiac muscle as targets of PLTX, according to the observed human symptoms. A NOEL of 300μg/kg can be estimated from this acute oral toxicity study.

Human risk associated with palytoxin exposure

Palytoxin (PTX) was first isolated from the zoanthid Palythoa toxica. Evaluation of PTX toxicity using various animal models determined that PTX was extremely potent through intravenous, intraperitoneal, and intratracheal exposure. PTX was less potent by direct intragastric exposure. PTX also caused significant, non-lethal effects through dermal and ocular exposure. PTX and PTX-like compounds have now been found in additional zoanthid species, red alga, a sea anemone, and several dinoflagellates. PTXs are found throughout certain reef associated food webs, including in fish and crabs responsible for human illness and death. Many of the organisms found to contain PTXs in the environment are also sold in the home aquarium trade, and recent evidence suggests poisonings have occurred through exposure to these organisms. Due to co-occurrence with other seafood toxins, such as ciguatoxins, saxitoxins, and tetrodotoxin, it has been difficult to assess the true risk of PTX poisoning through seafood consumption in humans, but limited cases have been well documented, some involving human fatalities. Recent evidence also suggests that humans are negatively impacted through PTX exposure by inhalation and dermal routes. Continued research into the distribution and occurrence of PTX and PTX-like compounds both in seafood and marine organisms sold in the aquarium trade appears warranted.

Marine toxins and the cytoskeleton: a new view of palytoxin toxicity

Palytoxin is a marine toxin first isolated from zoanthids (genus Palythoa), even though dinoflagellates of the genus Ostreopsis are the most probable origin of the toxin. Ostreopsis has a wide distribution in tropical and subtropical areas, but recently these dinoflagellates have also started to appear in the Mediterranean Sea. Two of the most remarkable properties of palytoxin are the large and complex structure (with different analogs, such as ostreocin-D or ovatoxin-a) and the extreme acute animal toxicity. The Na(+)/K(+)-ATPase has been proposed as receptor for palytoxin. The marine toxin is known to act on the Na(+) pump and elicit an increase in Na(+) permeability, which leads to depolarization and a secondary Ca(2+) influx, interfering with some functions of cells. Studies on the cellular cytoskeleton have revealed that the signaling cascade triggered by palytoxin leads to actin filament system distortion. The activity of palytoxin on the actin cytoskeleton is only partially associated with the cytosolic Ca(2+) changes; therefore, this ion represents an important factor in altering this structure, but it is not the only cause. The goal of the present minireview is to compile the findings reported to date about: (a) how palytoxin and analogs are able to modify the actin cytoskeleton within different cellular models; and (b) what signaling mechanisms could be involved in the modulation of cytoskeletal dynamics by palytoxin.

Palytoxin disrupts cardiac excitation-contraction coupling through interactions with P-type ion pumps.

Palytoxin is a coral toxin that seriously impairs heart function, but its effects on excitation-contraction (E-C) coupling have remained elusive. Therefore, we studied the effects of palytoxin on mechanisms involved in atrial E-C coupling. In field-stimulated cat atrial myocytes, palytoxin caused elevation of diastolic intracellular Ca(2+) concentration ([Ca(2+)](i)), a decrease in [Ca(2+)](i) transient amplitude, Ca(2+) alternans followed by [Ca(2+)](i) waves, and failures of Ca(2+) release. The decrease in [Ca(2+)](i) transient amplitude occurred despite high sarcoplasmic reticulum (SR) Ca(2+) load. In voltage-clamped myocytes, palytoxin induced a current with a linear current-voltage relationship (reversal potential approximately 5 mV) that was blocked by ouabain. Whole cell Ca(2+) current and ryanodine receptor Ca(2+) release channel function remained unaffected by the toxin. However, palytoxin significantly reduced Ca(2+) pumping of isolated SR vesicles. In current-clamped myocytes stimulated at 1 Hz, palytoxin induced a depolarization of the resting membrane potential that was accompanied by delayed afterdepolarizations. No major changes of action potential configuration were observed. The results demonstrate that palytoxin interferes with the function of the sarcolemmal Na(+)-K(+) pump and the SR Ca(2+) pump. The suggested mode of palytoxin toxicity in the atrium involves the conversion of Na(+)-K(+) pumps into nonselective cation channels as a primary event followed by depolarization, Na(+) accumulation, and Ca(2+) overload, which, in turn, causes arrhythmogenic [Ca(2+)](i) waves and delayed afterdepolarizations.